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Sample records for enhanced chondrogenic differentiation

  1. Enhancement of Matrix Metalloproteinase-2 (MMP-2 as a Potential Chondrogenic Marker during Chondrogenic Differentiation of Human Adipose-Derived Stem Cells

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    Yoshie Arai

    2016-06-01

    Full Text Available Human adipose-derived stem cells (hASCs have a capacity to undergo adipogenic, chondrogenic, and osteogenic differentiation. Recently, hASCs were applied to various fields including cell therapy for tissue regeneration. However, it is hard to predict the direction of differentiation of hASCs in real-time. Matrix metalloproteinases (MMPs are one family of proteolytic enzymes that plays a pivotal role in regulating the biology of stem cells. MMPs secreted by hASCs are expected to show different expression patterns depending on the differentiation state of hASCs because biological functions exhibit different patterns during the differentiation of stem cells. Here, we investigated proteolytic enzyme activity, especially MMP-2 activity, in hASCs during their differentiation. The activities of proteolytic enzymes and MMP-2 were higher during chondrogenic differentiation than during adipogenic and osteogenic differentiation. During chondrogenic differentiation, mRNA expression of MMP-2 and the level of the active form of MMP-2 were increased, which also correlated with Col II. It is concluded that proteolytic enzyme activity and the level of the active form of MMP-2 were increased during chondrogenic differentiation, which was accelerated in the presence of Col II protein. According to our findings, MMP-2 could be a candidate maker for real-time detection of chondrogenic differentiation of hASCs.

  2. Depletion of histone demethylase KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of stem cells from apical papilla

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    Dong, Rui [Laboratory of Molecular Signaling and Stem Cells Therapy, Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Capital Medical University School of Stomatology, Beijing 100050 (China); Yao, Rui [Department of Pediatrics, Stomatological Hospital of Nankai University, Tianjin 300041 (China); Du, Juan [Laboratory of Molecular Signaling and Stem Cells Therapy, Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Capital Medical University School of Stomatology, Beijing 100050 (China); Wang, Songlin [Molecular Laboratory for Gene Therapy and Tooth Regeneration, Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Capital Medical University School of Stomatology, Beijing 100050 (China); Department of Biochemistry and Molecular Biology, Capital Medical University School of Basic Medical Sciences, Beijing 100069 (China); Fan, Zhipeng, E-mail: zpfan@ccmu.edu.cn [Laboratory of Molecular Signaling and Stem Cells Therapy, Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction, Capital Medical University School of Stomatology, Beijing 100050 (China)

    2013-11-01

    Mesenchymal stem cells (MSCs) are a reliable resource for tissue regeneration, but the molecular mechanism underlying directed differentiation remains unclear; this has restricted potential MSC applications. The histone demethylase, lysine (K)-specific demethylase 2A (KDM2A), is evolutionarily conserved and ubiquitously expressed members of the JmjC-domain-containing histone demethylase family. A previous study determined that KDM2A can regulate the cell proliferation and osteo/dentinogenic differentiation of MSCs. It is not known whether KDM2A is involved in the other cell lineages differentiation of MSCs. Here, we show that depletion of KDM2A by short hairpin RNAs can enhance adipogenic and chondrogenic differentiation potentials in human stem cells from apical papilla (SCAPs). We found that the stemness-related genes, SOX2, and the embryonic stem cell master transcription factor, NANOG were significantly increased after silence of KDM2A in SCAPs. Moreover, we found that knock-down of the KDM2A co-factor, BCOR also up-regulated the mRNA levels of SOX2 and NANOG. Furthermore, Chromatin immunoprecipitation assays demonstrate that silence of KDM2A increased the histone H3 Lysine 4 (H3K4) trimethylation in the SOX2 and NANOG locus and regulates its expression. In conclusion, our results suggested that depletion of KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of SCAPs by up-regulated SOX2 and NANOG, BCOR also involved in this regulation as co-factor, and provided useful information to understand the molecular mechanism underlying directed differentiation in MSCs. - Highlights: • Depletion of KDM2A enhances adipogenic/chondrogenic differentiation in SCAPs. • Depletion of KDM2A enhances the differentiation of SCAPs by activate SOX2 and NANOG. • Silence of KDM2A increases histone H3 Lysine 4 trimethylation in SOX2 and NANOG. • BCOR is co-factor of KDM2A involved in the differentiation regulation.

  3. Depletion of histone demethylase KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of stem cells from apical papilla

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    Dong, Rui; Yao, Rui; Du, Juan; Wang, Songlin; Fan, Zhipeng

    2013-01-01

    Mesenchymal stem cells (MSCs) are a reliable resource for tissue regeneration, but the molecular mechanism underlying directed differentiation remains unclear; this has restricted potential MSC applications. The histone demethylase, lysine (K)-specific demethylase 2A (KDM2A), is evolutionarily conserved and ubiquitously expressed members of the JmjC-domain-containing histone demethylase family. A previous study determined that KDM2A can regulate the cell proliferation and osteo/dentinogenic differentiation of MSCs. It is not known whether KDM2A is involved in the other cell lineages differentiation of MSCs. Here, we show that depletion of KDM2A by short hairpin RNAs can enhance adipogenic and chondrogenic differentiation potentials in human stem cells from apical papilla (SCAPs). We found that the stemness-related genes, SOX2, and the embryonic stem cell master transcription factor, NANOG were significantly increased after silence of KDM2A in SCAPs. Moreover, we found that knock-down of the KDM2A co-factor, BCOR also up-regulated the mRNA levels of SOX2 and NANOG. Furthermore, Chromatin immunoprecipitation assays demonstrate that silence of KDM2A increased the histone H3 Lysine 4 (H3K4) trimethylation in the SOX2 and NANOG locus and regulates its expression. In conclusion, our results suggested that depletion of KDM2A enhanced the adipogenic and chondrogenic differentiation potentials of SCAPs by up-regulated SOX2 and NANOG, BCOR also involved in this regulation as co-factor, and provided useful information to understand the molecular mechanism underlying directed differentiation in MSCs. - Highlights: • Depletion of KDM2A enhances adipogenic/chondrogenic differentiation in SCAPs. • Depletion of KDM2A enhances the differentiation of SCAPs by activate SOX2 and NANOG. • Silence of KDM2A increases histone H3 Lysine 4 trimethylation in SOX2 and NANOG. • BCOR is co-factor of KDM2A involved in the differentiation regulation

  4. Overexpression of TGF-β1 enhances chondrogenic differentiation and proliferation of human synovium-derived stem cells

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    Kim, Yong Il; Ryu, Jae-Sung; Yeo, Jee Eun; Choi, Yun Jin; Kim, Yong Sang; Ko, Kinarm; Koh, Yong-Gon

    2014-01-01

    Highlights: • Continuous TGF-β1 overexpression in hSD-MSCs did not influence their phenotypes. • Retroviral-mediated transduction of TGFB1 in hSD-MSCs enhances cell proliferation. • TGF-β1 overexpression did not effect to adipo- or osteogenic potential of hSD-MSCs. • TGF-β1 overexpression in hSD-MSCs could stimulate and accelerate chondrogenesis. - Abstract: Transforming growth factor-beta (TGF-β) superfamily proteins play a critical role in proliferation, differentiation, and other functions of mesenchymal stem cells (MSCs). During chondrogenic differentiation of MSCs, TGF-β up-regulates chondrogenic gene expression by enhancing the expression of the transcription factor SRY (sex-determining region Y)-box9 (Sox9). In this study, we investigated the effect of continuous TGF-β1 overexpression in human synovium-derived MSCs (hSD-MSCs) on immunophenotype, differentiation potential, and proliferation rate. hSD-MSCs were transduced with recombinant retroviruses (rRV) encoding TGF-β1. The results revealed that continuous overexpression of TGF-β1 did not affect their phenotype as evidenced by flow cytometry and reverse transcriptase PCR (RT-PCR). In addition, continuous TGF-β1 overexpression strongly enhanced cell proliferation of hSD-MSCs compared to the control groups. Also, induction of chondrogenesis was more effective in rRV-TGFB-transduced hSD-MSCs as shown by RT-PCR for chondrogenic markers, toluidine blue staining and glycosaminoglycan (GAG)/DNA ratio. Our data suggest that overexpression of TGF-β1 positively enhances the proliferation and chondrogenic potential of hSD-MSCs

  5. Overexpression of TGF-β1 enhances chondrogenic differentiation and proliferation of human synovium-derived stem cells

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    Kim, Yong Il; Ryu, Jae-Sung; Yeo, Jee Eun; Choi, Yun Jin; Kim, Yong Sang [Center for Stem Cell and Arthritis Research, Department of Orthopedic Surgery, Yonsei Sarang Hospital, Seoul (Korea, Republic of); Ko, Kinarm [Center for Stem Cell Research, Institute of Advanced Biomedical Science, Konkuk University, Seoul 143-701 (Korea, Republic of); Koh, Yong-Gon, E-mail: yonseranglab@daum.net [Center for Stem Cell and Arthritis Research, Department of Orthopedic Surgery, Yonsei Sarang Hospital, Seoul (Korea, Republic of)

    2014-08-08

    Highlights: • Continuous TGF-β1 overexpression in hSD-MSCs did not influence their phenotypes. • Retroviral-mediated transduction of TGFB1 in hSD-MSCs enhances cell proliferation. • TGF-β1 overexpression did not effect to adipo- or osteogenic potential of hSD-MSCs. • TGF-β1 overexpression in hSD-MSCs could stimulate and accelerate chondrogenesis. - Abstract: Transforming growth factor-beta (TGF-β) superfamily proteins play a critical role in proliferation, differentiation, and other functions of mesenchymal stem cells (MSCs). During chondrogenic differentiation of MSCs, TGF-β up-regulates chondrogenic gene expression by enhancing the expression of the transcription factor SRY (sex-determining region Y)-box9 (Sox9). In this study, we investigated the effect of continuous TGF-β1 overexpression in human synovium-derived MSCs (hSD-MSCs) on immunophenotype, differentiation potential, and proliferation rate. hSD-MSCs were transduced with recombinant retroviruses (rRV) encoding TGF-β1. The results revealed that continuous overexpression of TGF-β1 did not affect their phenotype as evidenced by flow cytometry and reverse transcriptase PCR (RT-PCR). In addition, continuous TGF-β1 overexpression strongly enhanced cell proliferation of hSD-MSCs compared to the control groups. Also, induction of chondrogenesis was more effective in rRV-TGFB-transduced hSD-MSCs as shown by RT-PCR for chondrogenic markers, toluidine blue staining and glycosaminoglycan (GAG)/DNA ratio. Our data suggest that overexpression of TGF-β1 positively enhances the proliferation and chondrogenic potential of hSD-MSCs.

  6. Investigation of the optimal timing for chondrogenic priming of MSCs to enhance osteogenic differentiation in vitro as a bone tissue engineering strategy.

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    Freeman, F E; Haugh, M G; McNamara, L M

    2016-04-01

    Recent in vitro tissue engineering approaches have shown that chondrogenic priming of human bone marrow mesenchymal stem cells (MSCs) can have a positive effect on osteogenesis in vivo. However, whether chondrogenic priming is an effective in vitro bone regeneration strategy is not yet known. In particular, the appropriate timing for chondrogenic priming in vitro is unknown albeit that in vivo cartilage formation persists for a specific period before bone formation. The objective of this study is to determine the optimum time for chondrogenic priming of MSCs to enhance osteogenic differentiation by MSCs in vitro. Pellets derived from murine and human MSCs were cultured in six different media groups: two control groups (chondrogenic and osteogenic) and four chondrogenic priming groups (10, 14, 21 and 28 days priming). Biochemical analyses (Hoechst, sulfate glycosaminoglycan (sGAG), Alkaline Phosphate (ALP), calcium), histology (Alcian Blue, Alizarin Red) and immunohistochemistry (collagen types I, II and X) were performed on the samples at specific times. Our results show that after 49 days the highest amount of sGAG production occurred in MSCs chondrogenically primed for 21 days and 28 days. Moreover we found that chondrogenic priming of MSCs in vitro for specific amounts of time (14 days, 21 days) can have optimum influence on their mineralization capacity and can produce a construct that is mineralized throughout the core. Determining the optimum time for chondrogenic priming to enhance osteogenic differentiation in vitro provides information that might lead to a novel regenerative treatment for large bone defects, as well as addressing the major limitation of core degradation and construct failure. Copyright © 2013 John Wiley & Sons, Ltd.

  7. Peripheral blood aspirates overexpressing IGF-I via rAAV gene transfer undergo enhanced chondrogenic differentiation processes.

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    Frisch, Janina; Orth, Patrick; Rey-Rico, Ana; Venkatesan, Jagadeesh Kumar; Schmitt, Gertrud; Madry, Henning; Kohn, Dieter; Cucchiarini, Magali

    2017-11-01

    Implantation of peripheral blood aspirates induced towards chondrogenic differentiation upon genetic modification in sites of articular cartilage injury may represent a powerful strategy to enhance cartilage repair. Such a single-step approach may be less invasive than procedures based on the use of isolated or concentrated MSCs, simplifying translational protocols in patients. In this study, we provide evidence showing the feasibility of overexpressing the mitogenic and pro-anabolic insulin-like growth factor I (IGF-I) in human peripheral blood aspirates via rAAV-mediated gene transfer, leading to enhanced proliferative and chondrogenic differentiation (proteoglycans, type-II collagen, SOX9) activities in the samples relative to control (reporter rAAV-lacZ) treatment over extended periods of time (at least 21 days, the longest time-point evaluated). Interestingly, IGF-I gene transfer also triggered hypertrophic, osteo- and adipogenic differentiation processes in the aspirates, suggesting that careful regulation of IGF-I expression may be necessary to contain these events in vivo. Still, the current results demonstrate the potential of targeting human peripheral blood aspirates via therapeutic rAAV transduction as a novel, convenient tool to treat articular cartilage injuries. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  8. Anatomical region-dependent enhancement of 3-dimensional chondrogenic differentiation of human mesenchymal stem cells by soluble meniscus extracellular matrix.

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    Rothrauff, Benjamin B; Shimomura, Kazunori; Gottardi, Riccardo; Alexander, Peter G; Tuan, Rocky S

    2017-02-01

    Extracellular matrix (ECM) derived from decellularized tissues has been found to promote tissue neogenesis, most likely mediated by specific biochemical and physical signaling motifs that promote tissue-specific differentiation of progenitor cells. Decellularized ECM has been suggested to be efficacious for the repair of tissue injuries. However, decellularized meniscus contains a dense collagenous structure, which impedes cell seeding and infiltration and is not readily applicable for meniscus repair. In addition, the meniscus consists of two distinct anatomical regions that differ in vascularity and cellular phenotype. The purpose of this study was to explore the region-specific bioactivity of solubilized ECM derived from the inner and outer meniscal regions as determined in 2D and 3D cultures of adult mesenchymal stem cells (MSCs). When added as a medium supplement to 2D cultures of MSCs, urea-extracted fractions of the inner (imECM) and outer meniscal ECM (omECM) enhanced cell proliferation while imECM most strongly upregulated fibrochondrogenic differentiation on the basis of gene expression profiles. When added to 3D cultures of MSCs seeded in photocrosslinked methacrylated gelatin (GelMA) hydrogels, both ECM fractions upregulated chondrogenic differentiation as determined by gene expression and protein analyses, as well as elevated sulfated glycosaminoglycan sGAG content, compared to ECM-free controls. The chondrogenic effect at day 21 was most pronounced with imECM supplementation, but equivalent between ECM groups by day 42. Despite increased cartilage matrix, imECM and omECM constructs possessed compressive moduli similar to controls. In conclusion, soluble meniscal ECM may be considered for use as a tissue-specific reagent to enhance chondrogenesis for MSC-based 3D cartilage tissue engineering. The inner region of the knee meniscus is frequently injured and possesses a poor intrinsic healing capacity. Solubilized extracellular matrix (ECM) derived from

  9. Enhanced Chondrogenic Differentiation of Human Umbilical Cord Wharton's Jelly Derived Mesenchymal Stem Cells by GSK-3 Inhibitors.

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    Prapot Tanthaisong

    Full Text Available Articular cartilage is an avascular, alymphatic, and aneural system with very low regeneration potential because of its limited capacity for self-repair. Mesenchymal stem cells (MSCs are the preferred choice for cell-based therapies. Glycogen synthase kinase 3 (GSK-3 inhibitors are compounds that can induce the Wnt signaling pathway, which is involved in chondrogenesis and cartilage development. Here, we investigated the influence of lithium chloride (LiCl and SB216763 synergistically with TGF-β3 on chondrogenic differentiation in human mesenchymal stem cells derived from Wharton's jelly tissue (hWJ-MSCs. hWJ-MSCs were cultured and chondrogenic differentiation was induced in monolayer and pellet experiments using chondrogenic medium, chondrogenic medium supplemented with LiCl, or SB216763 for 4 weeks. After in vitro differentiation, cultured cells were examined for the expression of Sox9, ACAN, Col2a1, and β-catenin markers. Glycosaminoglycan (GAG accumulation was also examined by Alcian blue staining. The results indicated that SB216763 was more effective than LiCl as evidenced by a higher up-regulation of the expression of cartilage-specific markers, including Sox9, ACAN, Col2a1 as well as GAG accumulation. Moreover, collagen type II expression was strongly observed in cells cultured in the chondrogenic medium + SB216763 as evidenced by western blot analysis. Both treatments appeared to mediate the Wnt signaling pathway by up-regulating β-catenin gene expression. Further analyses showed that all treatments suppressed the progression of chondrocyte hypertrophy, determined by decreased expression of Col10a1 and Runx2. These results indicate that LiCl and SB216763 are potential candidates for further in vivo therapeutic trials and would be of great importance for cartilage regeneration.

  10. Enhancement of chondrogenic differentiation of rabbit mesenchymal stem cells by oriented nanofiber yarn-collagen type I/hyaluronate hybrid

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    Zheng, Xianyou; Wang, Wei; Liu, Shen; Wu, Jinglei; Li, Fengfeng; Cao, Lei; Liu, Xu-dong; Mo, Xiumei; Fan, Cunyi

    2016-01-01

    Cartilage defects cause joint pain and loss of mobility. It is crucial to induce the chondrogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) by both biological and structural signals in cartilage tissue engineering. Sponge-like scaffolds fabricated using native cartilage extracellular matrix components can induce the BMSC differentiation by biological signals and limited structural signals. In this study, an oriented poly(L-lactic acid)-co-poly(ε-caprolactone) P(LLA-CL)/collagen type I (Col-I) nanofiber yarn mesh, fabricated by dynamic liquid electrospinning served as a skeleton for a freeze-dried Col-I/hyaluronate (HA) chondral phase (SPONGE) containing both structural and biological signals to guide BMSC chondrogenic differentiation. In vitro results show that the Yarn Col-I/HA hybrid scaffold (Yarn-CH) promotes orientation, adhesion and proliferation of BMSCs better than SPONGE. Furthermore, BMSCs seeded on the Yarn-CH scaffold demonstrated a large increase in the glycosaminoglycan content and expression of collagen type II following a 21-day culture. - Highlights: • An oriented yarn was used as the skeleton of the sponge-like scaffold. • Both structural and biological signals were given for BMSC chondrogenic differentiation. • Yarn-CH promotes orientation and chondrogenesis differentiation of BMSCs. • Yarn-CH reproduces the superficial zone of the cartilage.

  11. Collagen-Coated Polytetrafluoroethane Membrane Inserts Enhances Chondrogenic Differentiation of Human Cord Blood Multi-Lineage Progenitor Cells

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    Munir, Samir; Søballe, Kjeld; Ulrich-Vinther, Michael

    culturing resulted in a multicellular layer tissue with formation of more cartilaginous tissue compared to micromass or CPP culture. In the membrane system MLPCs produced pellucid discs, 12 mm in diameter by 1 mm in thickness from 2x10^6 cells. The discs had hyaline-like cartilage extracellular matrix......Background: Articular chondrocytes and bone marrow-derived multipotent mesenchymal stromal cells (MSCs) are the favoured cells for cartilage tissue engineering. Umbilical cord blood has proven an alternative source of MSCs and moreover they may be more potent chondroprogenitor cells than bonemarrow...... with micromass or CPP cultures. Conclusions: In conclusion, we demonstrate that MLPCs possess’ chondrogenic potency, which increased when cultured scaffold-free on membrane inserts resulting in multicellular-layered hyaline-like cartilage tissue. Evaluating the effect of culturing pre-differentiated MLPCs on CPP...

  12. Amniotic Mesenchymal Stromal Cells Exhibit Preferential Osteogenic and Chondrogenic Differentiation and Enhanced Matrix Production Compared With Adipose Mesenchymal Stromal Cells.

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    Topoluk, Natasha; Hawkins, Richard; Tokish, John; Mercuri, Jeremy

    2017-09-01

    Therapeutic efficacy of various mesenchymal stromal cell (MSC) types for orthopaedic applications is currently being investigated. While the concept of MSC therapy is well grounded in the basic science of healing and regeneration, little is known about individual MSC populations in terms of their propensity to promote the repair and/or regeneration of specific musculoskeletal tissues. Two promising MSC sources, adipose and amnion, have each demonstrated differentiation and extracellular matrix (ECM) production in the setting of musculoskeletal tissue regeneration. However, no study to date has directly compared the differentiation potential of these 2 MSC populations. To compare the ability of human adipose- and amnion-derived MSCs to undergo osteogenic and chondrogenic differentiation. Controlled laboratory study. MSC populations from the human term amnion were quantified and characterized via cell counting, histologic assessment, and flow cytometry. Differentiation of these cells in comparison to commercially purchased human adipose-derived mesenchymal stromal cells (hADSCs) in the presence and absence of differentiation media was evaluated via reverse transcription polymerase chain reaction (PCR) for bone and cartilage gene transcript markers and histology/immunohistochemistry to examine ECM production. Analysis of variance and paired t tests were performed to compare results across all cell groups investigated. The authors confirmed that the human term amnion contains 2 primary cell types demonstrating MSC characteristics-(1) human amniotic epithelial cells (hAECs) and (2) human amniotic mesenchymal stromal cells (hAMSCs)-and each exhibited more than 90% staining for MSC surface markers (CD90, CD105, CD73). Average viable hAEC and hAMSC yields at harvest were 2.3 × 10 6 ± 3.7 × 10 5 and 1.6 × 10 6 ± 4.7 × 10 5 per milliliter of amnion, respectively. As well, hAECs and hAMSCs demonstrated significantly greater osteocalcin ( P = .025), aggrecan ( P

  13. IL-17 inhibits chondrogenic differentiation of human mesenchymal stem cells.

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    Masahiro Kondo

    Full Text Available OBJECTIVE: Mesenchymal stem cells (MSCs can differentiate into cells of mesenchymal lineages, such as osteoblasts and chondrocytes. Here we investigated the effects of IL-17, a key cytokine in chronic inflammation, on chondrogenic differentiation of human MSCs. METHODS: Human bone marrow MSCs were pellet cultured in chondrogenic induction medium containing TGF-β3. Chondrogenic differentiation was detected by cartilage matrix accumulation and chondrogenic marker gene expression. RESULTS: Over-expression of cartilage matrix and chondrogenic marker genes was noted in chondrogenic cultures, but was inhibited by IL-17 in a dose-dependent manner. Expression and phosphorylation of SOX9, the master transcription factor for chondrogenesis, were induced within 2 days and phosphorylated SOX9 was stably maintained until day 21. IL-17 did not alter total SOX9 expression, but significantly suppressed SOX9 phosphorylation in a dose-dependent manner. At day 7, IL-17 also suppressed the activity of cAMP-dependent protein kinase A (PKA, which is known to phosphorylate SOX9. H89, a selective PKA inhibitor, also suppressed SOX9 phosphorylation, expression of chondrogenic markers and cartilage matrix, and also decreased chondrogenesis. CONCLUSIONS: IL-17 inhibited chondrogenesis of human MSCs through the suppression of PKA activity and SOX9 phosphorylation. These results suggest that chondrogenic differentiation of MSCs can be inhibited by a mechanism triggered by IL-17 under chronic inflammation.

  14. Low-frequency, low-magnitude vibrations (LFLM enhances chondrogenic differentiation potential of human adipose derived mesenchymal stromal stem cells (hASCs

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    Krzysztof Marycz

    2016-02-01

    Full Text Available The aim of this study was to evaluate if low-frequency, low-magnitude vibrations (LFLM could enhance chondrogenic differentiation potential of human adipose derived mesenchymal stem cells (hASCs with simultaneous inhibition of their adipogenic properties for biomedical purposes. We developed a prototype device that induces low-magnitude (0.3 g low-frequency vibrations with the following frequencies: 25, 35 and 45 Hz. Afterwards, we used human adipose derived mesenchymal stem cell (hASCS, to investigate their cellular response to the mechanical signals. We have also evaluated hASCs morphological and proliferative activity changes in response to each frequency. Induction of chondrogenesis in hASCs, under the influence of a 35 Hz signal leads to most effective and stable cartilaginous tissue formation through highest secretion of Bone Morphogenetic Protein 2 (BMP-2, and Collagen type II, with low concentration of Collagen type I. These results correlated well with appropriate gene expression level. Simultaneously, we observed significant up-regulation of α3, α4, β1 and β3 integrins in chondroblast progenitor cells treated with 35 Hz vibrations, as well as Sox-9. Interestingly, we noticed that application of 35 Hz frequencies significantly inhibited adipogenesis of hASCs. The obtained results suggest that application of LFLM vibrations together with stem cell therapy might be a promising tool in cartilage regeneration.

  15. Expressions of pathologic markers in PRP based chondrogenic differentiation of human adipose derived stem cells.

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    Pakfar, Arezou; Irani, Shiva; Hanaee-Ahvaz, Hana

    2017-02-01

    Optimization of the differentiation medium through using autologous factors such as PRP is of great consideration, but due to the complex, variable and undefined composition of PRP on one hand and lack of control over the absolute regulatory mechanisms in in vitro conditions or disrupted and different mechanisms in diseased tissue microenvironments in in vivo conditions on the other hand, it is complicated and rather unpredictable to get the desired effects of PRP making it inevitable to monitor the possible pathologic or undesired differentiation pathways and therapeutic effects of PRP. Therefore, in this study the probable potential of PRP on inducing calcification, inflammation and angiogenesis in chondrogenically-differentiated cells was investigated. The expressions of chondrogenic, inflammatory, osteogenic and angiogenic markers from TGFβ or PRP-treated cells during chondrogenic differentiation of human adipose-derived stem cells (ADSCs) was evaluated. Expressions of Collagen II (Col II), Aggrecan, Sox9 and Runx2 were quantified using q-RT PCR. Expression of Col II and X was investigated by immunocytochemistry as well. Glycosaminoglycans (GAGs) production was also determined by GAG assay. Possible angiogenic/inflammatory potential was determined by quantitatively measuring the secreted VEGF, TNFα and phosphorylated VEGFR2 via ELISA. In addition, the calcification of the construct was monitored by measuring ALP activity and calcium deposition. Our data showed that PRP positively induced chondrogenesis; meanwhile the secretion of angiogenic and inflammatory markers was decreased. VEGFR2 phosphorylation and ALP activity had a decreasing trend, but tissue mineralization was enhanced upon treating with PRP. Although reduction in inflammatory/angiogenic potential of the chondrogenically differentiated constructs highlights the superior effectiveness of PRP in comparison to TGFβ for chondrogenic differentiation, yet further improvement of the PRP

  16. Human umbilical cord Wharton's jelly stem cells undergo enhanced chondrogenic differentiation when grown on nanofibrous scaffolds and in a sequential two-stage culture medium environment.

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    Fong, Chui-Yee; Subramanian, Arjunan; Gauthaman, Kalamegam; Venugopal, Jayarama; Biswas, Arijit; Ramakrishna, Seeram; Bongso, Ariff

    2012-03-01

    The current treatments used for osteoarthritis from cartilage damage have their disadvantages of donor site morbidity, complicated surgical interventions and risks of infection and graft rejection. Recent advances in tissue engineering have offered much promise in cartilage repair but the best cell source and in vitro system have not as yet been optimised. Human bone marrow mesenchymal stem cells (hBMSCs) have thus far been the cell of choice. However, we derived a unique stem cell from the human umbilical cord Wharton's jelly (hWJSC) that has properties superior to hBMSCs in terms of ready availability, prolonged stemness characteristics in vitro, high proliferation rates, wide multipotency, non-tumorigenicity and tolerance in allogeneic transplantation. We observed enhanced cell attachment, cell proliferation and chondrogenesis of hWJSCs over hBMSCs when grown on PCL/Collagen nanoscaffolds in the presence of a two-stage sequential complex/chondrogenic medium for 21 days. Improvement of these three parameters were confirmed via inverted optics, field emission scanning electron microscopy (FESEM), MTT assay, pellet diameters, Alcian blue histology and staining, glycosaminglycans (GAG) and hyaluronic acid production and expression of key chondrogenic genes (SOX9, Collagen type II, COMP, FMOD) using immunohistochemistry and real-time polymerase chain reaction (qRT-PCR). In separate experiments we demonstrated that the 16 ng/ml of basic fibroblast growth factor (bFGF) present in the complex medium may have contributed to driving chondrogenesis. We conclude that hWJSCs are an attractive stem cell source for inducing chondrogenesis in vitro when grown on nanoscaffolds and exposed sequentially first to complex medium and then followed by chondrogenic medium.

  17. Ultra-structural changes and expression of chondrogenic and hypertrophic genes during chondrogenic differentiation of mesenchymal stromal cells in alginate beads

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    Havva Dashtdar

    2016-03-01

    Full Text Available Chondrogenic differentiation of mesenchymal stromal cells (MSCs in the form of pellet culture and encapsulation in alginate beads has been widely used as conventional model for in vitro chondrogenesis. However, comparative characterization between differentiation, hypertrophic markers, cell adhesion molecule and ultrastructural changes during alginate and pellet culture has not been described. Hence, the present study was conducted comparing MSCs cultured in pellet and alginate beads with monolayer culture. qPCR was performed to assess the expression of chondrogenic, hypertrophic, and cell adhesion molecule genes, whereas transmission electron microscopy (TEM was used to assess the ultrastructural changes. In addition, immunocytochemistry for Collagen type II and aggrecan and glycosaminoglycan (GAG analysis were performed. Our results indicate that pellet and alginate bead cultures were necessary for chondrogenic differentiation of MSC. It also indicates that cultures using alginate bead demonstrated significantly higher (p < 0.05 chondrogenic but lower hypertrophic (p < 0.05 gene expressions as compared with pellet cultures. N-cadherin and N-CAM1 expression were up-regulated in second and third weeks of culture and were comparable between the alginate bead and pellet culture groups, respectively. TEM images demonstrated ultrastructural changes resembling cell death in pellet cultures. Our results indicate that using alginate beads, MSCs express higher chondrogenic but lower hypertrophic gene expression. Enhanced production of extracellular matrix and cell adhesion molecules was also observed in this group. These findings suggest that alginate bead culture may serve as a superior chondrogenic model, whereas pellet culture is more appropriate as a hypertrophic model of chondrogenesis.

  18. Novel biologically-inspired rosette nanotube PLLA scaffolds for improving human mesenchymal stem cell chondrogenic differentiation

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    Childs, Allie; Castro, Nathan J; Zhang, Lijie Grace; Hemraz, Usha D; Fenniri, Hicham

    2013-01-01

    Cartilage defects are a persistent issue in orthopedic tissue engineering where acute and chronic tissue damage stemming from osteoarthritis, trauma, and sport injuries, present a common and serious clinical problem. Unlike bone, cartilage repair continues to be largely intractable due to the tissue's inherently poor regenerative capacity. Thus, the objective of this study is to design a novel tissue engineered nanostructured cartilage scaffold via biologically-inspired self-assembling rosette nanotubes (RNTs) and biocompatible non-woven poly (l-lactic acid) (PLLA) for enhanced human bone marrow mesenchymal stem cell (hMSC) chondrogenic differentiation. Specifically, RNTs are a new class of biomimetic supramolecular nanomaterial obtained through the self-assembly of low-molecular-weight modified guanine/cytosine DNA base hybrids (the G∧C motif) in an aqueous environment. In this study, we synthesized a novel twin G∧C-based RNT (TB-RGDSK) functionalized with cell-favorable arginine–glycine–aspartic acid–serine–lysine (RGDSK) integrin binding peptide and a twin G∧C based RNT with an aminobutane linker molecule (TBL). hMSC adhesion, proliferation and chondrogenic differentiation were evaluated in vitro in scaffold groups consisting of biocompatible PLLA with TBL, 1:9 TB-RGDSK:TBL, and TB-RGDSK, respectively. Our results show that RNTs can remarkably increase total glycosaminoglycan, collagen, and protein production when compared to PLLA controls without nanotubes. Furthermore, the TB-RGDSK with 100% well-organized RGDSK peptides achieved the highest chondrogenic differentiation of hMSCs. The current in vitro study illustrated that RNT nanotopography and surface chemistry played an important role in enhancing hMSC chondrogenic differentiation thus making them promising for cartilage regeneration. (paper)

  19. The trans-well coculture of human synovial mesenchymal stem cells with chondrocytes leads to self-organization, chondrogenic differentiation, and secretion of TGFβ

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    Kubosch, Eva Johanna; Heidt, Emanuel; Bernstein, Anke

    2016-01-01

    BACKGROUND: Synovial mesenchymal stem cells (SMSC) possess a high chondrogenic differentiation potential, which possibly supports natural and surgically induced healing of cartilage lesions. We hypothesized enhanced chondrogenesis of SMSC caused by the vicinity of chondrocytes (CHDR). METHODS...

  20. Nicotine-induced chondrogenic differentiation of human bone marrow stromal cells in vitro.

    Science.gov (United States)

    Ying, Xiaozhou; Zhang, Wei; Cheng, Shaowen; Nie, Pengfei; Cheng, Xiaojie; Shen, Yue; Wang, Wei; Xue, Enxing; Chen, Qingyu; Kou, Dongquan; Peng, Lei; Zhang, Yu; Lu, Chuanzhu

    2012-11-01

    Nicotine has been reported that it has a dose-dependent effect on matrix mineralization by human bone marrow cells. However, there is no relevant research concerning on chondrogenic differentiation potential of bone marrow stromal stem cells (BMSCs) treated with nicotine in vitro. The aims of the study were to examine the effects of nicotine (0, 10(-7), 10(-6) and 10(-5) M) on the proliferation and chondrogenic differentiation of BMSCs from three healthy donors in vitro. BMSCs proliferation was analyzed by CCK8 assay and real-time polymerase chain reaction was used to assay the expression of type II collagen, aggrecan, type I collagen and type X collagen. The proteoglycan content was stained by Alcian blue, and the sulfated glycosaminoglycan (sGAG) content of BMSCs was quantified spectrofluorometrically using dimethylmethylene blue. The cell viability was not significantly impaired until up to a concentration of 10(-5) M nicotine. Nicotine promoted the proliferation and enhanced the expression of type II collagen at the level up to 10(-6) M (P < 0.05). The expression of aggrecan was reduced at the concentration of 10(-5) M nicotine at day 14 (P < 0.05), and there was no significant difference in aggrecan gene expression at 10(-7) and 10(-6) M nicotine levels compared to control group (n.s.). Also the fibroblastic and hypertrophic gene expressions were down-regulated in the chondrogenic medium with 10(-7)-10(-5) M nicotine (P < 0.05). It was implied that local application of nicotine at an appropriate concentration may be a promising approach for enhancing chondrogenic differentiation capacity of BMSCs in cell-based cartilage tissue engineering. Also these results indicate that nicotine maybe a potentially useful drug for the treatment of Osteoarthritis.

  1. An electromagnetic compressive force by cell exciter stimulates chondrogenic differentiation of bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Park, Sang-Hyug; Sim, Woo Young; Park, Sin Wook; Yang, Sang Sik; Choi, Byung Hyune; Park, So Ra; Park, Kwideok; Min, Byoung-Hyun

    2006-11-01

    In this study, we present a biological micro-electromechanical system and its application to the chondrogenic differentiation of rabbit bone marrow-derived mesenchymal stem cells (MSCs). Actuated by an electromagnetic force, the micro cell exciter was designed to deliver a cyclic compressive load (CCL) with various magnitudes. Two major parts in the system are an actuator and a cartridge-type chamber. The former has a permanent magnet and coil, and the latter is equipped with 7 sample dishes and 7 metal caps. Mixed with a 2.4% alginate solution, the alginate/MSC layers were positioned in the sample dishes; the caps contained chondrogenic defined medium without transforming growth factor-beta (TGF-beta). Once powered, the actuator coil-derived electromagnetic force pulled the metal caps down, compressing the samples. The cyclic load was given at 1-Hz frequency for 10 min twice a day. Samples in the dishes without a cap served as a control. The samples were analyzed at 3, 5, and 7 days after stimulation for cell viability, biochemical assays, histologic features, immunohistochemistry, and gene expression of the chondrogenic markers. Applied to the alginate/MSC layer, the CCL system enhanced the synthesis of cartilage-specific matrix proteins and the chondrogenic markers, such as aggrecan, type II collagen, and Sox9. We found that the micromechanically exerted CCL by the cell exciter was very effective in enhancing the chondrogenic differentiation of MSCs, even without using exogenous TGF-beta.

  2. Effects of BIO on proliferation and chondrogenic differentiation of mouse marrow derived mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Nasrin Fallah

    2013-06-01

    Full Text Available In vitroexpansion of mesenchymal stem cell (MSCs into large number is necessary fortheir application in cell-based treatment of articular cartilage defects. On the other hand,some studies have indicated that BIO (6-Bromoindirubin-3-Oxime possesses mitogeniceffects on cell culture. The objective of the present study was to examine the effect of BIO onin vitro expansion and chondrogenic differentiation of mouse marrow-derived MSCs. Theculture was established using bone marrow tissue obtained from 10 NMRI mice. MSC natureof the isolated cells was verified according to the minimal criteria proposed for MSC.Passaged-3 cells were seeded in 24-well culture plates and treated by 0.05, 0.01, 0.1, 1.0 and1.5 μM BIO forsevendays. The culture without BIO was taken as the control. At the end ofcultivation period, the cultures were examinedfor viable cell number which was then used tocalculate population doubling time (PDT. The BIO with higher proliferation-promoting effectwas investigated for its chondrogenic effect on MSC culture. There was significantly moreviable cells at the cultures treated by 0.1 μM BIO. At this culture the cells tended to doubletheir population in rapid rate (each 43.07 hr than the cells treated with the other BIOconcentrations (p< 0.05. Interestingly treatment of MSC chondrogenic culture with 0.1 μMBIO ledto the up-regulation of cartilage specific genes including aggrecan, collagen II andSox9. In conclusion BIO at 0.1 μM could enhance mouse MSC in vitro proliferation as well astheir chondrogenic differentiation. These findings would be of great importance for the fieldof regenerative medicine.

  3. Periodic heat shock accelerated the chondrogenic differentiation of human mesenchymal stem cells in pellet culture.

    Directory of Open Access Journals (Sweden)

    Jing Chen

    Full Text Available Osteoarthritis (OA is one of diseases that seriously affect elderly people's quality of life. Human mesenchymal stem cells (hMSCs offer a potential promise for the joint repair in OA patients. However, chondrogenic differentiation from hMSCs in vitro takes a long time (∼ 6 weeks and differentiated cells are still not as functionally mature as primary isolated chondrocytes, though chemical stimulations and mechanical loading have been intensively studied to enhance the hMSC differentiation. On the other hand, thermal stimulations of hMSC chondrogenesis have not been well explored. In this study, the direct effects of mild heat shock (HS on the differentiation of hMSCs into chondrocytes in 3D pellet culture were investigated. Periodic HS at 41 °C for 1 hr significantly increased sulfated glycosaminoglycan in 3D pellet culture at Day 10 of chondrogenesis. Immunohistochemical and Western Blot analyses revealed an increased expression of collagen type II and aggrecan in heat-shocked pellets than non heat-shocked pellets on Day 17 of chondrogenesis. In addition, HS also upregulated the expression of collagen type I and X as well as heat shock protein 70 on Day 17 and 24 of differentiation. These results demonstrate that HS accelerated the chondrogenic differentiation of hMSCs and induced an early maturation of chondrocytes differentiated from hMSCs. The results of this study will guide the design of future protocols using thermal treatments to facilitate cartilage regeneration with human mesenchymal stem cells.

  4. HIF-1α as a Regulator of BMP2-Induced Chondrogenic Differentiation, Osteogenic Differentiation, and Endochondral Ossification in Stem Cells

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    Nian Zhou

    2015-04-01

    Full Text Available Background/Aims: Joint cartilage defects are difficult to treat due to the limited self-repair capacities of cartilage. Cartilage tissue engineering based on stem cells and gene enhancement is a potential alternative for cartilage repair. Bone morphogenetic protein 2 (BMP2 has been shown to induce chondrogenic differentiation in mesenchymal stem cells (MSCs; however, maintaining the phenotypes of MSCs during cartilage repair since differentiation occurs along the endochondral ossification pathway. In this study, hypoxia inducible factor, or (HIF-1α, was determined to be a regulator of BMP2-induced chondrogenic differentiation, osteogenic differentiation, and endochondral bone formation. Methods: BMP2 was used to induce chondrogenic and osteogenic differentiation in stem cells and fetal limb development. After HIF-1α was added to the inducing system, any changes in the differentiation markers were assessed. Results: HIF-1α was found to potentiate BMP2-induced Sox9 and the expression of chondrogenesis by downstream markers, and inhibit Runx2 and the expression of osteogenesis by downstream markers in vitro. In subcutaneous stem cell implantation studies, HIF-1α was shown to potentiate BMP2-induced cartilage formation and inhibit endochondral ossification during ectopic bone/cartilage formation. In the fetal limb culture, HIF-1α and BMP2 synergistically promoted the expansion of the proliferating chondrocyte zone and inhibited chondrocyte hypertrophy and endochondral ossification. Conclusion: The results of this study indicated that, when combined with BMP2, HIF-1α induced MSC differentiation could become a new method of maintaining cartilage phenotypes during cartilage tissue engineering.

  5. Enamel Matrix Derivative has No Effect on the Chondrogenic Differentiation of Mesenchymal Stem Cells

    Energy Technology Data Exchange (ETDEWEB)

    Groeneveldt, Lisanne C.; Knuth, Callie; Witte-Bouma, Janneke [Department of Oral and Maxillofacial Surgery, Special Dental Care and Orthodontics, Erasmus University Medical Center, Rotterdam (Netherlands); O’Brien, Fergal J. [Tissue Engineering Research Group, Department of Anatomy, Royal College of Surgeons in Ireland, Dublin (Ireland); Wolvius, Eppo B.; Farrell, Eric, E-mail: e.farrell@erasmusmc.nl [Department of Oral and Maxillofacial Surgery, Special Dental Care and Orthodontics, Erasmus University Medical Center, Rotterdam (Netherlands)

    2014-09-02

    Background: Treatment of large bone defects due to trauma, tumor resection, or congenital abnormalities is challenging. Bone tissue engineering using mesenchymal stem cells (MSCs) represents a promising treatment option. However, the quantity and quality of engineered bone tissue are not sufficient to fill large bone defects. The aim of this study was to determine if the addition of enamel matrix derivative (EMD) improves in vitro chondrogenic priming of MSCs to ultimately improve in vivo MSC mediated endochondral bone formation. Methods: MSCs were chondrogenically differentiated in 2.0 × 10{sup 5} cell pellets in medium supplemented with TGFβ3 in the absence or presence of 1, 10, or 100 μg/mL EMD. Samples were analyzed for gene expression of RUNX2, Col II, Col X, and Sox9. Protein and glycoaminoglycan (GAG) production were also investigated via DMB assays, histology, and immunohistochemistry. Osteogenic and adipogenic differentiation capacity were also assessed. Results: The addition of EMD did not negatively affect chondrogenic differentiation of adult human MSCs. EMD did not appear to alter GAG production or expression of chondrogenic genes. Osteogenic and adipogenic differentiation were also unaffected though a trend toward decreased adipogenic gene expression was observed. Conclusion: EMD does not affect chondrogenic differentiation of adult human MSCs. As such the use of EMD in combination with chondrogenically primed MSCs for periodontal bone tissue repair is unlikely to have negative effects on MSC differentiation.

  6. Enamel Matrix Derivative has No Effect on the Chondrogenic Differentiation of Mesenchymal Stem Cells

    International Nuclear Information System (INIS)

    Groeneveldt, Lisanne C.; Knuth, Callie; Witte-Bouma, Janneke; O’Brien, Fergal J.; Wolvius, Eppo B.; Farrell, Eric

    2014-01-01

    Background: Treatment of large bone defects due to trauma, tumor resection, or congenital abnormalities is challenging. Bone tissue engineering using mesenchymal stem cells (MSCs) represents a promising treatment option. However, the quantity and quality of engineered bone tissue are not sufficient to fill large bone defects. The aim of this study was to determine if the addition of enamel matrix derivative (EMD) improves in vitro chondrogenic priming of MSCs to ultimately improve in vivo MSC mediated endochondral bone formation. Methods: MSCs were chondrogenically differentiated in 2.0 × 10 5 cell pellets in medium supplemented with TGFβ3 in the absence or presence of 1, 10, or 100 μg/mL EMD. Samples were analyzed for gene expression of RUNX2, Col II, Col X, and Sox9. Protein and glycoaminoglycan (GAG) production were also investigated via DMB assays, histology, and immunohistochemistry. Osteogenic and adipogenic differentiation capacity were also assessed. Results: The addition of EMD did not negatively affect chondrogenic differentiation of adult human MSCs. EMD did not appear to alter GAG production or expression of chondrogenic genes. Osteogenic and adipogenic differentiation were also unaffected though a trend toward decreased adipogenic gene expression was observed. Conclusion: EMD does not affect chondrogenic differentiation of adult human MSCs. As such the use of EMD in combination with chondrogenically primed MSCs for periodontal bone tissue repair is unlikely to have negative effects on MSC differentiation.

  7. Chondrogenic differentiation of ATDC5-cells under the influence of Mg and Mg alloy degradation

    Energy Technology Data Exchange (ETDEWEB)

    Martinez Sanchez, Adela H., E-mail: adela.martinez@hzg.de [Helmholtz-Zentrum Geesthacht, Institute of Material Research, Department for Structural Research on Macromolecules, Geesthacht (Germany); Feyerabend, Frank; Laipple, Daniel; Willumeit-Römer, Regine [Helmholtz-Zentrum Geesthacht, Institute of Material Research, Department for Structural Research on Macromolecules, Geesthacht (Germany); Weinberg, Annelie [Department of Orthopedics and Orthopedic Surgery, Medical University of Graz (Austria); Luthringer, Bérengère J.C. [Helmholtz-Zentrum Geesthacht, Institute of Material Research, Department for Structural Research on Macromolecules, Geesthacht (Germany)

    2017-03-01

    Biodegradable magnesium (Mg)-based materials are a potential alternative to permanent implants for application in children. Nevertheless effects of those materials on growth plate cartilage and chondrogenesis have not been previously evaluated. In vitro differentiation of ATDC5 cells was evaluated under the influence of pure Mg (PMg), Mg with 10 wt% of gadolinium (Mg-10Gd) and Mg with 2 wt% of silver (Mg-2Ag) degradation products (extracts) and direct cell culture on the materials. Gene expression showed an inhibitory effect on ATDC5 mineralization with the three extracts and a chondrogenic potential of Mg-10Gd. Cells cultured in Mg-10Gd and Mg-2Ag extracts showed the same proliferation and morphology than cells cultured in growth conditions. Mg-10Gd induced an increase in production of ECM and a bigger cell size, similar to the effects found with differentiation conditions. An increased metabolic activity was observed in cells cultured under the influence of Mg-10Gd extracts, indicated by an acidic pH during most of the culture period. After 7 days of culture on the materials, ATDC5 growth, distribution and ECM synthesis were higher on Mg-10Gd samples, followed by Mg-2Ag and PMg, which was influenced by the homogeneity and composition of the degradation layer. This study confirmed the tolerance of ATDC5 cells to Mg-based materials and a chondrogenic effect of Mg-10Gd. Further studies in vitro and in vivo are necessary to evaluate cell reactions to those materials, as well as the effects on bone growth and the biocompatibility of the alloying system in the body. - Highlights: • Degradation of PMg, and Mg-2Ag do not influence ATDC5 cells growth and chondrogenic redifferentiation. • Mg-10Gd enhances fast chondrogenic redifferentiation and expression of hyperthrophic markers on ATDC5 cells. • Further evaluation of the effects of PMg, Mg-10Gd and Mg-2Ag in vivo are necessary to confirm its potential for application in growing bones.

  8. Chondrogenic differentiation of ATDC5-cells under the influence of Mg and Mg alloy degradation

    International Nuclear Information System (INIS)

    Martinez Sanchez, Adela H.; Feyerabend, Frank; Laipple, Daniel; Willumeit-Römer, Regine; Weinberg, Annelie; Luthringer, Bérengère J.C.

    2017-01-01

    Biodegradable magnesium (Mg)-based materials are a potential alternative to permanent implants for application in children. Nevertheless effects of those materials on growth plate cartilage and chondrogenesis have not been previously evaluated. In vitro differentiation of ATDC5 cells was evaluated under the influence of pure Mg (PMg), Mg with 10 wt% of gadolinium (Mg-10Gd) and Mg with 2 wt% of silver (Mg-2Ag) degradation products (extracts) and direct cell culture on the materials. Gene expression showed an inhibitory effect on ATDC5 mineralization with the three extracts and a chondrogenic potential of Mg-10Gd. Cells cultured in Mg-10Gd and Mg-2Ag extracts showed the same proliferation and morphology than cells cultured in growth conditions. Mg-10Gd induced an increase in production of ECM and a bigger cell size, similar to the effects found with differentiation conditions. An increased metabolic activity was observed in cells cultured under the influence of Mg-10Gd extracts, indicated by an acidic pH during most of the culture period. After 7 days of culture on the materials, ATDC5 growth, distribution and ECM synthesis were higher on Mg-10Gd samples, followed by Mg-2Ag and PMg, which was influenced by the homogeneity and composition of the degradation layer. This study confirmed the tolerance of ATDC5 cells to Mg-based materials and a chondrogenic effect of Mg-10Gd. Further studies in vitro and in vivo are necessary to evaluate cell reactions to those materials, as well as the effects on bone growth and the biocompatibility of the alloying system in the body. - Highlights: • Degradation of PMg, and Mg-2Ag do not influence ATDC5 cells growth and chondrogenic redifferentiation. • Mg-10Gd enhances fast chondrogenic redifferentiation and expression of hyperthrophic markers on ATDC5 cells. • Further evaluation of the effects of PMg, Mg-10Gd and Mg-2Ag in vivo are necessary to confirm its potential for application in growing bones.

  9. The changing integrin expression and a role for integrin β8 in the chondrogenic differentiation of mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Vanessa L S LaPointe

    Full Text Available Many cartilage tissue engineering approaches aim to differentiate human mesenchymal stem cells (hMSCs into chondrocytes and develop cartilage in vitro by targeting cell-matrix interactions. We sought to better inform the design of cartilage tissue engineering scaffolds by understanding how integrin expression changes during chondrogenic differentiation. In three models of in vitro chondrogenesis, we studied the temporal change of cartilage phenotype markers and integrin subunits during the differentiation of hMSCs. We found that transcript expression of most subunits was conserved across the chondrogenesis models, but was significantly affected by the time-course of differentiation. In particular, ITGB8 was up-regulated and its importance in chondrogenesis was further established by a knockdown of integrin β8, which resulted in a non-hyaline cartilage phenotype, with no COL2A1 expression detected. In conclusion, we performed a systematic study of the temporal changes of integrin expression during chondrogenic differentiation in multiple chondrogenesis models, and revealed a role for integrin β8 in chondrogenesis. This work enhances our understanding of the changing adhesion requirements of hMSCs during chondrogenic differentiation and underlines the importance of integrins in establishing a cartilage phenotype.

  10. The Changing Integrin Expression and a Role for Integrin β8 in the Chondrogenic Differentiation of Mesenchymal Stem Cells

    Science.gov (United States)

    LaPointe, Vanessa L. S.; Verpoorte, Amanda; Stevens, Molly M.

    2013-01-01

    Many cartilage tissue engineering approaches aim to differentiate human mesenchymal stem cells (hMSCs) into chondrocytes and develop cartilage in vitro by targeting cell-matrix interactions. We sought to better inform the design of cartilage tissue engineering scaffolds by understanding how integrin expression changes during chondrogenic differentiation. In three models of in vitro chondrogenesis, we studied the temporal change of cartilage phenotype markers and integrin subunits during the differentiation of hMSCs. We found that transcript expression of most subunits was conserved across the chondrogenesis models, but was significantly affected by the time-course of differentiation. In particular, ITGB8 was up-regulated and its importance in chondrogenesis was further established by a knockdown of integrin β8, which resulted in a non-hyaline cartilage phenotype, with no COL2A1 expression detected. In conclusion, we performed a systematic study of the temporal changes of integrin expression during chondrogenic differentiation in multiple chondrogenesis models, and revealed a role for integrin β8 in chondrogenesis. This work enhances our understanding of the changing adhesion requirements of hMSCs during chondrogenic differentiation and underlines the importance of integrins in establishing a cartilage phenotype. PMID:24312400

  11. TrxR2 deficiencies promote chondrogenic differentiation and induce apoptosis of chondrocytes through mitochondrial reactive oxygen species

    International Nuclear Information System (INIS)

    Yan, Jidong; Xu, Jing; Fei, Yao; Jiang, Congshan; Zhu, Wenhua; Han, Yan; Lu, Shemin

    2016-01-01

    Thioredoxin reductase 2 (TrxR2) is a selenium (Se) containing protein. Se deficiency is associated with an endemic osteoarthropathy characterized by impaired cartilage formation. It is unclear whether TrxR2 have roles in cartilage function. We examined the effects of TrxR2 on chondrogenic ATDC5 cells through shRNA-mediated gene silencing of TrxR2. We demonstrated TrxR2 deficiencies could enhance chondrogenic differentiation and apoptosis of ATDC5 cells. TrxR2 deficiencies increased accumulation of cartilage glycosaminoglycans (GAGs) and mineralization. TrxR2 deficiencies also stimulated expression of extracellular (ECM) gene including Collagen II and Aggrecan. The enhanced chondrogenic properties were further confirmed by activation of Akt signaling which are required for chondrogenesis. In addition, TrxR2 deficiencies promoted chondrocyte proliferation through acceleration of cell cycle progression by increase in both S and G2/M phase cell distribution accompanied with induction of parathyroid hormone-related protein (PTHrP). Moreover, TrxR2 deficiencies induced chondrocyte death via apoptosis and increased cell sensitivity to exogenous oxidative stress. Furthermore, TrxR2 deficiencies induced emission of mitochondrial reactive oxygen species (ROS) without alteration of mitochondrial membrane potential and intracellular ATP content. Finally, treatment of TrxR2 deficiency cells with N-acetylcysteine (NAC) inhibited mitochondrial ROS production and chondrocyte apoptosis. NAC also prevented chondrogenic differentiation of TrxR2 deficiency cells by suppression of ECM gene expression, GAGs accumulation and mineralization, as well as attenuation of Akt signaling. Thus, TrxR2-mediated mitochondrial integrity is indispensable for chondrogenic differentiation of ATDC5 cells. TrxR2 deficiency-induced impaired proliferation and death of chondrocytes may be the pathological mechanism of the osteoarthropathy due to Se deficiency. Notably, this study also uncover the roles of

  12. TrxR2 deficiencies promote chondrogenic differentiation and induce apoptosis of chondrocytes through mitochondrial reactive oxygen species

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Jidong [Department of Human Anatomy, Histology and Embryology, School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an, Shaanxi 710061 (China); Xu, Jing [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an, Shaanxi 710061 (China); Fei, Yao [College of Life Sciences, Northwest University, Xi’an, Shaanxi Province 710069 (China); Jiang, Congshan; Zhu, Wenhua; Han, Yan [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an, Shaanxi 710061 (China); Lu, Shemin, E-mail: lushemin@xjtu.edu.cn [Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi’an Jiaotong University Health Science Center, Xi’an, Shaanxi 710061 (China); Key Laboratory of Environment and Genes Related to Diseases, Xi’an Jiaotong University, Ministry of Education of China (China)

    2016-05-15

    Thioredoxin reductase 2 (TrxR2) is a selenium (Se) containing protein. Se deficiency is associated with an endemic osteoarthropathy characterized by impaired cartilage formation. It is unclear whether TrxR2 have roles in cartilage function. We examined the effects of TrxR2 on chondrogenic ATDC5 cells through shRNA-mediated gene silencing of TrxR2. We demonstrated TrxR2 deficiencies could enhance chondrogenic differentiation and apoptosis of ATDC5 cells. TrxR2 deficiencies increased accumulation of cartilage glycosaminoglycans (GAGs) and mineralization. TrxR2 deficiencies also stimulated expression of extracellular (ECM) gene including Collagen II and Aggrecan. The enhanced chondrogenic properties were further confirmed by activation of Akt signaling which are required for chondrogenesis. In addition, TrxR2 deficiencies promoted chondrocyte proliferation through acceleration of cell cycle progression by increase in both S and G2/M phase cell distribution accompanied with induction of parathyroid hormone-related protein (PTHrP). Moreover, TrxR2 deficiencies induced chondrocyte death via apoptosis and increased cell sensitivity to exogenous oxidative stress. Furthermore, TrxR2 deficiencies induced emission of mitochondrial reactive oxygen species (ROS) without alteration of mitochondrial membrane potential and intracellular ATP content. Finally, treatment of TrxR2 deficiency cells with N-acetylcysteine (NAC) inhibited mitochondrial ROS production and chondrocyte apoptosis. NAC also prevented chondrogenic differentiation of TrxR2 deficiency cells by suppression of ECM gene expression, GAGs accumulation and mineralization, as well as attenuation of Akt signaling. Thus, TrxR2-mediated mitochondrial integrity is indispensable for chondrogenic differentiation of ATDC5 cells. TrxR2 deficiency-induced impaired proliferation and death of chondrocytes may be the pathological mechanism of the osteoarthropathy due to Se deficiency. Notably, this study also uncover the roles of

  13. The influence of scaffold microstructure on chondrogenic differentiation of mesenchymal stem cells

    International Nuclear Information System (INIS)

    Zhang, Jingjing; Ge, Zigang; Wu, Yingnan; Lee, Eng Hin; Thote, Tanushree; Yang, Zheng

    2014-01-01

    Different forms of biomaterials, including microspheres, sponges, hydrogels and nanofibres have been broadly used in cartilage regeneration; however, effects of internal structures of biomaterials on chondrogenesis of mesenchymal stem cells (MSCs) remain largely unexplored. Here we investigated the effect of physical microenvironments of sponges and hydrogels on chondrogenic differentiation of MSCs. MSCs, cultured in these two scaffold systems, were induced with TGF-β 3  in chondrogeneic differentiation medium and the chondrogenic differentiation was evaluated and compared after three weeks. MSCs in the sponges clustered with spindle morphologies, while they distributed homogenously with round morphologies in the hydrogel. The MSCs proliferated faster in the sponge compared to that in the hydrogel. Significantly higher glycosaminoglycan and collagen II were found in the sponges but not in the hydrogels. The different tissue formation ability of MSCs in these two systems could be attributed to the different metabolic requirements and the cellular events prerequisite in the chondrogenic process of MSCs. It is reasonable to conclude that sponges with relatively active microenvironments that facilitate cell–cell contacts and cell–matrix interaction are optimal for early stage of chondrogeneic differentiation. (paper)

  14. Micro-Topographies Promote Late Chondrogenic Differentiation Markers in the ATDC5 Cell Line

    NARCIS (Netherlands)

    Le, Bach Q.; Vasilevich, Aliaksei; Vermeulen, Steven; Hulshof, Frits; Stamatialis, Dimitrios F.; Van Blitterswijk, Clemens A.; De Boer, Jan

    2017-01-01

    Chemical and mechanical cues are well-established influencers of in vitro chondrogenic differentiation of ATDC5 cells. Here, we investigate the role of topographical cues in this differentiation process, a study not been explored before. Previously, using a library of surface micro-topographies we

  15. Micro-Topographies Promote Late Chondrogenic Differentiation Markers in the ATDC5 Cell Line.

    Science.gov (United States)

    Le, Bach Q; Vasilevich, Aliaksei; Vermeulen, Steven; Hulshof, Frits; Stamatialis, Dimitrios F; van Blitterswijk, Clemens A; de Boer, Jan

    2017-05-01

    Chemical and mechanical cues are well-established influencers of in vitro chondrogenic differentiation of ATDC5 cells. Here, we investigate the role of topographical cues in this differentiation process, a study not been explored before. Previously, using a library of surface micro-topographies we found some distinct patterns that induced alkaline phosphatase (ALP) production in human mesenchymal stromal cells. ALP is also a marker for hypertrophy, the end stage of chondrogenic differentiation preceding bone formation. Thus, we hypothesized that these patterns could influence end-stage chondrogenic differentiation of ATDC5 cells. In this study, we randomly selected seven topographies among the ALP influencing hits. Cells grown on these surfaces displayed varying nuclear shape and actin filament structure. When stimulated with insulin-transferrin-selenium (ITS) medium, nodule formation occurred and in some cases showed alignment to the topographical patterns. Gene expression analysis of cells growing on topographical surfaces in the presence of ITS medium revealed a downregulation of early markers and upregulation of late markers of chondrogenic differentiation compared to cells grown on a flat surface. In conclusion, we demonstrated that surface topography in addition to other cues can promote hypertrophic differentiation suitable for bone tissue engineering.

  16. Characterization of human adipose-derived stem cells and expression of chondrogenic genes during induction of cartilage differentiation.

    Science.gov (United States)

    Hamid, Adila A; Idrus, Ruszymah Bt Hj; Saim, Aminuddin Bin; Sathappan, Somasumdaram; Chua, Kien-Hui

    2012-01-01

    Understanding the changes in chondrogenic gene expression that are involved in the differentiation of human adipose-derived stem cells to chondrogenic cells is important prior to using this approach for cartilage repair. The aims of the study were to characterize human adipose-derived stem cells and to examine chondrogenic gene expression after one, two, and three weeks of induction. Human adipose-derived stem cells at passage 4 were evaluated by flow cytometry to examine the expression of surface markers. These adipose-derived stem cells were tested for adipogenic and osteogenic differentiation capacity. Ribonucleic acid was extracted from the cells for quantitative polymerase chain reaction analysis to determine the expression levels of chondrogenic genes after chondrogenic induction. Human adipose-derived stem cells were strongly positive for the mesenchymal markers CD90, CD73, CD44, CD9, and histocompatibility antigen and successfully differentiated into adipogenic and osteogenic lineages. The human adipose-derived stem cells aggregated and formed a dense matrix after chondrogenic induction. The expression of chondrogenic genes (collagen type II, aggrecan core protein, collagen type XI, COMP, and ELASTIN) was significantly higher after the first week of induction. However, a significantly elevated expression of collagen type X was observed after three weeks of chondrogenic induction. Human adipose-derived stem cells retain stem cell characteristics after expansion in culture to passage 4 and serve as a feasible source of cells for cartilage regeneration. Chondrogenesis in human adipose-derived stem cells was most prominent after one week of chondrogenic induction.

  17. Gene expression profile in human induced pluripotent stem cells: Chondrogenic differentiation in vitro, part A

    Science.gov (United States)

    Suchorska, Wiktoria Maria; Augustyniak, Ewelina; Richter, Magdalena; Trzeciak, Tomasz

    2017-01-01

    Human induced pluripotent stem cells (hiPSCs) offer promise in regenerative medicine, however more data are required to improve understanding of key aspects of the cell differentiation process, including how specific chondrogenic processes affect the gene expression profile of chondrocyte-like cells and the relative value of cell differentiation markers. The main aims of the present study were as follows: To determine the gene expression profile of chondrogenic-like cells derived from hiPSCs cultured in mediums conditioned with HC-402-05a cells or supplemented with transforming growth factor β3 (TGF-β3), and to assess the relative utility of the most commonly used chondrogenic markers as indicators of cell differentiation. These issues are relevant with regard to the use of human fibroblasts in the reprogramming process to obtain hiPSCs. Human fibroblasts are derived from the mesoderm and thus share a wide range of properties with chondrocytes, which also originate from the mesenchyme. Thus, the exclusion of dedifferentiation instead of chondrogenic differentiation is crucial. The hiPSCs were obtained from human primary dermal fibroblasts during a reprogramming process. Two methods, both involving embryoid bodies (EB), were used to obtain chondrocytes from the hiPSCs: EBs formed in a chondrogenic medium supplemented with TGF-β3 (10 ng/ml) and EBs formed in a medium conditioned with growth factors from HC-402-05a cells. Based on immunofluorescence and reverse transcription-quantiative polymerase chain reaction analysis, the results indicated that hiPSCs have the capacity for effective chondrogenic differentiation, in particular cells differentiated in the HC-402-05a-conditioned medium, which present morphological features and markers that are characteristic of mature human chondrocytes. By contrast, cells differentiated in the presence of TGF-β3 may demonstrate hypertrophic characteristics. Several genes [paired box 9, sex determining region Y-box (SOX) 5, SOX6

  18. Interleukin-6 inhibits early differentiation of ATDC5 chondrogenic progenitor cells.

    Science.gov (United States)

    Nakajima, Shoko; Naruto, Takuya; Miyamae, Takako; Imagawa, Tomoyuki; Mori, Masaaki; Nishimaki, Shigeru; Yokota, Shumpei

    2009-08-01

    Interleukin (IL)-6 is a causative agent of systemic juvenile idiopathic arthritis (sJIA), a chronic inflammatory disease complicated with severe growth impairment. Recent trials of anti-IL-6 receptor monoclonal antibody, tocilizumab, indicated that tocilizumab blocks IL-6/IL-6 receptor-mediated inflammation, and induces catch-up growth in children with sJIA. This study evaluates the effects of IL-6 on chondrogenesis by ATDC5 cells, a clonal murine chondrogenic cell line that provides an excellent model for studying endochondral ossification at growth plate. ATDC5 cells were examined for the expression of IL-6 receptor and gp130 by fluorescence-activated cell sorting analysis. Recombinant murine IL-6 was added to ATDC5 cultures to observe cell differentiation, using a quantitative RT-PCR for the chondrogenic differentiation markers type II collagen, aggrecan, and type X collagen. To block IL-6, the anti-mouse IL-6 receptor monoclonal antibody MR16-1 was added. As a result, the cells expressed IL-6 receptor and gp130. The expression of chondrogenic differentiation marker gene was reduced by IL-6, but this was abrogated by MR16-1. We conclude that IL-6 inhibits early chondrogenesis of ATDC5 cells suggesting that IL-6 may affect committed stem cells at a cellular level during chondrogenic differentiation of growth plate chondrocytes, and that IL-6 may be a cellular-level factor in growth impairment in sJIA.

  19. In vitro chondrogenic differentiation of human adipose-derived stem cells with silk scaffolds

    Directory of Open Access Journals (Sweden)

    Hyeon Joo Kim

    2012-12-01

    Full Text Available Human adipose-derived stem cells have shown chondrogenic differentiation potential in cartilage tissue engineering in combination with natural and synthetic biomaterials. In the present study, we hypothesized that porous aqueous-derived silk protein scaffolds would be suitable for chondrogenic differentiation of human adipose-derived stem cells. Human adipose-derived stem cells were cultured up to 6 weeks, and cell proliferation and chondrogenic differentiation were investigated and compared with those in conventional micromass culture. Cell proliferation, glycosaminoglycan, and collagen levels in aqueous-derived silk scaffolds were significantly higher than in micromass culture. Transcript levels of SOX9 and type II collagen were also upregulated in the cell–silk constructs at 6 weeks. Histological examination revealed that the pores of the silk scaffolds were filled with cells uniformly distributed. In addition, chondrocyte-specific lacunae formation was evident and distributed in the both groups. The results suggest the biodegradable and biocompatible three-dimensional aqueous-derived silk scaffolds provided an improved environment for chondrogenic differentiation compared to micromass culture.

  20. Platelet lysate induces chondrogenic differentiation of umbilical cord-derived mesenchymal stem cells.

    Science.gov (United States)

    Hassan, Ghmkin; Bahjat, Mohammad; Kasem, Issam; Soukkarieh, Chadi; Aljamali, Majd

    2018-01-01

    Articular cartilage has a poor capacity for self-repair, and thus still presents a major challenge in orthopedics. Mesenchymal stem cells (MSCs) are multipotent stem cells with the potential to differentiate into chondrocytes in the presence of transforming growth factor beta (TGF-β). Platelet lysate (PL) contains a relatively large number of growth factors, including TGF-β, and has been shown to ameliorate cartilage repair. Here, we investigated the ability of PL to direct chondrogenic differentiation of MSCs along with other standard differentiation components in a pellet culture system. We isolated and expanded MSCs from human umbilical cords using a PL-supplemented medium and characterized the cells based on immunophenotype and potential for differentiation to adipocytes and osteocytes. We further cultured MSCs as pellets in a chondrogenic-differentiation medium supplemented with PL. After 21 days, the pellets were processed for histological analysis and stained with alician blue and acridine orange. The expression of SOX9 was investigated using RT-PCR. MSCs maintained their stemness characteristics in the PL-supplemented medium. However, the distribution of cells in the pellets cultured in the PL-supplemented chondrogenic differentiation medium had a greater similarity to cartilage tissue-derived chondrocytes than to the negative control. The intense alician blue staining indicated an increased production of mucopolysaccharides in the differentiated pellets, which also showed elevated expression of SOX9 . Our data suggest that MSCs could be differentiated to chondrocytes in the presence of PL and absence of exogenous TGF-β. Further research needs to be conducted to understand the exact role and potential of PL in chondrogenic differentiation and chondrocyte regeneration.

  1. Characterization of lipidic markers of chondrogenic differentiation using mass spectrometry imaging.

    Science.gov (United States)

    Rocha, Beatriz; Cillero-Pastor, Berta; Eijkel, Gert; Bruinen, Anne L; Ruiz-Romero, Cristina; Heeren, Ron M A; Blanco, Francisco J

    2015-02-01

    Mesenchymal stem cells (MSC) are an interesting alternative for cell-based therapy of cartilage defects attributable to their capacity to differentiate toward chondrocytes in the process termed chondrogenesis. The metabolism of lipids has recently been associated with the modulation of chondrogenesis and also with the development of pathologies related to cartilage degeneration. Information about the distribution and modulation of lipids during chondrogenesis could provide a panel of putative chondrogenic markers. Thus, the discovery of new lipid chondrogenic markers could be highly valuable for improving MSC-based cartilage therapies. In this work, MS imaging was used to characterize the spatial distribution of lipids in human bone marrow MSCs during the first steps of chondrogenic differentiation. The analysis of MSC micromasses at days 2 and 14 of chondrogenesis by MALDI-MSI led to the identification of 20 different lipid species, including fatty acids, sphingolipids, and phospholipids. Phosphocholine, several sphingomyelins, and phosphatidylcholines were found to increase during the undifferentiated chondrogenic stage. A particularly detected lipid profile was verified by TOF secondary ion MS. Using this technology, a higher intensity of phosphocholine-related ions was observed in the peripheral region of the micromasses collected at day 14. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Development of a model system to analyze chondrogenic differentiation of mesenchymal stem cells

    Science.gov (United States)

    Ruedel, Anke; Hofmeister, Simone; Bosserhoff, Anja-Katrin

    2013-01-01

    High-density cell culture is widely used for the analysis of cartilage development of human mesenchymal stem cells (HMSCs) in vitro. Several cell culture systems, as micromass, pellet culture and alginate culture, are applied by groups in the field to induce chondrogenic differentiation of HMSCs. A draw back of all model systems is the high amount of cells necessary for the experiments. Further, handling of large experimental approaches is difficult due to culturing e.g. in 15 ml tubes. Therefore, we aimed to develop a new model system based on “hanging drop” cultures using 10 to 100 fold less cells. Here, we demonstrate that differentiation of chondrogenic cells was induced as previously shown in other model systems. Real time RT-PCR analysis demonstrated that Collagen type II and MIA/CD-RAP were upregulated during culturing whereas for induction of hypertrophic markers like Collagen type X and AP-2 epsilon treatment with TGF beta was needed. To further test the system, siRNA against Sox9 was used and effects on chondrogenic gene expression were evaluated. In summary, the hanging drop culture system was determined to be a promising tool for in vitro chondrogenic studies. PMID:24294400

  3. Human platelet lysate successfully promotes proliferation and subsequent chondrogenic differentiation of adipose-derived stem cells: a comparison with articular chondrocytes.

    Science.gov (United States)

    Hildner, F; Eder, M J; Hofer, K; Aberl, J; Redl, H; van Griensven, M; Gabriel, C; Peterbauer-Scherb, A

    2015-07-01

    Fetal calf serum (FCS) bears a potential risk for carrying diseases and eliciting immune reactions. Nevertheless, it still represents the gold standard as medium supplement in cell culture. In the present study, human platelet lysate (PL) was tested as an alternative to FCS for the expansion and subsequent chondrogenic differentiation of human adipose-derived stem cells (ASCs). ASCs were expanded with 10% FCS (group F) or 5% PL (group P). Subsequently, three-dimensional (3D) micromass pellets were created and cultured for 5 weeks in chondrogenic differentiation medium. Additionally, the de- and redifferentiation potential of human articular chondrocytes (HACs) was evaluated and compared to ASCs. Both HACs and ASCs cultured with PL showed strongly enhanced proliferation rates. Redifferentiation of HACs was possible for cells expanded up to 3.3 population doublings (PD). At this stage, PL-expanded HACs demonstrated better redifferentiation potential than FCS-expanded cells. ASCs could also be differentiated following extended passaging. Glycosaminoglycan (GAG) quantification and qRT-PCR of 10 cartilage related markers demonstrated a tendency for increased chondrogenic differentiation of PL-expanded ASCs compared to cells expanded with FCS. Histologically, collagen type II but also collagen type X was mainly present in group P. The present study demonstrates that PL strongly induces proliferation of ASCs, while the chondrogenic differentiation potential is retained. HACs also showed enhanced proliferation and even better redifferentiation when previously expanded with PL. This suggests that PL is superior to FCS as a supplement for the expansion of ASCs and HACs, particularly with regard to chondrogenic (re)differentiation. Copyright © 2013 John Wiley & Sons, Ltd.

  4. Generation of Col2a1-EGFP iPS cells for monitoring chondrogenic differentiation.

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    Taku Saito

    Full Text Available Induced pluripotent stem cells (iPSC are a promising cell source for cartilage regenerative medicine; however, the methods for chondrocyte induction from iPSC are currently developing and not yet sufficient for clinical application. Here, we report the establishment of a fluorescent indicator system for monitoring chondrogenic differentiation from iPSC to simplify screening for effective factors that induce chondrocytes from iPSC. We generated iPSC from embryonic fibroblasts of Col2a1-EGFP transgenic mice by retroviral transduction of Oct4, Sox2, Klf4, and c-Myc. Among the 30 clones of Col2a1-EGFP iPSC we established, two clones showed high expression levels of embryonic stem cell (ESC marker genes, similar to control ESC. A teratoma formation assay showed that the two clones were pluripotent and differentiated into cell types from all three germ layers. The fluorescent signal was observed during chondrogenic differentiation of the two clones concomitant with the increase in chondrocyte marker expression. In conclusion, Col2a1-EGFP iPSC are useful for monitoring chondrogenic differentiation and will contribute to research in cartilage regenerative medicine.

  5. TGF-β1 is Involved in Vitamin D-Induced Chondrogenic Differentiation of Bone Marrow-Derived Mesenchymal Stem Cells by Regulating the ERK/JNK Pathway

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    Xiaorui Jiang

    2017-08-01

    Full Text Available Background/Aims: Osteoarthritis (OA is characterized by degradation of cartilage, sole cell type of which is chondrocytes. Bone marrow-derived mesenchymal stem cells (BMSCs possess multipotency and can be directionally differentiated into chondrocytes under stimulation. This study was aimed to explore the possible roles of vitamin D and transforming growth factor-β1 (TGF-β1 in the chondrogenic differentiation of BMSCs. Methods: BMSCs were isolated from femurs and tibias of rats and characterized by flow cytometry. After stimulation with vitamin D, BMSC proliferation and migration were measured by Cell Counting Kit-8 (CCK-8 and Transwell assays, respectively. Chondrogenic differentiation was estimated through expression levels of specific markers by qRT-PCR and Western blot analysis. After stable transfection, the effects of aberrantly expressed TGF-β1 on vitamin D-induced alterations, including BMSC viability, migration and chondrogenic differentiation, were all evaluated utilizing CCK-8 assay, Transwell assay, qRT-PCR and Western blot analysis. Finally, the phosphorylation levels of key kinases in the extracellular signal-regulated kinase (ERK and c-Jun N-terminal kinase (JNK pathways were determined by Western blot analysis. Results: Vitamin D remarkably promoted BMSC viability, migration and chondrogenic differentiation. These alterations of BMSCs induced by vitamin D were reinforced by TGF-β1 overexpression while were reversed by TGF-β1 silencing. Additionally, the phosphorylation levels of ERK, JNK and c-Jun were enhanced by TGF-β1 overexpression but were reduced by TGF-β1 knockdown. Conclusion: Vitamin D promoted BMSC proliferation, migration and chondrogenic differentiation. TGF-β1 might be implicated in the vitamin D-induced alterations of BMSCs through regulating ERK/JNK pathway.

  6. Characterization of human adipose-derived stem cells and expression of chondrogenic genes during induction of cartilage differentiation

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    Adila A Hamid

    2012-01-01

    Full Text Available OBJECTIVES: Understanding the changes in chondrogenic gene expression that are involved in the differentiation of human adipose-derived stem cells to chondrogenic cells is important prior to using this approach for cartilage repair. The aims of the study were to characterize human adipose-derived stem cells and to examine chondrogenic gene expression after one, two, and three weeks of induction. MATERIALS AND METHODS: Human adipose-derived stem cells at passage 4 were evaluated by flow cytometry to examine the expression of surface markers. These adipose-derived stem cells were tested for adipogenic and osteogenic differentiation capacity. Ribonucleic acid was extracted from the cells for quantitative polymerase chain reaction analysis to determine the expression levels of chondrogenic genes after chondrogenic induction. RESULTS: Human adipose-derived stem cells were strongly positive for the mesenchymal markers CD90, CD73, CD44, CD9, and histocompatibility antigen and successfully differentiated into adipogenic and osteogenic lineages. The human adipose-derived stem cells aggregated and formed a dense matrix after chondrogenic induction. The expression of chondrogenic genes (collagen type II, aggrecan core protein, collagen type XI, COMP, and ELASTIN was significantly higher after the first week of induction. However, a significantly elevated expression of collagen type X was observed after three weeks of chondrogenic induction. CONCLUSION: Human adipose-derived stem cells retain stem cell characteristics after expansion in culture to passage 4 and serve as a feasible source of cells for cartilage regeneration. Chondrogenesis in human adiposederived stem cells was most prominent after one week of chondrogenic induction.

  7. Hypoxia-Inducible Factor 1 Is an Inductor of Transcription Factor Activating Protein 2 Epsilon Expression during Chondrogenic Differentiation

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    Stephan Niebler

    2015-01-01

    Full Text Available The transcription factor AP-2ε (activating enhancer-binding protein epsilon is expressed in cartilage of humans and mice. However, knowledge about regulatory mechanisms influencing AP-2ε expression is limited. Using quantitative real time PCR, we detected a significant increase in AP-2ε mRNA expression comparing initial and late stages of chondrogenic differentiation processes in vitro and in vivo. Interestingly, in these samples the expression pattern of the prominent hypoxia marker gene angiopoietin-like 4 (Angptl4 strongly correlated with that of AP-2ε suggesting that hypoxia might represent an external regulator of AP-2ε expression in mammals. In order to show this, experiments directly targeting the activity of hypoxia-inducible factor-1 (HIF1, the complex mediating responses to oxygen deprivation, were performed. While the HIF1-activating compounds 2,2′-dipyridyl and desferrioxamine resulted in significantly enhanced mRNA concentration of AP-2ε, siRNA against HIF1α led to a significantly reduced expression rate of AP-2ε. Additionally, we detected a significant upregulation of the AP-2ε mRNA level after oxygen deprivation. In sum, these different experimental approaches revealed a novel role for the HIF1 complex in the regulation of the AP-2ε gene in cartilaginous cells and underlined the important role of hypoxia as an important external regulatory stimulus during chondrogenic differentiation modulating the expression of downstream transcription factors.

  8. The effect of magnetic stimulation on the osteogenic and chondrogenic differentiation of human stem cells derived from the adipose tissue (hASCs)

    Energy Technology Data Exchange (ETDEWEB)

    Lima, João; Gonçalves, Ana I.; Rodrigues, Márcia T.; Reis, Rui L. [3Bs Research Group–Biomaterials, Biodegradables and Biomimetics, University of Minho, Guimarães (Portugal); ICVS/3Bs–PT Government Associate Laboratory, Braga/Guimarães (Portugal); Gomes, Manuela E., E-mail: megomes@dep.uminho.pt [3Bs Research Group–Biomaterials, Biodegradables and Biomimetics, University of Minho, Guimarães (Portugal); ICVS/3Bs–PT Government Associate Laboratory, Braga/Guimarães (Portugal)

    2015-11-01

    The use of magnetic nanoparticles (MNPs) towards the musculoskeletal tissues has been the focus of many studies, regarding MNPs ability to promote and direct cellular stimulation and orient tissue responses. This is thought to be mainly achieved by mechano-responsive pathways, which can induce changes in cell behavior, including the processes of proliferation and differentiation, in response to external mechanical stimuli. Thus, the application of MNP-based strategies in tissue engineering may hold potential to propose novel solutions for cell therapy on bone and cartilage strategies to accomplish tissue regeneration. The present work aims at studying the influence of MNPs on the osteogenic and chondrogenic differentiation of human adipose derived stem cells (hASCs). MNPs were incorporated in hASCs and cultured in medium supplemented for osteogenic and chondrogenic differentiation. Cultures were maintained up to 28 days with/without an external magnetic stimulus provided by a magnetic bioreactor, to determine if the MNPs alone could affect the osteogenic or chondrogenic phenotype of the hASCs. Results indicate that the incorporation of MNPs does not negatively affect the viability nor the proliferation of hASCs. Furthermore, Alizarin Red staining evidences an enhancement in extracellular (ECM) mineralization under the influence of an external magnetic field. Although not as evident as for osteogenic differentiation, Toluidine blue and Safranin-O stainings also suggest the presence of a cartilage-like ECM with glycosaminoglycans and proteoglycans under the magnetic stimulus provided. Thus, MNPs incorporated in hASCs under the influence of an external magnetic field have the potential to induce differentiation towards the osteogenic and chondrogenic lineages. - Highlights: • Cellular viability was not negatively influenced by the nanoparticles. • Chondrogenic medium influences more the synthesis of cartilage-like ECM than MNPs. • Synergetic effect among

  9. The effect of magnetic stimulation on the osteogenic and chondrogenic differentiation of human stem cells derived from the adipose tissue (hASCs)

    International Nuclear Information System (INIS)

    Lima, João; Gonçalves, Ana I.; Rodrigues, Márcia T.; Reis, Rui L.; Gomes, Manuela E.

    2015-01-01

    The use of magnetic nanoparticles (MNPs) towards the musculoskeletal tissues has been the focus of many studies, regarding MNPs ability to promote and direct cellular stimulation and orient tissue responses. This is thought to be mainly achieved by mechano-responsive pathways, which can induce changes in cell behavior, including the processes of proliferation and differentiation, in response to external mechanical stimuli. Thus, the application of MNP-based strategies in tissue engineering may hold potential to propose novel solutions for cell therapy on bone and cartilage strategies to accomplish tissue regeneration. The present work aims at studying the influence of MNPs on the osteogenic and chondrogenic differentiation of human adipose derived stem cells (hASCs). MNPs were incorporated in hASCs and cultured in medium supplemented for osteogenic and chondrogenic differentiation. Cultures were maintained up to 28 days with/without an external magnetic stimulus provided by a magnetic bioreactor, to determine if the MNPs alone could affect the osteogenic or chondrogenic phenotype of the hASCs. Results indicate that the incorporation of MNPs does not negatively affect the viability nor the proliferation of hASCs. Furthermore, Alizarin Red staining evidences an enhancement in extracellular (ECM) mineralization under the influence of an external magnetic field. Although not as evident as for osteogenic differentiation, Toluidine blue and Safranin-O stainings also suggest the presence of a cartilage-like ECM with glycosaminoglycans and proteoglycans under the magnetic stimulus provided. Thus, MNPs incorporated in hASCs under the influence of an external magnetic field have the potential to induce differentiation towards the osteogenic and chondrogenic lineages. - Highlights: • Cellular viability was not negatively influenced by the nanoparticles. • Chondrogenic medium influences more the synthesis of cartilage-like ECM than MNPs. • Synergetic effect among

  10. Curcumin inhibits cellular condensation and alters microfilament organization during chondrogenic differentiation of limb bud mesenchymal cells.

    Science.gov (United States)

    Kim, Dong Kyun; Kim, Song Ja; Kang, Shin Sung; Jin, Eun Jung

    2009-09-30

    Curcumin is a well known natural polyphenol product isolated from the rhizome of the plant Curcuma longa, anti-inflammatory agent for arthritis by inhibiting synthesis of inflammatory prostaglandins. However, the mechanisms by which curcumin regulates the functions of chondroprogenitor, such as proliferation, precartilage condensation, cytoskeletal organization or overall chondrogenic behavior, are largely unknown. In the present report, we investigated the effects and signaling mechanism of curcumin on the regulation of chondrogenesis. Treating chick limb bud mesenchymal cells with curcumin suppressed chondrogenesis by stimulating apoptotic cell death. It also inhibited reorganization of the actin cytoskeleton into a cortical pattern concomitant with rounding of chondrogenic competent cells and down-regulation of integrin beta1 and focal adhesion kinase (FAK) phosphorylation. Curcumin suppressed the phosphorylation of Akt leading to Akt inactivation. Activation of Akt by introducing a myristoylated, constitutively active form of Akt reversed the inhibitory actions of curcumin during chondrogenesis. In summary, for the first time, we describe biological properties of curcumin during chondrogenic differentiation of chick limb bud mesenchymal cells. Curcumin suppressed chondrogenesis by stimulating apoptotic cell death and down-regulating integrin-mediated reorganization of actin cytoskeleton via modulation of Akt signaling.

  11. Chondrogenic Differentiation of Mesenchymal Stem Cells: Challenges and Unfulfilled Expectations

    Science.gov (United States)

    Somoza, Rodrigo A.; Welter, Jean F.; Correa, Diego

    2014-01-01

    Articular cartilage repair and regeneration provides a substantial challenge in Regenerative Medicine because of the high degree of morphological and mechanical complexity intrinsic to hyaline cartilage due, in part, to its extracellular matrix. Cartilage remains one of the most difficult tissues to heal; even state-of-the-art regenerative medicine technology cannot yet provide authentic cartilage resurfacing. Mesenchymal stem cells (MSCs) were once believed to be the panacea for cartilage repair and regeneration, but despite years of research, they have not fulfilled these expectations. It has been observed that MSCs have an intrinsic differentiation program reminiscent of endochondral bone formation, which they follow after exposure to specific reagents as a part of current differentiation protocols. Efforts have been made to avoid the resulting hypertrophic fate of MSCs; however, so far, none of these has recreated a fully functional articular hyaline cartilage without chondrocytes exhibiting a hypertrophic phenotype. We reviewed the current literature in an attempt to understand why MSCs have failed to regenerate articular cartilage. The challenges that must be overcome before MSC-based tissue engineering can become a front-line technology for successful articular cartilage regeneration are highlighted. PMID:24749845

  12. The effect of dexamethasone and triiodothyronine on terminal differentiation of primary bovine chondrocytes and chondrogenically differentiated mesenchymal stem cells.

    Science.gov (United States)

    Randau, Thomas M; Schildberg, Frank A; Alini, Mauro; Wimmer, Matthias D; Haddouti, El-Mustapha; Gravius, Sascha; Ito, Keita; Stoddart, Martin J

    2013-01-01

    The newly evolved field of regenerative medicine is offering solutions in the treatment of bone or cartilage loss and deficiency. Mesenchymal stem cells, as well as articular chondrocytes, are potential cells for the generation of bone or cartilage. The natural mechanism of bone formation is that of endochondral ossification, regulated, among other factors, through the hormones dexamethasone and triiodothyronine. We investigated the effects of these hormones on articular chondrocytes and chondrogenically differentiated mesenchymal stem cells, hypothesizing that these hormones would induce terminal differentiation, with chondrocytes and differentiated stem cells being similar in their response. Using a 3D-alginate cell culture model, bovine chondrocytes and chondrogenically differentiated stem cells were cultured in presence of triiodothyronine or dexamethasone, and cell proliferation and extracellular matrix production were investigated. Collagen mRNA expression was measured by real-time PCR. Col X mRNA and alkaline phosphatase were monitored as markers of terminal differentiation, a prerequisite of endochondral ossification. The alginate culture system worked well, both for the culture of chondrocytes and for the chondrogenic differentiation of mesenchymal stem cells. Dexamethasone led to an increase in glycosaminoglycan production. Triiodothyronine increased the total collagen production only in chondrocytes, where it also induced signs of terminal differentiation, increasing both collagen X mRNA and alkaline phosphatase activity. Dexamethasone induced terminal differentiation in the differentiated stem cells. The immature articular chondrocytes used in this study seem to be able to undergo terminal differentiation, pointing to their possible role in the onset of degenerative osteoarthritis, as well as their potential for a cell source in bone tissue engineering. When chondrocyte-like cells, after their differentiation, can indeed be moved on towards terminal

  13. Paracrine and autocrine signals promoting full chondrogenic differentiation of a mesoblastic cell line.

    Science.gov (United States)

    Locker, Morgane; Kellermann, Odile; Boucquey, Marie; Khun, Huot; Huerre, Michel; Poliard, Anne

    2004-01-01

    The pluripotent mesoblastic C1 cell line was used under serum-free culture conditions to investigate how paracrine and autocrine signals cooperate to drive chondrogenesis. Sequential addition of two systemic hormones, dexamethasone and triiodothyronine, permits full chondrogenic differentiation. The cell intrinsic activation of the BMP signaling pathway and Sox9 expression occurring on mesoblastic condensation is insufficient for recruitment of the progenitors. Dexamethasone-dependent Sox9 upregulation is essential for chondrogenesis. Differentiation of lineage stem cells relies on cell autonomous regulations modulated by external signals. We used the pluripotent mesoblastic C1 cell line under serum-free culture conditions to investigate how paracrine and autocrine signals cooperate to induce differentiation of a precursor clone along the chondrogenic lineage. C1 cells, cultured as aggregates, were induced toward chondrogenesis by addition of 10(-7) M dexamethasone in serum-free medium. After 30 days, dexamethasone was replaced by 10 nM triiodothyronine to promote final hypertrophic conversion. Mature and hypertrophic phenotypes were characterized by immunocytochemistry using specific antibodies against types II and X collagens, respectively. Type II collagen, bone morphogenetic proteins (BMPs), BMP receptors, Smads, and Sox9 expression were monitored by reverse transcriptase-polymerase chain reaction (RT-PCR), Northern blot, and/or Western blot analysis. Once C1 cells have formed nodules, sequential addition of two systemic hormones is sufficient to promote full chondrogenic differentiation. In response to dexamethasone, nearly 100% of the C1 precursors engage in chondrogenesis and convert within 30 days into mature chondrocytes, which triggers a typical cartilage matrix. On day 25, a switch in type II procollagen mRNA splicing acted as a limiting step in the acquisition of the mature chondrocyte phenotype. On day 30, substitution of dexamethasone with

  14. The effects of dynamic and three-dimensional environments on chondrogenic differentiation of bone marrow stromal cells

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    Jung, Youngmee; Kim, Sang-Heon; Kim, Soo Hyun [Biomaterials Research Center, Korea Institute of Science and Technology, PO Box 131, Cheonryang, Seoul, 130-650 (Korea, Republic of); Kim, Young Ha, E-mail: soohkim@kist.re.k [Department of Materials Science and Engineering, Gwangju Institute of Science and Technology, 261 Cheomdan-gwagiro, Buk-gu, Gwangju 500-712 (Korea, Republic of)

    2009-10-15

    Articular cartilage is subjected to complex loading, which plays a major role in its growth, development and maintenance. Previously, we found that mechanical stimuli enhanced the development and function of engineered cartilage tissues in elastic mechano-active poly(lactide-co-caprolactone) (PLCL) scaffolds. In addition, it is well known that the three-dimensional spatial organization of cells and extracellular matrices in hydrogels is crucial to chondrogenesis. This study was conducted to enhance the chondrogenic differentiation of bone marrow stromal cells (BMSCs) in the hybrid scaffolds of fibrin gels and PLCL scaffolds in dynamic environments by compression. A highly elastic scaffold was fabricated from very elastic PLCL with 85% porosity and a 300-500{mu}m pore size using a gel-pressing method. A mixture of rabbit BMSCs and fibrin gels was then seeded onto the PLCL scaffolds and subjected to continuous compressive deformation of 5% strain at 0.1 Hz for 10 days in a chondrogenic medium containing 10 ng ml{sup -1} TGF-beta{sub 1}. The BMSCs-seeded scaffold constructs were then implanted subcutaneously into nude mice. As a control, the cell-PLCL scaffold constructs were cultured under dynamic conditions or the cell-PLCL/fibrin hybrid scaffold constructs and the cell-PLCL scaffold constructs were cultured under static conditions for 10 days in vitro. The results revealed that cells adhered onto the hybrid scaffolds of fibrin gels and PLCL scaffolds cultured under dynamic conditions. In addition, the accumulation of the extracellular matrix of cell-scaffold constructs, which was increased through mechanical stimulation, showed that chondrogenic differentiation was sustained and enhanced significantly in the stimulated hybrid scaffold constructs. Overall, the results of this study indicate that the proper periodic application of dynamic compression and the three-dimensional environments of the hybrid scaffolds composed of fibrin gels and elastic PLCL can encourage

  15. Gelatin-Based Hydrogels Promote Chondrogenic Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells In Vitro

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    Achim Salamon

    2014-02-01

    Full Text Available Due to the weak regeneration potential of cartilage, there is a high clinical incidence of articular joint disease, leading to a strong demand for cartilaginous tissue surrogates. The aim of this study was to evaluate a gelatin-based hydrogel for its suitability to support chondrogenic differentiation of human mesenchymal stem cells. Gelatin-based hydrogels are biodegradable, show high biocompatibility, and offer possibilities to introduce functional groups and/or ligands. In order to prove their chondrogenesis-supporting potential, a hydrogel film was developed and compared with standard cell culture polystyrene regarding the differentiation behavior of human mesenchymal stem cells. Cellular basis for this study were human adipose tissue-derived mesenchymal stem cells, which exhibit differentiation potential along the adipogenic, osteogenic and chondrogenic lineage. The results obtained show a promotive effect of gelatin-based hydrogels on chondrogenic differentiation of mesenchymal stem cells in vitro and therefore encourage subsequent in vivo studies.

  16. Gelatin-Based Hydrogels Promote Chondrogenic Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells In Vitro

    Science.gov (United States)

    Salamon, Achim; van Vlierberghe, Sandra; van Nieuwenhove, Ine; Baudisch, Frank; Graulus, Geert-Jan; Benecke, Verena; Alberti, Kristin; Neumann, Hans-Georg; Rychly, Joachim; Martins, José C.; Dubruel, Peter; Peters, Kirsten

    2014-01-01

    Due to the weak regeneration potential of cartilage, there is a high clinical incidence of articular joint disease, leading to a strong demand for cartilaginous tissue surrogates. The aim of this study was to evaluate a gelatin-based hydrogel for its suitability to support chondrogenic differentiation of human mesenchymal stem cells. Gelatin-based hydrogels are biodegradable, show high biocompatibility, and offer possibilities to introduce functional groups and/or ligands. In order to prove their chondrogenesis-supporting potential, a hydrogel film was developed and compared with standard cell culture polystyrene regarding the differentiation behavior of human mesenchymal stem cells. Cellular basis for this study were human adipose tissue-derived mesenchymal stem cells, which exhibit differentiation potential along the adipogenic, osteogenic and chondrogenic lineage. The results obtained show a promotive effect of gelatin-based hydrogels on chondrogenic differentiation of mesenchymal stem cells in vitro and therefore encourage subsequent in vivo studies. PMID:28788517

  17. Chondrogenic differentiation of mesenchymal stem cells in a leakproof collagen sponge

    International Nuclear Information System (INIS)

    Chen Guoping; Akahane, Daisuke; Kawazoe, Naoki; Yamamoto, Katsuyuki; Tateishi, Tetsuya

    2008-01-01

    A three-dimensional culture of mesenchymal stem cells (MSCs) in a porous scaffold has been developed as a promising strategy for cartilage tissue engineering. The chondrogenic differentiation of MSCs derived from human bone marrow was studied by culturing the cells in a novel scaffold constructed of leakproof collagen sponge. All the surfaces of the collagen sponge except the top were wrapped with a membrane that has pores smaller than the cells to protect against cell leakage during cell seeding. The cells adhered to the collagen, distributed evenly, and proliferated to fill the spaces in the sponge. Cell seeding efficiency was greater than 95%. The MSCs cultured in the collagen sponge in the presence of TGF-β3 and BMP6 expressed a high level of genes encoding type II and type X collagen, sox9, and aggrecan. Histological examination by HE staining indicated that the differentiated cells showed a round morphology. The extracellular matrices were positively stained by safranin O and toluidine blue. Immunostaining with anti-type II collagen and anti-cartilage proteoglycan showed that type II collagen and cartilage proteoglycan were detected around the cells. These results suggest the chondrogenic differentiation of MSCs when cultured in the collagen sponge in the presence of TGF-β3 and BMP6

  18. The effect of magnetic stimulation on the osteogenic and chondrogenic differentiation of human stem cells derived from the adipose tissue (hASCs)

    Science.gov (United States)

    Lima, João; Gonçalves, Ana I.; Rodrigues, Márcia T.; Reis, Rui L.; Gomes, Manuela E.

    2015-11-01

    The use of magnetic nanoparticles (MNPs) towards the musculoskeletal tissues has been the focus of many studies, regarding MNPs ability to promote and direct cellular stimulation and orient tissue responses. This is thought to be mainly achieved by mechano-responsive pathways, which can induce changes in cell behavior, including the processes of proliferation and differentiation, in response to external mechanical stimuli. Thus, the application of MNP-based strategies in tissue engineering may hold potential to propose novel solutions for cell therapy on bone and cartilage strategies to accomplish tissue regeneration. The present work aims at studying the influence of MNPs on the osteogenic and chondrogenic differentiation of human adipose derived stem cells (hASCs). MNPs were incorporated in hASCs and cultured in medium supplemented for osteogenic and chondrogenic differentiation. Cultures were maintained up to 28 days with/without an external magnetic stimulus provided by a magnetic bioreactor, to determine if the MNPs alone could affect the osteogenic or chondrogenic phenotype of the hASCs. Results indicate that the incorporation of MNPs does not negatively affect the viability nor the proliferation of hASCs. Furthermore, Alizarin Red staining evidences an enhancement in extracellular (ECM) mineralization under the influence of an external magnetic field. Although not as evident as for osteogenic differentiation, Toluidine blue and Safranin-O stainings also suggest the presence of a cartilage-like ECM with glycosaminoglycans and proteoglycans under the magnetic stimulus provided. Thus, MNPs incorporated in hASCs under the influence of an external magnetic field have the potential to induce differentiation towards the osteogenic and chondrogenic lineages.

  19. Chondrogenic differentiation of human mesenchymal stem cells cultured in a cobweb-like biodegradable scaffold

    International Nuclear Information System (INIS)

    Chen Guoping; Liu Dechang; Tadokoro, Mika; Hirochika, Rei; Ohgushi, Hajime; Tanaka, Junzo; Tateishi, Tetsuya

    2004-01-01

    Human mesenchymal stem cells (MSCs) were cultured in vitro in a cobweb-like biodegradable polymer scaffold: a poly(DL-lactic-co-glycolic acid)-collagen hybrid mesh in serum-free DMEM containing TGF-β3 for 1-10 weeks. The cells adhered to the hybrid mesh, distributed evenly, and proliferated to fill the spaces in the scaffold. The ability of the cells to express gene encoding type I collagen decreased, whereas its ability to express type II collagen and aggrecan increased. Histological examination by HE staining indicated that the cells showed fibroblast morphology at the early stage and became round after culture for 4 weeks. The cartilaginous matrices were positively stained by safranin O and toluidine blue. Immunostaining with anti-type II collagen and anti-cartilage proteoglycan showed that type II collagen and cartilage proteoglycan were detected around the cells. In addition, a homogeneous distribution of cartilaginous extracellular matrices was detected around the cells. These results suggest the chondrogenic differentiation of the mesenchymal stem cells in the hybrid mesh. The PLGA-collagen hybrid mesh enabled the aggregation of mesenchymal stem cells and provided a promotive microenvironment for the chondrogenic differentiation of the MSCs

  20. Fibroblast-Derived Extracellular Matrix Induces Chondrogenic Differentiation in Human Adipose-Derived Mesenchymal Stromal/Stem Cells in Vitro

    Directory of Open Access Journals (Sweden)

    Kevin Dzobo

    2016-08-01

    Full Text Available Mesenchymal stromal/stem cells (MSCs represent an area being intensively researched for tissue engineering and regenerative medicine applications. MSCs may provide the opportunity to treat diseases and injuries that currently have limited therapeutic options, as well as enhance present strategies for tissue repair. The cellular environment has a significant role in cellular development and differentiation through cell–matrix interactions. The aim of this study was to investigate the behavior of adipose-derived MSCs (ad-MSCs in the context of a cell-derived matrix so as to model the in vivo physiological microenvironment. The fibroblast-derived extracellular matrix (fd-ECM did not affect ad-MSC morphology, but reduced ad-MSC proliferation. Ad-MSCs cultured on fd-ECM displayed decreased expression of integrins α2 and β1 and subsequently lost their multipotency over time, as shown by the decrease in CD44, Octamer-binding transcription factor 4 (OCT4, SOX2, and NANOG gene expression. The fd-ECM induced chondrogenic differentiation in ad-MSCs compared to control ad-MSCs. Loss of function studies, through the use of siRNA and a mutant Notch1 construct, revealed that ECM-mediated ad-MSCs chondrogenesis requires Notch1 and β-catenin signaling. The fd-ECM also showed anti-senescence effects on ad-MSCs. The fd-ECM is a promising approach for inducing chondrogenesis in ad-MSCs and chondrogenic differentiated ad-MSCs could be used in stem cell therapy procedures.

  1. A reliable protocol for the isolation of viable, chondrogenically differentiated human mesenchymal stem cells from high-density pellet cultures.

    Science.gov (United States)

    Ullah, Mujib; Hamouda, Houda; Stich, Stefan; Sittinger, Michael; Ringe, Jochen

    2012-12-01

    Administration of chondrogenically differentiated mesenchymal stem cells (MSC) is discussed as a promising approach for the regenerative treatment of injured or diseased cartilage. The high-density pellet culture is the standard culture for chondrogenic differentiation, but cells in pellets secrete extracellular matrix (ECM) that they become entrapped in. Protocols for cell isolation from pellets often result in cell damage and dedifferentiation towards less differentiated MSC. Therefore, our aim was to develop a reliable protocol for the isolation of viable, chondrogenically differentiated MSC from high-density pellet cultures. Human bone marrow MSC were chondrogenically stimulated with transforming growth factor-β3, and the cartilaginous structure of the pellets was verified by alcian blue staining of cartilage proteoglycans, antibody staining of cartilage collagen type II, and quantitative real-time reverse-transcription polymerase chain reaction of the marker genes COL2A1 and SOX9. Trypsin and collagenases II and P were tested alone or in combination, and for different concentrations and times, to find a protocol for optimized pellet digestion. Whereas trypsin was not able to release viable cells, 90-min digestion with 300 U of collagenase II, 20 U of collagenase P, and 2 mM CaCl2 worked quite well and resulted in about 2.5×10(5) cells/pellet. The protocol was further optimized for the separation of released cells and ECM from each other. Cells were alcian blue and collagen type II positive and expressed COL2A1 and SOX9, verifying a chondrogenic character. However, they had different morphological shapes. The ECM was also uniformly alcian blue and collagen type II positive but showed different organizational and structural forms. To conclude, our protocol allows the reliable isolation of a defined number of viable, chondrogenically differentiated MSC from high-density pellet cultures. Such cells, as well as the ECM components, are of interest as

  2. Osteochondral defect repair using bilayered hydrogels encapsulating both chondrogenically and osteogenically pre-differentiated mesenchymal stem cells in a rabbit model

    NARCIS (Netherlands)

    Lam, J.; Lu, S.; Lee, E.J.; Trachtenberg, J.E.; Meretoja, V.V.; Dahlin, R.L.; van den Beucken, J.J.; Tabata, Y.; Wong, M.E.; Jansen, J.A.; Mikos, A.G.; Kasper, F.K.

    2014-01-01

    OBJECTIVE: To investigate the ability of cell-laden bilayered hydrogels encapsulating chondrogenically and osteogenically (OS) pre-differentiated mesenchymal stem cells (MSCs) to effect osteochondral defect repair in a rabbit model. By varying the period of chondrogenic pre-differentiation from 7

  3. Inhibition of TAK1 and/or JAK Can Rescue Impaired Chondrogenic Differentiation of Human Mesenchymal Stem Cells in Osteoarthritis-Like Conditions

    NARCIS (Netherlands)

    Beuningen, H.M. van; Vries-van Melle, M.L. de; Vitters, E.L.; Schreurs, W.; Berg, W.B. van den; Osch, G.J. van; Kraan, P.M. van der

    2014-01-01

    OBJECTIVE: To rescue chondrogenic differentiation of human mesenchymal stem cells (hMSCs) in osteoarthritic conditions by inhibition of protein kinases. METHODS: hMSCs were cultured in pellets. During early chondrogenic differentiation, these were exposed to osteoarthritic synovium-conditioned

  4. rFN/Cad-11-Modified Collagen Type II Biomimetic Interface Promotes the Adhesion and Chondrogenic Differentiation of Mesenchymal Stem Cells

    Science.gov (United States)

    Guo, Hongfeng; Zhang, Yuan; Li, Zhengsheng; Kang, Fei; Yang, Bo; Kang, Xia; Wen, Can; Yan, Yanfei; Jiang, Bo; Fan, Yujiang

    2013-01-01

    Properties of the cell-material interface are determining factors in the successful function of cells for cartilage tissue engineering. Currently, cell adhesion is commonly promoted through the use of polypeptides; however, due to their lack of complementary or modulatory domains, polypeptides must be modified to improve their ability to promote adhesion. In this study, we utilized the principle of matrix-based biomimetic modification and a recombinant protein, which spans fragments 7–10 of fibronectin module III (heterophilic motif ) and extracellular domains 1–2 of cadherin-11 (rFN/Cad-11) (homophilic motif ), to modify the interface of collagen type II (Col II) sponges. We showed that the designed material was able to stimulate cell proliferation and promote better chondrogenic differentiation of rabbit mesenchymal stem cells (MSCs) in vitro than both the FN modified surfaces and the negative control. Further, the Col II/rFN/Cad-11-MSCs composite stimulated cartilage formation in vivo; the chondrogenic effect of Col II alone was much less significant. These results suggested that the rFN/Cad-11-modified collagen type II biomimetic interface has dual biological functions of promoting adhesion and stimulating chondrogenic differentiation. This substance, thus, may serve as an ideal scaffold material for cartilage tissue engineering, enhancing repair of injured cartilage in vivo. PMID:23919505

  5. Characterization of Chondrogenic Gene Expression and Cartilage Phenotype Differentiation in Human Breast Adipose-Derived Stem Cells Promoted by Ginsenoside Rg1 In Vitro

    Directory of Open Access Journals (Sweden)

    Fang-Tian Xu

    2015-11-01

    Full Text Available Background/Aims: Investigating and understanding chondrogenic gene expression during the differentiation of human breast adipose-derived stem cells (HBASCs into chondrogenic cells is a prerequisite for the application of this approach for cartilage repair and regeneration. In this study, we aim to characterize HBASCs and to examine chondrogenic gene expression in chondrogenic inductive culture medium containing ginsenoside Rg1. Methods: Human breast adipose-derived stem cells at passage 3 were evaluated based on specific cell markers and their multilineage differentiation capacity. Cultured HBASCs were treated either with basic chondrogenic inductive conditioned medium alone (group A, control or with basic chondrogenic inductive medium plus 10 µg/ml (group B, 50 µg/ml (group C, or 100µg/ml ginsenoside Rg1 (group D. Cell proliferation was assessed using the CCK-8 assay for a period of 9 days. Two weeks after induction, the expression of chondrogenic genes (collagen type II, collagen type XI, ACP, COMP and ELASTIN was determined using real-time PCR in all groups. Results: The different concentrations of ginsenoside Rg1 that were added to the basic chondrogenic inductive culture medium promoted the proliferation of HBASCs at earlier stages (groups B, C, and D but resulted in chondrogenic phenotype differentiation and higher mRNA expression of collagen type II (CO-II, collagen type XI (CO-XI, acid phosphatase (ACP, cartilage oligomeric matrix protein (COMP and ELASTIN compared with the control (group A at later stages. The results reveal an obvious positive dose-effect relationship between ginsenoside Rg1 and the proliferation and chondrogenic phenotype differentiation of HBASCs in vitro. Conclusions: Human breast adipose-derived stem cells retain stem cell characteristics after expansion in culture through passage 3 and serve as a feasible source of cells for cartilage regeneration in vitro. Chondrogenesis in HBASCs was found to be prominent

  6. Hypoxic chondrogenic differentiation of human cord blood stem cells in structurally-graded polycaprolactone scaffolds

    DEFF Research Database (Denmark)

    Munir, Samir; Søballe, Kjeld; Ulrich-Vinther, Michael

    culturing resulted in a multicellular layer tissue with formation of more cartilaginous tissue compared to micromass or CPP culture. In the membrane system MLPCs produced pellucid discs, 12 mm in diameter by 1 mm in thickness from 2x10^6 cells. The discs had hyaline-like cartilage extracellular matrix......Background: Articular chondrocytes and bone marrow-derived multipotent mesenchymal stromal cells (MSCs) are the favoured cells for cartilage tissue engineering. Umbilical cord blood has proven an alternative source of MSCs and moreover they may be more potent chondroprogenitor cells than bonemarrow...... with micromass or CPP cultures. Conclusions: In conclusion, we demonstrate that MLPCs possess’ chondrogenic potency, which increased when cultured scaffold-free on membrane inserts resulting in multicellular-layered hyaline-like cartilage tissue. Evaluating the effect of culturing pre-differentiated MLPCs on CPP...

  7. Chondrogenic differentiation of ATDC5-cells under the influence of Mg and Mg alloy degradation.

    Science.gov (United States)

    Martinez Sanchez, Adela H; Feyerabend, Frank; Laipple, Daniel; Willumeit-Römer, Regine; Weinberg, Annelie; Luthringer, Bérengère J C

    2017-03-01

    Biodegradable magnesium (Mg)-based materials are a potential alternative to permanent implants for application in children. Nevertheless effects of those materials on growth plate cartilage and chondrogenesis have not been previously evaluated. In vitro differentiation of ATDC5 cells was evaluated under the influence of pure Mg (PMg), Mg with 10wt% of gadolinium (Mg-10Gd) and Mg with 2wt% of silver (Mg-2Ag) degradation products (extracts) and direct cell culture on the materials. Gene expression showed an inhibitory effect on ATDC5 mineralization with the three extracts and a chondrogenic potential of Mg-10Gd. Cells cultured in Mg-10Gd and Mg-2Ag extracts showed the same proliferation and morphology than cells cultured in growth conditions. Mg-10Gd induced an increase in production of ECM and a bigger cell size, similar to the effects found with differentiation conditions. An increased metabolic activity was observed in cells cultured under the influence of Mg-10Gd extracts, indicated by an acidic pH during most of the culture period. After 7days of culture on the materials, ATDC5 growth, distribution and ECM synthesis were higher on Mg-10Gd samples, followed by Mg-2Ag and PMg, which was influenced by the homogeneity and composition of the degradation layer. This study confirmed the tolerance of ATDC5 cells to Mg-based materials and a chondrogenic effect of Mg-10Gd. Further studies in vitro and in vivo are necessary to evaluate cell reactions to those materials, as well as the effects on bone growth and the biocompatibility of the alloying system in the body. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Chondroregulatory action of prolactin on proliferation and differentiation of mouse chondrogenic ATDC5 cells in 3-dimensional micromass cultures

    International Nuclear Information System (INIS)

    Seriwatanachai, Dutmanee; Krishnamra, Nateetip; Charoenphandhu, Narattaphol

    2012-01-01

    Highlights: ► Mouse chondrogenic ATDC5 cells expressed PRL receptor mRNAs and proteins. ► Low PRL concentration (10 ng/mL) increased chondrocyte viability and differentiation. ► Higher PRL concentrations (⩾100 ng/mL) decreased viability and increased apoptosis. -- Abstract: A recent investigation in lactating rats has provided evidence that the lactogenic hormone prolactin (PRL) increases endochondral bone growth and bone elongation, presumably by accelerating apoptosis of hypertrophic chondrocytes in the growth plate and/or subsequent chondrogenic matrix mineralization. Herein, we demonstrated the direct chondroregulatory action of PRL on proliferation, differentiation and apoptosis of chondrocytes in 3-dimensional micromass culture of mouse chondrogenic ATDC5 cell line. The results showed that ATDC5 cells expressed PRL receptor (PRLR) transcripts, and responded typically to PRL by downregulating PRLR expression. Exposure to a low PRL concentration of 10 ng/mL, comparable to the normal levels in male and non-pregnant female rats, increased chondrocyte viability, differentiation, proteoglycan accumulation, and mRNA expression of several chondrogenic differentiation markers, such as Sox9, ALP and Hspg2. In contrast, high PRL concentrations of ⩾100 ng/mL, comparable to the levels in pregnancy or lactation, decreased chondrocyte viability by inducing apoptosis, with no effect on chondrogenic marker expression. It could be concluded that chondrocytes directly but differentially responded to non-pregnant and pregnant/lactating levels of PRL, thus suggesting the stimulatory effect of PRL on chondrogenesis in young growing individuals, and supporting the hypothesis of hypertrophic chondrocyte apoptosis in the growth plate of lactating rats.

  9. Chondroregulatory action of prolactin on proliferation and differentiation of mouse chondrogenic ATDC5 cells in 3-dimensional micromass cultures

    Energy Technology Data Exchange (ETDEWEB)

    Seriwatanachai, Dutmanee [Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok (Thailand); Krishnamra, Nateetip [Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok (Thailand); Department of Physiology, Faculty of Science, Mahidol University, Bangkok (Thailand); Charoenphandhu, Narattaphol, E-mail: naratt@narattsys.com [Center of Calcium and Bone Research (COCAB), Faculty of Science, Mahidol University, Bangkok (Thailand); Department of Physiology, Faculty of Science, Mahidol University, Bangkok (Thailand)

    2012-03-30

    Highlights: Black-Right-Pointing-Pointer Mouse chondrogenic ATDC5 cells expressed PRL receptor mRNAs and proteins. Black-Right-Pointing-Pointer Low PRL concentration (10 ng/mL) increased chondrocyte viability and differentiation. Black-Right-Pointing-Pointer Higher PRL concentrations ( Greater-Than-Or-Slanted-Equal-To 100 ng/mL) decreased viability and increased apoptosis. -- Abstract: A recent investigation in lactating rats has provided evidence that the lactogenic hormone prolactin (PRL) increases endochondral bone growth and bone elongation, presumably by accelerating apoptosis of hypertrophic chondrocytes in the growth plate and/or subsequent chondrogenic matrix mineralization. Herein, we demonstrated the direct chondroregulatory action of PRL on proliferation, differentiation and apoptosis of chondrocytes in 3-dimensional micromass culture of mouse chondrogenic ATDC5 cell line. The results showed that ATDC5 cells expressed PRL receptor (PRLR) transcripts, and responded typically to PRL by downregulating PRLR expression. Exposure to a low PRL concentration of 10 ng/mL, comparable to the normal levels in male and non-pregnant female rats, increased chondrocyte viability, differentiation, proteoglycan accumulation, and mRNA expression of several chondrogenic differentiation markers, such as Sox9, ALP and Hspg2. In contrast, high PRL concentrations of Greater-Than-Or-Slanted-Equal-To 100 ng/mL, comparable to the levels in pregnancy or lactation, decreased chondrocyte viability by inducing apoptosis, with no effect on chondrogenic marker expression. It could be concluded that chondrocytes directly but differentially responded to non-pregnant and pregnant/lactating levels of PRL, thus suggesting the stimulatory effect of PRL on chondrogenesis in young growing individuals, and supporting the hypothesis of hypertrophic chondrocyte apoptosis in the growth plate of lactating rats.

  10. Poly(γ-Glutamic Acid) as an Exogenous Promoter of Chondrogenic Differentiation of Human Mesenchymal Stem/Stromal Cells.

    Science.gov (United States)

    Antunes, Joana C; Tsaryk, Roman; Gonçalves, Raquel M; Pereira, Catarina Leite; Landes, Constantin; Brochhausen, Christoph; Ghanaati, Shahram; Barbosa, Mário A; Kirkpatrick, C James

    2015-06-01

    Cartilage damage and/or aging effects can cause constant pain, which limits the patient's quality of life. Although different strategies have been proposed to enhance the limited regenerative capacity of cartilage tissue, the full production of native and functional cartilaginous extracellular matrix (ECM) has not yet been achieved. Poly(γ-glutamic acid) (γ-PGA), a naturally occurring polyamino acid, biodegradable into glutamate residues, has been explored for tissue regeneration. In this work, γ-PGA's ability to support the production of cartilaginous ECM by human bone marrow mesenchymal stem/stromal cells (MSCs) and nasal chondrocytes (NCs) was investigated. MSC and NC pellets were cultured in basal medium (BM), chondrogenic medium (CM), and CM-γ-PGA-supplemented medium (CM+γ-PGA) over a period of 21 days. Pellet size/shape was monitored with time. At 14 and 21 days of culture, the presence of sulfated glycosaminoglycans (sGAGs), type II collagen (Col II), Sox-9, aggrecan, type XI collagen (Col XI), type X collagen (Col X), calcium deposits, and type I collagen (Col I) was analyzed. After excluding γ-PGA's cytotoxicity, earlier cell condensation, higher sGAG content, Col II, Sox-9 (day 14), aggrecan, and Col X (day 14) production was observed in γ-PGA-supplemented MSC cultures, with no signs of mineralization or Col I. These effects were not evident with NCs. However, Sox-9 (at day 14) and Col X (at days 14 and 21) were increased, decreased, or absent, respectively. Overall, γ-PGA improved chondrogenic differentiation of MSCs, increasing ECM production earlier in culture. It is proposed that γ-PGA incorporation in novel biomaterials has a beneficial impact on future approaches for cartilage regeneration.

  11. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Z.H. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Li, X.L. [Department of Dermatology, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); He, X.J. [Department of Orthopedics, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Wu, B.J.; Xu, M. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Chang, H.M. [Department of Otolaryngology - Head and Neck Surgery, Affiliated Hospital of Xi' an Medical University, Xi' an (China); Zhang, X.H. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Xing, Z. [Department of Clinical Dentistry, Faculty of Dentistry, Center for Clinical Dental Research, University of Bergen, Bergen (Norway); Jing, X.H.; Kong, D.M.; Kou, X.H.; Yang, Y.Y. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China)

    2014-03-18

    SRY-related high-mobility-group box 9 (Sox9) gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs). After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering.

  12. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    Directory of Open Access Journals (Sweden)

    Z.H. Wang

    2014-04-01

    Full Text Available SRY-related high-mobility-group box 9 (Sox9 gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs. After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering.

  13. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    International Nuclear Information System (INIS)

    Wang, Z.H.; Li, X.L.; He, X.J.; Wu, B.J.; Xu, M.; Chang, H.M.; Zhang, X.H.; Xing, Z.; Jing, X.H.; Kong, D.M.; Kou, X.H.; Yang, Y.Y.

    2014-01-01

    SRY-related high-mobility-group box 9 (Sox9) gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs). After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering

  14. Extracellular Protease Inhibition Alters the Phenotype of Chondrogenically Differentiating Human Mesenchymal Stem Cells (MSCs) in 3D Collagen Microspheres.

    Science.gov (United States)

    Han, Sejin; Li, Yuk Yin; Chan, Barbara Pui

    2016-01-01

    Matrix remodeling of cells is highly regulated by proteases and their inhibitors. Nevertheless, how would the chondrogenesis of mesenchymal stem cells (MSCs) be affected, when the balance of the matrix remodeling is disturbed by inhibiting matrix proteases, is incompletely known. Using a previously developed collagen microencapsulation platform, we investigated whether exposing chondrogenically differentiating MSCs to intracellular and extracellular protease inhibitors will affect the extracellular matrix remodeling and hence the outcomes of chondrogenesis. Results showed that inhibition of matrix proteases particularly the extracellular ones favors the phenotype of fibrocartilage rather than hyaline cartilage in chondrogenically differentiating hMSCs by upregulating type I collagen protein deposition and type II collagen gene expression without significantly altering the hypertrophic markers at gene level. This study suggests the potential of manipulating extracellular proteases to alter the outcomes of hMSC chondrogenesis, contributing to future development of differentiation protocols for fibrocartilage tissues for intervertebral disc and meniscus tissue engineering.

  15. Extracellular Protease Inhibition Alters the Phenotype of Chondrogenically Differentiating Human Mesenchymal Stem Cells (MSCs in 3D Collagen Microspheres.

    Directory of Open Access Journals (Sweden)

    Sejin Han

    Full Text Available Matrix remodeling of cells is highly regulated by proteases and their inhibitors. Nevertheless, how would the chondrogenesis of mesenchymal stem cells (MSCs be affected, when the balance of the matrix remodeling is disturbed by inhibiting matrix proteases, is incompletely known. Using a previously developed collagen microencapsulation platform, we investigated whether exposing chondrogenically differentiating MSCs to intracellular and extracellular protease inhibitors will affect the extracellular matrix remodeling and hence the outcomes of chondrogenesis. Results showed that inhibition of matrix proteases particularly the extracellular ones favors the phenotype of fibrocartilage rather than hyaline cartilage in chondrogenically differentiating hMSCs by upregulating type I collagen protein deposition and type II collagen gene expression without significantly altering the hypertrophic markers at gene level. This study suggests the potential of manipulating extracellular proteases to alter the outcomes of hMSC chondrogenesis, contributing to future development of differentiation protocols for fibrocartilage tissues for intervertebral disc and meniscus tissue engineering.

  16. Glycosaminoglycan-Mimetic Signals Direct the Osteo/Chondrogenic Differentiation of Mesenchymal Stem Cells in a Three-Dimensional Peptide Nanofiber Extracellular Matrix Mimetic Environment.

    Science.gov (United States)

    Arslan, Elif; Guler, Mustafa O; Tekinay, Ayse B

    2016-04-11

    Recent efforts in bioactive scaffold development focus strongly on the elucidation of complex cellular responses through the use of synthetic systems. Designing synthetic extracellular matrix (ECM) materials must be based on understanding of cellular behaviors upon interaction with natural and artificial scaffolds. Hence, due to their ability to mimic both the biochemical and mechanical properties of the native tissue environment, supramolecular assemblies of bioactive peptide nanostructures are especially promising for development of bioactive ECM-mimetic scaffolds. In this study, we used glycosaminoglycan (GAG) mimetic peptide nanofiber gel as a three-dimensional (3D) platform to investigate how cell lineage commitment is altered by external factors. We observed that amount of fetal bovine serum (FBS) presented in the cell media had synergistic effects on the ability of GAG-mimetic nanofiber gel to mediate the differentiation of mesenchymal stem cells into osteogenic and chondrogenic lineages. In particular, lower FBS concentration in the culture medium was observed to enhance osteogenic differentiation while higher amount FBS promotes chondrogenic differentiation in tandem with the effects of the GAG-mimetic 3D peptide nanofiber network, even in the absence of externally administered growth factors. We therefore demonstrate that mesenchymal stem cell differentiation can be specifically controlled by the combined influence of growth medium components and a 3D peptide nanofiber environment.

  17. Effects of chitosan and bioactive glass modifications of knitted and rolled polylactide-based 96/4 L/D scaffolds on chondrogenic differentiation of adipose stem cells.

    Science.gov (United States)

    Ahtiainen, Katja; Sippola, Laura; Nurminen, Manu; Mannerström, Bettina; Haimi, Suvi; Suuronen, Riitta; Hyttinen, Jari; Ylikomi, Timo; Kellomäki, Minna; Miettinen, Susanna

    2015-01-01

    The performance of biodegradable knitted and rolled 3-dimensional (3D) polylactide-based 96/4 scaffolds modified with bioactive glass (BaG) 13-93, chitosan and both was compared with regard to the viability, proliferation and chondrogenic differentiation of rabbit adipose stem cells (ASCs). Scaffold porosities were determined by micro-computed tomography (μCT). Water absorption and degradation of scaffolds were studied during 28-day hydrolysis in Tris-buffer. Viability, number and differentiation of ASCs in PLA96/4 scaffolds were examined in vitro. The dimensions of the scaffolds were maintained during hydrolysis and mass loss was detected only in the BaG13-93 containing scaffolds. ASCs adhered and proliferated on each scaffold type. Cell aggregation and expression of chondral matrix components improved in all scaffold types in chondrogenic medium. Signs of hypertrophy were detected in the modified scaffolds but not in the plain PLA96/4 scaffold. Chondrogenic differentiation was most enhanced in the presence of chitosan. These findings indicate that the plain P scaffold provided a good 3D-matrix for ASC proliferation whereas the addition of chitosan to the PLA96/4 scaffold induced chondrogenic differentiation independent of the medium. Accordingly, a PLA96/4 scaffold modified by chitosan could provide a functional and bioactive basis for tissue-engineered chondral implants. Copyright © 2012 John Wiley & Sons, Ltd.

  18. Creating an Animal Model of Tendinopathy by Inducing Chondrogenic Differentiation with Kartogenin.

    Science.gov (United States)

    Yuan, Ting; Zhang, Jianying; Zhao, Guangyi; Zhou, Yiqin; Zhang, Chang-Qing; Wang, James H-C

    2016-01-01

    Previous animal studies have shown that long term rat treadmill running induces over-use tendinopathy, which manifests as proteoglycan accumulation and chondrocytes-like cells within the affected tendons. Creating this animal model of tendinopathy by long term treadmill running is however time-consuming, costly and may vary among animals. In this study, we used a new approach to develop an animal model of tendinopathy using kartogenin (KGN), a bio-compound that can stimulate endogenous stem/progenitor cells to differentiate into chondrocytes. KGN-beads were fabricated and implanted into rat Achilles tendons. Five weeks after implantation, chondrocytes and proteoglycan accumulation were found at the KGN implanted site. Vascularity as well as disorganization in collagen fibers were also present in the same site along with increased expression of the chondrocyte specific marker, collagen type II (Col. II). In vitro studies confirmed that KGN was released continuously from KGN-alginate in vivo beads and induced chondrogenic differentiation of tendon stem/progenitor cells (TSCs) suggesting that chondrogenesis after KGN-bead implantation into the rat tendons is likely due to the aberrant differentiation of TSCs into chondrocytes. Taken together, our results showed that KGN-alginate beads can be used to create a rat model of tendinopathy, which, at least in part, reproduces the features of over-use tendinopathy model created by long term treadmill running. This model is mechanistic (stem cell differentiation), highly reproducible and precise in creating localized tendinopathic lesions. It is expected that this model will be useful to evaluate the effects of various topical treatments such as NSAIDs and platelet-rich plasma (PRP) for the treatment of tendinopathy.

  19. Chondrogenic Differentiation of Defined Equine Mesenchymal Stem Cells Derived from Umbilical Cord Blood for Use in Cartilage Repair Therapy

    Directory of Open Access Journals (Sweden)

    Mélanie Desancé

    2018-02-01

    Full Text Available Cartilage engineering is a new strategy for the treatment of cartilage damage due to osteoarthritis or trauma in humans. Racehorses are exposed to the same type of cartilage damage and the anatomical, cellular, and biochemical properties of their cartilage are comparable to those of human cartilage, making the horse an excellent model for the development of cartilage engineering. Human mesenchymal stem cells (MSCs differentiated into chondrocytes with chondrogenic factors in a biomaterial appears to be a promising therapeutic approach for direct implantation and cartilage repair. Here, we characterized equine umbilical cord blood-derived MSCs (eUCB-MSCs and evaluated their potential for chondrocyte differentiation for use in cartilage repair therapy. Our results show that isolated eUCB-MSCs had high proliferative capacity and differentiated easily into osteoblasts and chondrocytes, but not into adipocytes. A three-dimensional (3D culture approach with the chondrogenic factors BMP-2 and TGF-β1 potentiated chondrogenic differentiation with a significant increase in cartilage-specific markers at the mRNA level (Col2a1, Acan, Snorc and the protein level (type II and IIB collagen without an increase in hypertrophic chondrocyte markers (Col10a1 and Mmp13 in normoxia and in hypoxia. However, these chondrogenic factors caused an increase in type I collagen, which can be reduced using small interfering RNA targeting Col1a2. This study provides robust data on MSCs characterization and demonstrates that eUCB-MSCs have a great potential for cartilage tissue engineering.

  20. Histone deacetylase 4 promotes TGF-beta1-induced synovium-derived stem cell chondrogenesis but inhibits chondrogenically differentiated stem cell hypertrophy.

    Science.gov (United States)

    Pei, Ming; Chen, Demeng; Li, Jingting; Wei, Lei

    2009-12-01

    The transforming growth factor-beta (TGF-beta) superfamily members play diverse roles in cartilage development and maintenance. TGF-beta up-regulates chondrogenic gene expression by enhancing transcription factor SRY (sex determining region Y)-box 9 (Sox9) and inhibits osteoblast differentiation by repressing runt-related transcription factor 2 (Runx2). Recently, histone deacetylases (HDACs) were reported to act as negative regulators of chondrocyte hypertrophy. It was speculated that HDAC4 may promote TGF-beta1-induced MSC chondrogenesis. In this study, the adenovirus-mediated HDAC4 gene (Ad.HDAC4) was utilized to infect synovium-derived stem cells (SDSCs). Adenovirus-mediated LacZ (Ad.LacZ) served as a control. The infected cells were centrifuged to form SDSC pellets followed by incubation in a serum-free chondrogenic medium for 15 days with or without 10ng/mL TGF-beta1. Transfection efficiency was determined in SDSCs using Ad.LacZ. Cytotoxicity was measured using lactate dehydrogenase assay. Histology, immunostaining, biochemical analysis, and real-time polymerase chain reaction were performed to assess chondrogenesis at protein and mRNA levels in infected SDSCs. Our data demonstrated that supplementation with TGF-beta1 could initiate and promote SDSC chondrogenesis; however, TGF-beta1 alone was insufficient to fully differentiate SDSCs into chondrocytes. Ad.HDAC4 could be efficiently transfected into SDSCs. Without TGF-beta1 treatment, HDAC4 had no effect on SDSC chondrogenesis; however, in the presence of TGF-beta1, HDAC4 could speed up and maintain a high level of chondrogenesis while down-regulating the hypertrophic marker - type X collagen expression. This study is the first report showing that HDAC4 overexpression promotes TGF-beta1-induced SDSC chondrogenesis but inhibits chondrogenically differentiated stem cell hypertrophy. The mechanism underlying this process needs further investigation.

  1. Long-Term Expansion, Enhanced Chondrogenic Potential, and Suppression of Endochondral Ossification of Adult Human MSCs via WNT Signaling Modulation

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    Roberto Narcisi

    2015-03-01

    Full Text Available Mesenchymal stem cells (MSCs are a potential source of chondrogenic cells for the treatment of cartilage disorders, but loss of chondrogenic potential during in vitro expansion and the propensity of cartilage to undergo hypertrophic maturation impede their therapeutic application. Here we report that the signaling protein WNT3A, in combination with FGF2, supports long-term expansion of human bone marrow-derived MSCs. The cells retained their chondrogenic potential and other phenotypic and functional properties of multipotent MSCs, which were gradually lost in the absence of WNT3A. Moreover, we discovered that endogenous WNT signals are the main drivers of the hypertrophic maturation that follows chondrogenic differentiation. Inhibition of WNT signals during differentiation prevented calcification and maintained cartilage properties following implantation in a mouse model. By maintaining potency during expansion and preventing hypertrophic maturation following differentiation, the modulation of WNT signaling removes two major obstacles that impede the clinical application of MSCs in cartilage repair.

  2. Inverse Regulation of Early and Late Chondrogenic Differentiation by Oxygen Tension Provides Cues for Stem Cell-Based Cartilage Tissue Engineering

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    Sophie Portron

    2015-01-01

    Full Text Available Background/Aims: Multipotent stem/stromal cells (MSC are considered promising for cartilage tissue engineering. However, chondrogenic differentiation of MSC can ultimately lead to the formation of hypertrophic chondrocytes responsible for the calcification of cartilage. To prevent the production of this calcified matrix at the articular site, the late hypertrophic differentiation of MSCs must be carefully controlled. Given that articular cartilage is avascular, we hypothesized that in addition to its stimulatory role in the early differentiation of chondrogenic cells, hypoxia may prevent their late hypertrophic conversion. Methods: Early and late chondrogenic differentiation were evaluated using human adipose MSC and murine ATDC5 cells cultured under either normoxic (21%O2 or hypoxic (5%O2 conditions. To investigate the effect of hypoxia on late chondrogenic differentiation, the transcriptional activity of hypoxia-inducible factor-1alpha (HIF-1α and HIF-2α were evaluated using the NoShift DNA-binding assay and through modulation of their activity (chemical inhibitor, RNA interference. Results: Our data demonstrate that low oxygen tension not only stimulates the early chondrogenic commitment of two complementary models of chondrogenic cells, but also inhibits their hypertrophic differentiation. Conclusion: These results suggest that hypoxia can be used as an instrumental tool to prevent the formation of a calcified matrix in MSC-based cartilage tissue engineering.

  3. Hydrostatic pressure promotes the proliferation and osteogenic/chondrogenic differentiation of mesenchymal stem cells: The roles of RhoA and Rac1

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    Yin-Hua Zhao

    2015-05-01

    Full Text Available Our previous studies have shown that hydrostatic pressure can serve as an active regulator for bone marrow mesenchymal stem cells (BMSCs. The current work further investigates the roles of cytoskeletal regulatory proteins Ras homolog gene family member A (RhoA and Ras-related C3 botulinum toxin substrate 1 (Rac1 in hydrostatic pressure-related effects on BMSCs. Flow cytometry assays showed that the hydrostatic pressure promoted cell cycle initiation in a RhoA- and Rac1-dependent manner. Furthermore, fluorescence assays confirmed that RhoA played a positive and Rac1 displayed a negative role in the hydrostatic pressure-induced F-actin stress fiber assembly. Western blots suggested that RhoA and Rac1 play central roles in the pressure-inhibited ERK phosphorylation, and Rac1 but not RhoA was involved in the pressure-promoted JNK phosphorylation. Finally, real-time polymerase chain reaction (PCR experiments showed that pressure promoted the expression of osteogenic marker genes in BMSCs at an early stage of osteogenic differentiation through the up-regulation of RhoA activity. Additionally, the PCR results showed that pressure enhanced the expression of chondrogenic marker genes in BMSCs during chondrogenic differentiation via the up-regulation of Rac1 activity. Collectively, our results suggested that RhoA and Rac1 are critical to the pressure-induced proliferation and differentiation, the stress fiber assembly, and MAPK activation in BMSCs.

  4. Hydrostatic pressure promotes the proliferation and osteogenic/chondrogenic differentiation of mesenchymal stem cells: The roles of RhoA and Rac1.

    Science.gov (United States)

    Zhao, Yin-Hua; Lv, Xin; Liu, Yan-Li; Zhao, Ying; Li, Qiang; Chen, Yong-Jin; Zhang, Min

    2015-05-01

    Our previous studies have shown that hydrostatic pressure can serve as an active regulator for bone marrow mesenchymal stem cells (BMSCs). The current work further investigates the roles of cytoskeletal regulatory proteins Ras homolog gene family member A (RhoA) and Ras-related C3 botulinum toxin substrate 1 (Rac1) in hydrostatic pressure-related effects on BMSCs. Flow cytometry assays showed that the hydrostatic pressure promoted cell cycle initiation in a RhoA- and Rac1-dependent manner. Furthermore, fluorescence assays confirmed that RhoA played a positive and Rac1 displayed a negative role in the hydrostatic pressure-induced F-actin stress fiber assembly. Western blots suggested that RhoA and Rac1 play central roles in the pressure-inhibited ERK phosphorylation, and Rac1 but not RhoA was involved in the pressure-promoted JNK phosphorylation. Finally, real-time polymerase chain reaction (PCR) experiments showed that pressure promoted the expression of osteogenic marker genes in BMSCs at an early stage of osteogenic differentiation through the up-regulation of RhoA activity. Additionally, the PCR results showed that pressure enhanced the expression of chondrogenic marker genes in BMSCs during chondrogenic differentiation via the up-regulation of Rac1 activity. Collectively, our results suggested that RhoA and Rac1 are critical to the pressure-induced proliferation and differentiation, the stress fiber assembly, and MAPK activation in BMSCs. Copyright © 2015. Published by Elsevier B.V.

  5. Monomeric, porous type II collagen scaffolds promote chondrogenic differentiation of human bone marrow mesenchymal stem cells in vitro

    Science.gov (United States)

    Tamaddon, M.; Burrows, M.; Ferreira, S. A.; Dazzi, F.; Apperley, J. F.; Bradshaw, A.; Brand, D. D.; Czernuszka, J.; Gentleman, E.

    2017-03-01

    Osteoarthritis (OA) is a common cause of pain and disability and is often associated with the degeneration of articular cartilage. Lesions to the articular surface, which are thought to progress to OA, have the potential to be repaired using tissue engineering strategies; however, it remains challenging to instruct cell differentiation within a scaffold to produce tissue with appropriate structural, chemical and mechanical properties. We aimed to address this by driving progenitor cells to adopt a chondrogenic phenotype through the tailoring of scaffold composition and physical properties. Monomeric type-I and type-II collagen scaffolds, which avoid potential immunogenicity associated with fibrillar collagens, were fabricated with and without chondroitin sulfate (CS) and their ability to stimulate the chondrogenic differentiation of human bone marrow-derived mesenchymal stem cells was assessed. Immunohistochemical analyses showed that cells produced abundant collagen type-II on type-II scaffolds and collagen type-I on type-I scaffolds. Gene expression analyses indicated that the addition of CS - which was released from scaffolds quickly - significantly upregulated expression of type II collagen, compared to type-I and pure type-II scaffolds. We conclude that collagen type-II and CS can be used to promote a more chondrogenic phenotype in the absence of growth factors, potentially providing an eventual therapy to prevent OA.

  6. Curcumin mediated suppression of nuclear factor-κB promotes chondrogenic differentiation of mesenchymal stem cells in a high-density co-culture microenvironment.

    Science.gov (United States)

    Buhrmann, Constanze; Mobasheri, Ali; Matis, Ulrike; Shakibaei, Mehdi

    2010-01-01

    Osteoarthritis (OA) and rheumatoid arthritis (RA) are characterised by joint inflammation and cartilage degradation. Although mesenchymal stem cell (MSC)-like progenitors are resident in the superficial zone of articular cartilage, damaged tissue does not possess the capacity for regeneration. The high levels of pro-inflammatory cytokines present in OA/RA joints may impede the chondrogenic differentiation of these progenitors. Interleukin (IL)-1β activates the transcription factor nuclear factor-κB (NF-κB), which in turn activates proteins involved in matrix degradation, inflammation and apoptosis. Curcumin is a phytochemical capable of inhibiting IL-1β-induced activation of NF-κB and expression of apoptotic and pro-inflammatory genes in chondrocytes. Therefore, the aim of the present study was to evaluate the influence of curcumin on IL-1β-induced NF-κB signalling pathway in MSCs during chondrogenic differentiation. MSCs were either cultured in a ratio of 1:1 with primary chondrocytes in high-density culture or cultured alone in monolayer with/without curcumin and/or IL-1β. We demonstrate that although curcumin alone does not have chondrogenic effects on MSCs, it inhibits IL-1β-induced activation of NF-κB, activation of caspase-3 and cyclooxygenase-2 in MSCs time and concentration dependently, as it does in chondrocytes. In IL-1β stimulated co-cultures, four-hour pre-treatment with curcumin significantly enhanced the production of collagen type II, cartilage specific proteoglycans (CSPGs), β1-integrin, as well as activating MAPKinase signaling and suppressing caspase-3 and cyclooxygenase-2. Curcumin treatment may help establish a microenvironment in which the effects of pro-inflammatory cytokines are antagonized, thus facilitating chondrogenesis of MSC-like progenitor cells in vivo. This strategy may support the regeneration of articular cartilage.

  7. Delta-like 1/fetal antigen-1 (Dlk1/FA1) is a novel regulator of chondrogenic cell differentiation via inhibition of the Akt kinase-dependent pathway

    DEFF Research Database (Denmark)

    Chen, Li; Qanie, Diyako; Jafari, Abbas

    2011-01-01

    Delta-like 1 (Dlk1, also known as fetal antigen-1, FA1) is a member of Notch/Delta family that inhibits adipocyte and osteoblast differentiation; however, its role in chondrogenesis is still not clear. Thus, we overexpressed Dlk1/FA1 in mouse embryonic ATDC5 cells and tested its effects...... on chondrogenic differentiation. Dlk1/FA1 inhibited insulin-induced chondrogenic differentiation as evidenced by reduction of cartilage nodule formation and gene expression of aggrecan, collagen Type II and X. Similar effects were obtained either by using Dlk1/FA1-conditioned medium or by addition of a purified......, secreted, form of Dlk1 (FA1) directly to the induction medium. The inhibitory effects of Dlk1/FA1 were dose-dependent and occurred irrespective of the chondrogenic differentiation stage: proliferation, differentiation, maturation, or hypertrophic conversion. Overexpression or addition of the Dlk1/FA1...

  8. Multilayered dense collagen-silk fibroin hybrid: a platform for mesenchymal stem cell differentiation towards chondrogenic and osteogenic lineages.

    Science.gov (United States)

    Ghezzi, Chiara E; Marelli, Benedetto; Donelli, Ilaria; Alessandrino, Antonio; Freddi, Giuliano; Nazhat, Showan N

    2017-07-01

    Type I collagen is a major structural and functional protein in connective tissues. However, collagen gels exhibit unstable geometrical properties, arising from extensive cell-mediated contraction. In an effort to stabilize collagen-based hydrogels, plastic compression was used to hybridize dense collagen (DC) with electrospun silk fibroin (SF) mats, generating multilayered DC-SF-DC constructs. Seeded mesenchymal stem cell (MSC)-mediated DC-SF-DC contraction, as well as growth and differentiation under chondrogenic and osteogenic supplements, were compared to those seeded in DC and on SF alone. The incorporation of SF within DC prevented extensive cell-mediated collagen gel contraction. The effect of the multilayered hybrid on MSC remodelling capacity was also evident at the transcription level, where the expression of matrix metalloproteinases and their inhibitor (MMP1, MMP2, MMP3, MMP13 and Timp1) by MSCs within DC-SF-DC were comparable to those on SF and significantly downregulated in comparison to DC, except for Timp1. Chondrogenic supplements stimulated extracellular matrix production within the construct, stabilizing its multilayered structure and promoting MSC chondrogenic differentiation, as indicated by the upregulation of the genes Col2a1 and Agg and the production of collagen type II. In osteogenic medium there was an upregulation in ALP and OP along with the presence of an apatitic phase, indicating MSC osteoblastic differentiation and matrix mineralization. In sum, these results have implications on the modulation of three-dimensional collagen-based gel structural stability and on the stimulation and maintenance of the MSC committed phenotype inherent to the in vitro formation of chondral tissue and bone, as well as on potential multilayered complex tissues. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  9. TiO2 coating promotes human mesenchymal stem cell proliferation without the loss of their capacity for chondrogenic differentiation

    International Nuclear Information System (INIS)

    Kaitainen, Salla; Lappalainen, Reijo; Mähönen, Anssi J; J Lammi, Mikko; Qu, Chengjuan; Kröger, Heikki

    2013-01-01

    Human mesenchymal stem cells (hMSCs) are used in applications, which may require a large amount of cells; therefore, efficient expansion of the cells is desired. We studied whether TiO 2 coating on plastic cell culture dishes could promote proliferation of hMSCs without adverse effects in chondrogenic differentiation. TiO 2 -films were deposited on polystyrene dishes and glass coverslips using an ultrashort pulsed laser deposition technique. Human MSCs from three donors were expanded on them until 95% confluence, and the cells were evaluated by morphology, immunocytochemistry and quantitative RT-PCR (qRT-PCR). The chondrogenic differentiation in pellets was performed after cultivation on TiO 2 -coated dishes. Chondrogenesis was evaluated by histological staining of proteoglycans and type II collagen, and qRT-PCR. Human MSC-associated markers STRO-1, CD44, CD90 and CD146 did not change after expansion on TiO 2 -coated coverslips. However, the cell number after a 48h-culture period was significantly higher on TiO 2 -coated culture dishes. Importantly, TiO 2 coating caused no significant differences in the proteoglycan and type II collagen staining of the pellets, or the expression of chondrocyte-specific genes in the chondrogenesis assay. Thus, the proliferation of hMSCs could be significantly increased when cultured on TiO 2 -coated dishes without weakening their chondrogenic differentiation capacity. The transparency of TiO 2 -films allows easy monitoring of the cell growth and morphology under a phase-contrast microscope. (paper)

  10. Platelet-Rich Plasma Preparation Types Show Impact on Chondrogenic Differentiation, Migration, and Proliferation of Human Subchondral Mesenchymal Progenitor Cells.

    Science.gov (United States)

    Kreuz, Peter Cornelius; Krüger, Jan Philipp; Metzlaff, Sebastian; Freymann, Undine; Endres, Michaela; Pruss, Axel; Petersen, Wolf; Kaps, Christian

    2015-10-01

    To evaluate the chondrogenic potential of platelet concentrates on human subchondral mesenchymal progenitor cells (MPCs) as assessed by histomorphometric analysis of proteoglycans and type II collagen. Furthermore, the migratory and proliferative effect of platelet concentrates were assessed. Platelet-rich plasma (PRP) was prepared using preparation kits (Autologous Conditioned Plasma [ACP] Kit [Arthrex, Naples, FL]; Regen ACR-C Kit [Regen Lab, Le Mont-Sur-Lausanne, Switzerland]; and Dr.PRP Kit [Rmedica, Seoul, Republic of Korea]) by apheresis (PRP-A) and by centrifugation (PRP-C). In contrast to clinical application, freeze-and-thaw cycles were subsequently performed to activate platelets and to prevent medium coagulation by residual fibrinogen in vitro. MPCs were harvested from the cortico-spongious bone of femoral heads. Chondrogenic differentiation of MPCs was induced in high-density pellet cultures and evaluated by histochemical staining of typical cartilage matrix components. Migration of MPCs was assessed using a chemotaxis assay, and proliferation activity was measured by DNA content. MPCs cultured in the presence of 5% ACP, Regen, or Dr.PRP formed fibrous tissue, whereas MPCs stimulated with 5% PRP-A or PRP-C developed compact and dense cartilaginous tissue rich in type II collagen and proteoglycans. All platelet concentrates significantly (ACP, P = .00041; Regen, P = .00029; Dr.PRP, P = .00051; PRP-A, P platelet concentrates but one (Dr.PRP, P = .63) showed a proliferative effect on MPCs, as shown by significant increases (ACP, P = .027; Regen, P = .0029; PRP-A, P = .00021; and PRP-C, P = .00069) in DNA content. Platelet concentrates obtained by different preparation methods exhibit different potentials to stimulate chondrogenic differentiation, migration, and proliferation of MPCs. Platelet concentrates obtained by commercially available preparation kits failed to induce chondrogenic differentiation of MPCs, whereas highly standardized PRP

  11. Curcumin inhibits cellular condensation and alters microfilament organization during chondrogenic differentiation of limb bud mesenchymal cells

    OpenAIRE

    Kim, Dongkyun; Kim, Song-Ja; Kang, Shin-Sung; Jin, Eun-Jung

    2009-01-01

    Curcumin is a well known natural polyphenol product isolated from the rhizome of the plant Curcuma longa, anti-inflammatory agent for arthritis by inhibiting synthesis of inflammatory prostaglandins. However, the mechanisms by which curcumin regulates the functions of chondroprogenitor, such as proliferation, precartilage condensation, cytoskeletal organization or overall chondrogenic behavior, are largely unknown. In the present report, we investigated the effects and signaling mechanism of ...

  12. Effects of Serial Passage on the Characteristics and Chondrogenic Differentiation of Canine Umbilical Cord Matrix Derived Mesenchymal Stem Cells

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    K. S. Lee

    2013-04-01

    Full Text Available Mesenchymal stem cells (MSCs are often known to have a therapeutic potential in the cell-mediated repair for fatal or incurable diseases. In this study, canine umbilical cord MSCs (cUC-MSCs were isolated from umbilical cord matrix (n = 3 and subjected to proliferative culture for 5 consecutive passages. The cells at each passage were characterized for multipotent MSC properties such as proliferation kinetics, expression patterns of MSC surface markers and self-renewal associated markers, and chondrogenic differentiation. In results, the proliferation of the cells as determined by the cumulative population doubling level was observed at its peak on passage 3 and stopped after passage 5, whereas cell doubling time dramatically increased after passage 4. Expression of MSC surface markers (CD44, CD54, CD61, CD80, CD90 and Flk-1, molecule (HMGA2 and pluripotent markers (sox2, nanog associated with self-renewal was negatively correlated with the number of passages. However, MSC surface marker (CD105 and pluripotent marker (Oct3/4 decreased with increasing the number of subpassage. cUC-MSCs at passage 1 to 5 underwent chondrogenesis under specific culture conditions, but percentage of chondrogenic differentiation decreased with increasing the number of subpassage. Collectively, the present study suggested that sequential subpassage could affect multipotent properties of cUC-MSCs and needs to be addressed before clinical applications.

  13. IGF-1 Gene Transfer to Human Synovial MSCs Promotes Their Chondrogenic Differentiation Potential without Induction of the Hypertrophic Phenotype

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    Yasutoshi Ikeda

    2017-01-01

    Full Text Available Mesenchymal stem cell- (MSC- based therapy is a promising treatment for cartilage. However, repair tissue in general fails to regenerate an original hyaline-like tissue. In this study, we focused on increasing the expression levels for insulin-like growth factor-1 (IGF-1 to improve repair tissue quality. The IGF-1 gene was introduced into human synovial MSCs with a lentiviral vector and examined the levels of gene expression and morphological status of MSCs under chondrogenic differentiation condition using pellet cultures. The size of the pellets derived from IGF-1-MSCs were significantly larger than those of the control group. The abundance of glycosaminoglycan (GAG was also significantly higher in the IGF-1-MSC group. The histology of the IGF-1-induced pellets demonstrated similarities to hyaline cartilage without exhibiting features of a hypertrophic chondrocyte phenotype. Expression levels for the Col2A1 gene and protein were significantly higher in the IGF-1 pellets than in the control pellets, but expression levels for Col10, MMP-13, ALP, and Osterix were not higher. Thus, IGF-1 gene transfer to human synovial MSCs led to an improved chondrogenic differentiation capacity without the detectable induction of a hypertrophic or osteogenic phenotype.

  14. Directing chondrogenic differentiation of mesenchymal stem cells with a solid-supported chitosan thermogel for cartilage tissue engineering

    International Nuclear Information System (INIS)

    Huang, Hongjie; Zhang, Xin; Hu, Xiaoqing; Dai, Linghui; Zhu, Jingxian; Man, Zhentao; Ao, Yingfang; Chen, Haifeng; Zhou, Chunyan

    2014-01-01

    Hydrogels are attractive for cartilage tissue engineering because of their high plasticity and similarity with the native cartilage matrix. However, one critical drawback of hydrogels for osteochondral repair is their inadequate mechanical strength. To address this limitation, we constructed a solid-supported thermogel comprising a chitosan hydrogel system and demineralized bone matrix. Scanning electron microscopy, the equilibrium scanning ratio, the biodegradation rate, biomechanical tests, biochemical assays, metabolic activity tests, immunostaining and cartilage-specific gene expression analysis were used to evaluate the solid-supported thermogel. Compared with pure hydrogel or demineralized matrix, the hybrid biomaterial showed superior porosity, equilibrium swelling and degradation rate. The hybrid scaffolds exhibited an increased mechanical strength: 75% and 30% higher compared with pure hydrogels and demineralized matrix, respectively. After three days culture, bone-derived mesenchymal stem cells (BMSCs) maintained viability above 90% in all three materials; however, the cell retention of the hybrid scaffolds was more efficient and uniform than the other materials. Matrix production and chondrogenic differentiation of BMSCs in the hybrid scaffolds were superior to its precursors, based on glycosaminoglycan quantification and hyaline cartilage marker expression after three weeks in culture. Its easy preparation, favourable biophysical properties and chondrogenic capacity indicated that this solid-supported thermogel could be an attractive biomaterial framework for cartilage tissue engineering. (paper)

  15. Differential expression of CCN-family members in primary human bone marrow-derived mesenchymal stem cells during osteogenic, chondrogenic and adipogenic differentiation

    Directory of Open Access Journals (Sweden)

    Hendrich Christian

    2005-03-01

    Full Text Available Abstract Background The human cysteine rich protein 61 (CYR61, CCN1 as well as the other members of the CCN family of genes play important roles in cellular processes such as proliferation, adhesion, migration and survival. These cellular events are of special importance within the complex cellular interactions ongoing in bone remodeling. Previously, we analyzed the role of CYR61/CCN1 as an extracellular signaling molecule in human osteoblasts. Since mesenchymal stem cells of bone marrow are important progenitors for various differentiation pathways in bone and possess increasing potential for regenerative medicine, here we aimed to analyze the expression of CCN family members in bone marrow-derived human mesenchymal stem cells and along the osteogenic, the adipogenic and the chondrogenic differentiation. Results Primary cultures of human mesenchymal stem cells were obtained from the femoral head of patients undergoing total hip arthroplasty. Differentiation into adipocytes and osteoblasts was done in monolayer culture, differentiation into chondrocytes was induced in high density cell pellet cultures. For either pathway, established differentiation markers and CCN-members were analyzed at the mRNA level by RT-PCR and the CYR61/CCN1 protein was analyzed by immunocytochemistry. RT-PCR and histochemical analysis revealed the appropriate phenotype of differentiated cells (Alizarin-red S, Oil Red O, Alcian blue, alkaline phosphatase; osteocalcin, collagen types I, II, IX, X, cbfa1, PPARγ, aggrecan. Mesenchymal stem cells expressed CYR61/CCN1, CTGF/CCN2, CTGF-L/WISP2/CCN5 and WISP3/CCN6. The CYR61/CCN1 expression decreased markedly during osteogenic differentiation, adipogenic differentiation and chondrogenic differentiation. These results were confirmed by immuncytochemical analyses. WISP2/CCN5 RNA expression declined during adipogenic differentiation and WISP3/CCN6 RNA expression was markedly reduced in chondrogenic differentiation. Conclusion The

  16. The effects of the WNT-signaling modulators BIO and PKF118-310 on the chondrogenic differentiation of human mesenchymal stem cells

    NARCIS (Netherlands)

    Huang, Xiaobin; Zhong, Leilei; Hendriks, Jan; Post, Janine N.; Karperien, Marcel

    2018-01-01

    Mesenchymal stem cells (MSCs) are multipotent cells, mainly from bone marrow, and an ideal source of cells in bone and cartilage tissue engineering. A study of the chondrogenic differentiation of MSCs is of particular interest for MSCs-based cartilage regeneration. In this study, we aimed to

  17. Molecular Validation of Chondrogenic Differentiation and Hypoxia Responsiveness of Platelet-Lysate Expanded Adipose Tissue–Derived Human Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Galeano-Garces, Catalina; Camilleri, Emily T.; Riester, Scott M.; Dudakovic, Amel; Larson, Dirk R.; Qu, Wenchun; Smith, Jay; Dietz, Allan B.; Im, Hee-Jeong; Krych, Aaron J.; Larson, A. Noelle; Karperien, Marcel; van Wijnen, Andre J.

    2017-01-01

    Objective: To determine the optimal environmental conditions for chondrogenic differentiation of human adipose tissue–derived mesenchymal stromal/stem cells (AMSCs). In this investigation we specifically investigate the role of oxygen tension and 3-dimensional (3D) culture systems. Design: Both

  18. Implication of C-type natriuretic peptide-3 signaling in glycosaminoglycan synthesis and chondrocyte hypertrophy during TGF-β1 induced chondrogenic differentiation of chicken bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Kocamaz, Erdogan; Gok, Duygu; Cetinkaya, Ayse; Tufan, A Cevik

    2012-10-01

    This study investigated the involvement of CNP-3, chick homologue for human C-type natriuretic peptide (CNP), in TGF-β1 induced chondrogenic differentiation of chicken bone marrow-derived mesenchymal stem cells (MSCs). Chondrogenic differentiation of MSCs in pellet cultures was induced by TGF-β1. Chondrogenic differentiation and glycosaminoglycan synthesis were analyzed on the basis of basic histology, collagen type II expression, and Alcian blue staining. Antibodies against CNP and NPR-B were used to block their function during these processes. Results revealed that expression of CNP-3 and NPR-B in MSCs were regulated by TGF-β1 in monolayer cultures at mRNA level. In pellet cultures of MSCs, TGF-β1 successfully induced chondrogenic differentiation and glycosaminoglycan synthesis. Addition of CNP into the TGF-β1 supplemented chondrogenic differentiation medium further induced the glycosaminoglycan synthesis and hypertrophy of differentiated chondrocytes in these pellets. Pellets induced with TGF-β1 and treated with antibodies against CNP and NPR-B, did show collagen type II expression, however, Alcian blue staining showing glycosaminoglycan synthesis was significantly suppressed. In conclusion, CNP-3/NPR-B signaling may strongly be involved in synthesis of glycosaminoglycans of the chondrogenic matrix and hypertrophy of differentiated chondrocytes during TGF-β1 induced chondrogenic differentiation of MSCs.

  19. Transfection of the IHH gene into rabbit BMSCs in a simulated microgravity environment promotes chondrogenic differentiation and inhibits cartilage aging.

    Science.gov (United States)

    Liu, Peng-Cheng; Liu, Kuan; Liu, Jun-Feng; Xia, Kuo; Chen, Li-Yang; Wu, Xing

    2016-09-27

    The effect of overexpressing the Indian hedgehog (IHH) gene on the chondrogenic differentiation of rabbit bone marrow-derived mesenchymal stem cells (BMSCs) was investigated in a simulated microgravity environment. An adenovirus plasmid encoding the rabbit IHH gene was constructed in vitro and transfected into rabbit BMSCs. Two large groups were used: conventional cell culture and induction model group and simulated microgravity environment group. Each large group was further divided into blank control group, GFP transfection group, and IHH transfection group. During differentiation induction, the expression levels of cartilage-related and cartilage hypertrophy-related genes and proteins in each group were determined. In the conventional model, the IHH transfection group expressed high levels of cartilage-related factors (Coll2 and ANCN) at the early stage of differentiation induction and expressed high levels of cartilage hypertrophy-related factors (Coll10, annexin 5, and ALP) at the late stage. Under the simulated microgravity environment, the IHH transfection group expressed high levels of cartilage-related factors and low levels of cartilage hypertrophy-related factors at all stages of differentiation induction. Under the simulated microgravity environment, transfection of the IHH gene into BMSCs effectively promoted the generation of cartilage and inhibited cartilage aging and osteogenesis. Therefore, this technique is suitable for cartilage tissue engineering.

  20. Potential Biomedical Application of Enzymatically Treated Alginate/Chitosan Hydrosols in Sponges—Biocompatible Scaffolds Inducing Chondrogenic Differentiation of Human Adipose Derived Multipotent Stromal Cells

    Directory of Open Access Journals (Sweden)

    Anna Zimoch-Korzycka

    2016-08-01

    Full Text Available Current regenerative strategies used for cartilage repair rely on biomaterial functionality as a scaffold for cells that may have potential in chondrogenic differentiation. The purpose of the research was to investigate the biocompatibility of enzymatically treated alginate/chitosan hydrosol sponges and their suitability to support chondrogenic differentiation of human adipose derived multipotent stromal cells (hASCs. The alginate/chitosan and enzyme/alginate/chitosan sponges were formed from hydrosols with various proportions and were used as a biomaterial in this study. Sponges were tested for porosity and wettability. The porosity of each sponge was higher than 80%. An equal dose of alginate and chitosan in the composition of sponges improved their swelling ability. It was found that equal concentrations of alginate and chitosan in hydrosols sponges assure high biocompatibility properties that may be further improved by enzymatic treatment. Importantly, the high biocompatibility of these biomaterials turned out to be crucial in the context of hydrosols’ pro-chondrogenic function. After exposure to the chondrogenic conditions, the hASCs in N/A/C and L/A/C sponges formed well developed nodules and revealed increased expression of collagen type II, aggrecan and decreased expression of collagen type I. Moreover, in these cultures, the reactive oxygen species level was lowered while superoxide dismutase activity increased. Based on the obtained results, we conclude that N/A/C and L/A/C sponges may have prospective application as hASCs carriers for cartilage repair.

  1. Macroautophagy and Selective Mitophagy Ameliorate Chondrogenic Differentiation Potential in Adipose Stem Cells of Equine Metabolic Syndrome: New Findings in the Field of Progenitor Cells Differentiation.

    Science.gov (United States)

    Marycz, Krzysztof; Kornicka, Katarzyna; Grzesiak, Jakub; Śmieszek, Agnieszka; Szłapka, Jolanta

    2016-01-01

    Equine metabolic syndrome (EMS) is mainly characterized by insulin resistance, obesity, and local or systemic inflammation. That unfriendly environment of adipose tissue has huge impact on stem cells population (ASC) residing within. In the present study, using molecular biology techniques and multiple imaging techniques (SEM, FIB-SEM, and confocal microscopy), we evaluated the impact of EMS on ASC viability and chondrogenic differentiation. Moreover, we visualized the mitochondrial network and dynamics in ASC CTRL and ASC EMS during control and chondrogenic conditions. In control conditions, ASC EMS were characterized by increased mitochondrial fission in comparison to ASC CTRL . We found that extensive remodeling of mitochondrial network including fusion and fission occurs during early step of differentiation. Moreover, we observed mitochondria morphology deterioration in ASC EMS . These conditions seem to cause autophagic shift in ASC EMS , as we observed increased accumulation of LAMP2 and formation of multiple autophagosomes in those cells, some of which contained dysfunctional mitochondria. "Autophagic" switch may be a rescue mechanism allowing ASC EMS to clear impaired by ROS proteins and mitochondria. Moreover it provides a precursors-to-macromolecules synthesis, especially during chondrogenesis. Our data indicates that autophagy in ASC EMS would be crucial for the quality control mechanisms and maintenance of cellular homeostasis ASC EMS allowing them to be in "stemness" status.

  2. Macroautophagy and Selective Mitophagy Ameliorate Chondrogenic Differentiation Potential in Adipose Stem Cells of Equine Metabolic Syndrome: New Findings in the Field of Progenitor Cells Differentiation

    Directory of Open Access Journals (Sweden)

    Krzysztof Marycz

    2016-01-01

    Full Text Available Equine metabolic syndrome (EMS is mainly characterized by insulin resistance, obesity, and local or systemic inflammation. That unfriendly environment of adipose tissue has huge impact on stem cells population (ASC residing within. In the present study, using molecular biology techniques and multiple imaging techniques (SEM, FIB-SEM, and confocal microscopy, we evaluated the impact of EMS on ASC viability and chondrogenic differentiation. Moreover, we visualized the mitochondrial network and dynamics in ASCCTRL and ASCEMS during control and chondrogenic conditions. In control conditions, ASCEMS were characterized by increased mitochondrial fission in comparison to ASCCTRL. We found that extensive remodeling of mitochondrial network including fusion and fission occurs during early step of differentiation. Moreover, we observed mitochondria morphology deterioration in ASCEMS. These conditions seem to cause autophagic shift in ASCEMS, as we observed increased accumulation of LAMP2 and formation of multiple autophagosomes in those cells, some of which contained dysfunctional mitochondria. “Autophagic” switch may be a rescue mechanism allowing ASCEMS to clear impaired by ROS proteins and mitochondria. Moreover it provides a precursors-to-macromolecules synthesis, especially during chondrogenesis. Our data indicates that autophagy in ASCEMS would be crucial for the quality control mechanisms and maintenance of cellular homeostasis ASCEMS allowing them to be in “stemness” status.

  3. Differential effects of BMP-2 and TGF-beta1 on chondrogenic differentiation of adipose derived stem cells

    DEFF Research Database (Denmark)

    Mehlhorn, A T; Niemeyer, P; Kaschte, K

    2007-01-01

    transcriptional regulation of Dlx-5, Msx-2 and Runx-2. MATERIALS AND METHODS: Encapsulated ASC were cultured for 14 days in medium containing TGF-beta1 and/or BMP-2. mRNA expression of the extracellular matrix molecules col2a1, cartilage oligomeric matrix protein, col10a1, alkaline phosphatase (AP......) and transcription factors Msx-2, Dlx-5 and Runx-2 was analysed. Release of glycosaminoglycans, collagen types II and X into the extracellular matrix was demonstrated. RESULTS: BMP-2 and TGF-beta1 induced a chondrogenic phenotype in ASC. Combined growth factor treatment had a synergistic effect on col10a1...

  4. Induction of Chondrogenic Differentiation of Human Mesenchymal Stem Cells by Biomimetic Gold Nanoparticles with Tunable RGD Density.

    Science.gov (United States)

    Li, Jingchao; Li, Xiaomeng; Zhang, Jing; Kawazoe, Naoki; Chen, Guoping

    2017-07-01

    Nanostructured materials have drawn a broad attention for their applications in biomedical fields. Ligand-modified nanomaterials can well mimic the dynamic extracellular matrix (ECM) microenvironments to regulate cell functions and fates. Herein, ECM mimetic gold nanoparticles (Au NPs) with tunable surface arginine-glycine-aspartate (RGD) density are designed and synthesized to induce the chondrogenic differentiation of human mesenchymal stem cells (hMSCs). The biomimetic Au NPs with an average size of 40 nm shows good biocompatibility without affecting the cell proliferation in the studied concentration range. The RGD motifs on Au NPs surface facilitate cellular uptake of NPs into monolayer hMSCs through integrin-mediated endocytosis. The biomimetic NPs have a promotive effect on cartilaginous matrix production and marker gene expression in cell pellet culture, especially for the biomimetic Au NPs with high surface RGD density. This study provides a novel strategy for fabricating biomimetic NPs to regulate cell differentiation, which holds great potentials in tissue engineering and biomedical applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Delta-like 1/Fetal Antigen-1 (Dlk1/FA1) Is a Novel Regulator of Chondrogenic Cell Differentiation via Inhibition of the Akt Kinase-dependent Pathway*

    Science.gov (United States)

    Chen, Li; Qanie, Diyako; Jafari, Abbas; Taipaleenmaki, Hanna; Jensen, Charlotte H.; Säämänen, Anna-Marja; Sanz, Maria Luisa Nueda; Laborda, Jorge; Abdallah, Basem M.; Kassem, Moustapha

    2011-01-01

    Delta-like 1 (Dlk1, also known as fetal antigen-1, FA1) is a member of Notch/Delta family that inhibits adipocyte and osteoblast differentiation; however, its role in chondrogenesis is still not clear. Thus, we overexpressed Dlk1/FA1 in mouse embryonic ATDC5 cells and tested its effects on chondrogenic differentiation. Dlk1/FA1 inhibited insulin-induced chondrogenic differentiation as evidenced by reduction of cartilage nodule formation and gene expression of aggrecan, collagen Type II and X. Similar effects were obtained either by using Dlk1/FA1-conditioned medium or by addition of a purified, secreted, form of Dlk1 (FA1) directly to the induction medium. The inhibitory effects of Dlk1/FA1 were dose-dependent and occurred irrespective of the chondrogenic differentiation stage: proliferation, differentiation, maturation, or hypertrophic conversion. Overexpression or addition of the Dlk1/FA1 protein to the medium strongly inhibited the activation of Akt, but not the ERK1/2, or p38 MAPK pathways, and the inhibition of Akt by Dlk1/FA1 was mediated through PI3K activation. Interestingly, inhibition of fibronectin expression by siRNA rescued the Dlk1/FA1-mediated inhibition of Akt, suggesting interaction of Dlk1/FA1 and fibronectin in chondrogenic cells. Our results identify Dlk1/FA1 as a novel regulator of chondrogenesis and suggest Dlk1/FA1 acts as an inhibitor of the PI3K/Akt pathways that leads to its inhibitory effects on chondrogenesis. PMID:21724852

  6. Use of the second harmonic generation microscopy to evaluate chondrogenic differentiation of mesenchymal stem cells for cartilage repair

    Science.gov (United States)

    Bordeaux-Rego, P.; Baratti, M. O.; Duarte, A. S. S.; Ribeiro, T. B.; Andreoli-Risso, M. F.; Vidal, B.; Miranda, J. B.; Adur, J.; de Thomaz, A. A.; Pelegati, V. B.; Costa, F. F.; Carvalho, H. F.; Cesar, C. L.; Luzo, A.; Olalla Saad, S. T.

    2012-03-01

    Articular cartilage injury remains one of the major concerns in orthopedic surgery. Mesenchymal stem cell (MSC) transplantation has been introduced to avoid some of the side effects and complications of current techniques.. With the aim to evaluate chondrogenic differentiation of mesenchymal stem cells, we used Second Harmonic Generation (SHG) microscopy to analyze the aggregation and orientation of collagen fibrils in the hyaline cartilage of rabbit knees. The experiment was performed using implants with type II collagen hydrogel (a biomaterial that mimics the microenvironment of the cartilage), one implant containing MSC and one other without MSC (control). After 10 weeks, the rabbit knees were dissected and fibril collagen distribution and spatial organization in the extracellular matrix of the lesions were verified by SHG. The result showed significant differences, whereas in histological sections of the cartilaginous lesions with MSC the collagen fibers are organized and regular; in the control sections the collagen fibers are more irregular, with absence of cells. A macroscopic analysis of the lesions confirmed this difference, showing a greater percentage of lesions filling in knees treated with MSC than in the knees used as controls. This study demonstrates that SHG microscopy will be an excellent tool to help in the evaluation of the effectiveness of MSC-based cell therapy for cartilage repair.

  7. Proliferation and chondrogenic differentiation of CD105-positive enriched rat synovium-derived mesenchymal stem cells in three-dimensional porous scaffolds

    International Nuclear Information System (INIS)

    Qi Jun; Chen Anmin; You Hongbo; Li Kunpeng; Zhang Di; Guo Fengjing

    2011-01-01

    Stem cell-based tissue engineering has provided an alternative strategy to treat cartilage lesions, and synovium-derived mesenchymal stem cells (SMSCs) are considered as a promising cell source for cartilage repair. In this study, the SMSCs were isolated from rat synovium, and CD105-positive (CD105 + ) cells were enriched using magnetic activated cell sorting. Sorted cells were subsequently seeded onto the chitosan-alginate composite three-dimensional (3D) porous scaffolds and cultured in chondrogenic culture medium in the presence of TGF-β 3 and BMP-2 for 2 weeks in vitro. After 2 weeks in culture, scanning electron microscopy results showed that cells attached and proliferated well on scaffolds, and secreted extracellular matrix were also observed. From day 7 to day 14, the total DNA and glucosaminoglycan content of the cells cultured in scaffolds were found to have increased significantly, and cell cycle analyses revealed that the percentage of cells in the S and G2/M phases increased and the percentage of cells in the G0/G1 phase decreased. Compared with non-sorted cells, the sorted cells cultured in scaffolds underwent more chondrogenic differentiation, as evidenced by higher expression of type II collagen and Sox9 at the protein and mRNA levels. The results suggest that CD105 + enriched SMSCs may be a potential cell source for cartilage tissue engineering, and the chitosan-alginate composite 3D porous scaffold could provide a favorable microenvironment for supporting proliferation and chondrogenic differentiation of cells.

  8. Scaffold composition affects cytoskeleton organization, cell-matrix interaction and the cellular fate of human mesenchymal stem cells upon chondrogenic differentiation.

    Science.gov (United States)

    Li, Yuk Yin; Choy, Tze Hang; Ho, Fu Chak; Chan, Pui Barbara

    2015-06-01

    The stem cell niche, or microenvironment, consists of soluble, matrix, cell and mechanical factors that together determine the cellular fates and/or differentiation patterns of stem cells. Collagen and glycosaminoglycans (GAGs) are important scaffolding materials that can mimic the natural matrix niche. Here, we hypothesize that imposing changes in the scaffold composition or, more specifically, incorporating GAGs into the collagen meshwork, will affect the morphology, cytoskeletal organization and integrin expression profiles, and hence the fate of human mesenchymal stem cells (MSCs) upon the induction of differentiation. Using chondrogenesis as an example, we microencapsulated MSCs in three scaffold systems that had varying matrix compositions: collagen alone (C), aminated collagen (AC) and aminated collagen with GAGs (ACG). We then induced the MSCs to differentiate toward a chondrogenic lineage, after which, we characterized the cell viability and morphology, as well as the level of cytoskeletal organization and the integrin expression profile. We also studied the fate of the MSCs by evaluating the major chondrogenic markers at both the gene and protein level. In C, MSC chondrogenesis was successfully induced and MSCs that spread in the scaffolds had a clear actin cytoskeleton; they expressed integrin α2β1, α5 and αv; promoted sox9 nuclear localization transcription activation; and upregulated the expression of chondrogenic matrix markers. In AC, MSC chondrogenesis was completely inhibited but the scaffold still supported cell survival. The MSCs did not spread and they had no actin cytoskeleton; did not express integrin α2 or αv; they failed to differentiate into chondrogenic lineage cells even on chemical induction; and there was little colocalization or functional interaction between integrin α5 and fibronectin. In ACG, although the MSCs did not express integrin α2, they did express integrin αv and there was strong co-localization and hence functional

  9. Alternative splicing targeting the hTAF4-TAFH domain of TAF4 represses proliferation and accelerates chondrogenic differentiation of human mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Jekaterina Kazantseva

    Full Text Available Transcription factor IID (TFIID activity can be regulated by cellular signals to specifically alter transcription of particular subsets of genes. Alternative splicing of TFIID subunits is often the result of external stimulation of upstream signaling pathways. We studied tissue distribution and cellular expression of different splice variants of TFIID subunit TAF4 mRNA and biochemical properties of its isoforms in human mesenchymal stem cells (hMSCs to reveal the role of different isoforms of TAF4 in the regulation of proliferation and differentiation. Expression of TAF4 transcripts with exons VI or VII deleted, which results in a structurally modified hTAF4-TAFH domain, increases during early differentiation of hMSCs into osteoblasts, adipocytes and chondrocytes. Functional analysis data reveals that TAF4 isoforms with the deleted hTAF4-TAFH domain repress proliferation of hMSCs and preferentially promote chondrogenic differentiation at the expense of other developmental pathways. This study also provides initial data showing possible cross-talks between TAF4 and TP53 activity and switching between canonical and non-canonical WNT signaling in the processes of proliferation and differentiation of hMSCs. We propose that TAF4 isoforms generated by the alternative splicing participate in the conversion of the cellular transcriptional programs from the maintenance of stem cell state to differentiation, particularly differentiation along the chondrogenic pathway.

  10. The Effects of the WNT-Signaling Modulators BIO and PKF118-310 on the Chondrogenic Differentiation of Human Mesenchymal Stem Cells

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    Xiaobin Huang

    2018-02-01

    Full Text Available Mesenchymal stem cells (MSCs are multipotent cells, mainly from bone marrow, and an ideal source of cells in bone and cartilage tissue engineering. A study of the chondrogenic differentiation of MSCs is of particular interest for MSCs-based cartilage regeneration. In this study, we aimed to optimize the conditions for the chrondogenic differentiation of MSCs by regulating WNT signaling using the small molecule WNT inhibitor PKF118-310 and activator BIO. Human mesenchymal stem cells (hMSCs were isolated from bone marrow aspirates and cultured in hMSCs proliferation medium. Pellet culture was subsequently established for three-dimensional chondrogenic differentiation of 5 weeks. WNT signaling was increased by the small molecule glycogen synthase kinase-3 inhibitor 6-bromoindirubin-3-oxim (BIO and decreased by the WNT inhibitor PKF118-310 (PKF. The effects of BIO and PKF on the chondrogenesis of hMSCs was examined by real-time PCR, histological methods, and ELISA. We found that activation of canonical WNT-signaling by BIO significantly downregulated the expression of cartilage-specific genes SOX9, COL2A1, and ACAN, and matrix metalloproteinase genes MMP1/3/9/13, but increased ADAMTS 4/5. Inhibition of WNT signaling by PKF increased the expression of SOX9, COL2A1, ACAN, and MMP9, but decreased MMP13 and ADAMTS4/5. In addition, a high level of WNT signaling induced the expression of hypertrophic markers COL10A1, ALPL, and RUNX2, the dedifferentiation marker COL1A1, and glycolysis genes GULT1 and PGK1. Deposition of glycosaminoglycan (GAG and collagen type II in the pellet matrix was significantly lost in the BIO-treated group and increased in the PKF-treated group. The protein level of COL10A1 was also highly induced in the BIO group. Interestingly, BIO decreased the number of apoptotic cells while PKF significantly induced apoptosis during chondrogenesis. The natural WNT antagonist DKK1 and the protein level of MMP1 in the pellet culture medium were

  11. The Effects of the WNT-Signaling Modulators BIO and PKF118-310 on the Chondrogenic Differentiation of Human Mesenchymal Stem Cells.

    Science.gov (United States)

    Huang, Xiaobin; Zhong, Leilei; Hendriks, Jan; Post, Janine N; Karperien, Marcel

    2018-02-13

    Mesenchymal stem cells (MSCs) are multipotent cells, mainly from bone marrow, and an ideal source of cells in bone and cartilage tissue engineering. A study of the chondrogenic differentiation of MSCs is of particular interest for MSCs-based cartilage regeneration. In this study, we aimed to optimize the conditions for the chrondogenic differentiation of MSCs by regulating WNT signaling using the small molecule WNT inhibitor PKF118-310 and activator BIO. Human mesenchymal stem cells (hMSCs) were isolated from bone marrow aspirates and cultured in hMSCs proliferation medium. Pellet culture was subsequently established for three-dimensional chondrogenic differentiation of 5 weeks. WNT signaling was increased by the small molecule glycogen synthase kinase-3 inhibitor 6-bromoindirubin-3-oxim (BIO) and decreased by the WNT inhibitor PKF118-310 (PKF). The effects of BIO and PKF on the chondrogenesis of hMSCs was examined by real-time PCR, histological methods, and ELISA. We found that activation of canonical WNT-signaling by BIO significantly downregulated the expression of cartilage-specific genes SOX9 , COL2A1, and ACAN , and matrix metalloproteinase genes MMP1/3/9/13, but increased ADAMTS 4/5 . Inhibition of WNT signaling by PKF increased the expression of SOX9 , COL2A1 , ACAN , and MMP9, but decreased MMP13 and ADAMTS4/5 . In addition, a high level of WNT signaling induced the expression of hypertrophic markers COL10A1, ALPL , and RUNX2, the dedifferentiation marker COL1A1 , and glycolysis genes GULT1 and PGK1 . Deposition of glycosaminoglycan (GAG) and collagen type II in the pellet matrix was significantly lost in the BIO-treated group and increased in the PKF-treated group. The protein level of COL10A1 was also highly induced in the BIO group. Interestingly, BIO decreased the number of apoptotic cells while PKF significantly induced apoptosis during chondrogenesis. The natural WNT antagonist DKK1 and the protein level of MMP1 in the pellet culture medium were

  12. New Insights into Osteogenic and Chondrogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells and Their Potential Clinical Applications for Bone Regeneration in Pediatric Orthopaedics

    Directory of Open Access Journals (Sweden)

    Nicola Giuliani

    2013-01-01

    Full Text Available Human mesenchymal stem cells (hMSCs are pluripotent adult stem cells capable of being differentiated into osteoblasts, adipocytes, and chondrocytes. The osteogenic differentiation of hMSCs is regulated either by systemic hormones or by local growth factors able to induce specific intracellular signal pathways that modify the expression and activity of several transcription factors. Runt-related transcription factor 2 (Runx2 and Wnt signaling-related molecules are the major factors critically involved in the osteogenic differentiation process by hMSCs, and SRY-related high-mobility-group (HMG box transcription factor 9 (SOX9 is involved in the chondrogenic one. hMSCs have generated a great interest in the field of regenerative medicine, particularly in bone regeneration. In this paper, we focused our attention on the molecular mechanisms involved in osteogenic and chondrogenic differentiation of hMSC, and the potential clinical use of hMSCs in osteoarticular pediatric disease characterized by fracture nonunion and pseudarthrosis.

  13. Different culture media affect growth characteristics, surface marker distribution and chondrogenic differentiation of human bone marrow-derived mesenchymal stromal cells.

    Science.gov (United States)

    Hagmann, Sebastien; Moradi, Babak; Frank, Sebastian; Dreher, Thomas; Kämmerer, Peer Wolfgang; Richter, Wiltrud; Gotterbarm, Tobias

    2013-07-30

    Bone marrow-derived mesenchymal stromal cells (BM-MSCs) play an important role in modern tissue engineering, while distinct variations of culture media compositions and supplements have been reported. Because MSCs are heterogeneous regarding their regenerative potential and their surface markers, these parameters were compared in four widely used culture media compositions. MSCs were isolated from bone marrow and expanded in four established cell culture media. MSC yield/1000 MNCs, passage time and growth index were observed. In P4, typical MSC surface markers were analysed by fluorescence cytometry. Additionally, chondrogenic, adipogenic and osteogenic differentiation potential were evaluated. Growth index and P0 cell yield varied importantly between the media. The different expansion media had a significant influence on the expression of CD10, CD90, CD105, CD140b CD146 and STRO-1. While no significant differences were observed regarding osteogenic and adipogenic differentiation, chondrogenic differentiation was superior in medium A as reflected by GAG/DNA content. The choice of expansion medium can have a significant influence on growth, differentiation potential and surface marker expression of mesenchymal stromal cells, which is of fundamental importance for tissue engineering procedures.

  14. -5p and -3p strands of miR-145 and miR-140 during mesenchymal stem cell chondrogenic differentiation.

    Science.gov (United States)

    Kenyon, Jonathan D; Sergeeva, Olga; Somoza, Rodrigo A; Li, Ming; Caplan, Arnold I; Khalil, Ahmad M; Lee, Zhenghong

    2018-04-20

    The chondrogenic differentiation of mesenchymal stem cells (MSCs) is mediated by transcription factors and small non-coding RNAs such as micro-RNAs (miRNAs). Each miRNA is initially transcribed as a long transcript, which matures to produce -5p and -3p strands. It is widely believed that the mature and functional miRNA from any given pre-miRNA, usually the -5p strand, is functional, while the opposing -3p strand is degraded. However, recent cartilage literature started to show functional -3p stands for a few miRNAs. This study aimed at examining both -5p and -3p strands of two key miRNAs miR-140 and miR-145 that are known to be involved in the chondrogenic differentiation of MSCs. The level (copy number) of both -5p and -3p strands of miR-145 and miR-140 along the timeline of MSC chondrogenic differentiation was determined by PCR. The gene expression profiles of several genes related to MSC chondrogenesis were compared with these miRNA profiles along the same timeline. While miR-145-3p is declining in step with miR-145-5p in pellet cultures during the process, the -3p strand is only 1% - 2% of the total miR-145 products. In contrast, the mature -3p and -5p products of miR-140 are found to increase with near equal molar expression throughout chondrogenic differentiation. Numerous genes are expressed by cartilage progenitor cells during development. One such target gene, Sox9 is a regulatory target of the dominant miR-145-5p, consistent with the data. Further experimental validations are warranted to confirm that ACAN, FOXO1 and RUNX3 as direct targets of miR-145-5p in the context of MSC chondrogenesis. Similarly, TRSP1 and ACAN are worth further validation as direct targets of miR-145-3p. For miR-140, SOX4 shall be further validated as a direct target of miR-140-5p while KLF4, PTHLH, and WNT5A can be validated as direct targets of miR-140-3p.

  15. Proliferation and chondrogenic differentiation of CD105-positive enriched rat synovium-derived mesenchymal stem cells in three-dimensional porous scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Qi Jun; Chen Anmin; You Hongbo; Li Kunpeng; Zhang Di; Guo Fengjing, E-mail: fjguo@tjh.tjmu.edu.cn [Department of Orthopedics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030 (China)

    2011-02-15

    Stem cell-based tissue engineering has provided an alternative strategy to treat cartilage lesions, and synovium-derived mesenchymal stem cells (SMSCs) are considered as a promising cell source for cartilage repair. In this study, the SMSCs were isolated from rat synovium, and CD105-positive (CD105{sup +}) cells were enriched using magnetic activated cell sorting. Sorted cells were subsequently seeded onto the chitosan-alginate composite three-dimensional (3D) porous scaffolds and cultured in chondrogenic culture medium in the presence of TGF-{beta}{sub 3} and BMP-2 for 2 weeks in vitro. After 2 weeks in culture, scanning electron microscopy results showed that cells attached and proliferated well on scaffolds, and secreted extracellular matrix were also observed. From day 7 to day 14, the total DNA and glucosaminoglycan content of the cells cultured in scaffolds were found to have increased significantly, and cell cycle analyses revealed that the percentage of cells in the S and G2/M phases increased and the percentage of cells in the G0/G1 phase decreased. Compared with non-sorted cells, the sorted cells cultured in scaffolds underwent more chondrogenic differentiation, as evidenced by higher expression of type II collagen and Sox9 at the protein and mRNA levels. The results suggest that CD105{sup +} enriched SMSCs may be a potential cell source for cartilage tissue engineering, and the chitosan-alginate composite 3D porous scaffold could provide a favorable microenvironment for supporting proliferation and chondrogenic differentiation of cells.

  16. Hypoxia Is a Critical Parameter for Chondrogenic Differentiation of Human Umbilical Cord Blood Mesenchymal Stem Cells in Type I/III Collagen Sponges

    Directory of Open Access Journals (Sweden)

    Tangni Gómez-Leduc

    2017-09-01

    Full Text Available Umbilical cord blood (UCB is an attractive alternative to bone marrow for isolation of mesenchymal stem cells (MSCs to treat articular cartilage defects. Here, we set out to determine the growth factors (bone morphogenetic protein 2 (BMP-2 and transforming growth factor-β (TGF-β1 and oxygen tension effects during chondrogenesis of human UCB-MSCs for cartilage engineering. Chondrogenic differentiation was induced using 3D cultures in type I/III collagen sponges with chondrogenic factors in normoxia (21% O2 or hypoxia (<5% O2 for 7, 14 and 21 days. Our results show that UCB-MSCs can be committed to chondrogenesis in the presence of BMP-2+TGF-β1. Normoxia induced the highest levels of chondrocyte-specific markers. However, hypoxia exerted more benefit by decreasing collagen X and matrix metalloproteinase-13 (MMP13 expression, two chondrocyte hypertrophy markers. However, a better chondrogenesis was obtained by switching oxygen conditions, with seven days in normoxia followed by 14 days in hypoxia, since these conditions avoid hypertrophy of hUCB-MSC-derived chondrocytes while maintaining the expression of chondrocyte-specific markers observed in normoxia. Our study demonstrates that oxygen tension is a key factor for chondrogenesis and suggests that UBC-MSCs 3D-culture should begin in normoxia to obtain a more efficient chondrocyte differentiation before placing them in hypoxia for chondrocyte phenotype stabilization. UCB-MSCs are therefore a reliable source for cartilage engineering.

  17. Chondrogenic properties of collagen type XI, a component of cartilage extracellular matrix.

    Science.gov (United States)

    Li, Ang; Wei, Yiyong; Hung, Clark; Vunjak-Novakovic, Gordana

    2018-08-01

    Cartilage extracellular matrix (ECM) has been used for promoting tissue engineering. However, the exact effects of ECM on chondrogenesis and the acting mechanisms are not well understood. In this study, we investigated the chondrogenic effects of cartilage ECM on human mesenchymal stem cells (MSCs) and identified the contributing molecular components. To this end, a preparation of articular cartilage ECM was supplemented to pellets of chondrogenically differentiating MSCs, pellets of human chondrocytes, and bovine articular cartilage explants to evaluate the effects on cell proliferation and the production of cartilaginous matrix. Selective enzymatic digestion and screening of ECM components were conducted to identify matrix molecules with chondrogenic properties. Cartilage ECM promoted MSC proliferation, production of cartilaginous matrix, and maturity of chondrogenic differentiation, and inhibited the hypertrophic differentiation of MSC-derived chondrocytes. Selective digestion of ECM components revealed a contributory role of collagens in promoting chondrogenesis. The screening of various collagen subtypes revealed strong chondrogenic effect of collagen type XI. Finally, collagen XI was found to promote production and inhibit degradation of cartilage matrix in human articular chondrocyte pellets and bovine articular cartilage explants. Our results indicate that cartilage ECM promotes chondrogenesis and inhibits hypertrophic differentiation in MSCs. Collagen type XI is the ECM component that has the strongest effects on enhancing the production and inhibiting the degradation of cartilage matrix. Copyright © 2018 Elsevier Ltd. All rights reserved.

  18. PDK2 promotes chondrogenic differentiation of mesenchymal stem cells by upregulation of Sox6 and activation of JNK/MAPK/ERK pathway

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    H. Wang

    Full Text Available This study was undertaken to clarify the role and mechanism of pyruvate dehydrogenase kinase isoform 2 (PDK2 in chondrogenic differentiation of mesenchymal stem cells (MSCs. MSCs were isolated from femurs and tibias of Sprague-Dawley rats, weighing 300-400 g (5 females and 5 males. Overexpression and knockdown of PDK2 were transfected into MSCs and then cell viability, adhesion and migration were assessed. Additionally, the roles of aberrant PDK2 in chondrogenesis markers SRY-related high mobility group-box 6 (Sox6, type ΙΙ procollagen gene (COL2A1, cartilage oligomeric matrix protein (COMP, aggrecan (AGC1, type ΙX procollagen gene (COL9A2 and collagen type 1 alpha 1 (COL1A1 were measured by quantitative reverse-transcription polymerase chain reaction (qRT-PCR. The expressions of c-Jun N-terminal kinase (JNK, p38 mitogen-activated protein kinase (MAPK and extracellular regulated protein kinase (ERK were measured. Overexpressing PDK2 promoted cell viability, adhesion and inhibited cell migration in MSCs (all P<0.05. qRT-PCR assay showed a potent increase in the mRNA expressions of all chondrogenesis markers in response to overexpressing PDK2 (P<0.01 or P<0.05. PDK2 overexpression also induced a significant accumulation in mRNA and protein expressions of JNK, p38MAPK and ERK in MSCs compared to the control (P<0.01 or P<0.05. Meanwhile, silencing PDK2 exerted the opposite effects on MSCs. This study shows a preliminary positive role and potential mechanisms of PDK2 in chondrogenic differentiation of MSCs. It lays the theoretical groundwork for uncovering the functions of PDK2 and provides a promising basis for repairing cartilage lesions in osteoarthritis.

  19. Impact of TGF-β family-related growth factors on chondrogenic differentiation of adipose-derived stem cells isolated from lipoaspirates and infrapatellar fat pads of osteoarthritic patients

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    E López-Ruiz

    2018-04-01

    Full Text Available The success of cell-based approaches for the treatment of cartilage defects requires an optimal autologous cell source with chondrogenic differentiation ability that maintains its differentiated properties and stability following implantation. The objective of this study was to compare the chondrogenic capacity of mesenchymal stem cells (MSCs isolated from lipoaspirates (ASCs and the infrapatellar fat pad (IFPSCs of osteoarthritic patients and treated with transforming growth factor (TGF-β family-related growth factors. Cells were cultured for 6 weeks in a 3D pellet culture system with the chimeric activin A/bone morphogenic protein (BMP-2 ligand (AB235, the chimeric nodal/BMP-2 ligand (NB260 or BMP-2. To investigate the stability of the new cartilage, ASCs-treated pellets were transplanted subcutaneously into severe combined immunodeficiency (SCID mice. Histological and immunohistochemical assessment confirmed that the growth factors induced cartilage differentiation in both isolated cell types. However, reverse transcription-quantitative PCR results showed that ASCs presented a higher chondrogenic potential than IFPSCs. In vivo results revealed that AB235-treated ASCs pellets were larger in size and could form stable cartilage-like tissue as compared to NB260-treated pellets, while BMP-2-treated pellets underwent calcification. The chondrogenic induction of ASCs by AB235 treatment was mediated by SMAD2/3 activation, as proved by immunofluorescence analysis. The results of this study indicated that the combination of ASCs and AB235 might lead to a cell-based cartilage regeneration treatment.

  20. Effects of in vitro low oxygen tension preconditioning of adipose stromal cells on their in vivo chondrogenic potential: application in cartilage tissue repair.

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    Sophie Portron

    Full Text Available PURPOSE: Multipotent stromal cell (MSC-based regenerative strategy has shown promise for the repair of cartilage, an avascular tissue in which cells experience hypoxia. Hypoxia is known to promote the early chondrogenic differentiation of MSC. The aim of our study was therefore to determine whether low oxygen tension could be used to enhance the regenerative potential of MSC for cartilage repair. METHODS: MSC from rabbit or human adipose stromal cells (ASC were preconditioned in vitro in control or chondrogenic (ITS and TGF-β medium and in 21 or 5% O2. Chondrogenic commitment was monitored by measuring COL2A1 and ACAN expression (real-time PCR. Preconditioned rabbit and human ASC were then incorporated into an Si-HPMC hydrogel and injected (i into rabbit articular cartilage defects for 18 weeks or (ii subcutaneously into nude mice for five weeks. The newly formed tissue was qualitatively and quantitatively evaluated by cartilage-specific immunohistological staining and scoring. The phenotype of ASC cultured in a monolayer or within Si-HPMC in control or chondrogenic medium and in 21 or 5% O2 was finally evaluated using real-time PCR. RESULTS/CONCLUSIONS: 5% O2 increased the in vitro expression of chondrogenic markers in ASC cultured in induction medium. Cells implanted within Si-HPMC hydrogel and preconditioned in chondrogenic medium formed a cartilaginous tissue, regardless of the level of oxygen. In addition, the 3D in vitro culture of ASC within Si-HPMC hydrogel was found to reinforce the pro-chondrogenic effects of the induction medium and 5% O2. These data together indicate that although 5% O2 enhances the in vitro chondrogenic differentiation of ASC, it does not enhance their in vivo chondrogenesis. These results also highlight the in vivo chondrogenic potential of ASC and their potential value in cartilage repair.

  1. Induction of mesenchymal stem cell chondrogenic differentiation and functional cartilage microtissue formation for in vivo cartilage regeneration by cartilage extracellular matrix-derived particles.

    Science.gov (United States)

    Yin, Heyong; Wang, Yu; Sun, Zhen; Sun, Xun; Xu, Yichi; Li, Pan; Meng, Haoye; Yu, Xiaoming; Xiao, Bo; Fan, Tian; Wang, Yiguo; Xu, Wenjing; Wang, Aiyuan; Guo, Quanyi; Peng, Jiang; Lu, Shibi

    2016-03-01

    We propose a method of preparing a novel cell carrier derived from natural cartilage extracellular matrix (ECM), designated cartilage ECM-derived particles (CEDPs). Through a series of processes involving pulverization, sieving, and decellularization, fresh cartilage was made into CEDPs with a median diameter of 263 ± 48 μm. Under microgravity culture conditions in a rotary cell culture system (RCCS), bone marrow stromal cells (BMSCs) can proliferate rapidly on the surface of CEDPs with high viability. Histological evaluation and gene expression analysis indicated that BMSCs were differentiated into mature chondrocytes after 21 days of culture without the use of exogenous growth factors. Functional cartilage microtissue aggregates of BMSC-laden CEDPs formed as time in culture increased. Further, the microtissue aggregates were directly implanted into trochlear cartilage defects in a rat model (CEDP+MSC group). Gait analysis and histological results indicated that the CEDP+MSC group obtained better and more rapid joint function recovery and superior cartilage repair compared to the control groups, in which defects were treated with CEDPs alone or only fibrin glue, at both 6 and 12 weeks after surgery. In conclusion, the innovative cell carrier derived from cartilage ECM could promote chondrogenic differentiation of BMSCs, and the direct use of functional cartilage microtissue facilitated cartilage regeneration. This strategy for cell culture, stem cell differentiation and one-step surgery using cartilage microtissue for cartilage repair provides novel prospects for cartilage tissue engineering and may have further broad clinical applications. We proposed a method to prepare a novel cell carrier derived from natural cartilage ECM, termed cartilage ECM-derived particles (CEDPs), which can support proliferation of MSCs and facilitate their chondrogenic differentiation. Further, the direct use of functional cartilage microtissue of MSC-laden CEDP aggregates for

  2. The Effect of Platelet Rich Plasma on Chondrogenic Differentiation of Human Adipose Derived Stem Cells in Transwell Culture

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    Mohammad Mardani

    2013-11-01

    : Our findings indicate that autologous PRP at an optimum concentration had beneficial effects on differentiation of hADSCs in Transwell culture. Further, in vivo studies are necessary to fully define the clinical implications of PRP.

  3. Impact of human platelet lysate on the expansion and chondrogenic capacity of cultured human chondrocytes for cartilage cell therapy.

    Science.gov (United States)

    Sykes, J G; Kuiper, J H; Richardson, J B; Roberts, S; Wright, K T; Kuiper, N J

    2018-05-01

    High hopes have been pinned on regenerative medicine strategies in order to prevent the progression of cartilage damage to osteoarthritis, particularly by autologous chondrocyte implantation (ACI). The loss of chondrocyte phenotype during in vitro monolayer expansion, a necessary step to obtain sufficient cell numbers, may be a key limitation in ACI. In this study, it was determined whether a shorter monolayer expansion approach could improve chondrogenic differentiation. The effects of two supplement types, foetal bovine serum (FBS) and Stemulate™ (a commercial source of human platelet lysate), on the expansion and re-differentiation potential of human chondrocytes, isolated from five individuals, were compared. Chondrocytes were expanded with 10 % FBS or 10 % Stemulate™. Pellets were cultured for 28 d in chondrogenic differentiation medium and assessed for the presence of cartilage matrix molecules and genes associated with chondrogenicity. Stemulate™ significantly enhanced the proliferation rate [average population doubling times: FBS, 25.07 ± 6.98 d (standard error of the mean, SEM) vs. Stemulate™, 13.10 ± 2.57 d (SEM)]. Sulphated glycosaminoglycans (sGAG), total collagen and qRT-PCR analyses of cartilage genes showed that FBS-expanded chondrocytes demonstrated significantly better chondrogenic capacity than Stemulate™-expanded chondrocytes. Histologically, FBS-expanded chondrocyte pellets appeared to be more stable, with a more intense staining for toluidine blue, indicating a greater chondrogenic capacity. Although Stemulate™ positively influenced chondrocyte proliferation, it had a negative effect on chondrogenic differentiation potential. This suggested that, in the treatment of cartilage defects, Stemulate™ might not be the ideal supplement for expanding chondrocytes (which maintained a chondrocyte phenotype) and, hence, for cell therapies (including ACI).

  4. Impact of human platelet lysate on the expansion and chondrogenic capacity of cultured human chondrocytes for cartilage cell therapy

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    JG Sykes

    2018-05-01

    Full Text Available High hopes have been pinned on regenerative medicine strategies in order to prevent the progression of cartilage damage to osteoarthritis, particularly by autologous chondrocyte implantation (ACI. The loss of chondrocyte phenotype during in vitro monolayer expansion, a necessary step to obtain sufficient cell numbers, may be a key limitation in ACI. In this study, it was determined whether a shorter monolayer expansion approach could improve chondrogenic differentiation. The effects of two supplement types, foetal bovine serum (FBS and Stemulate™ (a commercial source of human platelet lysate, on the expansion and re-differentiation potential of human chondrocytes, isolated from five individuals, were compared. Chondrocytes were expanded with 10 % FBS or 10 % Stemulate™. Pellets were cultured for 28 d in chondrogenic differentiation medium and assessed for the presence of cartilage matrix molecules and genes associated with chondrogenicity. Stemulate™ significantly enhanced the proliferation rate [average population doubling times: FBS, 25.07 ± 6.98 d (standard error of the mean, SEM vs. Stemulate™, 13.10 ± 2.57 d (SEM]. Sulphated glycosaminoglycans (sGAG, total collagen and qRT-PCR analyses of cartilage genes showed that FBS-expanded chondrocytes demonstrated significantly better chondrogenic capacity than Stemulate™-expanded chondrocytes. Histologically, FBS-expanded chondrocyte pellets appeared to be more stable, with a more intense staining for toluidine blue, indicating a greater chondrogenic capacity. Although Stemulate™ positively influenced chondrocyte proliferation, it had a negative effect on chondrogenic differentiation potential. This suggested that, in the treatment of cartilage defects, Stemulate™ might not be the ideal supplement for expanding chondrocytes (which maintained a chondrocyte phenotype and, hence, for cell therapies (including ACI.

  5. Incorporating pTGF-β1/calcium phosphate nanoparticles with fibronectin into 3-dimensional collagen/chitosan scaffolds: Efficient, sustained gene delivery to stem cells for chondrogenic differentiation

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    X Cao

    2012-02-01

    Full Text Available The objective of this study was to prepare a 3-dimensional nanoparticle gene delivery system (3D-NGDS based on collagen/chitosan scaffolds, in which plasmid transforming growth factor beta 1 (TGF-β1/calcium phosphate nanoparticles mixed with fibronectin (FN were used to transfect mesenchymal stem cells (MSCs. Scanning electron microscopy was used to characterise the microstructure of 3-dimensional collagen/chitosan scaffolds. An analysis performed to quantify the TGF-b1 concentrations in MSC cultures revealed that the MSCs transfected with the 3D-NGDS showed remarkably high levels of TGF-b1 over long periods, retaining a concentration of TGF-b1 of approximately 10 ng/mL within two weeks, with the highest level (12.6 ng/mL being observed on the 6th day. An immunohistochemistry analysis for collagen type II revealed that much higher production of collagen II from the 9th to 15th day was observed in the 3D-NGDS-transfected MSCs than that in MSCs transfected by the Lipofectamine 2000 method. The glycosaminoglycan content of the 3D-NGDS was comparable to those treated with TGF-β1 as well as TGF-β1 plus dexamethasone, and was significantly higher than those treated with free plasmid and Lipofectamine 2000. A remarkable type I collagen expression inhibition of the 3D-NGDS at day 21 was observed via ELISA. These results suggested that transfection with the 3D-NGDS could successfully induce MSC chondrogenic differentiation in vitro without dexamethasone. In summary, the 3D-NGDS could be developed into a promising alternative method to transfer exogenous nucleic acid to MSCs in clinical trials.

  6. Sox9-regulated miRNA-574-3p inhibits chondrogenic differentiation of mesenchymal stem cells.

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    David Guérit

    Full Text Available The aim of this study was to identify new microRNAs (miRNAs that are modulated during the differentiation of mesenchymal stem cells (MSCs toward chondrocytes. Using large scale miRNA arrays, we compared the expression of miRNAs in MSCs (day 0 and at early time points (day 0.5 and 3 after chondrogenesis induction. Transfection of premiRNA or antagomiRNA was performed on MSCs before chondrogenesis induction and expression of miRNAs and chondrocyte markers was evaluated at different time points during differentiation by RT-qPCR. Among miRNAs that were modulated during chondrogenesis, we identified miR-574-3p as an early up-regulated miRNA. We found that miR-574-3p up-regulation is mediated via direct binding of Sox9 to its promoter region and demonstrated by reporter assay that retinoid X receptor (RXRα is one gene specifically targeted by the miRNA. In vitro transfection of MSCs with premiR-574-3p resulted in the inhibition of chondrogenesis demonstrating its role during the commitment of MSCs towards chondrocytes. In vivo, however, both up- and down-regulation of miR-574-3p expression inhibited differentiation toward cartilage and bone in a model of heterotopic ossification. In conclusion, we demonstrated that Sox9-dependent up-regulation of miR-574-3p results in RXRα down-regulation. Manipulating miR-574-3p levels both in vitro and in vivo inhibited chondrogenesis suggesting that miR-574-3p might be required for chondrocyte lineage maintenance but also that of MSC multipotency.

  7. Sonic hedgehog protein promotes proliferation and chondrogenic differentiation of bone marrow-derived mesenchymal stem cells in vitro.

    Science.gov (United States)

    Warzecha, Jörg; Göttig, Stephan; Brüning, Christian; Lindhorst, Elmar; Arabmothlagh, Mohammad; Kurth, Andreas

    2006-10-01

    Sonic hedgehog (Shh) protein is known to be an important signaling protein in early embryonic development. Also, Shh is involved in the induction of early cartilaginous differentiation of mesenchymal cells in the limb and in the spine. The impact of Shh on adult stem cells, human bone marrow-derived mesenchymal stem cells (MSCs), was tested. The MSCs were treated either with recombinant Sonic hedgehog protein (r-Shh) or with transforming growth factor-beta 1 (TGF-beta(1)) as a positive control in vitro for 3 weeks. The effects on cartilaginous differentiation and proliferation were assayed. MSCs when treated with either Shh or TGF-beta(1) showed expression of cartilage markers aggrecan, Sox9, CEP-68, and collagen type II and X within 3 weeks. Only r-Shh-treated cells showed a very strong cell proliferation and much higher BrdU incorporation in cell assay systems. These are the first data that indicate an important role of Shh for the induction of cartilage production by MSCs in vitro.

  8. Mesenchymal stem cell-derived extracellular matrix enhances chondrogenic phenotype of and cartilage formation by encapsulated chondrocytes in vitro and in vivo.

    Science.gov (United States)

    Yang, Yuanheng; Lin, Hang; Shen, He; Wang, Bing; Lei, Guanghua; Tuan, Rocky S

    2018-03-15

    Mesenchymal stem cell derived extracellular matrix (MSC-ECM) is a natural biomaterial with robust bioactivity and good biocompatibility, and has been studied as a scaffold for tissue engineering. In this investigation, we tested the applicability of using decellularized human bone marrow derived MSC-ECM (hBMSC-ECM) as a culture substrate for chondrocyte expansion in vitro, as well as a scaffold for chondrocyte-based cartilage repair. hBMSC-ECM deposited by hBMSCs cultured on tissue culture plastic (TCP) was harvested, and then subjected to a decellularization process to remove hBMSCs. Compared with chondrocytes grown on TCP, chondrocytes seeded onto hBMSC-ECM exhibited significantly increased proliferation rate, and maintained better chondrocytic phenotype than TCP group. After being expanded to the same cell number and placed in high-density micromass cultures, chondrocytes from the ECM group showed better chondrogenic differentiation profile than those from the TCP group. To test cartilage formation ability, composites of hBMSC-ECM impregnated with chondrocytes were subjected to brief trypsin treatment to allow cell-mediated contraction, and folded to form 3-dimensional chondrocyte-impregnated hBMSC-ECM (Cell/ECM constructs). Upon culture in vitro in chondrogenic medium for 21 days, robust cartilage formation was observed in the Cell/ECM constructs. Similarly prepared Cell/ECM constructs were tested in vivo by subcutaneous implantation into SCID mice. Prominent cartilage formation was observed in the implanted Cell/ECM constructs 14 days post-implantation, with higher sGAG deposition compared to controls consisting of chondrocyte cell sheets. Taken together, these findings demonstrate that hBMSC-ECM is a superior culture substrate for chondrocyte expansion and a bioactive matrix potentially applicable for cartilage regeneration in vivo. Current cell-based treatments for focal cartilage defects face challenges, including chondrocyte dedifferentiation, need for

  9. Mangiferin Reduces the Inhibition of Chondrogenic Differentiation by IL-1β in Mesenchymal Stem Cells from Subchondral Bone and Targets Multiple Aspects of the Smad and SOX9 Pathways

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    Jeong-Eun Huh

    2014-09-01

    Full Text Available Mangiferin is a natural immunomodulator found in plants including mango trees. The effects of mangiferin on chondrogenesis and cartilage repair have not yet been reported. This study was designed to determine the effect of mangiferin on chondrogenic differentiation in IL-1β-stimulated mesenchymal stem cells (MSCs from subchondral bone and to explore the mechanisms underlying these effects. MSCs were isolated from the subchondral bone of rabbit and treated with mangiferin alone and/or interleukin-1β (IL-1β. Mangiferin induced chondrogenic differentiation in MSCs by upregulating transforming growth factor (TGF-β, bone morphogenetic protein (BMP-2, and BMP-4 and several key markers of chondrogenesis, including sex-determining region Y–box (SRY-box containing gene 9 (SOX9, type 2α1 collagen (Col2α1, cartilage link protein, and aggrecan. In IL-1β-stimulated MSCs, mangiferin significantly reversed the production of TGF-β, BMP-2, BMP-4, SOX9, Col2α1, cartilage link protein, and aggrecan, as well as matrix metalloproteinase (MMP-1, MMP-13, and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS5. Mangiferin upregulated the phosphorylation of Smad 2, Smad 3, Smad 1/5/8, and SOX9 in IL-1β-stimulated MSCs. In the presence of mangiferin, SOX9 siRNA suppressed the activation of Smad 2, Smad 3, Smad 1/5/8, aggrecan, and Col2α1 expression. In conclusion, mangiferin exhibits both chondrogenic and chondroprotective effects on damaged MSCs and mediates these effects by targeting multiple aspects of the Smad and SOX9 signaling pathways.

  10. Enhanced chondrogenesis of human nasal septum derived progenitors on nanofibrous scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Shafiee, Abbas [Department of Tissue Engineering, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Stem Cell biology and Tissue Engineering Departments, Stem Cell Technology Research Center, Tehran (Iran, Islamic Republic of); Institute of Health and Biomedical Innovation, Queensland University of Technology (QUT), Brisbane, QLD (Australia); Seyedjafari, Ehsan [Department of Biotechnology, College of Science, University of Tehran, Tehran (Iran, Islamic Republic of); Sadat Taherzadeh, Elham [Stem Cell biology and Tissue Engineering Departments, Stem Cell Technology Research Center, Tehran (Iran, Islamic Republic of); Dinarvand, Peyman [Stem Cell biology and Tissue Engineering Departments, Stem Cell Technology Research Center, Tehran (Iran, Islamic Republic of); The Edward A. Doisy Department of Biochemistry and Molecular Biology, Saint Louis University School of Medicine, Saint Louis, MO (United States); Soleimani, Masoud [Hematology Department, Faculty of Medical Science, Tarbiat Modares University, Tehran (Iran, Islamic Republic of); Ai, Jafar, E-mail: jafar_ai@tums.ac.ir [Department of Tissue Engineering, School of Advanced Technologies in Medicine, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of); Brain and Spinal Injury Research Center, Imam Hospital, Tehran University of Medical Sciences, Tehran (Iran, Islamic Republic of)

    2014-07-01

    Topographical cues can be exploited to regulate stem cell attachment, proliferation, differentiation and function in vitro and in vivo. In this study, we aimed to investigate the influence of different nanofibrous topographies on the chondrogenic differentiation potential of nasal septum derived progenitors (NSP) in vitro. Aligned and randomly oriented Ploy (L-lactide) (PLLA)/Polycaprolactone (PCL) hybrid scaffolds were fabricated via electrospinning. First, scaffolds were fully characterized, and then NSP were seeded on them to study their capacity to support stem cell attachment, proliferation and chondrogenic differentiation. Compared to randomly oriented nanofibers, aligned scaffolds showed a high degree of nanofiber alignment with much better tensile strength properties. Both scaffolds supported NSP adhesion, proliferation and chondrogenic differentiation. Despite the higher rate of cell proliferation on random scaffolds, a better chondrogenic differentiation was observed on aligned nanofibers as deduced from higher expression of chondrogenic markers such as collagen type II and aggrecan on aligned scaffolds. These findings demonstrate that electrospun constructs maintain NSP proliferation and differentiation, and that the aligned nanofibrous scaffolds can significantly enhance chondrogenic differentiation of nasal septum derived progenitors. - Highlights: • Electrospun nanofiber scaffolds with different topographies were fabricated. • Aligned nanofiber scaffolds had better tensile strength properties. • Nasal septum derived progenitors were cultured on nanofibrous scaffolds. • Both topographies support proliferation and chondrogenic differentiation. • Better chondrogenic differentiation was observed on aligned nanofibers.

  11. Enhanced chondrogenesis of human nasal septum derived progenitors on nanofibrous scaffolds

    International Nuclear Information System (INIS)

    Shafiee, Abbas; Seyedjafari, Ehsan; Sadat Taherzadeh, Elham; Dinarvand, Peyman; Soleimani, Masoud; Ai, Jafar

    2014-01-01

    Topographical cues can be exploited to regulate stem cell attachment, proliferation, differentiation and function in vitro and in vivo. In this study, we aimed to investigate the influence of different nanofibrous topographies on the chondrogenic differentiation potential of nasal septum derived progenitors (NSP) in vitro. Aligned and randomly oriented Ploy (L-lactide) (PLLA)/Polycaprolactone (PCL) hybrid scaffolds were fabricated via electrospinning. First, scaffolds were fully characterized, and then NSP were seeded on them to study their capacity to support stem cell attachment, proliferation and chondrogenic differentiation. Compared to randomly oriented nanofibers, aligned scaffolds showed a high degree of nanofiber alignment with much better tensile strength properties. Both scaffolds supported NSP adhesion, proliferation and chondrogenic differentiation. Despite the higher rate of cell proliferation on random scaffolds, a better chondrogenic differentiation was observed on aligned nanofibers as deduced from higher expression of chondrogenic markers such as collagen type II and aggrecan on aligned scaffolds. These findings demonstrate that electrospun constructs maintain NSP proliferation and differentiation, and that the aligned nanofibrous scaffolds can significantly enhance chondrogenic differentiation of nasal septum derived progenitors. - Highlights: • Electrospun nanofiber scaffolds with different topographies were fabricated. • Aligned nanofiber scaffolds had better tensile strength properties. • Nasal septum derived progenitors were cultured on nanofibrous scaffolds. • Both topographies support proliferation and chondrogenic differentiation. • Better chondrogenic differentiation was observed on aligned nanofibers

  12. Chondrogenic potential of mesenchymal stem cells from patients with rheumatoid arthritis and osteoarthritis: measurements in a microculture system.

    Science.gov (United States)

    Dudics, Valeria; Kunstár, Aliz; Kovács, János; Lakatos, Tamás; Géher, Pál; Gömör, Béla; Monostori, Eva; Uher, Ferenc

    2009-01-01

    Mesenchymal stem cells (MSCs) have the potential to differentiate into distinct mesenchymal tissues; including cartilage and bone, they can be an attractive cell source for cartilage tissue engineering approaches. Our objective here was to compare the in vitro chondrogenic potential of MSCs isolated from patients with rheumatoid arthritis (RA) and osteoarthritis (OA) with cells from normal donors. Marrow samples were removed during bone surgery and adherent cell cultures were established. The cells were then passed into a newly developed microaggregate culture system in a medium containing transforming growth factor beta3, insulin, dexamethasone and/or demineralized bone matrix. In vitro chondrogenic activity was measured as metabolic sulfate incorporation and type II collagen expression in pellet cultures. Culture-expanded MSCs from RA and OA patients did not differ significantly from the normal population with respect to their chondrogenic potential in vitro. Capability of total protein and proteoglycan synthesis as well as collagen II mRNA expression by cell aggregates was similar for all cell preparations in the presence of the appropriate growth and differentiation factors. Chondroprotective drugs such as chondroitin sulfate and glucosamine enhanced, whereas chloroquine inhibited chondrogenesis in normal donor-derived or patient-derived MSC cultures. Galectin-1, a beta-galactoside-binding protein with marked anti-inflammatory activity, stimulated the chondrogenic differentiation of mesenchymal cells in low (<2 microg/ml) concentration. These findings show that MSCs from RA and OA patients possess similar chondrogenic potential as MSCs isolated from healthy donors, therefore these cells may serve as a potential new prospect in cartilage replacement therapy. 2008 S. Karger AG, Basel.

  13. Chondrogenic potential of canine articular cartilage derived cells (cACCs

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    Nowak Urszula

    2016-01-01

    Full Text Available In the present paper, the potential of canine articular cartilage-derived cells (cACCs for chondrogenic differentiation was evaluated. The effectiveness of cACCs’ lineage commitment was analyzed after 14 days of culture in chondorgenic and non-chondrogenic conditions. Formation of proteoglycan-rich extracellular matrix was assessed using histochemical staining – Alcian Blue and Safranin-O, while elemental composition was determined by means of SEM-EDX. Additionally, ultrastructure of cACCs was evaluated using TEM. The expression of genes involved in chondrogenesis was monitored with quantitative Real Time PCR. Results obtained indicate that the potential of cACCs for cartilagous extracellular matrix formation may be maintained only in chondrogenic cultures. The formation of specific chondro-nodules was not observed in a non-chondrogenic culture environment. The analysis of cACCs’ ultrastructure, both in non-chondrogenic and chondrogenic cultures, revealed well-developed rough endoplasmatic reticulum and presence of mitochondria. The cACCs in chondrogenic medium shed an increased number of microvesicles. Furthermore, it was shown that the extracellular matrix of cACCs in chondrogenic cultures is rich in potassium and molybdenum. Additionally, it was determined that gene expression of collagen type II, aggrecan and SOX-9 was significantly increased during chondrogenic differentiation of cACCs. Results obtained indicate that the culture environment may significantly influence the cartilage phenotype of cACCs during long term culture.

  14. Anterior cruciate ligament-derived cells have high chondrogenic potential.

    Science.gov (United States)

    Furumatsu, Takayuki; Hachioji, Motomi; Saiga, Kenta; Takata, Naoki; Yokoyama, Yusuke; Ozaki, Toshifumi

    2010-01-01

    Anterior cruciate ligament (ACL)-derived cells have a character different from medial collateral ligament (MCL)-derived cells. However, the critical difference between ACL and MCL is still unclear in their healing potential and cellular response. The objective of this study was to investigate the mesenchymal differentiation property of each ligament-derived cell. Both ligament-derived cells differentiated into adipogenic, osteogenic, and chondrogenic lineages. In chondrogenesis, ACL-derived cells had the higher chondrogenic property than MCL-derived cells. The chondrogenic marker genes, Sox9 and alpha1(II) collagen (Col2a1), were induced faster in ACL-derived pellets than in MCL-derived pellets. Sox9 expression preceded the increase of Col2a1 in both pellet-cultured cells. However, the expression level of Sox9 and a ligament/tendon transcription factor Scleraxis did not parallel the increase of Col2a1 expression along with chondrogenic induction. The present study demonstrates that the balance between Sox9 and Scleraxis have an important role in the chondrogenic differentiation of ligament-derived cells. Copyright 2009 Elsevier Inc. All rights reserved.

  15. Oxygen tension regulates the osteogenic, chondrogenic and endochondral phenotype of bone marrow derived mesenchymal stem cells

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    Sheehy, Eamon J.; Buckley, Conor T. [Trinity Centre for Bioengineering, School of Engineering, Trinity College Dublin, Dublin 2 (Ireland); Kelly, Daniel J., E-mail: kellyd9@tcd.ie [Trinity Centre for Bioengineering, School of Engineering, Trinity College Dublin, Dublin 2 (Ireland)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Expansion in low oxygen enhances MSC proliferation and osteogenesis. Black-Right-Pointing-Pointer Differentiation in low oxygen enhances chondrogenesis and suppresses hypertrophy. Black-Right-Pointing-Pointer Oxygen can regulate the MSC phenotype for use in tissue engineering applications. -- Abstract: The local oxygen tension is a key regulator of the fate of mesenchymal stem cells (MSCs). The objective of this study was to investigate the effect of a low oxygen tension during expansion and differentiation on the proliferation kinetics as well as the subsequent osteogenic and chondrogenic potential of MSCs. We first hypothesised that expansion in a low oxygen tension (5% pO{sub 2}) would improve both the subsequent osteogenic and chondrogenic potential of MSCs compared to expansion in a normoxic environment (20% pO{sub 2}). Furthermore, we hypothesised that chondrogenic differentiation in a low oxygen environment would suppress hypertrophy of MSCs cultured in both pellets and hydrogels used in tissue engineering strategies. MSCs expanded at 5% pO{sub 2} proliferated faster forming larger colonies, resulting in higher cell yields. Expansion at 5% pO{sub 2} also enhanced subsequent osteogenesis of MSCs, whereas differentiation at 5% pO{sub 2} was found to be a more potent promoter of chondrogenesis than expansion at 5% pO{sub 2}. Greater collagen accumulation, and more intense staining for collagen types I and X, was observed in pellets maintained at 20% pO{sub 2} compared to 5% pO{sub 2}. Both pellets and hydrogels stained more intensely for type II collagen when undergoing chondrogenesis in a low oxygen environment. Differentiation at 5% pO{sub 2} also appeared to inhibit hypertrophy in both pellets and hydrogels, as demonstrated by reduced collagen type X and Alizarin Red staining and alkaline phosphatase activity. This study demonstrates that the local oxygen environment can be manipulated in vitro to either stabilise a

  16. Hydrostatic pressure acts to stabilise a chondrogenic phenotype in porcine joint tissue derived stem cells

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    T Vinardell

    2012-02-01

    Full Text Available Hydrostatic pressure (HP is a key component of the in vivo joint environment and has been shown to enhance chondrogenesis of stem cells. The objective of this study was to investigate the interaction between HP and TGF-β3 on both the initiation and maintenance of a chondrogenic phenotype for joint tissue derived stem cells. Pellets generated from porcine chondrocytes (CCs, synovial membrane derived stem cells (SDSCs and infrapatellar fat pad derived stem cells (FPSCs were subjected to 10 MPa of cyclic HP (4 h/day and different concentrations of TGF-β3 (0, 1 and 10 ng/mL for 14 days. CCs and stem cells were observed to respond differentially to both HP and TGF-β3 stimulation. HP in the absence of TGF-β3 did not induce robust chondrogenic differentiation of stem cells. At low concentrations of TGF-β3 (1 ng/mL, HP acted to enhance chondrogenesis of both SDSCs and FPSCs, as evident by a 3-fold increase in Sox9 expression and a significant increase in glycosaminoglycan accumulation. In contrast, HP had no effect on cartilage-specific matrix synthesis at higher concentrations of TGF-β3 (10 ng/mL. Critically, HP appears to play a key role in the maintenance of a chondrogenic phenotype, as evident by a down-regulation of the hypertrophic markers type X collagen and Indian hedgehog in SDSCs irrespective of the cytokine concentration. In the context of stem cell based therapies for cartilage repair, this study demonstrates the importance of considering how joint specific environmental factors interact to regulate not only the initiation of chondrogenesis, but also the development of a stable hyaline-like repair tissue.

  17. Comparative Analysis of Osteogenic/Chondrogenic Differentiation Potential in Primary Limb Bud-Derived and C3H10T1/2 Cell Line-Based Mouse Micromass Cultures

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    Róza Zákány

    2013-08-01

    Full Text Available Murine micromass models have been extensively applied to study chondrogenesis and osteogenesis to elucidate pathways of endochondral bone formation. Here we provide a detailed comparative analysis of the differentiation potential of micromass cultures established from either BMP-2 overexpressing C3H10T1/2 cells or mouse embryonic limb bud-derived chondroprogenitor cells, using micromass cultures from untransfected C3H10T1/2 cells as controls. Although the BMP-2 overexpressing C3H10T1/2 cells failed to form chondrogenic nodules, cells of both models expressed mRNA transcripts for major cartilage-specific marker genes including Sox9, Acan, Col2a1, Snorc, and Hapln1 at similar temporal sequence, while notable lubricin expression was only detected in primary cultures. Furthermore, mRNA transcripts for markers of osteogenic differentiation including Runx2, Osterix, alkaline phosphatase, osteopontin and osteocalcin were detected in both models, along with matrix calcification. Although the adipogenic lineage-specific marker gene FABP4 was also expressed in micromass cultures, Oil Red O-positive cells along with PPARγ2 transcripts were only detected in C3H10T1/2-derived micromass cultures. Apart from lineage-specific marker genes, pluripotency factors (Nanog and Sox2 were also expressed in these models, reflecting on the presence of various mesenchymal lineages as well as undifferentiated cells. This cellular heterogeneity has to be taken into consideration for the interpretation of data obtained by using these models.

  18. Bioinspired seeding of biomaterials using three dimensional microtissues induces chondrogenic stem cell differentiation and cartilage formation under growth factor free conditions

    NARCIS (Netherlands)

    Leijten, Jeroen Christianus Hermanus; Moreira Teixeira, Liliana; Bolander, J.; Ji, W.; Vanspauwen, B.; Lammertyn, J.; Schrooten, J.; Luyten, F.P.

    2016-01-01

    Cell laden biomaterials are archetypically seeded with individual cells and steered into the desired behavior using exogenous stimuli to control growth and differentiation. In contrast, direct cell-cell contact is instructive and even essential for natural tissue formation. Namely, microaggregation

  19. The Chondrogenic Induction Potential for Bone Marrow-Derived Stem Cells between Autologous Platelet-Rich Plasma and Common Chondrogenic Induction Agents: A Preliminary Comparative Study

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    Shan-zheng Wang

    2015-01-01

    Full Text Available The interests in platelet-rich plasma (PRP and their application in stem cell therapy have contributed to a better understanding of the basic biology of the prochondrogenesis effect on bone marrow-derived stem cells (BMSCs. We aimed at comparing the effect of autologous PRP with common chondrogenic induction agents (CCIAs on the chondrogenic differentiation of BMSCs. Rabbit BMSCs were isolated and characterized by flow cytometry and differentiated towards adipocytes and osteoblasts. The chondrogenic response of BMSCs to autologous PRP and CCIAs which included transforming growth factor-β1 (TGF-β1, dexamethasone (DEX, and vitamin C (Vc was examined by cell pellet culture. The isolated BMSCs after two passages highly expressed CD29 and CD44 but minimally expressed CD45. The osteogenic and adipogenic differentiation potentials of the isolated BMSCs were also confirmed. Compared with common CCIAs, autologous PRP significantly upregulated the chondrogenic related gene expression, including Col-2, AGC, and Sox-9. Osteogenic related gene expression, including Col-1 and OCN, was not of statistical significance between these two groups. Thus, our data shows that, compared with common chondrogenic induction agents, autologous PRP can be more effective in promoting the chondrogenesis of BMSCs.

  20. Cartilage Protective and Chondrogenic Capacity of WIN-34B, a New Herbal Agent, in the Collagenase-Induced Osteoarthritis Rabbit Model and in Progenitor Cells from Subchondral Bone

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    Jeong-Eun Huh

    2013-01-01

    Full Text Available We sought to determine the cartilage repair capacity of WIN-34B in the collagenase-induced osteoarthritis rabbit model and in progenitor cells from subchondral bone. The cartilage protective effect of WIN-34B was measured by clinical and histological scores, cartilage area, and proteoglycan and collagen contents in the collagenase-induced osteoarthritis rabbit model. The efficacy of chondrogenic differentiation of WIN-34B was assessed by expression of CD105, CD73, type II collagen, and aggrecan in vivo and was analyzed by the surface markers of progenitor cells, the mRNA levels of chondrogenic marker genes, and the level of proteoglycan, GAG, and type II collagen in vitro. Oral administration of WIN-34B significantly increased cartilage area, and this was associated with the recovery of proteoglycan and collagen content. Moreover, WIN-34B at 200 mg/kg significantly increased the expression of CD105, CD73, type II collagen, and aggrecan compared to the vehicle group. WIN-34B markedly enhanced the chondrogenic differentiation of CD105 and type II collagen in the progenitor cells from subchondral bone. Also, we confirmed that treatment with WIN-34B strongly increased the number of SH-2(CD105 cells and expression type II collagen in subchondral progenitor cells. Moreover, WIN-34B significantly increased proteoglycan, as measured by alcian blue staining; the mRNA level of type II α1 collagen, cartilage link protein, and aggrecan; and the inhibition of cartilage matrix molecules, such as GAG and type II collagen, in IL-1β-treated progenitor cells. These findings suggest that WIN-34B could be a potential candidate for effective anti-osteoarthritic therapy with cartilage repair as well as cartilage protection via enhancement of chondrogenic differentiation in the collagenase-induced osteoarthritis rabbit model and progenitor cells from subchondral bone.

  1. PRC1 Prevents Replication Stress during Chondrogenic Transit Amplification

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    Frank Spaapen

    2017-12-01

    Full Text Available Transit amplification (TA, a state of combined, rapid proliferative expansion and differentiation of stem cell-descendants, remains poorly defined at the molecular level. The Polycomb Repressive Complex 1 (PRC1 protein BMI1 has been localized to TA compartments, yet its exact role in TA is unclear. PRC1 proteins control gene expression, cell proliferation and DNA-damage repair. Coordination of such DNA-templated activities during TA is predicted to be crucial to support DNA replication and differentiation-associated transcriptional programming. We here examined whether chondrogenesis provides a relevant biological context for synchronized coordination of these chromatin-based tasks by BMI1. Taking advantage of a prominently featuring TA-phase during chondrogenesis in vitro and in vivo, we here report that TA is completely dependent on intact PRC1 function. BMI1-depleted chondrogenic progenitors rapidly accumulate double strand DNA breaks during DNA replication, present massive non-H3K27me3-directed transcriptional deregulation and fail to undergo chondrogenic TA. Genome-wide accumulation of Topoisomerase 2α and Geminin suggests a model in which PRC1 synchronizes replication and transcription during rapid chondrogenic progenitor expansion. Our combined data reveals for the first time a vital cell-autonomous role for PRC1 during chondrogenesis. We provide evidence that chondrocyte hyper-replication and hypertrophy represent a unique example of programmed senescence in vivo. These findings provide new perspectives on PRC1 function in development and disease.

  2. Direct induction of chondrogenic cells from human dermal fibroblast culture by defined factors.

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    Hidetatsu Outani

    Full Text Available The repair of large cartilage defects with hyaline cartilage continues to be a challenging clinical issue. We recently reported that the forced expression of two reprogramming factors (c-Myc and Klf4 and one chondrogenic factor (SOX9 can induce chondrogenic cells from mouse dermal fibroblast culture without going through a pluripotent state. We here generated induced chondrogenic (iChon cells from human dermal fibroblast (HDF culture with the same factors. We developed a chondrocyte-specific COL11A2 promoter/enhancer lentiviral reporter vector to select iChon cells. The human iChon cells expressed marker genes for chondrocytes but not fibroblasts, and were derived from non-chondrogenic COL11A2-negative cells. The human iChon cells formed cartilage but not tumors in nude mice. This approach could lead to the preparation of cartilage directly from skin in human, without going through pluripotent stem cells.

  3. Evaluation of insulin medium or chondrogenic medium on proliferation and chondrogenesis of ATDC5 cells.

    Science.gov (United States)

    Yao, Yongchang; Zhai, Zhichen; Wang, Yingjun

    2014-01-01

    The ATDC5 cell line is regarded as an excellent cell model for chondrogenesis. In most studies with ATDC5 cells, insulin medium (IM) was used to induce chondrogenesis while chondrogenic medium (CM), which was usually applied in chondrogenesis of mesenchymal stem cells (MSCs), was rarely used for ATDC5 cells. This study was mainly designed to investigate the effect of IM, CM, and growth medium (GM) on chondrogenesis of ATDC5 cells. ATDC5 cells were, respectively, cultured in IM, CM, and GM for a certain time. Then the proliferation and the chondrogenesis progress of cells in these groups were analyzed. Compared with CM and GM, IM promoted the proliferation of cells significantly. CM was effective for enhancement of cartilage specific markers, while IM induced the cells to express endochondral ossification related genes. Although GAG deposition per cell in CM group was significantly higher than that in IM and GM groups, the total GAG contents in IM group were the most. This study demonstrated that CM focused on induction of chondrogenic differentiation while IM was in favor of promoting proliferation and expression of endochondral ossification related genes. Combinational use of these two media would be more beneficial to bone/cartilage repair.

  4. Evaluation of Insulin Medium or Chondrogenic Medium on Proliferation and Chondrogenesis of ATDC5 Cells

    Directory of Open Access Journals (Sweden)

    Yongchang Yao

    2014-01-01

    Full Text Available Background. The ATDC5 cell line is regarded as an excellent cell model for chondrogenesis. In most studies with ATDC5 cells, insulin medium (IM was used to induce chondrogenesis while chondrogenic medium (CM, which was usually applied in chondrogenesis of mesenchymal stem cells (MSCs, was rarely used for ATDC5 cells. This study was mainly designed to investigate the effect of IM, CM, and growth medium (GM on chondrogenesis of ATDC5 cells. Methods. ATDC5 cells were, respectively, cultured in IM, CM, and GM for a certain time. Then the proliferation and the chondrogenesis progress of cells in these groups were analyzed. Results. Compared with CM and GM, IM promoted the proliferation of cells significantly. CM was effective for enhancement of cartilage specific markers, while IM induced the cells to express endochondral ossification related genes. Although GAG deposition per cell in CM group was significantly higher than that in IM and GM groups, the total GAG contents in IM group were the most. Conclusion. This study demonstrated that CM focused on induction of chondrogenic differentiation while IM was in favor of promoting proliferation and expression of endochondral ossification related genes. Combinational use of these two media would be more beneficial to bone/cartilage repair.

  5. Norepinephrine inhibition of mesenchymal stem cell and chondrogenic progenitor cell chondrogenesis and acceleration of chondrogenic hypertrophy.

    Science.gov (United States)

    Jenei-Lanzl, Zsuzsa; Grässel, Susanne; Pongratz, Georg; Kees, Frieder; Miosge, Nicolai; Angele, Peter; Straub, Rainer H

    2014-09-01

    Mesenchymal progenitor cell chondrogenesis is the biologic platform for the generation or regeneration of cartilage, but the external influence of the sympathetic nervous system on this process is not yet known. Sympathetic nerve fibers are present in articular tissue, and the sympathetic nervous system influences the musculoskeletal system by, for example, increasing osteoclastogenesis. This study was initiated to explore the role of the sympathetic neurotransmitter norepinephrine (NE) in mesenchymal stem cell (MSC)-dependent and cartilage progenitor cell (CPC)-dependent chondrogenesis. Using human MSCs or CPCs, chondrogenic differentiation was induced in the presence of NE, the specific β-adrenergic receptor (β-AR) agonist isoproterenol, and the specific β-AR antagonist nadolol. We studied sympathetic nerve fibers, tyrosine hydroxylase (TH) expression, catecholamine biosynthesis, and synovial fluid levels in human joints, as well as cartilage-specific matrix deposition during differentiation. TH+ sympathetic nerve fibers were present in the synovial tissue, meniscus, and subchondral bone marrow. In addition, synovial fluid from patients with knee trauma demonstrated high concentrations of NE. During MSC or CPC chondrogenesis, β-AR were expressed. Chondrogenic aggregates treated with NE or isoproterenol synthesized lower amounts of type II collagen and glycosaminoglycans. NE and isoproterenol treatment dose-dependently increased the levels of cartilage hypertrophy markers (type X collagen and matrix metalloproteinase 13). Nadolol reversed the inhibition of chondrogenesis and the up-regulation of cartilage hypertrophy. Our findings demonstrate NE-dependent inhibition of chondrogenesis and acceleration of hypertrophic differentiation. By inhibiting cartilage repair, these sympathetic influences can be important after joint trauma. These findings may be a basis for novel neurochondrogenic therapeutic options. Copyright © 2014 by the American College of

  6. Hierarchical scaffolds enhance osteogenic differentiation of human Wharton’s jelly derived stem cells

    International Nuclear Information System (INIS)

    Canha-Gouveia, Analuce; Rita Costa-Pinto, Ana; Martins, Albino M; Sousa, Rui A; Reis, Rui L; Neves, Nuno M; Silva, Nuno A; Salgado, António J; Sousa, Nuno; Faria, Susana

    2015-01-01

    Hierarchical structures, constituted by polymeric nano and microfibers, have been considered promising scaffolds for tissue engineering strategies, mainly because they mimic, in some way, the complexity and nanoscale detail observed in real organs. The chondrogenic potential of these scaffolds has been previously demonstrated, but their osteogenic potential is not yet corroborated. In order to assess if a hierarchical structure, with nanoscale details incorporated, is an improved scaffold for bone tissue regeneration, we evaluate cell adhesion, proliferation, and osteogenic differentiation of human Wharton’s jelly derived stem cells (hWJSCs), seeded into hierarchical fibrous scaffolds. Biological data corroborates that hierarchical fibrous scaffolds show an enhanced cell entrapment when compared to rapid prototyped scaffolds without nanofibers. Furthermore, upregulation of bone specific genes and calcium phosphate deposition confirms the successful osteogenic differentiation of hWJSCs on these scaffolds. These results support our hypothesis that a scaffold with hierarchical structure, in conjugation with hWJSCs, represents a possible feasible strategy for bone tissue engineering applications. (paper)

  7. Irisin Enhances Osteoblast Differentiation In Vitro

    Directory of Open Access Journals (Sweden)

    Graziana Colaianni

    2014-01-01

    Full Text Available It has been recently demonstrated that exercise activity increases the expression of the myokine Irisin in skeletal muscle, which is able to drive the transition of white to brown adipocytes, likely following a phenomenon of transdifferentiation. This new evidence supports the idea that muscle can be considered an endocrine organ, given its ability to target adipose tissue by promoting energy expenditure. In accordance with these new findings, we hypothesized that Irisin is directly involved in bone metabolism, demonstrating its ability to increase the differentiation of bone marrow stromal cells into mature osteoblasts. Firstly, we confirmed that myoblasts from mice subjected to 3 weeks of free wheel running increased Irisin expression compared to nonexercised state. The conditioned media (CM collected from myoblasts of exercised mice induced osteoblast differentiation in vitro to a greater extent than those of mice housed in resting conditions. Furthermore, the differentiated osteoblasts increased alkaline phosphatase and collagen I expression by an Irisin-dependent mechanism. Our results show, for the first time, that Irisin directly targets osteoblasts, enhancing their differentiation. This finding advances notable perspectives in future studies which could satisfy the ongoing research of exercise-mimetic therapies with anabolic action on the skeleton.

  8. Fingerprint image enhancement by differential hysteresis processing.

    Science.gov (United States)

    Blotta, Eduardo; Moler, Emilce

    2004-05-10

    A new method to enhance defective fingerprints images through image digital processing tools is presented in this work. When the fingerprints have been taken without any care, blurred and in some cases mostly illegible, as in the case presented here, their classification and comparison becomes nearly impossible. A combination of spatial domain filters, including a technique called differential hysteresis processing (DHP), is applied to improve these kind of images. This set of filtering methods proved to be satisfactory in a wide range of cases by uncovering hidden details that helped to identify persons. Dactyloscopy experts from Policia Federal Argentina and the EAAF have validated these results.

  9. Metabolic programming of mesenchymal stromal cells by oxygen tension directs chondrogenic cell fate

    NARCIS (Netherlands)

    Leijten, Jeroen Christianus Hermanus; Georgi, Nicole; Moreira Teixeira, Liliana; van Blitterswijk, Clemens; Post, Janine Nicole; Karperien, Hermanus Bernardus Johannes

    2014-01-01

    Actively steering the chondrogenic differentiation of mesenchymal stromal cells (MSCs) into either permanent cartilage or hypertrophic cartilage destined to be replaced by bone has not yet been possible. During limb development, the developing long bone is exposed to a concentration gradient of

  10. Impairment of the chondrogenic phase of endochondral ossification in vivo by inhibition of cyclooxygenase-2

    Directory of Open Access Journals (Sweden)

    MPF Janssen

    2017-10-01

    Full Text Available Many studies have reported on the effects of cyclooxygenase-2 (COX-2 inhibition on osteogenesis. However, far less is known about the effects of COX-2 inhibition on chondrogenic differentiation. Previous studies conducted by our group show that COX-2 inhibition influences in vitro chondrogenic differentiation. Importantly, this might have consequences on endochondral ossification processes occurring in vivo, such as bone fracture healing, growth plate development and ectopic generation of cartilage. The goal of our study was to investigate, in vivo, the effect of COX-2 inhibition by celecoxib on the cartilaginous phase of three different endochondral ossification scenarios. 10 mg/kg/day celecoxib or placebo were orally administered for 25 d to skeletally-immature New Zealand White rabbits (n = 6 per group. Endochondral ossification during fracture healing of a non-critical size defect in the ulna, femoral growth plate and ectopically-induced cartilaginous tissue were examined by radiography, micro-computed tomography (µ-CT, histology and gene expression analysis. Celecoxib treatment resulted in delayed bone fracture healing, alterations in growth plate development and progression of mineralisation. In addition, chondrogenic differentiation of ectopically-induced cartilaginous tissue was severely impaired by celecoxib. In conclusion, we found that celecoxib impaired the chondrogenic phase of endochondral ossification.

  11. Surface Markers for Chondrogenic Determination: A Highlight of Synovium-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Douglas D. Campbell

    2012-11-01

    Full Text Available Cartilage tissue engineering is a promising field in regenerative medicine that can provide substantial relief to people suffering from degenerative cartilage disease. Current research shows the greatest chondrogenic potential for healthy articular cartilage growth with minimal hypertrophic differentiation to be from mesenchymal stem cells (MSCs of synovial origin. These stem cells have the capacity for differentiation into multiple cell lineages related to mesenchymal tissue; however, evidence exists for cell surface markers that specify a greater potential for chondrogenesis than other differentiation fates. This review will examine relevant literature to summarize the chondrogenic differentiation capacities of tested synovium-derived stem cell (SDSC surface markers, along with a discussion about various other markers that may hold potential, yet require further investigation. With this information, a potential clinical benefit exists to develop a screening system for SDSCs that will produce the healthiest articular cartilage possible.

  12. Ihh enhances differentiation of CFK-2 chondrocytic cells and antagonizes PTHrP-mediated activation of PKA.

    Science.gov (United States)

    Deckelbaum, Ron A; Chan, George; Miao, Dengshun; Goltzman, David; Karaplis, Andrew C

    2002-07-15

    Indian Hedgehog (Ihh), a member of the hedgehog (HH) family of secreted morphogens, and parathyroid hormone-related peptide (PTHrP) are key regulators of cartilage cell (chondrocyte) differentiation. We have investigated, in vitro, the actions of HH signalling and its possible interplay with PTHrP using rat CFK-2 chondrocytic cells. Markers of chondrocyte differentiation [alkaline phosphatase (ALP) activity, and type II (Col2a1) and type X collagen (Col10a1) expression] were enhanced by overexpression of Ihh or its N-terminal domain (N-Ihh), effects mimicked by exogenous administration of recombinant N-terminal HH peptide. Moreover, a missense mutation mapping to the N-terminal domain of Ihh (W160G) reduces the capacity of N-Ihh to induce differentiation. Prolonged exposure of CFK-2 cells to exogenous N-Shh (5x10(-9) M) in the presence of PTHrP (10(-8) M) or forskolin (10(-7) M) resulted in perturbation of HH-mediated differentiation. In addition, overexpression of a constitutively active form of the PTHrP receptor (PTHR1 H223R) inhibited Ihh-mediated differentiation, implicating activation of protein kinase A (PKA) by PTHR1 as a probable mediator of the antagonistic effects of PTHrP. Conversely, overexpression of Ihh/N-Ihh or exogenous treatment with N-Shh led to dampening of PTHrP-mediated activation of PKA. Taken together, our data suggest that Ihh harbors the capacity to induce rather than inhibit chondrogenic differentiation, that PTHrP antagonizes HH-mediated differentiation through a PKA-dependent mechanism and that HH signalling, in turn, modulates PTHrP action through functional inhibition of signalling by PTHR1 to PKA.

  13. Actin depolymerization enhances adipogenic differentiation in human stromal stem cells.

    Science.gov (United States)

    Chen, Li; Hu, Huimin; Qiu, Weimin; Shi, Kaikai; Kassem, Moustapha

    2018-05-01

    Human stromal stem cells (hMSCs) differentiate into adipocytes that play a role in skeletal tissue homeostasis and whole body energy metabolism. During adipocyte differentiation, hMSCs exhibit significant changes in cell morphology suggesting changes in cytoskeletal organization. Here, we examined the effect of direct modulation of actin microfilament dynamics on adipocyte differentiation. Stabilizing actin filaments in hMSCs by siRNA-mediated knock down of the two main actin depolymerizing factors (ADFs): Cofilin 1 (CFL1) and Destrin (DSTN) or treating the cells by Phalloidin reduced adipocyte differentiation as evidenced by decreased number of mature adipocytes and decreased adipocyte specific gene expression (ADIPOQ, LPL, PPARG, FABP4). In contrast, disruption of actin cytoskeleton by Cytochalasin D enhanced adipocyte differentiation. Follow up studies revealed that the effects of CFL1 on adipocyte differentiation depended on the activity of LIM domain kinase 1 (LIMK1) which is the major upstream kinase of CFL1. Inhibiting LIMK by its specific chemical inhibitor LIMKi inhibited the phosphorylation of CFL1 and actin polymerization, and enhanced the adipocyte differentiation. Moreover, treating hMSCs by Cytochalasin D inhibited ERK and Smad2 signaling and this was associated with enhanced adipocyte differentiation. On the other hand, Phalloidin enhanced ERK and Smad2 signaling, but inhibited adipocyte differentiation which was rescued by ERK specific chemical inhibitor U0126. Our data provide a link between restructuring of hMSCs cytoskeleton and hMSCs lineage commitment and differentiation. Copyright © 2018 Elsevier B.V. All rights reserved.

  14. Actin depolymerization enhances adipogenic differentiation in human stromal stem cells

    DEFF Research Database (Denmark)

    Chen, Li; Hu, Huimin; Qiu, Weimin

    2018-01-01

    Human stromal stem cells (hMSCs) differentiate into adipocytes that play a role in skeletal tissue homeostasis and whole body energy metabolism. During adipocyte differentiation, hMSCs exhibit significant changes in cell morphology suggesting changes in cytoskeletal organization. Here, we examined...... differentiation as evidenced by decreased number of mature adipocytes and decreased adipocyte specific gene expression (ADIPOQ, LPL, PPARG, FABP4). In contrast, disruption of actin cytoskeleton by Cytochalasin D enhanced adipocyte differentiation. Follow up studies revealed that the effects of CFL1 on adipocyte...... differentiation depended on the activity of LIM domain kinase 1 (LIMK1) which is the major upstream kinase of CFL1. Inhibiting LIMK by its specific chemical inhibitor LIMKi inhibited the phosphorylation of CFL1 and actin polymerization, and enhanced the adipocyte differentiation. Moreover, treating h...

  15. Actin depolymerization enhances adipogenic differentiation in human stromal stem cells

    Directory of Open Access Journals (Sweden)

    Li Chen

    2018-05-01

    Full Text Available Human stromal stem cells (hMSCs differentiate into adipocytes that play a role in skeletal tissue homeostasis and whole body energy metabolism. During adipocyte differentiation, hMSCs exhibit significant changes in cell morphology suggesting changes in cytoskeletal organization. Here, we examined the effect of direct modulation of actin microfilament dynamics on adipocyte differentiation. Stabilizing actin filaments in hMSCs by siRNA-mediated knock down of the two main actin depolymerizing factors (ADFs: Cofilin 1 (CFL1 and Destrin (DSTN or treating the cells by Phalloidin reduced adipocyte differentiation as evidenced by decreased number of mature adipocytes and decreased adipocyte specific gene expression (ADIPOQ, LPL, PPARG, FABP4. In contrast, disruption of actin cytoskeleton by Cytochalasin D enhanced adipocyte differentiation. Follow up studies revealed that the effects of CFL1 on adipocyte differentiation depended on the activity of LIM domain kinase 1 (LIMK1 which is the major upstream kinase of CFL1. Inhibiting LIMK by its specific chemical inhibitor LIMKi inhibited the phosphorylation of CFL1 and actin polymerization, and enhanced the adipocyte differentiation. Moreover, treating hMSCs by Cytochalasin D inhibited ERK and Smad2 signaling and this was associated with enhanced adipocyte differentiation. On the other hand, Phalloidin enhanced ERK and Smad2 signaling, but inhibited adipocyte differentiation which was rescued by ERK specific chemical inhibitor U0126. Our data provide a link between restructuring of hMSCs cytoskeleton and hMSCs lineage commitment and differentiation. Keywords: Actin cytoskeleton, Actin depolymerizing factors, Adipocyte differentiation, Human stromal stem cells

  16. The canonical Wnt signaling pathway promotes chondrocyte differentiation in a Sox9-dependent manner

    International Nuclear Information System (INIS)

    Yano, Fumiko; Kugimiya, Fumitaka; Ohba, Shinsuke; Ikeda, Toshiyuki; Chikuda, Hirotaka; Ogasawara, Toru; Ogata, Naoshi; Takato, Tsuyoshi; Nakamura, Kozo; Kawaguchi, Hiroshi; Chung, Ung-il

    2005-01-01

    To better understand the role of the canonical Wnt signaling pathway in cartilage development, we adenovirally expressed a constitutively active (Canada) or a dominant negative (dn) form of lymphoid enhancer factor-1 (LEF-1), the main nuclear effector of the pathway, in undifferentiated mesenchymal cells, chondrogenic cells, and primary chondrocytes, and examined the expression of markers for chondrogenic differentiation and hypertrophy. caLEF-1 and LiCl, an activator of the canonical pathway, promoted both chondrogenic differentiation and hypertrophy, whereas dnLEF-1 and the gene silencing of β-catenin suppressed LiCl-promoted effects. To investigate whether these effects were dependent on Sox9, a master regulator of cartilage development, we stimulated Sox9-deficient ES cells with the pathway. caLEF-1 and LiCl promoted both chondrogenic differentiation and hypertrophy in wild-type, but not in Sox9-deficient, cells. The response of Sox9-deficient cells was restored by the adenoviral expression of Sox9. Thus, the canonical Wnt signaling pathway promotes chondrocyte differentiation in a Sox9-dependent manner

  17. Polymodal Transient Receptor Potential Vanilloid (TRPV Ion Channels in Chondrogenic Cells

    Directory of Open Access Journals (Sweden)

    Csilla Szűcs Somogyi

    2015-08-01

    Full Text Available Mature and developing chondrocytes exist in a microenvironment where mechanical load, changes of temperature, osmolarity and acidic pH may influence cellular metabolism. Polymodal Transient Receptor Potential Vanilloid (TRPV receptors are environmental sensors mediating responses through activation of linked intracellular signalling pathways. In chondrogenic high density cultures established from limb buds of chicken and mouse embryos, we identified TRPV1, TRPV2, TRPV3, TRPV4 and TRPV6 mRNA expression with RT-PCR. In both cultures, a switch in the expression pattern of TRPVs was observed during cartilage formation. The inhibition of TRPVs with the non-selective calcium channel blocker ruthenium red diminished chondrogenesis and caused significant inhibition of proliferation. Incubating cell cultures at 41 °C elevated the expression of TRPV1, and increased cartilage matrix production. When chondrogenic cells were exposed to mechanical load at the time of their differentiation into matrix producing chondrocytes, we detected increased mRNA levels of TRPV3. Our results demonstrate that developing chondrocytes express a full palette of TRPV channels and the switch in the expression pattern suggests differentiation stage-dependent roles of TRPVs during cartilage formation. As TRPV1 and TRPV3 expression was altered by thermal and mechanical stimuli, respectively, these are candidate channels that contribute to the transduction of environmental stimuli in chondrogenic cells.

  18. Chondrogenic potential of physically treated bovine cartilage matrix derived porous scaffolds on human dermal fibroblast cells.

    Science.gov (United States)

    Moradi, Ali; Ataollahi, Forough; Sayar, Katayoun; Pramanik, Sumit; Chong, Pan-Pan; Khalil, Alizan Abdul; Kamarul, Tunku; Pingguan-Murphy, Belinda

    2016-01-01

    Extracellular matrices have drawn attention in tissue engineering as potential biomaterials for scaffold fabrication because of their bioactive components. Noninvasive techniques of scaffold fabrication and cross-linking treatments are believed to maintain the integrity of bioactive molecules while providing proper architectural and mechanical properties. Cartilage matrix derived scaffolds are designed to support the maintenance of chondrocytes and provide proper signals for differentiation of chondroinducible cells. Chondroinductive potential of bovine articular cartilage matrix derived porous scaffolds on human dermal fibroblasts and the effect of scaffold shrinkage on chondrogenesis were investigated. An increase in sulfated glycosaminoglycans production along with upregulation of chondrogenic genes confirmed that physically treated cartilage matrix derived scaffolds have chondrogenic potential on human dermal fibroblasts. © 2015 Wiley Periodicals, Inc.

  19. The Distinct Effects of Estrogen and Hydrostatic Pressure on Mesenchymal Stem Cells Differentiation: Involvement of Estrogen Receptor Signaling.

    Science.gov (United States)

    Zhao, Ying; Yi, Fei-Zhou; Zhao, Yin-Hua; Chen, Yong-Jin; Ma, Heng; Zhang, Min

    2016-10-01

    This study aimed to investigate the differential and synergistic effects of mechanical stimulation and estrogen on the proliferation and osteogenic or chondrogenic differentiation potential of bone marrow mesenchymal stem cells (BMSCs) and the roles of estrogen receptor (ER) in them. BMSCs were isolated and cultured using the whole bone marrow adherence method, and flow cytometry was used to identify the surface marker molecules of BMSCs. Cells were pre-treated with 1 nM 17β-estradiol or 1 nM of the estrogen receptor antagonist tamoxifen. Then, the cells were stimulated with hydrostatic pressure. Assessment included flow cytometry analysis of the cell cycle; immunofluorescent staining for F-actin; protein quantification for MAPK protein; and mRNA analysis for Col I, OCN, OPN and BSP after osteogenic induction and Sox-9, Aggrecan and Col-II after chondrogenic induction. Hydrostatic pressure (90 kPa/1 h) and 1 nM 17β-estradiol enhanced the cellular proliferation ability and the cytoskeleton activity but without synergistic biological effects. Estrogen activated ERKs and JNKs simultaneously and promoted the osteogenic differentiation, whereas the pressure just caused JNK-1/2 activation and promoted the chondrogenic differentiation of BMSCs. Estrogen had antagonism effect on chondrogenic promotion of hydrostatic pressure. Mechanobiological effects of hydrostatic pressure are closely associated with ERα activity. MAPK molecules and F-actin were likely to be important mediator molecules in the ER-mediated mechanotransduction of BMSCs.

  20. Cell type dependent morphological adaptation in polyelectrolyte hydrogels governs chondrogenic fate.

    Science.gov (United States)

    Raghothaman, Deepak; Leong, Meng Fatt; Lim, Tze Chiun; Wan, Andrew C A; Ser, Zheng; Lee, Eng Hin; Yang, Zheng

    2016-04-04

    Repair of critical-size articular cartilage defects typically involves delivery of cells in biodegradable, 3D matrices. Differences in the developmental status of mesenchymal stem cells (MSCs) and terminally differentiated mature chondrocytes might be a critical factor in engineering appropriate 3D matrices for articular cartilage tissue engineering. This study examined the relationship between material-driven early cell morphological adaptations and chondrogenic outcomes, by studying the influence of aligned collagen type I (Col I) presentation on chondrocytes and MSC in interfacial polyelectrolyte complexation (IPC)-based hydrogels. In the absence of Col I, both chondrocytes and MSCs adopted rounded cell morphology and formed clusters, with chondrocyte clusters favoring the maintenance of hyaline phenotype, while MSC clusters differentiated to fibro-superficial zone-like chondrocytes. Encapsulated chondrocytes in IPC-Col I hydrogel adopted a fibroblastic morphology forming fibro-superficial zone-like phenotype, which could be reversed by inhibiting actin polymerization using cytochalasin D (CytD). In contrast, adoption of fibroblastic morphology by encapsulated MSCs in IPC-Col I facilitated superior chondrogenesis, generating a mature, hyaline neocartilage tissue. CytD treatment abrogated the elongation of MSCs and brought about a single cell-like state, resulting in insignificant chondrogenic differentiation, underscoring the essential requirement of providing matrix environments that are amenable to cell-cell interactions for robust MSC chondrogenic differentiation. Our study demonstrates that MSCs and culture-expanded chondrocytes favour differential microenvironmental niches and emphasizes the importance of designing biomaterials that meet cell type-specific requirements, in adopting chondrocyte or MSC-based approaches for regenerating hyaline, articular cartilage.

  1. Role of RHEB in Regulating Differentiation Fate of Mesenchymal Stem Cells for Cartilage and Bone Regeneration

    Directory of Open Access Journals (Sweden)

    Sajjad Ashraf

    2017-04-01

    Full Text Available Advances in mesenchymal stem cells (MSCs and cell replacement therapies are promising approaches to treat cartilage and bone defects since substantial differentiation capacities of MSCs match the demands of tissue regeneration. Our understanding of the dynamic process requiring indispensable differentiation of MSCs remains limited. Herein, we describe the role of RHEB (Ras homolog enriched in brain regulating gene signature for differentiation of human adipose derived mesenchymal stem cells (ASCs into chondrogenic, osteogenic, and adipogenic lineages. RHEB-overexpression increases the proliferation of the ASCs. RHEB enhances the chondrogenic differentiation of ASCs in 3D culture via upregulation of SOX9 with concomitant increase in glycosaminoglycans (GAGs, and type II collagen (COL2. RHEB increases the osteogenesis via upregulation of runt related transcription factor 2 (RUNX2 with an increase in the calcium and phosphate contents. RHEB also increases the expression of osteogenic markers, osteonectin and osteopontin. RHEB knockdown ASCs were incapable of expressing sufficient SRY (Sex determining region Y-box 9 (SOX9 and RUNX2, and therefore had decreased chondrogenic and osteogenic differentiation. RHEB-overexpression impaired ASCs differentiation into adipogenic lineage, through downregulation of CCAAT/enhancer binding protein beta (C/EBPβ. Conversely, RHEB knockdown abolished the negative regulation of adipogenesis. We demonstrate that RHEB is a novel regulator, with a critical role in ASCs lineage determination, and RHEB-modulated ASCs may be useful as a cell therapy for cartilage and bone defect treatments.

  2. Comparative study of the chondrogenic potential of human bone marrow stromal cells, neonatal chondrocytes and adult chondrocytes

    International Nuclear Information System (INIS)

    Saha, Sushmita; Kirkham, Jennifer; Wood, David; Curran, Stephen; Yang, Xuebin

    2010-01-01

    Research highlights: → This study has characterised three different cell types under conditions similar to those used for autologous chondrocyte implantation (ACI) for applications in cartilage repair/regeneration. → Compared for the first time the chondrogenic potential of neonatal chondrocytes with human bone marrow stromal cells (HBMSCs) and adult chondrocytes. → Demonstrated that adult chondrocytes hold greatest potential for use in ACI based on their higher proliferation rates, lower alkaline phosphatise activity and enhanced expression of chondrogenic genes. → Demonstrated the need for chondroinduction as a necessary pre-requisite to efficient chondrogenesis in vitro and, by extrapolation, for cell based therapy (e.g. ACI or cartilage tissue engineering). -- Abstract: Cartilage tissue engineering is still a major clinical challenge with optimisation of a suitable source of cells for cartilage repair/regeneration not yet fully addressed. The aims of this study were to compare and contrast the differences in chondrogenic behaviour between human bone marrow stromal cells (HBMSCs), human neonatal and adult chondrocytes to further our understanding of chondroinduction relative to cell maturity and to identify factors that promote chondrogenesis and maintain functional homoeostasis. Cells were cultured in monolayer in either chondrogenic or basal medium, recapitulating procedures used in existing clinical procedures for cell-based therapies. Cell doubling time, morphology and alkaline phosphatase specific activity (ALPSA) were determined at different time points. Expression of chondrogenic markers (SOX9, ACAN and COL2A1) was compared via real time polymerase chain reaction. Amongst the three cell types studied, HBMSCs had the highest ALPSA in basal culture and lowest ALPSA in chondrogenic media. Neonatal chondrocytes were the most proliferative and adult chondrocytes had the lowest ALPSA in basal media. Gene expression analysis revealed a difference in the

  3. Comparative study of the chondrogenic potential of human bone marrow stromal cells, neonatal chondrocytes and adult chondrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Saha, Sushmita [Biomaterials and Tissue Engineering Group, Leeds Dental Institute, University of Leeds, LS29LU (United Kingdom); Kirkham, Jennifer [Biomineralisation Group, Leeds Dental Institute, University of Leeds, LS29LU (United Kingdom); NIHR Leeds Musculoskeletal Biomedical Research Unit, University of Leeds, Chapel Allerton Hospital, Leeds LS74SA (United Kingdom); Wood, David [Biomaterials and Tissue Engineering Group, Leeds Dental Institute, University of Leeds, LS29LU (United Kingdom); Curran, Stephen [Smith and Nephew Research Centre, YO105DF (United Kingdom); Yang, Xuebin, E-mail: X.B.Yang@leeds.ac.uk [Biomaterials and Tissue Engineering Group, Leeds Dental Institute, University of Leeds, LS29LU (United Kingdom); NIHR Leeds Musculoskeletal Biomedical Research Unit, University of Leeds, Chapel Allerton Hospital, Leeds LS74SA (United Kingdom)

    2010-10-22

    Research highlights: {yields} This study has characterised three different cell types under conditions similar to those used for autologous chondrocyte implantation (ACI) for applications in cartilage repair/regeneration. {yields} Compared for the first time the chondrogenic potential of neonatal chondrocytes with human bone marrow stromal cells (HBMSCs) and adult chondrocytes. {yields} Demonstrated that adult chondrocytes hold greatest potential for use in ACI based on their higher proliferation rates, lower alkaline phosphatise activity and enhanced expression of chondrogenic genes. {yields} Demonstrated the need for chondroinduction as a necessary pre-requisite to efficient chondrogenesis in vitro and, by extrapolation, for cell based therapy (e.g. ACI or cartilage tissue engineering). -- Abstract: Cartilage tissue engineering is still a major clinical challenge with optimisation of a suitable source of cells for cartilage repair/regeneration not yet fully addressed. The aims of this study were to compare and contrast the differences in chondrogenic behaviour between human bone marrow stromal cells (HBMSCs), human neonatal and adult chondrocytes to further our understanding of chondroinduction relative to cell maturity and to identify factors that promote chondrogenesis and maintain functional homoeostasis. Cells were cultured in monolayer in either chondrogenic or basal medium, recapitulating procedures used in existing clinical procedures for cell-based therapies. Cell doubling time, morphology and alkaline phosphatase specific activity (ALPSA) were determined at different time points. Expression of chondrogenic markers (SOX9, ACAN and COL2A1) was compared via real time polymerase chain reaction. Amongst the three cell types studied, HBMSCs had the highest ALPSA in basal culture and lowest ALPSA in chondrogenic media. Neonatal chondrocytes were the most proliferative and adult chondrocytes had the lowest ALPSA in basal media. Gene expression analysis revealed

  4. Cartilage tumors. Pathology and radiomorphology; Chondrogene Knochentumoren. Pathologie und Radiomorphologie

    Energy Technology Data Exchange (ETDEWEB)

    Uhl, M. [RKK-Klinikum Freiburg, Klinik fuer Diagnostische und Interventionelle Radiologie, Kinderradiologie und Neuroradiologie SJK, Freiburg (Germany); Herget, G. [Universitaetsklinik Freiburg, Department Orthopaedie und Traumatologie, Freiburg (Germany); Kurz, P. [Universitaetsklinik Freiburg, Pathologisches Institut, Freiburg (Germany)

    2016-06-15

    Primary cartilage-forming tumors of the bone are frequent entities in the daily work of skeletal radiologists. This article describes the correlation of pathology and radiology in cartilage-forming skeletal tumors, in particular, enchondroma, osteochondroma, periosteal chondromas, chondroblastoma and various forms of chondrosarcoma. After reading, the radiologist should be able to deduce the different patterns of cartilage tumors on radiographs, CT, and MRI from the pathological aspects. Differentiation of enchondroma and chondrosarcoma is a frequent diagnostic challenge. Some imaging parameters, e. g., deep cortical scalloping (more than two thirds of the cortical thickness), cortical destruction, or a soft-tissue mass, are features of a sarcoma. Osteochondromas are bony protrusions with a continuous extension of bone marrow from the parent bone, the host cortical bone runs continuously from the osseous surface of the tumor into the shaft of the osteochondroma and the osteochondroma has a cartilage cap. Chondromyxoid fibromas are well-defined lytic and eccentric lesions of the metaphysis of the long bones, with nonspecific MRI findings. Chondroblastomas have a strong predilection for the epiphysis of long tubular bones and develop an intense perifocal bone marrow edema. Dedifferentiated chondrosarcomas are bimorphic lesions with a low-grade chondrogenic component and a high-grade noncartilaginous component. Most chondrogenic tumors have a predilection with regard to site and age at manifestation. (orig.) [German] Primaere knorpelbildende Tumoren sind haeufige Entitaeten in der taeglichen Arbeit des Radiologen. Der Beitrag beschreibt die Korrelation von Pathologie und Radiologie knorpelbildender Skeletttumoren, insbesondere von Enchondrom, Osteochondrom, periostalem Chondrom, Chondroblastom, und verschiedenen Varianten des Chondrosarkoms. Nach Lesen des Beitrags kann der Radiologe die verschiedenen typischen Muster knorpelbildender Tumoren im Roentgenbild

  5. An Enhanced Differential Evolution with Elite Chaotic Local Search

    Directory of Open Access Journals (Sweden)

    Zhaolu Guo

    2015-01-01

    Full Text Available Differential evolution (DE is a simple yet efficient evolutionary algorithm for real-world engineering problems. However, its search ability should be further enhanced to obtain better solutions when DE is applied to solve complex optimization problems. This paper presents an enhanced differential evolution with elite chaotic local search (DEECL. In DEECL, it utilizes a chaotic search strategy based on the heuristic information from the elite individuals to promote the exploitation power. Moreover, DEECL employs a simple and effective parameter adaptation mechanism to enhance the robustness. Experiments are conducted on a set of classical test functions. The experimental results show that DEECL is very competitive on the majority of the test functions.

  6. Significance of soluble growth factors in the chondrogenic response of human umbilical cord matrix stem cells in a porous three dimensional scaffold

    Directory of Open Access Journals (Sweden)

    RS Nirmal

    2013-11-01

    Full Text Available Stem cell based tissue engineering has emerged as a promising strategy for articular cartilage regeneration. Foetal derived mesenchymal stem cells (MSCs with their ease of availability, pluripotency and high expansion potential have been demonstrated to be an attractive cell source over adult MSCs. However, there is a need for optimisation of chondrogenic signals to direct the differentiation of these multipotent MSCs to chondrogenic lineage. In this study we have demonstrated the in vitro chondrogenesis of human umbilical cord matrix MSCs in three dimensional PVA-PCL (polyvinyl alcohol-polycaprolactone scaffolds in the presence of the individual growth factors TGFβ1, TGFβ3, IGF, BMP2 and their combination with BMP2. Gene expression, histology and immunohistology were evaluated after 28 d culture. The induced cells showed the feature of chondrocytes in their morphology and expression of typical chondrogenic extracellular matrix molecules. Moreover, the real-time PCR assay has shown the expression of gene markers of chondrogenesis, SOX9, collagen type II and aggrecan. The expression of collagen type I and collagen type X was also evaluated. This study has demonstrated the successful chondrogenic induction of human umbilical cord MSCs in 3D scaffolds. Interestingly, the growth factor combination of TGF-β3 and BMP-2 was found to be more effective for chondrogenesis as shown by the real-time PCR studies. The findings of this study suggest the importance of using growth factor combinations for successful chondrogenic differentiation of umbilical cord MSCs.

  7. γ-Oryzanol Enhances Adipocyte Differentiation and Glucose Uptake

    Directory of Open Access Journals (Sweden)

    Chang Hwa Jung

    2015-06-01

    Full Text Available Recent studies show that brown rice improves glucose intolerance and potentially the risk of diabetes, although the underlying molecular mechanisms remain unclear. One of the phytochemicals found in high concentration in brown rice is γ-oryzanol (Orz, a group of ferulic acid esters of phytosterols and triterpene alcohols. Here, we found that Orz stimulated differentiation of 3T3-L1 preadipocytes and increased the protein expression of adipogenic marker genes such as peroxisome proliferator-activated receptor gamma (PPAR-γ and CCAAT/enhanced binding protein alpha (C/EBPα. Moreover, Orz significantly increased the glucose uptake in insulin-resistant cells and translocation of glucose transporter type 4 (GLUT4 from the cytosol to the cell surface. To investigate the mechanism by which Orz stimulated cell differentiation, we examined its effects on cellular signaling of the mammalian target of rapamycin complex 1 (mTORC1, a central mediator of cellular growth and proliferation. The Orz treatment increased mTORC1 kinase activity based on phosphorylation of 70-kDa ribosomal S6 kinase 1 (S6K1. The effect of Orz on adipocyte differentiation was dependent on mTORC1 activity because rapamycin blocks cell differentiation in Orz-treated cells. Collectively, our results indicate that Orz stimulates adipocyte differentiation, enhances glucose uptake, and may be associated with cellular signaling mediated by PPAR-γ and mTORC1.

  8. γ-Oryzanol Enhances Adipocyte Differentiation and Glucose Uptake.

    Science.gov (United States)

    Jung, Chang Hwa; Lee, Da-Hye; Ahn, Jiyun; Lee, Hyunjung; Choi, Won Hee; Jang, Young Jin; Ha, Tae-Youl

    2015-06-15

    Recent studies show that brown rice improves glucose intolerance and potentially the risk of diabetes, although the underlying molecular mechanisms remain unclear. One of the phytochemicals found in high concentration in brown rice is γ-oryzanol (Orz), a group of ferulic acid esters of phytosterols and triterpene alcohols. Here, we found that Orz stimulated differentiation of 3T3-L1 preadipocytes and increased the protein expression of adipogenic marker genes such as peroxisome proliferator-activated receptor gamma (PPAR-γ) and CCAAT/enhanced binding protein alpha (C/EBPα). Moreover, Orz significantly increased the glucose uptake in insulin-resistant cells and translocation of glucose transporter type 4 (GLUT4) from the cytosol to the cell surface. To investigate the mechanism by which Orz stimulated cell differentiation, we examined its effects on cellular signaling of the mammalian target of rapamycin complex 1 (mTORC1), a central mediator of cellular growth and proliferation. The Orz treatment increased mTORC1 kinase activity based on phosphorylation of 70-kDa ribosomal S6 kinase 1 (S6K1). The effect of Orz on adipocyte differentiation was dependent on mTORC1 activity because rapamycin blocks cell differentiation in Orz-treated cells. Collectively, our results indicate that Orz stimulates adipocyte differentiation, enhances glucose uptake, and may be associated with cellular signaling mediated by PPAR-γ and mTORC1.

  9. Different Chondrogenic Potential among Human Induced Pluripotent Stem Cells from Diverse Origin Primary Cells

    Directory of Open Access Journals (Sweden)

    Yeri Alice Rim

    2018-01-01

    Full Text Available Scientists have tried to reprogram various origins of primary cells into human induced pluripotent stem cells (hiPSCs. Every somatic cell can theoretically become a hiPSC and give rise to targeted cells of the human body. However, there have been debates on the controversy about the differentiation propensity according to the origin of primary cells. We reprogrammed hiPSCs from four different types of primary cells such as dermal fibroblasts (DF, n=3, peripheral blood mononuclear cells (PBMC, n=3, cord blood mononuclear cells (CBMC, n=3, and osteoarthritis fibroblast-like synoviocytes (OAFLS, n=3. Established hiPSCs were differentiated into chondrogenic pellets. All told, cartilage-specific markers tended to express more by the order of CBMC > DF > PBMC > FLS. Origin of primary cells may influence the reprogramming and differentiation thereafter. In the context of chondrogenic propensity, CBMC-derived hiPSCs can be a fairly good candidate cell source for cartilage regeneration. The differentiation of hiPSCs into chondrocytes may help develop “cartilage in a dish” in the future. Also, the ideal cell source of hiPSC for chondrogenesis may contribute to future application as well.

  10. Ductile electroactive biodegradable hyperbranched polylactide copolymers enhancing myoblast differentiation.

    Science.gov (United States)

    Xie, Meihua; Wang, Ling; Guo, Baolin; Wang, Zhong; Chen, Y Eugene; Ma, Peter X

    2015-12-01

    Myotube formation is crucial to restoring muscular functions, and biomaterials that enhance the myoblast differentiation into myotubes are highly desirable for muscular repair. Here, we report the synthesis of electroactive, ductile, and degradable copolymers and their application in enhancing the differentiation of myoblasts to myotubes. A hyperbranched ductile polylactide (HPLA) was synthesized and then copolymerized with aniline tetramer (AT) to produce a series of electroactive, ductile and degradable copolymers (HPLAAT). The HPLA and HPLAAT showed excellent ductility with strain to failure from 158.9% to 42.7% and modulus from 265.2 to 758.2 MPa. The high electroactivity of the HPLAAT was confirmed by UV spectrometer and cyclic voltammogram measurements. These HPLAAT polymers also showed improved thermal stability and controlled biodegradation rate compared to HPLA. Importantly, when applying these polymers for myotube formation, the HPLAAT significantly improved the proliferation of C2C12 myoblasts in vitro compared to HPLA. Furthermore, these polymers greatly promoted myogenic differentiation of C2C12 cells as measured by quantitative analysis of myotube number, length, diameter, maturation index, and gene expression of MyoD and TNNT. Together, our study shows that these electroactive, ductile and degradable HPLAAT copolymers represent significantly improved biomaterials for muscle tissue engineering compared to HPLA. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Silk fibroin/gelatin–chondroitin sulfate–hyaluronic acid effectively enhances in vitro chondrogenesis of bone marrow mesenchymal stem cells

    International Nuclear Information System (INIS)

    Sawatjui, Nopporn; Damrongrungruang, Teerasak; Leeanansaksiri, Wilairat; Jearanaikoon, Patcharee; Hongeng, Suradej; Limpaiboon, Temduang

    2015-01-01

    Tissue engineering is becoming promising for cartilage repair due to the limited self-repair capacity of cartilage tissue. We previously fabricated and characterized a three-dimensional silk fibroin/gelatin–chondroitin sulfate–hyaluronic acid (SF–GCH) scaffold and showed that it could promote proliferation of human bone marrow mesenchymal stem cells (BM-MSCs). This study aimed to evaluate its biological performance as a new biomimetic material for chondrogenic induction of BM-MSCs in comparison to an SF scaffold and conventional pellet culture. We found that the SF–GCH scaffold significantly enhanced the proliferation and chondrogenic differentiation of BM-MSCs compared to the SF scaffold and pellet culture in which the production of sulfated glycoaminoglycan was increased in concordance with the up-regulation of chondrogenic-specific gene markers. Our findings indicate the significant role of SF–GCH by providing a supportive structure and the mimetic cartilage environment for chondrogenesis which enables cartilage regeneration. Thus, our fabricated SF–GCH scaffold may serve as a potential biomimetic material for cartilage tissue engineering. - Highlights: • SF–GCH scaffold enhances proliferation and chondrogenic differentiation of BM-MSCs. • SF–GCH acts as a supportive and biomimetic material for BM-MSC chondrogenesis. • SF–GCH is a potential biomimetic scaffold suitable for cartilage tissue engineering

  12. Silk fibroin/gelatin–chondroitin sulfate–hyaluronic acid effectively enhances in vitro chondrogenesis of bone marrow mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Sawatjui, Nopporn [Biomedical Sciences, Graduate School, Khon Kaen University, Khon Kaen 40002 (Thailand); Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen 40002 (Thailand); Damrongrungruang, Teerasak [Department of Oral Diagnosis, Faculty of Dentistry, Khon Kaen University, Khon Kaen 40002 (Thailand); Leeanansaksiri, Wilairat [Stem Cell Therapy and Transplantation Research Group, Suranaree University of Technology, Nakhon Ratchasima 30000 (Thailand); School of Microbiology, Suranaree University of Technology, Nakhon Ratchasima 30000 (Thailand); Jearanaikoon, Patcharee [Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen 40002 (Thailand); Hongeng, Suradej [Department of Pediatrics, Faculty of Medicine, Ramathibodi Hospital, Mahidol University, Bangkok 10400 (Thailand); Limpaiboon, Temduang, E-mail: temduang@kku.ac.th [Centre for Research and Development of Medical Diagnostic Laboratories, Faculty of Associated Medical Sciences, Khon Kaen University, Khon Kaen 40002 (Thailand)

    2015-07-01

    Tissue engineering is becoming promising for cartilage repair due to the limited self-repair capacity of cartilage tissue. We previously fabricated and characterized a three-dimensional silk fibroin/gelatin–chondroitin sulfate–hyaluronic acid (SF–GCH) scaffold and showed that it could promote proliferation of human bone marrow mesenchymal stem cells (BM-MSCs). This study aimed to evaluate its biological performance as a new biomimetic material for chondrogenic induction of BM-MSCs in comparison to an SF scaffold and conventional pellet culture. We found that the SF–GCH scaffold significantly enhanced the proliferation and chondrogenic differentiation of BM-MSCs compared to the SF scaffold and pellet culture in which the production of sulfated glycoaminoglycan was increased in concordance with the up-regulation of chondrogenic-specific gene markers. Our findings indicate the significant role of SF–GCH by providing a supportive structure and the mimetic cartilage environment for chondrogenesis which enables cartilage regeneration. Thus, our fabricated SF–GCH scaffold may serve as a potential biomimetic material for cartilage tissue engineering. - Highlights: • SF–GCH scaffold enhances proliferation and chondrogenic differentiation of BM-MSCs. • SF–GCH acts as a supportive and biomimetic material for BM-MSC chondrogenesis. • SF–GCH is a potential biomimetic scaffold suitable for cartilage tissue engineering.

  13. Bee venom enhances the differentiation of human regulatory T cells.

    Science.gov (United States)

    Caramalho, I; Melo, A; Pedro, E; Barbosa, M M P; Victorino, R M M; Pereira Santos, M C; Sousa, A E

    2015-10-01

    Venom-specific immunotherapy (VIT) is well recognized by its efficacy, and compelling evidence implicates regulatory T cells (Tregs) in the underlying tolerogenic mechanisms. Additionally, hymenoptera venom has for a long time been claimed to modulate immunity. Here, we investigated the putative role of bee venom (Bv) in human FOXP3-expressing Treg homeostasis and differentiation, irrespective of the donors' allergic status. We found that Bv significantly enhanced the differentiation of FOXP3-expressing cells both from conventional naïve CD4 T cells and mature CD4 thymocytes, a property that may contribute to the VIT's capacity to expand circulating Tregs in allergic individuals. We expect that our data enlightening the Treg-mediated immunomodulatory properties of Bv regardless of TCR specificity, to have application in other allergies, as well as in other clinical settings, such as autoimmunity and transplantation. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Mechanism involved in enhancement of osteoblast differentiation by hyaluronic acid

    International Nuclear Information System (INIS)

    Kawano, Michinao; Ariyoshi, Wataru; Iwanaga, Kenjiro; Okinaga, Toshinori; Habu, Manabu; Yoshioka, Izumi; Tominaga, Kazuhiro; Nishihara, Tatsuji

    2011-01-01

    Research highlights: → In this study was to investigate the effects of HA on osteoblast differentiation induced by BMP-2. → MG63 cells were incubated with BMP-2 and HA for various time periods. → Phosphorylation of Smad 1/5/8, p38, and ERK proteins was determined by western blot analysis. To elucidate the nuclear translocation of phosphorylated Smad 1/5/8, stimulated cells were subjected to immunofluorescence microscopy. → HA enhanced BMP-2 induces osteoblastic differentiation in MG63 cells via down-regulation of BMP-2 antagonists and ERK phosphorylation. -- Abstract: Objectives: Bone morphogenetic protein-2 (BMP-2) is expected to be utilized to fill bone defects and promote healing of fractures. However, it is unable to generate an adequate clinical response for use in bone regeneration. Recently, it was reported that glycosaminoglycans, including heparin, heparan sulfate, keratan sulfate, dermatan sulfate, chondroitin-4-sulfate, chondroitin-6-sulfate, and hyaluronic acid (HA), regulate BMP-2 activity, though the mechanism by which HA regulates osteogenic activities has not been fully elucidated. The aim of this study was to investigate the effects of HA on osteoblast differentiation induced by BMP-2. Materials and methods: Monolayer cultures of osteoblastic lineage MG63 cells were incubated with BMP-2 and HA for various time periods. To determine osteoblastic differentiation, alkaline phosphatase (ALP) activity in the cell lysates was quantified. Phosphorylation of Smad 1/5/8, p38, and ERK proteins was determined by Western blot analysis. To elucidate the nuclear translocation of phosphorylated Smad 1/5/8, stimulated cells were subjected to immunofluorescence microscopy. To further elucidate the role of HA in enhancement of BMP-2-induced Smad signaling, mRNA expressions of the BMP-2 receptor antagonists noggin and follistatin were detected using real-time RT-PCR. Results: BMP-2-induced ALP activation, Smad 1/5/8 phosphorylation, and nuclear translocation

  15. Mechanism involved in enhancement of osteoblast differentiation by hyaluronic acid

    Energy Technology Data Exchange (ETDEWEB)

    Kawano, Michinao [Division of Maxillofacial Diagnostic and Surgical Science, Department of Oral and Maxillofacial Surgery, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Ariyoshi, Wataru [Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Iwanaga, Kenjiro [Division of Maxillofacial Diagnostic and Surgical Science, Department of Oral and Maxillofacial Surgery, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Okinaga, Toshinori [Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Habu, Manabu [Division of Maxillofacial Diagnostic and Surgical Science, Department of Oral and Maxillofacial Surgery, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Yoshioka, Izumi [Division of Oral and Maxillofacial Surgery, Department of Medicine of Sensory and Motor Organs, University of Miyazaki, Kiyotake, Miyazaki 889-1692 (Japan); Tominaga, Kazuhiro [Division of Maxillofacial Diagnostic and Surgical Science, Department of Oral and Maxillofacial Surgery, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Oral Bioresearch Center, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Nishihara, Tatsuji, E-mail: tatsujin@kyu-dent.ac.jp [Division of Infections and Molecular Biology, Department of Health Promotion, Kyushu Dental College, Kitakyushu 803-8580 (Japan); Oral Bioresearch Center, Kyushu Dental College, Kitakyushu 803-8580 (Japan)

    2011-02-25

    Research highlights: {yields} In this study was to investigate the effects of HA on osteoblast differentiation induced by BMP-2. {yields} MG63 cells were incubated with BMP-2 and HA for various time periods. {yields} Phosphorylation of Smad 1/5/8, p38, and ERK proteins was determined by western blot analysis. To elucidate the nuclear translocation of phosphorylated Smad 1/5/8, stimulated cells were subjected to immunofluorescence microscopy. {yields} HA enhanced BMP-2 induces osteoblastic differentiation in MG63 cells via down-regulation of BMP-2 antagonists and ERK phosphorylation. -- Abstract: Objectives: Bone morphogenetic protein-2 (BMP-2) is expected to be utilized to fill bone defects and promote healing of fractures. However, it is unable to generate an adequate clinical response for use in bone regeneration. Recently, it was reported that glycosaminoglycans, including heparin, heparan sulfate, keratan sulfate, dermatan sulfate, chondroitin-4-sulfate, chondroitin-6-sulfate, and hyaluronic acid (HA), regulate BMP-2 activity, though the mechanism by which HA regulates osteogenic activities has not been fully elucidated. The aim of this study was to investigate the effects of HA on osteoblast differentiation induced by BMP-2. Materials and methods: Monolayer cultures of osteoblastic lineage MG63 cells were incubated with BMP-2 and HA for various time periods. To determine osteoblastic differentiation, alkaline phosphatase (ALP) activity in the cell lysates was quantified. Phosphorylation of Smad 1/5/8, p38, and ERK proteins was determined by Western blot analysis. To elucidate the nuclear translocation of phosphorylated Smad 1/5/8, stimulated cells were subjected to immunofluorescence microscopy. To further elucidate the role of HA in enhancement of BMP-2-induced Smad signaling, mRNA expressions of the BMP-2 receptor antagonists noggin and follistatin were detected using real-time RT-PCR. Results: BMP-2-induced ALP activation, Smad 1/5/8 phosphorylation, and

  16. The assessment of natural scaffolds ability in chondrogenic ...

    African Journals Online (AJOL)

    Arun Kumar Agnihotri

    1Stem Cell Laboratory, The Academic Center for Education, Culture and Research, ... between alginate and agarose groups in maintaining cells viable but, about chondrogenic .... Absorbance at 570 nm was measured on a ... as small cells with little cytoplasm and elliptic ... high density of cells and suitable interaction.

  17. Growth factor combination for chondrogenic induction from human mesenchymal stem cell

    International Nuclear Information System (INIS)

    Indrawattana, Nitaya; Chen Guoping; Tadokoro, Mika; Shann, Linzi H.; Ohgushi, Hajime; Tateishi, Tetsuya; Tanaka, Junzo; Bunyaratvej, Ahnond

    2004-01-01

    During the last decade, many strategies for cartilage engineering have been emerging. Stem cell induction is one of the possible approaches for cartilage engineering. The mesenchymal stem cells (MSCs) with their pluripotency and availability have been demonstrated to be an attractive cell source. It needs the stimulation with cell growth factors to make the multipluripotent MSCs differentiate into chondrogenic lineage. We have shown particular patterns of in vitro chondrogenesis induction on human bone marrow MSCs (hBMSCs) by cycling the growth factors. The pellet cultures of hBMSCs were prepared for chondrogenic induction. Growth factors: TGF-β3, BMP-6, and IGF-1 were used in combination for cell induction. Gene expression, histology, immunohistology, and real-time PCR methods were measured on days 21 after cell induction. As shown by histology and immunohistology, the induced cells have shown the feature of chondrocytes in their morphology and extracellular matrix in both inducing patterns of combination and cycling induction. Moreover, the real-time PCR assay has shown the expression of gene markers of chondrogenesis, collagen type II and aggrecan. This study has demonstrated that cartilage tissue can be created from bone marrow mesenchymal stem cells. Interestingly, the combined growth factors TGF-β3 and BMP-6 or TGF-β3 and IGF-1 were more effective for chondrogenesis induction as shown by the real-time PCR assay. The combination of these growth factors may be the important key for in vitro chondrogenesis induction

  18. GRESS, FORTRAN Pre-compiler with Differentiation Enhancement

    International Nuclear Information System (INIS)

    1999-01-01

    1 - Description of program or function: The GRESS FORTRAN pre-compiler (SYMG) and run-time library are used to enhance conventional FORTRAN-77 programs with analytic differentiation of arithmetic statements for automatic differentiation in either forward or reverse mode. GRESS 3.0 is functionally equivalent to GRESS 2.1. GRESS 2.1 is an improved and updated version of the previous released GRESS 1.1. Improvements in the implementation of a the CHAIN option have resulted in a 70 to 85% reduction in execution time and up to a 50% reduction in memory required for forward chaining applications. 2 - Method of solution: GRESS uses a pre-compiler to analyze FORTRAN statements and determine the mathematical operations embodied in them. As each arithmetic assignment statement in a program is analyzed, SYMG generates the partial derivatives of the term on the left with respect to each floating-point variable on the right. The result of the pre-compilation step is a new FORTRAN program that can produce derivatives for any REAL (i.e., single or double precision) variable calculated by the model. Consequently, GRESS enhances FORTRAN programs or subprograms by adding the calculation of derivatives along with the original output. Derivatives from a GRESS enhanced model can be used internally (e.g., iteration acceleration) or externally (e.g., sensitivity studies). By calling GRESS run-time routines, derivatives can be propagated through the code via the chain rule (referred to as the CHAIN option) or accumulated to create an adjoint matrix (referred to as the ADGEN option). A third option, GENSUB, makes it possible to process a subset of a program (i.e., a do loop, subroutine, function, a sequence of subroutines, or a whole program) for calculating derivatives of dependent variables with respect to independent variables. A code enhanced with the GENSUB option can use forward mode, reverse mode, or a hybrid of the two modes. 3 - Restrictions on the complexity of the problem: GRESS

  19. Chondrogenic potential of human mesenchymal stem cells and expression of Slug transcription factor.

    Science.gov (United States)

    Brini, Anna T; Niada, Stefania; Lambertini, Elisabetta; Torreggiani, Elena; Arrigoni, Elena; Lisignoli, Gina; Piva, Roberta

    2015-06-01

    The scientific literature rarely reports experimental failures or inconsistent outcomes in the induction of cell differentiation; however, researchers commonly experience poor or unsuccessful responses to differentiating agents when culturing stem cells. One way of investigating the underlying reasons for such responses is to look at the basal expression levels of specific genes in multipotent stem cells before the induction of differentiation. In addition to shedding light on the complex properties of stem cells and the molecular modulation of differentiation pathways, this strategy can also lead to the development of important time- and money-saving tools that aid the efficient selection of cellular specimens--in this case, stem cells that are more prone to differentiate towards specific lineages and are therefore more suitable for cell-based therapeutic protocols in regenerative medicine. To address this latter aspect, this study focused on understanding the reasons why some human mesenchymal stem cell (hMSC) samples are less efficient at differentiating towards chondrogenesis. This study shows that analysis of the basal expression levels of Slug, a negative regulator of chondrogenesis in hMSC, provides a rapid and simple tool for distinguishing stem cell samples with the potential to form a cartilage-like matrix, and that are therefore suitable for cartilage tissue engineering. It is shown that high basal levels of Slug prevent the chondrogenic differentiation of hMSCs, even in the presence of transforming growth factor-β and elevated levels of Sox9. Copyright © 2013 John Wiley & Sons, Ltd.

  20. TGF-ß1 enhances the BMP-2-induced chondrogenesis of bovine synovial explants and arrests downstream differentiation at an early stage of hypertrophy.

    Science.gov (United States)

    Shintani, Nahoko; Siebenrock, Klaus A; Hunziker, Ernst B

    2013-01-01

    Synovial explants furnish an in-situ population of mesenchymal stem cells for the repair of articular cartilage. Although bone morphogenetic protein 2 (BMP-2) induces the chondrogenesis of bovine synovial explants, the cartilage formed is neither homogeneously distributed nor of an exclusively hyaline type. Furthermore, the downstream differentiation of chondrocytes proceeds to the stage of terminal hypertrophy, which is inextricably coupled with undesired matrix mineralization. With a view to optimizing BMP-2-induced chondrogenesis, the modulating influences of fibroblast growth factor 2 (FGF-2) and transforming growth factor beta 1 (TGF-ß1) were investigated. Explants of bovine calf metacarpal synovium were exposed to BMP-2 (200 ng/ml) for 4 (or 6) weeks. FGF-2 (10 ng/ml) or TGF-ß1 (10 ng/ml) was introduced at the onset of incubation and was present either during the first week of culturing alone or throughout its entire course. FGF-2 enhanced the BMP-2-induced increase in metachromatic staining for glycosaminoglycans (GAGs) only when it was present during the first week of culturing alone. TGF-ß1 enhanced not only the BMP-2-induced increase in metachromasia (to a greater degree than FGF-2), but also the biochemically-assayed accumulation of GAGs, when it was present throughout the entire culturing period; in addition, it arrested the downstream differentiation of cells at an early stage of hypertrophy. These findings were corroborated by an analysis of the gene- and protein-expression levels of key cartilaginous markers and by an estimation of individual cell volume. TGF-ß1 enhances the BMP-2-induced chondrogenesis of bovine synovial explants, improves the hyaline-like properties of the neocartilage, and arrests the downstream differentiation of cells at an early stage of hypertrophy. With the prospect of engineering a mature, truly articular type of cartilage in the context of clinical repair, our findings will be of importance in fine-tuning the

  1. TGF-ß1 enhances the BMP-2-induced chondrogenesis of bovine synovial explants and arrests downstream differentiation at an early stage of hypertrophy.

    Directory of Open Access Journals (Sweden)

    Nahoko Shintani

    Full Text Available Synovial explants furnish an in-situ population of mesenchymal stem cells for the repair of articular cartilage. Although bone morphogenetic protein 2 (BMP-2 induces the chondrogenesis of bovine synovial explants, the cartilage formed is neither homogeneously distributed nor of an exclusively hyaline type. Furthermore, the downstream differentiation of chondrocytes proceeds to the stage of terminal hypertrophy, which is inextricably coupled with undesired matrix mineralization. With a view to optimizing BMP-2-induced chondrogenesis, the modulating influences of fibroblast growth factor 2 (FGF-2 and transforming growth factor beta 1 (TGF-ß1 were investigated.Explants of bovine calf metacarpal synovium were exposed to BMP-2 (200 ng/ml for 4 (or 6 weeks. FGF-2 (10 ng/ml or TGF-ß1 (10 ng/ml was introduced at the onset of incubation and was present either during the first week of culturing alone or throughout its entire course. FGF-2 enhanced the BMP-2-induced increase in metachromatic staining for glycosaminoglycans (GAGs only when it was present during the first week of culturing alone. TGF-ß1 enhanced not only the BMP-2-induced increase in metachromasia (to a greater degree than FGF-2, but also the biochemically-assayed accumulation of GAGs, when it was present throughout the entire culturing period; in addition, it arrested the downstream differentiation of cells at an early stage of hypertrophy. These findings were corroborated by an analysis of the gene- and protein-expression levels of key cartilaginous markers and by an estimation of individual cell volume.TGF-ß1 enhances the BMP-2-induced chondrogenesis of bovine synovial explants, improves the hyaline-like properties of the neocartilage, and arrests the downstream differentiation of cells at an early stage of hypertrophy. With the prospect of engineering a mature, truly articular type of cartilage in the context of clinical repair, our findings will be of importance in fine-tuning the

  2. Performance enhancement of implantable medical antenna using differential feed technique

    Directory of Open Access Journals (Sweden)

    Shankar Bhattacharjee

    2016-03-01

    Full Text Available The health care industry is continuously revolutionizing and advancing towards developing more efficient system suitable for human body. Today implantable devices have become a more interesting topic in health care services which primarily started with the pacemakers. Since then it is continuously evolving due to its non-invasive nature, instant monitoring and diagnosis, and periodic simulation. The main goal of these implantable devices is to efficiently monitor or inspect various ailments in the body and then transmits this to the server or base station. For proper communication between the implant and the base station, antenna design is of prime importance. In this paper MEMS based differentially fed dual band antenna has been proposed and can be used both in Medical Implant Communication Service (MICS band for transmission of data and industrial, scientific and medical (ISM band for wake-up purpose. The proposed antenna has been simulated for free space scenario and has been found to radiate in both MICS & ISM band with S11 of −17.62 dB and −14.31 dB respectively. Subsequently the antenna is inserted within a skin mimicking model with equivalent dielectric features and the results show variation in radiation characteristics between free space condition and within skin phantom. The design of the antenna has been optimized in such a way that minimum deviation occurs between the two conformal conditions. With the use of differential feeding technique performance of the antenna is quite enhanced in terms of various parameters when compared with single feed.

  3. Purinergic responses of chondrogenic stem cells to dynamic loading

    Directory of Open Access Journals (Sweden)

    Gađanski Ivana

    2013-01-01

    Full Text Available In habitually loaded tissues, dynamic loading can trigger ATP (adenosine 5’- triphosphate release to extracellular environment, and result in calcium signaling via ATP binding to purine P2 receptors1. In the current study we have compared purinergic responses (ATP release of two types of cells: bovine chondrocytes (bCHs and human mesenchymal stem cells (hMSC that were encapsulated in agarose and subjected to dynamic loading. Both cell types were cultured under chondrogenic conditions, and their responses to loading were evaluated by ATP release assay in combination with connexin (Cx-sensitive fluorescent dye (Lucifer Yellow - LY and a Cx-hemichannel blocker (Flufenamic acid - FFA. In response to dynamic loading, chondrogenic hMSCs released significantly higher amounts of ATP (5-fold in comparison to the bCHs early in culture (day 2. Triggering of LY uptake in the bCHs and hMSCs by dynamic loading implies opening of the Cx-hemichannels. However, the number of LY-positive cells in hMSC-constructs was 2.5-fold lower compared to the loaded bCH-constructs, suggesting utilization of additional mechanisms of ATP release. Cx-reactive sites were detected in both bCHs and hMSCs-constructs. FFA application led to reduced ATP release both in bCHs and hMSCs, which confirms the involvement of connexin hemichannels, with more prominent effects in bCHs than in hMSCs, further implying the existence of additional mechanism of ATP release in chondrogenic hMSCs. Taken together, these results indicate stronger purinergic response to dynamic loading of chondrogenic hMSCs than primary chondrocytes, by activation of connexin hemichannels and additional mechanisms of ATP release. [Projekat Ministrastva nauke Republike Srbije, ON174028 i br. III41007

  4. Differential evolution enhanced with multiobjective sorting-based mutation operators.

    Science.gov (United States)

    Wang, Jiahai; Liao, Jianjun; Zhou, Ying; Cai, Yiqiao

    2014-12-01

    Differential evolution (DE) is a simple and powerful population-based evolutionary algorithm. The salient feature of DE lies in its mutation mechanism. Generally, the parents in the mutation operator of DE are randomly selected from the population. Hence, all vectors are equally likely to be selected as parents without selective pressure at all. Additionally, the diversity information is always ignored. In order to fully exploit the fitness and diversity information of the population, this paper presents a DE framework with multiobjective sorting-based mutation operator. In the proposed mutation operator, individuals in the current population are firstly sorted according to their fitness and diversity contribution by nondominated sorting. Then parents in the mutation operators are proportionally selected according to their rankings based on fitness and diversity, thus, the promising individuals with better fitness and diversity have more opportunity to be selected as parents. Since fitness and diversity information is simultaneously considered for parent selection, a good balance between exploration and exploitation can be achieved. The proposed operator is applied to original DE algorithms, as well as several advanced DE variants. Experimental results on 48 benchmark functions and 12 real-world application problems show that the proposed operator is an effective approach to enhance the performance of most DE algorithms studied.

  5. In-vivo generation of bone via endochondral ossification by in-vitro chondrogenic priming of adult human and rat mesenchymal stem cells

    LENUS (Irish Health Repository)

    Farrell, Eric

    2011-01-31

    -glycerophosphate to the chondrogenic medium did not hamper this process. Using transgenic animals we also demonstrated that both host and donor cells played a role in bone formation. In conclusion these data indicate that in-vitro chondrogenic differentiation of human MSCs could lead to an alternative and superior approach for bone tissue engineering.

  6. Actin depolymerization enhances adipogenic differentiation in human stromal stem cells

    DEFF Research Database (Denmark)

    Chen, Li; Hu, Huimin; Qiu, Weimin

    2018-01-01

    Human stromal stem cells (hMSCs) differentiate into adipocytes that play a role in skeletal tissue homeostasis and whole body energy metabolism. During adipocyte differentiation, hMSCs exhibit significant changes in cell morphology suggesting changes in cytoskeletal organization. Here, we examined...... the effect of direct modulation of actin microfilament dynamics on adipocyte differentiation. Stabilizing actin filaments in hMSCs by siRNA-mediated knock down of the two main actin depolymerizing factors (ADFs): Cofilin 1 (CFL1) and Destrin (DSTN) or treating the cells by Phalloidin reduced adipocyte...

  7. Mesenchymal stem cells maintain TGF-beta-mediated chondrogenic phenotype in alginate bead culture

    DEFF Research Database (Denmark)

    Mehlhorn, A T; Schmal, H; Kaiser, S

    2006-01-01

    cultured in osteogenic medium after TGF-beta-mediated chondroinduction. Gene expression of col2a1, aggrecan, COMP, alkaline phosphatase (AP), and correlating protein synthesis was analyzed. After short-term stimulation with TGF-beta, MSCs maintained a chondrogenic phenotype. Chondrogenic gene expression...

  8. Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) Pathway Is Induced by Mechanical Load and Reduces the Activity of Hedgehog Signaling in Chondrogenic Micromass Cell Cultures

    Science.gov (United States)

    Juhász, Tamás; Szentléleky, Eszter; Szűcs Somogyi, Csilla; Takács, Roland; Dobrosi, Nóra; Engler, Máté; Tamás, Andrea; Reglődi, Dóra; Zákány, Róza

    2015-01-01

    Pituitary adenylate cyclase activating polypeptide (PACAP) is a neurohormone exerting protective function during various stress conditions either in mature or developing tissues. Previously we proved the presence of PACAP signaling elements in chicken limb bud-derived chondrogenic cells in micromass cell cultures. Since no data can be found if PACAP signaling is playing any role during mechanical stress in any tissues, we aimed to investigate its contribution in mechanotransduction during chondrogenesis. Expressions of the mRNAs of PACAP and its major receptor, PAC1 increased, while that of other receptors, VPAC1, VPAC2 decreased upon mechanical stimulus. Mechanical load enhanced the expression of collagen type X, a marker of hypertrophic differentiation of chondrocytes and PACAP addition attenuated this elevation. Moreover, exogenous PACAP also prevented the mechanical load evoked activation of hedgehog signaling: protein levels of Sonic and Indian Hedgehogs and Gli1 transcription factor were lowered while expressions of Gli2 and Gli3 were elevated by PACAP application during mechanical load. Our results suggest that mechanical load activates PACAP signaling and exogenous PACAP acts against the hypertrophy inducing effect of mechanical load. PMID:26230691

  9. Pituitary Adenylate Cyclase Activating Polypeptide (PACAP Pathway Is Induced by Mechanical Load and Reduces the Activity of Hedgehog Signaling in Chondrogenic Micromass Cell Cultures

    Directory of Open Access Journals (Sweden)

    Tamás Juhász

    2015-07-01

    Full Text Available Pituitary adenylate cyclase activating polypeptide (PACAP is a neurohormone exerting protective function during various stress conditions either in mature or developing tissues. Previously we proved the presence of PACAP signaling elements in chicken limb bud-derived chondrogenic cells in micromass cell cultures. Since no data can be found if PACAP signaling is playing any role during mechanical stress in any tissues, we aimed to investigate its contribution in mechanotransduction during chondrogenesis. Expressions of the mRNAs of PACAP and its major receptor, PAC1 increased, while that of other receptors, VPAC1, VPAC2 decreased upon mechanical stimulus. Mechanical load enhanced the expression of collagen type X, a marker of hypertrophic differentiation of chondrocytes and PACAP addition attenuated this elevation. Moreover, exogenous PACAP also prevented the mechanical load evoked activation of hedgehog signaling: protein levels of Sonic and Indian Hedgehogs and Gli1 transcription factor were lowered while expressions of Gli2 and Gli3 were elevated by PACAP application during mechanical load. Our results suggest that mechanical load activates PACAP signaling and exogenous PACAP acts against the hypertrophy inducing effect of mechanical load.

  10. R-spondin 2 facilitates differentiation of proliferating chondrocytes into hypertrophic chondrocytes by enhancing Wnt/β-catenin signaling in endochondral ossification

    International Nuclear Information System (INIS)

    Takegami, Yasuhiko; Ohkawara, Bisei; Ito, Mikako; Masuda, Akio; Nakashima, Hiroaki; Ishiguro, Naoki; Ohno, Kinji

    2016-01-01

    Endochondral ossification is a crucial process for longitudinal growth of bones. Differentiating chondrocytes in growth cartilage form four sequential zones of proliferation, alignment into column, hypertrophy, and substitution of chondrocytes with osteoblasts. Wnt/β-catenin signaling is essential for differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. R-spondin 2 (Rspo2), a member of R-spondin family, is an agonist for Wnt signaling, but its role in chondrocyte differentiation remains unknown. Here we report that growth cartilage of Rspo2-knockout mice shows a decreased amount of β-catenin and increased amounts collagen type II (CII) and Sox9 in the abnormally extended proliferating zone. In contrast, expression of collagen type X (CX) in the hypertrophic zone remains unchanged. Differentiating chondrogenic ATDC5 cells, mimicking proliferating chondrocytes, upregulate Rspo2 and its putative receptor, Lgr5, in parallel. Addition of recombinant human Rspo2 to differentiating ATDC5 cells decreases expressions of Col2a1, Sox9, and Acan, as well as production of proteoglycans. In contrast, lentivirus-mediated knockdown of Rspo2 has the opposite effect. The effect of Rspo2 on chondrogenic differentiation is mediated by Wnt/β-catenin signaling, and not by Wnt/PCP or Wnt/Ca 2+ signaling. We propose that Rspo2 activates Wnt/β-catenin signaling to reduce Col2a1 and Sox9 and to facilitate differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. - Highlights: • Rspo2 is a secreted activator of Wnt, and its knockout shows extended proliferating chondrocytes in endochondral ossification. • In proliferating chondrocytes of Rspo2-knockout mice, Sox9 and collagen type 2 are increased and β-catenin is decreased. • Rspo2 and its receptor Lgr5, as well as Sox9 and collagen type 2, are expressed in differentiating ATDC5 chondrogenic cells. • In ATDC5 cells, Rspo2 decreases expressions

  11. R-spondin 2 facilitates differentiation of proliferating chondrocytes into hypertrophic chondrocytes by enhancing Wnt/β-catenin signaling in endochondral ossification

    Energy Technology Data Exchange (ETDEWEB)

    Takegami, Yasuhiko [Division of Neurogenetics, Center of Neurological Disease and Cancer, Nagoya University Graduate School of Medicine, Nagoya (Japan); Department of Orthopaedic Surgery, Nagoya University School of Medicine, Nagoya (Japan); Ohkawara, Bisei; Ito, Mikako; Masuda, Akio [Division of Neurogenetics, Center of Neurological Disease and Cancer, Nagoya University Graduate School of Medicine, Nagoya (Japan); Nakashima, Hiroaki; Ishiguro, Naoki [Department of Orthopaedic Surgery, Nagoya University School of Medicine, Nagoya (Japan); Ohno, Kinji, E-mail: ohnok@med.nagoya-u.ac.jp [Division of Neurogenetics, Center of Neurological Disease and Cancer, Nagoya University Graduate School of Medicine, Nagoya (Japan)

    2016-04-22

    Endochondral ossification is a crucial process for longitudinal growth of bones. Differentiating chondrocytes in growth cartilage form four sequential zones of proliferation, alignment into column, hypertrophy, and substitution of chondrocytes with osteoblasts. Wnt/β-catenin signaling is essential for differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. R-spondin 2 (Rspo2), a member of R-spondin family, is an agonist for Wnt signaling, but its role in chondrocyte differentiation remains unknown. Here we report that growth cartilage of Rspo2-knockout mice shows a decreased amount of β-catenin and increased amounts collagen type II (CII) and Sox9 in the abnormally extended proliferating zone. In contrast, expression of collagen type X (CX) in the hypertrophic zone remains unchanged. Differentiating chondrogenic ATDC5 cells, mimicking proliferating chondrocytes, upregulate Rspo2 and its putative receptor, Lgr5, in parallel. Addition of recombinant human Rspo2 to differentiating ATDC5 cells decreases expressions of Col2a1, Sox9, and Acan, as well as production of proteoglycans. In contrast, lentivirus-mediated knockdown of Rspo2 has the opposite effect. The effect of Rspo2 on chondrogenic differentiation is mediated by Wnt/β-catenin signaling, and not by Wnt/PCP or Wnt/Ca{sup 2+} signaling. We propose that Rspo2 activates Wnt/β-catenin signaling to reduce Col2a1 and Sox9 and to facilitate differentiation of proliferating chondrocytes into hypertrophic chondrocytes in growth cartilage. - Highlights: • Rspo2 is a secreted activator of Wnt, and its knockout shows extended proliferating chondrocytes in endochondral ossification. • In proliferating chondrocytes of Rspo2-knockout mice, Sox9 and collagen type 2 are increased and β-catenin is decreased. • Rspo2 and its receptor Lgr5, as well as Sox9 and collagen type 2, are expressed in differentiating ATDC5 chondrogenic cells. • In ATDC5 cells, Rspo2 decreases

  12. Hippo Signaling Influences HNF4A and FOXA2 Enhancer Switching during Hepatocyte Differentiation

    Directory of Open Access Journals (Sweden)

    Olivia Alder

    2014-10-01

    Full Text Available Summary: Cell fate acquisition is heavily influenced by direct interactions between master regulators and tissue-specific enhancers. However, it remains unclear how lineage-specifying transcription factors, which are often expressed in both progenitor and mature cell populations, influence cell differentiation. Using in vivo mouse liver development as a model, we identified thousands of enhancers that are bound by the master regulators HNF4A and FOXA2 in a differentiation-dependent manner, subject to chromatin remodeling, and associated with differentially expressed target genes. Enhancers exclusively occupied in the embryo were found to be responsive to developmentally regulated TEAD2 and coactivator YAP1. Our data suggest that Hippo signaling may affect hepatocyte differentiation by influencing HNF4A and FOXA2 interactions with temporal enhancers. In summary, transcription factor-enhancer interactions are not only tissue specific but also differentiation dependent, which is an important consideration for researchers studying cancer biology or mammalian development and/or using transformed cell lines. : It is unclear how key transcription factors are critical for both lineage specification during embryonic development and maintenance of a differentiated, adult phenotype. By profiling the enhancer occupancy of the key transcription factors HNF4A and FOXA2 during mouse liver development, Alder et al. have found that YAP1 can influence enhancer interactions and target gene expression levels. Enhancer switching enables HNF4A and FOXA2 to fulfill distinct roles during organ development.

  13. A Variable Order Fractional Differential-Based Texture Enhancement Algorithm with Application in Medical Imaging.

    Directory of Open Access Journals (Sweden)

    Qiang Yu

    Full Text Available Texture enhancement is one of the most important techniques in digital image processing and plays an essential role in medical imaging since textures discriminate information. Most image texture enhancement techniques use classical integral order differential mask operators or fractional differential mask operators using fixed fractional order. These masks can produce excessive enhancement of low spatial frequency content, insufficient enhancement of large spatial frequency content, and retention of high spatial frequency noise. To improve upon existing approaches of texture enhancement, we derive an improved Variable Order Fractional Centered Difference (VOFCD scheme which dynamically adjusts the fractional differential order instead of fixing it. The new VOFCD technique is based on the second order Riesz fractional differential operator using a Lagrange 3-point interpolation formula, for both grey scale and colour image enhancement. We then use this method to enhance photographs and a set of medical images related to patients with stroke and Parkinson's disease. The experiments show that our improved fractional differential mask has a higher signal to noise ratio value than the other fractional differential mask operators. Based on the corresponding quantitative analysis we conclude that the new method offers a superior texture enhancement over existing methods.

  14. Cyclic hydrostatic pressure promotes a stable cartilage phenotype and enhances the functional development of cartilaginous grafts engineered using multipotent stromal cells isolated from bone marrow and infrapatellar fat pad.

    Science.gov (United States)

    Carroll, S F; Buckley, C T; Kelly, D J

    2014-06-27

    The objective of this study was to investigate how joint specific biomechanical loading influences the functional development and phenotypic stability of cartilage grafts engineered in vitro using stem/progenitor cells isolated from different source tissues. Porcine bone marrow derived multipotent stromal cells (BMSCs) and infrapatellar fat pad derived multipotent stromal cells (FPSCs) were seeded in agarose hydrogels and cultured in chondrogenic medium, while simultaneously subjected to 10MPa of cyclic hydrostatic pressure (HP). To mimic the endochondral phenotype observed in vivo with cartilaginous tissues engineered using BMSCs, the culture media was additionally supplemented with hypertrophic factors, while the loss of phenotype observed in vivo with FPSCs was induced by withdrawing transforming growth factor (TGF)-β3 from the media. The application of HP was found to enhance the functional development of cartilaginous tissues engineered using both BMSCs and FPSCs. In addition, HP was found to suppress calcification of tissues engineered using BMSCs cultured in chondrogenic conditions and acted to maintain a chondrogenic phenotype in cartilaginous grafts engineered using FPSCs. The results of this study point to the importance of in vivo specific mechanical cues for determining the terminal phenotype of chondrogenically primed multipotent stromal cells. Furthermore, demonstrating that stem or progenitor cells will appropriately differentiate in response to such biophysical cues might also be considered as an additional functional assay for evaluating their therapeutic potential. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. Complete human serum maintains viability and chondrogenic potential of human synovial stem cells: suitable conditions for transplantation.

    Science.gov (United States)

    Mizuno, Mitsuru; Katano, Hisako; Otabe, Koji; Komori, Keiichiro; Kohno, Yuji; Fujii, Shizuka; Ozeki, Nobutake; Horie, Masafumi; Tsuji, Kunikazu; Koga, Hideyuki; Muneta, Takeshi; Sekiya, Ichiro

    2017-06-13

    In our clinical practice, we perform transplantations of autologous synovial mesenchymal stem cells (MSCs) for cartilage and meniscus regenerative medicine. One of the most important issues to ensuring clinical efficacy involves the transport of synovial MSCs from the processing facility to the clinic. Complete human serum (100% human serum) is an attractive candidate material in which to suspend synovial MSCs for their preservation during transport. The purpose of this study was to investigate whether complete human serum maintained MSC viability and chondrogenic potential and to examine the optimal temperature conditions for the preservation of human synovial MSCs. Human synovium was harvested from the knees of 14 donors with osteoarthritis during total knee arthroplasty. Passage 2 synovial MSCs were suspended at 2 million cells/100 μL in Ringer's solution or complete human serum at 4, 13, and 37 °C for 48 h. These cells were analyzed for live cell rates, cell surface marker expression, metabolic activity, proliferation, and adipogenic, calcification, and chondrogenic differentiation potentials before and after preservation. After preservation, synovial MSCs maintained higher live cell rates in human serum than in Ringer's solution at 4 and 13 °C. Synovial MSCs preserved in human serum at 4 and 13 °C also maintained high ratios of propidium iodide - and annexin V - cells. MSC surface marker expression was not altered in cells preserved at 4 and 13 °C. The metabolic activities of cells preserved in human serum at 4 and 13 °C was maintained, while significantly reduced in other conditions. Replated MSCs retained their proliferation ability when preserved in human serum at 4 and 13 °C. Adipogenesis and calcification potential could be observed in cells preserved in each condition, whereas chondrogenic potential was retained only in cells preserved in human serum at 4 and 13 °C. The viability and chondrogenic potential of synovial MSCs were

  16. Long-term dynamic loading improves the mechanical properties of chondrogenic mesenchymal stem cell-laden hydrogel

    Directory of Open Access Journals (Sweden)

    AH Huang

    2010-02-01

    Full Text Available Mesenchymal stem cells (MSCs are an attractive cell source for cartilage tissue engineering given their ability to undergo chondrogenesis in 3D culture systems. Mechanical forces play an important role in regulating both cartilage development and MSC chondrogenic gene expression, however, mechanical stimulation has yet to enhance the mechanical properties of engineered constructs. In this study, we applied long-term dynamic compression to MSC-seeded constructs and assessed whether varying pre-culture duration, loading regimens and inclusion of TGF-beta3 during loading would influence functional outcomes and these phenotypic transitions. Loading initiated before chondrogenesis decreased functional maturation, although chondrogenic gene expression increased. In contrast, loading initiated after chondrogenesis and matrix elaboration further improved the mechanical properties of MSC-based constructs, but only when TGF-beta3 levels were maintained and under specific loading parameters. Although matrix quantity was not affected by dynamic compression, matrix distribution, assessed histologically and by FT-IRIS analysis, was significantly improved on the micro- (pericellular and macro- (construct expanse scales. Further, whole genome expression profiling revealed marked shifts in the molecular topography with dynamic loading. These results demonstrate, for the first time, that dynamic compressive loading initiated after a sufficient period of chondro-induction and with sustained TGF-beta exposure enhances matrix distribution and the mechanical properties of MSC-seeded constructs.

  17. Carbon nanopillars for enhanced stem cell differentiation and dopamine detection

    DEFF Research Database (Denmark)

    Bunea, Ada-Ioana; Amato, Letizia; Valsesia, Andrea

    of human neural stem cells (hNSCs) into dopaminergic neurons and that they can also be employed for detecting dopamine release from mature neurons attached to them [1]. Here, we report 3D carbon nanopillars, fabricated through colloidal lithography, with even more pronounced effect on the electrochemical......Parkinson’s disease is characterized by a deficit of dopamine in the brain, a neurotransmitter involved in the motor function. One of the future ideas for treatment is cell replacement therapy. Our group has previously shown that pyrolysed 3D carbon micropillars induce spontaneous differentiation...

  18. Differential evolution to enhance localization of mobile robots

    DEFF Research Database (Denmark)

    Lisowski, Michal; Fan, Zhun; Ravn, Ole

    2011-01-01

    . In addition, a novel mechanism for effective robot kidnap detection was proposed. Experiments were performed using computer simulations based on the odometer data and laser range finder measurements collected in advance by a robot in real-life. Experimental results showed that integrating DE enables MCL...... to provide more accurate robot pose estimations in shorter time while using fewer particles.......This paper focuses on the mobile robot localization problems: pose tracking, global localization and robot kidnap. Differential Evolution (DE) applied to extend Monte Carlo Localization (MCL) was investigated to better solve localization problem by increasing localization reliability and speed...

  19. Differential evolution optimization combined with chaotic sequences for image contrast enhancement

    Energy Technology Data Exchange (ETDEWEB)

    Santos Coelho, Leandro dos [Industrial and Systems Engineering Graduate Program, LAS/PPGEPS, Pontifical Catholic University of Parana, PUCPR Imaculada Conceicao, 1155, 80215-901 Curitiba, Parana (Brazil)], E-mail: leandro.coelho@pucpr.br; Sauer, Joao Guilherme [Industrial and Systems Engineering Graduate Program, LAS/PPGEPS, Pontifical Catholic University of Parana, PUCPR Imaculada Conceicao, 1155, 80215-901 Curitiba, Parana (Brazil)], E-mail: joao.sauer@gmail.com; Rudek, Marcelo [Industrial and Systems Engineering Graduate Program, LAS/PPGEPS, Pontifical Catholic University of Parana, PUCPR Imaculada Conceicao, 1155, 80215-901 Curitiba, Parana (Brazil)], E-mail: marcelo.rudek@pucpr.br

    2009-10-15

    Evolutionary Algorithms (EAs) are stochastic and robust meta-heuristics of evolutionary computation field useful to solve optimization problems in image processing applications. Recently, as special mechanism to avoid being trapped in local minimum, the ergodicity property of chaotic sequences has been used in various designs of EAs. Three differential evolution approaches based on chaotic sequences using logistic equation for image enhancement process are proposed in this paper. Differential evolution is a simple yet powerful evolutionary optimization algorithm that has been successfully used in solving continuous problems. The proposed chaotic differential evolution schemes have fast convergence rate but also maintain the diversity of the population so as to escape from local optima. In this paper, the image contrast enhancement is approached as a constrained nonlinear optimization problem. The objective of the proposed chaotic differential evolution schemes is to maximize the fitness criterion in order to enhance the contrast and detail in the image by adapting the parameters using a contrast enhancement technique. The proposed chaotic differential evolution schemes are compared with classical differential evolution to two testing images. Simulation results on three images show that the application of chaotic sequences instead of random sequences is a possible strategy to improve the performance of classical differential evolution optimization algorithm.

  20. Enhance and Maintain Chondrogenesis of Synovial Fibroblasts by Cartilage Extracellular Matrix Protein Matrilins

    Science.gov (United States)

    Pei, Ming; Luo, Junming; Chen, Qian

    2008-01-01

    Summary Objective Cartilage-specific extracellular matrix (ECM) proteins have been proposed to play key roles in modulating cellular phenotypes during chondrogenesis of mesenchymal stem cells. Matrilin (MATN) 1 and 3 are among the most up-regulated ECM proteins during chondrogenesis. The aim of this study was to analyze their roles in chondrogenesis of mesenchymal fibroblasts from synovium. Methods Primary synovial fibroblasts (SFBs) were purified from porcine synovium and incubated in pellet culture for 18 days. Chondrogenesis of SFB was analyzed by histological staining with safranin-O/fast green, and by quantifying glycosaminoglycans with dimethylmethylene blue assay. The mRNA levels of chondrogenic markers including collagen II, aggrecan, and Sox 9 were quantified by real-time RT-PCR, while the protein levels of Col II and matrilins were determined by western blot analysis. Results SFBs underwent chondrogenesis after incubation with TGF-β1 for three days; however, this process was attenuated during the subsequent incubation period. Expression of a MATN1 or 3 cDNA maintained and further enhanced chondrogenesis of SFBs as shown by increased cartilaginous matrix areas, elevated amount of glycosaminoglycans, and stimulated expression of chondrogenic markers. Conclusion Our findings suggest a novel function for MATN1 and 3 to maintain and enhance chondrogenesis of mesenchymal fibroblasts initiated by TGF-β. Our results also support a critical role of cartilage-specific ECM proteins to modulate cellular phenotypes in the microenvironment during chondrogenic differentiation. PMID:18282772

  1. Enhancing and maintaining chondrogenesis of synovial fibroblasts by cartilage extracellular matrix protein matrilins.

    Science.gov (United States)

    Pei, M; Luo, J; Chen, Q

    2008-09-01

    Cartilage-specific extracellular matrix (ECM) proteins have been proposed to play key roles in modulating cellular phenotypes during chondrogenesis of mesenchymal stem cells. Matrilin (MATN)1 and MATN3 are among the most up-regulated ECM proteins during chondrogenesis. The aim of this study was to analyze their roles in chondrogenesis of mesenchymal fibroblasts from synovium. Primary synovial fibroblasts (SFBs) were purified from porcine synovium and incubated in pellet culture for 18 days. Chondrogenesis of SFB was analyzed by histological staining with safranin-O/fast green, and by quantifying glycosaminoglycans (GAG) with dimethylmethylene blue assay. The mRNA levels of chondrogenic markers including collagen II, aggrecan, and Sox 9 were quantified by real-time reverse transcription polymerase chain reaction, while the protein levels of Col II and MATNs were determined by western blot analysis. SFBs underwent chondrogenesis after incubation with transforming growth factor-beta1 (TGF-beta1) for 3 days; however, this process was attenuated during the subsequent incubation period. Expression of a Matn1 or Matn3 cDNA maintained and further enhanced chondrogenesis of SFBs as shown by increased cartilaginous matrix areas, elevated amount of GAG, and stimulated expression of chondrogenic markers. Our findings suggest a novel function for MATN1 and MATN3 to maintain and enhance chondrogenesis of mesenchymal fibroblasts initiated by TGF-beta. Our results also support a critical role of cartilage-specific ECM proteins to modulate cellular phenotypes in the microenvironment during chondrogenic differentiation.

  2. Inhibition of TGF-β Signaling in SHED Enhances Endothelial Differentiation.

    Science.gov (United States)

    Xu, J G; Gong, T; Wang, Y Y; Zou, T; Heng, B C; Yang, Y Q; Zhang, C F

    2018-02-01

    Low efficiency of deriving endothelial cells (ECs) from adult stem cells hampers their utilization in tissue engineering studies. The purpose of this study was to investigate whether suppression of transforming growth factor beta (TGF-β) signaling could enhance the differentiation efficiency of dental pulp-derived stem cells into ECs. We initially used vascular endothelial growth factor A (VEGF-A) to stimulate 2 dental pulp-derived stem cells (dental pulp stem cells and stem cells from human exfoliated deciduous teeth [SHED]) and compared their differentiation capacity into ECs. We further evaluated whether the vascular endothelial growth factor receptor I (VEGF-RI)-specific ligand placental growth factor-1 (PlGF-1) could mediate endothelial differentiation. Finally, we investigated whether the TGF-β signaling inhibitor SB-431542 could enhance the inductive effect of VEGF-A on endothelial differentiation, as well as the underlying mechanisms involved. ECs differentiated from dental pulp-derived stem cells exhibited the typical phenotypes of primary ECs, with SHED possessing a higher endothelial differentiation potential than dental pulp stem cells. VEGFR1-specific ligand-PLGF exerted a negligible effect on SHED-ECs differentiation. Compared with VEGF-A alone, the combination of VEGF-A and SB-431542 significantly enhanced the endothelial differentiation of SHED. The presence of SB-431542 inhibited the phosphorylation of Suppressor of Mothers Against Decapentaplegic 2/3 (SMAD2/3), allowing for VEGF-A-dependent phosphorylation and upregulation of VEGFR2. Our results indicate that the combination of VEGF-A and SB-431542 could enhance the differentiation of dental pulp-derived stem cells into endothelial cells, and this process is mediated through enhancement of VEGF-A-VEGFR2 signaling and concomitant inhibition of TGF-β-SMAD2/3 signaling.

  3. Fibroblast growth factor-2 stimulates adipogenic differentiation of human adipose-derived stem cells

    International Nuclear Information System (INIS)

    Kakudo, Natsuko; Shimotsuma, Ayuko; Kusumoto, Kenji

    2007-01-01

    Adipose-derived stem cells (ASCs) have demonstrated a capacity for differentiating into a variety of lineages, including bone, cartilage, or fat, depending on the inducing stimuli and specific growth and factors. It is acknowledged that fibroblast growth factor-2 (FGF-2) promotes chondrogenic and inhibits osteogenic differentiation of ASCs, but thorough investigations of its effects on adipogenic differentiation are lacking. In this study, we demonstrate at the cellular and molecular levels the effect of FGF-2 on adipogenic differentiation of ASCs, as induced by an adipogenic hormonal cocktail consisting of 3-isobutyl-1-methylxanthine (IBMX), dexamethasone, insulin, and indomethacin. FGF-2 significantly enhances the adipogenic differentiation of human ASCs. Furthermore, in cultures receiving FGF-2 before adipogenic induction, mRNA expression of peroxisome proliferator-activated receptor γ2 (PPARγ2), a key transcription factor in adipogenesis, was upregulated. The results of FGF-2 supplementation suggest the potential applications of FGF-2 and ASCs in adipose tissue regeneration

  4. Impact of low oxygen tension on stemness, proliferation and differentiation potential of human adipose-derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Jane Ru; Pingguan-Murphy, Belinda; Wan Abas, Wan Abu Bakar [Department of Biomedical Engineering, Faculty of Engineering, University of Malaya, Lembah Pantai, 50603 Kuala Lumpur (Malaysia); Noor Azmi, Mat Adenan; Omar, Siti Zawiah [Department of Obstetrics and Gynaecology, Faculty of Medicine, University of Malaya, Lembah Pantai, 50603 Kuala Lumpur (Malaysia); Chua, Kien Hui [Department of Physiology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Wan Safwani, Wan Kamarul Zaman, E-mail: wansafwani@um.edu.my [Department of Biomedical Engineering, Faculty of Engineering, University of Malaya, Lembah Pantai, 50603 Kuala Lumpur (Malaysia)

    2014-05-30

    Highlights: • Hypoxia maintains the stemness of adipose-derived stem cells (ASCs). • ASCs show an increased proliferation rate under low oxygen tension. • Oxygen level as low as 2% enhances the chondrogenic differentiation potential of ASCs. • HIF-1α may regulate the proliferation and differentiation activities of ASCs under hypoxia. - Abstract: Adipose-derived stem cells (ASCs) have been found adapted to a specific niche with low oxygen tension (hypoxia) in the body. As an important component of this niche, oxygen tension has been known to play a critical role in the maintenance of stem cell characteristics. However, the effect of O{sub 2} tension on their functional properties has not been well determined. In this study, we investigated the effects of O{sub 2} tension on ASCs stemness, differentiation and proliferation ability. Human ASCs were cultured under normoxia (21% O{sub 2}) and hypoxia (2% O{sub 2}). We found that hypoxia increased ASC stemness marker expression and proliferation rate without altering their morphology and surface markers. Low oxygen tension further enhances the chondrogenic differentiation ability, but reduces both adipogenic and osteogenic differentiation potential. These results might be correlated with the increased expression of HIF-1α under hypoxia. Taken together, we suggest that growing ASCs under 2% O{sub 2} tension may be important in expanding ASCs effectively while maintaining their functional properties for clinical therapy, particularly for the treatment of cartilage defects.

  5. Platelet lysate enhances synovial fluid multipotential stromal cells functions: Implications for therapeutic use.

    Science.gov (United States)

    Altaie, Ala; Baboolal, Thomas G; Wall, Owen; Jones, Elena; McGonagle, Dennis

    2018-03-01

    Although intra-articular injection of platelet products is increasingly used for joint regenerative approaches, there are few data on their biological effects on joint-resident multipotential stromal cells (MSCs), which are directly exposed to the effects of these therapeutic strategies. Therefore, this study investigated the effect of platelet lysate (PL) on synovial fluid-derived MSCs (SF-MSCs), which in vivo have direct access to sites of cartilage injury. SF-MSCs were obtained during knee arthroscopic procedures (N = 7). Colony forming unit-fibroblast (CFU-F), flow-cytometric phenotyping, carboxyfluorescein succinimidyl ester-based immunomodulation for T-cell and trilineage differentiation assays were performed using PL and compared with standard conditions. PL-enhanced SF-MSC (PL-MSC) proliferation as CFU-F colonies was 1.4-fold larger, and growing cultures had shorter population-doubling times. PL-MSCs and fetal calf serum (FCS)-MSCs had the same immunophenotype and similar immunomodulation activities. In chondrogenic and osteogenic differentiation assays, PL-MSCs produced 10% more sulfated-glycosaminoglycan (sGAG) and 45% less Ca ++ compared with FCS-MSCs, respectively. Replacing chondrogenic medium transforming growth factor-β3 with 20% or 50% PL further increased sGAG production of PL-MSCs by 69% and 95%, respectively, compared with complete chondrogenic medium. Also, Dulbecco's Modified Eagle's Medium high glucose (HG-DMEM) plus 50% PL induced more chondrogenesis compared with HG-DMEM plus 10% FCS and was comparable to complete chondrogenic medium. This is the first study to assess SF-MSC responses to PL and provides biological support to the hypothesis that PL may be capable of modulating multiple functional aspects of joint resident MSCs with direct access to injured cartilage. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  6. NGSmethDB 2017: enhanced methylomes and differential methylation

    Science.gov (United States)

    Lebrón, Ricardo; Gómez-Martín, Cristina; Carpena, Pedro; Bernaola-Galván, Pedro; Barturen, Guillermo; Hackenberg, Michael; Oliver, José L.

    2017-01-01

    The 2017 update of NGSmethDB stores whole genome methylomes generated from short-read data sets obtained by bisulfite sequencing (WGBS) technology. To generate high-quality methylomes, stringent quality controls were integrated with third-part software, adding also a two-step mapping process to exploit the advantages of the new genome assembly models. The samples were all profiled under constant parameter settings, thus enabling comparative downstream analyses. Besides a significant increase in the number of samples, NGSmethDB now includes two additional data-types, which are a valuable resource for the discovery of methylation epigenetic biomarkers: (i) differentially methylated single-cytosines; and (ii) methylation segments (i.e. genome regions of homogeneous methylation). The NGSmethDB back-end is now based on MongoDB, a NoSQL hierarchical database using JSON-formatted documents and dynamic schemas, thus accelerating sample comparative analyses. Besides conventional database dumps, track hubs were implemented, which improved database access, visualization in genome browsers and comparative analyses to third-part annotations. In addition, the database can be also accessed through a RESTful API. Lastly, a Python client and a multiplatform virtual machine allow for program-driven access from user desktop. This way, private methylation data can be compared to NGSmethDB without the need to upload them to public servers. Database website: http://bioinfo2.ugr.es/NGSmethDB. PMID:27794041

  7. Differential Connexin Function Enhances Self-Renewal in Glioblastoma

    Directory of Open Access Journals (Sweden)

    Masahiro Hitomi

    2015-05-01

    Full Text Available The coordination of complex tumor processes requires cells to rapidly modify their phenotype and is achieved by direct cell-cell communication through gap junction channels composed of connexins. Previous reports have suggested that gap junctions are tumor suppressive based on connexin 43 (Cx43, but this does not take into account differences in connexin-mediated ion selectivity and intercellular communication rate that drive gap junction diversity. We find that glioblastoma cancer stem cells (CSCs possess functional gap junctions that can be targeted using clinically relevant compounds to reduce self-renewal and tumor growth. Our analysis reveals that CSCs express Cx46, while Cx43 is predominantly expressed in non-CSCs. During differentiation, Cx46 is reduced, while Cx43 is increased, and targeting Cx46 compromises CSC maintenance. The difference between Cx46 and Cx43 is reflected in elevated cell-cell communication and reduced resting membrane potential in CSCs. Our data demonstrate a pro-tumorigenic role for gap junctions that is dependent on connexin expression.

  8. NGSmethDB 2017: enhanced methylomes and differential methylation.

    Science.gov (United States)

    Lebrón, Ricardo; Gómez-Martín, Cristina; Carpena, Pedro; Bernaola-Galván, Pedro; Barturen, Guillermo; Hackenberg, Michael; Oliver, José L

    2017-01-04

    The 2017 update of NGSmethDB stores whole genome methylomes generated from short-read data sets obtained by bisulfite sequencing (WGBS) technology. To generate high-quality methylomes, stringent quality controls were integrated with third-part software, adding also a two-step mapping process to exploit the advantages of the new genome assembly models. The samples were all profiled under constant parameter settings, thus enabling comparative downstream analyses. Besides a significant increase in the number of samples, NGSmethDB now includes two additional data-types, which are a valuable resource for the discovery of methylation epigenetic biomarkers: (i) differentially methylated single-cytosines; and (ii) methylation segments (i.e. genome regions of homogeneous methylation). The NGSmethDB back-end is now based on MongoDB, a NoSQL hierarchical database using JSON-formatted documents and dynamic schemas, thus accelerating sample comparative analyses. Besides conventional database dumps, track hubs were implemented, which improved database access, visualization in genome browsers and comparative analyses to third-part annotations. In addition, the database can be also accessed through a RESTful API. Lastly, a Python client and a multiplatform virtual machine allow for program-driven access from user desktop. This way, private methylation data can be compared to NGSmethDB without the need to upload them to public servers. Database website: http://bioinfo2.ugr.es/NGSmethDB. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  9. Cartilage regeneration by chondrogenic induced adult stem cells in osteoarthritic sheep model.

    Science.gov (United States)

    Ude, Chinedu C; Sulaiman, Shamsul B; Min-Hwei, Ng; Hui-Cheng, Chen; Ahmad, Johan; Yahaya, Norhamdan M; Saim, Aminuddin B; Idrus, Ruszymah B H

    2014-01-01

    In this study, Adipose stem cells (ADSC) and bone marrow stem cells (BMSC), multipotent adult cells with the potentials for cartilage regenerations were induced to chondrogenic lineage and used for cartilage regenerations in surgically induced osteoarthritis in sheep model. Osteoarthritis was induced at the right knee of sheep by complete resection of the anterior cruciate ligament and medial meniscus following a 3-weeks exercise regimen. Stem cells from experimental sheep were culture expanded and induced to chondrogenic lineage. Test sheep received a single dose of 2 × 10(7) autologous PKH26-labelled, chondrogenically induced ADSCs or BMSCs as 5 mls injection, while controls received 5 mls culture medium. The proliferation rate of ADSCs 34.4 ± 1.6 hr was significantly higher than that of the BMSCs 48.8 ± 5.3 hr (P = 0.008). Chondrogenic induced BMSCs had significantly higher expressions of chondrogenic specific genes (Collagen II, SOX9 and Aggrecan) compared to chondrogenic ADSCs (P = 0.031, 0.010 and 0.013). Grossly, the treated knee joints showed regenerated de novo cartilages within 6 weeks post-treatment. On the International Cartilage Repair Society grade scores, chondrogenically induced ADSCs and BMSCs groups had significantly lower scores than controls (P = 0.0001 and 0.0001). Fluorescence of the tracking dye (PKH26) in the injected cells showed that they had populated the damaged area of cartilage. Histological staining revealed loosely packed matrixes of de novo cartilages and immunostaining demonstrated the presence of cartilage specific proteins, Collagen II and SOX9. Autologous chondrogenically induced ADSCs and BMSCs could be promising cell sources for cartilage regeneration in osteoarthritis.

  10. Cartilage regeneration by chondrogenic induced adult stem cells in osteoarthritic sheep model.

    Directory of Open Access Journals (Sweden)

    Chinedu C Ude

    Full Text Available OBJECTIVES: In this study, Adipose stem cells (ADSC and bone marrow stem cells (BMSC, multipotent adult cells with the potentials for cartilage regenerations were induced to chondrogenic lineage and used for cartilage regenerations in surgically induced osteoarthritis in sheep model. METHODS: Osteoarthritis was induced at the right knee of sheep by complete resection of the anterior cruciate ligament and medial meniscus following a 3-weeks exercise regimen. Stem cells from experimental sheep were culture expanded and induced to chondrogenic lineage. Test sheep received a single dose of 2 × 10(7 autologous PKH26-labelled, chondrogenically induced ADSCs or BMSCs as 5 mls injection, while controls received 5 mls culture medium. RESULTS: The proliferation rate of ADSCs 34.4 ± 1.6 hr was significantly higher than that of the BMSCs 48.8 ± 5.3 hr (P = 0.008. Chondrogenic induced BMSCs had significantly higher expressions of chondrogenic specific genes (Collagen II, SOX9 and Aggrecan compared to chondrogenic ADSCs (P = 0.031, 0.010 and 0.013. Grossly, the treated knee joints showed regenerated de novo cartilages within 6 weeks post-treatment. On the International Cartilage Repair Society grade scores, chondrogenically induced ADSCs and BMSCs groups had significantly lower scores than controls (P = 0.0001 and 0.0001. Fluorescence of the tracking dye (PKH26 in the injected cells showed that they had populated the damaged area of cartilage. Histological staining revealed loosely packed matrixes of de novo cartilages and immunostaining demonstrated the presence of cartilage specific proteins, Collagen II and SOX9. CONCLUSION: Autologous chondrogenically induced ADSCs and BMSCs could be promising cell sources for cartilage regeneration in osteoarthritis.

  11. Differentiation of focal liver lesions by contrast-enhanced MRI

    International Nuclear Information System (INIS)

    Heintz, P.; Ehrenheim, C.

    1989-01-01

    47 patients with liver tumours (haemangioma, focal nodular hyperplasia, hepatocellular carcinoma) underwent MRI of the liver before and after i.v. injection of 0.2 ml./kg. gadolinium-DTPA in addition to other imaging methods. The demarcation of focal nodular hyperplasia is not influenced by use of the contrast agent as it almost behaves like surrounding normal liver tissue, thus only indirectly facilitating its identification. With regard to liver haemangiomas that show the most intensive uptake of gadolinium-DTPA, the contrast enhanced image does not reach to contrast and sensitivity of a native T 2 -weighted SE image, especially in cases of small haemangiomas. The contrast agent is helpful, however, in the recognition of large cavernous haemangiomas that are partially fibrotic or thrombotic. Emphasis is given to the contrast agent in hepatomas: gadolinium-DTPA presents a pattern of uptake and distribution frequently found in hepatocellular carcinoma providing additional information on the delineation of internal tumour details. (orig.) [de

  12. Differentiation between tuberculosis and lymphoma in mediastinal lymph nodes: Evaluation with contrast-enhanced MDCT

    International Nuclear Information System (INIS)

    Tang, S.-S.; Yang, Z.-G.; Deng, W.; Shao, H.; Chen, J.; Wen, L.-Y.

    2012-01-01

    Aim: To determine the specific imaging criteria on contrast-enhanced multidetector computed tomography (MDCT) for differentiating between tuberculosis and lymphoma in mediastinal lymph nodes. Materials and methods: The anatomical distribution and enhancement patterns of mediastinal lymph nodes on contrast-enhanced MDCT were reviewed in 37 patients with tuberculosis and 54 patients with lymphoma. Of the patients with lymphoma, 18 had Hodgkin's disease and 36 had non-Hodgkin's lymphoma. Results: Region 10R was involved more often in tuberculosis than in Hodgkin's disease and non-Hodgkin's lymphoma. Region 6 had a higher tendency to be affected in Hodgkin's disease and non-Hodgkin's lymphoma compared with tuberculosis. Tuberculosis showed peripheral enhancement in 78% of cases, frequently with a multilocular appearance, compared to Hodgkin's disease and non-Hodgkin's lymphoma, which showed peripheral enhancement in only 6 and 3% of cases, respectively. Homogeneous enhancement was more commonly seen in lymphoma (83% for Hodgkin's disease, and 83% for non-Hodgkin's lymphoma) than in tuberculosis (8%). In the determination of tuberculosis, results showed that when a peripheral enhancement pattern was seen, sensitivity was 78%, specificity was 96%, and accuracy was 89%. In the determination of lymphoma, results showed that when a homogeneous enhancement pattern was seen, sensitivity was 83%, specificity was 92%, and accuracy was 87%. Conclusion: The findings of the present study indicate that specific anatomical distribution and enhancement patterns of lymphadenopathy shown on contrast-enhanced MDCT can be useful in differentiating tuberculosis from lymphoma of mediastinal lymph nodes.

  13. Differentiation of toxoplasmosis and lymphoma in HIV-positive patients with gadolinium-enhanced MR imaging

    International Nuclear Information System (INIS)

    Eisenberg, A.D.; Mani, J.R.; Norman, D.

    1990-01-01

    This paper determines whether gadolinium-enhanced MR imaging can be used to differentiate toxoplasmosis and lymphoma in patients with acquired immunodeficiency syndrome. One hundred fifty-nine lesions from 71 MR examinations of eight patients with lymphoma were evaluated for size, location, enhancement characteristics, lesion multiplicity. Multiple lesions occurred in 72% of toxoplasmosis and 75% of lymphoma cases. Toxoplasmosis lesions are smaller, with lesion most commonly between 1 and 2 cm, whereas lymphoma is most often between 2 and 3 cm. Except for a propensity for lymphoma to occur in the temporal lobes, no difference in lesion location was found. Both conditions usually markedly enhance, but the pattern of enhancement differs

  14. Maintaining the stability of nonlinear differential equations by the enhancement of HPM

    International Nuclear Information System (INIS)

    Hosein Nia, S.H.; Ranjbar, A.N.; Ganji, D.D.; Soltani, H.; Ghasemi, J.

    2008-01-01

    Homotopy perturbation method is an effective method to find a solution of a nonlinear differential equation. In this method, a nonlinear complex differential equation is transformed to a series of linear and nonlinear parts, almost simpler differential equations. These sets of equations are then solved iteratively. Finally, a linear series of the solutions completes the answer if the convergence is maintained. In this Letter, the need for stability verification is shown through some examples. Consequently, HPM is enhanced by a preliminary assumption. The idea is to keep the inherent stability of nonlinear dynamic, even the selected linear part is not

  15. The Infrapatellar Fat Pad as a Source of Perivascular Stem Cells with Increased Chondrogenic Potential for Regenerative Medicine.

    Science.gov (United States)

    Hindle, Paul; Khan, Nusrat; Biant, Leela; Péault, Bruno

    2017-01-01

    Perivascular stem cells (PSCs) are the natural ancestors of mesenchymal stem cells (MSCs) and are the stem cells responsible for homeostasis and repair in vivo. Prospectively identified and isolated PSCs have demonstrated increased plasticity and osteogenic potential. Cells from the infrapatellar fat pad (IFP) have demonstrated increased chondrogenic potential compared with those from subcutaneous fat. This research assessed the chondrogenic potential of IFP PSCs compared with MSCs from the IFP and bone marrow. Immunohistochemistry demonstrated the location of perivascular markers (CD146, CD34, neural/glial antigen 2 [NG2], platelet-derived growth factor receptor-β [PDGFRβ], and α-smooth muscle actin [α-SMA]) in relation to endothelial markers (CD31, CD144, von Willebrand factor [vWF]). Pericytes and adventitial cells were isolated from the stromal vascular fraction (3.8% and 21.2%, respectively) using flow cytometry with a viability of 88%. The mean numbers of pericytes and adventitial cells isolated were 4.6 ± 2.2 × 10 4 and 16.2 ± 3.2 × 10 4 , respectively, equating to 7.9 ± 4.4 × 10 3 and 20.8 ± 4.3 × 10 3 cells per gram of harvested tissue. Fluorescence-activated cell sorting demonstrated that cultured PSCs were CD44+CD90+CD105+; polymerase chain reaction and immunocytochemistry demonstrated that pericytes retained their CD146+ phenotype and expressed the pericyte markers PDGFRβ and NG2. Differentiation was confirmed using histochemical stains and genetic expression. Using a pellet model, the IFP PSCs and the MSCs generated significantly more extracellular matrix than bone marrow MSCs (p < .001 and p = .011, respectively). The IFP PSCs generated significantly more extracellular matrix than IFP MSCs (p = .002). Micromass culture demonstrated that differentiated PSCs were upregulated compared with MSCs for COL2A1, ACAN, and SOX9 expression by factors of 4.8 ± 1.3, 4.3 ± 0.9, and 7.0 ± 1.7, respectively. The IFP was a significantly better source

  16. Value of radio density determined by enhanced computed tomography for the differential diagnosis of lung masses

    International Nuclear Information System (INIS)

    Xie, Min

    2011-01-01

    Lung masses are often difficult to differentiate when their clinical symptoms and shapes or densities on computed tomography images are similar. However, with different pathological contents, they may appear differently on plain and enhanced computed tomography. Objectives: To determine the value of enhanced computed tomography for the differential diagnosis of lung masses based on the differences in radio density with and without enhancement. Patients and Methods: Thirty-six patients with lung cancer, 36 with pulmonary tuberculosis and 10 with inflammatory lung pseudo tumors diagnosed by computed tomography and confirmed by pathology in our hospital were selected. The mean ±SD radio densities of lung masses in the three groups of patients were calculated based on the results of plain and enhanced computed tomography. Results: There were no significant differences in the radio densities of the masses detected by plain computed tomography among patients with inflammatory lung pseudo tumors, tuberculosis and lung cancer (P> 0.05). However, there were significant differences (P< 0.01)between all the groups in terms of radio densities of masses detected by enhanced computed tomography. Conclusions: The radio densities of lung masses detected by enhanced computed tomography could potentially be used to differentiate between lung cancer, pulmonary tuberculosis and inflammatory lung pseudo tumors.

  17. Small Molecule-BIO Accelerates and Enhances Marrow-Derived Mesenchymal Stem Cell in Vitro Chondrogenesis

    Directory of Open Access Journals (Sweden)

    Mohamadreza Baghaban Eslaminejad

    2014-03-01

    Full Text Available Background: Hyaline cartilage defects exhibit a major challenge in the field of orthopedic surgery owing to its limited repair capacity. On the other hand, mesenchymal stem cells (MSCs are regarded as potent cells with a property of cartilage regeneration. We aimed to optimize marrow-derived MSC chondrogenic culture using a small bioactive molecule referred to as BIO. Methods: MSCs from the marrow of NMRI mice were extracted, culture-expanded, and characterized. Micro-mass culture was then established for chondrogenic differentiation (control group. The cultures of MSC in chondrogenic medium supplemented with 0.01, 0.05, 0.1, and 1 µM BIO were taken as the experimental groups. Cartilage differentiation was examined by both histological sections and real-time PCR for Sox9, aggrecan, and collagen II at different time points. Moreover, the involvement of the Wnt pathway was investigated. Results: Based on histological sections, there was seemingly more intense metachromatic matrix produced in the cultures with 0.01 µM BIO. In this experimental group, cartilage-specific genes tended to be upregulated at day 14 compared to day 21 of the control group, indicating the accelerating effect of BIO on cartilage differentiation. Overall, there was statistically a significant increase (P=0.01 in the expression level of cartilage-specific genes in cultures with 0.01 µM BIO (enhancing effects. These upregulations appeared to be mediated through the Wnt pathway evident from the significant upregulation of T-cell factor and beta-catenin molecules (P=0.01. Conclusion: Taken together, BIO at 0.01 µM could accelerate and enhance in vitro chondrogenesis of mouse marrow-derived MSCs. Please cite this article as: Baghaban Eslaminejad MR, Fallah N. Small Molecule-BIO Accelerates and Enhances Marrow-Derived Mesenchymal Stem Cell in Vitro Chondrogenesis. Iran J Med Sci. 2014;39(2:107-116.

  18. Fenofibrate Enhances the In Vitro Differentiation of Foxp3+ Regulatory T Cells in Mice

    Directory of Open Access Journals (Sweden)

    Zhou Zhou

    2012-01-01

    Full Text Available Foxp3+ regulatory T cells (Tregs play a critical role in maintaining immune self-tolerance. Reduced number and activity of Tregs are usually found in autoimmune and inflammatory diseases, and enhancing the differentiation of Tregs may be a promising therapeutic strategy. Some reports suggested an anti-inflammatory and anti-autoimmune potential for fenofibrate, a hypolipidemic drug used worldwide, whose lipid effects are mediated by the activation of peroxisome proliferator-activated receptor (PPAR. In the present paper, we found that fenofibrate dose-dependently increased transforming growth factor- and interleukin-2-induced Treg differentiation in vitro, by 1.96-fold from 0 to 20 M (12.59±1.34% to 24.69±3.03%, <0.05. Other PPAR activators, WY14643 (100 M, gemfibrozil (50 M, and bezafibrate (30 M, could not enhance Treg differentiation. In addition, PPAR could not upregulate the promoter activity of the Treg-specific transcription factor Foxp3. Fenofibrate might exert its function by enhancing Smad3 phosphorylation, a critical signal in Treg differentiation, via Akt suppression. Our work reveals a new PPAR independent anti-inflammatory mechanism of fenofibrate in up-regulating mouse Treg differentiation.

  19. Sphingosine 1-phosphate receptor activation enhances BMP-2-induced osteoblast differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Sato, Chieri [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Iwasaki, Tsuyoshi, E-mail: tsuyo-i@huhs.ac.jp [Division of Pharmacotherapy, Department of Pharmacy, School of Pharmacy, Hyogo University of Health Sciences, 1-3-6 Minatojima, Chuo-ku, Kobe 650-8530 (Japan); Kitano, Sachie; Tsunemi, Sachi; Sano, Hajime [Division of Rheumatology, Department of Internal Medicine, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan)

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer We investigated the role of S1P signaling for osteoblast differentiation. Black-Right-Pointing-Pointer Both S1P and FTY enhanced BMP-2-stimulated osteoblast differentiation by C2C12 cells. Black-Right-Pointing-Pointer S1P signaling enhanced BMP-2-stimulated Smad and ERK phosphorylation by C2C12 cells. Black-Right-Pointing-Pointer MEK/ERK signaling is a pathway underlying S1P signaling for osteoblast differentiation. -- Abstract: We previously demonstrated that sphingosine 1-phosphate (S1P) receptor-mediated signaling induced proliferation and prostaglandin productions by synovial cells from rheumatoid arthritis (RA) patients. In the present study we investigated the role of S1P receptor-mediated signaling for osteoblast differentiation. We investigated osteoblast differentiation using C2C12 myoblasts, a cell line derived from murine satellite cells. Osteoblast differentiation was induced by the treatment of bone morphogenic protein (BMP)-2 in the presence or absence of either S1P or FTY720 (FTY), a high-affinity agonist of S1P receptors. Osteoblast differentiation was determined by osteoblast-specific transcription factor, Runx2 mRNA expression, alkaline phosphatase (ALP) activity and osteocalcin production by the cells. Smad1/5/8 and extracellular signal-regulated kinase (ERK) 1/2 phosphorylation was examined by Western blotting. Osteocalcin production by C2C12 cells were determined by ELISA. Runx2 expression and ALP activity by BMP-2-stimulated C2C12 cells were enhanced by addition of either S1P or FTY. Both S1P and FTY enhanced BMP-2-induced ERK1/2 and Smad1/5/8 phosphorylation. The effect of FTY was stronger than that of S1P. S1P receptor-mediated signaling on osteoblast differentiation was inhibited by addition of mitogen-activated protein kinase/ERK kinase (MEK) 1/2 inhibitor, indicating that the S1P receptor-mediated MEK1/2-ERK1/2 signaling pathway enhanced BMP-2-Smad signaling. These results indicate that S1P

  20. Sphingosine 1-phosphate receptor activation enhances BMP-2-induced osteoblast differentiation

    International Nuclear Information System (INIS)

    Sato, Chieri; Iwasaki, Tsuyoshi; Kitano, Sachie; Tsunemi, Sachi; Sano, Hajime

    2012-01-01

    Highlights: ► We investigated the role of S1P signaling for osteoblast differentiation. ► Both S1P and FTY enhanced BMP-2-stimulated osteoblast differentiation by C2C12 cells. ► S1P signaling enhanced BMP-2-stimulated Smad and ERK phosphorylation by C2C12 cells. ► MEK/ERK signaling is a pathway underlying S1P signaling for osteoblast differentiation. -- Abstract: We previously demonstrated that sphingosine 1-phosphate (S1P) receptor-mediated signaling induced proliferation and prostaglandin productions by synovial cells from rheumatoid arthritis (RA) patients. In the present study we investigated the role of S1P receptor-mediated signaling for osteoblast differentiation. We investigated osteoblast differentiation using C2C12 myoblasts, a cell line derived from murine satellite cells. Osteoblast differentiation was induced by the treatment of bone morphogenic protein (BMP)-2 in the presence or absence of either S1P or FTY720 (FTY), a high-affinity agonist of S1P receptors. Osteoblast differentiation was determined by osteoblast-specific transcription factor, Runx2 mRNA expression, alkaline phosphatase (ALP) activity and osteocalcin production by the cells. Smad1/5/8 and extracellular signal-regulated kinase (ERK) 1/2 phosphorylation was examined by Western blotting. Osteocalcin production by C2C12 cells were determined by ELISA. Runx2 expression and ALP activity by BMP-2-stimulated C2C12 cells were enhanced by addition of either S1P or FTY. Both S1P and FTY enhanced BMP-2-induced ERK1/2 and Smad1/5/8 phosphorylation. The effect of FTY was stronger than that of S1P. S1P receptor-mediated signaling on osteoblast differentiation was inhibited by addition of mitogen-activated protein kinase/ERK kinase (MEK) 1/2 inhibitor, indicating that the S1P receptor-mediated MEK1/2-ERK1/2 signaling pathway enhanced BMP-2-Smad signaling. These results indicate that S1P receptor-mediated signaling plays a crucial role for osteoblast differentiation.

  1. Black ginseng activates Akt signaling, thereby enhancing myoblast differentiation and myotube growth

    Directory of Open Access Journals (Sweden)

    Soo-Yeon Lee

    2018-01-01

    Conclusion: BG enhances myoblast differentiation and myotube hypertrophy by activating Akt/mTOR/p70S6k axis. Thus, our study demonstrates that BG has promising potential to treat or prevent muscle loss related to aging or other pathological conditions, such as diabetes.

  2. Self-Biased Differential Rectifier with Enhanced Dynamic Range for Wireless Powering

    KAUST Repository

    Ouda, Mahmoud H.; Khalil, Waleed; Salama, Khaled N.

    2016-01-01

    A self-biased, cross-coupled, differential rectifier is proposed with enhanced power-conversion efficiency over an extended range of input power. A prototype is designed for UHF 433MHz RF power-harvesting applications and is implemented using 0.18μm

  3. Enhanced computational methods for quantifying the effect of geographic and environmental isolation on genetic differentiation

    NARCIS (Netherlands)

    Botta, Filippo; Eriksen, Casper; Fontaine, Michael Christophe; Guillot, Gilles

    2015-01-01

    In a recent paper, Bradburd et al. (2013) proposed a model to quantify the relative effect ofgeographic and environmental distance on genetic differentiation. Here, we enhance this method in several ways. 1. We modify the covariance model so as to fit better with mainstream geostatistical models and

  4. Enhancement of multiple cranial and spinal nerves in vanishing white matter: expanding the differential diagnosis.

    Science.gov (United States)

    Eluvathingal Muttikkal, Thomas Jose; Montealegre, Denia Ramirez; Matsumoto, Julie Ann

    2018-03-01

    Abnormal cranial or spinal nerve contrast enhancement on MRI in cases of suspected pediatric leukodystrophy is recognized as an important clue to the diagnosis of either metachromatic leukodystrophy or globoid cell leukodystrophy (Krabbe disease). We report a case of genetically confirmed childhood vanishing white matter with enhancement of multiple cranial and spinal nerves in addition to the more typical intracranial findings. This case expands the limited differential diagnosis of cranial nerve or spinal nerve enhancement in cases of suspected leukodystrophy and may aid in more efficient work-up and earlier diagnosis of vanishing white matter.

  5. Inhibiting actin depolymerization enhances osteoblast differentiation and bone formation in human stromal stem cells

    DEFF Research Database (Denmark)

    Chen, Li; Shi, Kaikai; Frary, Charles

    2015-01-01

    Remodeling of the actin cytoskeleton through actin dynamics is involved in a number of biological processes, but its role in human stromal (skeletal) stem cells (hMSCs) differentiation is poorly understood. In the present study, we demonstrated that stabilizing actin filaments by inhibiting gene...... expression of the two main actin depolymerizing factors (ADFs): Cofilin 1 (CFL1) and Destrin (DSTN) in hMSCs, enhanced cell viability and differentiation into osteoblastic cells (OB) in vitro, as well as heterotopic bone formation in vivo. Similarly, treating hMSC with Phalloidin, which is known to stabilize...... polymerized actin filaments, increased hMSCs viability and OB differentiation. Conversely, Cytocholasin D, an inhibitor of actin polymerization, reduced cell viability and inhibited OB differentiation of hMSC. At a molecular level, preventing Cofilin phosphorylation through inhibition of LIM domain kinase 1...

  6. Enhanced Differentiation of Human Embryonic Stem Cells Toward Definitive Endoderm on Ultrahigh Aspect Ratio Nanopillars

    DEFF Research Database (Denmark)

    Rasmussen, Camilla Holzmann; Reynolds, Paul M.; Petersen, Dorthe Roenn

    2016-01-01

    highlighted that the properties of the physical environment, such as substrate stiffness, affect cellular behavior. Here, mass-produced, injection molded polycarbonate nanopillars are presented, where the surface mechanical properties, i.e., stiffness, can be controlled by the geometric design...... of the ultrahigh aspect ratio nanopillars (stiffness can be reduced by 25.000X). It is found that tall nanopillars, yielding softer surfaces, significantly enhance the induction of defi nitive endoderm cells from pluripotent human embryonic stem cells, resulting in more consistent differentiation of a pure...... population compared to planar control. By contrast, further differentiation toward the pancreatic endoderm is less successful on “soft” pillars when compared to “stiff ” pillars or control, indicating differential cues during the different stages of differentiation. To accompany the mechanical properties...

  7. Differentiation of Enhancing Glioma and Primary Central Nervous System Lymphoma by Texture-Based Machine Learning.

    Science.gov (United States)

    Alcaide-Leon, P; Dufort, P; Geraldo, A F; Alshafai, L; Maralani, P J; Spears, J; Bharatha, A

    2017-06-01

    Accurate preoperative differentiation of primary central nervous system lymphoma and enhancing glioma is essential to avoid unnecessary neurosurgical resection in patients with primary central nervous system lymphoma. The purpose of the study was to evaluate the diagnostic performance of a machine-learning algorithm by using texture analysis of contrast-enhanced T1-weighted images for differentiation of primary central nervous system lymphoma and enhancing glioma. Seventy-one adult patients with enhancing gliomas and 35 adult patients with primary central nervous system lymphomas were included. The tumors were manually contoured on contrast-enhanced T1WI, and the resulting volumes of interest were mined for textural features and subjected to a support vector machine-based machine-learning protocol. Three readers classified the tumors independently on contrast-enhanced T1WI. Areas under the receiver operating characteristic curves were estimated for each reader and for the support vector machine classifier. A noninferiority test for diagnostic accuracy based on paired areas under the receiver operating characteristic curve was performed with a noninferiority margin of 0.15. The mean areas under the receiver operating characteristic curve were 0.877 (95% CI, 0.798-0.955) for the support vector machine classifier; 0.878 (95% CI, 0.807-0.949) for reader 1; 0.899 (95% CI, 0.833-0.966) for reader 2; and 0.845 (95% CI, 0.757-0.933) for reader 3. The mean area under the receiver operating characteristic curve of the support vector machine classifier was significantly noninferior to the mean area under the curve of reader 1 ( P = .021), reader 2 ( P = .035), and reader 3 ( P = .007). Support vector machine classification based on textural features of contrast-enhanced T1WI is noninferior to expert human evaluation in the differentiation of primary central nervous system lymphoma and enhancing glioma. © 2017 by American Journal of Neuroradiology.

  8. Osterix enhances proliferation and osteogenic potential of bone marrow stromal cells

    International Nuclear Information System (INIS)

    Tu Qisheng; Valverde, Paloma; Chen, Jake

    2006-01-01

    Osterix (Osx) is a zinc-finger-containing transcription factor that is expressed in osteoblasts of all endochondral and membranous bones. In Osx null mice osteoblast differentiation is impaired and bone formation is absent. In this study, we hypothesized that overexpression of Osx in murine bone marrow stromal cells (BMSC) would be able to enhance their osteoblastic differentiation and mineralization in vitro. Retroviral transduction of Osx in BMSC cultured in non-differentiating medium did not affect expression of Runx2/Cbfa1, another key transcription factor of osteoblast differentiation, but induced an increase in the expression of other markers associated with the osteoblastic lineage including alkaline phosphatase, bone sialoprotein, osteocalcin, and osteopontin. Retroviral transduction of Osx in BMSC also increased their proliferation, alkaline phosphatase activity, and ability to form bone nodules. These events occurred without significant changes in the expression of α1(II) procollagen or lipoprotein lipase, which are markers of chondrogenic and adipogenic differentiation, respectively

  9. Mediator Med23 deficiency enhances neural differentiation of murine embryonic stem cells through modulating BMP signaling.

    Science.gov (United States)

    Zhu, Wanqu; Yao, Xiao; Liang, Yan; Liang, Dan; Song, Lu; Jing, Naihe; Li, Jinsong; Wang, Gang

    2015-02-01

    Unraveling the mechanisms underlying early neural differentiation of embryonic stem cells (ESCs) is crucial to developing cell-based therapies of neurodegenerative diseases. Neural fate acquisition is proposed to be controlled by a 'default' mechanism, for which the molecular regulation is not well understood. In this study, we investigated the functional roles of Mediator Med23 in pluripotency and lineage commitment of murine ESCs. Unexpectedly, we found that, despite the largely unchanged pluripotency and self-renewal of ESCs, Med23 depletion rendered the cells prone to neural differentiation in different differentiation assays. Knockdown of two other Mediator subunits, Med1 and Med15, did not alter the neural differentiation of ESCs. Med15 knockdown selectively inhibited endoderm differentiation, suggesting the specificity of cell fate control by distinctive Mediator subunits. Gene profiling revealed that Med23 depletion attenuated BMP signaling in ESCs. Mechanistically, MED23 modulated Bmp4 expression by controlling the activity of ETS1, which is involved in Bmp4 promoter-enhancer communication. Interestingly, med23 knockdown in zebrafish embryos also enhanced neural development at early embryogenesis, which could be reversed by co-injection of bmp4 mRNA. Taken together, our study reveals an intrinsic, restrictive role of MED23 in early neural development, thus providing new molecular insights for neural fate determination. © 2015. Published by The Company of Biologists Ltd.

  10. Silk fibroin/gelatin-chondroitin sulfate-hyaluronic acid effectively enhances in vitro chondrogenesis of bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Sawatjui, Nopporn; Damrongrungruang, Teerasak; Leeanansaksiri, Wilairat; Jearanaikoon, Patcharee; Hongeng, Suradej; Limpaiboon, Temduang

    2015-01-01

    Tissue engineering is becoming promising for cartilage repair due to the limited self-repair capacity of cartilage tissue. We previously fabricated and characterized a three-dimensional silk fibroin/gelatin-chondroitin sulfate-hyaluronic acid (SF-GCH) scaffold and showed that it could promote proliferation of human bone marrow mesenchymal stem cells (BM-MSCs). This study aimed to evaluate its biological performance as a new biomimetic material for chondrogenic induction of BM-MSCs in comparison to an SF scaffold and conventional pellet culture. We found that the SF-GCH scaffold significantly enhanced the proliferation and chondrogenic differentiation of BM-MSCs compared to the SF scaffold and pellet culture in which the production of sulfated glycoaminoglycan was increased in concordance with the up-regulation of chondrogenic-specific gene markers. Our findings indicate the significant role of SF-GCH by providing a supportive structure and the mimetic cartilage environment for chondrogenesis which enables cartilage regeneration. Thus, our fabricated SF-GCH scaffold may serve as a potential biomimetic material for cartilage tissue engineering. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. SPIO-Enhanced MRI Findings of Well-Differentiated Hepatocellular Carcinomas: Correlation with MDCT Findings

    International Nuclear Information System (INIS)

    Kim, Seong Hyun; Lee, Won Jae; Lim, Hyo K.; Park, Cheol Keun

    2009-01-01

    This study was designed to assess superparamagnetic iron oxide (SPIO)-enhanced MRI findings of well-differentiated hepatocellular carcinomas (HCCs) correlated with their multidetector-row CT (MDCT) findings. Seventy-two patients with 84 pathologically proven well-differentiated HCCs underwent triple-phase MDCT and SPIO-enhanced MRI at a magnetic field strength of 1.5 Tesla (n = 49) and 3.0 Tesla (n = 23). Two radiologists in consensus retrospectively reviewed the CT and MR images for attenuation value and the signal intensity of each tumor. The proportion of hyperintense HCCs as depicted on SPIO-enhanced T2- or T2*-weighted images were compared in terms of tumor size ( 1 cm), five CT attenuation patterns based on arterial and equilibrium phases and magnetic field strength, by the use of univariate and multivariate analyses. Seventy-eight (93%) and 71 (85%) HCCs were identified by CT and on SPIO-enhanced T2- and T2*-weighted images, respectively. For the CT attenuation pattern, one (14%) of seven isodense-isodense, four (67%) of six hypodense- hypodense, four (80%) of five isodense-hypodense, 14 (88%) of 16 hyperdense- isodense and 48 (96%) of 50 hyperdense-hypodense HCCs were hyperintense (Cochran-Armitage test for trend, p 0.05). Most well-differentiated HCCs show hyperintensity on SPIOenhanced MRI, although the lesions show various CT attenuation patterns. The CT attenuation pattern is the main factor that affects the proportion of hyperintense well-differentiated HCCs as depicted on SPIO-enhanced MRI

  12. BAMBI Promotes C2C12 Myogenic Differentiation by Enhancing Wnt/β-Catenin Signaling

    Directory of Open Access Journals (Sweden)

    Qiangling Zhang

    2015-08-01

    Full Text Available Bone morphogenic protein and activin membrane-bound inhibitor (BAMBI is regarded as an essential regulator of cell proliferation and differentiation that represses transforming growth factor-β and enhances Wnt/β-catenin signaling in various cell types. However, its role in skeletal muscle remains largely unknown. In the current study, we found that the expression level of BAMBI peaked in the early differentiation phase of the C2C12 rodent myoblast cell line. Knockdown of BAMBI via siRNA inhibited C2C12 differentiation, indicated by repressed MyoD, MyoG, and MyHC expression as well as reductions in the differentiation and fusion indices. BAMBI knockdown reduced the activity of Wnt/β-catenin signaling, as characterized by the decreased nuclear translocation of β-catenin and the lowered transcription of Axin2, which is a well-documented target gene of the Wnt/β-catenin signaling pathway. Furthermore, treatment with LiCl, an activator of Wnt/β-catenin signaling, rescued the reduction in C2C12 differentiation caused by BAMBI siRNA. Taken together, our data suggest that BAMBI is required for normal C2C12 differentiation, and that its role in myogenesis is mediated by the Wnt/β-catenin pathway.

  13. Gadoxetate disodium-enhanced MR imaging: Differentiation between early-enhancing non-tumorous lesions and hypervascular hepatocellular carcinomas

    Energy Technology Data Exchange (ETDEWEB)

    Goshima, Satoshi, E-mail: gossy@par.odn.ne.jp [Department of Radiology, Gifu University Hospital, 1-1 Yanagido, Gifu 501-1193 (Japan); Kanematsu, Masayuki [Department of Radiology, Gifu University Hospital, 1-1 Yanagido, Gifu 501-1193 (Japan); Department of Radiology Services, Gifu University Hospital, 1-1 Yanagido, Gifu 501-1193 (Japan); Watanabe, Haruo; Kondo, Hiroshi; Mizuno, Nozomi; Kawada, Hiroshi [Department of Radiology, Gifu University Hospital, 1-1 Yanagido, Gifu 501-1193 (Japan); Shiratori, Yoshimune [Department of Medical Informatics, Gifu University School of Medicine, Gifu (Japan); Onozuka, Minoru [Department of Physiology and Neuroscience, Kanagawa Dental College, Yokosuka (Japan); Moriyama, Noriyuki [Research Center for Cancer Prevention and Screening, National Cancer Center Hospital, Tsukiji (Japan); Bae, Kyongtae T. [Department of Radiology, University of Pittsburgh Medical Center, Pittsburgh, PA (United States)

    2011-08-15

    Purpose: To retrospectively assess imaging features that help differentiate early-enhancing non-tumorous (EN) hepatic lesions from hepatocellular carcinomas (HCCs) on gadoxetate disodium-enhanced MR imaging. Materials and methods: Our institutional review board approved this retrospective study. We reviewed the studies of 158 patients (92 men and 65 women; age range: 29-91; mean age: 65.6 years) with chronic liver damage, who underwent gadoxetate disodium-enhanced MR imaging at 3T MR scanner. Hypervascular lesions identified during the hepatic artery phase were selected for a study cohort. The location, shape, size (maximum diameter and maximum area), and contrast enhancement signal intensity characteristics of the lesions were evaluated, then compared between the EN and HCC lesions. Results: A total of 65 EN lesions (range: 3-60 mm, mean: 13.6 {+-} 10.6 mm) from 35 patients and 33 HCCs (range: 9-61 mm, mean: 19.3 {+-} 12.6 mm) from 20 patients were identified. Lesions were more frequently round or oval in shape for HCCs (n = 29; 88%) than ENs (n = 26; 40%) (P < 0.01). Unexpectedly, some ENs (n = 12; 18%) showed hypointensity on hepatocyte-phase, and 6 (50%) of them were T2 hyperintense. For lesions smaller than 2 cm (9 ENs and 21 HCCs) on hepatic arterial-phase images, the mean area of hypointensity in hepatocyte-phase (54.2 {+-} 33.1 mm{sup 2}) was significantly smaller than those of the corresponding hyperintensity in hepatic arterial-phase (97.1 {+-} 42.0 mm{sup 2}) for EN lesions (P = 0.019), whereas no significant difference in area was found for HCCs. Conclusion: EN lesions may occasionally present with hypointensity during the hepatocyte-phase; presenting a diagnostic dilemma. In this situation, EN lesions may be differentiated from HCCs when a hypointense area in hepatocyte-phase is smaller than the corresponding hypervascular area in hepatic-arterial phase.

  14. Enhancement of Chiroptical Signals by Circular Differential Mie Scattering of Nanoparticles

    OpenAIRE

    SeokJae Yoo; Q-Han Park

    2015-01-01

    We enhance the weak optical signals of small chiral molecules via circular differential Mie scattering (CDMS) of nanoparticles immersed in them. CDMS is the preferential Mie scattering of left- and right-handed circularly polarized light by nanoparticles whose sizes are about the same as the wavelength of light. Solving the Mie scattering theory for chiral media, we find that the CDMS signal of the particle is linearly proportional to the chirality parameter κ of the molecules. This linear am...

  15. Collagen type II enhances chondrogenesis in adipose tissue-derived stem cells by affecting cell shape

    NARCIS (Netherlands)

    Lu, Z.; Doulabi, B.Z.; Huang, C.; Bank, R.A.; Helder, M.N.

    2010-01-01

    Ideally, biomaterials have inductive properties, favoring specific lineage differentiation. For chondrogenic induction, these properties have been attributed to collagen type II. However, the underlying mechanisms are largely unknown. This study aimed to investigate whether collagen type II favors

  16. Collagen Type II Enhances Chondrogenesis in Adipose Tissue-Derived Stem Cells by Affecting Cell Shape

    NARCIS (Netherlands)

    Lu, ZuFu; Doulabi, Behrouz Zandieh; Huang, ChunLing; Bank, Ruud A.; Helder, Marco N.

    Ideally, biomaterials have inductive properties, favoring specific lineage differentiation. For chondrogenic induction, these properties have been attributed to collagen type II. However, the underlying mechanisms are largely unknown. This study aimed to investigate whether collagen type II favors

  17. A paper-based scaffold for enhanced osteogenic differentiation of equine adipose-derived stem cells.

    Science.gov (United States)

    Petersen, Gayle F; Hilbert, Bryan J; Trope, Gareth D; Kalle, Wouter H J; Strappe, Padraig M

    2015-11-01

    We investigated the applicability of single layer paper-based scaffolds for the three-dimensional (3D) growth and osteogenic differentiation of equine adipose-derived stem cells (EADSC), with comparison against conventional two-dimensional (2D) culture on polystyrene tissue culture vessels. Viable culture of EADSC was achieved using paper-based scaffolds, with EADSC grown and differentiated in 3D culture retaining high cell viability (>94 %), similarly to EADSC in 2D culture. Osteogenic differentiation of EADSC was significantly enhanced in 3D culture, with Alizarin Red S staining and quantification demonstrating increased mineralisation (p < 0.0001), and an associated increase in expression of the osteogenic-specific markers alkaline phosphatase (p < 0.0001), osteopontin (p < 0.0001), and runx2 (p < 0.01). Furthermore, scanning electron microscopy revealed a spherical morphology of EADSC in 3D culture, compared to a flat morphology of EADSC in 2D culture. Single layer paper-based scaffolds provide an enhanced environment for the in vitro 3D growth and osteogenic differentiation of EADSC, with high cell viability, and a spherical morphology.

  18. Transudative vs exudative pleural effusions: differentiation using Gd-DTPA-enhanced MRI

    International Nuclear Information System (INIS)

    Frola, C.; Cantoni, S.; Turtulici, I.; Leoni, C.; Loria, F.; Gaeta, M.; Derchi, L.E.

    1997-01-01

    The aim of this study was to investigate the capability of Gd-DTPA-enhanced MRI to differentiate between exudative and transudative pleural effusions. An MRI examination was performed on 22 patients with different types of pleural effusion (10 transudative and 12 exudative effusions). T1-weighted SE images were obtained before and 20 min after administration of Gd-DTPA (0.1 mmol/kg). The degree of enhancement of pleural effusions was evaluated both by visual assessement and by quantitative analysis of images. None of 10 transudative effusions showed significative enhancement, whereas 10 of 12 exudative effusions showed enhancement (sensitivity 83 %, specificity 100 %, positive predictive value 100 %). The postcontrast signal intensity ratios (SIRs) of exudates were significantly higher than corresponding precontrast ratios (P = 0.0109) and the postcontrast SIRs of exudates were significantly higher than those of transudates (P = 0.0300). Exudative pleural effusions show a significant enhancement following administration of Gd-DTPA. We presume that this may be caused by increased pleural permeability and more rapid passage of a large amount of Gd-DTPA from the blood into the pleural fluid in case of exudative effusions. In our limited group of patients, signal enhancement proved the presence of an exudative effusion. Absence of signal enhancement suggests a transudate, but does not exclude an exudate. (orig.). With 5 figs., 2 tabs

  19. Extrinsic factors promoting insulin producing cell-differentiation and insulin expression enhancement-hope for diabetics.

    Science.gov (United States)

    Dave, Shruti

    2013-11-01

    Diabetes mellitus (DM) is considered to be an autoimmune disorder leading to destruction of beta-cells resulting in to a loss of blood sugar control. Attempts using many pharmacological compositions including exogenous insulin have failed to show tight control of glycemia and associated manifestations. Stem cells are considered a potential tool for the supply of insulin-producing cells (IPC) generation in vitro. Stem cell differentiation in to pancreatic lineages requires influence of both intrinsic and extrinsic factors. Application of islet growth factors is considered to be potential for enhancement of beta-cell replication, function and survival. Use of certain extrinsic factors is known to facilitate expression of transcription factors known to be important for beta-cell differentiation and production of insulin enabling IPC generation. Hierarchies of secreted signals and transcription factors have been identified by studies from several laboratories that guide cell differentiation in to IPC. This knowledge provides insights for in vitro IPC differentiation from stem cells. Current advancement in medical knowledge promises an insulin independency for DM patients. The review sheds light on few specific extrinsic factors which facilitate differentiation of stem cells in to IPC in vitro have been discussed; which can be proven as a potential therapeutic option for treatment of DM and associated diseases.

  20. Differentiation of recurrent breast cancer from radiation fibrosis with dynamic gadolinium-enhanced MR imaging

    International Nuclear Information System (INIS)

    Dao, T.H.; Campana, F.; Fourquet, A.; Rahmouni, A.

    1991-01-01

    This paper assesses the ability of dynamic gadolinium-enhanced MR imaging to differentiate radiation fibrosis from tumor recurrence of breast cancer after conservative treatment. Twenty-five women with previous breast cancer treated with radiation therapy underwent MR imaging examination. Tumor recurrence was suspected on palpation of masses (18 cases) or at mammography (7 cases). The MR imaging protocol was performed on a 0.5-T imager with a breast coil and included T1 and T2 spin-echo, short To inversion recovery (STIR), and dynamic gadolinium-enhanced T1-weighted sequenced to evaluate the hemokinetics of the lesion. Ratios of signal intensity of suspected lesions to that of fat, surrounding breast gland, and background noise were calculated. Percutaneous biopsies were performed in all cases after MR imaging. Curves of signal-to-noise ratio of recurrences (5 cases) showed an early enhancement within the first minutes after injection, although localized fibrosis (20 cases) was not significantly enhanced. T2 and STIR sequences were not contributive in differentiating fibrosis from tumor recurrence

  1. Differentiation between tuberculosis and primary tumors in the adrenal gland: evaluation with contrast-enhanced CT

    International Nuclear Information System (INIS)

    Yang, Zhi-Gang; Guo, Ying-Kun; Li, Yuan; Min, Peng-Qiu; Yu, Jian-Qun; Ma, En-Sen

    2006-01-01

    The aim of the present study is to determine imaging criteria for differentiating tuberculosis from primary tumors in the adrenal gland on contrast-enhanced CT. Non-contrast and contrast-enhanced CT features in 108 patients with adrenal tuberculosis (n=34) and primary tumor (n=74) were retrospectively assessed for the location, size, calcification and enhancement patterns. The primary tumors included 41 adenomas, 11 pheochromocytomas, 4 carcinomas, 3 lymphomas, 6 myelolipomas, 6 ganglioneuromas, 2 neurilemmomas and 1 ganglioneuroblastoma. Biochemical investigation was performed for all patients. Of the tuberculosis cases, 31 (91%) invaded with bilateral involvement, while 7 (9%) of the primary tumors invaded with bilateral involvement (P<0.001). Tuberculosis often showed calcification (20 of 34; 59%), whereas primary tumors infrequently showed calcification (6 of 74; 8%; P<0.001). Low attenuation in the center with peripheral rim enhancement was more commonly seen in tuberculosis (16 of 34; 47%) than in primary tumors (7 of 74; 9%; P<0.001). In the determination of tuberculosis, the highest sensitivity (91%) and accuracy (91%) were obtained with bilateral involvement, and the highest specificity (99%) was obtained with the contour preserved. In the determination of primary tumors using a combination of having unilateral involvement and being mass-like, the outcome was a sensitivity of 91%, specificity of 94% and accuracy of 92%. CT findings can differentiate tuberculosis from a primary tumor of the adrenal glands with high sensitivity and an acceptable specificity when combined with the endocrinological examination. (orig.)

  2. Enhancement of Chiroptical Signals by Circular Differential Mie Scattering of Nanoparticles.

    Science.gov (United States)

    Yoo, SeokJae; Park, Q-Han

    2015-09-25

    We enhance the weak optical signals of small chiral molecules via circular differential Mie scattering (CDMS) of nanoparticles immersed in them. CDMS is the preferential Mie scattering of left- and right-handed circularly polarized light by nanoparticles whose sizes are about the same as the wavelength of light. Solving the Mie scattering theory for chiral media, we find that the CDMS signal of the particle is linearly proportional to the chirality parameter κ of the molecules. This linear amplitude enhancement by CDMS of the particle holds, even for large particles, which have a retardation effect. We also demonstrate that the CDMS of a nanoparticle is sensitive to changes of molecular concentration, and that the nanoparticle can be utilized as a chiroptical biosensor detecting the concentration of analyte. We expect that the enhancement of molecular chiroptical signals by CDMS will pave the way for novel chiroptical spectroscopy using nanostructures.

  3. Exogenous transforming growth factor-β1 enhances smooth muscle differentiation in embryonic mouse jejunal explants.

    Science.gov (United States)

    Coletta, Riccardo; Roberts, Neil A; Randles, Michael J; Morabito, Antonino; Woolf, Adrian S

    2017-01-13

    An ex vivo experimental strategy that replicates in vivo intestinal development would in theory provide an accessible setting with which to study normal and dysmorphic gut biology. The current authors recently described a system in which mouse embryonic jejunal segments were explanted onto semipermeable platforms and fed with chemically defined serum-free media. Over 3 days in organ culture, explants formed villi and they began to undergo spontaneous peristalsis. As defined in the current study, the wall of the explanted gut failed to form a robust longitudinal smooth muscle (SM) layer as it would do in vivo over the same time period. Given the role of transforming growth factor β1 (TGFβ1) in SM differentiation in other organs, it was hypothesized that exogenous TGFβ1 would enhance SM differentiation in these explants. In vivo, TGFβ receptors I and II were both detected in embryonic longitudinal jejunal SM cells and, in organ culture, exogenous TGFβ1 induced robust differentiation of longitudinal SM. Microarray profiling showed that TGFβ1 increased SM specific transcripts in a dose dependent manner. TGFβ1 proteins were detected in amniotic fluid at a time when the intestine was physiologically herniated. By analogy with the requirement for exogenous TGFβ1 for SM differentiation in organ culture, the TGFβ1 protein that was demonstrated to be present in the amniotic fluid may enhance intestinal development when it is physiologically herniated in early gestation. Future studies of embryonic intestinal cultures should include TGFβ1 in the defined media to produce a more faithful model of in vivo muscle differentiation. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd. Copyright © 2017 The Authors Journal of Tissue Engineering and Regenerative Medicine Published by John Wiley & Sons, Ltd.

  4. Gadoxetic acid enhanced MRI for differentiation of FNH and HCA: a single centre experience

    Energy Technology Data Exchange (ETDEWEB)

    Grieser, Christian; Steffen, Ingo G.; Perez Fernandez, Carmen Maria; Hamm, Bernd; Denecke, Timm [Klinik fuer Radiologie, Campus Virchow-Klinikum, Charite - Universitaetsmedizin Berlin, Berlin (Germany); Kramme, Incken-Birthe; Blaeker, Hendrik; Kilic, Ergin [Institut fuer Pathologie, Campus Virchow-Klinikum, Charite - Universitaetsmedizin Berlin, Berlin (Germany); Seehofer, Daniel [Klinik fuer Allgemein, Viszeral- und Transplantationschirurgie, Campus Virchow-Klinikum, Charite - Universitaetsmedizin Berlin, Berlin (Germany); Schott, Eckart [Medizinische Klinik m.S. Hepatologie und Gastroenterologie, Campus Virchow-Klinikum, Charite - Universitaetsmedizin Berlin, Berlin (Germany)

    2014-06-15

    Evaluation of enhancement characteristics of histopathologically confirmed focal nodular hyperplasias (FNHs) and hepatocellular adenomas (HCAs) with gadoxetic acid-enhanced MRI. Sixty-eight patients with 115 histopathologically proven lesions (FNHs, n = 44; HCAs, n = 71) examined with gadoxetic acid-enhanced MRI were retrospectively enrolled (standard of reference: surgical resection, n = 53 patients (lesions: FNHs, n = 37; HCAs, n = 53); biopsy, n = 15 (lesions: FNHs, n = 7; HCAs, n = 18)). Two radiologists evaluated all MR images regarding morphological features as well as the vascular and hepatocyte-specific enhancement in consensus. For the hepatobiliary phase, relative enhancement of the lesions and lesion to liver enhancement were significantly lower for HCAs (mean, 48.7 (±48.4) % and 49.4 (±33.9) %) compared to FNHs (159.3 (±92.5) %; and 151.7 (±79) %; accuracy of 89 % and 90 %, respectively; P < 0.001). Visual strong uptake of FNHs vs. hypointensity of HCAs in the hepatobiliary phase resulted in an accuracy of 92 %. This parameter was superior to all other morphological and dynamic vascular criteria alone and in combination (accuracy, 54-85 %). For differentiation of FNHs and HCAs by means of MRI, gadoxetic acid uptake in the hepatobiliary phase was found to be superior to all other criteria alone and in combination. (orig.)

  5. Gadoxetic acid enhanced MRI for differentiation of FNH and HCA: a single centre experience

    International Nuclear Information System (INIS)

    Grieser, Christian; Steffen, Ingo G.; Perez Fernandez, Carmen Maria; Hamm, Bernd; Denecke, Timm; Kramme, Incken-Birthe; Blaeker, Hendrik; Kilic, Ergin; Seehofer, Daniel; Schott, Eckart

    2014-01-01

    Evaluation of enhancement characteristics of histopathologically confirmed focal nodular hyperplasias (FNHs) and hepatocellular adenomas (HCAs) with gadoxetic acid-enhanced MRI. Sixty-eight patients with 115 histopathologically proven lesions (FNHs, n = 44; HCAs, n = 71) examined with gadoxetic acid-enhanced MRI were retrospectively enrolled (standard of reference: surgical resection, n = 53 patients (lesions: FNHs, n = 37; HCAs, n = 53); biopsy, n = 15 (lesions: FNHs, n = 7; HCAs, n = 18)). Two radiologists evaluated all MR images regarding morphological features as well as the vascular and hepatocyte-specific enhancement in consensus. For the hepatobiliary phase, relative enhancement of the lesions and lesion to liver enhancement were significantly lower for HCAs (mean, 48.7 (±48.4) % and 49.4 (±33.9) %) compared to FNHs (159.3 (±92.5) %; and 151.7 (±79) %; accuracy of 89 % and 90 %, respectively; P < 0.001). Visual strong uptake of FNHs vs. hypointensity of HCAs in the hepatobiliary phase resulted in an accuracy of 92 %. This parameter was superior to all other morphological and dynamic vascular criteria alone and in combination (accuracy, 54-85 %). For differentiation of FNHs and HCAs by means of MRI, gadoxetic acid uptake in the hepatobiliary phase was found to be superior to all other criteria alone and in combination. (orig.)

  6. Adhesive and mechanical regulation of mesenchymal stem cell differentiation in human bone marrow and periosteum-derived progenitor cells

    Directory of Open Access Journals (Sweden)

    Jeroen Eyckmans

    2012-08-01

    It has previously been demonstrated that cell shape can influence commitment of human bone marrow-derived mesenchymal stem cells (hBMCs to adipogenic, osteogenic, chondrogenic, and other lineages. Human periosteum-derived cells (hPDCs exhibit multipotency similar to hBMCs, but hPDCs may offer enhanced potential for osteogenesis and chondrogenesis given their apparent endogenous role in bone and cartilage repair in vivo. Here, we examined whether hPDC differentiation is regulated by adhesive and mechanical cues comparable to that reported for hBMC differentiation. When cultured in the appropriate induction media, hPDCs at high cell seeding density demonstrated enhanced levels of adipogenic or chondrogenic markers as compared with hPDCs at low cell seeding density. Cell seeding density correlated inversely with projected area of cell spreading, and directly limiting cell spreading with micropatterned substrates promoted adipogenesis or chondrogenesis while substrates promoting cell spreading supported osteogenesis. Interestingly, cell seeding density influenced differentiation through both changes in cell shape and non-shape-mediated effects: density-dependent adipogenesis and chondrogenesis were regulated primarily by cell shape whereas non-shape effects strongly influenced osteogenic potential. Inhibition of cytoskeletal contractility by adding the Rho kinase inhibitor Y27632 further enhanced adipogenic differentiation and discouraged osteogenic differentiation of hPDCs. Together, our results suggest that multipotent lineage decisions of hPDCs are impacted by cell adhesive and mechanical cues, though to different extents than hBMCs. Thus, future studies of hPDCs and other primary stem cell populations with clinical potential should consider varying biophysical metrics for more thorough optimization of stem cell differentiation.

  7. ESC-Derived BDNF-Overexpressing Neural Progenitors Differentially Promote Recovery in Huntington's Disease Models by Enhanced Striatal Differentiation

    Directory of Open Access Journals (Sweden)

    Tina Zimmermann

    2016-10-01

    Full Text Available Huntington's disease (HD is characterized by fatal motoric failures induced by loss of striatal medium spiny neurons. Neuronal cell death has been linked to impaired expression and axonal transport of the neurotrophin BDNF (brain-derived neurotrophic factor. By transplanting embryonic stem cell-derived neural progenitors overexpressing BDNF, we combined cell replacement and BDNF supply as a potential HD therapy approach. Transplantation of purified neural progenitors was analyzed in a quinolinic acid (QA chemical and two genetic HD mouse models (R6/2 and N171-82Q on the basis of distinct behavioral parameters, including CatWalk gait analysis. Explicit rescue of motor function by BDNF neural progenitors was found in QA-lesioned mice, whereas genetic mouse models displayed only minor improvements. Tumor formation was absent, and regeneration was attributed to enhanced neuronal and striatal differentiation. In addition, adult neurogenesis was preserved in a BDNF-dependent manner. Our findings provide significant insight for establishing therapeutic strategies for HD to ameliorate neurodegenerative symptoms.

  8. Decreased hypertrophic differentiation accompanies enhanced matrix formation in co-cultures of outer meniscus cells with bone marrow mesenchymal stromal cells

    Science.gov (United States)

    2012-01-01

    Introduction The main objective of this study was to determine whether meniscus cells from the outer (MCO) and inner (MCI) regions of the meniscus interact similarly to or differently with mesenchymal stromal stem cells (MSCs). Previous study had shown that co-culture of meniscus cells with bone marrow-derived MSCs result in enhanced matrix formation relative to mono-cultures of meniscus cells and MSCs. However, the study did not examine if cells from the different regions of the meniscus interacted similarly to or differently with MSCs. Methods Human menisci were harvested from four patients undergoing total knee replacements. Tissue from the outer and inner regions represented pieces taken from one third and two thirds of the radial distance of the meniscus, respectively. Meniscus cells were released from the menisci after collagenase treatment. Bone marrow MSCs were obtained from the iliac crest of two patients after plastic adherence and in vitro culture until passage 2. Primary meniscus cells from the outer (MCO) or inner (MCI) regions of the meniscus were co-cultured with MSCs in three-dimensional (3D) pellet cultures at 1:3 ratio, respectively, for 3 weeks in the presence of serum-free chondrogenic medium containing TGF-β1. Mono-cultures of MCO, MCI and MSCs served as experimental control groups. The tissue formed after 3 weeks was assessed biochemically, histochemically and by quantitative RT-PCR. Results Co-culture of inner (MCI) or outer (MCO) meniscus cells with MSCs resulted in neo-tissue with increased (up to 2.2-fold) proteoglycan (GAG) matrix content relative to tissues formed from mono-cultures of MSCs, MCI and MCO. Co-cultures of MCI or MCO with MSCs produced the same amount of matrix in the tissue formed. However, the expression level of aggrecan was highest in mono-cultures of MSCs but similar in the other four groups. The DNA content of the tissues from co-cultured cells was not statistically different from tissues formed from mono-cultures of

  9. Differentiation of benign and malignant solitary pulmonary nodules : value of contrast-enhanced dynamic MR imaging

    International Nuclear Information System (INIS)

    Kim, Jeong Ho; Kim, Hyung Jin; Han, Heon; Lee, Hong Lyeol; Kim, Kwang Ho; Suh, Chang Hae

    1999-01-01

    To evaluate the usefulness of contrast-enhanced dynamic MR imaging for differentiation of benign and malignant solitary pulmonary nodules (SPNs). Twenty-three patients with histologically or radiologically provened SPNs smaller than 40mm (14 benign, 9 malignant) underwent MR examination using the breath-hold fast multiplanar spoiled gradient echo (FMPSPGR) technique. Pre-enhancement MR examination was followed by serial scans obtained at one-minute intervals, beginning one-minute after the onset of bolus injection of paramagnetic contrast agent for a total of five scans. Signal intensities of SPNs were measured from pre- and post-contrast enhanced MR images and peak percentage increase in signal intensity (p%SI) was calculated. Mean percentage increase in signal intensity (m%SI) was also calculated and the time-m%SI curve was plotted. The enhancement patterns of SPNs were classified as homogeneous, peripheral rim-like, inhomogeneous, or no (or minimal) enhancement. We compared differences in p%SI, the pattern of the time-m%SI curve, and the pattern of enhancement between benign and malignant SPNs. On dynamic MR images, alignant SPNs (n=9) showed a significantly higher p%SI than benign SPNs (n=14) (malignant : mean 120.6, range 81.8-171.6; benign : mean 29.5, range 3.7-78.9)(p<0.0001). With 80 p%SI as the threshold for malignancy-positive, both sensitivity and specificity were 100%. The m%SI of malignant SPNs rapidly increased at one minute after enhancement and decreased gradually thereafter, whereas that of benign SPNs increased more slowly to form a plateau. Eighty-nine percent (8/9) of malignant SPNs showed homogeneous enhancement. In contrast, among benign SPNs, peripheral rim-like enhancement and no (or minimal) enhancement occurred in the same proportion of cases : 50%(7/14). The superb demonstration of different enhancement characteristics obtained using dynamic contrast-enhanced MR imaging is useful to discriminate malignant from benign SPNs

  10. Apparent diffusion coefficient values and dynamic contrast enhancement patterns in differentiating seminomas from nonseminomatous testicular neoplasms

    International Nuclear Information System (INIS)

    Tsili, Athina C.; Sylakos, Anastasios; Ntorkou, Alexandra; Stavrou, Sotirios; Astrakas, Loukas G.; Sofikitis, Nikolaos; Argyropoulou, Maria I.

    2015-01-01

    Highlights: • Functional MRI in the characterization of testicular germ cell tumors was assessed. • ADC values proved useful in the characterization of testicular germ cell tumors. • Testicular germ cell tumors had similar enhancement patterns of dynamic MRI. - Abstract: Introduction: The aim of this study is to investigate the role of apparent diffusion coefficient (ADC) values and dynamic contrast enhancement (DCE) patterns in differentiating seminomas from nonseminomatous germ cell tumors (NSGCTs). Materials and methods: The MRI examinations of the scrotum of 26 men with histologically proven testicular GCTs were reviewed. DWI was performed in all patients, using a single shot, multi-slice spin-echo planar diffusion pulse sequence and b-values of 0 and 900 s/mm 2 . Subtraction DCE-MRI was performed in 20 cases using a 3D fast-field echo sequence after gadolinium administration. Time-signal intensity curves were created and semi-quantitative parameters (peak enhancement, time to peak, wash-in and wash-out rate) were calculated. The Student's t-test was used to compare the mean values of ADC, peak enhancement, time to peak, wash-in and wash-out rate between seminomas and NSGCTs. ROC analysis was also performed. Results: Histopathology disclosed the presence of 15 seminomas and 11 NSGCTs. The mean ± s.d. of ADC values (× 10 −3 mm 2 /s) of seminomas (0.59 ± 0.009) were significantly lower than those of NSGCTs (0.90 ± 0.33) (P = 0.01). The optimal ADC cut-off value was 0.68 × 10 −3 mm 2 /s. No differences between the two groups were observed for peak enhancement (P = 0.18), time to peak (P = 0.63) wash-in rate (P = 0.32) and wash-out rate (P = 0.18). Conclusions: ADC values may be used to preoperatively differentiate seminomas from NSGCTs

  11. Differentiation and fiber type-specific activity of a muscle creatine kinase intronic enhancer

    Directory of Open Access Journals (Sweden)

    Tai Phillip WL

    2011-07-01

    Full Text Available Abstract Background Hundreds of genes, including muscle creatine kinase (MCK, are differentially expressed in fast- and slow-twitch muscle fibers, but the fiber type-specific regulatory mechanisms are not well understood. Results Modulatory region 1 (MR1 is a 1-kb regulatory region within MCK intron 1 that is highly active in terminally differentiating skeletal myocytes in vitro. A MCK small intronic enhancer (MCK-SIE containing a paired E-box/myocyte enhancer factor 2 (MEF2 regulatory motif resides within MR1. The SIE's transcriptional activity equals that of the extensively characterized 206-bp MCK 5'-enhancer, but the MCK-SIE is flanked by regions that can repress its activity via the individual and combined effects of about 15 different but highly conserved 9- to 24-bp sequences. ChIP and ChIP-Seq analyses indicate that the SIE and the MCK 5'-enhancer are occupied by MyoD, myogenin and MEF2. Many other E-boxes located within or immediately adjacent to intron 1 are not occupied by MyoD or myogenin. Transgenic analysis of a 6.5-kb MCK genomic fragment containing the 5'-enhancer and proximal promoter plus the 3.2-kb intron 1, with and without MR1, indicates that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers (types I and IIa, respectively, but is not required for expression in fast-twitch muscle fibers (types IIb and IId. Conclusions In this study, we discovered that MR1 is critical for MCK expression in slow- and intermediate-twitch muscle fibers and that MR1's positive transcriptional activity depends on a paired E-box MEF2 site motif within a SIE. This is the first study to delineate the DNA controls for MCK expression in different skeletal muscle fiber types.

  12. Chaos Enhanced Differential Evolution in the Task of Evolutionary Control of Discrete Chaotic LOZI Map

    Directory of Open Access Journals (Sweden)

    Roman Senkerik

    2016-01-01

    Full Text Available In this paper, evolutionary technique Differential Evolution (DE is used for the evolutionary tuning of controller parameters for the stabilization of selected discrete chaotic system, which is the two-dimensional Lozi map. The novelty of the approach is that the selected controlled discrete dissipative chaotic system is used within Chaos enhanced heuristic concept as the chaotic pseudo-random number generator to drive the mutation and crossover process in the DE. The idea was to utilize the hidden chaotic dynamics in pseudo-random sequences given by chaotic map to help Differential evolution algorithm in searching for the best controller settings for the same chaotic system. The optimizations were performed for three different required final behavior of the chaotic system, and two types of developed cost function. To confirm the robustness of presented approach, comparisons with canonical DE strategy and PSO algorithm have been performed.

  13. Gold nanoparticles in injectable calcium phosphate cement enhance osteogenic differentiation of human dental pulp stem cells.

    Science.gov (United States)

    Xia, Yang; Chen, Huimin; Zhang, Feimin; Bao, Chongyun; Weir, Michael D; Reynolds, Mark A; Ma, Junqing; Gu, Ning; Xu, Hockin H K

    2018-01-01

    In this study, a novel calcium phosphate cement containing gold nanoparticles (GNP-CPC) was developed. Its osteogenic induction ability on human dental pulp stem cells (hDPSCs) was investigated for the first time. The incorporation of GNPs improved hDPSCs behavior on CPC, including better cell adhesion (about 2-fold increase in cell spreading) and proliferation, and enhanced osteogenic differentiation (about 2-3-fold increase at 14 days). GNPs endow CPC with micro-nano-structure, thus improving surface properties for cell adhesion and subsequent behaviors. In addition, GNPs released from GNP-CPC were internalized by hDPSCs, as verified by transmission electron microscopy (TEM), thus enhancing cell functions. The culture media containing GNPs enhanced the cellular activities of hDPSCs. This result was consistent with and supported the osteogenic induction results of GNP-CPC. In conclusion, GNP-CPC significantly enhanced the osteogenic functions of hDPSCs. GNPs are promising to modify CPC with nanotopography and work as bioactive additives thus enhance bone regeneration. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. The role of DNA methylation in catechol-enhanced erythroid differentiation of K562 cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xiao-Fei; Wu, Xiao-Rong; Xue, Ming; Wang, Yan; Wang, Jie; Li, Yang; Suriguga,; Zhang, Guang-Yao; Yi, Zong-Chun, E-mail: yizc@buaa.edu.cn

    2012-11-15

    Catechol is one of phenolic metabolites of benzene in vivo. Catechol is also widely used in pharmaceutical and chemical industries. In addition, fruits, vegetables and cigarette smoke also contain catechol. Our precious study showed that several benzene metabolites (phenol, hydroquinone, and 1,2,4-benzenetriol) inhibited erythroid differentiation of K562 cells. In present study, the effect of catechol on erythroid differentiation of K562 cells was investigated. Moreover, to address the role of DNA methylation in catechol-induced effect on erythroid differentiation in K562 cells, methylation levels of erythroid-specific genes were analyzed by Quantitative MassARRAY methylation analysis platform. Benzidine staining showed that exposure to catechol enhanced hemin-induced hemoglobin accumulation in K562 cells in concentration- and time-dependent manners. The mRNA expression of erythroid specific genes, including α-globin, β-globin, γ-globin, erythroid 5-aminolevulinate synthase, erythroid porphobilinogen deaminase, and transcription factor GATA-1 genes, showed a significant concentration-dependent increase in catechol-treated K562 cells. The exposure to catechol caused a decrease in DNA methylation levels at a few CpG sites in some erythroid specific genes including α-globin, β-globin and erythroid porphobilinogen deaminase genes. These results indicated that catechol improved erythroid differentiation potency of K562 cells at least partly via up-regulating transcription of some erythroid related genes, and suggested that inhibition of DNA methylation might be involved in up-regulated expression of some erythroid related genes. -- Highlights: ► Catechol enhanced hemin-induced hemoglobin accumulation. ► Exposure to catechol resulted in up-regulated expression of erythroid genes. ► Catechol reduced methylation levels at some CpG sites in erythroid genes.

  15. The role of DNA methylation in catechol-enhanced erythroid differentiation of K562 cells

    International Nuclear Information System (INIS)

    Li, Xiao-Fei; Wu, Xiao-Rong; Xue, Ming; Wang, Yan; Wang, Jie; Li, Yang; Suriguga,; Zhang, Guang-Yao; Yi, Zong-Chun

    2012-01-01

    Catechol is one of phenolic metabolites of benzene in vivo. Catechol is also widely used in pharmaceutical and chemical industries. In addition, fruits, vegetables and cigarette smoke also contain catechol. Our precious study showed that several benzene metabolites (phenol, hydroquinone, and 1,2,4-benzenetriol) inhibited erythroid differentiation of K562 cells. In present study, the effect of catechol on erythroid differentiation of K562 cells was investigated. Moreover, to address the role of DNA methylation in catechol-induced effect on erythroid differentiation in K562 cells, methylation levels of erythroid-specific genes were analyzed by Quantitative MassARRAY methylation analysis platform. Benzidine staining showed that exposure to catechol enhanced hemin-induced hemoglobin accumulation in K562 cells in concentration- and time-dependent manners. The mRNA expression of erythroid specific genes, including α-globin, β-globin, γ-globin, erythroid 5-aminolevulinate synthase, erythroid porphobilinogen deaminase, and transcription factor GATA-1 genes, showed a significant concentration-dependent increase in catechol-treated K562 cells. The exposure to catechol caused a decrease in DNA methylation levels at a few CpG sites in some erythroid specific genes including α-globin, β-globin and erythroid porphobilinogen deaminase genes. These results indicated that catechol improved erythroid differentiation potency of K562 cells at least partly via up-regulating transcription of some erythroid related genes, and suggested that inhibition of DNA methylation might be involved in up-regulated expression of some erythroid related genes. -- Highlights: ► Catechol enhanced hemin-induced hemoglobin accumulation. ► Exposure to catechol resulted in up-regulated expression of erythroid genes. ► Catechol reduced methylation levels at some CpG sites in erythroid genes.

  16. Dynamic contrast enhanced MRI in the differential diagnosis of adrenal adenomas and malignant adrenal masses

    International Nuclear Information System (INIS)

    Inan, Nagihan; Arslan, Arzu; Akansel, Gur; Anik, Yonca; Balci, N. Cem; Demirci, Ali

    2008-01-01

    Objective: To evaluate the value of dynamic MR imaging in the differential diagnosis of adrenal adenomas and malignant tumors, especially in cases with atypical adenomas. Materials and methods: Sixty-four masses (48 adenomas, 16 malignant tumors) were included in this prospective study. Signal loss of masses was evaluated using chemical shift MR imaging. Five dynamic series of T1-weighted spoiled gradient echo (FFE) images were obtained, with the acquisition starting simultaneously with i.v. contrast administration (0-100 s) followed by a T1-weighted FFE sequence in the late phase (5th minute). Contrast enhancement patterns in the early (25th second) and late (5th minute) phase images were evaluated. For the quantitative evaluation, signal intensity (SI)-time curves were obtained according to the SIs on the 0th, 25th, 50th 75th and 100th second. Also, the wash-in rate, maximum relative enhancement, time-to-peak, and wash-out of contrast at 100 s of masses in both groups were calculated. The statistical significance was determined by Mann-Whitney U test. To evaluate the diagnostic performance of the quantitative tests, receiver operating characteristic (ROC) analysis was performed. Results: Chemical shift MR imaging was able to differentiate 44 out of 48 adenomas (91.7%) from non-adenomas. The 4 adenomas (8.3%) which could not be differentiated from non-adenomas by this technique did not exhibit signal loss on out-of-phase images. With a cut-off value of 30, SI indices of adenomas had a sensitivity of 93.8%, specificity of 100% and a positive predictive value of 100%. On visual evaluation of dynamic MR imaging, early phase contrast enhancement patterns were homogeneous in 75% and punctate in 20,83% of the adenomas; while patchy in 56.25% and peripheral in 25% of the malignant tumors. On the late phase images 58.33% of the adenomas showed peripheral ring-shaped enhancement and 10.41% showed heterogeneous enhancement. All of the malignant masses showed heterogeneous

  17. Differentiation between early rheumatoid arthritis patients and healthy persons by conventional and dynamic contrast-enhanced magnetic resonance imaging

    DEFF Research Database (Denmark)

    Axelsen, Mette Bjørndal; Ejbjerg, B J; Hetland, M L

    2014-01-01

    OBJECTIVES: To identify the magnetic resonance imaging (MRI) parameter that best differentiates healthy persons and patients with early rheumatoid arthritis (RA), and to investigated responsiveness to treatment of various MRI parameters. METHOD: Conventional MRI and dynamic contrast-enhanced (DCE...

  18. A small-molecule/cytokine combination enhances hematopoietic stem cell proliferation via inhibition of cell differentiation.

    Science.gov (United States)

    Wang, Lan; Guan, Xin; Wang, Huihui; Shen, Bin; Zhang, Yu; Ren, Zhihua; Ma, Yupo; Ding, Xinxin; Jiang, Yongping

    2017-07-18

    Accumulated evidence supports the potent stimulating effects of multiple small molecules on the expansion of hematopoietic stem cells (HSCs) which are important for the therapy of various hematological disorders. Here, we report a novel, optimized formula, named the SC cocktail, which contains a combination of three such small molecules and four cytokines. Small-molecule candidates were individually screened and then combined at their optimal concentration with the presence of cytokines to achieve maximum capacity for stimulating the human CD34 + cell expansion ex vivo. The extent of cell expansion and the immunophenotype of expanded cells were assessed through flow cytometry. The functional preservation of HSC stemness was confirmed by additional cell and molecular assays in vitro. Subsequently, the expanded cells were transplanted into sublethally irradiated NOD/SCID mice for the assessment of human cell viability and engraftment potential in vivo. Furthermore, the expression of several genes in the cell proliferation and differentiation pathways was analyzed through quantitative polymerase chain reaction (qPCR) during the process of CD34 + cell expansion. The SC cocktail supported the retention of the immunophenotype of hematopoietic stem/progenitor cells remarkably well, by yielding purities of 86.6 ± 11.2% for CD34 + cells and 76.2 ± 10.5% for CD34 + CD38 - cells, respectively, for a 7-day culture. On day 7, the enhancement of expansion of CD34 + cells and CD34 + CD38 - cells reached a maxima of 28.0 ± 5.5-fold and 27.9 ± 4.3-fold, respectively. The SC cocktail-expanded CD34 + cells preserved the characteristics of HSCs by effectively inhibiting their differentiation in vitro and retained the multilineage differentiation potential in primary and secondary in vivo murine xenotransplantation trials. Further gene expression analysis suggested that the small-molecule combination strengthened the ability of the cytokines to enhance the Notch

  19. Enhanced computational methods for quantifying the effect of geographic and environmental isolation on genetic differentiation

    DEFF Research Database (Denmark)

    Botta, Filippo; Eriksen, Casper; Fontaine, Michaël C.

    2015-01-01

    1. In a recent paper, Bradburd et al. (Evolution, 67, 2013, 3258) proposed a model to quantify the relative effect of geographic and environmental distance on genetic differentiation. Here, we enhance this method in several ways. 2. We modify the covariance model so as to fit better with mainstre...... available as an R package called sunder. It takes as input georeferenced allele counts at the individual or population level for co-dominant markers. Program homepage: http://www2.imm.dtu.dk/~gigu/Sunder/....

  20. Effect of contrast-enhanced ultrasound on differential diagnosis of intrahepatic cholangiocarcinoma and arterial phase enhanced hepatic inflammatory lesions

    Institute of Scientific and Technical Information of China (English)

    Shanshan Yin; Qiuli Cui; Kun Yan; Wei Yang; Wei Wu; Liping Bao; Minhua Chen

    2017-01-01

    Objective:To investigate differential diagnosis between intrahepatic cholangiocarcinoma (ICC) and arterial phase enhanced hepatic inflammatory lesions in patients without liver cirrhosis using contrast-enhanced ultrasound (CEUS).Methods:ICC and hepatic inflammatory lesions cases with CEUS and pathological diagnosis between Sep 2013 and Oct 2016 were investigated retrospectively.Imaging features of conventional ultrasound and CEUS were analyzed.The parameters of time intensity curve (TIC),including the arrival time,peak intensity (PI) in the lesions,the starting time for washout,and the intensity difference at 3 min (△I3) after contrast agent infection between the lesion and the liver parenchyma,were compared between ICC and hepatic inflammatory lesions.Results:Twenty-five ICC and fifteen inflammatory patients were included in this study.Seventeen ICC (68.0%) and two inflammatory cases (13.3%) showed bile duct dilatation on conventional ultrasound.Using CEUS,three ICC cases (12.0%) were misdiagnosed as inflammatory lesions and three inflammatory lesions (20.0%) as ICC;two ICC (8.0%) and one inflammatory case (6.7%) could not be made definite diagnosis.Washout started at 34.5±3.5 s and 61.5± 12.9 s for ICC and inflammatory lesions respectively (P<0.001).The intensity difference between lesion and liver parenchyma at 3 min after contrast agent injection was 10.8±3.1 dB in ICC and 4.2±2.3 dB in inflammatory group (P<0.00 1).The sensitivity and specificity differentiating ICC and inflammatory lesions were 76% and 87% if the cut-offvalue of the intensity difference was 7.7 dB.Conclusions:Combined with TIC analysis,and particularly with the characteristic of the early-starting and obvious washout in ICC,CEUS can be useful in differential diagnosis between hepatic inflammatory lesions and ICC.

  1. Contrast-Enhanced Ultrasonography in Differential Diagnosis of Benign and Malignant Ovarian Tumors

    Science.gov (United States)

    Qiao, Jing-Jing; Yu, Jing; Yu, Zhe; Li, Na; Song, Chen; Li, Man

    2015-01-01

    Objective To evaluate the accuracy of contrast-enhanced ultrasonography (CEUS) in differential diagnosis of benign and malignant ovarian tumors. Methods The scientific literature databases PubMed, Cochrane Library and CNKI were comprehensively searched for studies relevant to the use of CEUS technique for differential diagnosis of benign and malignant ovarian cancer. Pooled summary statistics for specificity (Spe), sensitivity (Sen), positive and negative likelihood ratios (LR+/LR−), and diagnostic odds ratio (DOR) and their 95%CIs were calculated. Software for statistical analysis included STATA version 12.0 (Stata Corp, College Station, TX, USA) and Meta-Disc version 1.4 (Universidad Complutense, Madrid, Spain). Results Following a stringent selection process, seven high quality clinical trials were found suitable for inclusion in the present meta-analysis. The 7 studies contained a combined total of 375 ovarian cancer patients (198 malignant and 177 benign). Statistical analysis revealed that CEUS was associated with the following performance measures in differential diagnosis of ovarian tumors: pooled Sen was 0.96 (95%CI = 0.92∼0.98); the summary Spe was 0.91 (95%CI = 0.86∼0.94); the pooled LR+ was 10.63 (95%CI = 6.59∼17.17); the pooled LR− was 0.04 (95%CI = 0.02∼0.09); and the pooled DOR was 241.04 (95% CI = 92.61∼627.37). The area under the SROC curve was 0.98 (95% CI = 0.20∼1.00). Lastly, publication bias was not detected (t = −0.52, P = 0.626) in the meta-analysis. Conclusions Our results revealed the high clinical value of CEUS in differential diagnosis of benign and malignant ovarian tumors. Further, CEUS may also prove to be useful in differential diagnosis at early stages of this disease. PMID:25764442

  2. Privacy-Enhanced and Multifunctional Health Data Aggregation under Differential Privacy Guarantees.

    Science.gov (United States)

    Ren, Hao; Li, Hongwei; Liang, Xiaohui; He, Shibo; Dai, Yuanshun; Zhao, Lian

    2016-09-10

    With the rapid growth of the health data scale, the limited storage and computation resources of wireless body area sensor networks (WBANs) is becoming a barrier to their development. Therefore, outsourcing the encrypted health data to the cloud has been an appealing strategy. However, date aggregation will become difficult. Some recently-proposed schemes try to address this problem. However, there are still some functions and privacy issues that are not discussed. In this paper, we propose a privacy-enhanced and multifunctional health data aggregation scheme (PMHA-DP) under differential privacy. Specifically, we achieve a new aggregation function, weighted average (WAAS), and design a privacy-enhanced aggregation scheme (PAAS) to protect the aggregated data from cloud servers. Besides, a histogram aggregation scheme with high accuracy is proposed. PMHA-DP supports fault tolerance while preserving data privacy. The performance evaluation shows that the proposal leads to less communication overhead than the existing one.

  3. Privacy-Enhanced and Multifunctional Health Data Aggregation under Differential Privacy Guarantees

    Science.gov (United States)

    Ren, Hao; Li, Hongwei; Liang, Xiaohui; He, Shibo; Dai, Yuanshun; Zhao, Lian

    2016-01-01

    With the rapid growth of the health data scale, the limited storage and computation resources of wireless body area sensor networks (WBANs) is becoming a barrier to their development. Therefore, outsourcing the encrypted health data to the cloud has been an appealing strategy. However, date aggregation will become difficult. Some recently-proposed schemes try to address this problem. However, there are still some functions and privacy issues that are not discussed. In this paper, we propose a privacy-enhanced and multifunctional health data aggregation scheme (PMHA-DP) under differential privacy. Specifically, we achieve a new aggregation function, weighted average (WAAS), and design a privacy-enhanced aggregation scheme (PAAS) to protect the aggregated data from cloud servers. Besides, a histogram aggregation scheme with high accuracy is proposed. PMHA-DP supports fault tolerance while preserving data privacy. The performance evaluation shows that the proposal leads to less communication overhead than the existing one. PMID:27626417

  4. Bioactive glass ions as strong enhancers of osteogenic differentiation in human adipose stem cells.

    Science.gov (United States)

    Ojansivu, Miina; Vanhatupa, Sari; Björkvik, Leena; Häkkänen, Heikki; Kellomäki, Minna; Autio, Reija; Ihalainen, Janne A; Hupa, Leena; Miettinen, Susanna

    2015-07-01

    Bioactive glasses are known for their ability to induce osteogenic differentiation of stem cells. To elucidate the mechanism of the osteoinductivity in more detail, we studied whether ionic extracts prepared from a commercial glass S53P4 and from three experimental glasses (2-06, 1-06 and 3-06) are alone sufficient to induce osteogenic differentiation of human adipose stem cells. Cells were cultured using basic medium or osteogenic medium as extract basis. Our results indicate that cells stay viable in all the glass extracts for the whole culturing period, 14 days. At 14 days the mineralization in osteogenic medium extracts was excessive compared to the control. Parallel to the increased mineralization we observed a decrease in the cell amount. Raman and Laser Induced Breakdown Spectroscopy analyses confirmed that the mineral consisted of calcium phosphates. Consistently, the osteogenic medium extracts also increased osteocalcin production and collagen Type-I accumulation in the extracellular matrix at 13 days. Of the four osteogenic medium extracts, 2-06 and 3-06 induced the best responses of osteogenesis. However, regardless of the enhanced mineral formation, alkaline phosphatase activity was not promoted by the extracts. The osteogenic medium extracts could potentially provide a fast and effective way to differentiate human adipose stem cells in vitro. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  5. Thermo-responsive polymeric nanoparticles for enhancing neuronal differentiation of human induced pluripotent stem cells.

    Science.gov (United States)

    Seo, Hye In; Cho, Ann-Na; Jang, Jiho; Kim, Dong-Wook; Cho, Seung-Woo; Chung, Bong Geun

    2015-10-01

    We report thermo-responsive retinoic acid (RA)-loaded poly(N-isopropylacrylamide)-co-acrylamide (PNIPAM-co-Am) nanoparticles for directing human induced pluripotent stem cell (hiPSC) fate. Fourier transform infrared spectroscopy and (1)H nuclear magnetic resonance analysis confirmed that RA was efficiently incorporated into PNIAPM-co-Am nanoparticles (PCANs). The size of PCANs dropped with increasing temperatures (300-400 nm at room temperature, 80-90 nm at 37°C) due to its phase transition from hydrophilic to hydrophobic. Due to particle shrinkage caused by this thermo-responsive property of PCANs, RA could be released from nanoparticles in the cells upon cellular uptake. Immunocytochemistry and quantitative real-time polymerase chain reaction analysis demonstrated that neuronal differentiation of hiPSC-derived neuronal precursors was enhanced after treatment with 1-2 μg/ml RA-loaded PCANs. Therefore, we propose that this PCAN could be a potentially powerful carrier for effective RA delivery to direct hiPSC fate to neuronal lineage. The use of induced pluripotent stem cells (iPSCs) has been at the forefront of research in the field of regenerative medicine, as these cells have the potential to differentiate into various terminal cell types. In this article, the authors utilized a thermo-responsive polymer, Poly(N-isopropylacrylamide) (PNIPAM), as a delivery platform for retinoic acid. It was shown that neuronal differentiation could be enhanced in hiPSC-derived neuronal precursor cells. This method may pave a way for future treatment of neuronal diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Electrospun biomimetic scaffold of hydroxyapatite/chitosan supports enhanced osteogenic differentiation of mMSCs

    International Nuclear Information System (INIS)

    Peng Hongju; Feng Bei; Yuan Huihua; Zhang Yanzhong; Yin Zi; Liu Huanhuan; Chen Xiao; Ouyang Hongwei; Su Bo

    2012-01-01

    Engaging functional biomaterial scaffolds to regulate stem cell differentiation has drawn a great deal of attention in the tissue engineering and regenerative medicine community. In this study, biomimetic composite nanofibrous scaffolds of hydroxyapatite/chitosan (HAp/CTS) were prepared to investigate their capacity for inducing murine mesenchymal stem cells (mMSCs) to differentiate into the osteogenic lineage, in the absence and presence of an osteogenic supplementation (i.e., ascorbic acid, β-glycerol phosphate, and dexamethasone), respectively. Using electrospun chitosan (CTS) nanofibrous scaffolds as the control, cell morphology, growth, specific osteogenic genes expression, and quantified proteins secretion on the HAp/CTS scaffolds were sequentially examined and assessed. It appeared that the HAp/CTS scaffolds supported better attachment and proliferation of the mMSCs. Most noteworthy was that in the absence of the osteogenic supplementation, expression of osteogenic genes including collagen I (Col I), runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), and osteocalcin (OCN) were significantly upregulated in mMSCs cultured on the HAp/CTS nanofibrous scaffolds. Also increased secretion of the osteogenesis protein markers of alkaline phosphatase and collagen confirmed that the HAp/CTS nanofibrous scaffold markedly promoted the osteogenic commitment in the mMSCs. Moreover, the presence of osteogenic supplementation proved an enhanced efficacy of mMSC osteogenesis on the HAp/CTS nanofibrous scaffolds. Collectively, this study demonstrated that the biomimetic nanofibrous HAp/CTS scaffolds could support and enhance the adhesion, proliferation, and particularly osteogenic differentiation of the mMSCs. It also substantiated the potential of using biomimetic nanofibrous scaffolds of HAp/CTS for functional bone repair and regeneration applications. (paper)

  7. Enhanced reactive oxygen species overexpression by CuO nanoparticles in poorly differentiated hepatocellular carcinoma cells

    Science.gov (United States)

    Kung, Mei-Lang; Hsieh, Shu-Ling; Wu, Chih-Chung; Chu, Tian-Huei; Lin, Yu-Chun; Yeh, Bi-Wen; Hsieh, Shuchen

    2015-01-01

    Copper oxide nanoparticles (CuO NPs) are known to exhibit toxic effects on a variety of cell types and organs. To determine the oxidative impact of CuO NPs on hepatocellular carcinoma (HCC) cells, well-differentiated (HepG2) and poorly differentiated (SK-Hep-1) cells were exposed to CuO NPs. Cell viability assay showed that the median inhibition concentration (IC50) for SK-Hep-1 and HepG2 cells was 25 μg ml-1 and 85 μg ml-1, respectively. Cellular fluorescence intensity using DCFH-DA staining analysis revealed significant intracellular reactive oxygen species (ROS) generation of up to 242% in SK-Hep-1 cells, compared with 86% in HepG2 cells. HPLC analysis demonstrated that a CuO NP treatment caused cellular GSH depletion of 58% and a GSH/GSSG ratio decrease to ~0.1 in SK-Hep-1 cells. The oxidative stress caused by enhanced superoxide anion production was observed in both HepG2 (146%) and SK-Hep-1 (192%) cells. The Griess assay verified that CuO NPs induced NO production (170%) in SK-Hep-1 cells. Comet assay and western blot further demonstrated that CuO NPs induced severe DNA strand breakage (70%) in SK-Hep-1 cells and caused DNA damage via increased γ-H2AX levels. These results suggest that well-differentiated HepG2 cells possess a robust antioxidant defense system against CuO NP-induced ROS stress and exhibit more tolerance to oxidative stress. Conversely, poorly differentiated SK-Hep-1 cells exhibited a deregulated antioxidant defense system that allowed accumulation of CuO NP-induced ROS and resulted in severe cytotoxicity.Copper oxide nanoparticles (CuO NPs) are known to exhibit toxic effects on a variety of cell types and organs. To determine the oxidative impact of CuO NPs on hepatocellular carcinoma (HCC) cells, well-differentiated (HepG2) and poorly differentiated (SK-Hep-1) cells were exposed to CuO NPs. Cell viability assay showed that the median inhibition concentration (IC50) for SK-Hep-1 and HepG2 cells was 25 μg ml-1 and 85 μg ml-1, respectively

  8. Aculeatin, a coumarin derived from Toddalia asiatica (L.) Lam., enhances differentiation and lipolysis of 3T3-L1 adipocytes

    Energy Technology Data Exchange (ETDEWEB)

    Watanabe, Akio, E-mail: watanabea@jfrl.or.jp [Japan Food Research Laboratories, Osaka 567-0085 (Japan); Food and Biodynamic Chemistry Laboratory, Graduate School of Agricultural Science, Tohoku University, Miyagi 981-8555 (Japan); Kato, Tsuyoshi; Ito, Yusuke; Yoshida, Izumi; Harada, Teppei; Mishima, Takashi; Fujita, Kazuhiro; Watai, Masatoshi [Japan Food Research Laboratories, Osaka 567-0085 (Japan); Nakagawa, Kiyotaka; Miyazawa, Teruo [Food and Biodynamic Chemistry Laboratory, Graduate School of Agricultural Science, Tohoku University, Miyagi 981-8555 (Japan)

    2014-10-31

    Highlights: • Aculeatin promoted adipocyte differentiation. • Aculeatin improved glucose uptake. • Aculeatin enhanced adipocyte lipolysis. - Abstract: Toddalia asiatica (L.) Lam. (T. asiatica) has been utilized traditionally for medicinal purposes such as the treatment of diabetes. Currently, the extract is considered to be a good source of anti-diabetic agents, but the active compounds have yet to be identified. In this study, we investigated the effects of fractionated T. asiatica extracts on the differentiation of 3T3-L1 preadipocytes and identified aculeatin as a potential active agent. When 3T3-L1 preadipocytes were treated with aculeatin isolated from T. asiatica in the presence of insulin, aculeatin increased cellular triglyceride levels and glycerol-3-phosphate dehydrogenase activity. This indicated that aculeatin could enhance the differentiation of preadipocytes into adipocytes. Further analyses using a DNA microarray and real-time quantitative reverse-transcription PCR showed an increase in the expression of peroxisome proliferator-activated receptor-γ target genes (Pparg, Ap2, Cd36, Glut4 and Adipoq) by aculeatin, suggesting that aculeatin enhances the differentiation of 3T3-L1 cells by modulating the expression of genes critical for adipogenesis. Interestingly, after treatment of differentiated adipocytes with aculeatin, glucose uptake and lipolysis were enhanced. Overall, our results suggested that aculeatin is an active compound in T. asiatica for enhancing both differentiation and lipolysis of adipocytes, which are useful for the treatment of lipid abnormalities as well as diabetes.

  9. Adenovirus type 9 enhances differentiation and decreases cytokine release from preadipocytes.

    Science.gov (United States)

    Bil-Lula, Iwona; Sochocka, Marta; Zatońska, Katarzyna; Szuba, Andrzej; Sawicki, Grzegorz; Woźniak, Mieczysław

    2015-02-01

    The hypothesis was that preadipocytes would have intrinsically elevated propensity to differentiate into mature adipocytes due to AdV9 infection. To test this hypothesis, the metabolic and molecular mechanisms responsible for AdV9-induced adipogenesis were examined. An association between anti-AdV9 antibodies and human obesity was also identified. 3T3L1 cells were used as a surrogate model to analyze the preadipocyte proliferation, differentiation, and maturation. An expression of E4orf1, C/EBP-β, PPAR-γ, GAPDH, aP2, LEP and fatty acid synthase gene, intracellular lipid accumulation and cytokine release were assessed. The presence of anti-AdV antibodies, serum lipids, plasma leptin, and CRP was evaluated in 204 obese and non-obese patients. AdV9-infected cells accumulated more intracellular lipids in comparison to uninfected controls. AdV9 enhanced an expression of C/EBP-β and PPAR-γ leading to an increased differentiation of preadipocytes. Overexpression of aP2 and fatty acid synthase, and decreased expression of leptin confirmed an increased accumulation of intracellular lipids due to AdV infection. Secretion of TNF-α and IL-6 from AdV9-inoculated cells was decreased strongly. About 24.5% of prevalence of anti-AdV9 antibodies was reported in the study group. AdV9-infected subjects presented higher body weights, BMIs, WHR, and central obesity. The presence of anti-AdV9 antibodies was associated with changes in serum lipids level but neither elevated CRP nor decreased leptin levels were related to obesity due to AdV infection. Data obtained from this study provide the evidences that AdV9 is a second adenovirus, which has an influence on differentiation and lipid accumulation of 3T3L1 cells. © 2014 Wiley Periodicals, Inc.

  10. miR-34b/c Regulates Wnt1 and Enhances Mesencephalic Dopaminergic Neuron Differentiation

    Directory of Open Access Journals (Sweden)

    Roberto De Gregorio

    2018-04-01

    Full Text Available Summary: The differentiation of dopaminergic neurons requires concerted action of morphogens and transcription factors acting in a precise and well-defined time window. Very little is known about the potential role of microRNA in these events. By performing a microRNA-mRNA paired microarray screening, we identified miR-34b/c among the most upregulated microRNAs during dopaminergic differentiation. Interestingly, miR-34b/c modulates Wnt1 expression, promotes cell cycle exit, and induces dopaminergic differentiation. When combined with transcription factors ASCL1 and NURR1, miR-34b/c doubled the yield of transdifferentiated fibroblasts into dopaminergic neurons. Induced dopaminergic (iDA cells synthesize dopamine and show spontaneous electrical activity, reversibly blocked by tetrodotoxin, consistent with the electrophysiological properties featured by brain dopaminergic neurons. Our findings point to a role for miR-34b/c in neuronal commitment and highlight the potential of exploiting its synergy with key transcription factors in enhancing in vitro generation of dopaminergic neurons. : In this article, Bellenchi and colleagues show that the microRNA miR-34b/c is expressed in FACS-purified Pitx3-GFP+ neurons and promotes dopaminergic differentiation by negative modulating Wnt1 and the downstream WNT signaling pathway. Induced dopaminergic cells, expressing miR-34b/c, synthesize dopamine and show the electrophysiological properties featured by brain dopaminergic neurons. Keywords: microRNA, dopamine, mESC, miR34b/c, epiSC, transdifferentiation, Wnt1, Wnt pathway, reprogramming

  11. Schisandrae fructus enhances myogenic differentiation and inhibits atrophy through protein synthesis in human myotubes

    Directory of Open Access Journals (Sweden)

    Kim CH

    2016-05-01

    Full Text Available Cy Hyun Kim,1,2,* Jin-Hong Shin,1,3,* Sung Jun Hwang,1,2 Yung Hyun Choi,4 Dae-Seong Kim,1,3 Cheol Min Kim2,51Research Institute of Convergence of Biomedical Science and Technology, Pusan National University Yangsan Hospital, Yangsan, 2Center for Anti-Aging Industry, Pusan National University, Busan, 3Department of Neurology, Pusan National University Yangsan Hospital, Yangsan, 4Department of Biochemistry, Dong-eui University College of Korean Medicine, Busan, 5Department of Biomedical Informatics, Pusan National University School of Medicine, Yangsan, Republic of Korea*These authors contributed equally to this work Abstract: Schisandrae fructus (SF has recently been reported to increase skeletal muscle mass and inhibit atrophy in mice. We investigated the effect of SF extract on human myotube differentiation and its acting pathway. Various concentrations (0.1–10 µg/mL of SF extract were applied on human skeletal muscle cells in vitro. Myotube area and fusion index were measured to quantify myotube differentiation. The maximum effect was observed at 0.5 µg/mL of SF extract, enhancing differentiation up to 1.4-fold in fusion index and 1.6-fold in myotube area at 8 days after induction of differentiation compared to control. Phosphorylation of eukaryotic translation initiation factor 4E-binding protein 1 and 70 kDa ribosomal protein S6 kinase, which initiate translation as downstream of mammalian target of rapamycin pathway, was upregulated in early phases of differentiation after SF treatment. SF also attenuated dexamethasone-induced atrophy. In conclusion, we show that SF augments myogenic differentiation and attenuates atrophy by increasing protein synthesis through mammalian target of rapamycin/70 kDa ribosomal protein S6 kinase and eukaryotic translation initiation factor 4E-binding protein 1 signaling pathway in human myotubes. SF can be a useful natural dietary supplement in increasing skeletal muscle mass, especially in the aged

  12. Cyclic mechanical stretch enhances BMP9-induced osteogenic differentiation of mesenchymal stem cells.

    Science.gov (United States)

    Song, Yang; Tang, Yinhong; Song, Jinlin; Lei, Mingxing; Liang, Panpan; Fu, Tiwei; Su, Xudong; Zhou, Pengfei; Yang, Li; Huang, Enyi

    2018-04-01

    The purpose of this study was to investigate whether mechanical stretch can enhance the bone morphogenetic protein 9 (BMP9)-induced osteogenic differentiation in MSCs. Recombinant adenoviruses were used to overexpress the BMP9 in C3H10T1/2 MSCs. Cells were seeded onto six-well BioFlex collagen I-coated plates and subjected to cyclic mechanical stretch [6% elongation at 60 cycles/minute (1 Hz)] in a Flexercell FX-4000 strain unit for up to 12 hours. Immunostaining and confocal microscope were used to detect cytoskeleton organization. Cell cycle progression was checked by flow cytometry. Alkaline phosphatase activity was measured with a Chemiluminescence Assay Kit and was quantified with a histochemical staining assay. Matrix mineralization was examined by Alizarin Red S Staining. Mechanical stretch induces cytoskeleton reorganization and inhibits cell proliferation by preventing cells entry into S phase of the cell cycle. Although mechanical stretch alone does not induce the osteogenic differentiation of C3H10T1/2 MSCs, co-stimulation with mechanical stretch and BMP9 enhances alkaline phosphatase activity. The expression of key lineage-specific regulators (e.g., osteocalcin (OCN), SRY-related HMG-box 9, and runt-related transcription factor 2) is also increased after the co-stimulation, compared to the mechanical stretch stimulation along. Furthermore, mechanical stretch augments the BMP9-mediated bone matrix mineralization of C3H10T1/2 MSCs. Our results suggest that mechanical stretch enhances BMP9-induced osteoblastic lineage specification in C3H10T1/2 MSCs.

  13. Apparent diffusion coefficient values and dynamic contrast enhancement patterns in differentiating seminomas from nonseminomatous testicular neoplasms

    Energy Technology Data Exchange (ETDEWEB)

    Tsili, Athina C., E-mail: a_tsili@yahoo.gr [Department of Radiology, Medical School, University of Ioannina, 45110 Ioannina (Greece); Sylakos, Anastasios, E-mail: anasylakos@yahoo.gr [Department of Urology, Medical School, University of Ioannina, 45110 Ioannina (Greece); Ntorkou, Alexandra, E-mail: alexdorkou@yahoo.com [Department of Radiology, Medical School, University of Ioannina, 45110 Ioannina (Greece); Stavrou, Sotirios, E-mail: s.sotiris@yahoo.gr [Department of Urology, Medical School, University of Ioannina, 45110 Ioannina (Greece); Astrakas, Loukas G., E-mail: astrakas@uoi.gr [Department of Medical Physics, Medical School, University of Ioannina, 45110 Ioannina (Greece); Sofikitis, Nikolaos, E-mail: akrosnin@hotmail.com [Department of Urology, Medical School, University of Ioannina, 45110 Ioannina (Greece); Argyropoulou, Maria I., E-mail: margyrop@cc.uoi.gr [Department of Radiology, Medical School, University of Ioannina, 45110 Ioannina (Greece)

    2015-07-15

    Highlights: • Functional MRI in the characterization of testicular germ cell tumors was assessed. • ADC values proved useful in the characterization of testicular germ cell tumors. • Testicular germ cell tumors had similar enhancement patterns of dynamic MRI. - Abstract: Introduction: The aim of this study is to investigate the role of apparent diffusion coefficient (ADC) values and dynamic contrast enhancement (DCE) patterns in differentiating seminomas from nonseminomatous germ cell tumors (NSGCTs). Materials and methods: The MRI examinations of the scrotum of 26 men with histologically proven testicular GCTs were reviewed. DWI was performed in all patients, using a single shot, multi-slice spin-echo planar diffusion pulse sequence and b-values of 0 and 900 s/mm{sup 2}. Subtraction DCE-MRI was performed in 20 cases using a 3D fast-field echo sequence after gadolinium administration. Time-signal intensity curves were created and semi-quantitative parameters (peak enhancement, time to peak, wash-in and wash-out rate) were calculated. The Student's t-test was used to compare the mean values of ADC, peak enhancement, time to peak, wash-in and wash-out rate between seminomas and NSGCTs. ROC analysis was also performed. Results: Histopathology disclosed the presence of 15 seminomas and 11 NSGCTs. The mean ± s.d. of ADC values (× 10{sup −3} mm{sup 2}/s) of seminomas (0.59 ± 0.009) were significantly lower than those of NSGCTs (0.90 ± 0.33) (P = 0.01). The optimal ADC cut-off value was 0.68 × 10{sup −3} mm{sup 2}/s. No differences between the two groups were observed for peak enhancement (P = 0.18), time to peak (P = 0.63) wash-in rate (P = 0.32) and wash-out rate (P = 0.18). Conclusions: ADC values may be used to preoperatively differentiate seminomas from NSGCTs.

  14. In Vivo Evaluation of Biocompatibility and Chondrogenic Potential of a Cell-Free Collagen-Based Scaffold

    Directory of Open Access Journals (Sweden)

    Giovanna Calabrese

    2017-11-01

    Full Text Available Injured articular cartilage has a limited innate regenerative capacity, due to the avascular nature and low cellularity of the tissue itself. Although several approaches have been proposed to repair the joint cartilage, none of them has proven to be effective. The absence of suitable therapeutic options has encouraged tissue-engineering approaches combining specific cell types and biomaterials. In the present work, we have evaluated the potential of a cell-free Collagen I-based scaffold to promote the augmentation of cartilage-like phenotype after subcutaneous implantation in the mouse. Forty female mice were grafted subcutaneously with scaffolds, while four additional mice without scaffold were used as negative controls. The effects of scaffold were evaluated at 1, 2, 4, 8, or 16 weeks after implantation. Immunohistochemical analysis shows the expression of typical cartilage markers, including type-II Collagen, Aggrecan, Matrilin-1 and Sox 9. These data are also confirmed by qRT-PCR that further show that both COL2A1 and COL1A1 increase over time, but the first one increases more rapidly, thus suggesting a typical cartilage-like address. Histological analysis shows the presence of some pericellular lacunae, after 8 and 16 weeks. Results suggest that this scaffold (i is biocompatible in vivo, (ii is able to recruit host cells (iii induce chondrogenic differentiation of host cells. Such evidences suggest that this cell-free scaffold is promising and represents a potential approach for cartilage regeneration.

  15. CD14-negative isolation enhances chondrogenesis in synovial fibroblasts.

    Science.gov (United States)

    Bilgen, Bahar; Ren, Yuexin; Pei, Ming; Aaron, Roy K; Ciombor, Deborah McK

    2009-11-01

    Synovial membrane has been shown to contain mesenchymal stem cells. We hypothesized that an enriched population of synovial fibroblasts would undergo chondrogenic differentiation and secrete cartilage extracellular matrix to a greater extent than would a mixed synovial cell population (MSCP). The optimum doses of transforming growth factor beta 1 (TGF-beta1) and insulin-like growth factor 1 (IGF-1) for chondrogenesis were investigated. CD14-negative isolation was used to obtain a porcine cell population enriched in type-B synovial fibroblasts (SFB) from an MSCP. The positive cell surface markers in SFB were CD90, CD44, and cadherin-11. SFB and MSCP were cultured in the presence of 20 ng/mL TGF-beta1 for 7 days, and SFB were demonstrated to have higher chondrogenic potential. Further dose-response studies were carried out using the SFB cells and several doses of TGF-beta1 (2, 10, 20, and 40 ng/mL) and/or IGF-1 (1, 10, 100, and 500 ng/mL) for 14 days. TGF-beta1 supplementation was essential for chondrogenesis and prevention of cell death, whereas IGF-1 did not have a significant effect on the SFB cell number or glycosaminoglycan production. This study demonstrates that the CD14-negative isolation yields an enhanced cell population SFB that is more potent than MSCP as a cell source for cartilage tissue engineering.

  16. Evaluation of Insulin Medium or Chondrogenic Medium on Proliferation and Chondrogenesis of ATDC5 Cells

    OpenAIRE

    Yao, Yongchang; Zhai, Zhichen; Wang, Yingjun

    2014-01-01

    Background. The ATDC5 cell line is regarded as an excellent cell model for chondrogenesis. In most studies with ATDC5 cells, insulin medium (IM) was used to induce chondrogenesis while chondrogenic medium (CM), which was usually applied in chondrogenesis of mesenchymal stem cells (MSCs), was rarely used for ATDC5 cells. This study was mainly designed to investigate the effect of IM, CM, and growth medium (GM) on chondrogenesis of ATDC5 cells. Methods. ATDC5 cells were, respectively, cultured ...

  17. Self-Biased Differential Rectifier with Enhanced Dynamic Range for Wireless Powering

    KAUST Repository

    Ouda, Mahmoud H.

    2016-08-29

    A self-biased, cross-coupled, differential rectifier is proposed with enhanced power-conversion efficiency over an extended range of input power. A prototype is designed for UHF 433MHz RF power-harvesting applications and is implemented using 0.18μm CMOS technology. The proposed rectifier architecture is compared to the conventional cross-coupled rectifier. It demonstrates an improvement of more than 40% in the rectifier power conversion efficiency (PCE) and an input power range extension of more than 50% relative to the conventional crosscoupled rectifier. A sensitivity of -15.2dBm (30μW) input power for 1V output voltage and a peak power-conversion efficiency of 65% are achieved for a 50kω load. © 2004-2012 IEEE.

  18. Underwater Image Enhancement by Adaptive Gray World and Differential Gray-Levels Histogram Equalization

    Directory of Open Access Journals (Sweden)

    WONG, S.-L.

    2018-05-01

    Full Text Available Most underwater images tend to be dominated by a single color cast. This paper presents a solution to remove the color cast and improve the contrast in underwater images. However, after the removal of the color cast using Gray World (GW method, the resultant image is not visually pleasing. Hence, we propose an integrated approach using Adaptive GW (AGW and Differential Gray-Levels Histogram Equalization (DHE that operate in parallel. The AGW is applied to remove the color cast while DHE is used to improve the contrast of the underwater image. The outputs of both chromaticity components of AGW and intensity components of DHE are combined to form the enhanced image. The results of the proposed method are compared with three existing methods using qualitative and quantitative measures. The proposed method increased the visibility of underwater images and in most cases produces better quantitative scores when compared to the three existing methods.

  19. Improved Mobility Performance in LTE Co-Channel HetNets Through Speed Differentiated Enhancements

    DEFF Research Database (Denmark)

    Barbera, Simone; Michaelsen, Per Henrik; Säily, Mikko

    2012-01-01

    , requiring minimum assistance and signaling from the network. Extensive system level simulations are used to quantify the benefits. Results confirm that the proposed solutions offer improvements in several mobility key performance indicators such as radio link failure, number of handovers, offload to pico......This paper analyzes the mobility performance of LTE (Long Term Evolution) co-channel heterogeneous networks (HetNet) with macro and pico cells. Improved methods for differentiating offload and mobility robustness as a function of the UE (User Equipment) mobility are proposed. The suggested solution...... comprises two key elements, namely enhanced UE MSE (Mobility State Estimation), as well as optimized methods such that high speed users are primarily kept at the macro layer, while the offload to pico cells for low speed users is maximized. The proposed methods are designed as UE autonomous solutions...

  20. Surface-enhanced Raman spectroscopy for differentiation between benign and malignant thyroid tissues

    Science.gov (United States)

    Li, Zuanfang; Li, Chao; Lin, Duo; Huang, Zufang; Pan, Jianji; Chen, Guannan; Lin, Juqiang; Liu, Nenrong; Yu, Yun; Feng, Shangyuan; Chen, Rong

    2014-04-01

    The aim of this study was to evaluate the potential of applying silver nano-particle based surface-enhanced Raman scattering (SERS) to discriminate different types of human thyroid tissues. SERS measurements were performed on three groups of tissue samples including thyroid cancers (n = 32), nodular goiters (n = 20) and normal thyroid tissues (n = 25). Tentative assignments of the measured tissue SERS spectra suggest interesting cancer specific biomolecular differences. The principal component analysis (PCA) and linear discriminate analysis (LDA) together with the leave-one-out, cross-validated technique yielded diagnostic sensitivities of 92%, 75% and 87.5%; and specificities of 82.6%, 89.4% and 84.4%, respectively, for differentiation among normal, nodular and malignant thyroid tissue samples. This work demonstrates that tissue SERS spectroscopy associated with multivariate analysis diagnostic algorithms has great potential for detection of thyroid cancer at the molecular level.

  1. Enhanced Osteogenic Differentiation of Mesenchymal Stem Cells on Electrospun PES/PVA/PRP Nanofibrous Scaffolds.

    Science.gov (United States)

    Kashef-Saberi, Mahshid Sadat; Roodbari, Nasim Hayati; Parivar, Kazem; Vakilian, Saeid; Hanee-Ahvaz, Hana

    2018-03-28

    Over the last few decades, great advancements have been achieved in the field of bone tissue engineering (BTE). Containing a great number of growth factors needed in the process of osteogenesis, platelet rich plasma (PRP) has gained a great deal of attention. However, due to the contradictory results achieved in different studies, its effectiveness remains a mystery. Therefore, in this study, we investigated in vitro performance of co-electrospun PRP/poly ether sulfone/poly(vinyl) alcohol (PRP/PES/PVA) composite scaffolds for the osteogenic differentiation of human adipose-derived mesenchymal stem cells. The activated PRP was mixed with PVA solution to be used alongside PES solution for the electrospinning process. Fourier transform infrared spectroscopy, scanning electron microscopy and tensile tests were performed to evaluate the scaffolds. After confirmation of sustained release of protein, osteogenic potential of the co-electrospun PRP/polymer scaffolds was evaluated by measuring relative gene expression, calcium content, and alkaline phosphatase (ALP) activity. Alizarin red and Hematoxylin and Eosin staining were performed as well. The results of ALP activity and calcium content demonstrated the effectiveness of PRP when combined with PRP-incorporated scaffold in comparison with the other tested groups. In addition, the results of tensile mechanical testing indicated that addition of PRP improves the mechanical properties. Taking these results into account, it appears PES/PVA/PRP scaffold treated with PRP 5% enhances osteogenic differentiation most. In conclusion, incorporation of PRP into electrospun PES/PVA scaffold in this study had a positive influence on osteogenic differentiation of AdMSCs, and thus it may have great potential for BTE applications.

  2. A single enhancer regulating the differential expression of duplicated red-sensitive opsin genes in zebrafish.

    Directory of Open Access Journals (Sweden)

    Taro Tsujimura

    2010-12-01

    Full Text Available A fundamental step in the evolution of the visual system is the gene duplication of visual opsins and differentiation between the duplicates in absorption spectra and expression pattern in the retina. However, our understanding of the mechanism of expression differentiation is far behind that of spectral tuning of opsins. Zebrafish (Danio rerio have two red-sensitive cone opsin genes, LWS-1 and LWS-2. These genes are arrayed in a tail-to-head manner, in this order, and are both expressed in the long member of double cones (LDCs in the retina. Expression of the longer-wave sensitive LWS-1 occurs later in development and is thus confined to the peripheral, especially ventral-nasal region of the adult retina, whereas expression of LWS-2 occurs earlier and is confined to the central region of the adult retina, shifted slightly to the dorsal-temporal region. In this study, we employed a transgenic reporter assay using fluorescent proteins and P1-artificial chromosome (PAC clones encompassing the two genes and identified a 0.6-kb "LWS-activating region" (LAR upstream of LWS-1, which regulates expression of both genes. Under the 2.6-kb flanking upstream region containing the LAR, the expression pattern of LWS-1 was recapitulated by the fluorescent reporter. On the other hand, when LAR was directly conjugated to the LWS-2 upstream region, the reporter was expressed in the LDCs but also across the entire outer nuclear layer. Deletion of LAR from the PAC clones drastically lowered the reporter expression of the two genes. These results suggest that LAR regulates both LWS-1 and LWS-2 by enhancing their expression and that interaction of LAR with the promoters is competitive between the two genes in a developmentally restricted manner. Sharing a regulatory region between duplicated genes could be a general way to facilitate the expression differentiation in duplicated visual opsins.

  3. The role of catechol-O-methyltransferase in catechol-enhanced erythroid differentiation of K562 cells

    International Nuclear Information System (INIS)

    Suriguga,; Li, Xiao-Fei; Li, Yang; Yu, Chun-Hong; Li, Yi-Ran; Yi, Zong-Chun

    2013-01-01

    Catechol is widely used in pharmaceutical and chemical industries. Catechol is also one of phenolic metabolites of benzene in vivo. Our previous study showed that catechol improved erythroid differentiation potency of K562 cells, which was associated with decreased DNA methylation in erythroid specific genes. Catechol is a substrate for the catechol-O-methyltransferase (COMT)-mediated methylation. In the present study, the role of COMT in catechol-enhanced erythroid differentiation of K562 cells was investigated. Benzidine staining showed that exposure to catechol enhanced hemin-induced hemoglobin accumulation and induced mRNA expression of erythroid specific genes in K562 cells. Treatment with catechol caused a time- and concentration-dependent increase in guaiacol concentration in the medium of cultured K562 cells. When COMT expression was knocked down by COMT shRNA expression in K562 cells, the production of guaiacol significantly reduced, and the sensitivity of K562 cells to cytotoxicity of catechol significantly increased. Knockdown of COMT expression by COMT shRNA expression also eliminated catechol-enhanced erythroid differentiation of K562 cells. In addition, the pre-treatment with methyl donor S-adenosyl-L-methionine or its demethylated product S-adenosyl-L-homocysteine induced a significant increase in hemin-induced Hb synthesis in K562 cells and the mRNA expression of erythroid specific genes. These findings indicated that O-methylation catalyzed by COMT acted as detoxication of catechol and involved in catechol-enhanced erythroid differentiation of K562 cells, and the production of S-adenosyl-L-homocysteine partly explained catechol-enhanced erythroid differentiation. - Highlights: • Catechol enhanced hemin-induced hemoglobin accumulation. • COMT-catalyzed methylation acted as detoxication of catechol. • COMT involved in catechol-enhanced erythroid differentiation

  4. The role of catechol-O-methyltransferase in catechol-enhanced erythroid differentiation of K562 cells

    Energy Technology Data Exchange (ETDEWEB)

    Suriguga,; Li, Xiao-Fei; Li, Yang; Yu, Chun-Hong; Li, Yi-Ran; Yi, Zong-Chun, E-mail: yizc@buaa.edu.cn

    2013-12-15

    Catechol is widely used in pharmaceutical and chemical industries. Catechol is also one of phenolic metabolites of benzene in vivo. Our previous study showed that catechol improved erythroid differentiation potency of K562 cells, which was associated with decreased DNA methylation in erythroid specific genes. Catechol is a substrate for the catechol-O-methyltransferase (COMT)-mediated methylation. In the present study, the role of COMT in catechol-enhanced erythroid differentiation of K562 cells was investigated. Benzidine staining showed that exposure to catechol enhanced hemin-induced hemoglobin accumulation and induced mRNA expression of erythroid specific genes in K562 cells. Treatment with catechol caused a time- and concentration-dependent increase in guaiacol concentration in the medium of cultured K562 cells. When COMT expression was knocked down by COMT shRNA expression in K562 cells, the production of guaiacol significantly reduced, and the sensitivity of K562 cells to cytotoxicity of catechol significantly increased. Knockdown of COMT expression by COMT shRNA expression also eliminated catechol-enhanced erythroid differentiation of K562 cells. In addition, the pre-treatment with methyl donor S-adenosyl-L-methionine or its demethylated product S-adenosyl-L-homocysteine induced a significant increase in hemin-induced Hb synthesis in K562 cells and the mRNA expression of erythroid specific genes. These findings indicated that O-methylation catalyzed by COMT acted as detoxication of catechol and involved in catechol-enhanced erythroid differentiation of K562 cells, and the production of S-adenosyl-L-homocysteine partly explained catechol-enhanced erythroid differentiation. - Highlights: • Catechol enhanced hemin-induced hemoglobin accumulation. • COMT-catalyzed methylation acted as detoxication of catechol. • COMT involved in catechol-enhanced erythroid differentiation.

  5. Electrical Polarization of Titanium Surfaces for the Enhancement of Osteoblast Differentiation

    Science.gov (United States)

    Gittens, Rolando A.; Olivares-Navarrete, Rene; Rettew, Robert; Butera, Robert J.; Alamgir, Faisal M.; Boyan, Barbara D.; Schwartz, Zvi

    2014-01-01

    Electrical stimulation has been used clinically to promote bone regeneration in cases of fractures with delayed union or nonunion, with several in vitro and in vivo reports suggesting its beneficial effects on bone formation. However, the use of electrical stimulation of titanium (Ti) implants to enhance osseointegration is less understood, in part because of the few in vitro models that attempt to represent the in vivo environment. In this article, the design of a new in vitro system that allows direct electrical stimulation of osteoblasts through their Ti substrates without the flow of exogenous currents through the media is presented, and the effect of applied electrical polarization on osteoblast differentiation and local factor production was evaluated. A custom-made polycarbonate tissue culture plate was designed to allow electrical connections directly underneath Ti disks placed inside the wells, which were supplied with electrical polarization ranging from 100 to 500 mV to stimulate MG63 osteoblasts. Our results show that electrical polarization applied directly through Ti substrates on which the cells are growing in the absence of applied electrical currents may increase osteoblast differentiation and local factor production in a voltage-dependent manner. PMID:23996899

  6. Combined VEGF and LMP-1 delivery enhances osteoprogenitor cell differentiation and ectopic bone formation.

    Science.gov (United States)

    Wang, Xiuli; Cui, Fuai; Madhu, Vedavathi; Dighe, Abhijit S; Balian, Gary; Cui, Quanjun

    2011-02-01

    A novel strategy to enhance bone repair is to combine angiogenic factors and osteogenic factors. We combined vascular endothelial growth factor (VEGF) and LIM mineralization protein-1 (LMP-1) by using an internal ribosome entry site to link the genes within a single plasmid. We then evaluated the effects on osteoblastic differentiation in vitro and ectopic bone formation in vivo with a subcutaneously placed PLAGA scaffold loaded with a cloned mouse osteoprogenitor cell line, D1, transfected with plasmids containing VEGF and LMP-1 genes. The cells expressing both genes elevated mRNA expression of RunX2 and β-catenin and alkaline phosphatase activity compared to cells from other groups. In vivo, X-ray and micro-CT analysis of the retrieved implants revealed more ectopic bone formation at 2 and 3 weeks but not at 4 weeks compared to other groups. The results indicate that the combination of the therapeutic growth factors potentiates cell differentiation and may promote osteogenesis.

  7. Micro/Nano Structural Tantalum Coating for Enhanced Osteogenic Differentiation of Human Bone Marrow Stem Cells.

    Science.gov (United States)

    Ding, Ding; Xie, Youtao; Li, Kai; Huang, Liping; Zheng, Xuebin

    2018-04-03

    Recently, tantalum has been attracting much attention for its anticorrosion resistance and biocompatibility, and it has been widely used in surface modification for implant applications. To improve its osteogenic differentiation of human bone marrow stem cells (hBMSCs), a micro/nano structure has been fabricated on the tantalum coating surface through the combination of anodic oxidation and plasma spraying method. The morphology, composition, and microstructure of the modified coating were comprehensively studied by employing scanning electron microscopy (SEM), X-ray diffraction (XRD) as well as transmission electron microscopy (TEM). The effects of hierarchical structures as well as micro-porous structure of tantalum coating on the behavior for human bone marrow stem cells (hBMSCs) were evaluated and compared at both cellular and molecular levels in vitro. The experimental results show that a hierarchical micro/nano structure with Ta₂O₅ nanotubes spread onto a micro-scale tantalum coating has been fabricated successfully, which is confirmed to promote cell adhesion and spreading. Besides, the hierarchical micro/nano tantalum coating can provide 1.5~2.1 times improvement in gene expression, compared with the micro-porous tantalum coating. It demonstrates that it can effectively enhance the proliferation and differentiation of hBMSCs in vitro.

  8. Micro/Nano Structural Tantalum Coating for Enhanced Osteogenic Differentiation of Human Bone Marrow Stem Cells

    Directory of Open Access Journals (Sweden)

    Ding Ding

    2018-04-01

    Full Text Available Recently, tantalum has been attracting much attention for its anticorrosion resistance and biocompatibility, and it has been widely used in surface modification for implant applications. To improve its osteogenic differentiation of human bone marrow stem cells (hBMSCs, a micro/nano structure has been fabricated on the tantalum coating surface through the combination of anodic oxidation and plasma spraying method. The morphology, composition, and microstructure of the modified coating were comprehensively studied by employing scanning electron microscopy (SEM, X-ray diffraction (XRD as well as transmission electron microscopy (TEM. The effects of hierarchical structures as well as micro-porous structure of tantalum coating on the behavior for human bone marrow stem cells (hBMSCs were evaluated and compared at both cellular and molecular levels in vitro. The experimental results show that a hierarchical micro/nano structure with Ta2O5 nanotubes spread onto a micro-scale tantalum coating has been fabricated successfully, which is confirmed to promote cell adhesion and spreading. Besides, the hierarchical micro/nano tantalum coating can provide 1.5~2.1 times improvement in gene expression, compared with the micro-porous tantalum coating. It demonstrates that it can effectively enhance the proliferation and differentiation of hBMSCs in vitro.

  9. Enhancing proliferation and osteogenic differentiation of HMSCs on casein/chitosan multilayer films.

    Science.gov (United States)

    Li, Yan; Zheng, Zebin; Cao, Zhinan; Zhuang, Liangting; Xu, Yong; Liu, Xiaozhen; Xu, Yue; Gong, Yihong

    2016-05-01

    Creating a bioactive surface is important in tissue engineering. Inspired by the natural calcium binding property of casein (CA), multilayer films ((CA/CS)n) with chitosan (CS) as polycation were fabricated to enhance biomineralization, cell adhesion and differentiation. LBL self-assembly technique was used and the assembly process was intensively studied based on changes of UV absorbance, zeta potential and water contact angle. The increasing content of chitosan and casein with bilayers was further confirmed with XPS and TOF-SIMS analysis. To improve the biocompatibility, gelatin was surface grafted. In vitro mineralization test demonstrated that multilayer films had more hydroxyapatite crystal deposition. Human mesenchymal stem cells (HMSCs) were seeded onto these films. According to fluorescein diacetate (FDA) and cell cytoskeleton staining, MTT assay, expression of osteogenic marker genes, ALP activity, and calcium deposition quantification, it was found that these multilayer films significantly promoted HMSCs attachment, proliferation and osteogenic differentiation than TCPS control. Copyright © 2016. Published by Elsevier B.V.

  10. Effect of silver nanoparticles on human mesenchymal stem cell differentiation

    Directory of Open Access Journals (Sweden)

    Christina Sengstock

    2014-11-01

    Full Text Available Background: Silver nanoparticles (Ag-NP are one of the fastest growing products in nano-medicine due to their enhanced antibacterial activity at the nanoscale level. In biomedicine, hundreds of products have been coated with Ag-NP. For example, various medical devices include silver, such as surgical instruments, bone implants and wound dressings. After the degradation of these materials, or depending on the coating technique, silver in nanoparticle or ion form can be released and may come into close contact with tissues and cells. Despite incorporation of Ag-NP as an antibacterial agent in different products, the toxicological and biological effects of silver in the human body after long-term and low-concentration exposure are not well understood. In the current study, we investigated the effects of both ionic and nanoparticulate silver on the differentiation of human mesenchymal stem cells (hMSCs into adipogenic, osteogenic and chondrogenic lineages and on the secretion of the respective differentiation markers adiponectin, osteocalcin and aggrecan.Results: As shown through laser scanning microscopy, Ag-NP with a size of 80 nm (hydrodynamic diameter were taken up into hMSCs as nanoparticulate material. After 24 h of incubation, these Ag-NP were mainly found in the endo-lysosomal cell compartment as agglomerated material. Cytotoxicity was observed for differentiated or undifferentiated hMSCs treated with high silver concentrations (≥20 µg·mL−1 Ag-NP; ≥1.5 µg·mL−1 Ag+ ions but not with low-concentration treatments (≤10 µg·mL−1 Ag-NP; ≤1.0 µg·mL−1 Ag+ ions. Subtoxic concentrations of Ag-NP and Ag+ ions impaired the adipogenic and osteogenic differentiation of hMSCs in a concentration-dependent manner, whereas chondrogenic differentiation was unaffected after 21 d of incubation. In contrast to aggrecan, the inhibitory effect of adipogenic and osteogenic differentiation was confirmed by a decrease in the secretion of

  11. The potential application of LIPUS and PEMF on cartilage and chondrogenic differentiation : An in vitro study

    NARCIS (Netherlands)

    Tan, Lijun

    2016-01-01

    In chapter 2 a review is presented of the scientific literature on the use and mechanism of low-intensity pulsed ultrasound (LIPUS) and pulsed electromagnetic fields (PEMF) applied on articular cartilage, chondrocytes and chondrogenesis in order to compare these two non-invasive procedures regarding

  12. Effect of electromagnetic fields on the chondrogenic differentiation under microgravity conditions

    Data.gov (United States)

    National Aeronautics and Space Administration — A combination therapy of electromagnetic fields (EMF) and simulated microgravity (SMG) has not been examined in regenerative medicine of cartilage. In the present...

  13. Chondrogenic differentiation of human articular chondrocytes differs in biodegradable PGA/PLA scaffolds

    DEFF Research Database (Denmark)

    Zwingmann, Joern; Mehlhorn, Alexander T; Südkamp, Norbert

    2007-01-01

    Cartilage tissue engineering is applied clinically to cover and regenerate articular cartilage defects. Two bioresorbable nonwoven scaffolds, polyglycolic acid (PGA) and poly(lactic-co-glycolic acid) (PLGA) (90/10 copolymer of L-lactide and glycolide), were seeded with human chondrocytes after in...

  14. Transcriptional and epigenetic mechanisms underlying enhanced in vitro adipocyte differentiation by the brominated flame retardant BDE-47

    DEFF Research Database (Denmark)

    Kamstra, Jorke H; Hruba, Eva; Blumberg, Bruce

    2014-01-01

    Recent studies suggest that exposure to endocrine-disrupting compounds (EDCs) may play a role in the development of obesity. EDCs such as the flame retardant 2,2',4,4'-tetrabrominated diphenyl ether (BDE-47) have been shown to enhance adipocyte differentiation in the murine 3T3-L1 model. The mech......Recent studies suggest that exposure to endocrine-disrupting compounds (EDCs) may play a role in the development of obesity. EDCs such as the flame retardant 2,2',4,4'-tetrabrominated diphenyl ether (BDE-47) have been shown to enhance adipocyte differentiation in the murine 3T3-L1 model....... The mechanisms by which EDCs direct preadipocytes to form adipocytes are poorly understood. Here, we examined transcriptional and epigenetic mechanisms underlying the induction of in vitro adipocyte differentiation by BDE-47. Quantitative high content microscopy revealed concentration-dependent enhanced...

  15. Differential tissue expression of enhanced green fluorescent protein in 'green mice'.

    Science.gov (United States)

    Ma, De-Fu; Tezuka, Hideo; Kondo, Tetsuo; Sudo, Katsuko; Niu, Dong-Feng; Nakazawa, Tadao; Kawasaki, Tomonori; Yamane, Tetsu; Nakamura, Nobuki; Katoh, Ryohei

    2010-06-01

    In order to clarify tissue expression of enhanced green fluorescent protein (EGFP) in 'green mice' from a transgenic line having an EGFP cDNA under the control of a chicken beta-actin promoter and cytomegalovirus enhancer, we studied the expression of EGFP in various organs and tissues from these 'green mice' by immunohistochemistry with anti- EGFP antibody in conjunction with direct observation for EGFP fluorescence using confocal laser scanning microscopy. On immunohistochemical examination and on direct observation by confocal laser scanning microscopy, the level of EGFP expression varied among organs and tissues. EGFP expression was diffusely and strongly observed in the skin, pituitary, thyroid gland, parathyroid gland, heart, gall bladder, pancreas, adrenals and urinary bladder. There was only sporadic and weak expression of EGFP in the epithelium of the trachea, bronchus of the lung, stratified squamous epithelium and gastric glands of the stomach, hepatic bile ducts of the liver, glomeruli and renal tubules of the kidney and endo-metrial glands of the uterus. Furthermore, EGFP was only demonstrated within the goblet and paneth cells in the colon and small intestine, the tall columnar cells in the ductus epididymis, and the leydig cells in the testis. In conclusion, our results show that EGFP is differentially expressed in organs and tissues of 'green mice', which indicates that 'green mice' may prove useful for research involving transplantation and tissue clonality.

  16. Novel Differential Measurement of Natural and Added Phosphorus in Preserved versus Non-Enhanced Ham

    Science.gov (United States)

    Cupisti, Adamasco; Benini, Omar; Ferretti, Valerio; Gianfaldoni, Daniela; Kalantar-Zadeh, Kamyar

    2018-01-01

    Accurate assessment of the quantity and chemical type of phosphorus (P) content in processed meat products may have major clinical implications for management of kidney disease patients. We examined 40 lots of cooked ham including 20 without and 20 with P-containing preservatives. Novel spectro-photometrical methods were employed to measure total P and 3 different P subtypes, i.e., water-soluble (inorganic) P including added preservatives and natural P derived from phospholipids and phosphoproteins separately. Total Nitrogen and fat contents were assayed, as well. There was 66% more inorganic P in preserved vs. non-enhanced ham, i.e., 169±36 vs. 102±16 mg/100g (p0.05), indicating a small portion of unspecified P and/or undermeasurement Novel differential dietary P measurement detects added P-containing preservatives. Processed cooked ham has 66% more measurable inorganic P and 64% higher P-to-protein ratio than non-enhanced product. The contribution of processed food to global dietary phosphorus burden can negatively influence CKD outcome and counteract the efficacy of P-binder medications. PMID:22406120

  17. Enhanced Stem Cell Osteogenic Differentiation by Bioactive Glass Functionalized Graphene Oxide Substrates

    Directory of Open Access Journals (Sweden)

    Xiaoju Mo

    2016-01-01

    Full Text Available An unmet need in engineered bone regeneration is to develop scaffolds capable of manipulating stem cells osteogenesis. Graphene oxide (GO has been widely used as a biomaterial for various biomedical applications. However, it remains challenging to functionalize GO as ideal platform for specifically directing stem cell osteogenesis. Herein, we report facile functionalization of GO with dopamine and subsequent bioactive glass (BG to enhance stem cell adhesion, spreading, and osteogenic differentiation. On the basis of graphene, we obtained dopamine functionalized graphene oxide/bioactive glass (DGO/BG hybrid scaffolds containing different content of DGO by loading BG nanoparticles on graphene oxide surface using sol-gel method. To enhance the dispersion stability and facilitate subsequent nucleation of BG in GO, firstly, dopamine (DA was used to modify GO. Then, the modified GO was functionalized with bioactive glass (BG using sol-gel method. The adhesion, spreading, and osteoinductive effects of DGO/BG scaffold on rat bone marrow mesenchymal stem cells (rBMSCs were evaluated. DGO/BG hybrid scaffolds with different content of DGO could influence rBMSCs’ behavior. The highest expression level of osteogenic markers suggests that the DGO/BG hybrid scaffolds have great potential or elicit desired bone reparative outcome.

  18. Microtissues Enhance Smooth Muscle Differentiation and Cell Viability of hADSCs for Three Dimensional Bioprinting

    Directory of Open Access Journals (Sweden)

    Jin Yipeng

    2017-07-01

    points compared to the conventional single-cell bioprinting strategy (mean cell viability was 88.16 ± 3.98 vs. 61.76 ± 15% for microtissues and single-cells, respectively. These results provide a novel way to enhance the smooth muscle differentiation of hADSCs and a simple method to maintain better cell viability in 3D bioprinting.

  19. CCAAT/Enhancer Binding Protein β Regulates Expression of Indian Hedgehog during Chondrocytes Differentiation

    Science.gov (United States)

    Ushijima, Takahiro; Okazaki, Ken; Tsushima, Hidetoshi; Ishihara, Kohei; Doi, Toshio; Iwamoto, Yukihide

    2014-01-01

    Background CCAAT/enhancer binding protein β (C/EBPβ) is a transcription factor that promotes hypertrophic differentiation of chondrocytes. Indian hedgehog (Ihh) also stimulates the hypertrophic transition of chondrocytes. Furthermore, runt-related transcription factor-2 (RUNX2) was reported to regulate chondrocyte maturation during skeletal development and to directly regulate transcriptional activity of Ihh. In this study, we investigated whether the interaction of C/EBPβ and RUNX2 regulates the expression of Ihh during chondrocyte differentiation. Methodology/Results Immunohistochemistry of embryonic growth plate revealed that both C/EBPβ and Ihh were strongly expressed in pre-hypertrophic and hypertrophic chondrocytes. Overexpression of C/EBPβ by adenovirus vector in ATDC5 cells caused marked stimulation of Ihh and Runx2. Conversely, knockdown of C/EBPβ by lentivirus expressing shRNA significantly repressed Ihh and Runx2 in ATDC5 cells. A reporter assay revealed that C/EBPβ stimulated transcriptional activity of Ihh. Deletion and mutation analysis showed that the C/EBPβ responsive element was located between −214 and −210 bp in the Ihh promoter. An electrophoretic mobility shift assay (EMSA) and a chromatin immunoprecipitation (ChIP) assay also revealed the direct binding of C/EBPβ to this region. Moreover, reporter assays demonstrated that RUNX2 failed to stimulate the transcriptional activity of the Ihh promoter harboring a mutation at the C/EBPβ binding site. EMSA and ChIP assays showed that RUNX2 interacted to this element with C/EBPβ. Immunoprecipitation revealed that RUNX2 and C/EBPβ formed heterodimer complex with each other in the nuclei of chondrocytes. These data suggested that the C/EBPβ binding element is also important for RUNX2 to regulate the expression of Ihh. Ex vivo organ culture of mouse limbs transfected with C/EBPβ showed that the expression of Ihh and RUNX2 was increased upon ectopic C/EBPβ expression. Conclusions C

  20. CCAAT/enhancer binding protein β regulates expression of Indian hedgehog during chondrocytes differentiation.

    Directory of Open Access Journals (Sweden)

    Takahiro Ushijima

    Full Text Available CCAAT/enhancer binding protein β (C/EBPβ is a transcription factor that promotes hypertrophic differentiation of chondrocytes. Indian hedgehog (Ihh also stimulates the hypertrophic transition of chondrocytes. Furthermore, runt-related transcription factor-2 (RUNX2 was reported to regulate chondrocyte maturation during skeletal development and to directly regulate transcriptional activity of Ihh. In this study, we investigated whether the interaction of C/EBPβ and RUNX2 regulates the expression of Ihh during chondrocyte differentiation.Immunohistochemistry of embryonic growth plate revealed that both C/EBPβ and Ihh were strongly expressed in pre-hypertrophic and hypertrophic chondrocytes. Overexpression of C/EBPβ by adenovirus vector in ATDC5 cells caused marked stimulation of Ihh and Runx2. Conversely, knockdown of C/EBPβ by lentivirus expressing shRNA significantly repressed Ihh and Runx2 in ATDC5 cells. A reporter assay revealed that C/EBPβ stimulated transcriptional activity of Ihh. Deletion and mutation analysis showed that the C/EBPβ responsive element was located between -214 and -210 bp in the Ihh promoter. An electrophoretic mobility shift assay (EMSA and a chromatin immunoprecipitation (ChIP assay also revealed the direct binding of C/EBPβ to this region. Moreover, reporter assays demonstrated that RUNX2 failed to stimulate the transcriptional activity of the Ihh promoter harboring a mutation at the C/EBPβ binding site. EMSA and ChIP assays showed that RUNX2 interacted to this element with C/EBPβ. Immunoprecipitation revealed that RUNX2 and C/EBPβ formed heterodimer complex with each other in the nuclei of chondrocytes. These data suggested that the C/EBPβ binding element is also important for RUNX2 to regulate the expression of Ihh. Ex vivo organ culture of mouse limbs transfected with C/EBPβ showed that the expression of Ihh and RUNX2 was increased upon ectopic C/EBPβ expression.C/EBPβ and RUNX2 cooperatively stimulate

  1. Differentiating perforated from non-perforated appendicitis on contrast-enhanced magnetic resonance imaging

    International Nuclear Information System (INIS)

    Rosenbaum, Daniel G.; Kovanlikaya, Arzu; Askin, Gulce; Beneck, Debra M.

    2017-01-01

    The role of magnetic resonance imaging (MRI) in pediatric appendicitis is increasing; MRI findings predictive of appendiceal perforation have not been specifically evaluated. To assess the performance of MRI in differentiating perforated from non-perforated appendicitis. A retrospective review of pediatric patients undergoing contrast-enhanced MRI and subsequent appendectomy was performed, with surgicopathological confirmation of perforation. Appendiceal diameter and the following 10 MRI findings were assessed: appendiceal restricted diffusion, wall defect, appendicolith, periappendiceal free fluid, remote free fluid, restricted diffusion within free fluid, abscess, peritoneal enhancement, ileocecal wall thickening and ileus. Two-sample t-test and chi-square tests were used to analyze continuous and discrete data, respectively. Sensitivity and specificity for individual MRI findings were calculated and optimal thresholds for measures of accuracy were selected. Seventy-seven patients (mean age: 12.2 years) with appendicitis were included, of whom 22 had perforation. The perforated group had a larger mean appendiceal diameter and mean number of MRI findings than the non-perforated group (12.3 mm vs. 8.6 mm; 5.0 vs. 2.0, respectively). Abscess, wall defect and restricted diffusion within free fluid had the greatest specificity for perforation (1.00, 1.00 and 0.96, respectively) but low sensitivity (0.36, 0.25 and 0.32, respectively). The receiver operator characteristic curve for total number of MRI findings had an area under the curve of 0.92, with an optimal threshold of 3.5. A threshold of any 4 findings had the best ability to accurately discriminate between perforated and non-perforated cases, with a sensitivity of 82% and specificity of 85%. Contrast-enhanced MRI can differentiate perforated from non-perforated appendicitis. The presence of multiple findings increases diagnostic accuracy, with a threshold of any four findings optimally discriminating between

  2. Differentiating perforated from non-perforated appendicitis on contrast-enhanced magnetic resonance imaging.

    Science.gov (United States)

    Rosenbaum, Daniel G; Askin, Gulce; Beneck, Debra M; Kovanlikaya, Arzu

    2017-10-01

    The role of magnetic resonance imaging (MRI) in pediatric appendicitis is increasing; MRI findings predictive of appendiceal perforation have not been specifically evaluated. To assess the performance of MRI in differentiating perforated from non-perforated appendicitis. A retrospective review of pediatric patients undergoing contrast-enhanced MRI and subsequent appendectomy was performed, with surgicopathological confirmation of perforation. Appendiceal diameter and the following 10 MRI findings were assessed: appendiceal restricted diffusion, wall defect, appendicolith, periappendiceal free fluid, remote free fluid, restricted diffusion within free fluid, abscess, peritoneal enhancement, ileocecal wall thickening and ileus. Two-sample t-test and chi-square tests were used to analyze continuous and discrete data, respectively. Sensitivity and specificity for individual MRI findings were calculated and optimal thresholds for measures of accuracy were selected. Seventy-seven patients (mean age: 12.2 years) with appendicitis were included, of whom 22 had perforation. The perforated group had a larger mean appendiceal diameter and mean number of MRI findings than the non-perforated group (12.3 mm vs. 8.6 mm; 5.0 vs. 2.0, respectively). Abscess, wall defect and restricted diffusion within free fluid had the greatest specificity for perforation (1.00, 1.00 and 0.96, respectively) but low sensitivity (0.36, 0.25 and 0.32, respectively). The receiver operator characteristic curve for total number of MRI findings had an area under the curve of 0.92, with an optimal threshold of 3.5. A threshold of any 4 findings had the best ability to accurately discriminate between perforated and non-perforated cases, with a sensitivity of 82% and specificity of 85%. Contrast-enhanced MRI can differentiate perforated from non-perforated appendicitis. The presence of multiple findings increases diagnostic accuracy, with a threshold of any four findings optimally discriminating between

  3. Differentiating perforated from non-perforated appendicitis on contrast-enhanced magnetic resonance imaging

    Energy Technology Data Exchange (ETDEWEB)

    Rosenbaum, Daniel G.; Kovanlikaya, Arzu [New York-Presbyterian Hospital/Weill Cornell Medicine, Division of Pediatric Radiology, New York, NY (United States); Askin, Gulce [Weill Cornell Medical College, Division of Biostatistics and Epidemiology, New York, NY (United States); Beneck, Debra M. [New York-Presbyterian Hospital/Weill Cornell Medicine, Department of Pathology, New York, NY (United States)

    2017-10-15

    The role of magnetic resonance imaging (MRI) in pediatric appendicitis is increasing; MRI findings predictive of appendiceal perforation have not been specifically evaluated. To assess the performance of MRI in differentiating perforated from non-perforated appendicitis. A retrospective review of pediatric patients undergoing contrast-enhanced MRI and subsequent appendectomy was performed, with surgicopathological confirmation of perforation. Appendiceal diameter and the following 10 MRI findings were assessed: appendiceal restricted diffusion, wall defect, appendicolith, periappendiceal free fluid, remote free fluid, restricted diffusion within free fluid, abscess, peritoneal enhancement, ileocecal wall thickening and ileus. Two-sample t-test and chi-square tests were used to analyze continuous and discrete data, respectively. Sensitivity and specificity for individual MRI findings were calculated and optimal thresholds for measures of accuracy were selected. Seventy-seven patients (mean age: 12.2 years) with appendicitis were included, of whom 22 had perforation. The perforated group had a larger mean appendiceal diameter and mean number of MRI findings than the non-perforated group (12.3 mm vs. 8.6 mm; 5.0 vs. 2.0, respectively). Abscess, wall defect and restricted diffusion within free fluid had the greatest specificity for perforation (1.00, 1.00 and 0.96, respectively) but low sensitivity (0.36, 0.25 and 0.32, respectively). The receiver operator characteristic curve for total number of MRI findings had an area under the curve of 0.92, with an optimal threshold of 3.5. A threshold of any 4 findings had the best ability to accurately discriminate between perforated and non-perforated cases, with a sensitivity of 82% and specificity of 85%. Contrast-enhanced MRI can differentiate perforated from non-perforated appendicitis. The presence of multiple findings increases diagnostic accuracy, with a threshold of any four findings optimally discriminating between

  4. Endogenous collagen influences differentiation of human multipotent mesenchymal stromal cells

    NARCIS (Netherlands)

    Fernandes, H.; Mentink, A.; Bank, R.; Stoop, R.; Blitterswijk, C. van; Boer, J. de

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  5. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, Hugo; Mentink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  6. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, H.A.M.; Mentink-Leusink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  7. Value of fast dynamic enhanced MR imaging for the differential diagnosis between ovarian fibroma and subserosal leiomyoma

    International Nuclear Information System (INIS)

    Shan Jun; Wang Xiaomei; Xu Jianmin; Xu Hongli; Feng Xiaofeng; Gong Jingshan; Ling Rennan; Rao Zibin

    2004-01-01

    Objective: To study the value of dynamic enhanced MRI in the differential diagnosis between subserosal uterine leiomyoma and ovarian fibroma which were iso-signal and low signal ones on T 2 WI. Methods: Plain and dynamic enhanced MRI findings of 45 patients with histologically proved subserosal leiomyoma or ovarian fibroma were analyzed. There were 35 cases of subserosal leiomyoma and 10 cases of ovarian fibroma. The enhancement index of early phase (100 sec after the contrast medial administration) and the time to peak during 200 sec after the contrast administration (TTP 200 ) were analyzed. Results: On plain scanning, both subserosal leiomyoma and ovarian fibrioma appeared similarly as intrapelvic masses with low signal on T 2 WI. On Gadolinium dynamic enhancement, subserosal leiomyomas showed early and evident enhancement while ovarian fibroma showed delayed and moderate enhancement. The difference of the enhancement index of early phase and TTP 200 was statistically significant (P<0.001). Conclusion: Gadolinium dynamic enhanced MR imaging was helpful in the differential diagnosis between subserosal uterine leiomyoma and ovarian fibroma by the difference in the enhancement pattern

  8. The usefulness of contrast-enhanced sonography in the differential diagnostic of adrenal tumors

    International Nuclear Information System (INIS)

    Slonina, J.; Nienartowicz, E.; Malczewska, J.; Moron, K.; Kumar Agrawal, A.

    2006-01-01

    Introduction: The occurrence of gland tumors causes significant clinical problem. Non hormone-secreting tumors provide the most complicated diagnostic difficulties. The application of contrast-enhanced sonography could improve the vessels visualization and point out characteristic features of benign and malignant changes. The authors believe that this new method make possible the differential adrenal tumor diagnostic process more precise and increase the specificity of ultrasonography in the recognition of benign and malignant tumors. The aim of this study was to define the usefulness of contrasting agent Levovist in differential diagnostics of adrenal tumors and its influence on sensitivity and specificity of ultrasound examination and to establish patients qualification criteria for surgical procedures. Material and methods: Ultrasound examinations were made with the use of digital devise by GE Voluson 740, probe 4.6 MHz with Doppler options and volumetric probe 3D according to the following protocol: 26 patients with recognized adrenal tumor were qualified for the examination. Patients in the first stage of tumor vascularisation had Doppler examination with color (CD) and power Doppler (PD). Three-dimensional ultrasonography was used to improve visualization of vascularisation. In the final phase of the examination the patients were administrated of Levovist in the recommended by the producer dose: 2,5 g in the concentration of 400 mg/l. Results: 26 cases of adrenal gland tumours were subjected to analysis. In standard ultrasonographic examination focal changes in 25 patients were hipoechogenic focuses and in one case the focus was hyperechogenic. Heterogeneity of focuses was observed in 16 cases. In Doppler examination with color (CD) and power Doppler (PD) vascular blood flow was revealed within 12. After using contrasting agent Levovist vascular blood flow was achieved in 4 additional cases, which constituted 61% . Conclusions: 1. 3D ultrasound could be

  9. The use of contrast-enhanced color doppler ultrasound in the differentiation of retinal detachment from vitreous membrane

    International Nuclear Information System (INIS)

    Han, Sang Suk; Chang, Seung Kook; Yoon, Jung Hee; Lee, Young Joon

    2001-01-01

    To compare the clinical utility of contrast-enhanced color Doppler US in the differentiation of retinal detachment (RD) from vitreous membrane (VM) with that of various conventional US modalities, and to analyze the enhancement patterns in cases showing an enhancement effect. In 32 eyes examined over a recent two-year period, RD (n=14) and VM (n=18) were confirmed by surgery (n=28) or clinical follow-up (n=4). In all cases, gray-scale, color Doppler, and power Doppler US were performed prior to contrast injection, and after the intravenous injection of Levovist (Schering, Berlin) by hand for 30 seconds at a dose of 2.5 g and a concentration of 300 mg/mL via an antecubital vein, contrast-enhanced color Doppler US was performed. At Doppler US, the diagnostic criterion for RD and VM was whether or not color signals were visualized in membranous structures. Diagnostic accuracy was 78% at gray-scale US, 81% at color Doppler US, 59% at power Doppler US, and 97% at contrast-enhanced color Doppler US. The sensitivity of color Doppler US to color signals in RD increased from 57% to 93% after contrast enhancement. The enhancement patterns observed were signal accentuation (n=3), signal extension (n=2), signal addition (n=3), and new signal visualization (n=5). Contrast-enhanced color Doppler US was the most accurate US modality for differentiating RD from VM, showing a significantly increased signal detection rate in RD

  10. Attenuation of chondrogenic transformation in vascular smooth muscle by dietary quercetin in the MGP-deficient mouse model.

    Directory of Open Access Journals (Sweden)

    Kelly E Beazley

    Full Text Available Cartilaginous metaplasia of vascular smooth muscle (VSM is characteristic for arterial calcification in diabetes and uremia and in the background of genetic alterations in matrix Gla protein (MGP. A better understanding of the molecular details of this process is critical for the development of novel therapeutic approaches to VSM transformation and arterial calcification.This study aimed to identify the effects of bioflavonoid quercetin on chondrogenic transformation and calcification of VSM in the MGP-null mouse model and upon TGF-β3 stimulation in vitro, and to characterize the associated alterations in cell signaling.Molecular analysis revealed activation of β-catenin signaling in cartilaginous metaplasia in Mgp-/- aortae in vivo and during chondrogenic transformation of VSMCs in vitro. Quercetin intercepted chondrogenic transformation of VSM and blocked activation of β-catenin both in vivo and in vitro. Although dietary quercetin drastically attenuated calcifying cartilaginous metaplasia in Mgp-/- animals, approximately one-half of total vascular calcium mineral remained as depositions along elastic lamellae.Quercetin is potent in preventing VSM chondrogenic transformation caused by diverse stimuli. Combined with the demonstrated efficiency of dietary quercetin in preventing ectopic chondrogenesis in the MGP-null vasculature, these findings indicate a potentially broad therapeutic applicability of this safe for human consumption bioflavonoid in the therapy of cardiovascular conditions linked to cartilaginous metaplasia of VSM. Elastocalcinosis is a major component of MGP-null vascular disease and is controlled by a mechanism different from chondrogenic transformation of VSM and not sensitive to quercetin.

  11. Approximate rational Jacobi elliptic function solutions of the fractional differential equations via the enhanced Adomian decomposition method

    International Nuclear Information System (INIS)

    Song Lina; Wang Weiguo

    2010-01-01

    In this Letter, an enhanced Adomian decomposition method which introduces the h-curve of the homotopy analysis method into the standard Adomian decomposition method is proposed. Some examples prove that this method can derive successfully approximate rational Jacobi elliptic function solutions of the fractional differential equations.

  12. Differentiation between benign and malignant colon tumors using fast dynamic gadolinium-enhanced MR colonography; a feasibility study

    DEFF Research Database (Denmark)

    Achiam, M P; Andersen, L P H; Klein, M

    2010-01-01

    Colorectal cancer will present itself as a bowel obstruction in 16-23% of all cases. However, not all obstructing tumors are malignant and the differentiation between a benign and a malignant tumor can be difficult. The purpose of our study was to determine whether fast dynamic gadolinium-enhance...

  13. Incorporation of functionalized gold nanoparticles into nanofibers for enhanced attachment and differentiation of mammalian cells

    Directory of Open Access Journals (Sweden)

    Jung Dongju

    2012-06-01

    Full Text Available Abstract Background Electrospun nanofibers have been widely used as substrata for mammalian cell culture owing to their structural similarity to natural extracellular matrices. Structurally consistent electrospun nanofibers can be produced with synthetic polymers but require chemical modification to graft cell-adhesive molecules to make the nanofibers functional. Development of a facile method of grafting functional molecules on the nanofibers will contribute to the production of diverse cell type-specific nanofiber substrata. Results Small molecules, peptides, and functionalized gold nanoparticles were successfully incorporated with polymethylglutarimide (PMGI nanofibers through electrospinning. The PMGI nanofibers functionalized by the grafted AuNPs, which were labeled with cell-adhesive peptides, enhanced HeLa cell attachment and potentiated cardiomyocyte differentiation of human pluripotent stem cells. Conclusions PMGI nanofibers can be functionalized simply by co-electrospinning with the grafting materials. In addition, grafting functionalized AuNPs enable high-density localization of the cell-adhesive peptides on the nanofiber. The results of the present study suggest that more cell type-specific synthetic substrata can be fabricated with molecule-doped nanofibers, in which diverse functional molecules are grafted alone or in combination with other molecules at different concentrations.

  14. Constructing HVS-Based Optimal Substitution Matrix Using Enhanced Differential Evolution

    Directory of Open Access Journals (Sweden)

    Shu-Fen Tu

    2013-01-01

    Full Text Available Least significant bit (LSB substitution is a method of information hiding. The secret message is embedded into the last k bits of a cover-image in order to evade the notice of hackers. The security and stego-image quality are two main limitations of the LSB substitution method. Therefore, some researchers have proposed an LSB substitution matrix to address these two issues. Finding the optimal LSB substitution matrix can be conceptualized as a problem of combinatorial optimization. In this paper, we adopt a different heuristic method based on other researchers’ method, called enhanced differential evolution (EDE, to construct an optimal LSB substitution matrix. Differing from other researchers, we adopt an HVS-based measurement as a fitness function and embed the secret by modifying the pixel to a closest value rather than simply substituting the LSBs. Our scheme extracts the secret by modular operations as simple LSB substitution does. The experimental results show that the proposed embedding algorithm indeed improves imperceptibility of stego-images substantially.

  15. Application of enhanced discrete differential evolution approach to unit commitment problem

    International Nuclear Information System (INIS)

    Yuan Xiaohui; Su Anjun; Nie Hao; Yuan Yanbin; Wang Liang

    2009-01-01

    This paper proposes a discrete binary differential evolution (DBDE) approach to solve the unit commitment problem (UCP). The proposed method is enhanced by priority list based on the unit characteristics and heuristic search strategies to handle constraints effectively. The implementation of the proposed method for UCP consists of three stages. Firstly, the DBDE based on priority list is applied for unit scheduling when neglecting the minimum up/down time constraints. Secondly, repairing strategies are used to handle the minimum up/down time constraints and decommit excess spinning reserve units. Finally, heuristic unit substitution search and gray zone modification algorithm are used to improve optimal solution further. Furthermore, the effects of two crucial parameters on performance of the DBDE for solving UCP are studied as well. To verify the advantages of the method, the proposed method is tested and compared to the other methods on the systems with the number of units in the range of 10-100. Numerical results demonstrate that the proposed method is superior to other methods reported in the literature.

  16. Differential diagnosis of gastric cancer and gastritis: the role of contrast-enhanced ultrasound (CEUS).

    Science.gov (United States)

    Xue, Heng; Ge, Hui-Yu; Miao, Li-Ying; Wang, Shu-Min; Zhao, Bo; Wang, Jin-Rui; Cui, Li-Gang

    2017-03-01

    To evaluate the diagnostic performance of contrast-enhanced ultrasound (CEUS) in differential diagnosis of gastric cancer and gastritis, with histological results as reference standard. From September 2011 to August 2014, 82 patients (50 males and 32 females; mean age ± SD, 59.5 ± 15.0 years; range 19-91 years) with gastric cancer or gastritis were included in this Ethics Committee-approved prospective study. Conventional ultrasonography (US) and CEUS were applied to distinguish the two lesions, and both qualitative and quantitative features were evaluated. Of the 82 histopathologic-proven lesions, 58 were cancer and 24 were gastritis. For US, the gastric wall stratification was not preserved in about one-third of cancer (21/58, 36.2%) compared with gastritis (0/24, 0%) (p gastritis (19/24, 79.2%, p gastritis. CEUS has the potential to make the diagnosis more accurate.

  17. Uremic Toxins Enhance Statin-Induced Cytotoxicity in Differentiated Human Rhabdomyosarcoma Cells

    Directory of Open Access Journals (Sweden)

    Hitoshi Uchiyama

    2014-09-01

    Full Text Available The risk of myopathy and rhabdomyolysis is considerably increased in statin users with end-stage renal failure (ESRF. Uremic toxins, which accumulate in patients with ESRF, exert cytotoxic effects that are mediated by various mechanisms. Therefore, accumulation of uremic toxins might increase statin-induced cytotoxicity. The purpose of this study was to determine the effect of four uremic toxins—hippuric acid, 3-carboxy-4-methyl-5-propyl-2-furanpropionate, indole-3-acetic acid, and 3-indoxyl sulfate—on statin-induced myopathy. Differentiated rhabdomyosarcoma cells were pre-treated with the uremic toxins for seven days, and then the cells were treated with pravastatin or simvastatin. Cell viability and apoptosis were assessed by viability assays and flow cytometry. Pre-treatment with uremic toxins increased statin- but not cisplatin-induced cytotoxicity (p < 0.05 vs. untreated. In addition, the pre-treatment increased statin-induced apoptosis, which is one of the cytotoxic factors (p < 0.05 vs. untreated. However, mevalonate, farnesol, and geranylgeraniol reversed the effects of uremic toxins and lowered statin-induced cytotoxicity (p < 0.05 vs. untreated. These results demonstrate that uremic toxins enhance statin-induced apoptosis and cytotoxicity. The mechanism underlying this effect might be associated with small G-protein geranylgeranylation. In conclusion, the increased severity of statin-induced rhabdomyolysis in patients with ESRF is likely due to the accumulation of uremic toxins.

  18. The role of catechol-O-methyltransferase in catechol-enhanced erythroid differentiation of K562 cells.

    Science.gov (United States)

    Suriguga; Li, Xiao-Fei; Li, Yang; Yu, Chun-Hong; Li, Yi-Ran; Yi, Zong-Chun

    2013-12-15

    Catechol is widely used in pharmaceutical and chemical industries. Catechol is also one of phenolic metabolites of benzene in vivo. Our previous study showed that catechol improved erythroid differentiation potency of K562 cells, which was associated with decreased DNA methylation in erythroid specific genes. Catechol is a substrate for the catechol-O-methyltransferase (COMT)-mediated methylation. In the present study, the role of COMT in catechol-enhanced erythroid differentiation of K562 cells was investigated. Benzidine staining showed that exposure to catechol enhanced hemin-induced hemoglobin accumulation and induced mRNA expression of erythroid specific genes in K562 cells. Treatment with catechol caused a time- and concentration-dependent increase in guaiacol concentration in the medium of cultured K562 cells. When COMT expression was knocked down by COMT shRNA expression in K562 cells, the production of guaiacol significantly reduced, and the sensitivity of K562 cells to cytotoxicity of catechol significantly increased. Knockdown of COMT expression by COMT shRNA expression also eliminated catechol-enhanced erythroid differentiation of K562 cells. In addition, the pre-treatment with methyl donor S-adenosyl-L-methionine or its demethylated product S-adenosyl-L-homocysteine induced a significant increase in hemin-induced Hb synthesis in K562 cells and the mRNA expression of erythroid specific genes. These findings indicated that O-methylation catalyzed by COMT acted as detoxication of catechol and involved in catechol-enhanced erythroid differentiation of K562 cells, and the production of S-adenosyl-L-homocysteine partly explained catechol-enhanced erythroid differentiation. © 2013.

  19. Evaluation of dynamic enhanced CT scanning in the differentiation of adrenal lipid-poor adenomas with metastases

    International Nuclear Information System (INIS)

    Fang Xiangming; Hu Chunhong; Hu Xiaoyun; Chen Hongwei; Wu Liyuan; Zou Xinnong; Qian Pingyan

    2006-01-01

    Objective: To evaluate dynamic enhanced CT in differentiating adrenal metastases from adrenal lipid-poor adenomas(ALPA). Methods: Both plain and dynamic enhanced CT scanning was performed in 9 metastases with 13 masses and 28 lipid-poor adenoma with 30 masses. The types of time-density curve according to peak time(PT) and relative washout percentage(Washr) besides shape, size, margin, internal structure, surrounding status and enhanced pattern of each lesion were measuerd and compared between the two groups of metastases and ALPA. Results: There is difference between metastases and ALPA in the aspects of shape, density, neighboring structure and the type of enhancement. The type of TDC of matastases was characterized by fast-washin and fast-washout, which was quite differed from the type of TDC of ALPA characterized by fast-washin and slow-washout. According to this, the sensitiveity and specificity for differentiating metastases from ALPA were 96.7%, 92.3%. Conclusion: The types of TDC of dynamic enhanced CT is of great value in differentiating metastases from ALPA. (authors)

  20. Epigenomic analysis of primary human T cells reveals enhancers associated with TH2 memory cell differentiation and asthma susceptibility

    Science.gov (United States)

    Seumois, Grégory; Chavez, Lukas; Gerasimova, Anna; Lienhard, Matthias; Omran, Nada; Kalinke, Lukas; Vedanayagam, Maria; Ganesan, Asha Purnima V; Chawla, Ashu; Djukanović, Ratko; Ansel, K Mark; Peters, Bjoern; Rao, Anjana; Vijayanand, Pandurangan

    2014-01-01

    A characteristic feature of asthma is the aberrant accumulation, differentiation or function of memory CD4+ T cells that produce type 2 cytokines (TH2 cells). By mapping genome-wide histone modification profiles for subsets of T cells isolated from peripheral blood of healthy and asthmatic individuals, we identified enhancers with known and potential roles in the normal differentiation of human TH1 cells and TH2 cells. We discovered disease-specific enhancers in T cells that differ between healthy and asthmatic individuals. Enhancers that gained the histone H3 Lys4 dimethyl (H3K4me2) mark during TH2 cell development showed the highest enrichment for asthma-associated single nucleotide polymorphisms (SNPs), which supported a pathogenic role for TH2 cells in asthma. In silico analysis of cell-specific enhancers revealed transcription factors, microRNAs and genes potentially linked to human TH2 cell differentiation. Our results establish the feasibility and utility of enhancer profiling in well-defined populations of specialized cell types involved in disease pathogenesis. PMID:24997565

  1. Y-27632 enhances differentiation of blastocyst like cystic human embryoid bodies to endocrinologically active trophoblast cells on a biomimetic platform

    Directory of Open Access Journals (Sweden)

    Totey Satish M

    2009-09-01

    Full Text Available Abstract Trophoblast differentiation and formation of the placenta are important events linked to post-implantation embryonic development. Models mimicking the biology of trophoblast differentiation in a post-implantation maternal microenvironment are needed for understanding disorders like placental-ischemia or for applications in drug-screening, and would help in overcoming the ethical impasse on using human embryos for such research. Here we attempt to create such a model by using embryoid bodies (EBs and a biomimetic platform composed of a bilayer of fibronectin and gelatin on top of low-melting agarose. Using this model we test the hypothesis that cystic-EBs (day 30 that resemble blastocysts morphologically, are better sources as compared to noncytic EBs (day 10, for functional trophoblast differentiation; and that the Rho kinases inhibitor Y27632 can enhance this differentiation. Non/cytic EBs with/out Y27632 were grown on this platform for 28 days, and screened from secretion and expression of trophoblast and other lineage markers using ECLIA, RT-PCR, and Immunofluorescence. All EBs attached on this surface and rapidly proliferated into hCG and progesterone (P2 secreting functional trophoblast cells. However, the cells derived from cytic-EBs and cytic-EBs+ Y27632 showed the maximum secretion of these hormones and expressed IGF2, supporting our hypothesis. Also Y27632 reduced extraembryonic endoderm and trophoblast lineage differentiation from early noncystic-EBs, whereas, it specifically enhanced the induction of trophoblast and multinucleated syncitiotrophoblast differentiation from late cystic-EBs. In vivo trophoblast differentiation can be replicated in fibronectin based biomaterials, using cytic-EBs and by maneuvering the Rho-ROCK pathways. Response of EBs to a compound may vary temporally, and determination of their right stage is crucial for applications in directed-differentiation or drug-screening.

  2. The value of contrast-enhanced 64-row CT in differentiating benign from malignant serous ovarian neoplasms

    International Nuclear Information System (INIS)

    Dong Tianfa; Wu Meixian; Zhang Jiayun; Song Ting

    2009-01-01

    Objective: To assess the diagnostic value of contrast-enhanced 64-row CT scanning in deciding benign or malignant serous ovarian tumors. Methods: Fifty-eight cases of serous ovarian tumors proved pathologically were reviewed, including 25 malignant tumors, 25 benign, 8 borderline tumors. All patients underwent 64-row CT scanning, including plain scanning and contrast-enhance scanning. The tumors' shape, density, blood supply and enhancement features were evaluated. Results: Twenty-five cases of benign serous cystic adenoma were mostly unicameral, and showed a moderate mural enhancement only in 4 cases (16%) due to chronic pelvic infection and the others (21/25, 84%) had no of slight enhancement. Malignant tumors were cystic-solid mass with unclear margin, irregular shape and septa. Twenty-two cases of serous cystadenocarcinoma out of 25 cases (88%) appeared obvious enhancement and other 3 cases no enhancement. And 7 cases out of 8 (87.5%) borderlined serous cystadenomas showed different enhancement patterns. Conclusion: Benign ovarian serous neoplasms were mostly unicameral and no strong mural enhancement, suggesting a lack of blood supply. While, there were obvious enhancement in the ovarian serous cystadenocarcinoma and borderline serous cystadenoma with malignant potential. The 64-row CT is helpful for differentiating the nature of the serous ovarian neoplasm. (authors)

  3. Enhancement of ATRA-induced differentiation of neuroblastoma cells with LOX/COX inhibitors: an expression profiling study

    Directory of Open Access Journals (Sweden)

    Hermanova Marketa

    2010-05-01

    Full Text Available Abstract Background We performed expression profiling of two neuroblastoma cell lines, SK-N-BE(2 and SH-SY5Y, after combined treatment with all-trans retinoic acid (ATRA and inhibitors of lipoxygenases (LOX and cyclooxygenases (COX. This study is a continuation of our previous work confirming the possibility of enhancing ATRA-induced cell differentiation in these cell lines by the application of LOX/COX inhibitors and brings more detailed information concerning the mechanisms of the enhancement of ATRA-induced differentiation of neuroblastoma cells. Methods Caffeic acid, as an inhibitor of 5-lipoxygenase, and celecoxib, as an inhibitor on cyclooxygenase-2, were used in this study. Expression profiling was performed using Human Cancer Oligo GEArray membranes that cover 440 cancer-related genes. Results Cluster analyses of the changes in gene expression showed the concentration-dependent increase in genes known to be involved in the process of retinoid-induced neuronal differentiation, especially in cytoskeleton remodeling. These changes were detected in both cell lines, and they were independent of the type of specific inhibitors, suggesting a common mechanism of ATRA-induced differentiation enhancement. Furthermore, we also found overexpression of some genes in the same cell line (SK-N-BE(2 or SH-SY5Y after combined treatment with both ATRA and CA, or ATRA and CX. Finally, we also detected that gene expression was changed after treatment with the same inhibitor (CA or CX in combination with ATRA in both cell lines. Conclusions Obtained results confirmed our initial hypothesis of the common mechanism of enhancement in ATRA-induced cell differentiation via inhibition of arachidonic acid metabolic pathway.

  4. Enhancement of ATRA-induced differentiation of neuroblastoma cells with LOX/COX inhibitors: an expression profiling study.

    Science.gov (United States)

    Chlapek, Petr; Redova, Martina; Zitterbart, Karel; Hermanova, Marketa; Sterba, Jaroslav; Veselska, Renata

    2010-05-11

    We performed expression profiling of two neuroblastoma cell lines, SK-N-BE(2) and SH-SY5Y, after combined treatment with all-trans retinoic acid (ATRA) and inhibitors of lipoxygenases (LOX) and cyclooxygenases (COX). This study is a continuation of our previous work confirming the possibility of enhancing ATRA-induced cell differentiation in these cell lines by the application of LOX/COX inhibitors and brings more detailed information concerning the mechanisms of the enhancement of ATRA-induced differentiation of neuroblastoma cells. Caffeic acid, as an inhibitor of 5-lipoxygenase, and celecoxib, as an inhibitor on cyclooxygenase-2, were used in this study. Expression profiling was performed using Human Cancer Oligo GEArray membranes that cover 440 cancer-related genes. Cluster analyses of the changes in gene expression showed the concentration-dependent increase in genes known to be involved in the process of retinoid-induced neuronal differentiation, especially in cytoskeleton remodeling. These changes were detected in both cell lines, and they were independent of the type of specific inhibitors, suggesting a common mechanism of ATRA-induced differentiation enhancement. Furthermore, we also found overexpression of some genes in the same cell line (SK-N-BE(2) or SH-SY5Y) after combined treatment with both ATRA and CA, or ATRA and CX. Finally, we also detected that gene expression was changed after treatment with the same inhibitor (CA or CX) in combination with ATRA in both cell lines. Obtained results confirmed our initial hypothesis of the common mechanism of enhancement in ATRA-induced cell differentiation via inhibition of arachidonic acid metabolic pathway.

  5. Differentiation of focal-type autoimmune pancreatitis from pancreatic carcinoma: assessment by multiphase contrast-enhanced CT

    Energy Technology Data Exchange (ETDEWEB)

    Furuhashi, Naohiro; Suzuki, Kojiro; Sakurai, Yusuke; Naganawa, Shinji [Nagoya University Graduate School of Medicine, Department of Radiology, Nagoya (Japan); Ikeda, Mitsuru [Nagoya University Graduate School of Medicine, Department of Radiological Technology, Nagoya (Japan); Kawai, Yuichi [Japanese Red Cross Nagoya Daiichi Hospital, Department of Diagnostic Radiology, Nagoya (Japan)

    2015-05-01

    To evaluate the utility of multiphase contrast-enhanced computed tomography (CT) findings alone and in combination for differentiating focal-type autoimmune pancreatitis (f-AIP) from pancreatic carcinoma (PC). The study group comprised 22 f-AIP lesions and 61 PC lesions. Two radiologists independently evaluated CT findings. Frequencies of findings were compared between f-AIP and PC. Statistical, univariate and multivariate analyses were performed. Homogeneous enhancement during the portal phase (AIP, 59 % vs. PC, 3 %; P < 0.001), dotted enhancement during the pancreatic phase (50 % vs. 7 %; P < 0.001), duct-penetrating sign (46 % vs. 2 %; P < 0.001), enhanced duct sign (36 % vs. 2 %; P < 0.001) and capsule-like rim (46 % vs. 3 %; P < 0.001) were more frequently observed in AIP. Ring-like enhancement during the delayed phase (5 % vs. 46 %; P < 0.001) and peripancreatic strands with a length of at least 10 mm (5 % vs. 39 %; P = 0.001) were more frequently observed in PC. AIP was identified with 82 % sensitivity and 98 % specificity using four of these seven findings. Multivariate analysis revealed significant differences in dotted enhancement (P = 0.004), duct-penetrating sign (P < 0.001) and capsule-like rim (P = 0.007). The combination of CT findings may allow improvements in differentiating f-AIP from PC. (orig.)

  6. Influence of bone marrow-derived mesenchymal stem cells pre-implantation differentiation approach on periodontal regeneration in vivo.

    Science.gov (United States)

    Cai, Xinjie; Yang, Fang; Yan, Xiangzhen; Yang, Wanxun; Yu, Na; Oortgiesen, Daniel A W; Wang, Yining; Jansen, John A; Walboomers, X Frank

    2015-04-01

    The implantation of bone marrow-derived mesenchymal stem cells (MSCs) has previously been shown successful to achieve periodontal regeneration. However, the preferred pre-implantation differentiation strategy (e.g. maintenance of stemness, osteogenic or chondrogenic induction) to obtain optimal periodontal regeneration is still unknown. This in vivo study explored which differentiation approach is most suitable for periodontal regeneration. Mesenchymal stem cells were obtained from Fischer rats and seeded onto poly(lactic-co-glycolic acid)/poly(ɛ-caprolactone) electrospun scaffolds, and then pre-cultured under different in vitro conditions: (i) retention of multilineage differentiation potential; (ii) osteogenic differentiation approach; and (iii) chondrogenic differentiation approach. Subsequently, the cell-scaffold constructs were implanted into experimental periodontal defects of Fischer rats, with empty scaffolds as controls. After 6 weeks of implantation, histomorphometrical analyses were applied to evaluate the regenerated periodontal tissues. The chondrogenic differentiation approach showed regeneration of alveolar bone and ligament tissues. The retention of multilineage differentiation potential supported only ligament regeneration, while the osteogenic differentiation approach boosted alveolar bone regeneration. Chondrogenic differentiation of MSCs before implantation is a useful strategy for regeneration of alveolar bone and periodontal ligament, in the currently used rat model. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. SFRP2 enhances the osteogenic differentiation of apical papilla stem cells by antagonizing the canonical WNT pathway.

    Science.gov (United States)

    Jin, Luyuan; Cao, Yu; Yu, Guoxia; Wang, Jinsong; Lin, Xiao; Ge, Lihua; Du, Juan; Wang, Liping; Diao, Shu; Lian, Xiaomeng; Wang, Songlin; Dong, Rui; Shan, Zhaochen

    2017-01-01

    Exploring the molecular mechanisms underlying directed differentiation is helpful in the development of clinical applications of mesenchymal stem cells (MSCs). Our previous study on dental tissue-derived MSCs demonstrated that secreted frizzled-related protein 2 (SFRP2), a Wnt inhibitor, could enhance osteogenic differentiation in stem cells from the apical papilla (SCAPs). However, how SFRP2 promotes osteogenic differentiation of dental tissue-derived MSCs remains unclear. In this study, we used SCAPs to investigate the underlying mechanisms. SCAPs were isolated from the apical papilla of immature third molars. Western blot and real-time RT-PCR were applied to detect the expression of β-catenin and Wnt target genes. Alizarin Red staining, quantitative calcium analysis, transwell cultures and in vivo transplantation experiments were used to study the osteogenic differentiation potential of SCAPs. SFRP2 inhibited canonical Wnt signaling by enhancing phosphorylation and decreasing the expression of nuclear β-catenin in vitro and in vivo . In addition, the target genes of the Wnt signaling pathway, AXIN2 (axin-related protein 2) and MMP7 (matrix metalloproteinase-7), were downregulated by SFRP2 . WNT1 inhibited the osteogenic differentiation potential of SCAPs. SFRP2 could rescue this WNT1 -impaired osteogenic differentiation potential. The results suggest that SFRP2 could bind to locally present Wnt ligands and alter the balance of intracellular Wnt signaling to antagonize the canonical Wnt pathway in SCAPs. This elucidates the molecular mechanism underlying the SFRP2-mediated directed differentiation of SCAPs and indicates potential target genes for improving dental tissue regeneration.

  8. Long-Term Expandable SOX9+ Chondrogenic Ectomesenchymal Cells from Human Pluripotent Stem Cells

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    Katsutsugu Umeda

    2015-04-01

    Full Text Available Here we report the successful generation and long-term expansion of SOX9-expressing CD271+PDGFRα+CD73+ chondrogenic ectomesenchymal cells from the PAX3/SOX10/FOXD3-expressing MIXL1−CD271hiPDGFRαloCD73− neural crest-like progeny of human pluripotent stem cells in a chemically defined medium supplemented with Nodal/Activin/transforming growth factorβ (TGFβ inhibitor and fibroblast growth factor (FGF. When “primed” with TGFβ, such cells efficiently formed translucent cartilage particles, which were completely mineralized in 12 weeks in immunocompromized mice. The ectomesenchymal cells were expandable without loss of chondrogenic potential for at least 16 passages. They maintained normal karyotype for at least 10 passages and expressed genes representing embryonic progenitors (SOX4/12, LIN28A/B, cranial mesenchyme (ALX1/3/4, and chondroprogenitors (SOX9, COL2A1 of neural crest origin (SOX8/9, NGFR, NES. Ectomesenchyme is a source of many craniofacial bone and cartilage structures. The method we describe for obtaining a large quantity of human ectomesenchymal cells will help to model craniofacial disorders in vitro and potentially provide cells for the repair of craniofacial damage.

  9. The chondrogenic response to exercise in the proximal femur of normal and mdx mice

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    Nye David J

    2010-09-01

    Full Text Available Abstract Background Submaximal exercise is used in the management of muscular dystrophy. The effects of mechanical stimulation on skeletal development are well understood, although its effects on cartilage growth have yet to be investigated in the dystrophic condition. The objective of this study was to investigate the chondrogenic response to voluntary exercise in dystrophin-deficient mice. Methods Control and dystrophin-deficient (mdx mice were divided into sedentary and exercise-treated groups and tested for chondral histomorphometric differences at the proximal femur. Results Control mice ran 7 km/week further than mdx mice on average, but this difference was not statistically significant (P > 0.05. However, exercised control mice exhibited significantly enlarged femur head diameter, articular cartilage thickness, articular cartilage tissue area, and area of calcified cartilage relative to sedentary controls and exercised mdx mice (P Conclusions Mdx mice exhibit a reduced chondrogenic response to increased mechanical stimulation relative to controls. However, no significant reduction in articular dimensions was found, indicating loss of chondral tissue may not be a clinical concern with dystrophinopathy.

  10. The effect of dexamethasone and triiodothyronine on terminal differentiation of primary bovine chondrocytes and chondrogenically differentiated mesenchymal stem cells

    NARCIS (Netherlands)

    Randau, T.M.; Schildberg, F.A.; Alini, M.; Wimmer, M.D.; Haddouti, E.-M.; Gravius, S.; Ito, K.; Stoddart, M.J.

    2013-01-01

    The newly evolved field of regenerative medicine is offering solutions in the treatment of bone or cartilage loss and deficiency. Mesenchymal stem cells, as well as articular chondrocytes, are potential cells for the generation of bone or cartilage. The natural mechanism of bone formation is that of

  11. Hybrid Complexes of High and Low Molecular Weight Hyaluronans Highly Enhance HASCs Differentiation: Implication for Facial Bioremodelling

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    Antonietta Stellavato

    2017-11-01

    Full Text Available Background/Aims: Adipose-derived Stem Cells (ASCs are used in Regenerative Medicine, including fat grafting, recovery from local tissue ischemia and scar remodeling. The aim of this study was to evaluate hyaluronan based gel effects on ASCs differentiation and proliferation. Methods: Comparative analyses using high (H and low (L molecular weight hyaluronans (HA, hyaluronan hybrid cooperative complexes (HCCs, and high and medium cross-linked hyaluronan based dermal fillers were performed. Human ASCs were characterized by flow cytometry using CD90, CD34, CD105, CD29, CD31, CD45 and CD14 markers. Then, cells were treated for 7, 14 and 21 days with hyaluronans. Adipogenic differentiation was evaluated using Oil red-O staining and expression of leptin, PPAR-γ, LPL and adiponectin using qRT-PCR. Adiponectin was analyzed by immunofluorescence, PPAR-γ and adiponectin were analyzed using western blotting. ELISA assays for adiponectin and leptin were performed. Results: HCCs highly affected ASCs differentiation by up-regulating adipogenic genes and related proteins, that were also secreted in the culture medium. H-HA and L-HA induced a lower level of ASCs differentiation. Conclusion: HCCs-based formulations clearly enhance adipogenic differentiation and proliferation, when compared with linear HA and cross-linked hyaluronans. Injection of HCCs in subdermal fat compartment may recruit and differentiate stem cells in adipocytes, and considerably improving fat tissue renewal.

  12. Increased expression of enhancer of Zeste Homolog 2 (EZH2) differentiates squamous cell carcinoma from normal skin and actinic keratosis.

    Science.gov (United States)

    Xie, Qiang; Wang, Hongbei; Heilman, Edward R; Walsh, Michael G; Haseeb, M A; Gupta, Raavi

    2014-01-01

    Enhancer of Zeste Homolog 2 (EZH2) is a polycomb group protein that has been shown to be involved in the progression of multiple human cancers including melanoma. The expression of EZH2 in normal skin and in pre-malignant and malignant cutaneous squamous cell carcinoma (SCC) has not been studied. We examined the expression of EZH2 in normal skin, actinic keratosis (AK), SCC in situ, well-differentiated (SCC-WD), moderately-differentiated (SCC-MD) and poorly-differentiated SCC (SCC-PD) to ascertain whether EZH2 expression differentiates these conditions. Immunohistochemical staining for EZH2 was performed on formalin-fixed paraffin-embedded biopsies and a tissue microarray containing normal skin, AK, SCC in situ, and SCC of different grades. In comparison to the normal skin, EZH2 expression in actinic keratosis was increased (p=0.03). Similarly, EZH2 expression in all of the neoplastic conditions studied (SCC in situ, SCC-WD, SCC-MD and SCC-PD) was greatly increased in comparison to both the normal skin and actinic keratosis (p≤0.001). EZH2 expression increases incrementally from normal skin to AK and further to SCC, suggesting a role for EZH2 in the progression and differentiation of SCC. EZH2 expression may be used as a diagnostic marker for differentiating SCC from AK or normal skin.

  13. Enhanced differentiation of neural stem cells to neurons and promotion of neurite outgrowth by oxygen-glucose deprivation.

    Science.gov (United States)

    Wang, Qin; Yang, Lin; Wang, Yaping

    2015-06-01

    Stroke has become the leading cause of mortality worldwide. Hypoxic or ischemic insults are crucial factors mediating the neural damage in the brain tissue of stroke patients. Neural stem cells (NSCs) have been recognized as a promising tool for the treatment of ischemic stroke and other neurodegenerative diseases due to their inducible pluripotency. In this study, we aim to mimick the cerebral hypoxic-ischemic injury in vitro using oxygen-glucose deprivation (OGD) strategy, and evaluate the effects of OGD on the NSC's neural differentiation, as well as the differentiated neurite outgrowth. Our data showed that NSCs under the short-term 2h OGD treatment are able to maintain cell viability and the capability to form neurospheres. Importantly, this moderate OGD treatment promotes NSC differentiation to neurons and enhances the performance of the mature neuronal networks, accompanying increased neurite outgrowth of differentiated neurons. However, long-term 6h and 8h OGD exposures in NSCs lead to decreased cell survival, reduced differentiation and diminished NSC-derived neurite outgrowth. The expressions of neuron-specific microtubule-associated protein 2 (MAP-2) and growth associated protein 43 (GAP-43) are increased by short-term OGD treatments but suppressed by long-term OGD. Overall, our results demonstrate that short-term OGD exposure in vitro induces differentiation of NSCs while maintaining their proliferation and survival, providing valuable insights of adopting NSC-based therapy for ischemic stroke and other neurodegenerative disorders. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Interleukin-6 enhances human Ig production, but not as a terminal differentiation factor for B lymphocytes

    NARCIS (Netherlands)

    van Kooten, C.; van Oers, M. H.; Aarden, L. A.

    1990-01-01

    Most B-cell differentiation systems are complicated by the fact that they are both T-cell- and monocyte-dependent. Immobilized anti-CD3 antibodies induce monocyte-independent T-cell activation, allowing investigation of the role of interleukin-6 (IL6) in the process of B-cell differentiation. We

  15. Training with Differential Outcomes Enhances Discriminative Learning and Visuospatial Recognition Memory in Children Born Prematurely

    Science.gov (United States)

    Martinez, Lourdes; Mari-Beffa, Paloma; Roldan-Tapia, Dolores; Ramos-Lizana, Julio; Fuentes, Luis J.; Estevez, Angeles F.

    2012-01-01

    Previous studies have demonstrated that discriminative learning is facilitated when a particular outcome is associated with each relation to be learned. When this training procedure is applied (the differential outcome procedure; DOP), learning is faster and more accurate than when the more common non-differential outcome procedure is used. This…

  16. Assessment of MRI and dynamic contrast-enhanced MRI in the differential diagnosis of adenomatoid odontogenic tumor

    International Nuclear Information System (INIS)

    Asaumi, Jun-ichi; Yanagi, Yoshinobu; Konouchi, Hironobu; Hisatomi, Miki; Matsuzaki, Hidenobu; Shigehara, Hiroshi; Kishi, Kanji

    2004-01-01

    The radiographical differentiation of adenomatoid odontogenic tumor (AOT) from dentigerous cysts, calcifying odontogenic cysts, calcifying epithelial odontogenic tumors, odontogenic keratocysts and amelobastomas is sometimes difficult. We attempted to differentiate AOT from other lesions similar to AOT in radiographic findings using MRI. The MRI features of AOT in our three cases included homogeneous low SI in the cystic portion and homogeneous intermediate SI in the solid portion on T1WI, homogeneous high SI in the cystic portion and intermediate to slightly high SI in the solid portion on T2WI and enhancement of only the solid portion on CE-T1WI although none of the sequences included SI of calcifications. The contrast index curves in the three cases of AOT showed a gradual increase to 300 s, which signified a benign tumor. These MRI features were characteristic features of AOT and might be a basis for differentiating AOT from the above possible lesions in radiographic examinations

  17. Fetal Mesenchymal Stromal Cells Differentiating towards Chondrocytes Acquire a Gene Expression Profile Resembling Human Growth Plate Cartilage

    NARCIS (Netherlands)

    van Gool, S.A.; Emons, J.A.M.; Leijten, Jeroen Christianus Hermanus; Decker, E.; Sticht, C.; van Houwelingen, J.C.; Goeman, J.J.; Kleijburg, C.; Scherjon, S.; Gretz, N.; Wit, J.M.; Rappold, G.; Post, Janine Nicole; Karperien, Hermanus Bernardus Johannes

    2012-01-01

    Abstract We used human fetal bone marrow-derived mesenchymal stromal cells (hfMSCs) differentiating towards chondrocytes as an alternative model for the human growth plate (GP). Our aims were to study gene expression patterns associated with chondrogenic differentiation to assess whether

  18. [Value of contrast-enhanced ultrasound (CEUS) in the differential diagnosis between benign and malignant renal neoplasms].

    Science.gov (United States)

    Zhang, Sheng; Wang, Xiao-qing; Xin, Xiao-jie; Xu, Yong

    2013-05-01

    To investigate the value of contrast enhanced ultrasound (CEUS) imaging in the differential diagnosis between benign and malignant renal neoplasms. Two hundred and forty-five cases of renal space-occupying lesions confirmed by biopsy or surgical pathology were included in this study. The CEUS features of the renal space-occupying lesions, i.e., the enhancement degree, homogeneity of enhancement, washing-in and washing-out time and enhancement pattern, were retrospectively analyzed. There were 210 cases of malignant renal tumors and 35 cases of benign lesions. The CEUS modes of the malignant renal tumors included "quick in and quick out" 82 cases, "quick in and slow out" 64 cases, "slow in and quick out" 18 cases and "slow in and slow out" 46 cases; good enhancement 150 cases (71.4%) and inhomogeneous enhancement 180 cases (85.7%).Both the contrast agent filling defect area and solid component enhancement of solid-cystic tumors were important features of malignant renal tumors. In the 35 cases of benign lesions,the CEUS modes included "quick in and quick out" 4 cases, "quick in and slow out" 8 cases, "slow in and quick out" 10 cases and "slow in and slow out" 13 cases. Most of the benign tumors showed low enhancement 51.4% (18/35) and inhomogeneous enhancement 54.3% (19/35). There were significant differences between the malignant and benign renal neoplasms in CEUS mode, degree of enhancement and homogeneity of enhancement (P benign and malignant tumors were 77.1% and 83.8%, respectively, while the two-dimensional ultrasound diagnosis of benign and malignant tumors were 68.6% and 76.7%, respectively, with a significant difference (P benign and malignant renal lesions.

  19. Application of the enhanced homotopy perturbation method to solve the fractional-order Bagley-Torvik differential equation

    Energy Technology Data Exchange (ETDEWEB)

    Zolfaghari, M; Ghaderi, R; Sheikhol Eslami, A; Hosseinnia, S H; Sadati, J [Intelligent System Research Group, Faculty of Electrical and Computer Engineering, Babol, Noushirvani University of Technology, PO Box 47135-484, Babol (Iran, Islamic Republic of); Ranjbar, A [Golestan University, Gorgan (Iran, Islamic Republic of); Momani, S [Department of Mathematics, Mutah University, PO Box 7, Al-Karak (Jordan)], E-mail: h.hoseinnia@stu.nit.ac.ir, E-mail: a.ranjbar@nit.ac.ir, E-mail: shahermm@yahoo.com

    2009-10-15

    The enhanced homotopy perturbation method (EHPM) is applied for finding improved approximate solutions of the well-known Bagley-Torvik equation for three different cases. The main characteristic of the EHPM is using a stabilized linear part, which guarantees the stability and convergence of the overall solution. The results are finally compared with the Adams-Bashforth-Moulton numerical method, the Adomian decomposition method (ADM) and the fractional differential transform method (FDTM) to verify the performance of the EHPM.

  20. Application of the enhanced homotopy perturbation method to solve the fractional-order Bagley-Torvik differential equation

    International Nuclear Information System (INIS)

    Zolfaghari, M; Ghaderi, R; Sheikhol Eslami, A; Hosseinnia, S H; Sadati, J; Ranjbar, A; Momani, S

    2009-01-01

    The enhanced homotopy perturbation method (EHPM) is applied for finding improved approximate solutions of the well-known Bagley-Torvik equation for three different cases. The main characteristic of the EHPM is using a stabilized linear part, which guarantees the stability and convergence of the overall solution. The results are finally compared with the Adams-Bashforth-Moulton numerical method, the Adomian decomposition method (ADM) and the fractional differential transform method (FDTM) to verify the performance of the EHPM.

  1. Application of the enhanced homotopy perturbation method to solve the fractional-order Bagley-Torvik differential equation

    Science.gov (United States)

    Zolfaghari, M.; Ghaderi, R.; Sheikhol Eslami, A.; Ranjbar, A.; Hosseinnia, S. H.; Momani, S.; Sadati, J.

    2009-10-01

    The enhanced homotopy perturbation method (EHPM) is applied for finding improved approximate solutions of the well-known Bagley-Torvik equation for three different cases. The main characteristic of the EHPM is using a stabilized linear part, which guarantees the stability and convergence of the overall solution. The results are finally compared with the Adams-Bashforth-Moulton numerical method, the Adomian decomposition method (ADM) and the fractional differential transform method (FDTM) to verify the performance of the EHPM.

  2. Differentiating Focal Eosinophilic Infiltration from Metastasis in the Liver with Gadoxetic Acid-Enhanced Magnetic Resonance Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Mi Hee; Kim, Seong Hyun; Kim, Hee Jung; Lee, Min Woo; Lee, Won Jae [Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of)

    2011-08-15

    To determine the most useful findings of gadoxetic acid-enhanced 3.0 Tesla (T) MRI for differentiating focal eosinophilic infiltration (FEI) from hepatic metastasis with verification of their usefulness. Pathologically or clinically proven 39 FEIs from 25 patients and 79 hepatic metastases from 51 patients were included in the study. Gadoxetic acid-enhanced 3.0T MRI was performed in all cancer patients. Size differences measured between T2-weighted and hepatobiliary-phase images for lesions > 1 cm and morphologic findings (margin, shape, signal intensity on T1- and T2-weighted images, enhancement pattern on dynamic images, and target appearance on hepatobiliary-phase images) were compared between two groups via Student's t test as well as univariate and multivariate analyses. Diagnostic predictive values of two observers for differentiating two groups were assessed before (session 1) and after (session 2) recognition of results. Mean size difference (2.1 mm) in FEIs between the two images was significantly greater than for metastases (0.7 mm) (p < 0.05). An ill-defined margin and isointensity on T1-weighted images were independently significant morphologic findings (p < 0.05) for differentiating the two groups. All observers achieved a higher diagnostic accuracy in session 2 (97% and 98%) than session 1 (92% and 89%) with statistical significance in observer 2 (p < 0.05). All observers had significantly higher sensitivities (95%) and negative predictive values (NPVs) (98%) in session 2 than in session 1 (sensitivity, 74% in two observers; NPV, 89% and 88%) (p < 0.05). With the size change, an ill-defined margin and isointensity on T1-weighted images are the most useful findings for differentiating FEI from hepatic metastasis on gadoxetic acid-enhanced 3.0T MRI.

  3. Differentiation of mass-forming focal pancreatitis from pancreatic ductal adenocarcinoma: value of characterizing dynamic enhancement patterns on contrast-enhanced MR images by adding signal intensity color mapping

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Mimi [Hanyang University College of Medicine, Department of Radiology, Hanyang Medical Center, Seoul (Korea, Republic of); Jang, Kyung Mi [Sungkyunkwan University School of Medicine, Department of Radiology, Samsung Medical Center, Seoul (Korea, Republic of); Sungkyunkwan University School of Medicine, Department of Radiology and Center for Imaging Science, Samsung Medical Center, Seoul (Korea, Republic of); Kim, Jae-Hun; Jeong, Woo Kyoung; Kim, Seong Hyun; Kang, Tae Wook; Kim, Young Kon; Cha, Dong Ik [Sungkyunkwan University School of Medicine, Department of Radiology, Samsung Medical Center, Seoul (Korea, Republic of); Kim, Kyunga [Samsung Medical Center, Biostatics and Clinical Epidemiology Center, Research Institute for Future Medicine, Seoul (Korea, Republic of)

    2017-04-15

    To evaluate the value of dynamic enhancement patterns on contrast-enhanced MR images by adding signal intensity colour mapping (SICM) to differentiate mass-forming focal pancreatitis (MFFP) from pancreatic ductal adenocarcinoma (PDAC). Forty-one clinicopathologically proven MFFPs and 144 surgically confirmed PDACs were enrolled. Laboratory and MR imaging parameters were used to differentiate MFFP from PDAC. In particular, enhancement patterns on MR images adding SICM were evaluated. By using classification tree analysis (CTA), we determined the predictors for the differentiation of MFFP from PDAC. In the CTA, with all parameters except enhancement pattern on SICM images, ductal obstruction grade and T1 hypointensity grade of the pancreatic lesion were the first and second splitting predictor for differentiation of MFFP from PDAC, in order. By adding an enhancement pattern on the SICM images to CTA, the enhancement pattern was the only splitting predictor to differentiate MFFP from PDAC. The CTA model including enhancement pattern on SICM images has sensitivity of 78.0 %, specificity of 99.3 %, and accuracy of 94.6 % for differentiating MFFP from PDAC. The characterization of enhancement pattern for pancreatic lesions on contrast-enhanced MR images adding SICM would be helpful to differentiate MFFP from PDAC. (orig.)

  4. Enhanced proliferation and dopaminergic differentiation of ventral mesencephalic precursor cells by synergistic effect of FGF2 and reduced oxygen tension

    DEFF Research Database (Denmark)

    Jensen, Pia; Gramsbergen, Jan-Bert; Zimmer, Jens

    2011-01-01

    Effective numerical expansion of dopaminergic precursors might overcome the limited availability of transplantable cells in replacement strategies for Parkinson's disease. Here we investigated the effect of fibroblast growth factor-2 (FGF2) and FGF8 on expansion and dopaminergic differentiation o...... enzyme activity, which may explain the elevated dopamine levels. Our findings demonstrate that modulation of oxygen tension is a recognizable factor for in vitro expansion and dopaminergic differentiation of rat embryonic midbrain precursor cells....... of rat embryonic ventral mesencephalic neuroblasts cultured at high (20%) and low (3%) oxygen tension. More cells incorporated bromodeoxyuridine in cultures expanded at low as compared to high oxygen tension, and after 6 days of differentiation there were significantly more neuronal cells in low than......, switching FGF2-expanded cultures from low to high oxygen tension during the last two days of differentiation significantly enhanced dopamine release and intracellular dopamine levels as compared to all other treatment groups. In addition, the short-term exposure to high oxygen enhanced in situ assessed TH...

  5. The Hypoxia-Mimetic Agent Cobalt Chloride Differently Affects Human Mesenchymal Stem Cells in Their Chondrogenic Potential

    Directory of Open Access Journals (Sweden)

    Gabriella Teti

    2018-01-01

    Full Text Available Adult stem cells are a promising cell source for cartilage regeneration. They resided in a special microenvironment known as the stem-cell niche, characterized by the presence of low oxygen concentration. Cobalt chloride (CoCl2 imitates hypoxia in vitro by stabilizing hypoxia-inducible factor-alpha (HIF-1α, which is the master regulator in the cellular adaptive response to hypoxia. In this study, the influence of CoCl2 on the chondrogenic potential of human MSCs, isolated from dental pulp, umbilical cord, and adipose tissue, was investigated. Cells were treated with concentrations of CoCl2 ranging from 50 to 400 μM. Cell viability, HIF-1α protein synthesis, and the expression of the chondrogenic markers were analyzed. The results showed that the CoCl2 supplementation had no effect on cell viability, while the upregulation of chondrogenic markers such as SOX9, COL2A1, VCAN, and ACAN was dependent on the cellular source. This study shows that hypoxia, induced by CoCl2 treatment, can differently influence the behavior of MSCs, isolated from different sources, in their chondrogenic potential. These findings should be taken into consideration in the treatment of cartilage repair and regeneration based on stem cell therapies.

  6. Macrophage imaging by USPIO-enhanced MR for the differentiation of infectious osteomyelitis and aseptic vertebral inflammation

    Energy Technology Data Exchange (ETDEWEB)

    Bierry, Guillaume [University Hospital, Department of Radiology, Strasbourg (France); University Louis Pasteur, EA 3432, Strasbourg (France); Jehl, Francois [University Hospital of Strasbourg, Department of Bacteriology, Strasbourg (France); University Louis Pasteur, EA 3432, Strasbourg (France); Boehm, Nelly [University Louis Pasteur, Institute of Histology, Faculty of Medicine, Strasbourg (France); University Louis Pasteur, INSERM U666, Strasbourg (France); Robert, Philippe [Guerbet Research, Roissy (France); Dietemann, Jean-Louis; Kremer, Stephane [University Hospital, Department of Radiology, Strasbourg (France)

    2009-07-15

    The purpose of this study was to prospectively evaluate USPIO-enhanced MR imaging for the differentiation of vertebral infectious osteomyelitis and sterile inflammation. Vertebral osteomyelitis and sterile vertebral inflammation were induced in two groups of six rabbits each. MRI examinations were performed including unenhanced and gadolinium-enhanced fat-saturated SE T1w sequences. Once endplate enhancement was observed on the T1 gadolinium-enhanced MR sequence, a second MRI examination (SE T1w sequence) was performed 24 h after USPIO administration (45 {mu}mol Fe/kg). MR imaging was correlated with histopathological findings (macrophage immunostaining and Perls Prussian blue staining). On gadolinium-enhanced T1 sequences, a significant SNR increase in vertebral endplates was present in both groups without significant difference between the two groups (P = 0.26). On USPIO-enhanced T1 sequences, a significant SNR increase was only observed in the infection group (P = 0.03) with a significant difference in SNR between the infection and the sterile-inflammation groups (P = 0.002). Infected areas presented replacement of bone marrow by an intense macrophage infiltration, some being iron-loaded. Sterile inflammation showed a replacement of bone marrow by inflammatory tissue with only rare macrophages without any Perls blue staining. USPIO-enhanced MR imaging can distinguish infectious osteomyelitis from sterile vertebral inflammation due to different macrophage distributions in the two lesions. (orig.)

  7. Hypoxia Enhances Differentiation of Adipose Tissue-Derived Stem Cells toward the Smooth Muscle Phenotype

    Directory of Open Access Journals (Sweden)

    Fang Wang

    2018-02-01

    Full Text Available Smooth muscle differentiated adipose tissue-derived stem cells are a valuable resource for regeneration of gastrointestinal tissues, such as the gut and sphincters. Hypoxia has been shown to promote adipose tissue-derived stem cells proliferation and maintenance of pluripotency, but the influence of hypoxia on their smooth myogenic differentiation remains unexplored. This study investigated the phenotype and contractility of adipose-derived stem cells differentiated toward the smooth myogenic lineage under hypoxic conditions. Oxygen concentrations of 2%, 5%, 10%, and 20% were used during differentiation of adipose tissue-derived stem cells. Real time reverse transcription polymerase chain reaction and immunofluorescence staining were used to detect the expression of smooth muscle cells-specific markers, including early marker smooth muscle alpha actin, middle markers calponin, caldesmon, and late marker smooth muscle myosin heavy chain. The specific contractile properties of cells were verified with both a single cell contraction assay and a gel contraction assay. Five percent oxygen concentration significantly increased the expression levels of α-smooth muscle actin, calponin, and myosin heavy chain in adipose-derived stem cell cultures after 2 weeks of induction (p < 0.01. Cells differentiated in 5% oxygen conditions showed greater contraction effect (p < 0.01. Hypoxia influences differentiation of smooth muscle cells from adipose stem cells and 5% oxygen was the optimal condition to generate smooth muscle cells that contract from adipose stem cells.

  8. Inactivation of EGFR/AKT signaling enhances TSA-induced ovarian cancer cell differentiation.

    Science.gov (United States)

    Shao, Genbao; Lai, Wensheng; Wan, Xiaolei; Xue, Jing; Wei, Ye; Jin, Jie; Zhang, Liuping; Lin, Qiong; Shao, Qixiang; Zou, Shengqiang

    2017-05-01

    Ovarian tumor is one of the most lethal gynecologic cancers, but differentiation therapy for this cancer is poorly characterized. Here, we show that thrichostatin A (TSA), the well known inhibitor of histone deacetylases (HDACs), can induce cell differentiation in HO8910 ovarian cancer cells. TSA-induced cell differentiation is characterized by typical morphological change, increased expression of the differentiation marker FOXA2, decreased expression of the pluripotency markers SOX2 and OCT4, suppressing cell proliferation, and cell cycle arrest in the G1 phase. TSA also induces an elevated expression of cell cycle inhibitory protein p21Cip1 along with a decrease in cell cycle regulatory protein cyclin D1. Significantly, blockage of epidermal growth factor receptor (EGFR) signaling pathway with specific inhibitors of this signaling cascade promotes the TSA-induced differentiation of HO8910 cells. These results imply that the EGFR cascade inhibitors in combination with TSA may represent a promising differentiation therapy strategy for ovarian cancer.

  9. Melatonin enhances neural stem cell differentiation and engraftment by increasing mitochondrial function.

    Science.gov (United States)

    Mendivil-Perez, Miguel; Soto-Mercado, Viviana; Guerra-Librero, Ana; Fernandez-Gil, Beatriz I; Florido, Javier; Shen, Ying-Qiang; Tejada, Miguel A; Capilla-Gonzalez, Vivian; Rusanova, Iryna; Garcia-Verdugo, José M; Acuña-Castroviejo, Darío; López, Luis Carlos; Velez-Pardo, Carlos; Jimenez-Del-Rio, Marlene; Ferrer, José M; Escames, Germaine

    2017-09-01

    Neural stem cells (NSCs) are regarded as a promising therapeutic approach to protecting and restoring damaged neurons in neurodegenerative diseases (NDs) such as Parkinson's disease and Alzheimer's disease (PD and AD, respectively). However, new research suggests that NSC differentiation is required to make this strategy effective. Several studies have demonstrated that melatonin increases mature neuronal markers, which reflects NSC differentiation into neurons. Nevertheless, the possible involvement of mitochondria in the effects of melatonin during NSC differentiation has not yet been fully established. We therefore tested the impact of melatonin on NSC proliferation and differentiation in an attempt to determine whether these actions depend on modulating mitochondrial activity. We measured proliferation and differentiation markers, mitochondrial structural and functional parameters as well as oxidative stress indicators and also evaluated cell transplant engraftment. This enabled us to show that melatonin (25 μM) induces NSC differentiation into oligodendrocytes and neurons. These effects depend on increased mitochondrial mass/DNA/complexes, mitochondrial respiration, and membrane potential as well as ATP synthesis in NSCs. It is also interesting to note that melatonin prevented oxidative stress caused by high levels of mitochondrial activity. Finally, we found that melatonin enriches NSC engraftment in the ND mouse model following transplantation. We concluded that a combined therapy involving transplantation of NSCs pretreated with pharmacological doses of melatonin could efficiently restore neuronal cell populations in PD and AD mouse models depending on mitochondrial activity promotion. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  10. Omega-3 Polyunsaturated Fatty Acids Enhance Neuronal Differentiation in Cultured Rat Neural Stem Cells

    Directory of Open Access Journals (Sweden)

    Masanori Katakura

    2013-01-01

    Full Text Available Polyunsaturated fatty acids (PUFAs can induce neurogenesis and recovery from brain diseases. However, the exact mechanisms of the beneficial effects of PUFAs have not been conclusively described. We recently reported that docosahexaenoic acid (DHA induced neuronal differentiation by decreasing Hes1 expression and increasing p27kip1 expression, which causes cell cycle arrest in neural stem cells (NSCs. In the present study, we examined the effect of eicosapentaenoic acid (EPA and arachidonic acid (AA on differentiation, expression of basic helix-loop-helix transcription factors (Hes1, Hes6, and NeuroD, and the cell cycle of cultured NSCs. EPA also increased mRNA levels of Hes1, an inhibitor of neuronal differentiation, Hes6, an inhibitor of Hes1, NeuroD, and Map2 mRNA and Tuj-1-positive cells (a neuronal marker, indicating that EPA induced neuronal differentiation. EPA increased the mRNA levels of p21cip1 and p27kip1, a cyclin-dependent kinase inhibitor, which indicated that EPA induced cell cycle arrest. Treatment with AA decreased Hes1 mRNA but did not affect NeuroD and Map2 mRNA levels. Furthermore, AA did not affect the number of Tuj-1-positive cells or cell cycle progression. These results indicated that EPA could be involved in neuronal differentiation by mechanisms alternative to those of DHA, whereas AA did not affect neuronal differentiation in NSCs.

  11. Creation of an in vitro microenvironment to enhance human fetal synovium-derived stem cell chondrogenesis.

    Science.gov (United States)

    Li, Jingting; He, Fan; Pei, Ming

    2011-09-01

    Our aim was to assess the feasibility of the sequential application of extracellular matrix (ECM) and low oxygen to enhance chondrogenesis in human fetal synovium-derived stem cells (hfSDSCs). Human fetal synovial fibroblasts (hfSFs) were characterized and found to include hfSDSCs, as evidenced by their multi-differentiation capacity and the surface phenotype markers typical of mesenchymal stem cells. Passage-7 hfSFs were plated on either conventional plastic flasks (P) or ECM deposited by hfSFs (E) for one passage. Passage-8 hfSFs were then reseeded for an additional passage on either P or E. The pellets from expanded hfSFs were incubated in a serum-free chondrogenic medium supplemented with 10 ng/ml transforming growth factor-β3 under either normoxia (21% O(2); 21) or hypoxia (5% O(2); 5) for 14 days. Pellets were collected for evaluation of the treatments (EE21, EE5, EP21, EP5, PE21, PE5, PP21, and PP5) on expanded hfSF chondrogenesis by using histology, immunostaining, biochemistry, and real-time polymerase chain reaction. Our data suggest that, compared with seeding on conventional plastic flasks, hfSFs expanded on ECM exhibit a lower expression of senescence-associated β-galactosidase and an enhanced level of stage-specific embryonic antigen-4. ECM-expanded hfSFs also show increased cell numbers and an enhanced chondrogenic potential. Low oxygen (5% O(2)) during pellet culture enhances hfSF chondrogenesis. Thus, we demonstrate, for the first time, the presence of stem cells in hfSFs, and that modulation of the in vitro microenvironment can enhance hfSDSC chondrogenesis. hfSDSCs might represent a promising cell source for cartilage tissue engineering and regeneration.

  12. Deep Learning with Convolutional Neural Network for Differentiation of Liver Masses at Dynamic Contrast-enhanced CT: A Preliminary Study.

    Science.gov (United States)

    Yasaka, Koichiro; Akai, Hiroyuki; Abe, Osamu; Kiryu, Shigeru

    2018-03-01

    Purpose To investigate diagnostic performance by using a deep learning method with a convolutional neural network (CNN) for the differentiation of liver masses at dynamic contrast agent-enhanced computed tomography (CT). Materials and Methods This clinical retrospective study used CT image sets of liver masses over three phases (noncontrast-agent enhanced, arterial, and delayed). Masses were diagnosed according to five categories (category A, classic hepatocellular carcinomas [HCCs]; category B, malignant liver tumors other than classic and early HCCs; category C, indeterminate masses or mass-like lesions [including early HCCs and dysplastic nodules] and rare benign liver masses other than hemangiomas and cysts; category D, hemangiomas; and category E, cysts). Supervised training was performed by using 55 536 image sets obtained in 2013 (from 460 patients, 1068 sets were obtained and they were augmented by a factor of 52 [rotated, parallel-shifted, strongly enlarged, and noise-added images were generated from the original images]). The CNN was composed of six convolutional, three maximum pooling, and three fully connected layers. The CNN was tested with 100 liver mass image sets obtained in 2016 (74 men and 26 women; mean age, 66.4 years ± 10.6 [standard deviation]; mean mass size, 26.9 mm ± 25.9; 21, nine, 35, 20, and 15 liver masses for categories A, B, C, D, and E, respectively). Training and testing were performed five times. Accuracy for categorizing liver masses with CNN model and the area under receiver operating characteristic curve for differentiating categories A-B versus categories C-E were calculated. Results Median accuracy of differential diagnosis of liver masses for test data were 0.84. Median area under the receiver operating characteristic curve for differentiating categories A-B from C-E was 0.92. Conclusion Deep learning with CNN showed high diagnostic performance in differentiation of liver masses at dynamic CT. © RSNA, 2017 Online

  13. The value of gadoxetic acid-enhanced MRI for differentiation between hepatic microabscesses and metastases in patients with periampullary cancer

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Seo-Youn [Soonchunhyang University College of Medicine, Bucheon Hospital, Department of Radiology, Bucheon (Korea, Republic of); Kim, Young Kon; Cha, Dong Ik; Jeong, Woo Kyoung; Lee, Won Jae [Samsung Medical Center, Sungkyunkwan University School of Medicine, Department of Radiology and Center for Imaging Science, Seoul (Korea, Republic of); Min, Ji Hye [Samsung Medical Center, Sungkyunkwan University School of Medicine, Department of Radiology and Center for Imaging Science, Seoul (Korea, Republic of); Chungnam National University Hospital, Chungnam National University College of Medicine, Department of Radiology, Daejeon (Korea, Republic of)

    2017-10-15

    We aimed to identify features that differentiate hepatic microabscess from hepatic metastasis on gadoxetic acid-enhanced MRI in patients with periampullary cancer. We included 72 patients (31 patients with 83 hepatic microabscesses and 41 patients with 71 hepatic metastases) who had a history of periampullary cancer and underwent gadoxetic acid-enhanced MRI. Image analysis was performed for margin, signal intensity, rim enhancement, perilesional hyperaemia, pattern on DWI and dynamic phases, and size discrepancy between sequences by consensus of two observers. Multivariate analysis revealed that the following significant parameters favour microabscess: a history of bile duct cancer, perilesional hyperaemia, persistent arterial rim enhancement through the transitional phase (TP), and size discrepancy between T1WI and T2WI and between T1WI and hepatobiliary phase image (HBPI). The diagnostic accuracy for microabscess was highest (90.9%) when showing a size discrepancy ≥30% between T1WI and HBPI or persistent arterial rim enhancement through the TP. When the lesion was positive for both these variables, specificity reached 100%. The combination of a size discrepancy between T1WI and HBPI and persistent arterial rim enhancement through the TP represents a reliable MRI feature for distinguishing between hepatic microabscess and metastasis in patients with periampullary cancer. (orig.)

  14. Differential diagnosis between metastatic tumors and nonsolid benign lesions of the liver using ferucarbotran-enhanced MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Higashihara, Hiroki [Department of Radiology, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 5650871 (Japan)], E-mail: h-higashihara@radiol.med.osaka-u.ac.jp; Murakami, Takamichi [Department of Radiology, Kinki University School of Medicine 377-2 Oonohigashi, Osakasayama, Osaka 5898511 (Japan); Kim, Tonsok; Hori, Masatoshi; Onishi, Hiromitsu [Department of Radiology, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 5650871 (Japan); Nakata, Saki [Department of Radiology, Toyonaka Municipal Hospital, 4-14-1 Shibahara Chou, Toyonaka, Osaka 5608565 (Japan); Osuga, Keigo; Tomoda, Kaname; Nakamura, Hironobu [Department of Radiology, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 5650871 (Japan)

    2010-01-15

    Purpose: To evaluate ability of ferucarbotran-enhanced MR imaging (MRI) in differentiating metastases from nonsolid benign lesions of the liver according to signal-intensity characteristics. Materials and methods: Sixty-six consecutive patients, who had 138 focal hepatic lesions (26 cysts, 11 hemangiomas, and 101 metastases), underwent ferucarbotran-enhanced MRI. The signal-intensity pattern of each kind of lesion relative to the liver parenchyma on ferucarbotran-enhanced T2* and heavily T1-weighted gradient-echo images were assessed and categorized into the following three categories: high-intensity and iso-intensity, respectively (category A), high and low (category B), and iso- and low-intensity (category C). For category B, lesions were subdivided into two groups based on single-shot half-Fourier RARE images: category B1 (not significantly high-intensity) and category B2 (significantly high-intensity). Results: Category A had 11 hemangiomas and 2 metastatic tumors, category B1 had 97 metastatic tumors, category B2 had 2 metastatic tumors and 9 cysts, and category C had 17 cysts. When a tumor with a signal intensity of category A was considered to be hemangioma, category B1 metastasis, and category B2 and C cyst, the diagnostic accuracy for differentiating these lesions was 97% (134/138). Conclusion: The combination of signal-intensity pattern on ferucarbotran-enhanced T2*- and heavily T1-weighted gradient-echo MRI has ability to differentiate liver metastases from nonsolid benign lesions. However, T2-weighted single-shot half-Fourier RARE imaging should also be employed to achieve better performance.

  15. Bioreactor cultivation enhances NTEB formation and differentiation of NTES cells into cardiomyocytes.

    Science.gov (United States)

    Lü, Shuanghong; Liu, Sheng; He, Wenjun; Duan, Cuimi; Li, Yanmin; Liu, Zhiqiang; Zhang, Ye; Hao, Tong; Wang, Yanmeng; Li, Dexue; Wang, Changyong; Gao, Shaorong

    2008-09-01

    Autogenic embryonic stem cells established from somatic cell nuclear transfer (SCNT) embryos have been proposed as unlimited cell sources for cell transplantation-based treatment of many genetic and degenerative diseases, which can eliminate the immune rejection that occurs after transplantation. In the present study, pluripotent nuclear transfer ES (NTES) cell lines were successfully established from different strains of mice. One NTES cell line, NT1, with capacity of germline transmission, was used to investigate in vitro differentiation into cardiomyocytes. To optimize differentiation conditions for mass production of embryoid bodies (NTEBs) from NTES cells, a slow-turning lateral vessel (STLV) rotating bioreactor was used for culturing the NTES cells to produce NTEBs compared with a conventional static cultivation method. Our results demonstrated that the NTEBs formed in STLV bioreactor were more uniform in size, and no large necrotic centers with most of the cells in NTEBs were viable. Differentiation of the NTEBs formed in both the STLV bioreactor and static culture into cardiomyocytes was induced by ascorbic acid, and the results demonstrated that STLV-produced NTEBs differentiated into cardiomyocytes more efficiently. Taken together, our results suggested that STLV bioreactor provided a more ideal culture condition, which can facilitate the formation of better quality NTEBs and differentiation into cardiomyocytes more efficiently in vitro.

  16. Indian hedgehog gene transfer is a chondrogenic inducer of human mesenchymal stem cells

    Science.gov (United States)

    2012-01-01

    Introduction To date, no single most-appropriate factor or delivery method has been identified for the purpose of mesenchymal stem cell (MSC)-based treatment of cartilage injury. Therefore, in this study we tested whether gene delivery of the growth factor Indian hedgehog (IHH) was able to induce chondrogenesis in human primary MSCs, and whether it was possible by such an approach to modulate the appearance of chondrogenic hypertrophy in pellet cultures in vitro. Methods First-generation adenoviral vectors encoding the cDNA of the human IHH gene were created by cre-lox recombination and used alone or in combination with adenoviral vectors, bone morphogenetic protein-2 (Ad.BMP-2), or transforming growth factor beta-1 (Ad.TGF-β1) to transduce human bone-marrow derived MSCs at 5 × 102 infectious particles/cell. Thereafter, 3 × 105 cells were seeded into aggregates and cultured for 3 weeks in serum-free medium, with untransduced or marker gene transduced cultures as controls. Transgene expressions were determined by ELISA, and aggregates were analysed histologically, immunohistochemically, biochemically and by RT-PCR for chondrogenesis and hypertrophy. Results IHH, TGF-β1 and BMP-2 genes were equipotent inducers of chondrogenesis in primary MSCs, as evidenced by strong staining for proteoglycans, collagen type II, increased levels of glycosaminoglycan synthesis, and expression of mRNAs associated with chondrogenesis. IHH-modified aggregates, alone or in combination, also showed a tendency to progress towards hypertrophy, as judged by the expression of alkaline phosphatase and stainings for collagen type X and Annexin 5. Conclusion As this study provides evidence for chondrogenic induction of MSC aggregates in vitro via IHH gene delivery, this technology may be efficiently employed for generating cartilaginous repair tissues in vivo. PMID:22817660

  17. Intermittent, low dose carbon monoxide exposure enhances survival and dopaminergic differentiation of human neural stem cells

    DEFF Research Database (Denmark)

    Dreyer-Andersen, Nanna; Almeida, Ana Sofia; Jensen, Pia

    2018-01-01

    cells constitute an alternative source of cells for transplantation in Parkinson's disease, but efficient protocols for controlled dopaminergic differentiation need to be developed. Short-term, low-level carbon monoxide (CO) exposure has been shown to affect signaling in several tissues, resulting...... in both protection and generation of reactive oxygen species. The present study investigated the effect of CO produced by a novel CO-releasing molecule on dopaminergic differentiation of human neural stem cells. Short-term exposure to 25 ppm CO at days 0 and 4 significantly increased the relative content...... of β-tubulin III-immunoreactive immature neurons and tyrosine hydroxylase expressing catecholaminergic neurons, as assessed 6 days after differentiation. Also the number of microtubule associated protein 2-positive mature neurons had increased significantly. Moreover, the content of apoptotic cells...

  18. A Novel Atypical PKC-Iota Inhibitor, Echinochrome A, Enhances Cardiomyocyte Differentiation from Mouse Embryonic Stem Cells.

    Science.gov (United States)

    Kim, Hyoung Kyu; Cho, Sung Woo; Heo, Hye Jin; Jeong, Seung Hun; Kim, Min; Ko, Kyung Soo; Rhee, Byoung Doo; Mishchenko, Natalia P; Vasileva, Elena A; Fedoreyev, Sergey A; Stonik, Valentin A; Han, Jin

    2018-06-02

    Echinochrome A (EchA) is a marine bioproduct extracted from sea urchins having antioxidant, antimicrobial, anti-inflammatory, and chelating effects, and is the active component of the clinical drug histochrome. We investigated the potential use of Ech A for inducing cardiomyocyte differentiation from mouse embryonic stem cells (mESCs). We also assessed the effects of Ech A on mitochondrial mass, inner membrane potential (Δψm), reactive oxygen species generation, and levels of Ca 2+ . To identify the direct target of Ech A, we performed in vitro kinase activity and surface plasmon resonance binding assays. Ech A dose-dependently enhanced cardiomyocyte differentiation with higher beating rates. Ech A (50 μM) increased the mitochondrial mass and membrane potential but did not alter the mitochondrial superoxide and Ca 2+ levels. The in vitro kinase activity of the atypical protein kinase C-iota (PKCι) was significantly decreased by 50 μM of Ech A with an IC 50 for PKCι activity of 107 μM. Computational protein-ligand docking simulation results suggested the direct binding of Ech A to PKCι, and surface plasmon resonance confirmed the direct binding with a low K D of 6.3 nM. Therefore, Ech A is a potential drug for enhancing cardiomyocyte differentiation from mESCs through direct binding to PKCι and inhibition of its activity.

  19. MUM ENHANCERS are important for seed coat mucilage production and mucilage secretory cell differentiation in Arabidopsis thaliana.

    Science.gov (United States)

    Arsovski, Andrej A; Villota, Maria M; Rowland, Owen; Subramaniam, Rajagopal; Western, Tamara L

    2009-01-01

    Pollination triggers not only embryo development but also the differentiation of the ovule integuments to form a specialized seed coat. The mucilage secretory cells of the Arabidopsis thaliana seed coat undergo a complex differentiation process in which cell growth is followed by the synthesis and secretion of pectinaceous mucilage. A number of genes have been identified affecting mucilage secretory cell differentiation, including MUCILAGE-MODIFIED4 (MUM4). mum4 mutants produce a reduced amount of mucilage and cloning of MUM4 revealed that it encodes a UDP-L-rhamnose synthase that is developmentally up-regulated to provide rhamnose for mucilage pectin synthesis. To identify additional genes acting in mucilage synthesis and secretion, a screen for enhancers of the mum4 phenotype was performed. Eight mum enhancers (men) have been identified, two of which result from defects in known mucilage secretory cell genes (MUM2 and MYB61). Our results show that, in a mum4 background, mutations in MEN1, MEN4, and MEN5 lead to further reductions in mucilage compared to mum4 single mutants, suggesting that they are involved in mucilage synthesis or secretion. Conversely, mutations in MEN2 and MEN6 appear to affect mucilage release rather than quantity. With the exception of men4, whose single mutant exhibits reduced mucilage, none of these genes have a single mutant phenotype, suggesting that they would not have been identified outside the compromised mum4 background.

  20. Multi-layer thickness determination using differential-based enhanced Fourier transforms of X-ray reflectivity data

    Energy Technology Data Exchange (ETDEWEB)

    Poust, Benjamin [Department of Materials Science and Engineering, University of California, Los Angeles, CA (United States); Northrop Grumman Space Technology, Redondo Beach, CA (United States); Sandhu, Rajinder [Northrop Grumman Space Technology, Redondo Beach, CA (United States); Goorsky, Mark [Department of Materials Science and Engineering, University of California, Los Angeles, CA (United States)

    2009-08-15

    Layer thickness determination of single and multi-layer structures is achieved using a new method for generating Fourier transforms (FTs) of X-ray reflectivity data. This enhanced Fourier analysis is compared to other techniques in the determination of AlN layer thickness deposited on sapphire. In addition to demonstrably improved results, the results also agree with thicknesses determined using simulations and TEM measurements. The effectiveness of the technique is further demonstrated using the more complicated metamorphic epitaxial multi-layer AlSb/InAs structures deposited on GaAs. The approach reported here is based upon differentiating the specular intensity with respect to the vertical reciprocal space coordinate Q{sub Z}. In general, differentiation is far more effective at removing the sloping background present in reflectivity scans than logarithmic compression alone, average subtraction alone, or other methods. When combined with any of the other enhancement techniques, however, differentiation yields distinguishable discrete Fourier transform (DFT) power spectrum peaks for even the weakest and most truncated of sloping oscillations that are present in many reflectivity scans from multi-layer structures. (Abstract Copyright [2009], Wiley Periodicals, Inc.)

  1. Mechanisms in endocrinology: micro-RNAs: targets for enhancing osteoblast differentiation and bone formation.

    Science.gov (United States)

    Taipaleenmäki, Hanna; Bjerre Hokland, Lea; Chen, Li; Kauppinen, Sakari; Kassem, Moustapha

    2012-03-01

    Osteoblast differentiation and bone formation (osteogenesis) are regulated by transcriptional and post-transcriptional mechanisms. Recently, a novel class of regulatory factors termed micro-RNAs (miRNAs) has been identified as playing an important role in the regulation of many aspects of osteoblast biology including proliferation, differentiation, metabolism and apoptosis. Also, preliminary data from animal disease models suggest that targeting miRNAs in bone can be a novel approach to increase bone mass. This review highlights the current knowledge of miRNA biology and their role in bone formation and discusses their potential use in future therapeutic applications for metabolic bone diseases.

  2. Msx1-modulated muscle satellite cells retain a primitive state and exhibit an enhanced capacity for osteogenic differentiation

    International Nuclear Information System (INIS)

    Ding, Ke; Liu, Wen-ying; Zeng, Qiang; Hou, Fang; Xu, Jian-zhong; Yang, Zhong

    2017-01-01

    Multipotent muscle satellite cells (MuSCs) have been identified as potential seed cells for bone tissue engineering. However, MuSCs exhibit a rapid loss of stemness after in vitro culturing, thereby compromising their therapeutic efficiency. Muscle segment homeobox gene 1 (msx1) has been found to induce the dedifferentiation of committed progenitor cells, as well as terminally differentiated myotubes. In this study, a Tet-off retroviral gene delivery system was used to modulate msx1 expression. After ten passages, MuSCs that did not express msx-1 (e.g., the non-msx1 group) were compared with MuSCs with induced msx-1 expression (e.g., the msx1 group). The latter group exhibited a more juvenile morphology, it contained a significantly lower percentage of senescent cells characterized by positive β-galactosidase staining, and it exhibited increased proliferation and a higher proliferation index. Immunocytochemical stainings further detected a more primitive gene expression profile for the msx1 group, while osteogenic differentiation assays and ectopic bone formation assays demonstrated an improved capacity for the msx1 group to undergo osteogenic differentiation. These results suggest that transient expression of msx1 in MuSCs can retain a primitive state, thereby enhancing their capacity for osteogenic differentiation and restoring the potential for MuSCs to serve as seed cells for bone tissue engineering.

  3. Msx1-modulated muscle satellite cells retain a primitive state and exhibit an enhanced capacity for osteogenic differentiation.

    Science.gov (United States)

    Ding, Ke; Liu, Wen-Ying; Zeng, Qiang; Hou, Fang; Xu, Jian-Zhong; Yang, Zhong

    2017-03-01

    Multipotent muscle satellite cells (MuSCs) have been identified as potential seed cells for bone tissue engineering. However, MuSCs exhibit a rapid loss of stemness after in vitro culturing, thereby compromising their therapeutic efficiency. Muscle segment homeobox gene 1 (msx1) has been found to induce the dedifferentiation of committed progenitor cells, as well as terminally differentiated myotubes. In this study, a Tet-off retroviral gene delivery system was used to modulate msx1 expression. After ten passages, MuSCs that did not express msx-1 (e.g., the non-msx1 group) were compared with MuSCs with induced msx-1 expression (e.g., the msx1 group). The latter group exhibited a more juvenile morphology, it contained a significantly lower percentage of senescent cells characterized by positive β-galactosidase staining, and it exhibited increased proliferation and a higher proliferation index. Immunocytochemical stainings further detected a more primitive gene expression profile for the msx1 group, while osteogenic differentiation assays and ectopic bone formation assays demonstrated an improved capacity for the msx1 group to undergo osteogenic differentiation. These results suggest that transient expression of msx1 in MuSCs can retain a primitive state, thereby enhancing their capacity for osteogenic differentiation and restoring the potential for MuSCs to serve as seed cells for bone tissue engineering. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Msx1-modulated muscle satellite cells retain a primitive state and exhibit an enhanced capacity for osteogenic differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Ding, Ke, E-mail: dingke@med.uestc.edu.cn [Department of Pediatric Surgery, School of medicine, University of Electronic Science and Technology of China, Chengdu 610072 (China); Sichuan Academy of Medical Sciences & Sichuan Provincial People' s Hospital, Chengdu 610072 (China); Department of Orthopaedics, Southwest Hospital, Third Military Medical University, Chongqing 400038 (China); Liu, Wen-ying; Zeng, Qiang; Hou, Fang [Department of Pediatric Surgery, School of medicine, University of Electronic Science and Technology of China, Chengdu 610072 (China); Sichuan Academy of Medical Sciences & Sichuan Provincial People' s Hospital, Chengdu 610072 (China); Xu, Jian-zhong, E-mail: xjzspine@163.com [Department of Orthopaedics, Southwest Hospital, Third Military Medical University, Chongqing 400038 (China); Yang, Zhong, E-mail: zyang1999@163.com [Department of Clinical Hematology, Southwest Hospital, Third Military Medical University, Chongqing 400038 (China)

    2017-03-01

    Multipotent muscle satellite cells (MuSCs) have been identified as potential seed cells for bone tissue engineering. However, MuSCs exhibit a rapid loss of stemness after in vitro culturing, thereby compromising their therapeutic efficiency. Muscle segment homeobox gene 1 (msx1) has been found to induce the dedifferentiation of committed progenitor cells, as well as terminally differentiated myotubes. In this study, a Tet-off retroviral gene delivery system was used to modulate msx1 expression. After ten passages, MuSCs that did not express msx-1 (e.g., the non-msx1 group) were compared with MuSCs with induced msx-1 expression (e.g., the msx1 group). The latter group exhibited a more juvenile morphology, it contained a significantly lower percentage of senescent cells characterized by positive β-galactosidase staining, and it exhibited increased proliferation and a higher proliferation index. Immunocytochemical stainings further detected a more primitive gene expression profile for the msx1 group, while osteogenic differentiation assays and ectopic bone formation assays demonstrated an improved capacity for the msx1 group to undergo osteogenic differentiation. These results suggest that transient expression of msx1 in MuSCs can retain a primitive state, thereby enhancing their capacity for osteogenic differentiation and restoring the potential for MuSCs to serve as seed cells for bone tissue engineering.

  5. Inhibition of IKK/NF-κB Signaling Enhances Differentiation of Mesenchymal Stromal Cells from Human Embryonic Stem Cells.

    Science.gov (United States)

    Deng, Peng; Zhou, Chenchen; Alvarez, Ruth; Hong, Christine; Wang, Cun-Yu

    2016-04-12

    Embryonic stem cell-derived mesenchymal stromal cells (MSCs; also known as mesenchymal stem cells) represent a promising source for bone regenerative medicine. Despite remarkable advances in stem cell biology, the molecular mechanism regulating differentiation of human embryonic stem cells (hESCs) into MSCs remains poorly understood. Here, we report that inhibition of IκB kinase (IKK)/nuclear factor kappa B (NF-κB) signaling enhances differentiation of hESCs into MSCs by expediting the loss of pluripotent markers and increasing the expression of MSC surface markers. In addition, a significantly higher quantity of MSCs was produced from hESCs with IKK/NF-κB suppression. These isolated MSCs displayed evident multipotency with capacity to terminally differentiate into osteoblasts, chondrocytes, and adipocytes in vitro and to form bone in vivo. Collectively, our data provide important insights into the role of NF-κB in mesenchymal lineage specification during hESC differentiation, suggesting that IKK inhibitors could be utilized as an adjuvant in generating MSCs for cell-mediated therapies. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  6. High-Throughput Screening Enhances Kidney Organoid Differentiation from Human Pluripotent Stem Cells and Enables Automated Multidimensional Phenotyping.

    Science.gov (United States)

    Czerniecki, Stefan M; Cruz, Nelly M; Harder, Jennifer L; Menon, Rajasree; Annis, James; Otto, Edgar A; Gulieva, Ramila E; Islas, Laura V; Kim, Yong Kyun; Tran, Linh M; Martins, Timothy J; Pippin, Jeffrey W; Fu, Hongxia; Kretzler, Matthias; Shankland, Stuart J; Himmelfarb, Jonathan; Moon, Randall T; Paragas, Neal; Freedman, Benjamin S

    2018-05-15

    Organoids derived from human pluripotent stem cells are a potentially powerful tool for high-throughput screening (HTS), but the complexity of organoid cultures poses a significant challenge for miniaturization and automation. Here, we present a fully automated, HTS-compatible platform for enhanced differentiation and phenotyping of human kidney organoids. The entire 21-day protocol, from plating to differentiation to analysis, can be performed automatically by liquid-handling robots, or alternatively by manual pipetting. High-content imaging analysis reveals both dose-dependent and threshold effects during organoid differentiation. Immunofluorescence and single-cell RNA sequencing identify previously undetected parietal, interstitial, and partially differentiated compartments within organoids and define conditions that greatly expand the vascular endothelium. Chemical modulation of toxicity and disease phenotypes can be quantified for safety and efficacy prediction. Screening in gene-edited organoids in this system reveals an unexpected role for myosin in polycystic kidney disease. Organoids in HTS formats thus establish an attractive platform for multidimensional phenotypic screening. Copyright © 2018 Elsevier Inc. All rights reserved.

  7. Differentiation of prostate cancer from normal prostate tissue in an animal model: conventional MRI and dynamic contrast-enhanced MRI

    International Nuclear Information System (INIS)

    Gemeinhardt, O.; Prochnow, D.; Taupitz, M.; Hamm, B.; Beyersdorff, D.; Luedemann, L.; Abramjuk, C.

    2005-01-01

    Purpose: to differentiate orthotopically implanted prostate cancer from normal prostate tissue using magnetic resonance imaging (MRI) and Gd-DTPA-BMA-enhanced dynamic MRI in the rat model. Material and methods: tumors were induced in 15 rats by orthotopic implantation of G subline Dunning rat prostatic tumor cells. MRI was performed 56 to 60 days after tumor cell implantation using T1-weighted spin-echo, T2-weighted turbo SE sequences, and a 2D FLASH sequence for the contrast medium based dynamic study. The interstitial leakage volume, normalized permeability and the permeability surface area product of tumor and healthy prostate were determined quantitatively using a pharmacokinetic model. The results were confirmed by histologic examination. Results: axial T2-weighted TSE images depicted low-intensity areas suspicious for tumor in all 15 animals. The mean tumor volume was 46.5 mm3. In the dynamic study, the suspicious areas in all animals displayed faster and more pronounced signal enhancement than surrounding prostate tissue. The interstitial volume and the permeability surface area product of the tumors increased significantly by 420% (p<0.001) and 424% (p<0.001), respectively, compared to normal prostate tissue, while no significant difference was seen for normalized permeability alone. Conclusion: the results of the present study demonstrate that quantitative analysis of contrast-enhanced dynamic MRI data enables differentiation of small, slowly growing orthotopic prostate cancer from normal prostate tissue in the rat model. (orig.)

  8. Musicians' Enhanced Neural Differentiation of Speech Sounds Arises Early in Life: Developmental Evidence from Ages 3 to 30

    Science.gov (United States)

    Strait, Dana L.; O'Connell, Samantha; Parbery-Clark, Alexandra; Kraus, Nina

    2014-01-01

    The perception and neural representation of acoustically similar speech sounds underlie language development. Music training hones the perception of minute acoustic differences that distinguish sounds; this training may generalize to speech processing given that adult musicians have enhanced neural differentiation of similar speech syllables compared with nonmusicians. Here, we asked whether this neural advantage in musicians is present early in life by assessing musically trained and untrained children as young as age 3. We assessed auditory brainstem responses to the speech syllables /ba/ and /ga/ as well as auditory and visual cognitive abilities in musicians and nonmusicians across 3 developmental time-points: preschoolers, school-aged children, and adults. Cross-phase analyses objectively measured the degree to which subcortical responses differed to these speech syllables in musicians and nonmusicians for each age group. Results reveal that musicians exhibit enhanced neural differentiation of stop consonants early in life and with as little as a few years of training. Furthermore, the extent of subcortical stop consonant distinction correlates with auditory-specific cognitive abilities (i.e., auditory working memory and attention). Results are interpreted according to a corticofugal framework for auditory learning in which subcortical processing enhancements are engendered by strengthened cognitive control over auditory function in musicians. PMID:23599166

  9. Enhanced adipogenic differentiation of bovine bone marrow-derived mesenchymal stem cells

    Science.gov (United States)

    Until now, the isolation and characterization of bovine bone marrow-derived mesenchymal stem cells (bBM-MSCs) have not been established, which prompted us to optimize the differentiation protocol for bBM-MSCs. In this study, bBM-MSCs were freshly isolated from three 6-month-old cattle and used for p...

  10. Silicon nanowires enhanced proliferation and neuronal differentiation of neural stem cell with vertically surface microenvironment.

    Science.gov (United States)

    Yan, Qiuting; Fang, Lipao; Wei, Jiyu; Xiao, Guipeng; Lv, Meihong; Ma, Quanhong; Liu, Chunfeng; Wang, Wang

    2017-09-01

    Owing to its biocompatibility, noncytotoxicity, biodegradability and three-dimensional structure, vertically silicon nanowires (SiNWs) arrays are a promising scaffold material for tissue engineering, regenerative medicine and relevant medical applications. Recently, its osteogenic differentiation effects, reorganization of cytoskeleton and regulation of the fate on stem cells have been demonstrated. However, it still remains unknown whether SiNWs arrays could affect the proliferation and neuronal differentiation of neural stem cells (NSCs) or not. In the present study, we have employed vertically aligned SiNWs arrays as culture systems for NSCs and proved that the scaffold material could promote the proliferation and neuronal differentiation of NSCs while maintaining excellent cell viability and stemness. Immunofluorescence imaging analysis, Western blot and RT-PCR results reveal that NSCs proliferation and neuronal differentiation efficiency on SiNWs arrays are significant greater than that on silicon wafers. These results implicate SiNWs arrays could offer a powerful platform for NSCs research and NSCs-based therapy in the field of neural tissue engineering.

  11. MiR-34b/c Regulates Wnt1 and Enhances Mesencephalic Dopaminergic Neuron Differentiation

    NARCIS (Netherlands)

    De Gregorio, Roberto; Pulcrano, Salvatore; De Sanctis, Claudia; Volpicelli, Floriana; Guatteo, Ezia; von Oerthel, Lars; Latagliata, Emanuele Claudio; Esposito, Roberta; Piscitelli, Rosa Maria; Perrone-Capano, Carla; Costa, Valerio; Greco, Dario; Puglisi-Allegra, Stefano; Smidt, Marten P.; di Porzio, Umberto; Caiazzo, Massimiliano; Mercuri, Nicola Biagio; Li, Meng; Bellenchi, Gian Carlo

    2018-01-01

    The differentiation of dopaminergic neurons requires concerted action of morphogens and transcription factors acting in a precise and well-defined time window. Very little is known about the potential role of microRNA in these events. By performing a microRNA-mRNA paired microarray screening, we

  12. Differential diagnosis between pulmonary tuberculosis and lung abscess by contrast enhanced CT

    International Nuclear Information System (INIS)

    Kanauchi, Tetsu; Hoshi, Toshiko; Konno, Miyuki; Hando, Yumiko

    2001-01-01

    The contrast enhanced CT findings in 14 patients with active tuberculosis and 26 patients with lung abscess were retrospectively analyzed. Reflecting the difference of pathogenesis between tuberculosis and abscess, the findings are widely different. The findings suggesting pulmonary tuberculosis rather than lung abscess were as follows; multiple and irregular necrotic areas, positive CT angiogram sign, no marginal enhancement surrounding necrosis. Contrast enhanced CT may help to distinguish pulmonary tuberculosis from lung abscess, especially in cases of caseous pneumonia showing broad consolidations or mass-like shadows. (author)

  13. Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain.

    Science.gov (United States)

    Li, Xiaowei; Tzeng, Stephany Y; Liu, Xiaoyan; Tammia, Markus; Cheng, Yu-Hao; Rolfe, Andrew; Sun, Dong; Zhang, Ning; Green, Jordan J; Wen, Xuejun; Mao, Hai-Quan

    2016-04-01

    Strategies to enhance survival and direct the differentiation of stem cells in vivo following transplantation in tissue repair site are critical to realizing the potential of stem cell-based therapies. Here we demonstrated an effective approach to promote neuronal differentiation and maturation of human fetal tissue-derived neural stem cells (hNSCs) in a brain lesion site of a rat traumatic brain injury model using biodegradable nanoparticle-mediated transfection method to deliver key transcriptional factor neurogenin-2 to hNSCs when transplanted with a tailored hyaluronic acid (HA) hydrogel, generating larger number of more mature neurons engrafted to the host brain tissue than non-transfected cells. The nanoparticle-mediated transcription activation method together with an HA hydrogel delivery matrix provides a translatable approach for stem cell-based regenerative therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  14. Buckled structures and 5-azacytidine enhance cardiogenic differentiation of adipose-derived stem cells.

    Science.gov (United States)

    Ravichandran, Rajeswari; Venugopal, Jayarama Reddy; Mueller, Martina; Sundarrajan, Subramanian; Mukherjee, Shayanti; Pliska, Damian; Wintermantel, Erich; Ramakrishna, Seeram

    2013-12-01

    Myocardial infarction is caused after impairment of heart wall muscle following an immense cell loss and also when the myocardial tissue is lacking the inherent capacity to regenerate for normal functioning of myocardium. An immediate challenge in cardiac regeneration is to devise a strategy that leads to a reproducible degree of cardiac differentiation. We have speculated that ex vivo pretreatment of adipose-derived stem cells (ADSCs) using 5-azacytidine and a suitable patterned nanofibrous construct could lead to cardiomyogenic differentiation and results in superior biological and functional effects on cardiac regeneration of infarcted myocardium. Polyglycerol sebacate/gelatin fibers were fabricated by core/shell electrospinning with polyglycerol sebacate as the core material and gelatin as the shell material. Patterning of the core/shell fibers to form orthogonal and looped buckled nanostructures was achieved. Results demonstrated that the buckled fibers showing an orthogonal orientation and looped pattern had a Young's modulus of approximately 3.59 ± 1.58 MPa and 2.07 ± 0.44 MPa, respectively, which was comparable to that of native myocardium. The ADSCs cultured on these scaffolds demonstrated greater expression of the cardiac-specific marker proteins actinin, troponin and connexin 43, as well as characteristic multinucleation as shown by immunocytochemical and morphological analysis, indicating complete cardiogenic differentiation of ADSCs. In the natural milieu, cardiomyogenic differentiation probably involves multiple signaling pathways and we have postulated that a buckled structure combination of chemical treatment and environment-driven strategy induces cardiogenic differentiation of ADSCs. The combination of patterned buckled fibrous structures with stem cell biology may prove to be a productive device for myocardial infarction.

  15. Differentiation of intrahepatic mass-forming cholangiocarcinoma from hepatocellular carcinoma on gadoxetic acid-enhanced liver MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Rihyeon; Shin, Cheong-Il; Yoon, Jeong Hee; Joo, Ijin; Kim, Seong Ho; Hwang, Inpyeong [Seoul National University Hospital, Department of Radiology, Seoul (Korea, Republic of); Lee, Jeong Min; Han, Joon Koo [Seoul National University Hospital, Department of Radiology, Seoul (Korea, Republic of); Seoul National University Hospital, Institute of Radiation Medicine, Seoul (Korea, Republic of); Lee, Eun Sun; Choi, Byung Ihn [Chung-Ang University Hospital, Department of Radiology, Seoul (Korea, Republic of)

    2016-06-15

    To determine the different imaging features of intrahepatic mass-forming cholangiocarcinoma (IMCC) from hepatocellular carcinoma (HCC) on gadoxetic acid-enhanced magnetic resonance imaging (MRI). This retrospective study was institutional review board approved and the requirement for informed consent was waived. Patients who underwent gadoxetic acid-enhanced MRI with histologically confirmed IMCCs (n = 46) or HCCs (n = 58) were included. Imaging features of IMCCs and HCCs on gadoxetic acid-enhanced MRI including T2- and T1-weighted, diffusion weighted images, dynamic study and hepatobiliary phase (HBP) images were analyzed. Univariate and multivariate logistic regression analyses were performed to identify relevant differentiating features between IMCCs and HCCs. Multivariate analysis revealed heterogeneous T2 signal intensity and a hypointense rim on the HBP as suggestive findings of IMCCs and the wash-in and ''portal wash-out'' enhancement pattern as well as focal T1 high signal intensity foci as indicative of HCCs (all, p < 0.05). When we combined any three of the above four imaging features, we were able to diagnose IMCCs with 94 % (43/46) sensitivity and 86 % (50/58) specificity. Combined interpretation of enhancement characteristics including HBP images, morphologic features, and strict application of the ''portal wash-out'' pattern helped more accurate discrimination of IMCCs from HCCs. (orig.)

  16. Differential tissue expression of enhanced green fluorescent protein in ‘Green mice’

    OpenAIRE

    Ma, De-Fu; Tezuka, Hideo; Kondo, Tetsuo; Sudo, Katsuko; Niu, Dong-Feng; Nakazawa, Tadao; Kawasaki, Tomonori; Yamane, Tetsu; Nakamura, Nobuki; Katoh, Ryohei

    2010-01-01

    In order to clarify tissue expression of enhanced green fluorescent protein (EGFP) in ‘green mice’ from a transgenic line having an EGFP cDNA under the control of a chicken beta-actin promoter and cytomegalovirus enhancer, we studied the expression of EGFP in various organs and tissues from these ‘green mice’ by immunohistochemistry with anti- EGFP antibody in conjunction with direct observation for EGFP fluorescence using confocal laser scanning microscopy. On i...

  17. Retinoic acid-loaded polymeric nanoparticles enhance vascular regulation of neural stem cell survival and differentiation after ischaemia

    Science.gov (United States)

    Ferreira, R.; Fonseca, M. C.; Santos, T.; Sargento-Freitas, J.; Tjeng, R.; Paiva, F.; Castelo-Branco, M.; Ferreira, L. S.; Bernardino, L.

    2016-04-01

    Stroke is one of the leading causes of death and disability worldwide. However, current therapies only reach a small percentage of patients and may cause serious side effects. We propose the therapeutic use of retinoic acid-loaded nanoparticles (RA-NP) to safely and efficiently repair the ischaemic brain by creating a favourable pro-angiogenic environment that enhances neurogenesis and neuronal restitution. Our data showed that RA-NP enhanced endothelial cell proliferation and tubule network formation and protected against ischaemia-induced death. To evaluate the effect of RA-NP on vascular regulation of neural stem cell (NSC) survival and differentiation, endothelial cell-conditioned media (EC-CM) were collected. EC-CM from healthy RA-NP-treated cells reduced NSC death and promoted proliferation while EC-CM from ischaemic RA-NP-treated cells decreased cell death, increased proliferation and neuronal differentiation. In parallel, human endothelial progenitor cells (hEPC), which are part of the endogenous repair response to vascular injury, were collected from ischaemic stroke patients. hEPC treated with RA-NP had significantly higher proliferation, which further highlights the therapeutic potential of this formulation. To conclude, RA-NP protected endothelial cells from ischaemic death and stimulated the release of pro-survival, proliferation-stimulating factors and differentiation cues for NSC. RA-NP were shown to be up to 83-fold more efficient than free RA and to enhance hEPC proliferation. These data serve as a stepping stone to use RA-NP as vasculotrophic and neurogenic agents for vascular disorders and neurodegenerative diseases with compromised vasculature.

  18. Adiponectin enhances osteogenic differentiation in human adipose-derived stem cells by activating the APPL1-AMPK signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Tong; Wu, Yu-wei; Lu, Hui; Guo, Yuan [Second Dental Center, Peking University School and Hospital of Stomatology, Beijing (China); National Engineering Laboratory for Digital and Material Technology of Stomatology, Peking University School and Hospital of Stomatology, Beijing (China); Tang, Zhi-hui, E-mail: tang_zhihui@live.cn [Second Dental Center, Peking University School and Hospital of Stomatology, Beijing (China); National Engineering Laboratory for Digital and Material Technology of Stomatology, Peking University School and Hospital of Stomatology, Beijing (China)

    2015-05-29

    Human adipose-derived stem cells (hASCs) are multipotent progenitor cells with multi-lineage differentiation potential including osteogenesis and adipogenesis. While significant progress has been made in understanding the transcriptional control of hASC fate, little is known about how hASC differentiation is regulated by the autocrine loop. The most abundant adipocytokine secreted by adipocytes, adiponectin (APN) plays a pivotal role in glucose metabolism and energy homeostasis. Growing evidence suggests a positive association between APN and bone formation yet little is known regarding the direct effects of APN on hASC osteogenesis. Therefore, this study was designed to investigate the varied osteogenic effects and regulatory mechanisms of APN in the osteogenic commitment of hASCs. We found that APN enhanced the expression of osteoblast-related genes in hASCs, such as osteocalcin, alkaline phosphatase, and runt-related transcription factor-2 (Runx2, also known as CBFa1), in a dose- and time-dependent manner. This was further confirmed by the higher expression levels of alkaline phosphatase and increased formation of mineralization nodules, along with the absence of inhibition of cell proliferation. Importantly, APN at 1 μg/ml was the optimal concentration, resulting in maximum deposition of calcium nodules, and was significant superior to bone morphogenetic protein 2. Mechanistically, we found for the first time that APN increased nuclear translocation of the leucine zipper motif (APPL)-1 as well as AMP-activated protein kinase (AMPK) phosphorylation, which were reversed by pretreatment with APPL1 siRNA. Our results indicate that APN promotes the osteogenic differentiation of hASCs by activating APPL1-AMPK signaling, suggesting that manipulation of APN is a novel therapeutic target for controlling hASC fate. - Highlights: • Adiponectin enhances osteogenic differentiation in human adipose-derived stem cells. • The knock-down of APPL1 block the enhancement of

  19. Approximate Solutions of Delay Differential Equations with Constant and Variable Coefficients by the Enhanced Multistage Homotopy Perturbation Method

    Directory of Open Access Journals (Sweden)

    D. Olvera

    2015-01-01

    Full Text Available We expand the application of the enhanced multistage homotopy perturbation method (EMHPM to solve delay differential equations (DDEs with constant and variable coefficients. This EMHPM is based on a sequence of subintervals that provide approximate solutions that require less CPU time than those computed from the dde23 MATLAB numerical integration algorithm solutions. To address the accuracy of our proposed approach, we examine the solutions of several DDEs having constant and variable coefficients, finding predictions with a good match relative to the corresponding numerical integration solutions.

  20. Andrographolide Promotes Neural Differentiation of Rat Adipose Tissue-Derived Stromal Cells through Wnt/β-Catenin Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Yan Liang

    2017-01-01

    Full Text Available Adipose tissue-derived stromal cells (ADSCs are a high-yield source of pluripotent stem cells for use in cell-based therapies. We explored the effect of andrographolide (ANDRO, one of the ingredients of the medicinal herb extract on the neural differentiation of rat ADSCs and associated molecular mechanisms. We observed that rat ADSCs were small and spindle-shaped and expressed multiple stem cell markers including nestin. They were multipotent as evidenced by adipogenic, osteogenic, chondrogenic, and neural differentiation under appropriate conditions. The proportion of cells exhibiting neural-like morphology was higher, and neurites developed faster in the ANDRO group than in the control group in the same neural differentiation medium. Expression levels of the neural lineage markers MAP2, tau, GFAP, and β-tubulin III were higher in the ANDRO group. ANDRO induced a concentration-dependent increase in Wnt/β-catenin signaling as evidenced by the enhanced expression of nuclear β-catenin and the inhibited form of GSK-3β (pSer9. Thus, this study shows for the first time how by enhancing the neural differentiation of ADSCs we expect that ANDRO pretreatment may increase the efficacy of adult stem cell transplantation in nervous system diseases, but more exploration is needed.

  1. Performance Enhancement of Optical CDMA by Differential-Phase Method for Radio-over-Fiber Transmissions

    Directory of Open Access Journals (Sweden)

    Hsu-Chih Cheng

    2013-01-01

    Full Text Available The study proposes the differential-phase optical code-division multiple-access (OCDMA network for radio-over-fiber (RoF transmissions, and the characteristics are numerically analyzed. The network coder/decoders (codecs are structured on the basis of arrayed-waveguide-grating (AWG routers with complementary Walsh-Hadamard (CWH signature codes. In the proposed system, the network requires only two AWG routers to accomplish spectral encoding of radio base station (RBS and decoding of control station for the complementary keying, thus resulting in a simpler and low cost system. Performance analyses are evaluated with the dominant noise of phase-induced intensity noise (PIIN in spectral code OCDMA network. By the proposed AWG-based OCDMA with the differential-phase scheme, it is possible to establish interference-free and low crosstalk beat noise RoF systems.

  2. Retinoic acid and cAMP inhibit rat hepatocellular carcinoma cell proliferation and enhance cell differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Ionta, M. [Instituto de Ciências Biomédicas, Universidade Federal de Alfenas, Alfenas MG (Brazil); Departamento de Biologia Celular e do Desenvolvimento, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil); Rosa, M.C.; Almeida, R.B.; Freitas, V.M.; Rezende-Teixeira, P.; Machado-Santelli, G.M. [Departamento de Biologia Celular e do Desenvolvimento, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo SP (Brazil)

    2012-05-25

    Hepatocellular carcinoma (HCC) is the third highest cause of cancer death worldwide. In general, the disease is diagnosed at an advanced stage when potentially curative therapies are no longer feasible. For this reason, it is very important to develop new therapeutic approaches. Retinoic acid (RA) is a natural derivative of vitamin A that regulates important biological processes including cell proliferation and differentiation. In vitro studies have shown that RA is effective in inhibiting growth of HCC cells; however, responsiveness to treatment varies among different HCC cell lines. The objective of the present study was to determine if the combined use of RA (0.1 µM) and cAMP (1 mM), an important second messenger, improves the responsiveness of HCC cells to RA treatment. We evaluated the proliferative behavior of an HCC cell line (HTC) and the expression profile of genes related to cancer signaling pathway (ERK and GSK-3β) and liver differentiation [E-cadherin, connexin 26 (Cx26), and connexin 32 (Cx32)]. RA and cAMP were effective in inhibiting the proliferation of HTC cells independently of combined use. However, when a mixture of RA and cAMP was used, the signals concerning the degree of cell differentiation were increased. As demonstrated by Western blot, the treatment increased E-cadherin, Cx26, Cx32 and Ser9-GSK-3β (inactive form) expression while the expression of Cx43, Tyr216-GSK-3β (active form) and phosphorylated ERK decreased. Furthermore, telomerase activity was inhibited along treatment. Taken together, the results showed that the combined use of RA and cAMP is more effective in inducing differentiation of HTC cells.

  3. Enhancement of osteoblastic differentiation in alginate gel beads with bioactive octacalcium phosphate particles

    International Nuclear Information System (INIS)

    Endo, Kosei; Anada, Takahisa; Suzuki, Osamu; Yamada, Masumi; Seki, Minoru; Sasaki, Keiichi

    2015-01-01

    The present study investigated whether alginate (Alg) hydrogel microbeads have a role in maintaining mouse bone marrow stromal ST-2 cells and release the cells after being stimulated by synthetic octacalcium phosphate (OCP), which is a mineral crystal capable of stimulating osteoblastic differentiation during a conversion process to hydroxyapatite (HA). The ST-2 cell suspension in the alginate solution, which contained various concentrations of OCP granules with diameters less than 53 μm, was extruded drop-wise into a stirred gelation solution containing BaCl 2 using an encapsulator with nitrogen gas stream. The Alg-microbeads (Alg/OCP · ST-2 microbeads) that were generated, which had a diameter of approximately 400 μm, were incubated for up to 14 d and then assessed for osteoblastic differentiation. Alg-microbeads with cells were also incubated to identify the possible conversion from OCP to HA. Osteoblast differentiation markers in ST-2 cells, alkaline phosphatase (ALP) and collagen type I, were up-regulated in the presence of higher amounts of OCP. X-ray diffraction analysis and Fourier transform infrared spectroscopy confirmed that the OCP tended to convert to HA over time, suggesting that the OCP in Alg-microbeads interacts three-dimensionally with ST-2 cells and stimulates its osteoblastic differentiation. The release of ST-2 cells from the microbeads was also estimated. ST-2 cells were identified outside of the microbeads, although the cell number tended to decrease with increasing OCP. These results suggest that Alg/OCP microbeads could be used as a vehicle to activate osteoblastic cells and deliver them to sites where bone regeneration is needed. (paper)

  4. Micro/Nano Structural Tantalum Coating for Enhanced Osteogenic Differentiation of Human Bone Marrow Stem Cells

    OpenAIRE

    Ding Ding; Youtao Xie; Kai Li; Liping Huang; Xuebin Zheng

    2018-01-01

    Recently, tantalum has been attracting much attention for its anticorrosion resistance and biocompatibility, and it has been widely used in surface modification for implant applications. To improve its osteogenic differentiation of human bone marrow stem cells (hBMSCs), a micro/nano structure has been fabricated on the tantalum coating surface through the combination of anodic oxidation and plasma spraying method. The morphology, composition, and microstructure of the modified coating were co...

  5. Evaluation of dynamic contrast-enhanced T1-weighted perfusion MRI in the differentiation of tumor recurrence from radiation necrosis

    DEFF Research Database (Denmark)

    Larsen, Anne Vibeke Andrée; Simonsen, Helle J; Law, Ian

    2013-01-01

    INTRODUCTION: To investigate if perfusion measured with dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) can be used to differentiate radiation necrosis from tumor recurrence in patients with high-grade glioma. METHODS: The study was approved by the institutional review board...... to measure cerebral blood volume (CBV), blood-brain barrier (BBB) permeability and cerebral blood flow (CBF). Subjects also underwent FDG-PET and lesions were classified as either metabolically active or inactive. Follow-up clinical MRI and lesion histology in case of additional tissue resection was used...... to determine whether lesions were regressing or progressing. RESULTS: Fourteen enhancing lesions could be classified as progressing (11) or regressing (three). An empirical threshold of 2.0 ml/100 g for CBV allowed detection of regressing lesions with a sensitivity of 100 % and specificity of 100 %. FDG-PET...

  6. Osteoblast-secreted collagen upregulates paracrine Sonic hedgehog signaling by prostate cancer cells and enhances osteoblast differentiation

    Directory of Open Access Journals (Sweden)

    Zunich Samantha M

    2012-07-01

    Full Text Available Abstract Background Induction of osteoblast differentiation by paracrine Sonic hedgehog (Shh signaling may be a mechanism through which Shh-expressing prostate cancer cells initiate changes in the bone microenvironment and promote metastases. A hallmark of osteoblast differentiation is the formation of matrix whose predominant protein is type 1 collagen. We investigated the formation of a collagen matrix by osteoblasts cultured with prostate cancer cells, and its effects on interactions between prostate cancer cells and osteoblasts. Results In the presence of exogenous ascorbic acid (AA, a co-factor in collagen synthesis, mouse MC3T3 pre-osteoblasts in mixed cultures with human LNCaP prostate cancer cells or LNCaP cells modified to overexpress Shh (LNShh cells formed collagen matrix with distinct fibril ultrastructural characteristics. AA increased the activity of alkaline phosphatase and the expression of the alkaline phosphatase gene Akp2, markers of osteoblast differentiation, in MC3T3 pre-osteoblasts cultured with LNCaP or LNShh cells. However, the AA-stimulated increase in Akp2 expression in MC3T3 pre-osteoblasts cultured with LNShh cells far exceeded the levels observed in MC3T3 cells cultured with either LNCaP cells with AA or LNShh cells without AA. Therefore, AA and Shh exert a synergistic effect on osteoblast differentiation. We determined whether the effect of AA on LNShh cell-induced osteoblast differentiation was mediated by Shh signaling. AA increased the expression of Gli1 and Ptc1, target genes of the Shh pathway, in MC3T3 pre-osteoblasts cultured with LNShh cells to at least twice their levels without AA. The ability of AA to upregulate Shh signaling and enhance alkaline phosphatase activity was blocked in MC3T3 cells that expressed a dominant negative form of the transcription factor GLI1. The AA-stimulated increase in Shh signaling and Shh-induced osteoblast differentiation was also inhibited by the specific collagen synthesis

  7. Clinical application of proton magnetic resonance spectroscopy in differential diagnosis of intracranial lesions with ring-like enhancement

    International Nuclear Information System (INIS)

    Lai Ying; Cheng Kailiang; Zhang Mengchao; Liu Yunxia; Wang Wei

    2009-01-01

    Objective: To study the value of clinical application of 1 H proton magnetic resonance spectroscopy ( 1 H MRS) in the differential diagnosis of intracranial lesions with ring-like enhancement. Methods: 28 cases were diagnosed of intracranial lesions with ring-like enhancement by clinical examination and pathologic test. A total of 28 ratios cases included 6 cases high grade glioma, 10 cases of metastatic carcinoma (n=10) and 12 cases of brain abscess, after examined with 1HMRS, the ratios of various metabolites in focal center, enhancement ring,perifocal edema region and normal control group were detected and compared. Results: The ratios of NAA/Cho, Cho/Cr and NAA/Cr in focal center had no significantly differences between high grade glioma and metastatic carcinoma (P>0.05). The peak of NAA was significantly different between high grade glioma and metastatic carcinoma (P 0.05). The peak of AA was characteristic of brain abscess. The ratio of Cho/Cr 0 in brain abscess was significantly lower than those in high grade glioma and metastatic carcinoma (P 0 denoted the Cho content of contralateral normal brain region). These results accorded with the result of pathological examination. Conclusion: 1 HMRS can improve the diagnostic accuracy of intracranial lesions with ring-like enhancement. (authors)

  8. High molecular weight hyaluronic acid increases the differentiation potential of the murine chondrocytic ATDC5 cell line.

    Science.gov (United States)

    Sato, Eiichi; Ando, Takashi; Ichikawa, Jiro; Okita, Genki; Sato, Nobutaka; Wako, Masanori; Ohba, Tetsuro; Ochiai, Satoshi; Hagino, Tetsuo; Jacobson, Richard; Haro, Hirotaka

    2014-12-01

    Osteoarthritis (OA) is a group of common, chronic, and painful inflammatory joint diseases. One important finding in OA patients is a remarkable decrease in the molecular weight of hyaluronic acid (HA) in the synovial fluid of affected joints. Therapeutic HA is available to patients in most parts of the world as a viscosupplementation product for the treatment of OA. Previous clinical reports show that high molecular weight HA (HMWHA) more effectively relieves pain than low molecular weight HA (LMWHA). However, the mechanism behind this finding remains unclear. In this study, we investigated whether a LMWHA (Low-0.9 MDa) and two types of HMWHA (High-1.9 MDa and 6 MDa) differentially affected chondroregulatory action. We tested this using ATDC5 cell, a murine chondrocytic cell line widely used in culture systems to study chondrogenic differentiation. We found that HMWHA, especially hylan G-F 20 (High-6 MDa), significantly induced aggrecan and proteoglycan accumulation, nodule formation, and mRNA expression of chondrogenic differentiation markers in a time- and dose-dependent manner. In addition, we showed that HMWHA prevented TNF-α induced inhibition of chondrogenic differentiation, with no effect on cell proliferation or viability. These results reveal that HMWHA significantly promotes chondrogenic differentiation of ATDC5 cells in vitro, and suggest that HMWHA plays a significant chondroregulatory role in vivo. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  9. Hydroxylation of methylated DNA by TET1 in chondrocyte differentiation of C3H10T1/2 cells

    Directory of Open Access Journals (Sweden)

    Ryo Ito

    2016-03-01

    Full Text Available DNA methylation is closely involved in the regulation of cellular differentiation, including chondrogenic differentiation of mesenchymal stem cells. Recent studies showed that Ten–eleven translocation (TET family proteins converted 5-methylcytosine (5mC to 5-hydroxymethylcytosine, 5-formylcytosine and 5carboxylcytosine by oxidation. These reactions constitute potential mechanisms for active demethylation of methylated DNA. However, the relationship between the DNA methylation patterns and the effects of TET family proteins in chondrocyte differentiation is still unclear. In this study, we showed that DNA hydroxylation of 5mC was increased during chondrocytic differentiation of C3H10T1/2 cells and that the expression of Tet1 was particularly enhanced. Moreover, knockdown experiments revealed that the downregulation of Tet1 expression caused decreases in chondrogenesis markers such as type 2 and type 10 collagens. Furthermore, we found that TET proteins had a site preference for hydroxylation of 5mC on the Insulin-like growth factor 1 (Igf1 promoter in chondrocytes. Taken together, we showed that the expression of Tet1 was specifically facilitated in chondrocyte differentiation and Tet1 can regulate chondrocyte marker gene expression presumably through its hydroxylation activity for DNA.

  10. Differentiation between tuberculosis and leukemia in abdominal and pelvic lymph nodes: evaluation with contrast-enhanced multidetector computed tomography

    Directory of Open Access Journals (Sweden)

    Ge Zhang

    2015-03-01

    Full Text Available PURPOSE: To compare the characteristics of tubercular vs. leukemic involvement of abdominopelvic lymph nodes using multidetector computed tomography (CT. MATERIALS AND METHODS: We retrospectively reviewed multidetector computed tomography features including lymph node size, shape, enhancement patterns, and anatomical distribution, in 106 consecutive patients with newly diagnosed, untreated tuberculosis (55 patients; 52% or leukemia (51 patients; 48%. In patients with leukemia, 32 (62.7% had chronic lymphocytic leukemia, and 19 (37.3% had acute leukemias; of these, 10 (19.6% had acute myeloid leukemia, and 9 (17.6% had acute lymphocytic leukemia. RESULTS: The lower para-aortic (30.9% for tuberculosis, 63.2% for acute leukemias and 87.5% for chronic lymphocytic leukemia and inguinal (9.1% for tuberculosis, 57.9% for acute leukemias and 53.1% for chronic lymphocytic leukemia lymph nodes were involved more frequently in the three types of leukemia than in tuberculosis (both with p <0.017. Tuberculosis showed peripheral enhancement, frequently with a multilocular appearance, in 43 (78.2% patients, whereas patients with leukemia (78.9% for acute myeloid leukemia and acute lymphocytic leukemia, 87.5% for chronic lymphocytic leukemia demonstrated predominantly homogeneous enhancement (both with p <0.017. For the diagnosis of tuberculosis, the analysis showed that a peripheral enhancement pattern had a sensitivity of 78.2%, a specificity of 100%, and an accuracy of 88.7%. For the diagnosis of leukemia, the analysis showed that a homogeneous enhancement pattern was associated with a sensitivity of 84.3%, a specificity of 94.5%, and an accuracy of 89.6%. CONCLUSION: Our findings indicate that the anatomical distribution and enhancement patterns of lymphadenopathy seen on multidetector computed tomography are useful for differentiating between untreated tuberculosis and leukemia of the abdominopelvic lymph nodes.

  11. Serum-Induced Differentiation of Glioblastoma Neurospheres Leads to Enhanced Migration/Invasion Capacity That Is Associated with Increased MMP9.

    Directory of Open Access Journals (Sweden)

    Justin V Joseph

    Full Text Available Glioblastoma (GBM is a highly infiltrative brain tumor in which cells with properties of stem cells, called glioblastoma stem cells (GSCs, have been identified. In general, the dominant view is that GSCs are responsible for the initiation, progression, invasion and recurrence of this tumor. In this study, we addressed the question whether the differentiation status of GBM cells is associated with their invasive capacity. For this, several primary GBM cell lines were used, cultured either as neurospheres known to enrich for GSCs or in medium supplemented with 10% FCS that promotes differentiation. The differentiation state of the cells was confirmed by determining the expression of stem cell and differentiation markers. The migration/invasion potential of these cells was tested using in vitro assays and intracranial mouse models. Interestingly, we found that serum-induced differentiation enhanced the invasive potential of GBM cells, which was associated with enhanced MMP9 expression. Chemical inhibition of MMP9 significantly reduced the invasive potential of differentiated cells in vitro. Furthermore, the serum-differentiated cells could revert back to an undifferentiated/stem cell state that were able to form neurospheres, although with a reduced efficiency as compared to non-differentiated counterparts. We propose a model in which activation of the differentiation program in GBM cells enhances their infiltrative potential and that depending on microenvironmental cues a significant portion of these cells are able to revert back to an undifferentiated state with enhanced tumorigenic potential. Thus, effective therapy should target both GSCs and differentiated offspring and targeting of differentiation-associated pathways may offer therapeutic opportunities to reduce invasive growth of GBM.

  12. Glucose enhancement of recognition memory: differential effects on effortful processing but not aspects of 'remember-know' responses.

    Science.gov (United States)

    Scholey, Andrew; Macpherson, Helen; Sünram-Lea, Sandra; Elliott, Jade; Stough, Con; Kennedy, David

    2013-01-01

    The administration of a glucose drink has been shown to enhance cognitive performance with effect sizes comparable with those from pharmaceutical interventions in human trials. In the memory domain, it is currently debated whether glucose facilitation of performance is due to differential targeting of hippocampal memory or whether task effort is a more important determinant. Using a placebo-controlled, double-blind, crossover 2(Drink: glucose/placebo) × 2(Effort: ± secondary task) design, 20 healthy young adults' recognition memory performance was measured using the 'remember-know' procedure. Two high effort conditions (one for each drink) included secondary hand movements during word presentation. A 25 g glucose or 30 mg saccharine (placebo) drink was consumed 10 min prior to the task. The presence of a secondary task resulted in a global impairment of memory function. There were significant Drink × Effort interactions for overall memory accuracy but no differential effects for 'remember' or 'know' responses. These data suggest that, in some circumstances, task effort may be a more important determinant of the glucose facilitation of memory effect than hippocampal mediation. This article is part of a Special Issue entitled 'Cognitive Enhancers'. Copyright © 2012 Elsevier Ltd. All rights reserved.

  13. Dioxinodehydroeckol Enhances the Differentiation of Osteoblasts by Regulating the Expression of Phospho-Smad1/5/8

    Directory of Open Access Journals (Sweden)

    Byul-Nim Ahn

    2016-09-01

    Full Text Available Lack of bone formation-related health problems are a major problem for the aging population in the modern world. As a part of the ongoing trend of developing natural substances that attenuate osteoporotic bone loss conditions, dioxinodehydroeckol (DHE from edible brown alga Ecklonia cava was tested for its effects on osteoblastogenic differentiation in MC3T3-E1 pre-osteoblasts. DHE was observed to successfully enhance osteoblast differentiation, as indicated by elevated cell proliferation, alkaline phosphatase activity, intracellular cell mineralization, along with raised levels of osteoblastogenesis indicators at the concentration of 20 μM. Results suggested a possible intervening of DHE on the bone morphogenetic protein (BMP signaling pathway, according to elevated protein levels of BMP-2, collagen-I, and Smads. In addition, the presence of DHE was also able to raise the phosphorylated extracellular signal–regulated kinase (ERK and c-Jun N-terminal kinase (JNK levels which are also activated by the BMP signaling pathway. In conclusion, DHE is suggested to be a potential bioactive compound against bone loss that could enhance osteoblastogenesis with a suggested BMP pathway interaction.

  14. Intermittent, low dose carbon monoxide exposure enhances survival and dopaminergic differentiation of human neural stem cells

    DEFF Research Database (Denmark)

    Dreyer-Andersen, Nanna; Almeida, Ana Sofia; Jensen, Pia

    2018-01-01

    Exploratory studies using human fetal tissue have suggested that intrastriatal transplantation of dopaminergic neurons may become a future treatment for patients with Parkinson's disease. However, the use of human fetal tissue is compromised by ethical, regulatory and practical concerns. Human stem...... cells constitute an alternative source of cells for transplantation in Parkinson's disease, but efficient protocols for controlled dopaminergic differentiation need to be developed. Short-term, low-level carbon monoxide (CO) exposure has been shown to affect signaling in several tissues, resulting...... in Parkinson's disease....

  15. Airline seat inventory control benefiting from currency differentials to enhance revenues

    Directory of Open Access Journals (Sweden)

    C.E. Love

    2003-12-01

    Full Text Available The purpose of this paper is to develop an airline seat inventory control model which will capitalise on currency differentials that exist between city pairs. The approach taken here is to maximise Expected Marginal Seat Revenues as proposed by Belobaba for non-nested fare classes. the basic Expected Marginal Seat Revenue model is extended to explicitly include the effects of overbooking. Data from the South African Airways return flight between Cape Town and London is utilised to demonstrate the model.

  16. Electroporation-mediated transfer of SOX trio genes (SOX-5, SOX-6, and SOX-9) to enhance the chondrogenesis of mesenchymal stem cells.

    Science.gov (United States)

    Kim, Hye-Joung; Im, Gun-Il

    2011-12-01

    The purpose of this study was to test the hypothesis that the SOX trio genes (SOX-5, SOX-6, and SOX-9) have a lower level of expression during the chondrogenic differentiation of mesenchymal stem cells (MSCs) compared with chondrocytes and that the electroporation-mediated gene transfer of SOX trio promotes chondrogenesis from human MSCs. An in vitro pellet culture was carried out using MSCs or chondrocytes at passage 3 and analyzed after 7 and 21 days. Then, MSCs were transfected with SOX trio genes and analyzed for the expression of chondrogenic markers after 21 days of in vitro culture. Without transforming growth factor-β1, the untransfected MSCs had a lower level of SOX trio gene and protein expression than chondrocytes. However, the level of SOX-9 gene expression increased in MSCs when treated with transforming growth factor-β1. GAG level significantly increased 7-fold in MSCs co-transfected with SOX trio, which was corroborated by Safranin-O staining. SOX trio co-transfection significantly increased COL2A1 gene and protein and decreased COL10A1 protein in MSCs. It is concluded that the SOX trio have a significantly lower expression in human MSCs than in chondrocytes and that the electroporation-mediated co-transfection of SOX trio enhances chondrogenesis and suppresses hypertrophy of human MSCs.

  17. Differentiation between benign and malignant colon tumors using fast dynamic gadolinium-enhanced MR colonography; a feasibility study

    Energy Technology Data Exchange (ETDEWEB)

    Achiam, M.P., E-mail: achiam1@dadlnet.d [Department of Diagnostic Radiology, Copenhagen University Hospital Herlev, Herlev Ringvej, DK-2730 Herlev (Denmark); Department of Surgical Gastroenterology D, Copenhagen University Hospital Herlev, Herlev Ringvej, DK-2730 Herlev (Denmark); Department of Diagnostic Sciences, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3C, DK-2200 Copenhagen (Denmark); Andersen, L.P.H.; Klein, M. [Department of Surgical Gastroenterology D, Copenhagen University Hospital Herlev, Herlev Ringvej, DK-2730 Herlev (Denmark); Department of Diagnostic Sciences, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3C, DK-2200 Copenhagen (Denmark); Logager, V.; Chabanova, E.; Thomsen, H.S. [Department of Diagnostic Radiology, Copenhagen University Hospital Herlev, Herlev Ringvej, DK-2730 Herlev (Denmark); Department of Diagnostic Sciences, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3C, DK-2200 Copenhagen (Denmark); Rosenberg, J. [Department of Surgical Gastroenterology D, Copenhagen University Hospital Herlev, Herlev Ringvej, DK-2730 Herlev (Denmark); Department of Diagnostic Sciences, Faculty of Health Sciences, University of Copenhagen, Blegdamsvej 3C, DK-2200 Copenhagen (Denmark)

    2010-06-15

    Background: Colorectal cancer will present itself as a bowel obstruction in 16-23% of all cases. However, not all obstructing tumors are malignant and the differentiation between a benign and a malignant tumor can be difficult. The purpose of our study was to determine whether fast dynamic gadolinium-enhanced MR imaging combined with MR colonography could be used to differentiate a benign from a malignant obstructing colon tumor. Methods: Patients with benign colon tumor stenosis, based on diverticulitis, were asked to participate in the study. The same number of patients with verified colorectal cancer was included. Both groups had to be scheduled for surgery to be included. Two blinded observers analyzed the tumors on MR by placing a region of interest in the tumor and a series of parameters were evaluated, e.g. wash-in, wash-out and time-to-peak. Results: 14 patients were included. The wash-in and wash-out rates were significantly different between the benign and malignant tumors, and a clear distinction between benign and malignant disease was therefore possible by looking only at the MR data. Furthermore, MR colography evaluating the rest of the colon past the stenosis was possible with all patients. Conclusion: The results showed the feasibility of using fast dynamic gadolinium-enhanced MR imaging to differentiate between benign and malignant colonic tumors. With a high intra-class correlation and significant differences found on independent segments of the tumor, the method appears to be reproducible. Furthermore, the potential is big in performing a full preoperative colon evaluation even in patients with obstructing cancer. Trial number: (NCT00114829).

  18. Differentiation between benign and malignant colon tumors using fast dynamic gadolinium-enhanced MR colonography; a feasibility study

    International Nuclear Information System (INIS)

    Achiam, M.P.; Andersen, L.P.H.; Klein, M.; Logager, V.; Chabanova, E.; Thomsen, H.S.; Rosenberg, J.

    2010-01-01

    Background: Colorectal cancer will present itself as a bowel obstruction in 16-23% of all cases. However, not all obstructing tumors are malignant and the differentiation between a benign and a malignant tumor can be difficult. The purpose of our study was to determine whether fast dynamic gadolinium-enhanced MR imaging combined with MR colonography could be used to differentiate a benign from a malignant obstructing colon tumor. Methods: Patients with benign colon tumor stenosis, based on diverticulitis, were asked to participate in the study. The same number of patients with verified colorectal cancer was included. Both groups had to be scheduled for surgery to be included. Two blinded observers analyzed the tumors on MR by placing a region of interest in the tumor and a series of parameters were evaluated, e.g. wash-in, wash-out and time-to-peak. Results: 14 patients were included. The wash-in and wash-out rates were significantly different between the benign and malignant tumors, and a clear distinction between benign and malignant disease was therefore possible by looking only at the MR data. Furthermore, MR colography evaluating the rest of the colon past the stenosis was possible with all patients. Conclusion: The results showed the feasibility of using fast dynamic gadolinium-enhanced MR imaging to differentiate between benign and malignant colonic tumors. With a high intra-class correlation and significant differences found on independent segments of the tumor, the method appears to be reproducible. Furthermore, the potential is big in performing a full preoperative colon evaluation even in patients with obstructing cancer. Trial number: (NCT00114829).

  19. Surface modification of TiO{sub 2} nanotubes with osteogenic growth peptide to enhance osteoblast differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Lai, Min, E-mail: minlai@jsnu.edu.cn [School of Life Science, Jiangsu Normal University, Xuzhou, Jiangsu 221116 (China); Jin, Ziyang [School of Life Science, Jiangsu Normal University, Xuzhou, Jiangsu 221116 (China); Su, Zhiguo [Department of Pharmacy, The Affiliated hospital of Qingdao University, Qingdao, Shandong 266555 (China)

    2017-04-01

    To investigate the influence of surface-biofunctionalized substrates on osteoblast behavior, a layer of aligned TiO{sub 2} nanotubes with a diameter of around 70 nm was fabricated on titanium surface by anodization, and then osteogenic growth peptide (OGP) was conjugated onto TiO{sub 2} nanotubes through the intermediate layer of polydopamine. The morphology, composition and wettability of different surfaces were characterized by field-emission scanning electron microscopy (FE-SEM), atomic force microscopy (AFM), X-ray photoelectron spectroscopy (XPS) and contact angle measurements, respectively. The effects of OGP-modified TiO{sub 2} nanotube substrates on the morphology, proliferation and differentiation of osteoblasts were examined in vitro. Immunofluorescence staining revealed that the OGP-functionalized TiO{sub 2} nanotubes were favorable for cell spreading. However, there was no significant difference in cell proliferation observed among the different groups. Cells grown onto OGP-functionalized TiO{sub 2} nanotubes showed significantly higher (p < 0.05 or p < 0.01) levels of alkaline phosphatase (ALP) and mineralization after 4, 7 and 14 days of culture, respectively. Cells grown on OGP-functionalized TiO{sub 2} nanotubes had significantly higher (p < 0.05 or p < 0.01) expression of osteogenic-related genes including runt related transcription factor 2 (Runx2), ALP, collagen type I (Col I), osteopontin (OPN) and osteocalcin (OC) after 14 days of culture. These data suggest that surface functionalization of TiO{sub 2} nanotubes with OGP was beneficial for cell spreading and differentiation. This study provides a novel platform for the development and fabrication of titanium-based implants that enhance the propensity for osseointegration between the native tissue and implant interface. - Highlights: • The OGP functionalized TiO{sub 2} nanotube substrates were successfully fabricated through a direct and effective method. • The OGP functionalized substrates

  20. Enhanced performance for differential detection in coherent Brillouin optical time-domain analysis sensors

    Science.gov (United States)

    Shao, Liyang; Zhang, Yunpeng; Li, Zonglei; Zhang, Zhiyong; Zou, Xihua; Luo, Bin; Pan, Wei; Yan, Lianshan

    2016-11-01

    Logarithmic detectors (LogDs) have been used in coherent Brillouin optical time-domain analysis (BOTDA) sensors to reduce the effect of phase fluctuation, demodulation complexities, and measurement time. However, because of the inherent properties of LogDs, a DC component at the level of hundreds of millivolts that prohibits high-gain signal amplification (SA) could be generated, resulting in unacceptable data acquisition (DAQ) inaccuracies and decoding errors in the process of prototype integration. By generating a reference light at a level similar to the probe light, differential detection can be applied to remove the DC component automatically using a differential amplifier before the DAQ process. Therefore, high-gain SA can be employed to reduce quantization errors. The signal-to-noise ratio of the weak Brillouin gain signal is improved from ˜11.5 to ˜21.8 dB. A BOTDA prototype is implemented based on the proposed scheme. The experimental results show that the measurement accuracy of the Brillouin frequency shift (BFS) is improved from ±1.9 to ±0.8 MHz at the end of a 40-km sensing fiber.

  1. The differential outcomes procedure enhances adherence to treatment: A simulated study with healthy adults

    Directory of Open Access Journals (Sweden)

    Michael eMolina

    2015-11-01

    Full Text Available Memory for medical recommendations is a prerequisite for good adherence to treatment, and therefore to ameliorate the negative effects of the disease, a problem that mainly affects people with memory deficits. We conducted a simulated study to test the utility of a procedure (the differential outcomes procedure, DOP that may improve adherence to treatment by increasing the patient’s learning and retention of medical recommendations regarding medication. The DOP requires the structure of a conditional discriminative learning task in which correct choice responses to specific stimulus-stimulus associations are reinforced with a particular reinforcer or outcome. In two experiments, participants had to learn and retain in their memory the pills that were associated with particular disorders. To assess whether the DOP improved long-term retention of the learned disorder/pill associations, participants were asked to perform two recognition memory tests, 1 hour and 1 week after completing the learning phase. The results showed that compared with the standard non-differential outcomes procedure (NOP, the DOP produced better learning and long-term retention of the previously learned associations. These findings suggest that the DOP can be used as a useful complementary technique in intervention programs targeted at increasing adherence to clinical recommendations.

  2. Laminin and biomimetic extracellular elasticity enhance functional differentiation in mammary epithelia

    Energy Technology Data Exchange (ETDEWEB)

    Alcaraz, Jordi; Xu, Ren; Mori, Hidetoshi; Nelson, Celeste M.; Mroue, Rana; Spencer, Virginia A.; Brownfield, Doug; Radisky, Derek C.; Bustamante, Carlos; Bissell, Mina J.

    2008-10-20

    In the mammary gland, epithelial cells are embedded in a 'soft' environment and become functionally differentiated in culture when exposed to a laminin-rich extracellular matrix gel. Here, we define the processes by which mammary epithelial cells integrate biochemical and mechanical extracellular cues to maintain their differentiated phenotype. We used single cells cultured on top of gels in conditions permissive for {beta}-casein expression using atomic force microscopy to measure the elasticity of the cells and their underlying substrata. We found that maintenance of {beta}-casein expression required both laminin signalling and a 'soft' extracellular matrix, as is the case in normal tissues in vivo, and biomimetic intracellular elasticity, as is the case in primary mammary epithelial organoids. Conversely, two hallmarks of breast cancer development, stiffening of the extracellular matrix and loss of laminin signalling, led to the loss of {beta}-casein expression and non-biomimetic intracellular elasticity. Our data indicate that tissue-specific gene expression is controlled by both the tissues unique biochemical milieu and mechanical properties, processes involved in maintenance of tissue integrity and protection against tumorigenesis.

  3. Study on enhancing mutagenic efficacy by irradiating rice plant before ear differentiation

    International Nuclear Information System (INIS)

    Guo Guangrong; Yi Weiping; Chen Xuebin; Liu; Wuquan; Sun Zhiyuan

    1992-06-01

    Rice plants were irradiated externally with 60 Co γ-ray or internally with 32 P β-beam at the time just before ear differentiation. The mutation frequency, shape of mutated sector, hereditary variation of the mutant of Japonica rice, hereditary pattern of the mutant characters and the application of useful mutants were investigated. The results showed: 1. the irradiation treatment at the time before ear differentiation can greatly raise the mutation frequency. The mutation frequency in creased to 21.67% for early maturity, 11.01% for dwarfness and 12.92% for late maturity respectively (counted on the basis of ear number) under the external irradiation of 60 Co γ-ray. The mutation frequencies for early, late and dwarfness maturities irradiated internally by 32 P β-beam were 15.5%, 61.6% and 26.34% respectively (counted on the basis of grain number), or 46.87%, 91.3% and 65.22% respectively (counted on the basis of ear number). 2. Mutants, that are rarely existed, of Japonica rice were obtained. 3. The segregation of posterity for some mutant characters was not in Mendel's regularity, the relationships between dominance and recessive for some mutant characters were reversed. 4. The prospect for the application of useful mutants in the rice breeding is encouraging

  4. Chondrogenic Effect of Intra-articular Hypertonic-Dextrose (Prolotherapy) in Severe Knee Osteoarthritis.

    Science.gov (United States)

    Topol, Gastón Andrés; Podesta, Leandro Ariel; Reeves, Kenneth Dean; Giraldo, Marcia Mallma; Johnson, Lanny L; Grasso, Raul; Jamín, Alexis; Clark, Tom; Rabago, David

    2016-11-01

    Dextrose injection is reported to improve knee osteoarthritis (KOA)-related clinical outcomes, but its effect on articular cartilage is unknown. A chondrogenic effect of dextrose injection has been proposed. To assess biological and clinical effects of intra-articular hypertonic dextrose injections (prolotherapy) in painful KOA. Case series with blinded arthroscopic evaluation before and after treatment. Physical medicine and day surgery practice. Symptomatic KOA for at least 6 months, arthroscopy-confirmed medial compartment exposed subchondral bone, and temporary pain relief with intra-articular lidocaine injection. Four to 6 monthly 10-mL intra-articular injections with 12.5% dextrose. Visual cartilage growth assessment of 9 standardized medial condyle zones in each of 6 participants by 3 arthroscopy readers masked to pre-/postinjection status (total 54 zones evaluated per reader); biopsy of a cartilage growth area posttreatment, evaluated using hematoxylin and eosin and Safranin-O stains, quantitative polarized light microscopy, and immunohistologic cartilage typing; self-reported knee specific quality of life using the Western Ontario McMaster University Osteoarthritis Index (WOMAC, 0-100 points). Six participants (1 female and 5 male) with median age of 71 years, WOMAC composite score of 57.5 points, and a 9-year pain duration received a median of 6 dextrose injections and follow-up arthroscopy at 7.75 months (range 4.5-9.5 months). In 19 of 54 zone comparisons, all 3 readers agreed that the posttreatment zone showed cartilage growth compared with the pretreatment zone. Biopsy specimens showed metabolically active cartilage with variable cellular organization, fiber parallelism, and cartilage typing patterns consistent with fibro- and hyaline-like cartilage. Compared with baseline status, the median WOMAC score improved 13 points (P = .013). Self-limited soreness after methylene blue instillation was noted. Positive clinical and chondrogenic effects were seen

  5. Ring-Like Enhancement of Hepatocellular Carcinoma in Gadoxetic Acid-Enhanced Multiphasic Hepatic Arterial Phase Imaging With Differential Subsampling With Cartesian Ordering.

    Science.gov (United States)

    Ichikawa, Shintaro; Motosugi, Utaroh; Oishi, Naoki; Shimizu, Tatsuya; Wakayama, Tetsuya; Enomoto, Nobuyuki; Matsuda, Masanori; Onishi, Hiroshi

    2018-04-01

    The aim of this study was to evaluate the efficacy of multiphasic hepatic arterial phase (HAP) imaging using DISCO (differential subsampling with Cartesian ordering) in increasing the confidence of diagnosis of hepatocellular carcinoma (HCC). This retrospective study was approved by the institutional review board, and the requirement for informed patient consent was waived. Consecutive patients (from 2 study periods) with malignant liver nodules were examined by gadoxetic acid-enhanced magnetic resonance imaging using either multiphasic (6 phases; n = 135) or single (n = 230) HAP imaging, which revealed 519 liver nodules other than benign ones (HCC, 497; cholangiocarcinoma, 11; metastases, 10; and malignant lymphoma, 1). All nodules were scored in accordance with the Liver Imaging Reporting and Data System (LI-RADS v2014), with or without consideration of ring-like enhancement in multiphasic HAP images as a major feature. In the multiphasic HAP group, 178 of 191 HCCs were scored as LR-3 to LR-5 (3 [1.69%], 85 [47.8%], and 90 [50.6%], respectively). Upon considering ring-like enhancement in multiphasic HAP images as a major feature, 5 more HCCs were scored as LR-5 (95 [53.4%]), which was a significantly more confident diagnosis than that with single HAP images (295 of 306 HCCs scored as LR-3 to LR-5: 13 [4.41%], 147 [49.8%], and 135 [45.8%], respectively; P = 0.0296). There was no significant difference in false-positive or false-negative diagnoses between the multiphasic and single HAP groups (P = 0.8400 and 0.1043, respectively). Multiphasic HAP imaging can improve the confidence of diagnosis of HCCs in gadoxetic acid-enhanced magnetic resonance imaging.

  6. Enhanced Cooperation, EMU Reforms and Their Implications for Differentiation in the European Union

    Directory of Open Access Journals (Sweden)

    Kubin Tomasz

    2017-10-01

    Full Text Available Initially, before the entry into force of the Maastricht Treaty, differences in integration between members of the European Communities (EC; later the European Union were relatively few and usually temporary in nature. The Schengen Agreement, the Maastricht Treaty and the Treaty of Amsterdam, and the possibility of establishing enhanced cooperation meant that the problem was becoming more and more important in the functioning of the EU—both in theory and in practice.

  7. An Algorithm of Image Heterogeneity with Contrast-Enhanced Ultrasound in Differential Diagnosis of Solid Thyroid Nodules.

    Science.gov (United States)

    Jin, Lifang; Xu, Changsong; Xie, Xueqian; Li, Fan; Lv, Xiuhong; Du, Lianfang

    2017-01-01

    Enhancement heterogeneity on contrast-enhanced ultrasonography (CEUS) is used to differentiate between benign and malignant thyroid nodules. In this study, we used an algorithm to quantify enhancement heterogeneity of solid thyroid nodules on CEUS. The heterogeneity value (HV) is calculated as standard deviation/mean intensity × 100 (using Adobe Photoshop). The heterogeneity ratio (HR) is calculated as the ratio of the HV of the nodule to that of the surrounding parenchyma. Three phases-ascending, peak and descending phases-were studied. HV values at ascending (HV a ) and peak (HV p ) phases were significantly higher in malignant nodules than in benign nodules (95.57 ± 43.87 vs. 73.06 ± 44.04, p = 0.009, and 32.53 ± 10.73 vs. 26.44 ± 8.25, p = 0.002, respectively). HR a , HR p and HR d were significantly higher in malignant nodules than in benign nodules (1.93 ± 1.03 vs. 1.00 ± 0.47, p = 0.000, 1.43 ± 0.51 vs. 1.09 ± 0.28, p = 0.000, and 1.33 ± 0.40 vs. 1.08 ± 0.33, p = 0.001, respectively). HR a achieved optimal diagnostic performance on receiver operating characteristic curve analysis. The algorithm used for assessment of image heterogeneity on CEUS examination may be a useful adjunct to conventional ultrasound for differential diagnosis of solid thyroid nodules. Copyright © 2016 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

  8. Acute ingestion of different macronutrients differentially enhances aspects of memory and attention in healthy young adults.

    Science.gov (United States)

    Jones, Emma K; Sünram-Lea, Sandra I; Wesnes, Keith A

    2012-02-01

    The role of carbohydrates on mood and cognition is fairly well established, however research examining the behavioural effects of the other macronutrients is limited. The current study compared the effects of a 25 g glucose drink to energetically matched protein and fat drinks and an inert placebo. Following a blind, placebo-controlled, randomised crossover design, 18 healthy young adults consumed drinks containing fat, glucose, protein and placebo. Cognitive performance was examined at baseline and again 15- and 60 min post drink. Mood was assessed at baseline and then 10-, 35- and 80 min post drink. Attention and speed were enhanced 15 min following fat or glucose ingestion and working memory was enhanced 15 min following protein ingestion. Sixty minutes post drink memory enhancements were observed after protein and memory impairment was observed following glucose. All drinks increased ratings of alertness. The findings suggest that macronutrients: (i) have different windows of opportunity for effects (ii) target different cognitive domains. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. Differential permeation of piroxicam-loaded PLGA micro/nanoparticles and their in vitro enhancement

    International Nuclear Information System (INIS)

    Shankarayan, Raju; Kumar, Sumit; Mishra, Prashant

    2013-01-01

    Piroxicam is a non-steroidal anti-inflammatory drug used for the treatment of musculoskeletal pain. The main problem encountered when piroxicam is administered orally is its gastric side-effect (ulcer, bleeding and holes in the stomach). Transmucosal delivery and encapsulation of piroxicam in biodegradable particles offer potential advantages over conventional oral delivery. The present study was aimed to develop an alternative to piroxicam-delivery which could overcome the direct contact of the drug at the mucosal membrane and its permeation through the mucosal membrane was studied. To achieve this, the piroxicam was encapsulated in Poly (lactide-co-glycolide) (PLGA) microparticles (size 1–4 μm, encapsulation efficiency 80–85 %) and nanoparticles (size 151.6 ± 28.6 nm, encapsulation efficiency 92.17 ± 3.08 %). Various formulation process parameters were optimised for the preparation of piroxicam-loaded PLGA nanoparticles of optimal size and encapsulation efficiency. Transmucosal permeability of piroxicam-loaded PLGA micro- and nanoparticles through the porcine oesophageal mucosa was studied. Using fluorescently labelled PLGA micro- and nanoparticles, size-dependent permeation was demonstrated. Furthermore, the effect of different permeation enhancers on the flux rate and permeability coefficient for the permeation of nanoparticles was investigated. The results suggested that amongst the permeation enhancers used the most efficient enhancement of permeation was observed with 10 mM sodium dodecyl sulphate.

  10. Differential permeation of piroxicam-loaded PLGA micro/nanoparticles and their in vitro enhancement

    Energy Technology Data Exchange (ETDEWEB)

    Shankarayan, Raju; Kumar, Sumit; Mishra, Prashant, E-mail: pmishra@dbeb.iitd.ac.in [Indian Institute of Technology Delhi, Department of Biochemical Engineering and Biotechnology (India)

    2013-03-15

    Piroxicam is a non-steroidal anti-inflammatory drug used for the treatment of musculoskeletal pain. The main problem encountered when piroxicam is administered orally is its gastric side-effect (ulcer, bleeding and holes in the stomach). Transmucosal delivery and encapsulation of piroxicam in biodegradable particles offer potential advantages over conventional oral delivery. The present study was aimed to develop an alternative to piroxicam-delivery which could overcome the direct contact of the drug at the mucosal membrane and its permeation through the mucosal membrane was studied. To achieve this, the piroxicam was encapsulated in Poly (lactide-co-glycolide) (PLGA) microparticles (size 1-4 {mu}m, encapsulation efficiency 80-85 %) and nanoparticles (size 151.6 {+-} 28.6 nm, encapsulation efficiency 92.17 {+-} 3.08 %). Various formulation process parameters were optimised for the preparation of piroxicam-loaded PLGA nanoparticles of optimal size and encapsulation efficiency. Transmucosal permeability of piroxicam-loaded PLGA micro- and nanoparticles through the porcine oesophageal mucosa was studied. Using fluorescently labelled PLGA micro- and nanoparticles, size-dependent permeation was demonstrated. Furthermore, the effect of different permeation enhancers on the flux rate and permeability coefficient for the permeation of nanoparticles was investigated. The results suggested that amongst the permeation enhancers used the most efficient enhancement of permeation was observed with 10 mM sodium dodecyl sulphate.

  11. Differential permeation of piroxicam-loaded PLGA micro/nanoparticles and their in vitro enhancement

    Science.gov (United States)

    Shankarayan, Raju; Kumar, Sumit; Mishra, Prashant

    2013-03-01

    Piroxicam is a non-steroidal anti-inflammatory drug used for the treatment of musculoskeletal pain. The main problem encountered when piroxicam is administered orally is its gastric side-effect (ulcer, bleeding and holes in the stomach). Transmucosal delivery and encapsulation of piroxicam in biodegradable particles offer potential advantages over conventional oral delivery. The present study was aimed to develop an alternative to piroxicam-delivery which could overcome the direct contact of the drug at the mucosal membrane and its permeation through the mucosal membrane was studied. To achieve this, the piroxicam was encapsulated in Poly (lactide- co-glycolide) (PLGA) microparticles (size 1-4 μm, encapsulation efficiency 80-85 %) and nanoparticles (size 151.6 ± 28.6 nm, encapsulation efficiency 92.17 ± 3.08 %). Various formulation process parameters were optimised for the preparation of piroxicam-loaded PLGA nanoparticles of optimal size and encapsulation efficiency. Transmucosal permeability of piroxicam-loaded PLGA micro- and nanoparticles through the porcine oesophageal mucosa was studied. Using fluorescently labelled PLGA micro- and nanoparticles, size-dependent permeation was demonstrated. Furthermore, the effect of different permeation enhancers on the flux rate and permeability coefficient for the permeation of nanoparticles was investigated. The results suggested that amongst the permeation enhancers used the most efficient enhancement of permeation was observed with 10 mM sodium dodecyl sulphate.

  12. Dual Function of Glucosamine in Gelatin/Hyaluronic Acid Cryogel to Modulate Scaffold Mechanical Properties and to Maintain Chondrogenic Phenotype for Cartilage Tissue Engineering.

    Science.gov (United States)

    Chen, Chih-Hao; Kuo, Chang-Yi; Wang, Yan-Jie; Chen, Jyh-Ping

    2016-11-23

    Glucosamine (GlcN) fulfills many of the requirements as an ideal component in scaffolds used in cartilage tissue engineering. The incorporation of GlcN in a gelatin/hyaluronic acid (GH) cryogel scaffold could provide biological cues in maintaining the phenotype of chondrocytes. Nonetheless, substituting gelatin with GlcN may also decrease the crosslinking density and modulate the mechanical properties of the cryogel scaffold, which may be beneficial as physical cues for chondrocytes in the scaffold. Thus, we prepared cryogel scaffolds containing 9% GlcN (GH-GlcN9) and 16% GlcN (GH-GlcN16) by carbodiimide-mediated crosslinking reactions at -16 °C. The crosslinking density and the mechanical properties of the cryogel matrix could be tuned by adjusting the content of GlcN used during cryogel preparation. In general, incorporation of GlcN did not influence scaffold pore size and ultimate compressive strain but increased porosity. The GH-GlcN16 cryogel showed the highest swelling ratio and degradation rate in hyaluronidase and collagenase solutions. On the contrary, the Young's modulus, storage modulus, ultimate compressive stress, energy dissipation level, and rate of stress relaxation decreased by increasing the GlcN content in the cryogel. The release of GlcN from the scaffolds in the culture medium of chondrocytes could be sustained for 21 days for GH-GlcN16 in contrast to only 7 days for GH-GlcN9. In vitro cell culture experiments using rabbit articular chondrocytes revealed that GlcN incorporation affected cell proliferation, morphology, and maintenance of chondrogenic phenotype. Overall, GH-GlcN16 showed the best performance in maintaining chondrogenic phenotype with reduced cell proliferation rate but enhanced glycosaminoglycans (GAGs) and type II collagen (COL II) secretion. Quantitative real-time polymerase chain reaction also showed time-dependent up-regulation of cartilage-specific marker genes (COL II, aggrecan and Sox9) for GH-GlcN16. Implantation of

  13. A resolution-enhancing image reconstruction method for few-view differential phase-contrast tomography

    Science.gov (United States)

    Guan, Huifeng; Anastasio, Mark A.

    2017-03-01

    It is well-known that properly designed image reconstruction methods can facilitate reductions in imaging doses and data-acquisition times in tomographic imaging. The ability to do so is particularly important for emerging modalities such as differential X-ray phase-contrast tomography (D-XPCT), which are currently limited by these factors. An important application of D-XPCT is high-resolution imaging of biomedical samples. However, reconstructing high-resolution images from few-view tomographic measurements remains a challenging task. In this work, a two-step sub-space reconstruction strategy is proposed and investigated for use in few-view D-XPCT image reconstruction. It is demonstrated that the resulting iterative algorithm can mitigate the high-frequency information loss caused by data incompleteness and produce images that have better preserved high spatial frequency content than those produced by use of a conventional penalized least squares (PLS) estimator.

  14. Quantitative contrast-enhanced harmonic EUS in differential diagnosis of focal pancreatic masses (with videos)

    DEFF Research Database (Denmark)

    Săftoiu, Adrian; Vilmann, Peter; Dietrich, Christoph F

    2015-01-01

    BACKGROUND: The role of EUS with contrast agents can be expanded through the use of time-intensity curve (TIC) analysis and computer-aided interpretation. OBJECTIVE: To validate the use of parameters derived from TIC analysis in an artificial neural network (ANN) classification model designed......-guided FNA (EUS-FNA), TIC analysis, and ANN processing. MAIN OUTCOME MEASUREMENTS: Sensitivity, specificity, positive and negative predictive values (PPV, NPV) for EUS-FNA, CEH-EUS, and the ANN. RESULTS: After excluding all of the recordings that did not meet the technical and procedural criteria, 112 cases...... rise time, mean transit time, and time to peak. For the ANN, sensitivity was 94.64%, specificity 94.44%, PPV 97.24%, and NPV 89.47%. LIMITATIONS: Only PC and CP lesions were included. CONCLUSION: Parameters obtained through TIC analysis can differentiate between PC and CP cases and can be used...

  15. Face and voice as social stimuli enhance differential physiological responding in a Concealed Information Test

    Directory of Open Access Journals (Sweden)

    Wolfgang eAmbach

    2012-11-01

    Full Text Available Attentional, intentional, and motivational factors are known to influence the physiological responses in a Concealed Information Test (CIT. Although concealing information is essentially a social action closely related to motivation, CIT studies typically rely on testing participants in an environment lacking of social stimuli: Subjects interact with a computer while sitting alone in an experimental room. To address this gap, we examined the influence of social stimuli on the physiological responses in a CIT.Seventy-one participants underwent a mock-crime experiment with a modified CIT. In a between-subjects design, subjects were either questioned acoustically by a pre-recorded male voice presented together with a virtual male experimenter’s uniform face or by a text field on the screen, which displayed the question devoid of face and voice. Electrodermal activity (EDA, respiration line length (RLL, phasic heart rate (pHR, and finger pulse waveform length (FPWL were registered. The Psychopathic Personality Inventory - Revised (PPI-R was administered in addition. The differential responses of RLL, pHR, and FPWL to probe vs. irrelevant items were greater in the condition with social stimuli than in the text condition; interestingly, the differential responses of EDA did not differ between conditions. No modulatory influence of the PPI-R sum or subscale scores was found.The results emphasize the relevance of social aspects in the process of concealing information and in its detection. Attentional demands as well as the participants’ motivation to avoid detection might be the important links between social stimuli and physiological responses in the CIT.

  16. Combined 3 Tesla MRI Biomarkers Improve the Differentiation between Benign vs Malignant Single Ring Enhancing Brain Masses.

    Directory of Open Access Journals (Sweden)

    Simone Salice

    Full Text Available To evaluate whether the combination of imaging biomarkers obtained by means of different 3 Tesla (3T Magnetic Resonance Imaging (MRI advanced techniques can improve the diagnostic accuracy in the differentiation between benign and malignant single ring-enhancing brain masses.14 patients presenting at conventional 3T MRI single brain mass with similar appearance as regard ring enhancement, presence of peri-lesional edema and absence of hemorrhage signs were included in the study. All lesions were histologically proven: 5 pyogenic abscesses, 6 glioblastomas, and 3 metastases. MRI was performed at 3 Tesla and included Diffusion Weighted Imaging (DWI, Dynamic Susceptibility Contrast -Perfusion Weighted Imaging (DSC-PWI, Magnetic Resonance Spectroscopy (MRS, and Diffusion Tensor Imaging (DTI. Imaging biomarkers derived by those advanced techniques [Cerebral Blood Flow (CBF, relative Cerebral Blood Volume (rCBV, relative Main Transit Time (rMTT, Choline (Cho, Creatine (Cr, Succinate, N-Acetyl Aspartate (NAA, Lactate (Lac, Lipids, relative Apparent Diffusion Coefficient (rADC, and Fractional Anisotropy (FA] were detected by two experienced neuroradiologists in joint session in 4 areas: Internal Cavity (IC, Ring Enhancement (RE, Peri-Lesional edema (PL, and Contralateral Normal Appearing White Matter (CNAWM. Significant differences between benign (n = 5 and malignant (n = 9 ring enhancing lesions were tested with Mann-Withney U test. The diagnostic accuracy of MRI biomarkers taken alone and MRI biomarkers ratios were tested with Receiver Operating Characteristic (ROC analysis with an Area Under the Curve (AUC ≥ 0.9 indicating a very good diagnostic accuracy of the variable.Five MRI biomarker ratios achieved excellent accuracy: IC-rADC/PL-NAA (AUC = 1, IC-rADC/IC-FA (AUC = 0.978, RE-rCBV/RE-FA (AUC = 0.933, IC-rADC/RE-FA (AUC = 0.911, and IC-rADC/PL-FA (AUC = 0.911. Only IC-rADC achieved a very good diagnostic accuracy (AUC = 0.909 among MRI biomarkers

  17. Mentalizing skills do not differentiate believers from non-believers, but credibility enhancing displays do.

    Directory of Open Access Journals (Sweden)

    David L R Maij

    Full Text Available The ability to mentalize has been marked as an important cognitive mechanism enabling belief in supernatural agents. In five studies we cross-culturally investigated the relationship between mentalizing and belief in supernatural agents with large sample sizes (over 67,000 participants in total and different operationalizations of mentalizing. The relative importance of mentalizing for endorsing supernatural beliefs was directly compared with credibility enhancing displays-the extent to which people observed credible religious acts during their upbringing. We also compared autistic with neurotypical adolescents. The empathy quotient and the autism-spectrum quotient were not predictive of belief in supernatural agents in all countries (i.e., The Netherlands, Switzerland and the United States, although we did observe a curvilinear effect in the United States. We further observed a strong influence of credibility enhancing displays on belief in supernatural agents. These findings highlight the importance of cultural learning for acquiring supernatural beliefs and ask for reconsiderations of the importance of mentalizing.

  18. * Tissue-Specific Extracellular Matrix Enhances Skeletal Muscle Precursor Cell Expansion and Differentiation for Potential Application in Cell Therapy.

    Science.gov (United States)

    Zhang, Deying; Zhang, Yong; Zhang, Yuanyuan; Yi, Hualin; Wang, Zhan; Wu, Rongpei; He, Dawei; Wei, Guanghui; Wei, Shicheng; Hu, Yun; Deng, Junhong; Criswell, Tracy; Yoo, James; Zhou, Yu; Atala, Anthony

    2017-08-01

    Skeletal muscle precursor cells (MPCs) are considered a key candidate for cell therapy in the treatment of skeletal muscle dysfunction due to injury, disease, or age. However, expansion of a sufficient number of functional skeletal muscle cells in vitro from a small tissue biopsy has been challenging due to changes in phenotypic expression of these cells under traditional culture conditions. Thus, the aim of the study was to develop a better culture system for the expansion and myo-differentiation of MPCs that could further be used for therapy. For this purpose, we developed an ideal method of tissue decellularization and compared the ability of different matrices to support MPC growth and differentiation. Porcine-derived skeletal muscle and liver and kidney extracellular matrix (ECM) were generated by decellularization methods consisting of distilled water, 0.2 mg/mL DNase, or 5% fetal bovine serum. Acellular matrices were further homogenized, dissolved, and combined with a hyaluronic acid-based hydrogel decorated with heparin (ECM-HA-HP). The cell proliferation and myogenic differentiation capacity of human MPCs were assessed when grown on gel alone, ECM, or each ECM-HA-HP substrate. Human MPC proliferation was significantly enhanced when cultured on the ECM-HA-HP substrates compared to the other substrates tested, with the greatest proliferation on the muscle ECM-HA-HP (mECM-HA-HP) substrate. The number of differentiated myotubes was significantly increased on the mECM-HA-HP substrate compared to the other gel-ECM substrates, as well as the numbers of MPCs expressing specific myogenic cell markers (i.e., myosin, desmin, myoD, and myf5). In conclusion, skeletal mECM-HA-HP as a culture substrate provided an optimal culture microenvironment potentially due to its similarity to the in vivo environment. These data suggest a potential use of skeletal muscle-derived ECM gel for the expansion and differentiation of human MPCs for cell-based therapy for skeletal muscle

  19. Enhanced lipase recovery through RSM integrated differential evolutionary approach from the fermented biomass

    Directory of Open Access Journals (Sweden)

    Vijay Kumar Garlapati

    2013-10-01

    Full Text Available The aim of this work was to apply a modeling integrated optimisation approach for a complex, highly nonlinear system for an extracellular lipase extraction process. The model was developed using mutation, crossover and selection variables of Differential Evolution (DE based on central composite design of Response Surface Methodology. The experimentally validated model was optimized by DE, a robust evolutionary optimization tool. A maximum lipase activity of 134.13 U/gds (more than 36.28 U/gds compared to one variable at a time approach was observed with the DE-stated optimum values of 25.01% dimethyl sulfoxide concentration, 40 mM buffer, 128.52 min soaking time and 35ºC with the DE control parameters, namely number of population, generations, crossover operator and scaling factor as 20, 50, 0.5 and 0.25, respectively. The use of DE approach improved the optimization capability and decision speed, resulting in an improved yield of 36.28 U/gds compared to the one variable at a time approach for the extracellular lipase activity under the non-optimized conditions. The developed mathematical model and optimization were generic in nature, which seemed to be useful for the scale-up studies of maximum recovery of lipase from the fermented biomass.

  20. Using Differential Evolution to Optimize Learning from Signals and Enhance Network Security

    Energy Technology Data Exchange (ETDEWEB)

    Harmer, Paul K [Air Force Institute of Technology; Temple, Michael A [Air Force Institute of Technology; Buckner, Mark A [ORNL; Farquhar, Ethan [ORNL

    2011-01-01

    Computer and communication network attacks are commonly orchestrated through Wireless Access Points (WAPs). This paper summarizes proof-of-concept research activity aimed at developing a physical layer Radio Frequency (RF) air monitoring capability to limit unauthorizedWAP access and mprove network security. This is done using Differential Evolution (DE) to optimize the performance of a Learning from Signals (LFS) classifier implemented with RF Distinct Native Attribute (RF-DNA) fingerprints. Performance of the resultant DE-optimized LFS classifier is demonstrated using 802.11a WiFi devices under the most challenging conditions of intra-manufacturer classification, i.e., using emissions of like-model devices that only differ in serial number. Using identical classifier input features, performance of the DE-optimized LFS classifier is assessed relative to a Multiple Discriminant Analysis / Maximum Likelihood (MDA/ML) classifier that has been used for previous demonstrations. The comparative assessment is made using both Time Domain (TD) and Spectral Domain (SD) fingerprint features. For all combinations of classifier type, feature type, and signal-to-noise ratio considered, results show that the DEoptimized LFS classifier with TD features is uperior and provides up to 20% improvement in classification accuracy with proper selection of DE parameters.

  1. Using meta-differential evolution to enhance a calculation of a continuous blood glucose level.

    Science.gov (United States)

    Koutny, Tomas

    2016-09-01

    We developed a new model of glucose dynamics. The model calculates blood glucose level as a function of transcapillary glucose transport. In previous studies, we validated the model with animal experiments. We used analytical method to determine model parameters. In this study, we validate the model with subjects with type 1 diabetes. In addition, we combine the analytic method with meta-differential evolution. To validate the model with human patients, we obtained a data set of type 1 diabetes study that was coordinated by Jaeb Center for Health Research. We calculated a continuous blood glucose level from continuously measured interstitial fluid glucose level. We used 6 different scenarios to ensure robust validation of the calculation. Over 96% of calculated blood glucose levels fit A+B zones of the Clarke Error Grid. No data set required any correction of model parameters during the time course of measuring. We successfully verified the possibility of calculating a continuous blood glucose level of subjects with type 1 diabetes. This study signals a successful transition of our research from an animal experiment to a human patient. Researchers can test our model with their data on-line at https://diabetes.zcu.cz. Copyright © 2016 The Author. Published by Elsevier Ireland Ltd.. All rights reserved.

  2. Throughput Fairness Enhancement Using Differentiated Channel Access in Heterogeneous Sensor Networks

    Directory of Open Access Journals (Sweden)

    James Jong Hyuk Park

    2011-06-01

    Full Text Available Nowadays, with wireless sensor networks (WSNs being widely applied to diverse applications, heterogeneous sensor networks (HSNs, which can simultaneously support multiple sensing tasks in a common sensor field, are being considered as the general form of WSN system deployment. In HSNs, each application generates data packets with a different size, thereby resulting in fairness issues in terms of the network performance. In this paper, we present the design and performance evaluation of a differentiated channel access scheme (abbreviated to DiffCA to resolve the fairness problem in HSNs. DiffCA achieves fair performance among the application groups by providing each node with an additional backoff counter, whose value varies according to the size of the packets. A mathematical model based on the discrete time Markov chain is presented and is analyzed to measure the performance of DiffCA. The numerical results show that the performance degradation of disadvantaged application groups can be effectively compensated for by DiffCA. Simulation results are given to verify the accuracy of the numerical model.

  3. κ-Carrageenan Enhances the Biomineralization and Osteogenic Differentiation of Electrospun Polyhydroxybutyrate and Polyhydroxybutyrate Valerate Fibers.

    Science.gov (United States)

    Goonoo, Nowsheen; Khanbabaee, Behnam; Steuber, Marc; Bhaw-Luximon, Archana; Jonas, Ulrich; Pietsch, Ullrich; Jhurry, Dhanjay; Schönherr, Holger

    2017-05-08

    Novel electrospun materials for bone tissue engineering were obtained by blending biodegradable polyhydroxybutyrate (PHB) or polyhydroxybutyrate valerate (PHBV) with the anionic sulfated polysaccharide κ-carrageenan (κ-CG) in varying ratios. In both systems, the two components phase separated as shown by FTIR, DSC and TGA. According to the contact angle data, κ-CG was localized preferentially at the fiber surface in PHBV/κ-CG blends in contrast to PHB/κ-CG, where the biopolymer was mostly found within the fiber. In contrast to the neat polyester fibers, the blends led to the formation of much smaller apatite crystals (800 nm vs 7 μm). According to the MTT assay, NIH3T3 cells grew in higher density on the blend mats in comparison to neat polyester mats. The osteogenic differentiation potential of the fibers was determined by SaOS-2 cell culture for 2 weeks. Alizarin red-S staining suggested an improved mineralization on the blend fibers. Thus, PHBV/κ-CG fibers resulted in more pronounced bioactive and osteogenic properties, including fast apatite-forming ability and deposition of nanosized apatite crystals.

  4. Throughput fairness enhancement using differentiated channel access in heterogeneous sensor networks.

    Science.gov (United States)

    Kim, Eui-Jik; Shon, Taeshik; Park, James Jong Hyuk; Jeong, Young-Sik

    2011-01-01

    Nowadays, with wireless sensor networks (WSNs) being widely applied to diverse applications, heterogeneous sensor networks (HSNs), which can simultaneously support multiple sensing tasks in a common sensor field, are being considered as the general form of WSN system deployment. In HSNs, each application generates data packets with a different size, thereby resulting in fairness issues in terms of the network performance. In this paper, we present the design and performance evaluation of a differentiated channel access scheme (abbreviated to DiffCA) to resolve the fairness problem in HSNs. DiffCA achieves fair performance among the application groups by providing each node with an additional backoff counter, whose value varies according to the size of the packets. A mathematical model based on the discrete time Markov chain is presented and is analyzed to measure the performance of DiffCA. The numerical results show that the performance degradation of disadvantaged application groups can be effectively compensated for by DiffCA. Simulation results are given to verify the accuracy of the numerical model.

  5. Glucose administration prior to a divided attention task improves tracking performance but not word recognition: evidence against differential memory enhancement?

    Science.gov (United States)

    Scholey, Andrew B; Sünram-Lea, Sandra I; Greer, Joanna; Elliott, Jade; Kennedy, David O

    2009-01-01

    The cognition-enhancing effects of glucose administration to humans have been well-documented; however, it remains unclear whether this effect preferentially targets episodic memory or other cognitive domains. The effect of glucose on the allocation of attentional resources during memory encoding was assessed using a sensitive dual-attention paradigm. One hundred and twenty volunteers (mean age 21.60, SD 4.89, 77 females) took part in this randomised, double-blind, placebo-controlled, parallel groups study where each consumed a 25-g glucose drink or a placebo. Half of the participants in each drink condition attempted to track a moving on-screen target during auditory word presentation. The distance between the cursor and the tracking target was used as an index of attentional cost during encoding. Effects of drink and tracking on recognition memory and drink on tracking performance were assessed. Self-rated appetite and mood were co-monitored. Co-performing the tracking task significantly impaired memory performance irrespective of drink condition. In the placebo-tracking condition, there was a cost to tracking m