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Sample records for ctgf mrna expression

  1. Effects of 17β-estradiol on the expressions of CTGF and PAIP-1 in MG-63 cells

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    翟木绪

    2006-01-01

    Objective To observe the expressions of connective tissue growth factor (CTGF) and polyadenylate-binding protein interacting protein-1 (PAIP-1) mRNA during MG-63 cell proliferation and differentiation, and to investigate the effect of 17β-estradiol (E2) on the expressions of CTGF and PAIP-1 mRNA. Methods The expressions of typeⅠcollagen, alkaline phosphatase (ALP) and osteocalcin mRNA were determined by semiquantitative RT-PCR. Cultured cells were stained with

  2. Alteration of Connective Tissue Growth Factor (CTGF) Expression in Orbital Fibroblasts from Patients with Graves' Ophthalmopathy.

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    Tsai, Chieh-Chih; Wu, Shi-Bei; Chang, Pei-Chen; Wei, Yau-Huei

    2015-01-01

    Graves' ophthalmopathy (GO) is a disfiguring and sometimes blinding disease, which is characterized by inflammation and swelling of orbital tissues, with fibrosis and adipogenesis being predominant features. The aim of this study is to investigate whether the expression levels of fibrosis-related genes, especially that of connective tissue growth factor (CTGF), are altered in orbital fibroblasts of patients with GO. The role of oxidative stress in the regulation of CTGF expression in GO orbital fibroblasts is also examined. By a SYBR Green-based real time quantitative PCR (RT-QPCR), we demonstrated that the mRNA expression levels of fibronectin, apolipoprotein J, and CTGF in cultured orbital fibroblasts from patients with GO were significantly higher than those of age-matched normal controls (p = 0.007, 0.037, and 0.002, respectively). In addition, the protein expression levels of fibronectin, apolipoprotein J, and CTGF analyzed by Western blot were also significantly higher in GO orbital fibroblasts (p = 0.046, 0.032, and 0.008, respectively) as compared with the control. Furthermore, after treatment of orbital fibroblasts with a sub-lethal dose of hydrogen peroxide (200 μM H2O2), we found that the H2O2-induced increase of CTGF expression was more pronounced in the GO orbital fibroblasts as compared with those in normal controls (20% vs. 7%, p = 0.007). Importantly, pre-incubation with antioxidants including N-acetylcysteine (NAC) and vitamin C, respectively, resulted in significant attenuation of the induction of CTGF in GO orbital fibroblasts in response to H2O2 (p = 0.004 and 0.015, respectively). Taken together, we suggest that oxidative stress plays a role in the alteration of the expression of CTGF in GO orbital fibroblasts that may contribute to the pathogenesis and progression of GO. Antioxidants may be used in combination with the therapeutic agents for effective treatment of GO.

  3. Expression of CYR61, CTGF, VEGF-C, VEGFR-2 mRNA in Bone Marrow of Leukemia Patients and Its Clinical Significance%白血病患者骨髓CYR61、CTGF、VEGF-C、VEGFR-2mRNA的表达及其临床意义

    Institute of Scientific and Technical Information of China (English)

    徐宁; 李静; 李喆; 李志芹; 韩海燕; 贺其图; 卢燕; 韩轩茂; 马宏杰; 张冬霞; 刘学文; 袁晓俊; 贾国荣

    2011-01-01

    The study was aimed to detect the levels of CYR61, CTGF, VEGF-C, VEGFR-2 mRNA in bone marrow (BM) of leukemia patients and investigate the interaction of CYR61, CTGF, VEGF-C, VEGFR-2 protiens in occurrence, development, infiltration and metastasis of leukemia and its clinical significance, to find a new tumor marker for diagnosis and treatment of leukemia with some new directions. 74 patients with leukemia were enrolled in this study, 38 out of them were males and 36 were females, aged from 6 to 77 years old with the median age of 45 years old. In the control group, 7 males and 5 females, aged from 16 to 78 years old with the median age of 46. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) was used to detect the levels of CYR61, CTGF, VEGF-C, VEGFR-2 mRNA. The results showed that the levels of CYR61, CTGF, VEGF-C, VEGFR-2 mRNA in BM of newly diagnosed patients with acute and chronic leukemia of each group were significantly higher as compared with the control group (p < 0.05). The levels of CYR61, CTGF mRNA in acute leukemia remission group were significantly higher than those in control group (p = 0.039, 0.025). The level of CTGF mRNA was highest in B-ALL group, and was higher than that in AML, CML, CLL, T-ALL groups (p =0. 002, 0. 034, 0. 002, 0. 010). In AML group, mRNA expressions of CYR61 and CTGF, CYR61 and VEGF-C, CTGF and VEGFR-2 were positively correlated (r =0. 452, 0. 466, 0. 464; p = 0.045, 0.038, 0.039), and in CML group mRNA expression of CYR61 and VEGF-C was positively correlated (r =0.882,p =0.000). The expression levels of VEGF-C, VEGFR-2 mRNA in acute leukemia patients with extramedullary infiltration were higher than those in acute leukemia patients without extramedullary infiltration (p =0. 028, 0. 047). VEGF-C mRNA expression and the original cell counts in AML group were positively correlated ( r = 0. 418, p = 0. 034). It is concluded that CYR61, CTGF, VEGF-C and VEGFR-2 interact each other in the pathogenesis of

  4. Mechanical stretch increases CCN2/CTGF expression in anterior cruciate ligament-derived cells

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    Miyake, Yoshiaki [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Furumatsu, Takayuki, E-mail: matino@md.okayama-u.ac.jp [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Kubota, Satoshi; Kawata, Kazumi [Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Ozaki, Toshifumi [Department of Orthopaedic Surgery, Science of Functional Recovery and Reconstruction, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan); Takigawa, Masaharu [Department of Biochemistry and Molecular Dentistry, Okayama University Graduate School of Medicine, Dentistry, and Pharmaceutical Sciences, 2-5-1 Shikata-cho, Kita-ku, Okayama (Japan)

    2011-06-03

    Highlights: {yields} CCN2/CTGF localizes to the ligament-to-bone interface, but is not to the midsubstance region of human anterior cruciate ligament (ACL). {yields} Mechanical stretch induces higher increase of CCN2/CTGF gene expression and protein secretion in ACL interface cells compared with ACL midsubstance cells. {yields} CCN2/CTGF treatment stimulates the proliferation of ACL interface cells. -- Abstract: Anterior cruciate ligament (ACL)-to-bone interface serves to minimize the stress concentrations that would arise between two different tissues. Mechanical stretch plays an important role in maintaining cell-specific features by inducing CCN family 2/connective tissue growth factor (CCN2/CTGF). We previously reported that cyclic tensile strain (CTS) stimulates {alpha}1(I) collagen (COL1A1) expression in human ACL-derived cells. However, the biological function and stress-related response of CCN2/CTGF were still unclear in ACL fibroblasts. In the present study, CCN2/CTGF was observed in ACL-to-bone interface, but was not in the midsubstance region by immunohistochemical analyses. CTS treatments induced higher increase of CCN2/CTGF expression and secretion in interface cells compared with midsubstance cells. COL1A1 expression was not influenced by CCN2/CTGF treatment in interface cells despite CCN2/CTGF stimulated COL1A1 expression in midsubstance cells. However, CCN2/CTGF stimulated the proliferation of interface cells. Our results suggest that distinct biological function of stretch-induced CCN2/CTGF might regulate region-specific phenotypes of ACL-derived cells.

  5. Expression and clinical significance of connective tissue growth factor (CTGF) in Graves' ophthalmopathy.

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    Huang, Yi-Ming; Chang, Pei-Chen; Wu, Shi-Bei; Kau, Hui-Chuan; Tsai, Chieh-Chih; Liu, Catherine Jui-Ling; Wei, Yau-Huei

    2017-05-01

    To examine the expression of connective tissue growth factor (CTGF) in human cultured orbital fibroblasts from patients with Graves' ophthalmopathy (GO) and investigate whether a correlation exists between the presence of CTGF protein and clinical parameters of the disease. The protein expression levels of CTGF were analysed by western blots in cultured orbital fibroblasts from 10 patients with GO and 7 age-matched normal controls. Associations between the protein expression of CTGF and the clinical factors of GO, including clinical demographics, thyroid function, clinical activity score (CAS) and ophthalmopathy index (OI), was evaluated. The mean protein expression levels of CTGF in the GO orbital fibroblasts were significantly higher than those of normal controls (ppathophysiology of GO. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  6. CTGF increases matrix metalloproteinases expression and subsequently promotes tumor metastasis in human osteosarcoma through down-regulating miR-519d.

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    Tsai, Hsiao-Chi; Su, Hong-Lin; Huang, Chun-Yin; Fong, Yi-Chin; Hsu, Chin-Jung; Tang, Chih-Hsin

    2014-06-15

    Osteosarcoma, the most common primary malignant bone tumor, shows potent capacity for local invasion and distant metastasis. Connective tissue growth factor (CTGF/CCN2), a secreted protein, binds to integrins, modulates invasive behavior of certain human cancer cells. Effect of CTGF in metastasis of human osteosarcoma is unknown. We found overexpression of CTGF increasing matrix metalloproteinases (MMPs)-2 and MMP-3 expression as well as promoting cell migration. MicroRNA (miRNA) analysis of CTGF-overexpressed osteosarcoma versus control cells probed mechanisms of CTGF-mediated promotion of migration. Among miRNAs regulated by CTGF, miR-519d was most downregulated after CTGF treatment. Co-transfection with miR-519d mimic reversed CTGF-mediated MMPs expression and cell migration. Also, MEK and ERK inhibitors or mutants reduced CTGF-increased cell migration and miR-519d suppression. By contrast, knockdown of CTGF diminished lung metastasis in vivo. Clinical samples indicate CTGF expression as linked with clinical stage and tumor metastasis. Taken together, data show CTGF elevating MMPs expression and subsequently promoting tumor metastasis in human osteosarcoma, down-regulating miR-519d via MEK and ERK pathways, making CTGF a new molecular therapeutic target in osteosarcoma metastasis.

  7. CTGF increases matrix metalloproteinases expression and subsequently promotes tumor metastasis in human osteosarcoma through down-regulating miR-519d

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    Tsai, Hsiao-Chi; Su, Hong-Lin; Huang, Chun-Yin; Fong, Yi-Chin; Hsu, Chin-Jung; Tang, Chih-Hsin

    2014-01-01

    Osteosarcoma, the most common primary malignant bone tumor, shows potent capacity for local invasion and distant metastasis. Connective tissue growth factor (CTGF/CCN2), a secreted protein, binds to integrins, modulates invasive behavior of certain human cancer cells. Effect of CTGF in metastasis of human osteosarcoma is unknown. We found overexpression of CTGF increasing matrix metalloproteinases (MMPs)-2 and MMP-3 expression as well as promoting cell migration. MicroRNA (miRNA) analysis of CTGF-overexpressed osteosarcoma versus control cells probed mechanisms of CTGF-mediated promotion of migration. Among miRNAs regulated by CTGF, miR-519d was most downregulated after CTGF treatment. Co-transfection with miR-519d mimic reversed CTGF-mediated MMPs expression and cell migration. Also, MEK and ERK inhibitors or mutants reduced CTGF-increased cell migration and miR-519d suppression. By contrast, knockdown of CTGF diminished lung metastasis in vivo. Clinical samples indicate CTGF expression as linked with clinical stage and tumor metastasis. Taken together, data show CTGF elevating MMPs expression and subsequently promoting tumor metastasis in human osteosarcoma, down-regulating miR-519d via MEK and ERK pathways, making CTGF a new molecular therapeutic target in osteosarcoma metastasis. PMID:25003330

  8. Connective-Tissue Growth Factor (CTGF/CCN2 Induces Astrogenesis and Fibronectin Expression of Embryonic Neural Cells In Vitro.

    Directory of Open Access Journals (Sweden)

    Fabio A Mendes

    Full Text Available Connective-tissue growth factor (CTGF is a modular secreted protein implicated in multiple cellular events such as chondrogenesis, skeletogenesis, angiogenesis and wound healing. CTGF contains four different structural modules. This modular organization is characteristic of members of the CCN family. The acronym was derived from the first three members discovered, cysteine-rich 61 (CYR61, CTGF and nephroblastoma overexpressed (NOV. CTGF is implicated as a mediator of important cell processes such as adhesion, migration, proliferation and differentiation. Extensive data have shown that CTGF interacts particularly with the TGFβ, WNT and MAPK signaling pathways. The capacity of CTGF to interact with different growth factors lends it an important role during early and late development, especially in the anterior region of the embryo. ctgf knockout mice have several cranio-facial defects, and the skeletal system is also greatly affected due to an impairment of the vascular-system development during chondrogenesis. This study, for the first time, indicated that CTGF is a potent inductor of gliogenesis during development. Our results showed that in vitro addition of recombinant CTGF protein to an embryonic mouse neural precursor cell culture increased the number of GFAP- and GFAP/Nestin-positive cells. Surprisingly, CTGF also increased the number of Sox2-positive cells. Moreover, this induction seemed not to involve cell proliferation. In addition, exogenous CTGF activated p44/42 but not p38 or JNK MAPK signaling, and increased the expression and deposition of the fibronectin extracellular matrix protein. Finally, CTGF was also able to induce GFAP as well as Nestin expression in a human malignant glioma stem cell line, suggesting a possible role in the differentiation process of gliomas. These results implicate ctgf as a key gene for astrogenesis during development, and suggest that its mechanism may involve activation of p44/42 MAPK signaling

  9. Inhibition of connective tissue growth factor (CTGF/CCN2) expression decreases the survival and myogenic differentiation of human rhabdomyosarcoma cells.

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    Croci, Stefania; Landuzzi, Lorena; Astolfi, Annalisa; Nicoletti, Giordano; Rosolen, Angelo; Sartori, Francesca; Follo, Matilde Y; Oliver, Noelynn; De Giovanni, Carla; Nanni, Patrizia; Lollini, Pier-Luigi

    2004-03-01

    Connective tissue growth factor (CTGF/CCN2), a cysteine-rich protein of the CCN (Cyr61, CTGF, Nov) family of genes, emerged from a microarray screen of genes expressed by human rhabdomyosarcoma cells. Rhabdomyosarcoma is a soft tissue sarcoma of childhood deriving from skeletal muscle cells. In this study, we investigated the role of CTGF in rhabdomyosarcoma. Human rhabdomyosarcoma cells of the embryonal (RD/12, RD/18, CCA) and the alveolar histotype (RMZ-RC2, SJ-RH4, SJ-RH30), rhabdomyosarcoma tumor specimens, and normal skeletal muscle cells expressed CTGF. To determine the function of CTGF, we treated rhabdomyosarcoma cells with a CTGF antisense oligonucleotide or with a CTGF small interfering RNA (siRNA). Both treatments inhibited rhabdomyosarcoma cell growth, suggesting the existence of a new autocrine loop based on CTGF. CTGF antisense oligonucleotide-mediated growth inhibition was specifically due to a significant increase in apoptosis, whereas cell proliferation was unchanged. CTGF antisense oligonucleotide induced a strong decrease in the level of myogenic differentiation of rhabdomyosarcoma cells, whereas the addition of recombinant CTGF significantly increased the proportion of myosin-positive cells. CTGF emerges as a survival and differentiation factor and could be a new therapeutic target in human rhabdomyosarcoma.

  10. Activation of the connective tissue growth factor (CTGF-transforming growth factor β 1 (TGF-β 1 axis in hepatitis C virus-expressing hepatocytes.

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    Tirumuru Nagaraja

    Full Text Available BACKGROUND: The pro-fibrogenic cytokine connective tissue growth factor (CTGF plays an important role in the development and progression of fibrosis in many organ systems, including liver. However, its role in the pathogenesis of hepatitis C virus (HCV-induced liver fibrosis remains unclear. METHODS: In the present study, we assessed CTGF expression in HCV-infected hepatocytes using replicon cells containing full-length HCV genotype 1 and the infectious HCV clone JFH1 (HCV genotype 2 by real-time PCR, Western blot analysis and confocal microscopy. We evaluated transforming growth factor β1 (TGF-β1 as a key upstream mediator of CTGF production using neutralizing antibodies and shRNAs. We also determined the signaling molecules involved in CTGF production using various immunological techniques. RESULTS: We demonstrated an enhanced expression of CTGF in two independent models of HCV infection. We also demonstrated that HCV induced CTGF expression in a TGF-β1-dependent manner. Further dissection of the molecular mechanisms revealed that CTGF production was mediated through sequential activation of MAPkinase and Smad-dependent pathways. Finally, to determine whether CTGF regulates fibrosis, we showed that shRNA-mediated knock-down of CTGF resulted in reduced expression of fibrotic markers in HCV replicon cells. CONCLUSION: Our studies demonstrate a central role for CTGF expression in HCV-induced liver fibrosis and highlight the potential value of developing CTGF-based anti-fibrotic therapies to counter HCV-induced liver damage.

  11. Antisense CTGF inhibits the expression of CTGF in hypertrophic scar of rabbit ears%药物抑制兔耳病理性瘢痕中结缔组织生长因子的实验

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    王少华; 李健; 王燕华; 吕建平; 李记森; 王勇

    2011-01-01

    Objective To investigate a medicine which can inhibit the expression of connective tissue growth factor (CTGF) in hypertrophic scar of rabbit ears. Methods 24 bigears white rabbits were used to establish a model of hypertrophic scar of rabbit ears, which was randomly divided into four groups. The hypertrophic scar was injected intralesionally with antisense CTGF (group A), betamethason (group B), triamcinolone acetonide (group C) and physiological saline (group D). Some scar tissue samples were sectioned in every group when the scar was treated after 7, 14, 30, and 60 days, respectively. The expression of CTGF mRNA in the scar was assessed by in situ hybridization and hematoxylin and eosin stain (HE stain) in every samples. Results The expression of CTGF mRNA and the counting of fibroblasts decreased in group A, which showed statistical difference as compared with groups B, C and D. Conclusions The results suggesz that antisense CTGF is able to inhibit the proliferation process of hypertrophic scar in rabbit ears and to remarkably decrease the degree of fiborsis in the scar.%目的 探讨能有效抑制结缔组织生长因子(CTGF)的药物,以便为治疗病理性瘢痕提供依据.方法 选择24只大耳白兔建立兔耳病理性瘢痕模型,随机分为4组:A组注射CTGF反义寡核苷酸,B组注射复方倍他米松,C组注射醋酸曲安奈得,D组为对照组,仅注射生理盐水.通过原位杂交法分别检测瘢痕组织中不同治疗组不同时段的CTGF表达,并通过苏木精-伊红(HE)染色法检测瘢痕组织中不同治疗组不同时段的成纤维细胞数.结果 A、B、C 3组在同一时段其CTGF表达均比D组低,差异有统计学意义(P<0.05),A组较B、C两组CTGF表达低,差异也具有统计学意义(P<0.05).B、C两组之间CTGF表达差异无统计学意义(P>0.05).成纤维细胞计数结果与以上结果基本一致.结论 CTGF反义寡核苷酸、复方倍他米松、醋酸曲安奈得均可抑制病理性瘢痕

  12. The Expression and the Mechanism of Enhanced Anti-apoptosis of CTGF in Human Esophageal Cancer Cell Line Eca109%CTGF 在食管癌组织中表达及其促进凋亡抵抗的作用机制

    Institute of Scientific and Technical Information of China (English)

    黄芳

    2014-01-01

    目的:研究CTGF在人食管癌细胞中的表达情况及其对人食管癌细胞系Eca109细胞凋亡的影响和可能机制。方法应用Real Time PCR获取编码CTGF全长的cDNA,经免疫组织化学染色,经Western Blot检测食管癌中CT-GF mRNA及蛋白的表达情况;应用pcDNA3.1质粒和慢病毒载体分别上调和下调Eca109细胞中CTGF的表达,应用流式细胞仪检测细胞凋亡比率;Western Blot检测凋亡相关蛋白BCl-XL 表达。结果与正常组织比较,食管癌组织中CT-GF mRNA及蛋白的表达水平均明显增高,免疫组化结果显示食管癌标本中CTGF显色较正常对照组织强。转染pcD-NA3.1 CTGF质粒后,有效增强了Eca109细胞中CTGF蛋白的表达和凋亡的抵抗作用,且抗凋亡蛋白BCl-XL 表达水平上调。用RNAi逆转录病毒转染后,有效下调了内源CTGF的表达,对INF-γ介导的凋亡抵抗作用较对照组下降,与对照组相比,BCl-XL 蛋白表达水平亦下调。结论在食管癌中CTGF表达水平明显增高,并且可以通过上调抗凋亡蛋白BCl-XL 表达从而抑制细胞凋亡。%Objective To observe the CTGF expression in the sample of ESCC and study the effects and mechanism of CTGF on cell apoptosis of Eca 109 cell.Methods Application of Real Time PCR to obtain full-length cDNA encoding CTGF ,af-ter immunohistochemical staining ,the expression of CTGF Western Blot for the detection of esophageal cancer specimens in mR -NA and protein;the application of pcDNA3.1 plasmid and lentiviral vectors were up-regulated and down regulated CTGF expres-sion in Eca109 cells,flow cytometry was used to detect the expression of apoptosis ratio;Western Blot for the detection of apoptosis related protein BCl-XL .Results Compared with the control ,the expression level of CTGF mRNA and protein in ESCC were sig-nificant higher .In cancerous tissues ,CTGF immunoreactivity was more intense than in corresponding normal tissues .The expres-sion of

  13. Evaluation of genistein ability to modulate CTGF mRNA/protein expression, genes expression of TGFβ isoforms and expression of selected genes regulating cell cycle in keloid fibroblasts in vitro.

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    Jurzak, Magdalena; Adamczyk, Katarzyna; Antończak, Paweł; Garncarczyk, Agnieszka; Kuśmierz, Dariusz; Latocha, Małgorzata

    2014-01-01

    RT-QPCR was used to estimate transcription level of selected genes in normal and keloid fibroblasts treated with genistein. Secreted/cell-associated CTGF protein was evaluated in cell growth's medium by ELISA. Total protein quantification was evaluated by fluorimetric assay in cells llsates (Quant-iT TM Protein Assay Kit). It was found that TGFβ1, β2 and β3 genes expression are decreased by genistein. Genistein suppresses the expression of CTGF mRNA and CTGF protein in a concentration dependent manner, p53 and p21 genes expression are modulated by genistein in concentration dependent manner. The agent also modulates BAX/BCL-2 ratio in examined cells in vitro.

  14. Growth differentiation factor 8 suppresses cell proliferation by up-regulating CTGF expression in human granulosa cells.

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    Chang, Hsun-Ming; Pan, Hui-Hui; Cheng, Jung-Chien; Zhu, Yi-Min; Leung, Peter C K

    2016-02-15

    Connective tissue growth factor (CTGF) is a matricellular protein that plays a critical role in the development of ovarian follicles. Growth differentiation factor 8 (GDF8) is mainly, but not exclusively, expressed in the mammalian musculoskeletal system and is a potent negative regulator of skeletal muscle growth. The aim of this study was to investigate the effects of GDF8 and CTGF on the regulation of cell proliferation in human granulosa cells and to examine its underlying molecular determinants. Using dual inhibition approaches (inhibitors and small interfering RNAs), we have demonstrated that GDF8 induces the up-regulation of CTGF expression through the activin receptor-like kinase (ALK)4/5-mediated SMAD2/3-dependent signaling pathways. In addition, the increase in CTGF expression contributes to the GDF8-induced suppressive effect on granulosa cell proliferation. Our findings suggest that GDF8 and CTGF may play critical roles in the regulation of proliferative events in human granulosa cells. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  15. Inverse expression of cystein-rich 61 (Cyr61/CCN1) and connective tissue growth factor (CTGF/CCN2) in borderline tumors and carcinomas of the ovary.

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    Bartel, Frank; Balschun, Katharina; Gradhand, Elise; Strauss, Hans G; Dittmer, Jürgen; Hauptmann, Steffen

    2012-09-01

    Members of the CCN [cystein-rich 61 (Cyr61)/connective tissue growth factor (CTGF)/nephroblastoma (NOV)] protein family are involved in the regulation of cellular proliferation, apoptosis, and migration and are also assumed to play a role in carcinogenesis. Therefore, we performed a retrospective study to investigate the immunohistochemical expression of both Cyr61 and CTGF in 92 borderline tumors (BOTs) and 107 invasive carcinomas of the ovary (IOCs). To determine their diagnostic and prognostic value, we correlated protein expression with clinicopathologic factors including overall and disease-free survival. Cyr61 and CTGF were found to be inversely expressed in both BOTs and IOCs, with a stronger expression of Cyr61 in IOCs. Moreover, Cyr61 was found to be preferentially expressed in high-grade serous carcinomas, whereas CTGF was found more frequently in low-grade serous carcinomas. Weak Cyr61 levels correlated with both low estrogen receptor and p53 expression (P=0.038, P=0.04, respectively). However, no association was observed between CTGF, estrogen receptor, and p53 expression levels in IOCs. Regarding prognosis, Cyr61 was found to be of no value, but the loss of CTGF was found to be associated with a poor prognosis in multivariate analysis of overall (relative risk 2.8; P=0.050) and disease-free (relative risk 2.3; P=0.031) survival. Cyr61 and CTGF are inversely expressed in BOTs and IOCs, and loss of CTGF independently indicates poor prognosis in IOCs.

  16. PRS-CTGF-siRNA attenuates matrix production and the expression of vascular endothelial growth factors induced by transforming growth factor β1 in cultured human peritoneal mesothelial cells%PRS-CTGF-siRNA对TGF-β1诱导的人腹膜间皮细胞细胞外基质及VEGF表达的影响

    Institute of Scientific and Technical Information of China (English)

    肖力; 刘伏友; 彭佑铭; 段绍斌; 刘虹; 刘映红; 凌光辉

    2008-01-01

    .Results Both mRNA and protein expressions of CTGF,FN,Col I,laminin (LN) and VEGF were significantly increased in HPMC with 5 μg/L TGF-β1 stimulation (P<0.01,respectively).CTGF,FN,Col I,LN mRNA and protein and VEGF mRNA expression stimulated by TGF-β1 were significantly decreased in HPMC infected with PRS-CTGF-siRNA1~4 retrovirus vectors (P<0.01,respectively).The inhibitory rates on CTGF were 69.3%,22.2%,27.4% and 38.8%,respectively (P<0.01).At the same time,there was also a significant reduction of VEGF protein expression in HPMC infected with PRS-CTGF-siRNA1 vector (P<0.01).There was no significant difference in HPMC infected with PRS void vector. Conclusion CTGF siRNA delivered by PRS retrovirus vector can effectively inhibit the enhancement of extracellular matrix and VEGF expression stimulated by TGF-β1 in HPMC.

  17. [Influence of ASODN to the human tenon's fibroblasts in expressing CTGF induced by transforming growth factor beta2].

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    Hu, Yi-Zhen; Wang, Yu-Hong; Cao, Yang; Zhang, Ming-Chang

    2008-02-01

    To investigate the effect of connective tissue growth factor's antisense oligonucleotides (ASODN) on the growth of human tenon' s capsule fibroblasts (HTF) induced by transforming growth factor beta2 (TGF-beta2) in vitro. It was a experimental study. HTF was collected from glaucoma patients and cultured. The 5-6 passage was used for experiments. The HTF induced by TGF-beta2 was divided into the following groups: N group: normal HTF; T group: HTF induced by TGF-beta2; A group: CTGF ASODN antisense:5'-TACTGGCGGCGGTCAT-3' encapsulated with liposome; S group: sense 5'-ATGACCGCCGCCAGTA-3' encapsulated with liposome; D group: HTF encapsulated with liposome only. The activity of HTF treated by different concentrations of liposome was detected using methylthianolyldiphenyl tetrazolium bromide (MT) colorimetry. The expression of CTGF was detected by reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry assays. The expression of fibronectin (Fn) was examined by Western blot and immunocytochemistry assays. Liposome-ASODN (A group) significantly (F=15.25, 204.88, 19.73, 90.00; P HTF induced by TGF-beta2 compared with S and D group. However, Liposome alone (T group) has no significant impact in HTF growth compared with T group (t = 0.90, 2.32, 0.75, 2.11; P > 0.05). CTGF-ASODN inhibits the CTGF and Fn expression of HTF induced by TGF-beta2, which may delay the formation of scar in glaucoma filtering surgery.

  18. Angiotensin II increases CTGF expression via MAPKs/TGF-{beta}1/TRAF6 pathway in atrial fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Jun [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Liu, Xu, E-mail: xkliuxu@yahoo.cn [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Wang, Quan-xing, E-mail: shmywqx@126.com [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Tan, Hong-wei [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China); Guo, Meng [National Key Laboratory of Medical Immunology, Second Military Medical University, Shanghai (China); Jiang, Wei-feng; Zhou, Li [Department of Cardiology, Shanghai Chest Hospital, Shanghai Jiaotong University School of medicine, Shanghai (China)

    2012-10-01

    The activation of transforming growth factor-{beta}1(TGF-{beta}1)/Smad signaling pathway and increased expression of connective tissue growth factor (CTGF) induced by angiotensin II (AngII) have been proposed as a mechanism for atrial fibrosis. However, whether TGF{beta}1/non-Smad signaling pathways involved in AngII-induced fibrogenetic factor expression remained unknown. Recently tumor necrosis factor receptor associated factor 6 (TRAF6)/TGF{beta}-associated kinase 1 (TAK1) has been shown to be crucial for the activation of TGF-{beta}1/non-Smad signaling pathways. In the present study, we explored the role of TGF-{beta}1/TRAF6 pathway in AngII-induced CTGF expression in cultured adult atrial fibroblasts. AngII (1 {mu}M) provoked the activation of P38 mitogen activated protein kinase (P38 MAPK), extracellular signal-regulated kinase 1/2(ERK1/2) and c-Jun NH(2)-terminal kinase (JNK). AngII (1 {mu}M) also promoted TGF{beta}1, TRAF6, CTGF expression and TAK1 phosphorylation, which were suppressed by angiotensin type I receptor antagonist (Losartan) as well as p38 MAPK inhibitor (SB202190), ERK1/2 inhibitor (PD98059) and JNK inhibitor (SP600125). Meanwhile, both TGF{beta}1 antibody and TRAF6 siRNA decreased the stimulatory effect of AngII on TRAF6, CTGF expression and TAK1 phosphorylation, which also attenuated AngII-induced atrial fibroblasts proliferation. In summary, the MAPKs/TGF{beta}1/TRAF6 pathway is an important signaling pathway in AngII-induced CTGF expression, and inhibition of TRAF6 may therefore represent a new target for reversing Ang II-induced atrial fibrosis. -- Highlights: Black-Right-Pointing-Pointer MAPKs/TGF{beta}1/TRAF6 participates in AngII-induced CTGF expression in atrial fibroblasts. Black-Right-Pointing-Pointer TGF{beta}1/TRAF6 participates in AngII-induced atrial fibroblasts proliferation. Black-Right-Pointing-Pointer TRAF6 may represent a new target for reversing Ang II-induced atrial fibrosis.

  19. Relationship between hepatic CTGF expression and routine blood tests at the time of liver transplantation for biliary atresia: hope or hype for a biomarker of hepatic fibrosis

    Directory of Open Access Journals (Sweden)

    Haafiz A

    2011-04-01

    Full Text Available Allah Haafiz1, Christian Farrington1, Joel Andres1, Saleem Islam21Hepatology and Liver Transplantation, Division of Pediatric Gastroenterology, Hepatology and Nutrition, 2Division of Pediatric Surgery, University of Florida College of Medicine, Gainesville, FL, USABackground: Progressive hepatic fibrosis (HF is a prominent feature of biliary atresia (BA, the most common indication for liver transplantation (LT in children. Despite its importance in BA, HF is not evaluated in routine patient care because the invasiveness of liver biopsy makes histologic monitoring of fibrosis unfeasible. Therefore, the identification of noninvasive markers to assess HF is desirable especially in children.Purpose: The main goal of this pilot project was to establish an investigational framework correlating hepatic expression of fibrogenic markers with routine blood tests in BA.Methods: Using liver explants from patients with BA (n = 26, immune-expression of connective tissue growth factor (CTGF, a key fibrogenic cytokine was determined using horseradish-labeled antibodies. Expression intensities of lobular (L-CTGF and portal (P-CTGF CTGF were determined by using ImageJ software. These CTGF intensities were correlated with blood tests performed at the time of LT. Correlation coefficients were determined for each blood test variable versus mean L-CTGF and P-CTGF expression intensities. A P-value of less than 0.05 was considered statistically significant.Results: All patients had end-stage liver disease and persistent cholestasis at the time of LT. Kendall tau (t rank correlation coefficient for L-CTGF and white blood cell (WBC was inversed (—0.52; P ≤ 0.02. Similar but statistically nonsignificant inverse relationships were noted between L-CTGF and prothrombin time (PT (—0.15; P ≤ 0.4, international normalized ratio (INR (—0.14; P ≤ 0.5, and platelet count (—0.36; P ≤ 0.09. Inversed (t rank correlation coefficients were also evident between P-CTGF

  20. Increased Expression of Connective Tissue Growth Factor (CTGF) in Multiple Organs After Exposure of Non-Human Primates (NHP) to Lethal Doses of Radiation.

    Science.gov (United States)

    Zhang, Pei; Cui, Wanchang; Hankey, Kim G; Gibbs, Allison M; Smith, Cassandra P; Taylor-Howell, Cheryl; Kearney, Sean R; MacVittie, Thomas J

    2015-11-01

    Exposure to sufficiently high doses of ionizing radiation is known to cause fibrosis in many different organs and tissues. Connective tissue growth factor (CTGF/CCN2), a member of the CCN family of matricellular proteins, plays an important role in the development of fibrosis in multiple organs. The aim of the present study was to quantify the gene and protein expression of CTGF in a variety of organs from non-human primates (NHP) that were previously exposed to potentially lethal doses of radiation. Tissues from non-irradiated NHP and NHP exposed to whole thoracic lung irradiation (WTLI) or partial-body irradiation with 5% bone marrow sparing (PBI/BM5) were examined by real-time quantitative reverse transcription PCR, western blot, and immunohistochemistry. Expression of CTGF was elevated in the lung tissues of NHP exposed to WTLI relative to the lung tissues of the non-irradiated NHP. Increased expression of CTGF was also observed in multiple organs from NHP exposed to PBI/BM5 compared to non-irradiated NHP; these included the lung, kidney, spleen, thymus, and liver. These irradiated organs also exhibited histological evidence of increased collagen deposition compared to the control tissues. There was significant correlation of CTGF expression with collagen deposition in the lung and spleen of NHP exposed to PBI/BM5. Significant correlations were observed between spleen and multiple organs on CTGF expression and collagen deposition, respectively, suggesting possible crosstalk between spleen and other organs. These data suggest that CTGF levels are increased in multiple organs after radiation exposure and that inflammatory cell infiltration may contribute to the elevated levels of CTGF in multiple organs.

  1. Combined expression of CTGF and tissue inhibitor of metalloprotease-1 promotes synthesis of proteoglycan and collagen type Ⅱ in rhesus monkey lumbar intervertebral disc cells in vitro

    Institute of Scientific and Technical Information of China (English)

    LIU Yong; KONG Jie; CHEN Bo-hua; HU You-gu

    2010-01-01

    Background Low back pain has emerged as a widespread disease often caused by intervertebral disc degeneration.This study aimed to establish an in vitro cell culture model of rhesus monkey lumbar intervertebral discs and to investigate the effect of combined connective tissue growth factor (CTGF) and tissue inhibitor of metalloprotease-1(TIMP-1) expression mediated by adeno-associated virus (AAV) on collagen type Ⅱ and proteoglycan levels.The purpose of these investigations was to explore potential methods for relieving the degeneration of lumbar intervertebral disc cells.Methods Rhesus monkey lumbar intervertebral disc nucleus pulposus cells (NPCs) were isolated by enzyme digestion,cultured, and transduced with rAAV2-CTGF-IRES-TIMP-1, rAAV2-CTGF, or rAAV2-TIMP-1 at a multiplicity of infection (MOl) of 106.The expression of collagen type Ⅱ and proteoglycan was measured using RT-PCR and Western blotting.The synthetic rate of proteoglycan was measured using 35S incorporation.Results Rhesus monkey lumbar intervertebral disc NPCs were transduced with rAAV2-CTGF-IRES-TIMP-1,rAAV2-CTGF, and rAAV2-TIMP-1 and the transduced genes were expressed and detected.Compared to the control,CTGF promoted the synthesis of collagen type Ⅱ and proteoglycan.TIMP-1 showed an enhancing effect on the expression of proteoglycan but no effect on collagen type Ⅱ.Expression of both genes in rhesus monkey lumbar intervertebral disc NPCs significantly enhances the synthesis of proteoglycan and collagen type Ⅱ.Conclusions Single gene transduction of CTGF or TIMP-1 can enhanced synthesis of proteoglycan.CTGF expression can also enhance collagen type Ⅱ protein synthesis.Combined transduction of both CTGF and TIMP1 can significantly promote the expression of proteoglycan and collagen type Ⅱ to levels greater than transduction of a single gene alone.Our study provides a good basis for multi-gene therapy to treat lumbar intervertebral disc degeneration.

  2. SOX9 directly Regulates CTGF/CCN2 Transcription in Growth Plate Chondrocytes and in Nucleus Pulposus Cells of Intervertebral Disc.

    Science.gov (United States)

    Oh, Chun-do; Yasuda, Hideyo; Zhao, Weiwei; Henry, Stephen P; Zhang, Zhaoping; Xue, Ming; de Crombrugghe, Benoit; Chen, Di

    2016-07-20

    Several lines of evidence indicate that connective tissue growth factor (CTGF/CCN2) stimulates chondrocyte proliferation and maturation. Given the fact that SOX9 is essential for several steps of the chondrocyte differentiation pathway, we asked whether Ctgf (Ccn2) is the direct target gene of SOX9. We found that Ctgf mRNA was down-regulated in primary sternal chondrocytes from Sox9(flox/flox) mice infected with Ad-CMV-Cre. We performed ChIP-on-chip assay using anti-SOX9 antibody, covering the Ctgf gene from 15 kb upstream of its 5'-end to 10 kb downstream of its 3'-end to determine SOX9 interaction site. One high-affinity interaction site was identified in the Ctgf proximal promoter by ChIP-on-chip assay. An important SOX9 regulatory element was found to be located in -70/-64 region of the Ctgf promoter. We found the same site for SOX9 binding to the Ctgf promoter in nucleus pulposus (NP) cells. The loss of Sox9 in growth plate chondrocytes in knee joint and in NP cells in intervertebral disc led to the decrease in CTGF expression. We suggest that Ctgf is the direct target gene of SOX9 in chondrocytes and NP cells. Our study establishes a strong link between two regulatory molecules that have a major role in cartilaginous tissues.

  3. Simvastatin inhibits transforming growth factor-β1-induced expression of type I collagen, CTGF, and α-SMA in keloid fibroblasts.

    Science.gov (United States)

    Mun, Je-Ho; Kim, Young-Mi; Kim, Byung-Soo; Kim, Jae-Ho; Kim, Moon-Bum; Ko, Hyun-Chang

    2014-01-01

    Simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme-A reductase inhibitor, is used to reduce cholesterol levels. Accumulating evidence has revealed the immunomodulatory and anti-inflammatory effects of simvastatin that prevent cardiovascular diseases. In addition, the beneficial effects of statins on fibrosis of various organs have been reported. However, the functional effect of statins on dermal fibrosis of keloids has not yet been explored. The objective of this study was to determine whether simvastatin could affect dermal fibrosis associated with keloids. We examined the effect of simvastatin on transforming growth factor (TGF)-β1-induced production of type I collagen, connective tissue growth factor (CTGF or CCN2), and α-smooth muscle actin (α-SMA). Keloid fibroblasts were cultured and exposed to different concentrations of simvastatin in the presence of TGF-β1, and the effects of simvastatin on TGF-β1-induced collagen and CTGF production in keloid fibroblasts were determined. The type I collagen, CTGF, and α-SMA expression levels and the Smad2 and Smad3 phosphorylation levels were assessed by Western blotting. The effect of simvastatin on cell viability was evaluated by assessing the colorimetric conversion of 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide. Simvastatin suppressed TGF-β1-induced type I collagen, CTGF, and α-SMA production in a concentration-dependent manner. The TGF-β1-induced Smad2 and Smad3 phosphorylation levels were abrogated by simvastatin pretreatment. The inhibition of type I collagen, CTGF, and α-SMA expression by simvastatin was reversed by geranylgeranyl pyrophosphate, suggesting that the simvastatin-induced cellular responses were due to inhibition of small GTPase Rho involvement. A RhoA activation assay showed that preincubation with simvastatin significantly blocked TGF-β1-induced RhoA activation. The Rho-associated coiled kinase inhibitor Y27632 abrogated TGF-β1-induced production of type I collagen

  4. Increased Epithelial Expression of CTGF and S100A7 with Elevated Subepithelial Expression of IL-1β in Trachomatous Trichiasis.

    Directory of Open Access Journals (Sweden)

    Tamsyn Derrick

    2016-06-01

    Full Text Available To characterize the histological appearance and expression of pro-inflammatory mediators, growth factors, matrix metalloproteinases and biomarkers of epithelial-mesenchymal transition (EMT in healthy control and trachomatous trichiasis (TT conjunctival tissue.Conjunctival biopsies were taken from 20 individuals with TT and from 16 individuals with healthy conjunctiva, which served as controls. Study participants were of varying ethnicity and were living in a trachoma-endemic region of northern Tanzania. Formalin-fixed paraffin-embedded tissue sections were stained using hematoxylin and eosin or by immunohistochemistry using antibodies against: IL-1β, IL-6, IL-17A, IL-22, CXCL5, S100A7, cleaved caspase 1 (CC1, PDGF, CTGF, TGFβ2, MMP7, MMP9, E-cadherin, vimentin, and αSMA.Tissue from TT cases had a greater inflammatory cell infiltrate relative to controls and greater disruption of collagen structure. CTGF and S100A7 were more highly expressed in the epithelium and IL-1β was more highly expressed in the substantia propria of TT cases relative to controls. Latent TGFβ2 was slightly more abundant in the substantia propria of control tissue. No differences were detected between TT cases and controls in the degree of epithelial atrophy, the number of myofibroblasts or expression of EMT biomarkers.These data indicate that the innate immune system is active in the immunopathology of trachoma, even in the absence of clinical inflammation. CTGF might provide a direct link between inflammation and fibrosis and could be a suitable target for therapeutic treatment to halt the progression of trachomatous scarring.

  5. Novel angiogenic inhibitor DN-9693 that inhibits post-transcriptional induction of connective tissue growth factor (CTGF/CCN2) by vascular endothelial growth factor in human endothelial cells.

    Science.gov (United States)

    Kondo, Seiji; Tanaka, Noriko; Kubota, Satoshi; Mukudai, Yoshiki; Yosimichi, Gen; Sugahara, Toshio; Takigawa, Masaharu

    2006-01-01

    Connective tissue growth factor (CTGF/CCN2) is a potent angiogenic factor. In this report, we describe for the first time that vascular endothelial growth factor (VEGF)-mediated induction of the ctgf/ccn2 gene was a post-transcriptional event that was inhibited by a novel angiogenic inhibitor, DN-9693, in human umbilical vein endothelial cells. Steady-state mRNA levels of ctgf/ccn2 were remarkably increased by VEGF in a concentration-dependent manner, whereas the activity of the ctgf/ccn2 promoter was not responsive to VEGF as confirmed by a reporter gene assay and quantitative real-time PCR analysis. By employing a RNA degradation assay, we eventually found that the observed increase in the ctgf/ccn2 mRNA level was due to an increased stability of the mRNA induced by VEGF. DN-9693 at a dose of 0.1 to 2 ng/mL did not affect basal levels of ctgf/ccn2 mRNA; however, enhancement of ctgf/ccn2 mRNA expression by VEGF was specifically inhibited by DN-9693. Of importance, the inhibitory effects could be also ascribed to post-transcriptional regulation, because the VEGF-mediated increase in stability of ctgf/ccn2 mRNA was suppressed by DN-9693. Furthermore, we investigated the effects of DN-9693 on VEGF-induced activation of three subgroups of mitogen-activated protein kinase pathways and found that DN-9693 blocked the activation of these pathways by VEGF. These results suggest that VEGF increases ctgf/ccn2 mRNA stability through mitogen-activated protein kinase-mediated intracellular signaling cascade(s), which can be inhibited posttranscriptionally by a novel angiogenic inhibitor, DN-9693, in human umbilical vein endothelial cells.

  6. Expression of TGF-β1 and CTGF Is Associated with Fibrosis of Denervated Sternocleidomastoid Muscles in Mice.

    Science.gov (United States)

    Liu, Fei; Tang, Weifang; Chen, Donghui; Li, Meng; Gao, Yinna; Zheng, Hongliang; Chen, Shicai

    2016-01-01

    Injury to the recurrent laryngeal nerve often leads to permanent vocal cord paralysis, which has a significant negative impact on the quality of life. Long-term denervation can induce laryngeal muscle fibrosis, which obstructs the muscle recovery after laryngeal reinnervation. However, the mechanisms of fibrosis remain unclear. In this study, we aimed to analyze the changes in the expression of fibrosis-related factors, including transforming growth factor-β1 (TGF-β1), connective tissue growth factor (CTGF), and α-smooth muscle actin (α-SMA) in denervated skeletal muscles using a mouse model of accessory nerve transection. Because of the small size, we used sternocleidomastoid muscles instead of laryngeal muscles for denervation experiments. Masson's trichrome staining showed that the grade of atrophy and fibrosis of muscles became more severe with time, but showed a plateau at 4 weeks after denervation, followed by a slow decrease. Quantitative assessment and immunohistochemistry showed that TGF-β1 expression peaked at 1 week after denervation (p muscle cells were detected at 1 week after denervation, peaked at 2 weeks (p muscle fibrosis. They may induce the differentiation of myoblasts into myofibroblasts, as characterized by the activation of α-SMA. These findings may provide insights on key pathological processes in denervated skeletal muscle fibrosis and develop novel therapeutic strategies.

  7. Antiproliferative factor regulates connective tissue growth factor (CTGF/CCN2) expression in T24 bladder carcinoma cells

    Science.gov (United States)

    Matika, Christina A.; Wasilewski, Melissa; Arnott, John A.; Planey, Sonia Lobo

    2012-01-01

    Antiproliferative factor (APF) is a sialoglycopeptide elevated in the urine of patients with interstitial cystitis (IC)—a chronic, painful bladder disease of unknown etiology. APF inhibits the proliferation of normal bladder epithelial and T24 bladder carcinoma cells in vitro by binding to cytoskeleton-associated protein 4 (CKAP4) and altering the transcription of genes involved in proliferation, cellular adhesion, and tumorigenesis; however, specific molecular mechanisms and effector genes that control APF's antiproliferative effects are unknown. In this study, we found that there was a 7.5-fold up-regulation of connective tissue growth factor (CTGF/CCN2) expression in T24 bladder carcinoma cells treated with APF. Western blot revealed a dose-dependent increase in CCN2 protein levels, with secretion into the culture medium after APF treatment. CCN2 overexpression enhanced APF's antiproliferative activity, whereas CCN2 knockdown diminished APF-induced p53 expression. Using a luciferase reporter construct, we found that APF treatment resulted in fivefold activation of the CCN2 proximal promoter and, of importance, that small interfering RNA–mediated knockdown of CKAP4 inhibited CCN2 upregulation. In addition, we demonstrate that CKAP4 translocates to the nucleus and binds to the CCN2 proximal promoter in an APF-dependent manner, providing evidence that CCN2 regulation by APF involves CKAP4 nuclear translocation and binding to the CCN2 promoter. PMID:22438586

  8. Cyclical cell stretching of skin-derived fibroblasts downregulates connective tissue growth factor (CTGF) production.

    Science.gov (United States)

    Kanazawa, Yuichiro; Nomura, Jun; Yoshimoto, Shinya; Suzuki, Toshikazu; Kita, Kazuko; Suzuki, Nobuo; Ichinose, Masaharu

    2009-01-01

    Delayed healing of skin wounds can be caused by wound instability, whereas appropriate massage or exercise prevents sclerosis and scar contracture. However, the mechanism by which wound healing is related to mechanical stress has not been fully elucidated. The present study aimed to identify whether mechanical stretching of fibroblasts reduces their production of extracellular matrix. We transferred skin fibroblasts into collagen-coated elastic silicone chambers, cultured them on a stretching apparatus, and used RT-PCR to examine the effects of mechanical stretching on the expression levels of 17 genes related to extracellular matrix production and growth factor secretion. We found that connective tissue growth factor (CTGF) was downregulated after 24 hr of cell stretching. Specifically, the CTGF mRNA and protein levels were 50% and 48% of the control levels, respectively. These findings suggest that cyclic stretching of fibroblasts contributes to anti-fibrotic processes by reducing CTGF production.

  9. The role of CTGF in diabetic retinopathy.

    Science.gov (United States)

    Klaassen, Ingeborg; van Geest, Rob J; Kuiper, Esther J; van Noorden, Cornelis J F; Schlingemann, Reinier O

    2015-04-01

    Connective tissue growth factor (CTGF, CCN2) contributes to fibrotic responses in diabetic retinopathy, both before clinical manifestations occur in the pre-clinical stage of diabetic retinopathy (PCDR) and in proliferative diabetic retinopathy (PDR), the late clinical stage of the disease. CTGF is a secreted protein that modulates the actions of many growth factors and extracellular matrix (ECM) proteins, leading to tissue reorganization, such as ECM formation and remodeling, basal lamina (BL) thickening, pericyte apoptosis, angiogenesis, wound healing and fibrosis. In PCDR, CTGF contributes to thickening of the retinal capillary BL and is involved in loss of pericytes. In this stage, CTGF expression is induced by advanced glycation end products, and by growth factors such as vascular endothelial growth factor (VEGF) and transforming growth factor (TGF)-β. In PDR, the switch from neovascularization to a fibrotic phase - the angio-fibrotic switch - in PDR is driven by CTGF, in a critical balance with vascular endothelial growth factor (VEGF). We discuss here the roles of CTGF in the pathogenesis of DR in relation to ECM remodeling and wound healing mechanisms, and explore whether CTGF may be a potential novel therapeutic target in the clinical management of early as well as late stages of DR.

  10. Expression and clinical significance of NF-κB, CTGF and OPN in mononuclear cells in peripheral blood as well as renal tissues in patients with IgA nephropathy

    Institute of Scientific and Technical Information of China (English)

    Cheng-Luo Hao; Chang-Bin Liao

    2016-01-01

    Objective:To study the expression and clinical significance of NF-κB, CTGF and OPN in mononuclear cells in peripheral blood as well as renal tissues in patients with IgA nephropathy. Methods:A total of 25 nephropathy patients diagnosed with IgA nephropathy and 25 patients receiving nephrectomy due to trauma or tumor in our hospital were studied. Peripheral blood and kidney tissues were collected to test NF-κB, CTGF, OPN, T-bet, GATA-3, RORγT and Foxp3 expressions.Results:CTGF and OPN percentages in peripheral blood mononuclear cells and kidney tissues of nephropathy patients were higher than those of the control group. NF-κB, CTGF and OPN expressions were significantly higher in M1, E1, S1 group patients’ peripheral blood mononuclear cells and renal tissues than those in M0, E1 and S1 group. T-bet, GATA-3 and RORγT expressions in nephropathy patients’ peripheral blood were significantly higher than those in the control group, and were positively correlated with NF-κB, CTGF and OPN expressions. The expression of Foxp3 was significantly lower than that of control group, and was negatively correlated with NF-κB, CTGF and OPN expressions.Conclusions: The expression of NF-κB, CTGF and OPN in peripheral blood mononuclear cells and renal tissue in patients with IgA nephropathy is abnormally high and can evaluate the prognosis of the disease and the differentiation of CD4+T cells.

  11. Construction of Retrovirus Expressing CTGF siRNA Mediated by PRS Retrovirus Vector%PRS载体介导的表达CTGF-SiRNA逆转录病毒载体的构建

    Institute of Scientific and Technical Information of China (English)

    肖力; 刘伏友; 彭佑铭; 袁芳; 陈俊香

    2006-01-01

    [目的]构建表达人结缔组织生长因子(CTGF)小分子干扰RNA(small interfering RNA,SiRNA)的PRS-CTGF-SiRNA逆转录病毒载体.[方法]根据SiRNA靶序列设计要求及PRS逆转录病毒载体特点分别设计含64 bp DNA序列的4对寡核苷酸,并在体外合成.PRS载体用BgⅡ、HindⅢ双酶切完全后,分别与退火的4对寡核苷酸进行连接,连接后去载体自连,构建成表达人CTGF小分子干扰RNA的PRS-CTGF-SiRNA逆转录病毒载体.[结果]经酶切、连接后构建成的质粒称之为PRS-CTGF-SiRNA1~4,经酶切与测序证实构建成功,无任何碱基突变.[结论]成功构建了能表达人CTGF-SiRNA的PRS-CTGF-SiRNA逆转录病毒载体,为腹膜透析腹膜纤维化防治的体内、外研究提供一种可能有效的方法.

  12. Locally expressed IGF1 propeptide improves mouse heart function in induced dilated cardiomyopathy by blocking myocardial fibrosis and SRF-dependent CTGF induction

    Directory of Open Access Journals (Sweden)

    Melissa Touvron

    2012-07-01

    Cardiac fibrosis is critically involved in the adverse remodeling accompanying dilated cardiomyopathies (DCMs, which leads to cardiac dysfunction and heart failure (HF. Connective tissue growth factor (CTGF, a profibrotic cytokine, plays a key role in this deleterious process. Some beneficial effects of IGF1 on cardiomyopathy have been described, but its potential role in improving DCM is less well characterized. We investigated the consequences of expressing a cardiac-specific transgene encoding locally acting IGF1 propeptide (muscle-produced IGF1; mIGF1 on disease progression in a mouse model of DCM [cardiac-specific and inducible serum response factor (SRF gene disruption] that mimics some forms of human DCM. Cardiac-specific mIGF1 expression substantially extended the lifespan of SRF mutant mice, markedly improved cardiac functions, and delayed both DCM and HF. These protective effects were accompanied by an overall improvement in cardiomyocyte architecture and a massive reduction of myocardial fibrosis with a concomitant amelioration of inflammation. At least some of the beneficial effects of mIGF1 transgene expression were due to mIGF1 counteracting the strong increase in CTGF expression within cardiomyocytes caused by SRF deficiency, resulting in the blockade of fibroblast proliferation and related myocardial fibrosis. These findings demonstrate that SRF plays a key role in the modulation of cardiac fibrosis through repression of cardiomyocyte CTGF expression in a paracrine fashion. They also explain how impaired SRF function observed in human HF promotes fibrosis and adverse cardiac remodeling. Locally acting mIGF1 efficiently protects the myocardium from these adverse processes, and might thus represent a therapeutic avenue to counter DCM.

  13. Endothelin-1 induces connective tissue growth factor expression in cardiomyocytes.

    Science.gov (United States)

    Recchia, Anna Grazia; Filice, Elisabetta; Pellegrino, Daniela; Dobrina, Aldo; Cerra, Maria Carmela; Maggiolini, Marcello

    2009-03-01

    Endothelin (ET)-1 is a vasoconstrictor involved in cardiovascular diseases. Connective tissue growth factor/CCN2 (CTGF) is a fibrotic mediator overexpressed in human atherosclerotic lesions, myocardial infarction, and hypertension. In different cell types CTGF regulates cell proliferation/apoptosis, migration, and extracellular matrix (ECM) accumulation and plays important roles in angiogenesis, chondrogenesis, osteogenesis, tissue repair, cancer and fibrosis. In the present study, we investigated the ET-1 signaling which triggers CTGF expression in cultured adult mouse atrial-muscle HL-1 cells used as a model system. ET-1 activated the CTGF promoter and induced CTGF expression at both mRNA and protein levels. Real-time PCR analysis revealed CTGF induction also in isolated rat heart preparations perfused with ET-1. Several intracellular signals elicited by ET-1 via ET receptors and even Epidermal Growth Factor Receptor (EGFR) contributed to the up-regulation of CTGF, including ERK activation and induction of the AP-1 components c-fos and c-jun, as also evaluated by ChIP analysis. Moreover, in cells treated with ET-1 the expression of ECM component decorin was abolished by CTGF silencing, indicating that CTGF is involved in ET-1 induced ECM accumulation not only in a direct manner but also through downstream effectors. Collectively, our data indicate that CTGF could be a mediator of the profibrotic effects of ET-1 in cardiomyocytes. CTGF inhibitors should be considered in setting a comprehensive pharmacological approach towards ET-1 induced cardiovascular diseases.

  14. Effects of matrine on collagen proliferation and TNF-α, TGF-β1 and CTGF in atrial tissues of dogs with persistent atrial fibrillation

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    You-ping Dai

    2013-12-01

    Full Text Available Objective:To study the effects of matrine (mat on collagen synthesis and expression of tumor necrosis factoralpha (TNF-α, and transforming growth factor-β1 (TGF-β1 and connective tissue growth factor (CTGF in atrial tissues of dogs with persistent atrial fibrillation (AF. Methods : Ten healthy beagle dogs were randomly divided into two groups: AF group (n=5 and AF/Mat group (n=5, using right ventricular pacing to establish AF model. The collagen volume fraction (CVF in atrial tissue were detected by sirius red staining to determine the level of fabrication. The level of TNF-α, TGF-β1 and CTGF were detected by immunohisto-chemistry. The mRNA expression level of TNF-α, TGF-β1 and CTGF were detected by reverse transcription-polymerase chain reaction (RT-PCR. Results:  Compared with the AF group, the fabriation level of AF/Mat was decreased obviously (P<0.05, the expression levels of TNF-α, TGF-β1 and CTGF were decreased, and the mRNA expression level were decreased significantly in atrial tissues (P<0.05 and P<0.01. Conclusion: Matrine may inhibits fabrosis in atrial tissues through inhibition collagen proliferation and expression of TNF-α, TGF-β1 and CTGF.

  15. Effect of gene CTGF transfection on the expression of MMP-2 and MMP-9 and proliferation in human cervical cancer cells%CTGF基因转染对宫颈癌细胞MMP-2、MMP-9表达及细胞增殖的影响

    Institute of Scientific and Technical Information of China (English)

    肖蔚; 焦霞; 钱华; 崔永安; 林梅; 王薇; 周彤敏; 窦荣荣; 于鸿

    2012-01-01

    Objective: To investigate the effect of connective tissue growth factor ( CTGF ) on the proliferation in human cervical cancer cell line Hela, with a focus on the expression of matrix metalloproteinase-2( MMP-2 ) and matrix metalloproteinase-9( MMP-9 ), and explore the underlying mechanism for the role of CTGF in the development of cervical cancer. Methods: pcDNA3. 0-CTGF and pcDNA3.0 were transfected into Hela cells through lipofectamine and positive clones which were screened by G418. Fluorescence quan-titive polymerase chain reaction( FQ-PCR ) and Western blot were employed to identify mRNA and protein expression of CTGF in Hela cells, respectively. The expression of MMP-2 and MMP-9 in positive clones was detected by FQ-PCR and Western blot. Cell viability was assessed by dimethylthiazoldiphenyl-tetrazolium-bromide ( MTT ) method. Results: Positive clone C-16 with CTGF over-expression were successfully established. Compared with non-transfected control group, the expression of MMP-2 and MMP-9 in C-16 were increased significantly,and the proliferation level of C-16 was increased significantly. Conclusion: CTGF transfection could effectively enhance the expression of MMP-2 and MMP-9 and the proliferation in Hela cells which suggested a potential role for CTGF in gene therapy of cervical cancer.%目的:研究结缔组织生长因子(connective tissue growth factor,CTGF)基因对人宫颈癌Hela细胞基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)表达及细胞增殖的影响,探讨CTGF在宫颈癌侵袭和转移中的作用机制.方法:经脂质体介导将含有CTGF重组表达质粒转染人宫颈癌Hela细胞株,用G418筛选阳性细胞克隆及实时荧光定量PCR、蛋白质印迹鉴定;采用实时荧光定量PCR和蛋白质印迹法检测阳性克隆细胞MMP-2及MMP-9的表达;噻唑盐(MTT)比色法检测阳性细胞克隆的增殖活性.结果:成功建立稳定高表达CTGF的阳性Hela细胞克隆,证实其MMP-2、MMP-9表达及细

  16. EFFECTS OF CIGARETTE SMOKE EXTRACT ON PROLIFERATION AND CTGF EXPRESSION OF HUMAN PULMONARY ARTERY SMOOTH MUSCLE CELLS%结缔组织生长因子在香烟提取物致人肺动脉平滑肌细胞增殖中的作用研究

    Institute of Scientific and Technical Information of China (English)

    卞维仕; 康振兴; 武善霞; 田锋; 杜庆伟

    2016-01-01

    Objective In order to explor the effects of cigarette smoke extract (CSE) on cell proliferation and the role of connective tissue growth factor (CTGF) .Methods In this study ,Cultured hPASMCs were divided into six groups :A :the normal control group ,B :hypoxia control group ,C :CSE group ,D :CSE and rhTGF‐β1 group ,E :CSE and CTGF sense oligonucleotide group ,F :CSE and CTGF antisense oligo‐nucleotide group .Then ,the expression of CTGF mRNA and protein of the supernatant of cells ,collagen content of cell culture supernatant .Results In Cell proliferation assay ,C group was more increased than in group A respectively ,Compared with D and C group , D group was more outstanding ,F groupwas decreased significantly ,compared with C group ( P 0 .05) .The results of CTGF mRNA ex‐pression and the content of type Ⅷ collagen in the su‐pernatant of cell culture were showed similar tendency to the protein expression .Conclusion CSE can in‐duce hPASMC proliferation ,CTGF may play an important role in it .%目的:探讨香烟提取物(CSE)对人肺动脉平滑肌细胞(hPASMC)增殖的影响及结缔组织生长因子(CTGF)在其中的作用。方法培养的hPASMC随机分为正常对照组(A)、缺氧对照组(B)、CSE组(C)、CSE+rhTGF‐β1组(D)、CSE+CTGF正义寡核苷酸组(E)、CSE+CTGF反义寡核苷酸组(F),分别检测细胞增殖、CTGF mRNA及蛋白表达、细胞培养上清液中I型胶原含量。结果细胞增殖检测,C组与A组比较明显升高,D组与C组比较,D组升高更卓著,F较C组下降显著( P均<0.01);E和C组之间无显著性差异( P >0.05)。hPASMC内CTGF蛋白表达量C组较A组明显升高,D较C组升高更明显,F较C组下降显著( P均<0.01);E组与C组之间无显著性差异( P >0.05);CTGF mRNA表达结果及细胞培养上清液中I型胶原含量均与蛋白表达有相似趋势。结论 CSE能诱导hPASMC增殖

  17. Atorvastatin Attenuates Bleomycin-Induced Pulmonary Fibrosis via Suppressing iNOS Expression and the CTGF (CCN2/ERK Signaling Pathway

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    Bo Zhu

    2013-12-01

    Full Text Available Pulmonary fibrosis is a progressive and fatal lung disorder with high mortality rate. To date, despite the fact that extensive research trials are ongoing, pulmonary fibrosis continues to have a poor response to available medical therapy. Statins, 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, known for its broad pharmacological activities, remains a remedy against multiple diseases. The present study investigated the antifibrotic potential of atorvastatin against bleomycin-induced lung fibrosis and to further explore the possible underlying mechanisms. Our results showed that atorvastatin administration significantly ameliorated the bleomycin mediated histological alterations and blocked collagen deposition with parallel reduction in the hydroxyproline level. Atorvastatin reduced malondialdehyde (MDA level and lung indices. Atorvastatin also markedly decreased the expression of inducible nitric oxide synthase (iNOS in lung tissues and, thus, prevented nitric oxide (NO release in response to bleomycin challenge. Furthermore, atorvastatin exhibited target down-regulation of connective tissue growth factor (CTGF (CCN2 and phosphorylation extracellular regulated protein kinases (p-ERK expression. Taken together, atorvastatin significantly ameliorated bleomycin-induced pulmonary fibrosis in rats, via the inhibition of iNOS expression and the CTGF (CCN2/ERK signaling pathway. The present study provides evidence that atorvastatin may be a potential therapeutic reagent for the treatment of lung fibrosis.

  18. PRS-CTGF-SiRNA逆转录病毒载体及包装细胞株的构建%Construction of retrovirus expressing CTGF siRNA and its steady packaging cell line mediated by PRS retrovirus vector

    Institute of Scientific and Technical Information of China (English)

    肖力; 刘伏友; 肖平; 彭佑铭; 段绍斌; 刘虹; 刘映红

    2007-01-01

    目的 用PRS逆转录病毒载体构建介导人结缔组织生长因子(CTGF)小分子干扰RNA(small interfering RNA,sRNA)表达的PRS-CTGF-SiRNA逆转录病毒载体,并建立高效、稳定表达CTGF-SiRNA的包装细胞株.方法 根据siRNA靶序列设计要求及PRS逆转录病毒载体特点分别设计含64bp DNA序列的4对寡核苷酸,并在体外合成.PRS载体用BglⅡ、HindⅢ双酶切完全后,分别与退火的4对寡核苷酸进行连接,构建成表达CTGF小分子干扰RNA的pRS-CTGF-SiRNA逆转录病毒载体.用脂质体2000介导转染PT67包装细胞,经抗性筛选,挑单克隆,扩大培养,用NIH3T3细胞测定病毒滴度,筛选出高效产毒细胞株.结果 经酶切、连接后构建成的质粒称之为PRS-CTGF-SiRNA1~4,经酶切与测序证实构建成功,无任何碱基突变.经脂质体介导转染包装细胞、抗性筛选和NIH3T3测定病毒滴度,筛选出具有高病毒滴度的细胞集落,扩大培养后建立CTGF-SiRNA高效表达的包装细胞株.结论 成功构建了能表达CTGFsiRNA的PRS-CTGF-SiRNA逆转录病毒载体,并建立了稳定、高效生产有活性的的CTGF-SiRNA产毒包装细胞株.

  19. CTGF对体外人晶状体上皮细胞表达MMP-2、TIMP-2的影响%Effect of CTGF on Expression of MMP-2 ,TIMP-2 of Human Lens Epithelial Cells Incubated in vitro

    Institute of Scientific and Technical Information of China (English)

    谢伟英; 徐国兴

    2013-01-01

    目的 研究结缔组织生长因子(CTGF)对体外培养人晶状体上皮细胞(HLECs)基质金属蛋白酶-2(MMP-2)及基质金属蛋白酶抑制剂-2(TIMP-2)表达的影响.方法 采用免疫细胞化学染色法及反转录-聚合酶链反应(RT-PCR)法检测不同浓度CTGF对HLECs诱导MMP-2及TIMP-2的表达.结果 在无CTGF刺激的情况下,HLECs基本不表达MMP-2及TIMP-2;随着CTGF浓度增加,HLECs表达MMP-2增加,并且这种作用随浓度的增加而增强,每两组之间的比较差异均有统计学意义(P<0.01或P<0.05);对TIMP-2的表达无明显影响(P>0.05).结论 CTGF能诱导HLECs表达MMP-2,对HLECs表达TIMP-2无影响;MMP-2/TIMP-2的比例失调,引起ECM的代谢紊乱,可能是后发性白内障的发病机制之一.%Objective To investigate the effect of connective tissue growth factor( CTGF) on the expression of matrix metalloprotein-ases -2(MMP -2) and tissue inhibitor of matrix metalloproteinases -2(TIMP -2) of human lens epithelial cells(HLECs) in vitro,thus to evaluate the role of CTGF in the pathogenesis of post - cataract. Methods The expression of MMP - 2 and TIMP - 2 was detected by immunocytochemical stain and RT - PCR when HLECs were incubated for 24h. Results In a dose - dependent manner,CTGF increased MMP -2 mRNA and protein expression in HLECs(P 0. 05 ) . Conclusion CTGF can induce the expression of MMP - 2 in HLECs, but has no effect on TIMP - 2. This leads to in balance between MMP -2 and TIMP -2. This might be an important pathway leading to post - cataract.

  20. Effect of high glucose, angiotensin Ⅱ and receptor antagonist Losartan on the expression of connective tissue growth factor in cultured mesangial cells

    Institute of Scientific and Technical Information of China (English)

    黄颂敏; 刘芳; 沙朝晖; 付平; 杨一帆; 徐勇; 周海燕

    2003-01-01

    Objective To observe the effect of high glucose, angiotensin Ⅱ (AngⅡ) and Losartan on the expression of connective tissue growth factor (CTGF) mRNA in cultured mesangial cells (MCs). Methods MCs of SD rats were isolated and cultured. High glucose (30 mmol/L) and AngⅡ (10-9, 10 7, and 10-5 mol/L) were added to the medium for 72 hours to observe the influence on CTGF mRNA expression. Losartan of 10-5 mol/L and AngⅡ of 10-5 mol/L were added to the medium to observe the effects of Losartan on CTGF mRNA expression stimulated by AngⅡ. The expressions of CTGF mRNA were detected by reverse transcriptase polymerase chain reaction (RT-PCR).Results RT-PCR showed that high glucose and AngⅡ up-regulated the expression of CTGF mRNA, and AngⅡ stimulated the expression in a dose-dependent manner. Expression of CTGF mRNA induced by AngⅡwas partially suppressed by 10-5mol/L Losartan (P<0.05).Conclusions High glucose and AngⅡ can enhance the expression of CTGF mRNA and thus be involved in the process of renal fibrosis. Losartan can have a partial fibrogenesis-inhibiting effect, with implications for the treatment of renal fibrosis.

  1. Coffee attenuates fibrosis by decreasing the expression of TGF-β and CTGF in a murine model of liver damage.

    Science.gov (United States)

    Arauz, Jonathan; Moreno, Marina Galicia-; Cortés-Reynosa, Pedro; Salazar, Eduardo Pérez; Muriel, Pablo

    2013-09-01

    This study was performed to evaluate the antifibrotic properties of coffee in a model of liver damage induced by repeated administration of thioacetamide (TAA) in male Wistar rats. In this study, cirrhosis was induced by chronic TAA administration and the effects of co-administration of conventional caffeinated coffee or decaffeinated coffee (CC, DC, respectively) for 8 weeks were evaluated. TAA administration elevated serum alkaline phosphatase (AP), γ-glutamyl transpeptidase (γ-GTP) and alanine aminotransferase (ALAT), liver lipid peroxidation, collagen content, depleted liver glycogen and glutathione peroxidase (GPx) activity. Additionally increased levels of a number of proteins were detected including transforming growth factor-beta (TGF-β), connective tissue growth factor (CTGF) and alpha-smooth muscle actin (α-SMA), and matrix metalloproteinase (MMP)-2, 9 and 13. Coffee suppressed most of the changes produced by TAA. Histopathological analysis was in agreement with biochemical and molecular findings. These results indicate that coffee attenuates experimental cirrhosis; the action mechanisms are probably associated with its antioxidant properties and mainly by its ability to block the elevation of the profibrogenic cytokine TGF-β and its downstream effector CTGF. Various components of coffee that have been related to such a favorable effect include caffeine, coffee oils kahweol, cafestol and antioxidant substances; however, no definite evidence for the role of these components has been established. These results support earlier findings suggesting a beneficial effect of coffee on the liver. However, more basic clinical studies must be performed to confirm this hypothesis. Copyright © 2012 John Wiley & Sons, Ltd.

  2. Connective Tissue Growth Factor Expression in Human Bronchial Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Amrita DOSANJH

    2006-01-01

    Connective tissue growth factor (CTGF) is a cysteine-rich protein that promotes extracellular matrix deposition. CTGF is selectively induced by transforming growth factor β and des-Arg kallidin in lung fibroblasts and increases steady-state mRNA levels of α type I collagen, 5α-integrin and fibronectin in fibroblasts. Bronchial epithelial cells have been proposed to functionally interact with lung fibroblasts. We therefore investigated if bronchial epithelial cells are able to synthesize CTGF. Human bronchial epithelial cells were grown to subconfluence in standard growth media. Proliferating cells grown in small airway growth media were harvested following starvation for up to 24 h. Expression of CTGF transcripts was measured by PCR. Immunocytochemistry was also completed using a commercially available antibody.The cells expressed readily detectable CTGF transcripts. Starvation of these cells resulted in a quantitative decline of CTGF transcripts. Direct sequencing of the PCR product identified human CTGF. Immunocytochemistry confirmed intracellular CTGF in the cells and none in negative control cells. We conclude that bronchial epithelial cells could be a novel source of CTGF. Bronchial epithelial cell-derived CTGF could thus directly influence the deposition of collagen in certain fibrotic lung diseases.

  3. Mechanical tension increases CCN2/CTGF expression and proliferation in gingival fibroblasts via a TGFβ-dependent mechanism.

    Directory of Open Access Journals (Sweden)

    Fen Guo

    Full Text Available Unlike skin, oral gingival do not scar in response to tissue injury. Fibroblasts, the cell type responsible for connective tissue repair and scarring, are exposed to mechanical tension during normal and pathological conditions including wound healing and fibrogenesis. Understanding how human gingival fibroblasts respond to mechanical tension is likely to yield valuable insights not only into gingival function but also into the molecular basis of scarless repair. CCN2/connective tissue growth factor is potently induced in fibroblasts during tissue repair and fibrogenesis. We subjected gingival fibroblasts to cyclical strain (up to 72 hours using the Flexercell system and showed that CCN2 mRNA and protein was induced by strain. Strain caused the rapid activation of latent TGFβ, in a fashion that was reduced by blebbistatin and FAK/src inhibition, and the induction of endothelin (ET-1 mRNA and protein expression. Strain did not cause induction of α-smooth muscle actin or collagen type I mRNAs (proteins promoting scarring; but induced a cohort of pro-proliferative mRNAs and cell proliferation. Compared to dermal fibroblasts, gingival fibroblasts showed reduced ability to respond to TGFβ by inducing fibrogenic mRNAs; addition of ET-1 rescued this phenotype. Pharmacological inhibition of the TGFβ type I (ALK5 receptor, the endothelin A/B receptors and FAK/src significantly reduced the induction of CCN2 and pro-proliferative mRNAs and cell proliferation. Controlling TGFβ, ET-1 and FAK/src activity may be useful in controlling responses to mechanical strain in the gingiva and may be of value in controlling fibroproliferative conditions such as gingival hyperplasia; controlling ET-1 may be of benefit in controlling scarring in response to injury in the skin.

  4. MicroRNA signature in wound healing following excimer laser ablation: role of miR-133b on TGFβ1, CTGF, SMA, and COL1A1 expression levels in rabbit corneal fibroblasts.

    Science.gov (United States)

    Robinson, Paulette M; Chuang, Tsai-Der; Sriram, Sriniwas; Pi, Liya; Luo, Xiao Ping; Petersen, Bryon E; Schultz, Gregory S

    2013-10-23

    The role of microRNA (miRNA) regulation in corneal wound healing and scar formation has yet to be elucidated. This study analyzed the miRNA expression pattern involved in corneal wound healing and focused on the effect of miR-133b on expression of several profibrotic genes. Laser-ablated mouse corneas were collected at 0 and 30 minutes and 2 days. Ribonucleic acid was collected from corneas and analyzed using cell differentiation and development miRNA PCR arrays. Luciferase assay was used to determine whether miR-133b targeted the 3' untranslated region (UTR) of transforming growth factor β1 (TGFβ1) and connective tissue growth factor (CTGF) in rabbit corneal fibroblasts (RbCF). Quantitative real-time PCR (qRT-PCR) and Western blots were used to determine the effect of miR-133b on CTGF, smooth muscle actin (SMA), and collagen (COL1A1) in RbCF. Migration assay was used to determine the effect of miR-133b on RbCF migration. At day 2, 37 of 86 miRNAs had substantial expression fold changes. miR-133b had the greatest fold decrease at -14.33. Pre-miR-133b targeted the 3' UTR of CTGF and caused a significant decrease of 38% (P wound healing, suggesting novel miRNA targets to reduce scar formation.

  5. PLGA-PEG-PLGA microspheres as a delivery vehicle for antisense oligonucleotides to CTGF: Implications on post-surgical peritoneal adhesion prevention

    Science.gov (United States)

    Azeke, John Imuetinyan-Jesu, Jr.

    Abdominal adhesions are the aberrant result of peritoneal wound healing commonly associated with surgery and inflammation. A subject of a large number of studies since the first half of the last century, peritoneal adhesion prevention has, for the most part, evaded the scientific community and continues to cost Americans an estimated $2-4 billion annually. It is known that transforming growth factor-beta (TGF-beta) plays a key role in the wound healing cascade; however, suppression of this multifunctional growth factor's activity may have more harmful consequences than can be tolerated. As a result, much attention has fallen on connective tissue growth factor (CTGF), a downstream mediator of TGF-beta's fibrotic action. It has been demonstrated in several in vitro models, that the suppression of CTGF hinders fibroblast proliferation, a necessary condition for fibrosis. Furthermore, antisense oligonucleotides (antisense oligos, AO) to CTGF have been shown to knock down CTGF mRNA levels by specifically hindering the translation of CTGF protein. Antisense technologies have met with a great deal of excitement as a viable means of preventing diseases such as adhesions by hindering protein translation at the mRNA level. However, the great challenge associated with the use of these drugs lies in the short circulation time when administered "naked". Viral delivery systems, although excellent platforms in metabolic studies, are not ideal for diagnostic use because of the inherent danger associated with viral vectors. Microparticles made of biodegradable polymers have therefore presented themselves as a viable means of delivering these drugs to target cells over extended periods. Herein, we present two in vivo studies confirming the up-regulation of TGF-beta protein and CTGF mRNA following injury to the uterine tissues of female rats. We were able to selectively knockdown post-operative CTGF protein levels following surgery, however, our observations led us to conclude that

  6. CTGF与TGF-β1在胃癌组织中的表达及其意义%EXPRESSION OF CTGF AND TGF-β1 AND THEIR SIGNIFICANCE IN GASTRIC CANCER

    Institute of Scientific and Technical Information of China (English)

    赵刚; 窦拉加; 张成武; 李菊英

    2009-01-01

    Objective To investigate the expression and significance of Conective tissue growth factor(CTGF)and Tranforming grouth factor-β1(TGF-β1)in gastric carcinoma.Methods The expression of CTGF and TGF-β1 in 50 patients with gastric cancer was examined by immunohistochemistry.Results The expression of CTGF and TGF-β1 was higher in GC group than that of in normal (P<0.05);Expression of CTGF were highly correlated with the carcinoma's degree of differentiation,lymph node metastasis and clinical stage (P<0.05);Expression of TGF-β1 were highly correlated with the carcinoma's degree of differentiation and lymph node metastasis (P<0.05) too;And CTGF and TGF-β1 overexpression were positively correlated in gastric carcinoma(rs=0.355,P=0.012).Conclusion All of CTGF and TGF-β1were over-expressed in gastric carcinoma,and correlated with the differentiation and tlymph node metastasis.It is considered that detection of CTGF and TGF-β1 may serve as fairly good index for early diagnosis and evaluating the prognosis of gastric carcinoma.%目的 探讨结缔组织生长因子与转化生长因子-β1在胃癌组织中的表达及其意义.方法 利用免疫组织化学方法检测50例胃癌组织和正常组织中CTGF、TGF-β1的表达,并分析其与胃癌临床病理因素之间的关系和意义.结果 胃癌组织中CTGF、TGF-β1的阳性表达率均明显高于正常组织(P<0.05);CTGF的高表达与胃癌的分化程度、淋巴结转移与否、临床分期均密切相关(P<0.05);TGF-β1的高表达与胃癌的分化程度和淋巴结转移密切相关(P<0.05);相关分析显示在胃癌组织中CTGF和TGF-β1的表达呈正相关关系(rs=0.355,P=0.012).结论 CTGF和TGF-β1在胃癌组织中均呈现高表达,并且与胃癌的分化程度、淋巴结转移与否有关,两者联合检测可以作为胃癌早期诊断和预后判断的指标.

  7. MicroRNA-26b inhibits metastasis of osteosarcoma via targeting CTGF and Smad1.

    Science.gov (United States)

    Duan, Guoqing; Ren, Chunfeng; Zhang, Yuanmin; Feng, Shiqing

    2015-08-01

    Downregulation of miR-26b has been found in various cancers, but it has never been investigated in osteosarcoma. In this study, we demonstrated downregulation of miR-26b in osteosarcoma tissues, negatively correlated with the expression of connective tissue growth factor (CTGF) and Smad1. Luciferase reporter assay confirmed the interaction of miR-26b with the 3' untranslated regions (UTRs) of CTGF and Smad1. Transfection of miR-26b in osteosarcoma cells suppressed the expression of CTGF and Smad1, suggesting CTGF and Smad1 as direct targets of miR-26b. Overexpression of miR-26b inhibited the migration of osteosarcoma cells, which was reversed by overexpression of CTGF or Smad1. Knockdown of CTGF by small interfering RNA (siRNA) interference blocked the activation of Smad1, ERK1/2, and MMP2, which was opposite to the overexpression of CTGF. Differently, Smad1 did not significantly affect CTGF level, but mediated ERK1/2 phosphorylation and MMP2 activation. Furthermore, miR-26b inhibited lung metastasis of osteosarcoma in vivo. Our data indicated that downregulation of miR-26b in osteosarcoma elevated the levels of CTGF and Smad1, facilitating osteosarcoma metastasis.

  8. [Preliminary study of the expression of connective tissue growth factor in papillary muscles of the patients with rheumatic heart disease].

    Science.gov (United States)

    Wang, Y N; Li, T; Gu, J R; Yu, B Y

    2016-04-19

    To investigate the expression and the effect of connective tissue growth factor (CTGF) on rheumatic myocardial fibrosis of rheumatic heart disease (RHD). The papillary muscles samples were obtained from patients with RHD during mitral valve replacement.The expression of TGF-β1, CTGF mRNA and CTGF protein were detected with semiquantitative RT-PCR technique and immunohistochemistry technologyin the papillary muscles cell from 41RHD patients and 20 normal papillary muscles samples.The area of myocardial fibrosis was measured by imaging analysis system. SPSS package was used to analyze the relationship between the expression of CTGF and the area of myocardial fibrosis. Compared with normal controls (PU 2.4±0.9), the mean level of CTGF protein expression in the papillary muscles samples of the RHD patients (PU 44.7±6.0) was significantly increased(Pfibrosis (r=0.856, Pfibrosis of RHD.

  9. A pilot trial assessing urinary gene expression profiling with an mRNA array for diabetic nephropathy.

    Directory of Open Access Journals (Sweden)

    Min Zheng

    Full Text Available BACKGROUND: The initiation and progression of diabetic nephropathy (DN is complex. Quantification of mRNA expression in urinary sediment has emerged as a novel strategy for studying renal diseases. Considering the numerous molecules involved in DN development, a high-throughput platform with parallel detection of multiple mRNAs is needed. In this study, we constructed a self-assembling mRNA array to analyze urinary mRNAs in DN patients with aims to reveal its potential in searching novel biomarkers. METHODS: mRNA array containing 88 genes were fabricated and its performance was evaluated. A pilot study with 9 subjects including 6 DN patients and 3 normal controls were studied with the array. DN patients were assigned into two groups according to their estimate glomerular rate (eGFR: DNI group (eGFR>60 ml/min/1.73 m(2, n = 3 and DNII group (eGFR<60 ml/min/1.73 m(2, n = 3. Urinary cell pellet was collected from each study participant. Relative abundance of these target mRNAs from urinary pellet was quantified with the array. RESULTS: The array we fabricated displayed high sensitivity and specificity. Moreover, the Cts of Positive PCR Controls in our experiments were 24±0.5 which indicated high repeatability of the array. A total of 29 mRNAs were significantly increased in DN patients compared with controls (p<0.05. Among these genes, α-actinin4, CDH2, ACE, FAT1, synaptopodin, COL4α, twist, NOTCH3 mRNA expression were 15-fold higher than those in normal controls. In contrast, urinary TIMP-1 mRNA was significantly decreased in DN patients (p<0.05. It was shown that CTGF, MCP-1, PAI-1, ACE, CDH1, CDH2 mRNA varied significantly among the 3 study groups, and their mRNA levels increased with DN progression (p<0.05. CONCLUSION: Our pilot study demonstrated that mRNA array might serve as a high-throughput and sensitive tool for detecting mRNA expression in urinary sediment. Thus, this primary study indicated that mRNA array probably could be a

  10. Role of CTGF/CCN2 in reparative dentinogenesis in human dental pulp.

    Science.gov (United States)

    Muromachi, Koichiro; Kamio, Naoto; Matsumoto, Takashi; Matsushima, Kiyoshi

    2012-03-01

    Connective tissue growth factor/CCN family 2 (CTGF/CCN2) has been considered to participate in tooth development. To date, the expression and role of CTGF/CCN2 in reparative dentinogenesis have been unclear. Our previous study revealed that matrix metalloproteinase-3 (MMP-3) stimulates cell migration via CTGF/CCN2 expression and secretion in human dental pulp cells, and that this is dependent on dynamin-related endocytosis and independent of protease activity. The objective of the present study was to determine the expression of CTGF/CCN2 in reparative dentin in human carious teeth and to examine the effect of CTGF/CCN2 on mineralization in cultured human dental pulp cells. Minimal expression of CTGF/CCN2 was evident in odontoblasts subjacent to the dentin-pulp junction in healthy teeth, whereas strong expression was detected in odontoblast-like cells lining the reparative dentin subjacent to dental caries. In human dental pulp cells, CTGF/CCN2 promoted mineralization but failed to induce proliferation, suggesting that this molecule has the ability to induce the differentiation of human dental pulp cells. Taken together, the data suggest that CTGF/CCN2 is likely involved in reparative dentinogenesis through formation of hard tissue in human carious teeth.

  11. Cartilage-specific over-expression of CCN family member 2/connective tissue growth factor (CCN2/CTGF) stimulates insulin-like growth factor expression and bone growth.

    Science.gov (United States)

    Tomita, Nao; Hattori, Takako; Itoh, Shinsuke; Aoyama, Eriko; Yao, Mayumi; Yamashiro, Takashi; Takigawa, Masaharu

    2013-01-01

    Previously we showed that CCN family member 2/connective tissue growth factor (CCN2) promotes the proliferation, differentiation, and maturation of growth cartilage cells in vitro. To elucidate the specific role and molecular mechanism of CCN2 in cartilage development in vivo, in the present study we generated transgenic mice overexpressing CCN2 and analyzed them with respect to cartilage and bone development. Transgenic mice were generated expressing a ccn2/lacZ fusion gene in cartilage under the control of the 6 kb-Col2a1-enhancer/promoter. Changes in cartilage and bone development were analyzed histologically and immunohistologically and also by micro CT. Primary chondrocytes as well as limb bud mesenchymal cells were cultured and analyzed for changes in expression of cartilage-related genes, and non-transgenic chondrocytes were treated in culture with recombinant CCN2. Newborn transgenic mice showed extended length of their long bones, increased content of proteoglycans and collagen II accumulation. Micro-CT analysis of transgenic bones indicated increases in bone thickness and mineral density. Chondrocyte proliferation was enhanced in the transgenic cartilage. In in vitro short-term cultures of transgenic chondrocytes, the expression of col2a1, aggrecan and ccn2 genes was substantially enhanced; and in long-term cultures the expression levels of these genes were further enhanced. Also, in vitro chondrogenesis was strongly enhanced. IGF-I and IGF-II mRNA levels were elevated in transgenic chondrocytes, and treatment of non-transgenic chondrocytes with recombinant CCN2 stimulated the expression of these mRNA. The addition of CCN2 to non-transgenic chondrocytes induced the phosphorylation of IGFR, and ccn2-overexpressing chondrocytes showed enhanced phosphorylation of IGFR. Our data indicates that the observed effects of CCN2 may be mediated in part by CCN2-induced overexpression of IGF-I and IGF-II. These findings indicate that CCN2-overexpression in transgenic

  12. Expressions of cysteine-rich61, connective tissue growth factor and Nov genes in hepatocellular carcinoma and their clinical significance

    Institute of Scientific and Technical Information of China (English)

    Zhi-Jun Zeng; Lian-Yue Yang; Xiang Ding; Wei Wang

    2004-01-01

    AIM: To investigate the expression of cysteine-rich61 (Cyr61),connective tissue growth factor (CTGF) and nephroblastoma overexpressed gene (Nov) in hepatocellular carcinoma (HCC),and to evaluate the relationship between Cyr61, CTGF and Nov genes expression with invasion and metastasis of HCC.METHODS: Thirty-one HCC specimens were divided into small hepatocellular carcinoma (SHCC), nodular hepatocellular carcinoma (NHCC), solitary large hepatocellular carcinoma (SLHCC) according to their diameter and number of nodes. Reverse transcription polymerse chain reaction (RT-PCR) was used to detect the mRNA expression levels of Cyr61, CTGF and Nov genes in 31 resected specimens of hepatocellular carcinoma and para-cancerous normal liver tissues semi-quantitatively and the relation between their expression levels and clinical pathological parameters were compared.RESULTS: The expressions of Cyr61 and CTGF mRNA in carcinoma tissues were significantly higher than those in para-cancerous normal liver tissues (P<0.01). The expressions of Cyr61 and CTGF mRNA in HCC with venous invasion were higher than those in HCC without venous invasion. CTGF expression in HCC Edmondson's grade Ⅲ-Ⅳ was significantly higher than that in HCC Edmondson's grade Ⅰ-Ⅱ (P = 0.022). There was no obvious correlation between Nov mRNA and clinical-pathological features.Compared to NHCC, SLHCC had better cell differentiation,easier capsule formation, less microscopic venous invasion,milder liver cirrhosis. The expressions of Cyr61 and CTGF mRNA in NHCC were significantly higher than those in SLHCC and SHCC.CONCLUSION: Cyr61 and CTGF genes may play an important role in hepatocellular carcinogenesis and correlate with recurrence and metastasis of hepatocellular carcinoma.SLHCC has better biological behaviors than NHCC.

  13. CTGF mediates Smad-dependent transforming growth factor β signaling to regulate mesenchymal cell proliferation during palate development.

    Science.gov (United States)

    Parada, Carolina; Li, Jingyuan; Iwata, Junichi; Suzuki, Akiko; Chai, Yang

    2013-09-01

    Transforming growth factor β (TGF-β) signaling plays crucial functions in the regulation of craniofacial development, including palatogenesis. Here, we have identified connective tissue growth factor (Ctgf) as a downstream target of the TGF-β signaling pathway in palatogenesis. The pattern of Ctgf expression in wild-type embryos suggests that it may be involved in key processes during palate development. We found that Ctgf expression is downregulated in both Wnt1-Cre; Tgfbr2(fl/fl) and Osr2-Cre; Smad4(fl/fl) palates. In Tgfbr2 mutant embryos, downregulation of Ctgf expression is associated with p38 mitogen-activated protein kinase (MAPK) overactivation, whereas loss of function of Smad4 itself leads to downregulation of Ctgf expression. We also found that CTGF regulates its own expression via TGF-β signaling. Osr2-Cre; Smad4(fl/fl) mice exhibit a defect in cell proliferation similar to that of Tgfbr2 mutant mice, as well as cleft palate. We detected no alteration in bone morphogenetic protein (BMP) downstream targets in Smad4 mutant palates, suggesting that the reduction in cell proliferation is due to defective transduction of TGF-β signaling via decreased Ctgf expression. Significantly, an exogenous source of CTGF was able to rescue the cell proliferation defect in both Tgfbr2 and Smad4 mutant palates. Collectively, our data suggest that CTGF regulates proliferation as a mediator of the canonical pathway of TGF-β signaling during palatogenesis.

  14. High lib mRNA expression in breast carcinomas.

    Science.gov (United States)

    Satoh, Kazuki; Hata, Mitsumi; Yokota, Hiroshi

    2004-06-30

    Lib, first identified as a novel beta-amyloid responsive gene in rat astrocytes, has an extracellular domain of 15 leucine-rich repeats (LRRs) followed by a transmembrane domain and a short cytoplasmic region. It is a distinctly inducible gene and is thought to play a key role in inflammatory states via the LRR extracellular motif, an ideal structural framework for protein-protein and protein-matrix interactions. To evaluate potential roles of Lib, we screened various tumors for Lib expression. Lib mRNA expression was high and uniquely expressed in breast tumor tissues, compared to paired normal breast tissues. Lib mRNA was localized in the ductal carcinoma cells and Lib protein displayed a homophilic association on the surface of cultured cells. These data suggest that Lib may play a role in the progression of breast carcinomas and may be a diagnostic marker for breast tumors.

  15. CTGF siRNA 联合电针治疗抑制脊髓损伤胶质瘢痕形成%The study on CTGF siRNA combined with electro-acupuncture for inhibition of formation of glial scar following spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    张政; 房清敏; 杨成; 孙兆忠; 耿晓鹏; 朱吉超; 王大巍; 邢建强; 孙金川

    2015-01-01

    Objective To investigate the anti‐CTGF small interferon RNA (siRNA ) combined with electro‐acupuncture for inhibition of formation of glial scar following spinal cord injury (SCI) .Methods Adult female SD rats were randomly divided into SCI group ,electro‐acupuncture(EA) group ,CTGF siRNA interference (CTGF) group ,and CTGF siRNA interference com‐bined with electro -acupuncture (CTGF + EA) group .They were sampled at 3 d ,7 d and 14 d after injury ,respectively .The changes of glial scar following spinal cord injury were observed by using immunofluorescence histochemistry and reverse tran‐scriptase polymerase chain reaction (RT‐PCR) .The changes in the expression of GFAP ,Vimentin ,fibronectin (FN) and lami‐nin (LN) ,and mRNA expression of GFAP ,Vimentin ,FN and LN were detected ,respectively .Results The SCI group showed reactive astrocyte proliferation with increased number ;the expression levels of GFAP ,Vimentin ,FN and LN were increased . The expression levels of GFAP ,Vimentin ,FN and LN were decreased in the EA group、CTGF group and CTGF+ EA group .. In CTGF + EA group ,the expression levels of GFAP ,Vimentin ,FN and LN were significantly decreased .RT‐PCR results showed that the mRNA expression of GFAP ,Vimentin ,FN and LN in EA group、CTGF group and CTGF + EA group were all decreased compared with the SCI group (P<0.05);the expression of GFAP ,Vimentin ,FN and LN in the CTGF + EA group were also decreased compared with the EA group、CTGF group (P<0.05) .Conclusion After spinal cord injury ,CTGF siRNA combined with electro‐acupuncture can effectively inhibit the expression of GFAP ,Vimentin ,FN and LN ,and can inhibit the formation of glial scar to a certain extent ,contributing to the recovery of neurological function .%目的:探讨抗结缔组织生长因子(CTGF)小干扰 RNA(smal interferon RNA ,siRNA)联合电针治疗对脊髓损伤(spinal cord injury ,SCI)胶质瘢痕形成的影响。方法选用成年雌

  16. Vibrational force alters mRNA expression in osteoblasts

    Science.gov (United States)

    Tjandrawinata, R. R.; Vincent, V. L.; Hughes-Fulford, M.

    1997-01-01

    Serum-deprived mouse osteoblastic (MC3T3E1) cells were subjected to a vibrational force modeled by NASA to simulate a space shuttle launch (7.83 G rms). The mRNA levels for eight genes were investigated to determine the effect of vibrational force on mRNA expression. The mRNA levels of two growth-related protooncogenes, c-fos and c-myc, were up-regulated significantly within 30 min after vibration, whereas those of osteocalcin as well as transforming growth factor-beta1 were decreased significantly within 3 h after vibration. No changes were detected in the levels of beta-actin, histone H4, or cytoplasmic phospholipase A2 after vibration. No basal levels of cyclooxygenase-2 expression were detected. In addition, the extracellular concentrations of prostaglandin E2 (PGE2), a potent autocrine/paracrine growth factor in bone, were not significantly altered after vibration most likely due to the serum deprivation state of the osteoblasts. In comparison with the gravitational launch profile, vibrational-induced changes in gene expression were greater both in magnitude and number of genes activated. Taken together, these data suggest that the changes in mRNA expression are due to a direct mechanical effect of the vibrational force on the osteoblast cells and not to changes in the local PGE2 concentrations. The finding that launch forces induce gene expression is of utmost importance since many of the biological experiments do not dampen vibrational loads on experimental samples. This lack of dampening of vibrational forces may partially explain why 1-G onboard controls sometimes do not reflect 1-G ground controls. These data may also suggest that scientists use extra ground controls that are exposed to launch forces, have these forces dampened on launched samples, or use facilities such as Biorack that provide an onboard 1-G centrufuge in order to control for space shuttle launch forces.

  17. CTGF promotes osteosarcoma angiogenesis by regulating miR-543/angiopoietin 2 signaling.

    Science.gov (United States)

    Wang, Li-Hong; Tsai, Hsiao-Chi; Cheng, Yu-Che; Lin, Chih-Yang; Huang, Yuan-Li; Tsai, Chun-Hao; Xu, Guo-Hong; Wang, Shih-Wei; Fong, Yi-Chin; Tang, Chih-Hsin

    2017-04-10

    Osteosarcoma is the most common primary solid tumor of bone. It has a high metastatic potential and occurs predominantly in adolescents and young adults. Angiopoietin 2 (Angpt2) is a key regulator in tumor angiogenesis, facilitating tumor growth and metastasis. Connective tissue growth factor (CTGF, also known as CCN2), is a cysteine-rich protein that has been reported to promote metastasis of osteosarcoma. However, the effect of CTGF on Angpt2 regulation and angiogenesis in human osteosarcoma remains largely unknown. We found that overexpression of CTGF in osteosarcoma cells increased Angpt2 production and induced angiogenesis, in vitro and in vivo. Our findings demonstrate that CTGF-enhanced Angpt2 expression and angiogenesis is mediated by the phospholipase C (PLC)/protein kinase C (PKCδ) signaling pathway. Moreover, endogenous microRNA-543 (miR-543) expression was negatively regulated by CTGF via the PLC/PKCδ pathway. We also provide evidence showing clinical significance between CTGF, Angpt2, and miR-543 as well as tumor staging in human osteosarcoma tissue. CTGF may serve as a therapeutic target in the process of osteosarcoma metastasis and angiogenesis.

  18. The analysis of OMC’s abnormal and CTGF’s over expression in sinus operation%OMC 解剖异常与局部 CTGF 表达异常的关系分析

    Institute of Scientific and Technical Information of China (English)

    王毅; 习国平; 刘志明

    2014-01-01

    objective our test is discover the correlation between the ctGf’s express and the abnormal of oMc in the sinus endoscope surgery. through this test ,we can learn ctGf’s role in the correlation between the oMc’ abnormal and inflammation in part. Methods Patients were divided to 2 groups according to abnormal of oMc. Group a include those patients whose oMc is abnormal.Group B include those patients whose oMc is normal. through immunohistochemistry ,and percentage of positive cells were calculated under the high power lens .analyse the optical density through the imageJ analyse system .the data was analyse by sPss statistics software. results the ctGf masculine rate and optical density of group a are higher than group B. conclusion ctGf’s over expression is correlation with oMc’s abnormal,this perhaps can discover ctGf’s over expression is one of the reason that oMc’s abnormal can cause nasosinusitis.%目的:旨在通过研究鼻内窥镜手术中窦口鼻道复合体(ostiomeatalex oMc)解剖结构变异的程度和局部结缔组织生长因子(connective tissue growth factor,ctGf)表达的强度之间的关系。从而了解 ctGf 在 oMc 结构异常和局部炎性病变之间所起作用。方法将手术患者根据 oMc 变异程度分为两组,oMc 变异患者为 a 组,无明显变异者为B 组。通过免疫组织化学法对两组样本进行处理,通过对两组黏膜上皮细胞和粘膜下腺体细胞阳性细胞计数(%)及对 imageJ 光密度测定,sPss 统计软件进行统计学分析。结果 a 组患者黏膜细胞及黏膜下腺体细胞阳性率及光密度值均高于 B 组患者。结论 oMc 结构异常和局部 ctGf 表达有关,从而表明局部 ctGf 增多可能是 oMc 引起局部炎性增生性病变原因之一。

  19. Role of connective growth factor in plasminogen activator inhibitor-1 and fibronectin expression induced by transforming growth factor β1 in renal tubular cells

    Institute of Scientific and Technical Information of China (English)

    张春; 孟宪芳; 朱忠华; 杨晓; 邓安国

    2004-01-01

    Background Connective tissue growth factor (CTGF) contributes greatly to renal tubulointerstitial fibrosis, which is the final event leading to end-stage renal failure. This study was designed to investigate the effects of CTGF antisense oligodeoxynucleotides (ODNs) on the expressions of plasminogen activator inhibitor-1 (PAI-1) and fibronectin in renal tubular cells induced by transforming growth factor β1 (TGF-β1) in addition to the role of CTGF in the accumulation and degradation of renal extracellular matrix (ECM).Methods A human proximal tubular epithelial cell line (HKC) was cultured in vitro. Cationic lipid-mediated CTGF antisense ODNs were transfected into HKC cells. After HKC cells were stimulated with TGF-β1 (5 μg/L), the mRNA levels of PAI-1 and fibronectin were measured by RT-PCR. Intracellular PAI-1 protein synthesis was assessed by flow cytometry. The secreted PAI-1 and fibronectin in the medium were determined by Western blot and ELISA, respectively.Results TGF-β1 was found to induce tubular CTGF, PAI-1, and fibronectin mRNA expression. PAI-1 and fibronectin mRNA expression induced by TGF-β1 was significantly inhibited by CTGF antisense ODNs. CTGF antisense ODNs also inhibited intracellular PAI-1 protein synthesis and lowered the levels of PAI-1 and fibronectin protein secreted into the medium.Conclusions CTGF may play a crucial role in the accumulation and degradation of excessive ECM during tubulointerstitial fibrosis, and transfecting CTGF antisense ODNs may be an effective way to prevent renal fibrosis.

  20. Control of gene expression during T cell activation: alternate regulation of mRNA transcription and mRNA stability

    Directory of Open Access Journals (Sweden)

    Gorospe Myriam

    2005-05-01

    Full Text Available Abstract Background Microarray technology has become highly valuable for identifying complex global changes in gene expression patterns. The effective correlation of observed changes in gene expression with shared transcription regulatory elements remains difficult to demonstrate convincingly. One reason for this difficulty may result from the intricate convergence of both transcriptional and mRNA turnover events which, together, directly influence steady-state mRNA levels. Results In order to investigate the relative contribution of gene transcription and changes in mRNA stability regulation to standard analyses of gene expression, we used two distinct microarray methods which individually measure nuclear gene transcription and changes in polyA mRNA gene expression. Gene expression profiles were obtained from both polyA mRNA (whole-cell and nuclear run-on (newly transcribed RNA across a time course of one hour following the activation of human Jurkat T cells with PMA plus ionomycin. Comparative analysis revealed that regulation of mRNA stability may account for as much as 50% of all measurements of changes in polyA mRNA in this system, as inferred by the absence of any corresponding regulation of nuclear gene transcription activity for these groups of genes. Genes which displayed dramatic elevations in both mRNA and nuclear run-on RNA were shown to be inhibited by Actinomycin D (ActD pre-treatment of cells while large numbers of genes regulated only through altered mRNA turnover (both up and down were ActD-resistant. Consistent patterns across the time course were observed for both transcribed and stability-regulated genes. Conclusion We propose that regulation of mRNA stability contributes significantly to the observed changes in gene expression in response to external stimuli, as measured by high throughput systems.

  1. Inhibition effect of small interfering RNA of connective tissue growth factor on the expression of vascular endothelial growth factor and connective tissue growth factor in cultured human peritoneal mesothelial cells

    Institute of Scientific and Technical Information of China (English)

    LIU Fu-you; FU Xiao; ZHU Jian-lian; XIAO Li; PENG You-ming; DUAN Shao-bin; LIU Hong; LIU Ying-hong; LING Gui-hui; YUAN Fang; CHEN Jun-xiang

    2007-01-01

    Background The peritoneum response to peritoneal dialysis can lead to fibrosis. The transforming growth factor β1(TGF-β1) plays a key role in regulating tissue repair and remodelling after injury. Connective tissue growth factor (CTGF),a downstream mediator of TGF-β1 inducing fibrosis, has been implicated in peritoneal fibrosis. Vascular endothelial growth factor (VEGF) plays a key role in angiogenesis that can hasten peritoneal fibrosis. In this study, we investigated the effect of small interfering RNA (siRNA) of CTGF by pRETRO-SUPER (PRS) retrovirus vector on the expression of CTGF and VEGF in human peritoneal mesothelial cells.Methods Retrovirus producing CTGF siRNA were constructed from the inverted oligonucleotides and transferred into packaging cell line PT67 with lipofectamine, and the virus supernatant was used to infect human peritoneal mesothelial cell (HPMC). The cells were divided into seven groups: low glucose DMEM, low glucose DMEM + TGF-β1 5 ng/ml, low glucose DMEM + TGF-β1 5 ng/ml + PRS-CTGF-siRNA1-4 and low glucose DMEM + TGF-β1 5 ng/ml + PRS. The expression of CTGF and VEGF were measured by semiquantitative RT-PCR and Western blot.Results Low levels of CTGF and VEGF were detected in confluent HPMCs. Following stimulation with TGF-β1, the levels of CTGF and VEGF were significantly upregulated (P<0.01). Introduction of PRS-CTGF-siRNA1-4 resulted in the significant reduction of CTGF mRNA and protein, and VEGF mRNA (P<0.01), especially in groups PRS-CTGF-siRNA1 and PRS-CTGF-siRNA4. The introduction of PRS void vector did not have these effects (P>0.05).Conclusions The expression of CTGF siRNA mediated by PRS retrovirus vector can effectively reduce the level of CTGF and VEGF induced by TGF-β1 in cultured HPMCs. This study may provide potential therapeutic strategies to prevent the peritoneal fibrosis.

  2. Selenoprotein P mRNA expression in human hepatic tissues

    Institute of Scientific and Technical Information of China (English)

    Chun-Li Li; Ke-Jun Nan; Tao Tian; Chen-Guang Sui; Yan-Fang Liu

    2007-01-01

    AIM: To investigate the expression of Selenoprotein P mRNA (SePmRNA) in tissues of normal liver, liver cirrhosis and hepatocellular carcinoma (HCC), and its relationship with HCC occurrence and development.METHODS: The expression of SePmRNA in tissues of normal liver, liver cirrhosis and HCC were detected by in situ hybridization using a cDNA probe.RESULTS: The enzyme digesting products of pBluescript-Human Selenoprotein P were evaluated by electrophoresis.The positive expression of SePmRNA was found in the tissues of normal liver,liver cirrhosis and HCC.The expression of SeP mRNA was found in hepatic interstitial substance,especially in endothelial cells and lymphocytes of vasculature.The positive rate of SePmRNA in normal liver tissue was 84.6% (11/13) and the positive signals appeared in the nucleus and cytoplasm,mostly in the nucleolus,and the staining granules were larger in the nucleolus and around the nucleus.The positive rate of SePmRNA in liver cirrhosis tissue was 45.O% (9/20) and the positive signals were mainly in the nucleolus and cytoplasm,being less around the nucleus and inner nucleus than that in normal liver tissue. The positive rate of SePmRNA in HCC tissue was 30.0% (9/30) and the positive signals were in the cytoplasm, but less in the nucleus.CONCLUSION: SePmRNA expression in the tissues of normal liver and HCC is significantly different (84.6% vs 30.0%, P = 0.003), suggesting that SeP might play a role in the occurrence and development of HCC.

  3. Increased IL-10 mRNA and IL-23 mRNA expression in multiple sclerosis

    DEFF Research Database (Denmark)

    Krakauer, Martin; Sorensen, P; Khademi, M

    2008-01-01

    of the regulatory cytokine IL-10. The elevated IL-23 mRNA levels in MS patients are noteworthy in view of the newly discovered IL-23-driven Th17 T-cell subset, which is crucial in animal models of MS. Since IFN-beta therapy resulted in decreased IL-23 mRNA levels, the Th17 axis could be another target of IFN...

  4. Highest trkB mRNA expression in the entorhinal cortex among hippocampal subregions in the adult rat: contrasting pattern with BDNF mRNA expression.

    Science.gov (United States)

    Tokuyama, W; Hashimoto, T; Li, Y X; Okuno, H; Miyashita, Y

    1998-11-20

    Brain-derived neurotrophic factor (BDNF) and its receptor, TrkB, regulate synaptic functions in the hippocampus of the adult rodent. In previous studies, in situ hybridization methods have been used to evaluate regional differences in BDNF and trkB mRNA expression levels in hippocampal subregions. However, these studies have failed to reach consensus regarding the regional differences in the mRNA expression levels. In the present study, we quantitated mRNA expression levels using two different methods, ribonuclease protection assays and a quantitative reverse-transcription polymerase chain reaction technique, in four hippocampal subregions: the entorhinal cortex, dentate gyrus (DG), CA3 and CA1. These two methods yielded the same results. We found that BDNF and trkB mRNA expression levels did not covary in the four subregions. BDNF and full length trkB (trkB FL) mRNA in the entorhinal cortex and the DG show contrasting expression patterns. The expression level of BDNF mRNA was highest in the DG among the hippocampal subregions and low in the entorhinal cortex and the CA1, whereas the trkB FL mRNA expression level was highest in the entorhinal cortex, low in the DG and lowest in the CA3. These results suggest regional differences in BDNF/TrkB signaling for maintenance and modifiability of neuronal connections in the hippocampal formation.

  5. Connective tissue growth factor (CTGF/CCN2) in haemophilic arthropathy and arthrofibrosis: a histological analysis.

    Science.gov (United States)

    Jiang, J; Leong, N L; Khalique, U; Phan, T M; Lyons, K M; Luck, J V

    2016-11-01

    Joint haemorrhage is the principal clinical manifestation of haemophilia frequently leading to advanced arthropathy and arthrofibrosis, resulting in severe disability. The degree and prevalence of arthrofibrosis in hemophilic arthropathy is more severe than in other forms of arthropathy. Expression of connective tissue growth factor (CTGF) has been linked to many fibrotic diseases, but has not been studied in the context of haemophilic arthropathy. We aim to compare synovial tissues histologically from haemophilia and osteoarthritis patients with advanced arthropathy in order to compare expression of proteins that are possibly aetiologic in the development of arthrofibrosis. Human synovial tissues were obtained from 10 haemophilia and 10 osteoarthritis patients undergoing joint surgery and processed for histology and immunohistochemistry. All samples from haemophilia patients had synovitis with hypertrophy and hyperplasia of synovial villi. Histologically, synovial tissues contained hyperplastic villi with increased cellularity and abundant haemosiderin- and ferritin-pigmented macrophage-like cells (HMCs), with a perivascular localization in the sub-surface layer. CTGF staining was observed in the surface layer and sub-surface layer in all haemophilia patients, exclusively co-localizing with HMCs. Quantification showed that the extent of CTGF-positive areas was correlated with the degree of detection of HMCs. CTGF was not observed in any of the samples from osteoarthritis patients. Using histological analysis, we showed that CTGF expression is elevated in haemophilia patients with arthrofibrosis and absent in patients with osteoarthritis. Additionally, we found that CTGF is always associated with haemosiderin-pigmented macrophage-like cells, which suggests that CTGF is produced by synovial A cells following the uptake of blood breakdown products. © 2016 John Wiley & Sons Ltd.

  6. Angiotensin (1-7) inhibits angiotensin Ⅱ-stimulated expression of connective tissue growth factor mRNA in hepatic stellate cells%血管紧张素(1-7)对血管紧张素Ⅱ诱导肝星状细胞表达结缔组织生长因子的抑制作用

    Institute of Scientific and Technical Information of China (English)

    李旭; 黄茂梁; 黄珊; 张文雍; 宁佐伟; 孟莹

    2012-01-01

    mRNA的表达具有抑制作用.%Objective To explore the angiotensin peptide [Ang (1-7)]-mediated inhibition of Ang Ⅱ in human hepatic stellate cells (HSCs) and determine the involvement of the ACE2-Ang (1-7)-Mas axis.Methods The human HSC line,LX2,was used in all experiments,and divided into control (unstimulated)and Ang Ⅱ-stimulated (10-6 mol/L) groups.The Ang Ⅱ-stimulated cells were further divided among several pretreatment (prior to Ang Ⅱ) groups:ROCK-inhibited (Y27632 blocking agent,10-6mol/L); irbesartan-inhibited (AT-1 receptor antagonist,10-6mol/L); and Mas receptor-inhibited (A779 Mas receptor antagonist,10-6mol/L).To explore the potential inhibitory effects of various Ang family members,the Ang Ⅱ-stimulated and pre-treated LX2 cells were exposed to Ang (1-7) (10-6 mol/L) for 24h.Western blot,reverse transcription-polymerase chain reaction (RT-PCR),and QuantiGene assay were used to assess changes in protein and mRNA expression levels of RhoA,ROCK,and connective tissue growth factor (CTGF).Results Compared with the control group,Ang Ⅱ-stimulated cells showed significantly increased levels of RhoA protein (0.337±0.074 vs.0.870±0.093),ROCK2 mRNA (0.747±0.061 vs.0.368±0.023),and CTGF mRNA (0.262±0.007 vs.0.578±0.028) (all,P<0.01).Pre-treatment with irbesartan 0r Y27632 eliminated these responses.Ang (1-7) inhibited the Ang Ⅱ-stimulated up-regulation of RhoA,ROCK,and CTGF.Conclusion Ang (1-7) can inhibit the Ang Ⅱ-stimulated up-regulation of RhoA,ROCK and CTGF in hepatic stellate cells,indicating that the ACE2-Ang (1-7)-Mas axis,an important branch of the renin-Ang-aldosterone system is involved in the occurrence and development of liver fibrosis.

  7. Exogenous mRNA encoding tetanus or botulinum neurotoxins expressed in Aplysia neurons

    NARCIS (Netherlands)

    Mochida, Sumiko; Poulain, Bernard; Eisel, Ulrich; Binz, Thomas; Kurazono, Hisao; Niemann, Heiner; Tauc, Ladislav; Bullock, Theodore H.

    1990-01-01

    Injection of exogenous mRNA purified from various tissue preparations into cellular translation systems such as Xenopus oocytes has allowed expression of complex proteins (e.g., receptors for neurotransmitters). No evidence for expression of injected exogenous mRNA, however, has been reported in

  8. Neurotrophin-3 mRNA expression in rat intrafusal muscle fibres after denervation and reinnervation

    NARCIS (Netherlands)

    Copray, JCVM; Brouwer, N

    1997-01-01

    We have studied the regulation of the expression of neurotrophin-3 (NT-3) mRNA in neonatal and adult rat muscle spindles after denervation and after denervation followed by reinnervation. Denervation of the intrafusal fibres did not result in an upregulation of the NT-3 mRNA expression but decreased

  9. Exogenous mRNA encoding tetanus or botulinum neurotoxins expressed in Aplysia neurons

    NARCIS (Netherlands)

    Mochida, Sumiko; Poulain, Bernard; Eisel, Ulrich; Binz, Thomas; Kurazono, Hisao; Niemann, Heiner; Tauc, Ladislav; Bullock, Theodore H.

    1990-01-01

    Injection of exogenous mRNA purified from various tissue preparations into cellular translation systems such as Xenopus oocytes has allowed expression of complex proteins (e.g., receptors for neurotransmitters). No evidence for expression of injected exogenous mRNA, however, has been reported in ter

  10. CTGF siRNA ameliorates tubular cell apoptosis and tubulointerstitial fibrosis in obstructed mouse kidneys in a Sirt1-independent manner

    Directory of Open Access Journals (Sweden)

    Ren Y

    2015-07-01

    Full Text Available Yunzhuo Ren,* Chunyang Du,* Li Yan, Jingying Wei, Haijiang Wu, Yonghong Shi, Huijun Duan Department of Pathology, Hebei Medical University, Shijiazhuang, Hebei, People’s Republic of China *These authors contributed equally to this work Abstract: Transforming growth factor-β1 (TGF-β1 plays an important role in the pathogenesis and progression of chronic kidney disease. Connective tissue growth factor (CTGF is a critical fibrogenic mediator of TGF-β1. Mammalian sirtuin 1 (Sirt1 is reported to attenuate renal fibrosis by inhibiting the TGF-β1 pathway. This study was designed to detect whether the delivery of CTGF siRNA in vivo directly ameliorates renal fibrosis. Furthermore, the relationship with Sirt1 underlying the protective effect of CTGF siRNA on interstitial fibrosis and apoptosis was explored. Here, we report that the expressions of CTGF and TGF-β1 were increased while Sirt1 expression and activity were both dramatically decreased in mouse kidneys with unilateral ureteral obstruction. Recombinant human TGF-β1 treatment in HK-2 cells increased CTGF levels and remarkably decreased Sirt1 levels and was accompanied by apoptosis and release of fibrosis-related factors. Recombinant human CTGF stimulation also directly induced apoptosis and fibrosis. The CTGF siRNA plasmid ameliorated tubular cell apoptosis and tubulointerstitial fibrosis, but did not affect Sirt1 expression and activity both in vivo and in vitro. Furthermore, overexpression of Sirt1 abolished TGF-β1-induced cell apoptosis and fibrosis, while Sirt1 overexpression suppressed CTGF expression via stimulation by TGF-β1. This study provides evidence that treatment strategies involving the delivery of siRNA targeting potentially therapeutic transgenes may be efficacious. Our results suggest that the decrease in Sirt1 is associated with the upregulated expression of CTGF in renal fibrosis, and may aid in the design of new therapies for the prevention of renal fibrosis. Keywords

  11. Effect of MLP on Expression of CTGF in Renal Tissue of Diabetic Nephropathy Rats%桑叶多糖对DN大鼠肾脏CTGF表达的影响

    Institute of Scientific and Technical Information of China (English)

    宋铁军; 李齐; 薛继婷; 韩智学; 刘洪凤; 王桂云

    2014-01-01

    目的:观察桑叶多糖(mulberry leaves polysaccharide,MLP)对糖尿病肾病(diabetic nephropathy,DN)大鼠肾组织结缔组织生长因子(connective tissue growth factor,CTGF)基因表达的影响,探讨MLP抗肾纤维化的作用机制.方法:将Wistar大鼠随机分为正常对照组、模型组、MLP高、中、低剂量治疗组.采用一次性腹腔注射STZ的方法建立DN大鼠模型.给予不同剂量MLP进行治疗8w后,检测24h UAlb和尿蛋白含量;通过RT-PCR法测定肾脏中CTGF mRNA表达.结果:MLP能显著降低DN大鼠24h UAlb和尿蛋白含量(P<0.01),下调CTGF mRNA表达(P<0.01).结论:MLP可减少DN大鼠尿蛋白、24h UAlb,降低肾脏CTGF mRNA的表达.

  12. Negative regulation of neuromedin U mRNA expression in the rat pars tuberalis by melatonin.

    Directory of Open Access Journals (Sweden)

    Sayaka Aizawa

    Full Text Available The pars tuberalis (PT is part of the anterior pituitary gland surrounding the median eminence as a thin cell layer. The characteristics of PT differ from those of the pars distalis (PD, such as cell composition and gene expression, suggesting that the PT has a unique physiological function compared to the PD. Because the PT highly expresses melatonin receptor type 1, it is considered a mediator of seasonal and/or circadian signals of melatonin. Expression of neuromedin U (NMU that is known to regulate energy balance has been previously reported in the rat PT; however, the regulatory mechanism of NMU mRNA expression and secretion in the PT are still obscure. In this study, we examined both the diurnal change of NMU mRNA expression in the rat PT and the effects of melatonin on NMU in vivo. In situ hybridization and quantitative PCR analysis of laser microdissected PT samples revealed that NMU mRNA expression in the PT has diurnal variation that is high during the light phase and low during the dark phase. Furthermore, melatonin administration significantly suppressed NMU mRNA expression in the PT in vivo. On the other hand, 48 h fasting did not have an effect on PT-NMU mRNA expression, and the diurnal change of NMU mRNA expression was maintained. We also found the highest expression of neuromedin U receptor type 2 (NMUR2 mRNA in the third ventricle ependymal cell layer, followed by the arcuate nucleus and the spinal cord. These results suggest that NMU mRNA expression in the PT is downregulated by melatonin during the dark phase and shows diurnal change. Considering that NMU mRNA in the PT showed the highest expression level in the brain, PT-NMU may act on NMUR2 in the brain, especially in the third ventricle ependymal cell layer, with a circadian rhythm.

  13. Glucocorticoids modulate BDNF mRNA expression in the rat hippocampus after traumatic brain injury.

    Science.gov (United States)

    Grundy, P L; Patel, N; Harbuz, M S; Lightman, S L; Sharples, P M

    2000-10-20

    Brain-derived neurotrophic factor (BDNF) expression in rat hippocampus is increased after experimental traumatic brain injury (TBI) and may be neuroprotective. Glucocorticoids are important regulators of brain neurotrophin levels and are often prescribed following TBI. The effect of adrenalectomy (ADX) on the expression of BDNF mRNA in the hippocampus after TBI has not been investigated to date. We used fluid percussion injury (FPI) and in situ hybridization to evaluate the expression of BDNF mRNA in the hippocampus 4 h after TBI in adrenal-intact or adrenalectomized rats (with or without corticosterone replacement). FPI and ADX independently increased expression of BDNF mRNA. In animals undergoing FPI, prior ADX caused further elevation of BDNF mRNA and this upregulation was prevented by corticosterone replacement in ADX rats. These findings suggest that glucocorticoids are involved in the modulation of the BDNF mRNA response to TBI.

  14. Interleukin-21 mRNA expression during virus infections

    DEFF Research Database (Denmark)

    Holm, Christian; Nyvold, C.G.; Paludan, Søren Riis;

    2006-01-01

    and activational effects of IL-21 on different leukocytes come into play in vivo in an immune response has so far not been fully investigated. We show here for the first time in vivo, that IL-21 mRNA is produced in the spleen when mice are challenged with herpes simplex virus type 2 (HSV-2) or lymphocytic...... choriomeningitis virus (LCMV). We show in HSV-2 challenged mice that this production takes place in CD4+ T cell fractions and is absent in CD4+ T cell-depleted fractions. We also show that the peak of IL-21 mRNA production in both the HSV-2 and LCMV-challenged mice coincides with the onset of the adaptive immune...

  15. Interleukin-21 mRNA expression during virus infections

    DEFF Research Database (Denmark)

    Holm, Christian; Nyvold, Charlotte Guldborg; Paludan, Søren Riis

    2006-01-01

    and activational effects of IL-21 on different leukocytes come into play in vivo in an immune response has so far not been fully investigated. We show here for the first time in vivo, that IL-21 mRNA is produced in the spleen when mice are challenged with herpes simplex virus type 2 (HSV-2) or lymphocytic...... choriomeningitis virus (LCMV). We show in HSV-2 challenged mice that this production takes place in CD4+ T cell fractions and is absent in CD4+ T cell-depleted fractions. We also show that the peak of IL-21 mRNA production in both the HSV-2 and LCMV-challenged mice coincides with the onset of the adaptive immune...... response. Thus, our data suggest a role for IL-21 in the early stages of adaptive immune response against virus infections....

  16. Interleukin-21 mRNA expression during virus infections

    DEFF Research Database (Denmark)

    Holm, Christian; Nyvold, C.G.; Paludan, Søren Riis

    2006-01-01

    and activational effects of IL-21 on different leukocytes come into play in vivo in an immune response has so far not been fully investigated. We show here for the first time in vivo, that IL-21 mRNA is produced in the spleen when mice are challenged with herpes simplex virus type 2 (HSV-2) or lymphocytic...... choriomeningitis virus (LCMV). We show in HSV-2 challenged mice that this production takes place in CD4+ T cell fractions and is absent in CD4+ T cell-depleted fractions. We also show that the peak of IL-21 mRNA production in both the HSV-2 and LCMV-challenged mice coincides with the onset of the adaptive immune...... response. Thus, our data suggest a role for IL-21 in the early stages of adaptive immune response against virus infections....

  17. 酒精性肝纤维化模型大鼠CTGF mRNA和丙二醛的表达及黄芪注射液的干预作用%The expression of CTGFmRNA and MDA in rats with alcoholic hepatic fibrosis and the Intervention of Huangqi injection on them

    Institute of Scientific and Technical Information of China (English)

    张超贤; 郭晓凤; 郭李柯

    2010-01-01

    目的 研究CTGF mRNA和MDA在酒精性肝纤维化(AHF)模型大鼠中的表达及黄芪注射液的干预作用.方法 SD大鼠45只,随机分为正常对照组、模型组、黄芪沣射液组.以乙醇灌胃诱导大鼠肝纤维化模型,治疗组造模同时给予黄芪注射液尾静脉注射,共16周.之后处死大鼠取肝组织标本,光镜观察肝组织的病理变化,放射免疫法测定肝组织丙二醛(MDA)含量,逆转录-聚合酶链反应(RT-PCR)法检测肝组织结缔组织生长因子(CTGF)mRNA表达.结果 与模型组比较,治疗组肝小叶结构破坏明显减轻,肝脏胶原纤维增生亦明显减轻,纤维条索疏松变窄.肝细胞水肿好转,变性改善,炎细胞浸润减少(P<0.01).与模型组比较,肝组织胶原面积、MDA及CTGF mRNA表达显著下降(P<0.01),Masson染色胶原纤维面积和MDA、CTGF mRNA表达呈正相关关系(R_1=0.571,P<0.05;R_2=0.558,P<0.05).结论 黄芪注射液对慢性肝损伤有保护作用,这与其抑制肝组织中CTGF mRNA的表达和抗脂质过氧化作用有关.%Objective To explore the expression of CTGFmRNA and MDA in rats with alcoholic hepatic fibrosis and intervention of Huangqi injection on them.Methods 45 male SD rats were randomly divided into three groups:a normal group,a model group,and a Huangqi injection group.Alcohol was intragastricly administrated for 16 weeks to induce the model of hepatic fibrosis.At the same time of modeling,The Huangqi injection was injected into tail vein of rats in the Huangqi injection group.The rats were killed after 16 weeks.The histomorphylogic structure of the liver tissues were observed under optical microscope;The levels of MDA in liver tissue were determined by radioimmunoassay,and the expressions of connective tissue growth factor(CTGF)mRNA were measured by reverse transcriptase-polymerase chain reaction(RT-PCR).Results Compared with the model group.the destructions and proliferations of collagen fibers were lightened,fiber cords

  18. COX-2 mRNA expression in esophageal squamous cell carcinoma (ESCC) and effect by NSAID.

    Science.gov (United States)

    Liu, X; Li, P; Zhang, S-T; You, H; Jia, J-D; Yu, Z-L

    2008-01-01

    To investigate cyclooxygenase-2 (COX-2) mRNA expression in human esophageal squamous cell carcinoma and the effect of a non-steroidal anti-inflammatory drug (NSAID) on it, in order to explore the mechanism of COX-2 in esophageal squamous cell carcinoma (ESCC) carcinogenesis and the ability of NSAID to prevent or treat ESCC. Frozen specimens of human ESCC and adjacent normal esophageal squamous epithelium pairs (n = 22) were examined for COX-2 mRNA expression by reverse-transcription polymerase chain reaction (RT-PCR). After incubation with aspirin (a non-selective COX inhibitor) or Nimesulide (a selective COX-2 inhibitor), the proliferation status of two human esophageal squamous cancer cell lines, EC-9706 and EC-109, was quantified by 3-(4,5-dimethyl-thiazol-2yl)-2,5-diphenyltetrazolium bromide assay. The expression of COX-2 mRNA in these cells was detected by RT-PCR. COX-2 mRNA was expressed in 12 of 22 (54.5%) ESCC tissue samples, but it was undetectable in all the specimens of adjacent normal esophageal squamous epithelium COX-2 mRNA expression. Both aspirin (5-20 mmol/L) and Nimesulide (0.1-0.8 mmol/L) inhibited EC-9706 cell line proliferation and suppressed its COX-2 mRNA expression dose-dependently. However, only aspirin (5-20 mmol/L) could inhibit proliferation in the EC-109 cell line and suppress COX-2 mRNA expression. Nimesulide (0.1-0.8 mmol/L) could neither inhibit EC-109 cell growth nor suppress COX-2 mRNA expression. COX-2 mRNA expression is a frequent phenomenon in human ESCC tissue samples and plays an important role in the carcinogenesis of ESCC. NSAID may be useful in the chemoprevention and therapy of human ESCC and its effects are likely to be mediated by modulating COX-2 activity.

  19. Expression of hippocampal adrenergic receptor mRNA in a rat model of depression

    Institute of Scientific and Technical Information of China (English)

    Jianbin Zhang; Lingling Wang; Xinjun Wang; Jingfeng Jiang; Xiaoren Xiang; Tianjun Wang

    2011-01-01

    Adrenergic receptor dysfunction is suggested as a potential cause of hippocampal vulnerability to stress-related pathology. We examined mRNA expression of adrenergic receptor (AR) subtypes α1-AR, α2-AR, and β1-AR in hippocampal subregions (CA1, CA3, dentate gyrus) using in situ hybridization in a depression model induced by chronic unpredictable mild stress and social isolation. α1-AR mRNA expression was significantly increased in the CA3 and dentate gyrus, β1-AR mRNA was significantly increased in the CA1, and α2-AR mRNA remained unchanged in all regions of depression rats compared with controls. Thus, different AR subtypes exhibit a differing pattern of mRNA expression in various hippocampal subregions following depression.

  20. Dynamic Analysis of the Expression of the TGFβ/SMAD2 Pathway and CCN2/CTGF during Early Steps of Tooth Development

    Science.gov (United States)

    Pacheco, Marcos S.; Reis, Alice H.; Aguiar, Diego P.; Lyons, Karen M.; Abreu, José G.

    2009-01-01

    Background/Aims CCN2 is present during tooth development. However, the relationship between CCN2 and the transforming growth factor β (TGFβ)/SMAD2/3 signaling cascade during early stages of tooth development is unclear. Here, we compare the expression of CCN2 and TGFβ/SMAD2/3 components during tooth development, and analyze the functioning of TGFβ/SMAD2/3 in wild-type (WT) and Ccn2 null (Ccn2−/−) mice. Methods Coronal sections of mice on embryonic day (E)11.5, E12.5, E13.5, E14.5 and E18.5 from WT and Ccn2−/− were immunoreacted to detect CCN2 and components of the TGFβ signaling pathway and assayed for 5′-bromo-2′-deoxyuridine immunolabeling and proliferating cell nuclear antigen immunostaining. Results CCN2 and TGFβ signaling components such as TGFβ1, TGFβ receptor II, SMADs2/3 and SMAD4 were expressed in inducer tissues during early stages of tooth development. Proliferation analysis in these areas showed that epithelial cells proliferate less than mesenchymal cells from E11.5 to E13.5, while at E14.5 they proliferate more than mesenchymal cells. We did not find a correlation between functioning of the TGFβ1 cascade and CCN2 expression because Ccn2−/− mice showed neither a reduction in SMAD2 phosphorylation nor a difference in cell proliferation. Conclusion CCN2 and the TGFβ/SMAD2/3 signaling pathway are active in signaling centers of tooth development where proliferation is dynamic, but these mechanisms may act independently. PMID:18089935

  1. Dietary glycerol for quail: association between productive performance and COX III mRNA expression.

    Science.gov (United States)

    Silva, S C C; Gasparino, E; Batista, E; Tanamati, F; Vesco, A P D; Lala, B; de Oliveira, D P

    2016-05-25

    This study was carry out to evaluate mRNA expression of mitochondrial cytochrome c oxidase III in the Pectoralis superficialis muscle of 28-day-old quails fed diets containing 0, 8, and 12% glycerol. Total RNA was extracted (N = 10) and cDNA was amplified using specifics primers for qRT-PCR. Feed efficiency and feed intake were evaluated. COX III mRNA expression in breast muscle was higher in the group fed with 12% glycerol (0.863 AU); no differences were observed in the expression of this gene between the muscle of animals fed diets without glycerol (0.357 AU) and 8% glycerol (0.415 AU). Quails that showed greater COX III mRNA expression also showed the lowest feed efficiency. These results show that there is a difference in COX III mRNA expression in breast muscle of 28-day-old quail fed diets different concentrations of glycerol.

  2. Impaired expression of glycogen synthase mRNA in skeletal muscle of NIDDM patients

    DEFF Research Database (Denmark)

    Vestergaard, H; Bjørbaek, C; Andersen, P H

    1991-01-01

    with NIDDM were accompanied by a 39% reduction (P less than 0.02) in the steady state level of GS mRNA per microgram DNA of muscle. In both diabetic and control subjects, the mRNA expression of GS was unaffected after euglycemic-hyperinsulinemic clamp for 4 h. With single-stranded conformation polymorphism...

  3. Impaired expression of glycogen synthase mRNA in skeletal muscle of NIDDM patients

    DEFF Research Database (Denmark)

    Vestergaard, H; Bjørbaek, C; Andersen, P H

    1991-01-01

    with NIDDM were accompanied by a 39% reduction (P less than 0.02) in the steady state level of GS mRNA per microgram DNA of muscle. In both diabetic and control subjects, the mRNA expression of GS was unaffected after euglycemic-hyperinsulinemic clamp for 4 h. With single-stranded conformation polymorphism...

  4. Expression of local renin and angiotensinogen mRNA in cirrhotic portal hypertensive patient

    Institute of Scientific and Technical Information of China (English)

    Li Zhang; Zhen Yang; Bao-Min Shi; Da-Peng Li; Chong-Yun Fang; Fa-Zu Qiu

    2003-01-01

    AIM: To investigate the expression of local renin and angiotensinogen mRNA in cirrhotic portal hypertensive patients.METHODS: The expression of local renin and angiotensinogen mRNA in the liver, splenic artery and vein of PH patients was detected by RT-PCR analysis.RESULTS: Expression of local renin mRNA in the liver of control group was (0.19±0.11), significantly lower than that in splenic artery(0.45±0.17)or splenic vein(0.39±0.12)respectively, (P<0.05). Expression of local angiotensinogen mRNA in the liver was (0.64±0.21), significantly higher than that in splenic artery(0.41±0.15) or in splenic vein (0.35±0.18)respectively, (P<0.05). Expression of local renin mRNA in the liver, splenic artery and vein of PH group was (0.78±0.28),(0.86±0.35) and (0.81±0.22) respectively, significantly higher than that in the control group, (P<0.05). Expression of local angiotensinogen mRNA in the liver, splenic artery and vein of PH group was (0.96±0.25), (0.83±0.18) and (0.79±0.23)respectively, significantly higher than that in the control group,(P<0.05). There was no significant difference between the liver, splenic artery and vein in the expression of local renin or local angiotensinogen mRNA in PH group, (P<0.05).CONCLUSION: In normal subjects the expression of local renin and angiotensinogen mRNA was organ specific, but with increase of the expression of LRAS, the organ-specificity became lost in cirrhotic patients. LRAS may contribute to increased resistance of portal vein with liver and formation of splanchnic vasculopathy.

  5. Astrocyte cultures derived from human brain tissue express angiotensinogen mRNA

    Energy Technology Data Exchange (ETDEWEB)

    Milsted, A.; Barna, B.P.; Ransohoff, R.M.; Brosnihan, K.B.; Ferrario, C.M. (Cleveland Clinic Foundation, OH (USA))

    1990-08-01

    The authors have identified human cultured cell lines that are useful for studying angiotensinogen gene expression and its regulation in the central nervous system. A model cell system of human central nervous system origin expressing angiotensinogen has not previously been available. Expression of angiotensinogen mRNA appears to be a basal property of noninduced human astrocytes, since astrocytic cell lines derived from human glioblastomas or nonneoplastic human brain tissue invariably produced angiotensinogen mRNA. In situ hybridization histochemistry revealed that angiotensinogen mRNA production was not limited to a subpopulation of astrocytes because >99% of cells in these cultures contained angiotensinogen mRNA. These cell lines will be useful in studies of the molecular mechanisms controlling angiotensin synthesis and the role of biologically active angiotensin in the human brain by allowing the authors to examine regulation of expression of the renin-angiotensin system in human astrocyte cultures.

  6. Lipoprotein Lipase mRNA expression in different tissues of farm ...

    African Journals Online (AJOL)

    Lipoprotein Lipase mRNA expression in different tissues of farm animals. ... Lipoprotein lipase (LPL) controls triacylglycerol partitioning between adipose tissues and muscles, so it is important enzyme for ... Article Metrics. Metrics Loading .

  7. Differential expression of IGF-1 mRNA isoforms in colorectal carcinoma and normal colon tissue.

    Science.gov (United States)

    Kasprzak, Aldona; Szaflarski, Witold; Szmeja, Jacek; Andrzejewska, Małgorzata; Przybyszewska, Wiesława; Kaczmarek, Elżbieta; Koczorowska, Maria; Kościński, Tomasz; Zabel, Maciej; Drews, Michał

    2013-01-01

    The insulin-like growth factor (IGF)-1 gene consists of 6 exons resulting in the expression of 6 variant forms of mRNA (IA, IB, IC, IIA, IIB and IIC) due to an alternative splicing. The mechanisms of IGF-1 gene splicing and the role of local expression manifested by IGF-1 mRNA variants in colorectal carcinoma (CRC) have not been extensively investigated. Therefore, the aim of our study was to analyse the expression of IGF-1 mRNA isoforms [A, B, C, P1 (class I) and P2 (class II)], as well as the protein expression in CRC and control samples isolated from 28 patients. The expression of Ki-67 was also analysed and clinical data were obtained. For this purpose, we used quantitative real-time PCR (qPCR) and immunocytochemistry. The expression of mRNAs coding for all splicing isoforms of IGF-1 was observed in every tissue sample studied, with a significantly lower expression noted in the CRC as compared to the control samples. The cytoplasmic expression of IGF-1 protein was found in 50% of the CRC and in ~40% of the non-tumor tissues; however, no significant quantitative inter-group differences were observed. The expression of the IGF-1 gene in the 2 groups of tissues was controlled by the P1 and P2 promoters in a similar manner. No significant differences were detected in the expression of the IGF-1 A and B isoforms; however, their expression was significantly higher compared to that of isoform C. No significant differences were observed between the expression of Ki-67 mRNA in the CRC and control tissue even though the expression of the Ki-67 protein was higher in the CRC compared to the control samples. Ki-67 protein expression was associated with the macroscopic and microscopic aspects of CRC. A significant positive correlation was found between the local production of total mRNA and isoform A and the expression of Ki-67 mRNA, although only in the non-tumor tissues. In CRC samples, the local expression of the total IGF-1 mRNA and all splicing isoforms of IGF-1 mRNA

  8. Methylation Inactivation Mechanism of Parkin Gene mRNA Expression in Nasopharyngeal Carcinoma

    Institute of Scientific and Technical Information of China (English)

    Ni Haifeng; Jiang Bo; Zhou Zhen; Li Yong; Huang Guangwu

    2014-01-01

    Objective:To investigate the methylation inactivation and the clinical signiifcance of Parkin gene mRNA expression in nasopharyngeal carcinoma (NPC). Methods: The methylation-speciifc PCR (MSP) and semi-quantitative reverse transcription PCR (RT-PCR) were used to detect methylation and the mRNA expression level of Parkin gene in 54 cases of NPC tissues and 16 cases of normal nasopharyngeal epithelial (NNE) tissues.The mRNA expression of Parkin gene in two NPC cell lines (CNE1 and CNE2) were detected before and after treatment with the methyltransferase inhibitor 5-aza-2-deoxycytidine so as to analyze the effects of Parkin gene methylation and demethylation on Parkin gene mRNA expression and the relationship between Parkin gene mRNA expression and clinical factors. Results: The methylation frequency of Parkin gene in human NPC tissues was 62.96% (34/54), but didn’t happen in any of 16 cases of NNE tissues. The mRNA expression level was (0.3430±0.4947) in 54 cases of NPC tissues and (1.0052±0.4911) in NNE tissues, showing that the mRNA expression level of NPC tissues was significantly down-regulated (P0.05), but was closely related to lymph node metastasis (P<0.05). Conclusion:Parkin gene mRNA expression, serving as a cancer suppressor gene in the occurrence and development of NPC, is inactivated and regulated by methylation, which also has a negative correlation with lymph node metastasis and could be considered as the judgment of predictive index of clinical prognosis of NPC.

  9. Increased IL-10 mRNA and IL-23 mRNA expression in multiple sclerosis: interferon-beta treatment increases IL-10 mRNA expression while reducing IL-23 mRNA expression

    DEFF Research Database (Denmark)

    Krakauer, M.; Sorensen, P.; Khademi, M.

    2008-01-01

    of the regulatory cytokine IL-10. The elevated IL-23 mRNA levels in MS patients are noteworthy in view of the newly discovered IL-23-driven Th17 T-cell subset, which is crucial in animal models of MS. Since IFN-beta therapy resulted in decreased IL-23 mRNA levels, the Th17 axis could be another target of IFN...

  10. Carboxylesterase 1 gene duplication and mRNA expression in adipose tissue are linked to obesity and metabolic function

    DEFF Research Database (Denmark)

    Friedrichsen, Martin; Poulsen, Pernille; Wojtaszewski, Jørgen

    2013-01-01

    involved in the control of mRNA expression. Here, we investigated mRNA expression level in adipose tissue and its association with measures of adiposity and metabolic function in a population of elderly twins. Furthermore, the heritability of mRNA expression level in adipose tissue and the effect of gene...

  11. An Experimental Study on the Expression of Heme Oxygenase-2 mRNA in Hirschsprung's Disease

    Institute of Scientific and Technical Information of China (English)

    朱珉; 魏明发; 刘芳

    2002-01-01

    Summary: In order to investigate the relationship between the expression of heme oxygenase-2 (HO-2) mRNA and the pathogenesis of Hirschsprung's disease (HD), total ribonucleic acid (RNA) was extracted in the aganglionic and ganglionic segments of colon respectively from 15 cases of HD. The single-stranded cDNA of HO-2 was synthesized and further amplified by reverse transcription-poly merase chain reaction (RT-PCR). The expression of HO-2 mRNA was normal in ganglionic seg ments, but absent in aganglionic segments. It is concluded that the absence of HO-2 mRNA expres sion may be an important mechanism responsible for HD.

  12. Connecting protein and mRNA burst distributions for stochastic models of gene expression

    CERN Document Server

    Elgart, Vlad; Fenley, Andrew T; Kulkarni, Rahul V

    2011-01-01

    The intrinsic stochasticity of gene expression can lead to large variability in protein levels for genetically identical cells. Such variability in protein levels can arise from infrequent synthesis of mRNAs which in turn give rise to bursts of protein expression. Protein expression occurring in bursts has indeed been observed experimentally and recent studies have also found evidence for transcriptional bursting, i.e. production of mRNAs in bursts. Given that there are distinct experimental techniques for quantifying the noise at different stages of gene expression, it is of interest to derive analytical results connecting experimental observations at different levels. In this work, we consider stochastic models of gene expression for which mRNA and protein production occurs in independent bursts. For such models, we derive analytical expressions connecting protein and mRNA burst distributions which show how the functional form of the mRNA burst distribution can be inferred from the protein burst distributio...

  13. TGF beta 2 mRNA expression and pregnancy failure in mice.

    Science.gov (United States)

    Gorivodsky, M; Torchinsky, A; Zemliak, I; Savion, S; Fein, A; Toder, V

    1999-08-01

    We describe here a pattern of transforming growth factor (TGF) beta2 mRNA expression at the fetomaternal interface in mice with high rate of resorptions as well as its expression following maternal immunopotentiation. TGF beta 2 mRNA expression was evaluated in the uteroplacental units of mice with spontaneous (CBA/J x DBA/2J mouse combination) or cyclophosphamide (CP)-induced pregnancy loss. The effect of immunopotentiation on TGF beta 2 mRNA expression was determined in CP-treated females who underwent nonspecific immunostimulation with xenogeneic (rat) leukocytes. A quantitative analysis of TGF beta 2 mRNA level was performed using RNase protection assay. Distribution of TGF beta 2 mRNA transcripts at the fetomaternal interface was studied by in situ hybridization analysis. RNase protection analysis revealed four TGF beta 2 specific mRNA forms (330, 270, 230, and 170 bp) in the uteroplacental units of mice with either normal or decreased reproductive performance. A significant decrease (about 50%) in the level of TGF beta 2 mRNA was registered in the uteroplacental unit of mice with pregnancy loss as compared to the control mice. TGF beta 2 transcripts were abundant in the uterine epithelium and stroma. A specific hybridization signal was detected also in metrial gland cells and it was found to be substantially lower in CP-treated as compared to intact mice. In the resorbing uteroplacental unit, the expression of TGF beta 2 mRNA was completely lost in the uterine epithelium, and the number of TGF beta 2 mRNA-positive metrial gland cells was lower as compared to the control. Immunopotentiation decreased the resorption rate in mice with CP-induced pregnancy loss and caused a dramatic increase in TGF beta 2 mRNA expression: the level of TGF beta 2 mRNA was found to be higher by 2.0-3.2 fold in the uteroplacental unit of immunized as compared to nonimmunized CP-treated mice. These data suggest that distortion of TGF beta 2 expression at the fetomaternal interface

  14. Expression of TLR9 and Its mRNA in the Lesions of Lichen Planus

    Institute of Scientific and Technical Information of China (English)

    LI Jiawen; CHEN Jing; TAN Zhijian; LIU Houjun; LIU Zhixiang

    2007-01-01

    To investigate the role of toll-like receptor 9 (TLR9) in the pathogenesis of lichen planus,the expressions of TLR9 and its mRNA in the lesional skin of lichen planus were detected by immunohistochemical technique (SP) and RT-PCR. As control, normal skin of healthy volunteers was also tested. The immunohistochemical study showed that the expression of TLR9 in the lesional skin of lichen planus was significantly higher than that in the normal controls. The results of RT-PCR showed that both skin lesions and normal controls had TLR9 expression. In skin lesions, the expression level of TLR9 mRNA was 1.6075±0.0930, which was significantly higher than that in normal controls (P<0.001). These findings indicated that up-regulated expression of TLR9 and its mRNA might be involved in the pathogenesis of lichen planus.

  15. Histone gene expression and histone mRNA 3' end structure in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Pettitt Jonathan

    2007-06-01

    Full Text Available Abstract Background Histone protein synthesis is essential for cell proliferation and required for the packaging of DNA into chromatin. In animals, histone proteins are provided by the expression of multicopy replication-dependent histone genes. Histone mRNAs that are processed by a histone-specific mechanism to end after a highly conserved RNA hairpin element, and lack a poly(A tail. In vertebrates and Drosophila, their expression is dependent on HBP/SLBP that binds to the RNA hairpin element. We showed previously that these cis and trans acting regulators of histone gene expression are conserved in C. elegans. Here we report the results of an investigation of the histone mRNA 3' end structure and of histone gene expression during C. elegans development. Results Sequence analysis of replication-dependent histone genes revealed the presence of several highly conserved sequence elements in the 3' untranslated region of histone pre-mRNAs, including an RNA hairpin element and a polyadenylation signal. To determine whether in C. elegans histone mRNA 3' end formation occurs at this polyadenylation signal and results in polyadenylated histone mRNA, we investigated the mRNA 3' end structure of histone mRNA. Using poly(A selection, RNAse protection and sequencing of histone mRNA ends, we determined that a majority of C. elegans histone mRNAs lack a poly(A tail and end three to six nucleotides after the hairpin structure, after an A or a U, and have a 3' OH group. RNAi knock down of CDL-1, the C. elegans HBP/SLBP, does not significantly affect histone mRNA levels but severely depletes histone protein levels. Histone gene expression varies during development and is reduced in L3 animals compared to L1 animals and adults. In adults, histone gene expression is restricted to the germ line, where cell division occurs. Conclusion Our findings indicate that the expression of C. elegans histone genes is subject to control mechanisms similar to the ones in other

  16. mRNA expression of Rho GTPase-related signaling molecules during rat hippocampal development

    Institute of Scientific and Technical Information of China (English)

    Guoqing Guo; Jifeng Zhang; Li Xin; Jing Chen; Weizai Shen; Lin Yuan; Shizhen Zhong

    2009-01-01

    BACKGROUND:Rho GTPase family members have been shown to participate in neurite growth by regulating the neuronal cytoskeleton.However,there are very few reports of developmental roles of signaling molecules related to Rho GTPases.OBJECTIVE:To investigate messenger ribonucleic acid mRNA expression of signaling molecules associated with Rho GTPases,including Rho-A,Rac-1,collapsin response mediator protein 1(CRMP-1),and tubulin β3 (Tub β3) during rat hippocampus development.DESIGN,TIME AND SETTING:A non-randomized,controlled,animal experiment,based on different developmental stages of the rat hippocampus,was performed at the Guangdong Key Laboratory of Tissue Construction and Detection,Institute of Clinical Anatomy,Southern Medical University between December 2005 and July 2007.MATERIALS:Trizol reagent was purchased from Invitrogen,USA.RNA PCR kit (AMV) Ver 3.0 and 150 bp DNA Ladder Marker were purchased from TaKaRa,Japan.Unless otherwise specified,all other reagents were purchased from Sigma-Aldrich,USA.METHODS:Twenty-five Sprague Dawley rats were assigned to five groups (n=5) according to developmental stages:embryonic (embryonic 15 days),neonatal (postnatal 5 days),juvenile (postnatal 1 month),adult (postnatal 3 months),and senile (postnatal 18 months).MAIN OUTCOME MEASURES:Detection of mRNA expression of Rho-A,Rac-1,CRMP-1,and Tub β3 during various hippocampal developmental stages by reverse-transcription polymerase chain reaction.RESULTS:Hippocampal mRNA expression of Rho-A,as well as Rac-1,reached peak levels at embryonic,juvenile,and senile stages,and was relatively less during neonatal and adult stages.mRNA expression of Rac-1 was greater than Rho-A during each hippocampal developmental stage.CRMP-1 mRNA expression levels were as follows:embryonic>neonatal>juvenile>adult<senile,while Tub β3 mRNA expression was embryonic>neonatal>juvenile>adult=senile.CONCLUSION:Rho-A and Rac-1 shared similar expression profiles,which demonstrated similar

  17. Single-cell detection of mRNA expression using nanofountain-probe electroporated molecular beacons.

    Science.gov (United States)

    Giraldo-Vela, Juan P; Kang, Wonmo; McNaughton, Rebecca L; Zhang, Xuemei; Wile, Brian M; Tsourkas, Andrew; Bao, Gang; Espinosa, Horacio D

    2015-05-01

    New techniques for single-cell analysis enable new discoveries in gene expression and systems biology. Time-dependent measurements on individual cells are necessary, yet the common single-cell analysis techniques used today require lysing the cell, suspending the cell, or long incubation times for transfection, thereby interfering with the ability to track an individual cell over time. Here a method for detecting mRNA expression in live single cells using molecular beacons that are transfected into single cells by means of nanofountain probe electroporation (NFP-E) is presented. Molecular beacons are oligonucleotides that emit fluorescence upon binding to an mRNA target, rendering them useful for spatial and temporal studies of live cells. The NFP-E is used to transfect a DNA-based beacon that detects glyceraldehyde 3-phosphate dehydrogenase and an RNA-based beacon that detects a sequence cloned in the green fluorescence protein mRNA. It is shown that imaging analysis of transfection and mRNA detection can be performed within seconds after electroporation and without disturbing adhered cells. In addition, it is shown that time-dependent detection of mRNA expression is feasible by transfecting the same single cell at different time points. This technique will be particularly useful for studies of cell differentiation, where several measurements of mRNA expression are required over time.

  18. Effect of taurine on mRNA expression of thioredoxin interacting protein in Caco-2 cells.

    Science.gov (United States)

    Gondo, Yusuke; Satsu, Hideo; Ishimoto, Yoko; Iwamoto, Taku; Shimizu, Makoto

    2012-09-28

    Taurine (2-aminoethanesulfonic acid), a sulfur-containing β-amino acid, plays an important role in several essential biological processes; although, the underlying mechanisms for these regulatory functions remain to be elucidated, especially at the genetic level. We investigated the effects of taurine on the gene expression profile in Caco-2 cells using DNA microarray. Taurine increased the mRNA expression of thioredoxin interacting protein (TXNIP), which is involved in various metabolisms and diseases. β-Alanine or γ-aminobutyric acid (GABA), which are structurally or functionally related to taurine, did not increase TXNIP mRNA expression. These suggest the expression of TXNIP mRNA is induced specifically by taurine. β-Alanine is also known to be a substrate of taurine transporter (TAUT) and competitively inhibits taurine uptake. Inhibition of taurine uptake by β-alanine eliminated the up-regulation of TXNIP, which suggests TAUT is involved in inducing TXNIP mRNA expression. The up-regulation of TXNIP mRNA expression by taurine was also observed at the protein level. Furthermore, taurine significantly increased TXNIP promoter activity. Our present study demonstrated the taurine-specific phenomenon of TXNIP up-regulation, which sheds light on the physiological function of taurine. Copyright © 2012 Elsevier Inc. All rights reserved.

  19. 丝胶对糖尿病大鼠肾脏细胞外基质代谢相关因子PDGF和CTGF表达的调节作用%Regulating Effects of Sericin on ECM Related Factors PDGF and CTGF Expression in Kidney of Diabetes Mellitus Rats

    Institute of Scientific and Technical Information of China (English)

    宋成军; 苗光新; 李健; 付文亮; 杨振军; 陈志宏

    2013-01-01

    Objective To investigate the regulating effects of sericin on expression of extracellular matrix (ECM) related factors plateletderived growth factor (PDGF) and connective tissue growth factor (CTGF) in kidney of type 2 diabetes mellitus rats.Methods A total of 48 adult healthy male SD rats were randomly divided into 4 groups (n =12):normal control group,diabetes mellitus model group,sericin treatment group and positive control group.Type 2 diabetes mellitus rat model was established by injecting streptozotocin into the peritoneal cavity continuously,and then the rats were respectively lavaged with sericin (2.4 g·kg 1·d-1) in sericin treatment group and metformin (55.33 mg·kg-1·d-1) in positive control group.The type Ⅰ collagen (Ⅰ-C) and fibronectin (FN) content in serum of rats were detected by ELISA.Western Blotting and RT-PCR were used to determine the expression of PDGF and CTGF in kidney.Results Compared with rats in diabetes mellitus model group,the 24 hours urine protein,Ⅰ-C and FN content in serum of rats in sericin treatment group was decreased obviously; the expression of PDGF and CTGF in rat kidney decreased significantly in sericin treatment group (P < 0.01,P < 0.05).Conclusion Sericin can recover the balance of ECM synthesis and degradation by down-regulation of the PDGF and CTGF expression in kidney of diabetes mellitus rats,which further delay the development of glomerulosclemsis and renal interstitial fibrosis and protect kidney injury during diabetes mellitus.%目的 探讨丝胶对2型糖尿病大鼠肾脏细胞外基质(ECM)代谢相关因子血小板衍生生长因子(PDGF)和结缔组织生长因子(CTGF)表达的调节作用.方法 健康成年雄性SD大鼠48只,随机分为正常对照组、糖尿病模型组、丝胶治疗组和阳性对照组(n=12).链脲佐菌素腹腔注射建立2型糖尿病大鼠模型,丝胶治疗组大鼠按照2.4 g·kg-1·d-1的剂量灌胃给予丝胶、阳性对照组大鼠按照55.33 mg·kg·d-1的

  20. mRNA EXPRESSION OF PTEN AND VEGF GENES IN EPITHELIAL OVARIAN CANCER

    Institute of Scientific and Technical Information of China (English)

    陈颖; 赵雨杰; 郑华川; 杨雪飞; 汪桂兰; 辛彦

    2003-01-01

    Objective: To investigate the mRNA expression of PTEN and vascular endothelial growth factor (VEGF) genes in ovarian cancer. Methods:We examined mRNA expression of PTEN and VEGF165 in normal ovary (n=5), ovarian cyst (n=5), ovarian borderline tumor (n=9), epithelial ovarian cancer (n=60) and ovarian cancer cell line (CAOV-3) by RT-PCR. Their expressions were compared with clinicopathological features of ovarian cancer. The relationship between their expressions was concerned in all ovarian samples as well. Results:mRNA expression level of PTEN gene was significantly lower in ovarian borderline tumor or ovarian cancer than that in normal ovary or ovarian cyst(P<0.05). It was negatively correlated with clinicopathological staging(P<0.05),whereas positively with histological differentiation (P<0.05). mRNA expression level of PTEN gene was significantly lower in ovarian endometrioid cancer than ovarian serous or mucinous cancer(P<0.05). mRNA expression level of VEGF165 gene was significantly higher in ovarian cancer than that in normal ovary or ovarian cyst(P<0.05). It was positively correlated with clinicopathological staging(P<0.05), whereas negatively with histological differentiation (P<0.05). mRNA expression level of VEGF165 gene was significantly higher in ovarian serous cancer than in other ovarian epithelial cancers (P<0.05). mRNA expression of VEGF165 gene was inversely correlated with mRNA expression level of PTEN gene. Conclusion:Down-regulated expression of PTEN and up-regulated expression of VEGF were considered as two important events in tumorigenesis of ovarian cancer and could be used as molecular markers to indicate the pathobiological behaviors of ovarian cancer. Decreased PTEN expression and increased VEGF expression were closely associated with tumorigenesis and pathobiological behaviors of ovarian endometrioid and serous cancer respectively. Reduced expression of PTEN gene might be involved in carcinogenesis and progression of ovarian cancer by

  1. BUB1 mRNA is significantly co-expressed with AURKA and AURKB mRNA in advanced-stage ovarian serous carcinoma.

    Science.gov (United States)

    Davidson, Ben; Nymoen, Dag Andre; Elgaaen, Bente Vilming; Staff, Anne Cathrine; Tropé, Claes G; Kærn, Janne; Reich, Reuven; Falkenthal, Thea E Hetland

    2014-06-01

    The objective of this study was to investigate the expression and clinical role of the spindle checkpoint kinase budding uninhibited by benzimidazole 1 (Bub1) in primary and metastatic advanced-stage ovarian serous carcinoma. BUB1 mRNA expression was analyzed in 178 tumors (88 effusions, 38 primary carcinomas, and 52 solid metastases) from 144 patients with advanced-stage disease using quantitative real-time polymerase chain reaction (PCR). Bub1 protein expression by Western blotting was studied in 63 carcinomas (30 effusions and 33 solid lesions). BUB1 mRNA expression at different anatomic sites was studied for association with clinicopathologic parameters, including chemotherapy resistance and survival. BUB1 mRNA was universally expressed in serous carcinomas, irrespective of anatomic site. BUB1 mRNA levels were uniformly low in six ovarian surface epithelium specimens analyzed for comparative purposes. Bub1 protein was expressed in 22/30 effusions and 28/33 solid lesions. BUB1 mRNA expression was significantly higher in chemo-naïve primary carcinomas and solid metastases compared to specimens obtained following neoadjuvant chemotherapy (p cancer. BUB1 mRNA levels are lower following chemotherapy exposure in solid lesions, though its presence is unrelated to clinical behavior including response to chemotherapy and survival. BUB1 is co-expressed with AURKA and AURKB suggesting biological relationship between these spindle cell components.

  2. Tuning protein expression using synonymous codon libraries targeted to the 5' mRNA coding region

    DEFF Research Database (Denmark)

    Goltermann, Lise; Borch Jensen, Martin; Bentin, Thomas

    2011-01-01

    In bacteria, the 5' mRNA coding region plays an important role in determining translation output. Here, we report synthetic sequences that when placed in the 5'-mRNA coding region, leading to recombinant proteins containing short N-terminal extensions, virtually abolish, enhance or produce...... intermediate expression levels of green fluorescent protein in Escherichia coli. At least in one case, no apparent effect on protein stability was observed, pointing to RNA level effects as the principal reason for the observed expression differences. Targeting a synonymous codon library to the 5' coding...... and hence is important to recombinant and, most certainly, endogenous gene expression....

  3. [The expression of human telomerase reverse transcriptase mRNA and its significance in acute leukemia].

    Science.gov (United States)

    Meng, Xiao-Li; Lin, Mao-Fang; Jin, Jie

    2003-02-01

    To investigate the expression of hTERT mRNA in bone marrow mononuclear cells (MNCs) from acute leukemia patients, the method of semi-quntitative RT-PCR was used to examine the expression of hTERT mRNA in marrow MNCs, and the telomerase activity of marrow MNCs was determined with the method of TRAP-PCR-ELISA by using a commercial kit. The results indicated that the expression of hTERT mRNA of marrow MNCs in 30 untreated AL patients was markedly higher than that in 12 CR cases (0.71 +/- 0.34 vs 0.43 +/- 0.25, P < 0.05) and 6 normal volunteers (0.71 +/- 0.34 vs 0.22 +/- 0.21, P < 0.01), respectively. Telomerase activity of marrow MNCs in 30 untreated AL patients was significantly higher than that in 12 CR cases (0.235 +/- 0.395 vs 0.012 +/- 0.015, P = 0.007). Moreover, there was a positive correlation between the hTERT mRNA synthesis and telomerase activity in AL cells (r = 0.421, P < 0.01). The pencentage of blast cells in marrow smear of the untreated AL patients was positively correlated with both the expression of hTERT mRNA and the telomerase activity of bone marrow MNCs (r = 0.457, P < 0.05 and r = 0.411, P < 0.05), respectively. It is concluded that the expression of hTERT mRNA in bone marrow MNCs from untreated AL patients was correlated with their telomerase activity. It is suggested that the expression of hTERT mRNA leukemic cells indicates their higher proliferation ability.

  4. Amygdala kindling increases fear responses and decreases glucocorticoid receptor mRNA expression in hippocampal regions.

    Science.gov (United States)

    Kalynchuk, Lisa E; Meaney, Michael J

    2003-12-01

    Amygdala kindling dramatically increases fearful behavior in rats. Because kindling-induced fear increases in magnitude as rats receive more stimulations, kindling provides an excellent model for studying the nature and neural mechanisms of fear sensitization. In the present experiment, we studied whether the development of kindling-induced fear is related to changes in glucocorticoid receptor (GR) mRNA expression in various brain regions. Rats received 20, 60 or 100 amygdala kindling stimulations or 100 sham stimulations. One day after the final stimulation, their fearful behavior was assessed in an unfamiliar open field. Then, the rats were sacrificed and their brains were processed for in situ hybridization of GR mRNA expression. We found that compared with the sham-stimulated rats, the rats that received 60 or 100 kindling stimulations were significantly more fearful in the open field and also had significantly less GR mRNA expression in the dentate gyrus and CA1 subfield of the hippocampus. Importantly, the changes in fearful behavior were significantly correlated with the changes in GR mRNA expression. These results suggest that alterations in GR mRNA expression in hippocampal regions may play a role in the development of kindling-induced fear.

  5. Effect of Interventional Therapy on the Expression of Survivin mRNA in Cervical Cancer.

    Science.gov (United States)

    Wang, Shao-Guang; Mu, Nan; Sun, Hai-Yan

    2017-08-01

    To examine the expression of survivin mRNA in cervical squamous cancer at different time points during interventional therapy in order to find the optimal time point for continual treatment. Fifty patients with stage IB2-IIB cervical squamous cancer receiving transarterial infusion chemotherapy and chemoembolization were included in the present study. The expression of survivin mRNA in cancer samples before and after interventional therapy (on days 7, 14 and 21) were examined by reverse transcription-polymerase chain reaction. The expression of survivin mRNA in cancer samples before the interventional therapy was significantly higher than that of any time point after the interventional therapy (ptreatment, the expression of survivin mRNA decreased until day 14. The expression of survivin mRNA was inhibited by the interventional therapy. It seems that day 14 after interventional treatment is the right time point for continuation of treatment. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  6. Drosophila glutamate receptor mRNA expression and mRNP particles.

    Science.gov (United States)

    Ganesan, Subhashree; Karr, Julie E; Featherstone, David E

    2011-01-01

    The processes controlling glutamate receptor expression early in synaptogenesis are poorly understood. Here, we examine glutamate receptor (GluR) subunit mRNA expression and localization in Drosophila embryonic/larval neuromuscular junctions (NMJs). We show that postsynaptic GluR subunit gene expression is triggered by contact from the presynaptic nerve, approximately halfway through embryogenesis. After contact, GluRIIA and GluRIIB mRNA abundance rises quickly approximately 20-fold, then falls within a few hours back to very low levels. Protein abundance, however, gradually increases throughout development. At the same time that mRNA levels decrease following their initial spike, GluRIIA, GluRIIB, and GluRIIC subunit mRNA aggregates become visible in the cytoplasm of postsynaptic muscle cells. These mRNA aggregates do not colocalize with eIF4E, but nevertheless presumably represent mRNP particles of unknown function. Multiplex FISH shows that different GluR subunit mRNAs are found in different mRNPs. GluRIIC mRNPs are most common, followed by GluRIIA and then GluRIIB mRNPs. GluR mRNP density is not increased near NMJs, for any subunit; if anything, GluR mRNP density is highest away from NMJs and near nuclei. These results reveal some of the earliest events in postsynaptic development and provide a foundation for future studies of GluR mRNA biology.

  7. Myxovirus Resistance Protein A mRNA Expression Kinetics in Multiple Sclerosis Patients Treated with IFNβ

    Science.gov (United States)

    Libertinova, Jana; Meluzinova, Eva; Tomek, Ales; Horakova, Dana; Kovarova, Ivana; Matoska, Vaclav; Kumstyrova, Simona; Zajac, Miroslav; Hyncicova, Eva; Liskova, Petra; Houzvickova, Eva; Martinkovic, Lukas; Bojar, Martin; Havrdova, Eva; Marusic, Petr

    2017-01-01

    Introduction Interferon-β (IFNß) is the first-line treatment for relapsing-remitting multiple sclerosis. Myxovirus resistance protein A (MxA) is a marker of IFNß bioactivity, which may be reduced by neutralizing antibodies (NAbs) against IFNß. The aim of the study was to analyze the kinetics of MxA mRNA expression during long-term IFNβ treatment and assess its predictive value. Methods A prospective, observational, open-label, non-randomized study was designed in multiple sclerosis patients starting IFNß treatment. MxA mRNA was assessed prior to initiation of IFNß therapy and every three months subsequently. NAbs were assessed every six months. Assessment of relapses was scheduled every three months during 24 months of follow up. The disease activity was correlated to the pretreatment baseline MxA mRNA value. In NAb negative patients, clinical status was correlated to MxA mRNA values. Results 119 patients were consecutively enrolled and 107 were included in the final analysis. There was no correlation of MxA mRNA expression levels between baseline and month three. Using survival analysis, none of the selected baseline MxA mRNA cut off points allowed prediction of time to first relapse on the treatment. In NAb negative patients, mean MxA mRNA levels did not significantly differ in patients irrespective of relapse status. Conclusion Baseline MxA mRNA does not predict the response to IFNß treatment or the clinical status of the disease and the level of MxA mRNA does not correlate with disease activity in NAb negative patients. PMID:28081207

  8. Developmental expression of parvalbumin mRNA in the cerebral cortex and hippocampus of the rat.

    Science.gov (United States)

    de Lecea, L; del Río, J A; Soriano, E

    1995-08-01

    Parvalbumin (PARV) belongs to the family of calcium-binding proteins bearing the EF hand domain. Immunocytochemical studies in the cerebral cortex have demonstrated that neurons containing PARV include two types of GABAergic interneurons, namely, basket and axo-axonic chandelier cells. The present study examines the onset and pattern of PARV mRNA expression during the development of rat neocortex and hippocampus by means of 'in situ' hybridization with an oligonucleotide probe corresponding to rat PARV cDNA. In animals aged P0-P6 no signal was detected above background in neocortex or hippocampus. At P8, a few cortical cells displayed a number of silver grains just above background levels. By P10 PARV mRNA-expressing cells in the neocortex were detected almost exclusively in layer V of somatosensory, frontal and cingulate cortices. At P12 PARV mRNA was mainly detected in layers IV, V and VIa. By P14 there was a marked overall increase in the entire neocortex, including layer II-III, both in the number of cells and in their intensity of labelling. Further maturation in the pattern of PARV mRNA concentration was observed between P16 and P21. In the hippocampus low hybridization was observed at P10-P12. In subsequent stages both the number of positive cells and the intensity of labelling increased steadily. No clear-cut radial gradients for the expression of PARV mRNA were observed in the hippocampal region. Our results show that the developmental radial gradient followed by PARV mRNA expression in the neocortex does not follow an 'inside-out' gradient, consistent with previous immunocytochemical findings. Taken together, these data indicate that the developmental sequence followed by the PARV protein directly reflects mRNA abundance and suggest that PARV mRNA expression correlates with the functional maturation of cortical interneurons.

  9. Quantitation of HDAC1 mRNA expression in invasive carcinoma of the breast

    Institute of Scientific and Technical Information of China (English)

    Zhenhuan Zhang; Hirotaka Iwase; Hiroko Yamashita; Tatsuya Toyama; Hiroshi Sugiura; Yoshiaki Ando; Keiko Mita; Maho Hamaguchi; Yasuo Hara; Shunzo Kobayashi

    2006-01-01

    Estrogen is well-established as a mitogenic factor implicated in the tumorigenesis and progression of breast cancer via its binding to the estrogen receptor a(ERα). Recent data indicate that chromatin inactivation mediated by histone deacetylation(HDAC) and DNA methylation is a critical component of ERα silencing in human breast cancer cells. The aim of this study was to determine the expression of the HDAC1 gene in malignant human breast tissue and to correlate our observations with available clinical information. In the present study, the level of expression of HDAC1 mRNA was assessed by LightCycler-based quantitative real-time reverse transcriptase (RT)-PCR analvsis in 162 cases of invasive carcinoma of the breast. Associations between HDAC1 mRNA expression and different clinicopathological factors were sought. It was found that HDAC1 mRNA was expressed at significantly higher levels in tumors from patients over 50 years of age and in those tumors without axillary lymph node involvement, that are less than 2 cm, that are of a non-high histological grade, that are HER2 negative and that are ERα/PgR positive. Patients with tumors displaying high levels of HDAC1 mRNA expression tended to have a better prognosis in terms of both disease-free and overall survival. However, univariate and multivariate analysis did not show HDAC1 mRNA expression level to be an independent prognostic factor for either disease-free or overall survival. These results imply that HDAC1 mRNA expression could have potential as an endocrine response marker and may have prognostic implications for breast cancer progression.

  10. Expression of Npas4 mRNA in telencephalic areas of adult and postnatal mouse brain

    Directory of Open Access Journals (Sweden)

    Joanne C Damborsky

    2015-11-01

    Full Text Available The transcription factor neuronal PAS domain-containing protein 4 (Npas4 is an inducible immediate early gene which regulates the formation of inhibitory synapses, and could have a significant regulatory role during cortical circuit formation. However, little is known about basal Npas4 mRNA expression during postnatal development. Here, postnatal and adult mouse brain sections were processed for isotopic in situ hybridization using an Npas4 specific cRNA antisense probe. In adults, Npas4 mRNA was found in the telencephalon with very restricted or no expression in diencephalon or mesencephalon. In most telencephalic areas, including the anterior olfactory nucleus (AON, piriform cortex, neocortex, hippocampus, dorsal caudate putamen (CPu, septum and basolateral amygdala nucleus (BLA, basal Npas4 expression was detected in scattered cells which exhibited strong hybridization signal. In embryonic and neonatal brain sections, Npas4 mRNA expression signals were very low. Starting at postnatal day 5 (P5, transcripts for Npas4 were detected in the AON, CPu and piriform cortex. At P8, additional Npas4 hybridization was found in CA1 and CA3 pyramidal layer, and in primary motor cortex. By P13, robust mRNA expression was located in layers IV and VI of all sensory cortices, frontal cortex and cingulate cortex. After onset of expression, postnatal spatial mRNA distribution was similar to that in adults, with the exception of the CPu, where Npas4 transcripts became gradually restricted to the most dorsal part. In conclusion, the spatial distribution of Npas4 mRNA is mostly restricted to telencephalic areas, and the temporal expression increases with developmental age during postnatal development, which seem to correlate with the onset of activity-driven excitatory transmission.

  11. Expression of Nogo-A mRNA after injury of the rat central nervous system

    Institute of Scientific and Technical Information of China (English)

    Xigao Guo; Yang Guo; Tao Huang

    2008-01-01

    BACKGROUND: Nogo protein has been identified as an inhibitor of axonal growth, which was highly expressed in central nervous system; however, there are only a few studies on changes of Nogo-A expression following central nervous system injury.OBJECTIVE: To investigate the dynamic expression of Nogo-A mRNA after rat central nervous system injury.DESIGN: Randomized controlled animal study.MATERIALS: Thirty-five rats were randomly divided into two groups, normal animal group (n = 5) and model group (n = 30). The model group was then divided into six subgroups at six time points: 12, 24 hours and 3, 9, 15, and 21 days post-injury, with five rats in each subgroup.METHODS: The left parietal lobe of rats was contused by free-fall strike, and total RNA was extracted from the entire brain tissue. Semi-quantitative RT-PCR was used to detect Nogo-A mRNA expression, and the ratio between expression of the target gene and glyceraldehyde phosphate dehydrogenase was used to determine the relative expression level.MAIN OUTCOME MEASURES: To determine whether Nogo-A mRNA expression was higher than usual following brain injury.RESULTS: The level of Nogo-A mRNA started to increase 12 hours after injury (P 0.05).CONCLUSION: After injury of the central nervous system, Nogo-A may play a pivotal role in obstructing regeneration of the nerve.

  12. Visfatin mRNA expression in human subcutaneous adipose tissue is regulated by exercise

    DEFF Research Database (Denmark)

    Frydelund-Larsen, Lone; Åkerström, Thorbjörn; Nielsen, Søren

    2006-01-01

    Visfatin [pre-beta-cell colony-enhancing factor (PBEF)] is a novel adipokine that is produced by adipose tissue, skeletal muscle, and liver and has insulin-mimetic actions. Regular exercise enhances insulin sensitivity. In the present study, we therefore examined visfatin mRNA expression...... in abdominal subcutaneous adipose tissue and skeletal muscle biopsies obtained from healthy young men at time points 0, 3, 4.5, 6, 9, and 24 h in relation to either 3 h of ergometer cycle exercise at 60% of Vo(2 max) or rest. Adipose tissue visfatin mRNA expression increased threefold at the time points 3, 4.......5, and 6 h in response to exercise (n = 8) compared with preexercise samples and compared with the resting control group (n = 7, P = 0.001). Visfatin mRNA expression in skeletal muscle was not influenced by exercise. The exercise-induced increase in adipose tissue visfatin was, however, not accompanied...

  13. Detection of MDR1 mRNA expression with optimized gold nanoparticle beacon

    Science.gov (United States)

    Zhou, Qiumei; Qian, Zhiyu; Gu, Yueqing

    2016-03-01

    MDR1 (multidrug resistance gene) mRNA expression is a promising biomarker for the prediction of doxorubicin resistance in clinic. However, the traditional technical process in clinic is complicated and cannot perform the real-time detection mRNA in living single cells. In this study, the expression of MDR1 mRNA was analyzed based on optimized gold nanoparticle beacon in tumor cells. Firstly, gold nanoparticle (AuNP) was modified by thiol-PEG, and the MDR1 beacon sequence was screened and optimized using a BLAST bioinformatics strategy. Then, optimized MDR1 molecular beacons were characterized by transmission electron microscope, UV-vis and fluorescence spectroscopies. The cytotoxicity of MDR1 molecular beacon on L-02, K562 and K562/Adr cells were investigated by MTT assay, suggesting that MDR1 molecular beacon was low inherent cytotoxicity. Dark field microscope was used to investigate the cellular uptake of hDAuNP beacon assisted with ultrasound. Finally, laser scanning confocal microscope images showed that there was a significant difference in MDR1 mRNA expression in K562 and K562/Adr cells, which was consistent with the results of q-PCR measurement. In summary, optimized MDR1 molecular beacon designed in this study is a reliable strategy for detection MDR1 mRNA expression in living tumor cells, and will be a promising strategy for in guiding patient treatment and management in individualized medication.

  14. Acute digoxin loading reduces ABCA8A mRNA expression in the mouse liver.

    Science.gov (United States)

    Wakaumi, Michi; Ishibashi, Kenichi; Ando, Hitoshi; Kasanuki, Hiroshi; Tsuruoka, Shuichi

    2005-12-01

    Human ABCA8, a new member of the ATP binding cassette (ABC) transporter family, transports certain lipophilic drugs, such as digoxin. To investigate the roles of this transporter, we cloned a mouse homologue of ABCA8, from a mouse heart cDNA library, named ABCA8a. The deduced mouse ABCA8a protein is 66% identical with that of human ABCA8 and possesses features common to the ABC superfamily. It was found that ABCA8a was mainly expressed in the liver and heart, similar to human ABCA8. We further evaluated the effect of acute digoxin (a substrate for ABCA8) intoxication on the mRNA expression of ABCA8 using northern blotting with a 3' non-coding region as a probe to avoid cross-hybridization with other ABCA genes. Following acute digoxin infusion, the mRNA expression of ABCA8 was significantly reduced in the liver 12-24 h after injection (14.7% of vehicle treatment), but not in the heart and kidney. Real-time quantitative polymerase chain reaction analysis confirmed the reduction in ABCA8a mRNA. Similar reductions in ABCA5, ABCA7, ABCA8b and ABCA9 mRNA were also observed. A comparable amount of digitoxin did not affect ABCA8a mRNA expression in the liver. The results suggest that ABCA8 may play a role in digoxin metabolism in the liver.

  15. The in vitro effect of desflurane preconditioning on endothelial adhesion molecules and mRNA expression.

    Science.gov (United States)

    Biao, Zhu; Zhanggang, Xue; Hao, Jiang; Changhong, Miao; Jing, Cang

    2005-04-01

    Lower expression of intercellular adhesion molecule-1 (ICAM-1), vascular adhesion molecule-1 (VCAM-1), and E-selectin may be responsible for attenuated ischemic-reperfusion neutrophil adhesion to vascular endothelium. Desflurane reduces ischemia-reperfusion injury. Therefore, we assessed whether desflurane affects the protein expression of ICAM-1 and E-selectin and mRNA expression of ICAM-1 and VCAM-1 of human umbilical venous endothelial cells (HUVEC) stimulated with tumor necrosis factor-alpha (TNF-alpha). HUVEC were preconditioned for 60 min with 1 minimum alveolar concentration desflurane before stimulating with TNF-alpha. Protein expression of adhesion molecules ICAM-1 and E-selectin of HUVEC were evaluated via immunocytochemical techniques combined with image cytometry. ICAM-1 and VCAM-1 mRNA expression of HUVEC were determined via reverse transcription-polymerase chain reaction. Desflurane not only reduced the protein expression of ICAM-1 and E-selectin but also ICAM-1 and VCAM-1 mRNA expression of the HUVEC. The adhesion rate of neutrophils with desflurane-treated HUVEC was slower. The decreased neutrophil adhesion on the desflurane-treated HUVEC correlated well with the decrease in adhesion molecule expression. These results show that desflurane affects the expression of adhesion molecules involved in the multistep process of neutrophil recruitment. Desflurane related ischemia-reperfusion injury reduction correlates well with expression inhibition of ICAM-1, VCAM-1, and E-selectin that mediates neutrophil rotation and firm adhesion on the vascular endothelium.

  16. EXPRESSION AND SIGNIFICANCE OF SURVIVIN mRNA IN LUNG CANCER TISSUE MICROARRAY DETECTED BY FISH

    Institute of Scientific and Technical Information of China (English)

    Xin-yun Wang; Xing-ye Wu; Zhi Yao; Yan Li; Ting Liu; Hai-yan Zheng; Cong-zhong Zhu; Cui-yun Sun; Ai-xiang Wang; Min Zhao

    2005-01-01

    Objective To investigate the expression of Survivin mRNA in lung cancer tissue microarray (TMA) by fluorescence in situ hybridization (FISH) method, and determine the role and significance of it in lung cancer genesis and progress. Methods The expression of Survivin mRNA was detected by FISH method and TMA technology. Fifty-four cases of lung cancer and 10 cases of normal lung tissue were examined. Results Survivin mRNA was expressed in 66.7% (36/54) of lung cancer; the positive ratio of lung cancer was significantly higher than that of normal lung tissue (0/10; x2= 15.238, P < 0.05). The positive ratio of Survivin mRNA was significantly higher in poor differentiated cancer (20/24, 83.3%) than moderate and well differentiated cancer (16/30, 53.3%; x2= 5.40, P <0.05). The positive ratio of Survivin mRNA was significantly higher in group with lymph node metastasis (27/32, 84.4%) than without lymph node metastasis (9/22, 40.9%; x2= 11.084, P < 0.05). The positive ratio of Survivin mRNA was significantly higher in stage Ⅲ-Ⅳ(12/13, 92.3%) than stage Ⅰ - Ⅱ (24/41, 58.5%; x2= 5.066, P < 0.05). Conclusion Survivin mRNA highly expresses in lung cancer, which is related to the progress and malignant behavior. Survivin may play a promoting role in lung cancer genesis and progress and provide a basis for estimating prognosis and treatment.

  17. Effects of Bevacizumab on CTGF and PEDF in proliferative membrane in patients with PDR

    Directory of Open Access Journals (Sweden)

    Lei Sun

    2017-06-01

    Full Text Available AIM: To investigate the effect of intravitreal injection of anti-vascular endothelial growth factor(VEGFdrug Bevacizumab on connective tissue growth factor(CTGFand pigment epithelium derived factor(PEDFin proliferative membranes in patients with proliferative diabetic retinopathy(PDR.METHODS: From January 2015 to December 2016, 117 patients(126 eyeswith PDR were analyzed using the case-control study. The selected cases were randomly divided into two groups, respectively, Group A of 60 cases(63 eyesand Group B of 57 cases(63 eyes. The Group A was treated with vitrectomy alone, while the Group B received intravitreal injection of 0.05mL/1.25mg bevacizumab before vitrectomy. The peeled epiretinal membranes in patients were stained and then the histopathological changes were observed for primitive cells and angiogenesis, the expression of CTGF and PEDF were tested. RESULTS: The expression of CTGF and PEDF in the proliferative membrane of the two groups were expressed in the cytoplasm. The Group A showed positive expression in 38 cases, the positive rate was 60.3%. Compared with Group A, the positive expression rate of CTGF in Group B(92.1%was significantly higher(PP>0.05. CONCLUSION: In patients with PDR after intravitreal injection of bevacizumab, the retinal neovascularization is significantly reduced, which is conducive to vitrectomy. The positive expression rate of CTGF significantly increased in patients with PDR after intravitreal injection of bevacizumab, but there was no significant change in the expression of PEDF in the anterior membrane.

  18. Gammaherpesviral gene expression and virion composition are broadly controlled by accelerated mRNA degradation.

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    Emma Abernathy

    2014-01-01

    Full Text Available Lytic gammaherpesvirus infection restricts host gene expression by promoting widespread degradation of cytoplasmic mRNA through the activity of the viral endonuclease SOX. Though generally assumed to be selective for cellular transcripts, the extent to which SOX impacts viral mRNA stability has remained unknown. We addressed this issue using the model murine gammaherpesvirus MHV68 and, unexpectedly, found that all stages of viral gene expression are controlled through mRNA degradation. Using both comprehensive RNA expression profiling and half-life studies we reveal that the levels of the majority of viral mRNAs but not noncoding RNAs are tempered by MHV68 SOX (muSOX activity. The targeting of viral mRNA by muSOX is functionally significant, as it impacts intracellular viral protein abundance and progeny virion composition. In the absence of muSOX-imposed gene expression control the viral particles display increased cell surface binding and entry as well as enhanced immediate early gene expression. These phenotypes culminate in a viral replication defect in multiple cell types as well as in vivo, highlighting the importance of maintaining the appropriate balance of viral RNA during gammaherpesviral infection. This is the first example of a virus that fails to broadly discriminate between cellular and viral transcripts during host shutoff and instead uses the targeting of viral messages to fine-tune overall gene expression.

  19. MicroRNA-145 Inhibits Cell Migration and Invasion and Regulates Epithelial-Mesenchymal Transition (EMT) by Targeting Connective Tissue Growth Factor (CTGF) in Esophageal Squamous Cell Carcinoma.

    Science.gov (United States)

    Han, Qiang; Zhang, Hua-Yong; Zhong, Bei-Long; Wang, Xiao-Jing; Zhang, Bing; Chen, Hua

    2016-10-23

    BACKGROUND This study investigated the mechanism of miR-145 in targeting connective tissue growth factor (CTGF), which affects the proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) of ESCC cells. MATERIAL AND METHODS A total of 50 ESCC tissues and their corresponding normal adjacent esophageal tissue samples were collected. Then, miR-145 expression in both ESCC clinical specimens and cell lines was detected using quantitative real-time PCR. CTGF protein was detected using immunohistochemistry. Dual luciferase reporter gene assay was employed to assess the effect of miR-145 on the 3'UTR luciferase activity of CTGF. Eca109 cells were transfected with miR-145 mimics and CTGF siRNA, respectively, and changes in cellular proliferation, migration, and invasion were detected via MTT assay, wound-healing assay, and Transwell assay, respectively. Western blotting assay was used to detect the expression of marker genes related to EMT. RESULTS MiR-145 was significantly down-regulated in ESCC tissues and cell lines compared with normal tissues and cell lines (Ptissues was than in normal adjacent esophageal tissues (Ptissues and cell lines, while the protein expression of CTGF exhibited the opposite trend. MiR-145 inhibited the proliferation, migration, invasiveness, and the EMT process of ESCC cells through targeted regulation of CTGF expression.

  20. Prediction of microRNAs affecting mRNA expression during retinal development

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    Cogliati Tiziana

    2010-01-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are small RNA molecules (~22 nucleotides which have been shown to play an important role both in development and in maintenance of adult tissue. Conditional inactivation of miRNAs in the eye causes loss of visual function and progressive retinal degeneration. In addition to inhibiting translation, miRNAs can mediate degradation of targeted mRNAs. We have previously shown that candidate miRNAs affecting transcript levels in a tissue can be deduced from mRNA microarray expression profiles. The purpose of this study was to predict miRNAs which affect mRNA levels in developing and adult retinal tissue and to confirm their expression. Results Microarray expression data from ciliary epithelial retinal stem cells (CE-RSCs, developing and adult mouse retina were generated or downloaded from public repositories. Analysis of gene expression profiles detected the effects of multiple miRNAs in CE-RSCs and retina. The expression of 20 selected miRNAs was confirmed by RT-PCR and the cellular distribution of representative candidates analyzed by in situ hybridization. The expression levels of miRNAs correlated with the significance of their predicted effects upon mRNA expression. Highly expressed miRNAs included miR-124, miR-125a, miR-125b, miR-204 and miR-9. Over-expression of three miRNAs with significant predicted effects upon global mRNA levels resulted in a decrease in mRNA expression of five out of six individual predicted target genes assayed. Conclusions This study has detected the effect of miRNAs upon mRNA expression in immature and adult retinal tissue and cells. The validity of these observations is supported by the experimental confirmation of candidate miRNA expression and the regulation of predicted target genes following miRNA over-expression. Identified miRNAs are likely to be important in retinal development and function. Misregulation of these miRNAs might contribute to retinal degeneration and disease

  1. Effects of Ginseng and Echinacea on Cytokine mRNA Expression in Rats

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    Deniz Uluışık

    2012-01-01

    Full Text Available The aim of the study was to determine the effect of ginseng and echinacea on the mRNA expression of IL-10, TNF-α, and TGF-β1 in healthy rats. Six-week-old male Fischer 344 rats (n=48 were used. The animals were divided into three equal groups, as follows: control (C; ginseng (G; echinacea (E. While the C group was fed a standard rat diet (Purina ad libitum for a period of 40 days, the G and E groups animals received the same diet containing 0.5 g/kg of Panax ginseng root powder and 0.75 g/kg of Echinacea purpurea root powder, respectively. Blood samples were obtained from 8 rats in each group after 20 and 40 days of treatment, and the mRNA expression of IL-10, TNF-α, and TGF-β1 was determined. After 20 days of treatment, the expression of IL-10 mRNA in the G group was different from the C group (P<0.05; however, after 40 days of treatment, there was no difference between the groups. There was no difference after 20 and 40 days of treatment between the groups with respect to the expression of TGF-β1 mRNA. After 20 days of treatment, the expression of TNF-α mRNA in the E group was higher (P<0.05 than the C group. After 40 days of treatment, the expression of TNF-α mRNA was similar in all of the groups. Based on the current study, the increase in expression of IL-10 mRNA in the G group and the increase in expression of TNF-α mRNA in the E group support the use of these plants for purposes of modulating the immune system. However, a more detailed study regarding the effects of ginseng and echinacea on these cytokines and other cytokines is needed.

  2. Differential Gene Expression in Retina of Myopic Chicken Eyes Using mRNA Differential Display

    Institute of Scientific and Technical Information of China (English)

    ShenHX; ZhangQJ

    1999-01-01

    Purpose:To study differentially expressed genes in retina of experimental myopic chicken.Methods:Experimental myopia in chicken was induced by form-deprivatin.The mRNA in chicen retina was analyzed by using differential display.Results:Experimental myopia was successfully induced in chicken through form-deprivation.Differentially expressed gene fragments were detected in retina of chicken with myopic evelopment and recovery as compared with normal controld.Conclusion:The differential display of mRNA may be a useful way in cloning myopic-related genes.

  3. Angiotensin II receptor mRNA expression and vasoconstriction in human coronary arteries

    DEFF Research Database (Denmark)

    Wackenfors, Angelica; Pantev, Emil; Emilson, Malin;

    2004-01-01

    Angiotensin II is a potent vasoconstrictor that is implicated in the pathogenesis of hypertension, heart failure and atherosclerosis. In the present study, angiotensin II receptor mRNA expression levels were quantified by real-time polymerase chain reaction and the vasocontractile responses...... to angiotensin II were characterised by in vitro pharmacology in endothelium-denuded human coronary arteries. Angiotensin II type 1 (AT(1)) and type 2 (AT(2)) receptor mRNA expression levels were significantly down-regulated in arteries from patients with heart failure as compared to controls. The angiotensin II...

  4. Effect of Exercise on the Expression of Adiponectin mRNA and GLUT4 mRNA in Type 2 Diabetic Rats

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To investigate the impact of exercise on the expression of adiponectin and GLUT4 mR NA in type 2 diabetic rats, type 2 diabetic rat model was made. The diabetic rats were treated with swimming training for 8 weeks. The expression of adiponectin mRNA in perirenal fat and GLUT4mRNA in skeletal muscles were assessed by reverse transcription polymerase chain reaction (RT PCR) and the levels of blood glucose, serum insulin, and blood lipid were measured. Our results showed that the expression of adiponectin mRNA and GLUT4 mRNA in diabetic model group was decreased by 45 % (P<0.01), 43 % (P<0.01) respectively. The gene expression of adiponectin and GLUT4 was increased significantly in swimming group (P<0.05 and P<0.01, respectively).Compared with the model group, fasting insulin, TG, TC and FFA were decreased significantly in the training group (P<0.05 or P<0.01) as compared with model group. It is concluded that exercise can promote the expression of adiponectin mRNA and GLUT4 mRNA in type 2 diabetic rats,which may be one of the mechanisms responsible for the amelioration of insulin resistance in the rats.

  5. EFFECT OF ANGIOTENSIN Ⅱ AND ITS RECEPIOR ANTAGONIST ON EXPRESSION OF CONNECTIVE TISSUEGROWTH FACTOR IN MESANGIAL CELLS%血管紧张素Ⅱ及其受体拮抗剂在系膜细胞培养中对结缔组织生长因子表达的影响

    Institute of Scientific and Technical Information of China (English)

    黄颂敏; 刘芳; 刘小菁; 刘瑞

    2000-01-01

    OBJECTIVE Connective tissue growth factor (CTGF) has been implicated in the process of renal fibrosis by stimulatingrenal cell proliferation and accumulation of extracellular matrix. The effect might be influenced by angiotensin Ⅱ(Ang Ⅱ) or thusby angiotensin Ⅱ receptor antagonist. In this study,we observe the effect ofAng Ⅱ and Losartan (an Ang Ⅱ receptor antagonist)on the expression of CTGF and CTGF mRNA in cultured mesangial cells(MCs). METHODOLOGY Mesangial cells of SDrots were isolated and cultured. Ang Ⅱ of different concentration( 10-9, 10-7, 10-5 mol/L) were added to the medium for 72hours to observe the influence of CTGF mRNA expression.Losartan of 10-5mol/L and Ang Ⅱ of 10-7 mol/L were added to themedium to observe the effects of Losartan on Ang Ⅱ stimulated CTGF mRNA expression. RESULTS Ang Ⅱ was found to theexpression of MCs CTGF mRNA in a dose-dependent manner. Expression of CTGF mRNA induced by Ang Ⅱ was partially sup-pressed by Losartan at the conccntration of 10-7 mol/L. CONCLUSION Ang Ⅱ can enhance MCs' expression of CTGF mR-NA and thus is involved in the process of renal fibrosis. Losantan can partially suppress Ang Ⅱ's effect of stimulating CTGF mRNAexpression in cultured MCs, and thus implication in the ueatment of renal fibrosis.%目的:研究血管紧张素Ⅱ(AngⅡ)及其受体拮抗剂Losartan对系膜细胞(MCs)结缔组织生长因子(CT-GF)表达的影响方法:分离培养SD大鼠肾小球系膜细胞。培养的系膜细胞中分别加入不同浓度的AngⅡ(10-、10-7、10-5mol/L),及10-7mol/L AngⅡ+10-5mol/L Losartan,作用72h后,采用反转录-聚合酶链反应(RT-PCR)技术测定MCs CTGF mRNA水平变化。结果:AngⅡ能促进MCs CTGF mRNA表达,且呈剂量依赖性;Losartan能部分降低AngⅡ对CTGF mRNA表达的诱导。结论:AngⅡ可能通过促进MCs CTGF的表达而促进了肾纤维化的进展;Losartan可以部分抑制AngⅡ对CTGF mRNA表达的诱导,从而可

  6. mRNA expression of adipocytokines and glucocorticoid-related genes are associated with downregulation of E-cadherin mRNA in colorectal adenocarcinomas.

    Science.gov (United States)

    Størkson, Ragnhild H; Aamodt, Rolf; Vetvik, Katja Kannisto; Pietilainen, Kirsi; Bukholm, Geir; Jonsdottir, Kristin; Vollan, Hilde S; Sonerud, Tonje; Lüders, Torben; Jacobsen, Morten B; Bukholm, Ida R K

    2012-08-01

    There is a consistently reported relationship between the incidence of colon cancer and obesity. It is thought that adipose tissue, particularly visceral fat, which secretes systemic factors that alter immunological, metabolic and endocrine milieu and promotes insulin resistance by producing adipocytokines, is important in cancer progression. Systemic high concentrations of adipocytokines, such as tumour necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), and glucocorticoid metabolism-related genes have been associated with gastrointestinal cancer. However, limited information exists about the expression of these cytokines within tumour tissue. mRNA expression of TNF-α, IL-6,IL-8, IL-10, IL-1RN, glucocorticoid receptor alpha (GR-α), 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), plasminogen activator inhibitor-1 (PAI-1), Slug, vimentin, Snail and E-cadherin was analysed in paired samples of tumour tissue and normal mucosa in 60 surgical patients for Dukes B and C colorectal adenocarcinomas using quantitative reverse transcription PCR and microarray technology. The mRNA expression level of analysed genes was compared between tumour tissue and normal mucosa from the same patients, and a correlation to mRNA expression of E-cadherin in the same tissue samples was also performed. A highly significant difference in mRNA expression level of several of the analysed genes was observed between tumour tissue and the normal intestinal mucosa. Inverse correlation between mRNA expression of 11βHSD1, IL-6, GR-α and PAI-1 on one hand and mRNA expression of E-cadherin on the other hand was observed. Results show that the adipocytokines and glucocorticoid metabolism-related genes are overexpressed in colorectal adenocarcinomas, and expression of these genes is associated with the downregulation of E-cadherin mRNA, connecting these genes to carcinogenesis and progression of colorectal cancer.

  7. Adaptive and maladaptive expression of the mRNA regulatory protein HuR

    Institute of Scientific and Technical Information of China (English)

    Suman; Govindaraju; Beth; S; Lee

    2013-01-01

    The RNA-binding proteins involved in regulation of mRNA post-transcriptional processing and translation control the fates of thousands of mRNA transcripts and basic cellular processes. The best studied of these, HuR, is well characterized as a mediator of mRNA stability and translation, and more recently, as a factor in nuclear functions such as pre-mRNA splicing. Due to HuR’s role in regulating thousands of mRNA transcripts, including those for other RNA-binding proteins, HuR can act as a master regulator of cell survival and proliferation. HuR itself is subject to multiple post-translationa modifications including regulation of its nucleocytoplasmic distribution. However, the mechanisms that govern HuR levels in the cell have only recently begun to be defined. These mechanisms are critical to cell health, as it has become clear in recent years that aberrant expression of HuR can lead alternately to decreased cell viability or to promotion of pathological proliferation and invasiveness. HuR is expressed as alternate mRNAs that vary in their untranslated regions, leading to differences in transcript stability and translatability. Multiple transcription factors and modulators of mRNA stability that regulate HuR mRNA expression have been identified. In addition, translation of HuR is regulated by numerous microRNAs, several of which have been demonstrated to have anti-tumor properties due to their suppression of HuR expression. This review summarizes the current state of knowledge of the factors that regulate HuR expression, along with the circumstances under which these factors contribute to cancer and inflammation.

  8. Heparanase mRNA expression and point mutation in hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Xiao-Peng Chen; Yin-Bib Liu; Jing Rui; Shu-You Peng; Cheng-Hong Peng; Zi-Yan Zhou; Liang-Hui Shi; Hong-Wei Shen; Bin Xu

    2004-01-01

    AIM: To explore the expression of heparanase mRNA and point mutation in hepatocellular carcinoma (HCC).METHODS: Reverse transcription polymerase chain reaction was used to measure the expression of heparanase mRNA in the primary tumor tissues and surrounding liver tissues of 33 HCC patients. T-A cloning and sequencing were used to detect whether there was any mutation in the amplified PCR products.RESULTS: The expression of heparanase mRNA was positive in 16 primary tumor tissues of HCC, and the positive rate was 48.5%, which was significantly higher than that in the surrounding liver parenchyma (P<0.01). The positive rate for heparanase gene in high-tendency to metastatic recurrence group (71.4%, 10/14) was obviously higher than that in low-tendency to metastatic recurrence group(31.6%, 6/19) (P= 0.023). The positive rate for heparanase gene in patients with metastatic recurrence during postoperative follow-up (78.6%, 11/14) was also significantly higher than that in those without metastatic recurrence (21.4%, 3/14)(P = 0.003). Sequence analysis of the HPA PCR products was made in 7 patients, and 2-point mutations were found in 4 patients, one of which was sense mutation, neither base insertion nor deletion was detected. The mutation rate was 57.1% (4/7).CONCLUSION: The expression rate of heparanase mRNA increases in HCC, and HPA mRNA may be one of the reliable markers for the metastatic activity gained by the liver tumor cells and could be used clinically in predicting metastatic recurrence of HCC. Point mutation may be one of the causes for enhanced heparanase mRNA expression.

  9. How is mRNA expression predictive for protein expression? A correlation study on human circulating monocytes

    Institute of Scientific and Technical Information of China (English)

    Yanfang Guo; Yuan Chen; Hui Jiang; Lijun Tan; Jingyun Xie; Xuezhen Zhu; Songping Liang; Hongwen Deng; Peng Xiao; Shufeng Lei; Feiyan Deng; Gary Guishan Xiao; Yaozhong Liu; Xiangding Chen; Liming Li; Shan Wu

    2008-01-01

    A key assumption in studying mRNA expression is that it is informative in the prediction of protein expression. However,only limited studies have explored the mRNA-protein expression correlation in yeast or human tissues and the results have been relatively inconsistent. We carried out correlation analyses on mRNA-protein expressions in freshly isolated human circulating monocytes from 30 unrelated women. The expressed proteins for 71 genes were quantified and identified by 2-D electrophoresis coupled with mass spectrometry. The corresponding mRNA expressions were quantified by Affymetrix gene chips. Significant correlation (r=0.235, P<0.0001) was observed for the whole dataset including all studied genes and all samples. The correlations varied in different biological categories of gene ontology. For example, the highest correlation was achieved for genes of the extracellular region in terms of cellular component (r=0.643, P<0.0001) and the lowest correlation was obtained for genes of regulation (r=0.099, P=0.213) in terms of biological process. In the genome, half of the samples showed significant positive correlation for the 71 genes and significant correlation was found between the average mRNA and the average protein expression levels in all samples (r=0.296, P<0.01). However, at the study group level, only five studied genes had significant positive correlation across all the samples. Our results showed an overall positive correlation between mRNA and protein expression levels.However, the moderate and varied correlations suggest that mRNA expression might be sometimes useful, but certainly far from perfect, in predicting protein expression levels.

  10. The influence of eccentric exercise on mRNA expression of skeletal muscle regulators.

    Science.gov (United States)

    Jensky, Nicole E; Sims, Jennifer K; Rice, Judd C; Dreyer, Hans C; Schroeder, E Todd

    2007-11-01

    To evaluate change in myostatin, follistatin, MyoD and SGT mRNA gene expression using eccentric exercise to study mechanisms of skeletal muscle hypertrophy. Young (28+/-5 years) and older (68+/-6 years) men participated in a bout of maximal single-leg eccentric knee extension on an isokinetic dynamometer at 60 degrees /s: six sets, 12-16 maximal eccentric repetitions. Muscle biopsies of the vastus lateralis were obtained from the dominant leg before exercise and 24 h after exercise. Paired t tests were used to compare change (pre versus post-exercise) for normalized gene expression in all variables. Independent t tests were performed to test group differences (young vs. older). A probability level of Peccentric exercise. Similarly, we did not observe significant change in myostatin (-3.83+/-8.8; P=0.23), follistatin (-2.66+/-5.2; P=0.17), MyoD (-0.13+/-3.1; P=0.90), or SGT (-1.6+/-3.5; P=0.19) mRNA expression in older subjects. Furthermore, the non-significant changes in mRNA expression were not different between young and older subjects, P>0.23 for all variables. Our data suggests that a single bout of maximal eccentric exercise does not alter myostatin, follistatin, MyoD or SGT mRNA gene expression in young or older subjects.

  11. TDP-43 suppresses tau expression via promoting its mRNA instability.

    Science.gov (United States)

    Gu, Jianlan; Wu, Feng; Xu, Wen; Shi, Jianhua; Hu, Wen; Jin, Nana; Qian, Wei; Wang, Xinglong; Iqbal, Khalid; Gong, Cheng-Xin; Liu, Fei

    2017-06-02

    In the brains of individuals with Alzheimer's disease (AD) and chronic traumatic encephalopathy, tau pathology is accompanied usually by intracellular aggregation of transactive response DNA-binding protein 43 (TDP-43). However, the role of TDP-43 in tau pathogenesis is not understood. Here, we investigated the role of TDP-43 in tau expression in vitro and in vivo. We found that TDP-43 suppressed tau expression by promoting its mRNA instability through the UG repeats of its 3΄-untranslated region (3΄-UTR). The C-terminal region of TDP-43 was required for this function. Neurodegenerative diseases-causing TDP-43 mutations affected tau mRNA instability differentially, in that some promoted and others did not significantly affect tau mRNA instability. The expression levels of tau and TDP-43 were inverse in the frontal cortex and the cerebellum. Accompanied with cytoplasmic accumulation of TDP-43, tau expression was elevated in TDP-43M337V transgenic mouse brains. The level of TDP-43, which is decreased in AD brains, was found to correlate negatively with the tau level in human brain. Our findings indicate that TDP-43 suppresses tau expression by promoting the instability of its mRNA. Down-regulation of TDP-43 may be involved in the tau pathology in AD and related neurodegenerative disorders. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  12. mRNA Expression of Ovine Angiopoietin-like Protein 4 Gene in Adipose Tissues.

    Science.gov (United States)

    Zhang, Jing; Jing, Jiong-Jie; Jia, Xia-Li; Qiao, Li-Ying; Liu, Jian-Hua; Liang, Chen; Liu, Wen-Zhong

    2016-05-01

    Angiopoietin-like protein 4 (ANGPTL4) is involved in a variety of functions, including lipoprotein metabolism and angiogenesis. To reveal the role of ANGPTL4 in fat metabolism of sheep, ovine ANGPTL4 mRNA expression was analyzed in seven adipose tissues from two breeds with distinct tail types. Forty-eight animals with the gender ratio of 1:1 for both Guangling Large Tailed (GLT) and Small Tailed Han (STH) sheep were slaughtered at 2, 4, 6, 8, 10, and 12 months of age, respectively. Adipose tissues were collected from greater and lesser omental, subcutaneous, retroperitoneal, perirenal, mesenteric, and tail fats. Ontogenetic mRNA expression of ANGPTL4 in these adipose tissues from GTL and STH was studied by quantitative real time polymerase chain reaction. The results showed that ANGPTL4 mRNA expressed in all adipose tissues studied with the highest in subcutaneous and the lowest in mesenteric fat depots. Months of age, tissue and breed are the main factors that significantly influence the mRNA expression. These results provide new insights into ovine ANGPTL4 gene expression and clues for its function mechanism.

  13. Heme Oxygenase-1 mRNA Expression in Egyptian Patients With Chronic Liver Disease

    Directory of Open Access Journals (Sweden)

    Abeer El-Sayed Abd El-Wahab

    2012-04-01

    Full Text Available Background: Chronic liver disease (CLD is a global medical problem. This disease is associated with increased hepatic oxidative stress. One of the antioxidant enzymes that protect cells against this stress is heme oxygenase-1 (HO-1.Objectives: This study aimed to investigate the mRNA expression of HO-1 in Egyptian patients with CLD and its relation to oxidative stress biomarkers.Patients and Methods: Levels of serum ferritin, carboxyhemoglobin, malondialdehyde (MDA, and erythrocyte-reduced glutathione (GSH were measured, and HO-1 mRNA expression was detected in 45 CLD patients (15 with nonalcoholic steatohepatitis [NASH], 15 with chronic hepatitis C, and 15 with liver cirrhosis and 15 healthy controls.Results: HO-1 mRNA expression was increased in patients with NASH, chronic hepatitis C, and liver cirrhosis compared to controls. The expression in cirrhotic patients was significantly higher than that in patients with NASH and chronic hepatitis C. Compared to controls, patients with NASH, chronic hepatitis C, and liver cirrhosis had higher levels of ferritin, carboxyhemoglobin, and MDA and lower levels of GSH. HO-1 mRNA expression was positively correlated with levels of carboxyhemoglobin, serum ferritin, and serum MDA and negatively correlated with levels of erythrocyte GSH in CLD patients.Conclusions: HO-1 mRNA expression was significantly increased in CLD patients, and the increase reflected the severity of the disease. The significant relationship between the increased HO-1 expression and oxidative stress biomarkers in patients with CLD suggests that HO-1 may play an important role in protecting the liver from oxidative stress-dependent damage. Therefore, induction of HO-1 could be a novel therapeutic option for CLD.

  14. Expression of cell cycle regulating factor mRNA in small cell lung cancer xenografts

    DEFF Research Database (Denmark)

    Krarup, M; Poulsen, H S; Spang-Thomsen, M

    1998-01-01

    We have investigated the expression of cyclins, cyclin dependent kinases (CDK), and CDK inhibitors (CKI) at the mRNA level in a panel of small-cell lung cancer (SCLC) cell lines in vitro and in vivo as xenografts in nude mice. The results showed that the cell lines expressed varying amounts of most...... cyclin and CDK's but only a few of the cell lines expressed cyclin D1 and/or D2 and some lacked expression of CDK6. Most cell lines expressed mRNA for the CKI's but two cell lines lacked expression of P15INK4B and p16INK4A. The mRNA expression differed for a few of the cell lines regarding cyclin D2...... and CDK6 when in vitro and in vivo data were compared. Two of the cell lines that express the retinoblastoma (Rb) protein had no sign of a deregulated Rb pathway but further studies at the protein level are necessary to demonstrate whether these two cell lines should have a normal Rb pathway or whether...

  15. The expression of TRPA1 mRNA in the rat brain

    Institute of Scientific and Technical Information of China (English)

    Peng Du; Shua Li; Jinyu Zheng; Zhi-yuan Yu; Minjie Xie; Wei Wang

    2006-01-01

    Objective: To investigate the distribution of TRPA1 (one kind of the TRP-like ion channel family) channel in the hippocampus and cerebral cortex of rat. Methods: RT-PCR was used to amplify the fragment of TRPA1 in the DRG (dorsal root ganglion), hippocampus and cerebral cortex of adult SD rat. In situ hybridization staining was used to show the distribution of TRPA1 mRNA in the hippocampus and cerebral cortex of adult rat brain. Results: Both RT-PCR and in situ hybridization staining showed that TRPA1 mRNA was expressed in hippocampus and cerebral cortex of the adult rat brain. Conclusion: Ourresults suggest that there is expression of TRPA1 mRNA both in the hippocampus and cerebral cortex of the adult rat brain.

  16. CYP1A mRNA expression in redeye mullets (Liza haematocheila) from Bohai Bay, China.

    Science.gov (United States)

    An, Lihui; Hu, Jianying; Yang, Min; Zheng, Binghui; Wei, An; Shang, Jingjing; Zhao, Xingru

    2011-04-01

    Induction of cytochrome P4501A (CYP1A) has been used as a biomarker in fish for monitoring aromatic and organic contaminants. In this study, a partial of CYP1A gene in redeye mullet (Liza haematocheila) was isolated and sequenced, and then a real-time quantitative reverse-transcription polymerase chain reaction assay was developed for quantification of CYP1A mRNA normalized to β-actin. The developed method was applied to detect CYP1A mRNA expression in redeye mullets collected from Nandaihe (reference site) and Dashentang (impacted site) in Bohai Bay, China. CYP1A mRNA expression values were significantly elevated in redeye mullets from Dashentang compared to a reference site--Nandaihe, which was correlated with the contents of different environmentally relevant pollutants in tissues, particularly with PCBs and PBDEs.

  17. Chamber-dependent expression of brain natriuretic peptide and its mRNA in normal and diabetic pig heart

    DEFF Research Database (Denmark)

    Christoffersen, Christina; Goetze, Jens P; Bartels, Emil D;

    2002-01-01

    -induced diabetes doubled the expression of BNP mRNA in porcine atrial myocardium (P=0.03), but was without effect on BNP mRNA in the ventricular myocardium. The data suggest that BNP mRNA processing and proBNP storage differ between the atrial and ventricular myocardium. The results also imply that diabetes...

  18. Eosinophil cationic protein mRNA expression in children with bronchial asthma.

    Science.gov (United States)

    Yu, H Y; Li, X Y; Cai, Z F; Li, L; Shi, X Z; Song, H X; Liu, X J

    2015-11-13

    Studies have shown that eosinophils are closely related to pathogenesis of bronchial asthma. Eosinophils release eosinophil cationic protein (ECP), which plays an important role in infection and allergic reactions. Serum ECP mRNA expression in children with bronchial asthma has not been adequately investigated. We analyzed serum ECP mRNA expression in 63 children with bronchial asthma and 21 healthy children by using reverse-transcriptase polymerase chain reaction to understand the role of ECP in children with bronchial asthma. The children with bronchial asthma were segregated into acute-phase and stable-phase groups, based on the severity of the illness. Serum ECP mRNA expression in children with bronchial asthma (0.375 ± 0.04) was significantly higher than that in healthy controls (0.20 ± 0.02; P children in the acute-phase group showed higher ECP mRNA expression level (0.44 ± 0.06) than those in the stable-phase (0.31 ± 0.03) and healthy control groups (0.20 ± 0.02; P children with bronchial asthma.

  19. Endothelin converting-enzyme-1 mRNA expression in human cardiovascular disease

    NARCIS (Netherlands)

    Bohnemeier, H; Pinto, YM; Horkay, F; Toth, M; Juhasz-Nagy, A; Orzechowski, HD; Bohm, M; Paul, M

    1998-01-01

    Endothelin-1 converting-enzyme (ECE-1) cleaves the precursor, big-endothelin-1, to the active peptide endothelin-1. The aim of this study was to investigate whether ECE-1 mRNA expression is modified in human cardiovascular disease. Tissue samples from the left human atrium were analyzed for ECE-1 ex

  20. Real time imaging of mRNA expression dynamics in live cells using protein complementation methods

    Science.gov (United States)

    Meller, Amit

    2009-03-01

    Traditional methods for mRNA quantification in cells, such as northern blots, quantitative PCR or microarrays assays, require cell lysis and therefore do not preserve its dynamics. These methods cannot be used to probe the spatio-temporal localization of mRNA in cells, which provide useful information for a wide range biomolecular process, including RNA metabolizim, expression kinetics and RNA interference. To probe mRNA dynamics in live prokaryotic and eukaryotic cells, we develop a method, which exploit the strong affinity of the eukaryotic initiation factor 4A (eIF4A) to specific RNA aptamers. Two parts of the eIF4A are fused to a split Green Fluorescence Protein (GFP), and are expressed in the cells at high abundance. However, only when the RNA apatmer is also present, the two protein parts complement and become fluorescent. Thus, the fluorescent background remains low, allowing us to directly image the expression of mRNA molecules in live e-coli cells from its early onset, over hours. We find that the expression kinetics can be classified in one out of at least three forms, which also display distinct spatial distributions. I will discuss the possible biological origin for these distributions and their time evolution.

  1. Expression of TGF-β1 and CTGF in rats with experimental diabetic macroangiopathy and intervention role of traditional Chinese medicine%实验性糖尿病鼠大血管病变TGF-β1和CTGF的表达及中药的干预作用

    Institute of Scientific and Technical Information of China (English)

    王军; 徐阳; 袁向科; 张学勇; 丁志明

    2012-01-01

    [Objective] To explore the pathogenesis mechanism of macroangiopathy in diabetic patients trough observing the expression of TGF-β1 and CTGF in rats with experimental diabetic macroangiopathy and decide the target in preventing and treating the macroan-giopathy in patients with diabetes through observing the preventing effect of TCM. [Methods] Among eighty male SD rats seventy were induced with diabetes (DM). They were divided into 5 groups: model group (n=14), high, medium and low dose of traditional Chinese medicine groups (n=14), melbine group (n—14). All rats in these six groups were giving with ordinary feed for 12 weeks. After that, arteria cruralis was obtained and the site and change of expression of CTGF and TGF-β1 in which were observed. [Results] TGF-β1 was mainly expressed in the cytoplasm. It is strong positive in model group. In Chinese medicine intervention groups the expression of TGF-β1 was declined obviously (P<0.01). Melbine has influence on the expression of TGF-β1, but the comparison with high dose of traditional Chinese medicine group has significant difference (P<0.05). A large of brown-lyellow material with strong positive reaction (CTGF) could be seen in cytochylema in submucosa of blood vessels of model group. But the CTGF expression was all significantly lowered in cell of blood vessels in drug intervention groups (P<0.01). The difference of expression between high dose group and Western medicine control group was statistically different. [Conclusion] 1 )The onset of diabetic angiopathy may be concerned with fibrosis. 2) Modified Taohe Chengqi Decoction can reduce the expression of Fibrosis factor TGF-β1 and CTGF in rats with experimental diabetic macroangiopathy. It can improve macroangiopathy in DM and has a dose-ldependent relationship.%[目的]通过观测实验性糖尿病鼠大血管病变中转化生长因子(TGF-β1)和结缔组织生长因子(CTGF)相关表达,揭示糖尿病大血管病变的发病机制.并通过中药

  2. Relationship between expression of somatostatin receptors subtype 2 mRNA and estrogen and progesterone receptors in breast cancer

    Institute of Scientific and Technical Information of China (English)

    曾希志; 姚榛祥

    2003-01-01

    Objectives To observe the expression of somatostatin receptor subtype 2 (SSTR2) mRNA, and investigate the relationship between the expression of SSTR2 mRNA and the expressions of estrogen and progesterone receptors (ERs and PRs) in benign and malignant breast tissues.Methods Samples from a total of 23 breast carcinomas, 16 mammary hyperplasias, and 9 mammary fibroadenomas were analyzed. SSTR2 mRNA expression was examined by in situ hybridization using multiphase oligoprobes. ER and PR expressions were detected by immunohistochemical staining. A computerized image analysis system was utilized to estimate the relative content of SSTR2 mRNA.Results The rate of expression (87.0%) and relative content (0.47) of SSTR2 mRNA in breast cancer were higher than those in benign breast tissue (64%,0.26) (P<0.05). SSTR2 mRNA expression was closely correlated with ER and PR expressions in breast cancer (P<0.05). SSTR2 mRNA was also positively correlated with ER expression in benign breast tissues.Conclusions SSTR2 mRNA expression is higher or in benign breast tissues than in malignant ones. There is a significant positive correlation between SSTR2 mRNA and ER and PR expressions. Combined antiestrogen and somatostatin analogue in treatment of ER-positive breast cancers should be further investigated.

  3. Tissue-specific mRNA expression profiles of human solute carrier 35 transporters.

    Science.gov (United States)

    Nishimura, Masuhiro; Suzuki, Satoshi; Satoh, Tetsuo; Naito, Shinsaku

    2009-01-01

    Pairs of forward and reverse primers and TaqMan probes specific to each of 23 human solute carrier 35 (SLC35) transporters were prepared. The mRNA expression level of each target transporter was analyzed in total RNA from single and pooled specimens of adult human tissues (adipose tissue, adrenal gland, bladder, bone marrow, brain, cerebellum, colon, heart, kidney, liver, lung, mammary gland, ovary, pancreas, peripheral leukocytes, placenta, prostate, retina, salivary gland, skeletal muscle, small intestine, smooth muscle, spinal cord, spleen, stomach, testis, thymus, thyroid gland, tonsil, trachea, and uterus), from pooled specimens of fetal human tissues (brain, heart, kidney, liver, spleen, and thymus), and from three human cell lines (HeLa cell line ATCC#: CCL-2, human cell line Hep G2, and human breast carcinoma cell line MDA-435) by real-time reverse transcription PCR using an Applied Biosystems 7500 Fast Real-Time PCR System. The mRNA expression of SLC35As, SLC35Bs, SLC35Cs, SLC35D1, SLC35D2, SLC35Es, and SLC35F5 was found to be ubiquitous in both adult and fetal tissues. SLC35D3 mRNA was expressed at the highest levels in the adult retina. SLC35F1 mRNA was expressed at high levels in the adult and fetal brain. SLC35F2 mRNA was expressed at the highest levels in the adult salivary gland. Both SLC35F3 and SLC35F4 mRNAs were expressed at the highest levels in the adult cerebellum. Further, individual differences in the mRNA expression levels of human SLC35 transporters in the liver were also evaluated. Our newly determined expression profiles were used to study the gene expression in 31 adult human tissues, 6 fetal human tissues, and 3 cell lines, and tissues with high transcriptional activity for human SLC35 transporters were identified. These results are expected to be valuable for research concerning the clinical diagnosis of disease.

  4. IGF-1 mRNA expression of adult rat thyroid cell cultured in vitro

    Institute of Scientific and Technical Information of China (English)

    HE Feng-ping(何凤屏); YIN Rui-xing(尹瑞兴); XUAN Su(冼苏); JEAN Joss

    2003-01-01

    Objective:To investigate the law of age-related changes of insulin-like growth factor-1(IGF-1)expression of rat thyroid cells cultured in vitro.Methods:Rat thyroid of different age(10,45,65,100,150 weeks)was isolated and thyrocytes cultured.Total RNA was extracted in different rat age group when thyroid cells had been cultured for two weeks,mRNA IGF-1 expression was measured with reverse-transcription polymerase chain reaction(RT-PCR)in each group and compared.Results:Quantity of total RNA in thyroid cells decreased with ageing when the rat thyroid cells had been cultured for 2 weeks.There is significant difference among groups(P < 0.05).Expression of IGF-1 mRNA could be detected in thyroid cells of different age cultured in vitro.Quantity of IGF-1 mRNA expression by RTPCR analysis increased from 10 to 45 weeks old,and then decreased with ageing.Conclusion:Rat thyroid cells from different age cultured in vitro can express IGF-1 mRNA.Quantity of total RNA in thyroid cells cultured in vitro decreased with aging.IGF-1 mRNA expression was correlated to age(r =0.401,P <0.05).

  5. Analysis of correlations between protein complex and protein-protein interaction and mRNA expression

    Institute of Scientific and Technical Information of China (English)

    CAI Lun; XUE Hong; LU Hongchao; ZHAO Yi; ZHU Xiaopeng; BU Dongbo; LING Lunjiang; CHEN Runsheng

    2003-01-01

    Protein-protein interaction is a physical interaction of two proteins in living cells. In budding yeast Saccharomyces cerevisiae, large-scale protein-protein interaction data have been obtained through high-throughput yeast two-hybrid systems (Y2H) and protein complex purification techniques based on mass-spectrometry. Here, we collect 11855 interactions between total 2617 proteins. Through seriate genome-wide mRNA expression data, similarity between two genes could be measured. Protein complex data can also be obtained publicly and can be translated to pair relationship that any two proteins can only exist in the same complex or not. Analysis of protein complex data, protein-protein interaction data and mRNA expression data can elucidate correlations between them. The results show that proteins that have interactions or similar expression patterns have a higher possibility to be in the same protein complex than randomized selected proteins, and proteins which have interactions and similar expression patterns are even more possible to exist in the same protein complex. The work indicates that comprehensive integration and analysis of public large-scale bioinformatical data, such as protein complex data, protein-protein interaction data and mRNA expression data, may help to uncover their relationships and common biological information underlying these data. The strategies described here may help to integrate and analyze other functional genomic and proteomic data, such as gene expression profiling, protein-localization mapping and large-scale phenotypic data, both in yeast and in other organisms.

  6. Investigation of HSP60 gene expression in mRNA level in heart at dilated cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Riabenko D. V.

    2009-02-01

    Full Text Available The expression of HSP60 in the mRNA level in human hearts at the end-stage of dilated cardiomyopathy (DCM as well as in the hearts of mice with disease model similar to human DCM was investigated. We observed a significant increase in the Hsp60 mRNA level at the beginning of the disease and decrease to a normal level at the end stage. As the Hsp60 level was increased during the disease up to the end stage we can presume some changes in the regulation of Hsp60 synthesis or its degradation at DCM progression

  7. Epigenetic Regulation of Dopamine Transporter mRNA Expression in Human Neuroblastoma Cells

    Science.gov (United States)

    Green, Ashley L.; Hossain, Muhammad M.; Tee, Siew C.; Zarbl, Helmut; Guo, Grace L.; Richardson, Jason R.

    2016-01-01

    The dopamine transporter (DAT) is a key regulator of dopaminergic neurotransmission. As such, proper regulation of DAT expression is important to maintain homeostasis, and disruption of DAT expression can lead to neurobehavioral dysfunction. Based on genomic features within the promoter of the DAT gene, there is potential for DAT expression to be regulated through epigenetic mechanisms, including DNA methylation and histone acetylation. However, the relative contribution of these mechanisms to DAT expression has not been empirically determined. Using pharmacologic and genetic approaches, we demonstrate that inhibition of DNA methyltransferase (DNMT) activity increased DAT mRNA approximately 1.5–2 fold. This effect was confirmed by siRNA knockdown of DNMT1. Likewise, the histone deacetylase (HDAC) inhibitors valproate and butyrate also increased DAT mRNA expression, but the response was much more robust with expression increasing over tenfold. Genetic knockdown of HDAC1 by siRNA also increased DAT expression, but not to the extent seen with pharmacological inhibition, suggesting additional isoforms of HDAC or other targets may contribute to the observed effect. Together, these data identify the relative contribution of DNMTs and HDACs in regulating expression. These finding may aid in understanding the mechanistic basis for changes in DAT expression in normal and pathophysiological states. PMID:25963949

  8. Expression of SART-1 mRNA in canine squamous cell carcinomas.

    Science.gov (United States)

    Takaishi, Yumi; Yoshida, Yukari; Nakagaki, Kazuhide; Fujita, Michio; Taniguchi, Akiko; Orima, Hiromitsu

    2008-12-01

    SART-1, a squamous cell carcinoma (SCC) antigen recognized by cytotoxic T lymphocytes, has been useful in human cancer therapy. The SART-1(259) peptide is a potential candidate for vaccine. The present study examined an orthologue of the mRNA coding this peptide in canine SCCs. Specimens were obtained from seven canine patients with SCC, and the mRNA was isolated from the samples. The SART-1 and beta-actin genes were amplified by reverse-transcription polymerase chain reaction, using the isolated mRNA as a template. Canine SART-1 was amplified in six of the seven specimens, while beta-actin was detected in all the samples. In dogs, carcinomas expressing SART-1 could be a target for cytotoxic T lymphocyte mediated immunotherapy.

  9. Expression of connexin 43 mRNA and protein in developing follicles of prepubertal porcine ovaries

    Science.gov (United States)

    Melton, C.M.; Zaunbrecher, G.M.; Yoshizaki, G.; Patio, R.; Whisnant, S.; Rendon, A.; Lee, V.H.

    2001-01-01

    A major form of cell-cell communication is mediated by gap junctions, aggregations of intercellular channels composed of connexins (Cxs), which are responsible for exchange of low molecular weight (hybridization revealed that Cx43 mRNA was detectable in granulosa cells of primary follicles but undetectable in dormant primordial follicles. The intensity of the signal increased with follicular growth and was greatest in the large antral follicles. Immunohistochemical evaluation indicated that Cx43 protein expression correlated with the presence of Cx43 mRNA. These results indicate that substantial amounts of Cx43 are first expressed in granulosa cells following activation of follicular development and that this expression increases throughout follicular growth and maturation. These findings suggest an association between the enhancement of intercellular gap-junctional communication and onset of follicular growth. ?? 2001 Elsevier Science Inc. All rights reserved.

  10. Adrenocorticotropin receptors: Functional expression from rat adrenal mRNA in Xenopus laevis oocytes

    Energy Technology Data Exchange (ETDEWEB)

    Mertz, L.M.; Catt, K.J. (National Inst. of Health, Bethesda, MD (United States))

    1991-10-01

    The adrenocorticotropin (ACTH) receptor, which binds corticotropin and stimulates adenylate cyclase and steroidogenesis in adrenocortical cells, was expressed in Xenopus laevis oocytes microinjected with rat adrenal poly(A){sup +} RNA. Expression of the ACTH receptor in individual stage 5 and 6 oocytes was monitored by radioimmunoassay of ligand-stimulated cAMP production. Injection of 5-40 ng of adrenal mRNA caused dose-dependent increases in ACTH-responsive cAMP production. Size fractionation of rat adrenal poly(A){sup +}RNA by sucrose density-gradient centrifugation revealed that mRNA encoding the ACTH receptor was present in the 1.1-to 2.0-kilobase fraction. These data indicate that ACTH receptors can be expressed from adrenal mRNA in Xenopus oocytes and are fully functional in terms of ligand specificity and signal generation. The extracellular cAMP response to ACTH is a sensitive and convenient index of receptor expression. This system should permit more complete characterization and expression cloning of the ACTH receptor.

  11. Oligodendrocyte transcription factor 1 mRNA and protein expression in organotypic rat brain slices

    Institute of Scientific and Technical Information of China (English)

    Hong Cui; Lijun Yang; Dezhuang Huang; Wandong Zhang; Weijuan Han; Yanqing Yao; Wenxing Jiang

    2010-01-01

    Numerous studies have confirmed that oligodendrocyte transcription factor 1 (Olig-1) is vital for myelin repair. However, the effects of hypoxia and ischemia on Olig-1 expression remain unknown.In this study, Olig-1 mRNA and protein expressions were analyzed by in situ hybridization and immunohistochemistry, to determine the expression profile of Olig-1 in rat brain slices exposed to hypoxia and ischemia. Brains were obtained from 2-day-old Sprague-Dawley rats, and sections were randomly assigned to control and hypoxia/ischemia groups. Hematoxylin-eosin staining revealed karyorrhexis and karyopyknosis in cells from the hypoxia/ischemia group. Under electron microscopy, mitochondria swelling and neuropil edema were observed in the hypoxia/ischemia group. Olig-1 mRNA and protein expressions were increased at 1 day after hypoxia and ischemia treatment. These results suggest that in situ hybridization and immunohistochemistry could be used simultaneously to detect mRNA and protein expression in brain slices.

  12. Maternal mRNA expression levels of H19 are inversely associated with risk of macrosomia.

    Science.gov (United States)

    Jiang, Hua; Yu, Yang; Xun, Pengcheng; Zhang, Jun; Luo, Guanghua; Wang, Qiuwei

    2014-06-29

    To investigate the associations between the mRNA levels of H19 in term placenta and risk of macrosomia. Term placentas were collected from 37 macrosomia and 37 matched neonates with normal birth weight (controls) born in Changzhou Women and Children Health Hospital, Jiangsu province, P. R. China from March 1 to June 30, 2008. The mRNA levels of H19 in those placentas were measured by real-time polymerase chain reaction (PCR). Simple and multiple logistic regression models were used to explore the risk factors in the development of macrosomia. All analyses were performed using Stata 10.0 (StataCorp, College Station, Texas, USA). The average H19 mRNA level of the macrosomia group was 1.450 ±0.456 while in the control group it was 2.080 ±1.296. Based on the result of Student's t test, there was a significant difference in H19 mRNA level between the macrosomia group and the control group (p = 0.008). After controlling for potential confounders, the multivariable adjusted odds ratio (OR) of macrosomia for those in the highest tertile of H19 mRNA level was 0.12 (95% CI: 0.02-0.59) when compared to those in the lowest tertile (p for linear trend = 0.009). The term placental H19 mRNA levels were inversely related to the occurrence of macrosomia. Our findings suggest that the low expression of H19 mRNA may contribute to the development of macrosomia.

  13. Gallium nitrate regulates rat osteoblast expression of osteocalcin protein and mRNA levels.

    Science.gov (United States)

    Guidon, P T; Salvatori, R; Bockman, R S

    1993-01-01

    Gallium nitrate, a group IIIa metal salt, has been found to be clinically effective for the treatment of accelerated bone resorption in cancer-related hypercalcemia and Paget's disease. Here we report the effects of gallium nitrate on osteocalcin mRNA and protein levels on the rat osteoblast-like cell line ROS 17/2.8. Gallium nitrate reduced both constitutive and vitamin D3-stimulated osteocalcin protein levels in culture medium by one-half and osteocalcin mRNA levels to one-third to one-tenth of control. Gallium nitrate also inhibited vitamin D3 stimulation of osteocalcin and osteopontin mRNA levels but did not affect constitutive osteopontin mRNA levels. Among several different metals examined, gallium was unique in its ability to reduce osteocalcin mRNA levels without decreasing levels of other mRNAs synthesized by ROS 17/2.8 cells. The effects of gallium nitrate on osteocalcin mRNA and protein synthesis mimic those seen when ROS 17/2.8 cells are exposed to transforming growth factor beta 1 (TGF beta 1); however, TGF-beta 1 was not detected in gallium nitrate-treated ROS 17/2.8 cell media. Use of the RNA polymerase II inhibitor 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole demonstrated that gallium nitrate did not alter the stability of osteocalcin mRNA. Transient transfection assays using the rat osteocalcin promoter linked to the bacterial reporter gene chloramphenicol acetyltransferase indicated that gallium nitrate blocked reporter gene expression stimulated by the osteocalcin promoter. This is the first reported effect of gallium nitrate on isolated osteoblast cells.

  14. Hemizygous deletion of CTGF/CCN2 does not suffice to prevent fibrosis of the severely injured kidney

    NARCIS (Netherlands)

    Falke, L.L.; Dendooven, A.; Leeuwis, J.W.; Nguyen, T.Q.; Geest, R.J. van; Giezen, D.M. van der; Broekhuizen, R.; Lyons, K.; Stoop, R.; Kemperman, H.; Schlingemann, R.; Joles, J.A.; Goldschmeding, R.

    2012-01-01

    Background: Connective Tissue Growth Factor (CTGF/CCN2) is an important mediator of kidney fibrosis. Previous observations indicated that attenuation of CCN2 expression sufficed to alleviate early kidney damage. However, little is known about the role of CCN2 in fibrosis of severely damaged and more

  15. TELOMERASE ACTIVITY AND hTERT mRNA EXPRESSION IN ACUTE LEUKEMIA

    Institute of Scientific and Technical Information of China (English)

    何冬梅; 张洹

    2004-01-01

    Objective: To investigate the clinical implications of telomerase activity and human telomerase reverse transcriptase (hTERT) expression as useful diagnostic marker in acute leukemia. Methods: Expression of hTERT was detected by reverse transcription- polymerase chain reaction (RT-PCR) in 24 cases with acute leukemia and in 12 normal persons. Quantitative levels of telomerase activity were examined by polymerase chain reaction enzyme-linked immunoassay (PCR-ELISA). Results: In the bone marrow and peripheral blood of 24 acute leukemia, telomerase activity was detected in 75% of the samples, with absorbances (A) of 0.538(0.062 and 0.463(0.054, respectively. Whereas in 12 normal peripheral blood, telomerase activity had only a positive rate of 8.3%, with A value of 0.16(0.012. telomerase activities in the bone marrow and peripheral blood of acute leukemia were significantly higher than in normal control (P<0.05). RT-PCR analysis revealed that hTERT mRNA was expressed in 79.17%(19/24) of acute leukemia, but in only 1 of 12 normal peripheral blood. In 24 acute leukemias, 17 cases had both positive telomerse activity and hTERT mRNA expression. The expression of hTERT mRNA is correlated with telomerase activity (P<0.01). Conclusion: Telomerase and hTERT mRNA could be useful in diagnosis of acute leukemia. hTERT gene expression was strongly associated with telomerase activity in acute leukemia.

  16. Rhythmic expression of Nocturnin mRNA in multiple tissues of the mouse

    Directory of Open Access Journals (Sweden)

    Green Carla B

    2001-05-01

    Full Text Available Abstract Background Nocturnin was originally identified by differential display as a circadian clock regulated gene with high expression at night in photoreceptors of the African clawed frog, Xenopus laevis. Although encoding a novel protein, the nocturnin cDNA had strong sequence similarity with a C-terminal domain of the yeast transcription factor CCR4, and with mouse and human ESTs. Since its original identification others have cloned mouse and human homologues of nocturnin/CCR4, and we have cloned a full-length cDNA from mouse retina, along with partial cDNAs from human, cow and chicken. The goal of this study was to determine the temporal pattern of nocturnin mRNA expression in multiple tissues of the mouse. Results cDNA sequence analysis revealed a high degree of conservation among vertebrate nocturnin/CCR4 homologues along with a possible homologue in Drosophila. Northern analysis of mRNA in C3H/He and C57/Bl6 mice revealed that the mNoc gene is expressed in a broad range of tissues, with greatest abundance in liver, kidney and testis. mNoc is also expressed in multiple brain regions including suprachiasmatic nucleus and pineal gland. Furthermore, mNoc exhibits circadian rhythmicity of mRNA abundance with peak levels at the time of light offset in the retina, spleen, heart, kidney and liver. Conclusion The widespread expression and rhythmicity of mNoc mRNA parallels the widespread expression of other circadian clock genes in mammalian tissues, and suggests that nocturnin plays an important role in clock function or as a circadian clock effector.

  17. Embryonic and Postnatal Expression of Aryl Hydrocarbon Receptor mRNA in Mouse Brain

    Science.gov (United States)

    Kimura, Eiki; Tohyama, Chiharu

    2017-01-01

    Aryl hydrocarbon receptor (AhR), a member of the basic helix-loop-helix-Per-Arnt-Sim transcription factor family, plays a critical role in the developing nervous system of invertebrates and vertebrates. Dioxin, a ubiquitous environmental pollutant, avidly binds to this receptor, and maternal exposure to dioxin has been shown to impair higher brain functions and dendritic morphogenesis, possibly via an AhR-dependent mechanism. However, there is little information on AhR expression in the developing mammalian brain. To address this issue, the present study analyzed AhR mRNA expression in the brains of embryonic, juvenile, and adult mice by reverse transcription (RT)-PCR and in situ hybridization. In early brain development (embryonic day 12.5), AhR transcript was detected in the innermost cortical layer. The mRNA was also expressed in the hippocampus, cerebral cortex, cerebellum, olfactory bulb, and rostral migratory stream on embryonic day 18.5, postnatal days 3, 7, and 14, and in 12-week-old (adult) mice. Hippocampal expression was abundant in the CA1 and CA3 pyramidal and dentate gyrus granule cell layers, where expression level of AhR mRNA in 12-week old is higher than that in 7-day old. These results reveal temporal and spatial patterns of AhR mRNA expression in the mouse brain, providing the information that may contribute to the elucidation of the physiologic and toxicologic significance of AhR in the developing brain. PMID:28223923

  18. Expression of Plasmacytoid Dendritic Cells, IRF-7, IFN-α mRNA in the Lesions of Psoriasis Vulgaris

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    To investigate the expression of plasmacytoid dendritic cells (pDCs), interferon regulatory factor-7 (IRF-7) and interferon alpha (IFN- α ) mRNA in skin lesions of patients with psoriasis vulgaris, the expressions of plasmacytoid dendritic cells, IRF-7, IFN-α mRNA in the lesional skin of psoriasis vulgaris were detected by immunohistochemical technique (SP) and RT-PCR. Normal skin of healthy volunteers, serving as control, was also tested. The immunohistochemical study showed that the expression of pDCs in the psoriatic lesions was significantly higher than that in the normal controls. RT-PCR showed that the mRNA expression of IRF-7 was much higher than that in normal controls, but no difference in the expression of IFN-α mRNA was found between two groups. Our findings indicate that up-regulated expression of pDCs, IRF-7mRNA might be involved in the pathogenesis of psoriasis.

  19. Acute stress increases neuropsin mRNA expression in the mouse hippocampus through the glucocorticoid pathway.

    Science.gov (United States)

    Harada, Akiko; Shiosaka, Sadao; Ishikawa, Yasuyuki; Komai, Shoji

    2008-05-01

    Stress affects synaptic plasticity and may alter various types of behaviour, including anxiety or memory formation. In the present study, we examined the effects of acute stress (1 h restraint with or without tail-shock) on mRNA levels of a plasticity-related serine protease neuropsin (NP) in the hippocampus using semiquantitative RT-PCR and in situ hybridization. We found that NP mRNA expression was dramatically increased shortly after exposure to the acute restraint tail-shock stress and remained at high level for at least 24 h. The level of NP mRNA would be correlated to the elevated plasma concentration of the glucocorticoid corticosterone (CORT) and to the stress intensity. Application of CORT either onto primary cultured hippocampal neurons (5 nM) or in vivo to adrenalectomized (ADX) mice (10 mg/kg B.W., s.c.) mimicked the effect of stress and significantly elevated NP mRNA. These results suggest that the upregulation of NP mRNA after stress is CORT-dependent and point to a role for neuropsin in stress-induced neuronal plasticity.

  20. Alpha-synuclein mRNA expression in oligodendrocytes in MSA.

    Science.gov (United States)

    Asi, Yasmine T; Simpson, Julie E; Heath, Paul R; Wharton, Stephen B; Lees, Andrew J; Revesz, Tamas; Houlden, Henry; Holton, Janice L

    2014-06-01

    Multiple system atrophy (MSA) is a progressive neurodegenerative disease presenting clinically with parkinsonian, cerebellar, and autonomic features. α-Synuclein (αsyn), encoded by the gene SNCA, is the main constituent of glial cytoplasmic inclusion (GCI) found in oligodendrocytes in MSA, but the methods of its accumulation have not been established. The aim of this study is to investigate alterations in regional and cellular SNCA mRNA expression in MSA as a possible substrate for GCI formation. Quantitative reverse transcription polymerase chain reaction (qPCR) was performed on postmortem brain samples from 15 MSA, 5 IPD, and 5 control cases to investigate regional expression in the frontal and occipital regions, dorsal putamen, pontine base, and cerebellum. For cellular expression analysis, neurons and oligodendrocytes were isolated by laser-capture microdissection from five MSA and five control cases. SNCA mRNA expression was not significantly different between the MSA, IPD and control cases in all regions (multilevel model, P = 0.14). After adjusting for group effect, the highest expression was found in the occipital cortex while the lowest was in the putamen (multilevel model, P MSA oligodendrocytes expressed more SNCA than control oligodendrocytes and expression in MSA neurons was slightly lower than that in controls, however, these results did not reach statistical significance. We have demonstrated regional variations in SNCA expression, which is higher in cortical than subcortical regions. This study is the first to demonstrate SNCA mRNA expression by oligodendrocytes in human postmortem tissue using qPCR and, although not statistically significant, could suggest that this may be increased in MSA compared to controls.

  1. Connective tissue and its growth factor CTGF distinguish the morphometric and molecular remodeling of the bladder in a model of neurogenic bladder.

    Science.gov (United States)

    Altuntas, Cengiz Z; Daneshgari, Firouz; Izgi, Kenan; Bicer, Fuat; Ozer, Ahmet; Sakalar, Cagri; Grimberg, Kerry O; Sayin, Ismail; Tuohy, Vincent K

    2012-11-01

    We previously reported that mice with experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis (MS), develop profound urinary bladder dysfunction. Because neurogenic bladder in MS patients causes marked bladder remodeling, we next examined morphometric and molecular alterations of the bladder in EAE mice. EAE was created in female SJL/J mice by immunization with the p139-151 encephalitogenic peptide of myelin proteolipid protein in complete Freund's adjuvant, along with intraperitoneal injections of Bordetella pertussis toxin. Seventy days after immunization, mice were scored for the level of neurological impairment and then killed. Spinal cord sections were assessed for demyelination, inflammation, and T cell infiltration; the composition of the bladder tissue was measured quantitatively; and gene expression of markers of tissue remodeling and fibrosis was assessed. A significant increase in the bladder weight-to-body weight ratio was observed with increasing neurological impairment, and morphometric analysis showed marked bladder remodeling with increased luminal area and tissue hypertrophy. Despite increased amounts of all tissue components (urothelium, smooth muscle, and connective tissue), the ratio of connective tissue to muscle increased significantly in EAE mice compared with control mice. Marked increases in mRNA expression of collagen type I α(2), tropoelastin, transforming growth factor-β3, and connective tissue growth factor (CTGF) were observed in EAE mice, as were decreased levels of mRNAs for smooth muscle myosin heavy chain, nerve growth factors, and muscarinic and purinergic receptors. Our results suggest that bladder remodeling corresponding to EAE severity may be due to enhanced expression of CTGF and increased growth of connective tissue.

  2. Rapamycin regulates connective tissue growth factor expression of lung epithelial cells via phosphoinositide 3-kinase.

    Science.gov (United States)

    Xu, Xuefeng; Wan, Xuan; Geng, Jing; Li, Fei; Yang, Ting; Dai, Huaping

    2013-09-01

    The pathogenesis of idiopathic pulmonary fibrosis (IPF) remains largely unknown. It is believed that IPF is mainly driven by activated alveolar epithelial cells that have a compromised migration capacity, and that also produce substances (such as connective tissue growth factor, CTGF) that contribute to fibroblast activation and matrix protein accumulation. Because the mechanisms regulating these processes are unclear, the aim of this study was to determine the role of rapamycin in regulating epithelial cell migration and CTGF expression. Transformed epithelial cell line A549 and normal human pulmonary alveolar or bronchial epithelial cells were cultured in regular medium or medium containing rapamycin. Real time reverse transcriptase polymerase chain reaction was employed to determine CTGF mRNA expression. Western blotting and an enzyme-linked immunosorbent assay were used for detecting CTGF protein. Wound healing and migration assays were used to determine the cell migration potential. Transforming growth factor (TGF)-β type I receptor (TβRI) inhibitor, SB431542 and phosphoinositide 3-kinase (PI3K) inhibitor, LY294002 were used to determine rapamycin's mechanism of action. It was found that treatment of A549 and normal human alveolar or bronchial epithelial cells with rapamycin significantly promoted basal or TGF-β1 induced CTGF expression. LY294002, not SB431542 attenuated the promotional effect of rapamycin on CTGF expression. Cell mobility was not affected by rapamycin in wound healing and migration assays. These data suggest rapamycin has a profibrotic effect in vitro and underscore the potential of combined therapeutic approach with PI3K and mammalian target of rapamycin inhibitors for the treatment of animal or human lung fibrosis.

  3. Differential expression of melanopsin mRNA and protein in Brown Norwegian rats.

    Science.gov (United States)

    Hannibal, Jens; Georg, Birgitte; Fahrenkrug, Jan

    2013-01-01

    Melanopsin is expressed in a subpopulation of retinal ganglion cells rendering these cells intrinsically photosensitive (ipRGCs). The ipRGCs are the primary RGCs mediating light entrainment of the circadian clock and control of the pupillary light reflex, light regulated melatonin secretion and negative masking behaviour. Previous studies have demonstrated that melanopsin expression in albino rats is regulated by light and darkness. The present study was undertaken to study the influence of light and darkness during the circadian day and after extended periods of constant light and darkness on melanopsin expression in the pigmented retina of the Brown Norwegian rat (Rattus norvegicus). The diurnal and circadian expressions were examined in retinal extracts from rats euthanized every 4 h during a 24 h light/dark (LD) and a 24 h dark cycle (DD) using quantitative real-time PCR and Western blotting. To study whether light regulates melanopsin expression, rats were sacrificed after being placed in either constant light (LL) or darkness for 3 or 21 d. Flat mount retinas from animals kept during either LL or DD were also examined by immunohistochemistry. Melanopsin mRNA expression displayed a significant rhythmic change during the LD cycle with peak expression around dusk and nadir at dawn. Melanopsin protein also changed over the LD cycle with peak expression at the end of the night and nadir at dusk. Rhythmic expression of melanopsin mRNA but not melanopsin protein was found in constant darkness. After 3 or 21 d in either LL or DD melanopsin mRNA expression was unaltered. Melanopsin protein was at the same high level after 3 and 21 d in DD, whereas a significant decrease was found after prolonging the light period for 3 or 21 d. The change in melanopsin protein was primarily due to change in immunoreactivity in the dendritic processes. In conclusion we found that light and darkness are important for regulation of melanopsin protein expression whereas input from a

  4. Local IGFBP-3 mRNA expression, apoptosis and risk of colorectal adenomas

    Directory of Open Access Journals (Sweden)

    Omofoye Oluwaseun

    2008-05-01

    Full Text Available Abstract Background IGF binding protein-3 (IGFBP-3 regulates the bioavailability of insulin-like growth factors I and II, and has both anti-proliferative and pro-apoptotic properties. Elevated plasma IGFBP-3 has been associated with reduced risk of colorectal cancer (CRC, but the role of tissue IGFBP-3 is not well defined. We evaluated the association between tissue or plasma IGFBP-3 and risk of colorectal adenomas or low apoptosis. Methods Subjects were consenting patients who underwent a clinically indicated colonoscopy at UNC Hospitals and provided information on diet and lifestyle. IGFBP-3 mRNA in normal colon was assessed by real time RT-PCR. Plasma IGFBP-3 was measured by ELISA and apoptosis was determined by morphology on H & E slides. Logistic regression was used to compute odds ratio (OR and 95% confidence intervals. Results We observed a modest correlation between plasma IGFBP-3 and tissue IGFBP-3 expression (p = 0.007. There was no significant association between plasma IGFBP-3 and adenomas or apoptosis. Tissue IGFBP-3 mRNA expression was significantly lower in cases than controls. Subjects in the lowest three quartiles of tissue IGFBP-3 gene expression were more likely to have adenomas. Consistent with previous reports, low apoptosis was significantly associated with increased risk of adenomas (p = 0.003. Surprisingly, local IGFBP-3 mRNA expression was inversely associated with apoptosis. Conclusion Low expression of IGFBP-3 mRNA in normal colonic mucosa predicts increased risk of adenomas. Our findings suggest that local IGFBP-3 in the colon may directly increase adenoma risk but IGFBP-3 may act through a pathway other than apoptosis to influence adenoma risk.

  5. Cytokine mRNA expression in lesions in cats with chronic gingivostomatitis.

    Science.gov (United States)

    Harley, R; Helps, C R; Harbour, D A; Gruffydd-Jones, T J; Day, M J

    1999-07-01

    Semiquantitative reverse transcription-PCR assays were developed to measure feline interleukin-2 (IL-2), IL-4, IL-5, IL-6, IL-10, and IL-12 (p35 & p40); gamma interferon (IFN-gamma); and glyceraldehyde-3-phosphate dehydrogenase mRNA concentrations in biopsies of feline oral mucosa. Biopsies were collected from 30 cats with chronic gingivostomatitis (diseased) prior to each cat receiving one of four treatments. In 23 cases replicate biopsies were collected 3 months after treatment commenced. Biopsies were also analyzed from 11 cats without clinical disease (nondiseased). Expression of IL-2, IL-10, IL-12 (p35 and p40), and IFN-gamma was detected in most nondiseased biopsies, while IL-6 was detected in a minority, and IL-4 and IL-5 were both undetectable. Compared to nondiseased cats, the diseased population showed a significant increase in the relative mRNA expression of IL-2, IL-4, IL-6, IL-10, IL-12 (p35 and p40), and IFN-gamma. In contrast, IL-5 mRNA expression was unchanged and was only detected in one case. No significant relationship was demonstrable between the change in relative expression of specific cytokine mRNA and the change in clinical severity of the local mucosal lesions over the treatment period. The results demonstrate that the normal feline oral mucosa is biased towards a predominantly (Th) type 1 profile of cytokine expression and that during the development of lesions seen in feline chronic gingivostomatitis there is a shift in the cytokine profile from a type 1 to a mixed type 1 and type 2 response.

  6. Changes of glucocorticoid receptor mRNA expression in basolateral amygdale-kindled rats

    Institute of Scientific and Technical Information of China (English)

    BAO Guan-shui; CHENG Xu-qin; HUA Yin; WANG Zhe-dong; LIU Zhen-guo

    2011-01-01

    Background Glucocorticoid receptor (GR) is believed to be a major factor in brain maturation and in modulation of a series of brain activity.Hippocampal neurons are abundant in glucocorticoid receptor,and there is significant change in GR expression under certain pathological state.Epilepsy is a special pathological state of the central nervous system.This study aimed to explore the role of GR in epilepsy by observing the change and functions of GR in hippocampus with a basolateral amygdale-electrical kindled rat epilepsy model.Methods Firstly,we established the basolateral amygdale-electrical kindled rat epilepsy model.Then GR mRNA expression in the hippocampus was assayed by semi-quantitative reverse transcription-PCR in this experiment.In addition,the processes of epileptic seizures were observed and electroencephalograms were recorded.One-way analysis of variance (ANOVA) was employed for comparing means of multiple groups,followed Fisher's least significant difference (LSD) for paired comparison.Results The rats were successfully kindled after an average of (13.50±3.99) times electrical stimulation,in which it was showed that GR mRNA expression reduced obviously as compared with the control group and the sham groups (P<0.001).The down-regulation of GR mRNA expression was abated or reversed by some anti-epilepsy drugs (P <0.001 compared with the epilepsy group),accompanied by attenuation of seizures and improvement of electroencephalograms.Conclusions Down-regulation of hippocampal GR mRNA expression may be related to the kindling.Anti-epilepsy drugs exposure can retard this change.

  7. Substance P mRNA expression in the rat spinal cord following selective brachial plexus injury

    Institute of Scientific and Technical Information of China (English)

    Na Liu; Longju Chen; Feng Li; Wutian Wu

    2008-01-01

    BACKGROUND: The neuropeptide, substance P, has various bioactivities and is widely distributed in the central nervous system. Substance P participates in neural transmission in the spinal cord and plays an important role in regeneration and repair of nerve injury.OBJECTIVE: To investigate substance P mRNA expression in the anterior horn of the spinal cord following brachial plexus injury.DESIGN, TIME AND SETTING: A molecular cell biology randomized controlled study was performed at the Department of Anatomy, Zhongshan Medical College, Sun Yat-sen University and the DaAn Gene Laboratory in May 2005.MATERIALS: A total of 29 adult male Sprague Dawley rats were randomly assigned to a control group (n=5) and an injury group (n = 24).METHODS: The injury group was divided into three subgroups. In subgroup A, the right seventh cervical vertebra (C7) anterior root was avulsed, and the residual nerve root at the distal end was removed. In subgroup B, the right C7 anterior root was avulsed, and the right C5 first thoracic vertebrae (TO posterior root was incised. Thus afferent pathways of the posterior root that connected with the anterior horn motor neurons were blocked. In subgroup C, the right C7 anterior root was avulsed, and a right C5-6 hemisection was performed. Thus the descending fiber pathways of the cortex that connected with anterior horn motor neurons were blocked. In the control group, the C5-T1 vertebral plate was opened, and then the skin was sutured.MAIN OUTCOME MEASURE: Substance P mRNA expression in the anterior horn of the spinal cord was quantified using fluorescent quantitative reverse transcription-polymerase chain reaction.RESULTS: Substance P mRNA expression was low in the anterior horn of the rat spinal cord in the control group. Substance P mRNA expression in the anterior horn of the spinal cord was upregulated and was significantly higher in the injury group compared with the control group (P < 0.01 ). Substance P mRNA expression was highest in

  8. Orotate phosphoribosyl transferase mRNA expression and the response of cholangiocarcinoma to 5-fluorouracil

    Institute of Scientific and Technical Information of China (English)

    Chariya Hahnvajanawong; Jariya Chaiyagool; Wunchana Seubwai; Vajarabhongsa Bhudhisawasdi; Nisana Namwat; Narong Khuntikeo; Banchob Sripa

    2012-01-01

    AIM:To determine whether expression of certain enzymes related to 5-fluorouracil (5-FU) metabolism predicts 5-FU chemosensitivity in cholangiocarcinoma (CCA).METHODS:The histoculture drug response assay (HDRA) was performed using surgically resected CCA tissues.Tumor cell viability was determined morphologically with hematoxylin and eosin-and terminal deoxynucleotide transferase-mediated dUTP nick-end labeling-stained tissues.The mRNA expression of thymidine phosphorylase (TP),orotate phosphoribosyl transferase (OPRT),thymidylate synthase (TS),and dihydropyrimidine dehydrogenase (DPD) was determined with realtime reverse transcriptase-polymerase chain reaction.The levels of gene expression and the sensitivity to 5-FU were evaluated.RESULTS:Twenty-three CCA tissues were obtained from patients who had been diagnosed with intrahepatic CCA and who underwent surgical resection at Srinagarind Hospital,Khon Kaen University from 2007 to 2009.HDRA was used to determine the response of these CCA tissues to 5-FU.Based on the dose-response curve,200 μg/mL 5-FU was selected as the test concentration.The percentage of inhibition index at the median point was selected as the cut-off point to differentiate the responding and non-responding tumors to 5-FU.When the relationship between TP,OPRT,TS and DPD mRNA expression levels and the sensitivity of CCA tissues to 5-FU was examined,only OPRT mRNA expression was significantly correlated with the response to 5-FU.The mean expression level of OPRT was significantly higher in the responder group compared to the non-responder group (0.41 ± 0.25 vs 0.22 ± 0.12,P < 0.05).CONCLUSION:OPRT mRNA expression may be a useful predictor of 5-FU chemosensitivity of CCA.Whether OPRT mRNA could be used to predict the success of 5-FU chemotherapy in CCA patients requires confirmation in patients.

  9. Anesthesia for euthanasia influences mRNA expression in healthy mice and after traumatic brain injury.

    Science.gov (United States)

    Staib-Lasarzik, Irina; Kriege, Oliver; Timaru-Kast, Ralph; Pieter, Dana; Werner, Christian; Engelhard, Kristin; Thal, Serge C

    2014-10-01

    Tissue sampling for gene expression analysis is usually performed under general anesthesia. Anesthetics are known to modulate hemodynamics, receptor-mediated signaling cascades, and outcome parameters. The present study determined the influence of anesthetic paradigms typically used for euthanization and tissue sampling on cerebral mRNA expression in mice. Naïve mice and animals with acute traumatic brain injury induced by controlled cortical impact (CCI) were randomized to the following euthanasia protocols (n=10-11/group): no anesthesia (NA), 1 min of 4 vol% isoflurane in room air (ISO), 3 min of a combination of 5 mg/kg midazolam, 0.05 mg/kg fentanyl, and 0.5 mg/kg medetomidine intraperitoneally (COMB), or 3 min of 360 mg/kg chloral hydrate intraperitoneally (CH). mRNA expression of actin-1-related gene (Act1), FBJ murine osteosarcoma viral oncogene homolog B (FosB), tumor necrosis factor alpha (TNFα), heat shock protein beta-1 (HspB1), interleukin (IL)-6, tight junction protein 1 (ZO-1), IL-1ß, cyclophilin A, micro RNA 497 (miR497), and small cajal body-specific RNA 17 were determined by real-time polymerase chain reaction (PCR) in hippocampus samples. In naïve animals, Act1 expression was downregulated in the CH group compared with NA. FosB expression was downregulated in COMB and CH groups compared with NA. CCI reduced Act1 and FosB expression, whereas HspB1 and TNFα expression increased. After CCI, HspB1 expression was significantly higher in ISO, COMB, and CH groups, and TNFα expression was elevated in ISO and COMB groups. MiR497, IL-6, and IL-1ß were upregulated after CCI but not affected by anesthetics. Effects were independent of absolute mRNA copy numbers. The data demonstrate that a few minutes of anesthesia before tissue sampling are sufficient to induce immediate mRNA changes, which seem to predominate in the early-regulated gene cluster. Anesthesia-related effects on gene expression might explain limited reproduciblity of real

  10. Expression and its Clinical Signiifcance of CK19 mRNA in Peripheral Blood of Patients with Gastric Cancer

    Institute of Scientific and Technical Information of China (English)

    Zhu Minghui; Zhu Li; Qin Youjuan; Chen Lin

    2014-01-01

    Objective:To investigate the expression of targeted gene CK19 mRNA in peripheral blood of the patients with gastric cancer and its correlation with biological behaviors of gastric cancer. Methods: CK19 mRNA in peripheral blood of 56 patients with gastric cancer was detected by RT-PCR method. Meanwhile, the peripheral blood of 12 healthy volunteers and 12 gastric cancer tissues were respectively selected as negative and positive controls. The correlation between CK19 mRNA expression in peripheral blood of the patients with gastric cancer and clinical pathological characteristics was analyzed. Results: The positive rate of CK19 mRNA in peripheral blood of 56 patients with gastric cancer was 58.9% (33/56). The expression rate of CK19 mRNA in 12 gastric cancer tissues came up to 100.0% (12/12), whereas CK19 mRNA in peripheral blood of 12 healthy volunteers was expressed negatively. CK19 mRNA expression was signiifcantly related to the clinical staging of gastric cancer and lymphatic metastasis (P0.05). Conclusion: The application of RT-PCR was highly sensitive and speciifc in detecting the CK19 mRNA in peripheral blood of patients with gastric cancer, in which CK19 mRNA is expected to be a tumor marker for judging the metastasis and recurrence and evaluating the efifcacyof gastric cancer.

  11. UCP2 mRNA expression is dependent on glucose metabolism in pancreatic islets

    DEFF Research Database (Denmark)

    Dalgaard, Louise Torp

    2012-01-01

    Uncoupling Protein 2 (UCP2) is expressed in the pancreatic β-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism f...... down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients....

  12. Progressive APOBEC3B mRNA expression in distant breast cancer metastases

    Science.gov (United States)

    Dalm, Simone U.; de Weerd, Vanja; Moelans, Cathy B.; ter Hoeve, Natalie; van Diest, Paul J.; Martens, John W. M.; van Deurzen, Carolien H. M.

    2017-01-01

    Background APOBEC3B was recently identified as a gain-of-function enzymatic source of mutagenesis, which may offer novel therapeutic options with molecules that specifically target this enzyme. In primary breast cancer, APOBEC3B mRNA is deregulated in a substantial proportion of cases and its expression is associated with poor prognosis. However, its expression in breast cancer metastases, which are the main causes of breast cancer-related death, remained to be elucidated. Patients and methods RNA was isolated from 55 primary breast cancers and paired metastases, including regional lymph node (N = 20) and distant metastases (N = 35). APOBEC3B mRNA levels were measured by RT-qPCR. Expression levels of the primary tumors and corresponding metastases were compared, including subgroup analysis by estrogen receptor (ER/ESR1) status. Results Overall, APOBEC3B mRNA levels of distant metastases were significantly higher as compared to the corresponding primary breast tumor (P = 0.0015), an effect that was not seen for loco-regional lymph node metastases (P = 0.23). Subgroup analysis by ER-status showed that increased APOBEC3B levels in distant metastases were restricted to metastases arising from ER-positive primary breast cancers (P = 0.002). However, regarding ER-negative primary tumors, only loco-regional lymph node metastases showed increased APOBEC3B expression when compared to the corresponding primary tumor (P = 0.028). Conclusion APOBEC3B mRNA levels are significantly higher in breast cancer metastases as compared to the corresponding ER-positive primary tumors. This suggests a potential role for APOBEC3B in luminal breast cancer progression, and consequently, a promising role for anti-APOBEC3B therapies in advanced stages of this frequent form of breast cancer. PMID:28141868

  13. DDR2 polymorphisms and mRNA expression in lung cancers of Japanese patients

    OpenAIRE

    Sasaki, Hidefumi; SHITARA, MASAYUKI; YOKOTA, KEISUKE; OKUDA, KATSUHIRO; HIKOSAKA, YU; MORIYAMA, SATORU; YANO, MOTOKI; Fujii, Yoshitaka

    2012-01-01

    Discoidin domain receptor 2, DDR2, is a tyrosine kinase receptor for fibrillar collagen that is involved in postnatal development, tissue repair and primary and metastatic cancer progression. Recently, mutations in the DDR2 kinase gene were identified in squamous cell lung cancer from large-scale Sanger sequencing. The present study investigated the DDR2 gene mutations and mRNA expression in surgically treated non-small cell lung cancer (NSCLC) of squamous histology cases. The presence or abs...

  14. Basal keratin expression in breast cancer by quantification of mRNA and by immunohistochemistry

    Directory of Open Access Journals (Sweden)

    Pluciennik Elzbieta

    2010-04-01

    Full Text Available Abstract Definitions of basal-like breast cancer phenotype vary, and microarray-based expression profiling analysis remains the gold standard for the identification of these tumors. Immunohistochemical identification of basal-like carcinomas is hindered with a fact, that on microarray level not all of them express basal-type cytokeratin 5/6, 14 and 17. We compared expression of cytokeratin 5, 14 and 17 in 115 patients with operable breast cancer estimated by real-time RT-PCR and immunohistochemistry. Despite the method of dichotomization and statistical analysis, there were cases with discordant results comparing immunohistochemistry and RT-PCR analysis. For dichotomisation based on quartiles and ROC, 14% of cases were negative on immunohistochemical examination for CK5/6, but presented high CK5 mRNA levels. There were also 48-55% cases, which were CK5/6-immunopositive, but were negative by mRNA examination. Similar discordances were observed for CK14 and CK17. Basal keratin mRNAs did not correlate with ER mRNA levels, while immunohistochemistry produced significant relationship with ER status. Our observation suggest that both method may produce different results in a small proportion of cases. Discordance between immunohistochemistry and RT-PCR may confound attempts to establish a simple methods for identification of basal-like tumors.

  15. Effects of Ginseng and Echinacea on Cytokine mRNA Expression in Rats

    Science.gov (United States)

    Uluışık, Deniz; Keskin, Ercan

    2012-01-01

    The aim of the study was to determine the effect of ginseng and echinacea on the mRNA expression of IL-10, TNF-α, and TGF-β1 in healthy rats. Six-week-old male Fischer 344 rats (n = 48) were used. The animals were divided into three equal groups, as follows: control (C); ginseng (G); echinacea (E). While the C group was fed a standard rat diet (Purina) ad libitum for a period of 40 days, the G and E groups animals received the same diet containing 0.5 g/kg of Panax ginseng root powder and 0.75 g/kg of Echinacea purpurea root powder, respectively. Blood samples were obtained from 8 rats in each group after 20 and 40 days of treatment, and the mRNA expression of IL-10, TNF-α, and TGF-β1 was determined. After 20 days of treatment, the expression of IL-10 mRNA in the G group was different from the C group (P echinacea on these cytokines and other cytokines is needed. PMID:22666172

  16. Hormone and metabolic factors associated with leptin mRNA expression in pre- and postmenopausal women.

    Science.gov (United States)

    Fajardo, Martha E; Malacara, Juan M; Martínez-Rodríguez, Herminia G; Barrera-Saldaña, Hugo A

    2004-06-01

    Recent information has extended leptin's action, beyond the control of appetite, to various sites of metabolic regulation. To better understand leptin's role we studied its production in subcutaneous and visceral fat compartments before and after menopause. During elective abdominal surgery, biopsies of subcutaneous and omental tissues were taken from 20 women at pre- (BMI 28.4 +/- 4.5 kg/m2) and 10 at postmenopause (BMI 30.6 +/- 7.7 kg/m2). In both groups serum leptin levels were similar, and highly correlated with BMI. In subcutaneous adipose tissue, leptin mRNA expression was significantly higher in pre- than in postmenopausal women (50.4 +/- 20.5 amol/microg total RNA versus 34.5 +/- 24.9 amol/microg total RNA, respectively). Leptin mRNA expression in subcutaneous tissue was independently correlated with fasting glucose (R = 0.89, P < 0.006) at premenopause, and with serum estradiol (R = 0.77, P < 0.04) at postmenopause. Leptin mRNA expression in visceral fat was correlated with DHEAS (R = 0.86, P < 0.001), at premenopause. These results indicate that in both compartments, leptin production is sensitive to different but overlapping stimuli, conveying information about energy availability to central and peripheral sites under different conditions of estrogen exposure.

  17. Expression of cytokine mRNA during immuno—modulation of murine suppressor macrophages

    Institute of Scientific and Technical Information of China (English)

    FENGWEIGUO; ZHONGLIANGCHANG

    1998-01-01

    In order to analyze the mechanism of immunomodulation by LPS on murine peritoneal suppressor macrophages,we have,using RNase protection assay,checked the changes of mRNA expression pattern of several cytokine genes during the immuno-modulation.It has been found that,after treating peritoneal suppressor macrophages with LPS,mRNAs of IL-12 p35,IL-12 p40,IL-6 and IFN-γ are newly appeared,while those of IL-1α,IL-1β and IL-1Ra are increaseb and those of other cytokines,like TGF-β1 and MIF are not changed at all.It seems certain that those cytokines,whose expression is increased by LPS stimulation,may be responsible for the functional changes of suppressor macrophages during immuno-modulation.Among these changes,the appearance of IL-12 mRNA may play a critical role,and,in this regard,the synergetic effect betewwn IFN-γ and LPS on the increase of IL-12 p35 and IL-12 p40 mRNA expression is an interesting finding.

  18. Distinct prognostic values of four-Notch-receptor mRNA expression in ovarian cancer.

    Science.gov (United States)

    Zhou, Xinling; Teng, Lingling; Wang, Min

    2016-05-01

    Notch signaling pathway includes ligands and Notch receptors, which are frequently deregulated in several human malignancies including ovarian cancer. Aberrant activation of Notch signaling has been linked to ovarian carcinogenesis and progression. In the current study, we used the "Kaplan-Meier plotter" (KM plotter) database, in which updated gene expression data and survival information from a total of 1306 ovarian cancer patients were used to access the prognostic value of four Notch receptors in ovarian cancer patients. Hazard ratio (HR), 95 % confidence intervals, and log-rank P were calculated. Notch1 messenger RNA (mRNA) high expression was not found to be correlated to overall survival (OS) for all ovarian cancer, as well as in serous and endometrioid cancer patients followed for 20 years. However, Notch1 mRNA high expression is significantly associated with worsen OS in TP53 wild-type ovarian cancer patients, while it is significantly associated with better OS in TP53 mutation-type ovarian cancer patients. Notch2 mRNA high expression was found to be significantly correlated to worsen OS for all ovarian cancer patients, as well as in grade II ovarian cancer patients. Notch3 mRNA high expression was found to be significantly correlated to better OS for all ovarian cancer patients, but not in serous cancer patients and endometrioid cancer patients. Notch4 mRNA high expression was not found to be significantly correlated to OS for all ovarian cancer patients, serous cancer patients, and endometrioid cancer patients. These results indicate that there are distinct prognostic values of four Notch receptors in ovarian cancer. This information will be useful for better understanding of the heterogeneity and complexity in the molecular biology of ovarian cancer and for developing tools to more accurately predict their prognosis. Based on our results, Notch1 could be a potential drug target of TP53 wild-type ovarian cancer and Notch2 could be a potential drug

  19. Regulation and function of FTO mRNA expression in human skeletal muscle and subcutaneous adipose tissue

    DEFF Research Database (Denmark)

    Grunnet, Louise G; Nilsson, Emma; Ling, Charlotte

    2009-01-01

    and adipose tissue, and their influence on in vivo glucose and fat metabolism. Research Design and Methods. The FTO rs9939609 polymorphism was genotyped in two twin cohorts: 1) 298 elderly twins aged 62-83 years with glucose tolerance ranging from normal to type 2 diabetes and 2) 196 young (25-32 years...... expression was not influenced by FTO rs9939609 genotype. FTO mRNA expression in skeletal muscle was regulated by age and sex, whereas age and BMI were predictors of adipose tissue FTO mRNA expression. FTO mRNA expression in adipose tissue was associated with an atherogenic lipid profile. In skeletal muscle...

  20. Sequencing of mRNA identifies re-expression of fetal splice variants in cardiac hypertrophy.

    Science.gov (United States)

    Ames, E G; Lawson, M J; Mackey, A J; Holmes, J W

    2013-09-01

    Cardiac hypertrophy has been well-characterized at the level of transcription. During cardiac hypertrophy, genes normally expressed primarily during fetal heart development are re-expressed, and this fetal gene program is believed to be a critical component of the hypertrophic process. Recently, alternative splicing of mRNA transcripts has been shown to be temporally regulated during heart development, leading us to consider whether fetal patterns of splicing also reappear during hypertrophy. We hypothesized that patterns of alternative splicing occurring during heart development are recapitulated during cardiac hypertrophy. Here we present a study of isoform expression during pressure-overload cardiac hypertrophy induced by 10 days of transverse aortic constriction (TAC) in rats and in developing fetal rat hearts compared to sham-operated adult rat hearts, using high-throughput sequencing of poly(A) tail mRNA. We find a striking degree of overlap between the isoforms expressed differentially in fetal and pressure-overloaded hearts compared to control: forty-four percent of the isoforms with significantly altered expression in TAC hearts are also expressed at significantly different levels in fetal hearts compared to control (Phypertrophy and fetal heart development are significantly enriched for genes involved in cytoskeletal organization, RNA processing, developmental processes, and metabolic enzymes. Our data strongly support the concept that mRNA splicing patterns normally associated with heart development recur as part of the hypertrophic response to pressure overload. These findings suggest that cardiac hypertrophy shares post-transcriptional as well as transcriptional regulatory mechanisms with fetal heart development. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. Identification of phylogenetically conserved enhancer elements implicated in pancreas development in the WISP1 and CTGF orthologs.

    Science.gov (United States)

    Kapasa, M; Serafimidis, I; Gavalas, A; Kossida, S

    2008-11-01

    WISP1 and CTGF are members of the CCN family of growth factors encoding extracellular matrix proteins participating in several developmental and tumorigenic processes. Both are induced by the WNT signaling pathway, and microarray data suggest that expression of WISP1 and CTGF is repressed by Neurogenin3 (Ngn3 (NEUROG3)), a transcription factor directing specification of the endocrine pancreas. Single-cell reverse transcription polymerase chain reaction analysis suggested that this was a cell autonomous effect. To identify possible common regulatory networks involved in WISP1 and CTGF gene expression, their genomic regions were searched for common transcription factor motifs using a combination of in silico approaches and documented knowledge concerning pancreas development. This analysis revealed the presence of a conserved enhancer in both CTGF and WISP1 regulatory regions in 10 species covering a wide evolutionary distance. This enhancer contains binding sites for Ngn1/3 (NEUROG1/3) and transcription factors that are critically involved in pancreas development. Furthermore, it contained binding sites for three additional transcription factor families, which may indicate novel players are involved in this process.

  2. Proprotein convertase 1 mRNA and protein expression in ischemic rat cortex after reperfusion

    Institute of Scientific and Technical Information of China (English)

    Shuqin Zhan; An Zhou; Jingquan Lan; Tao Yang

    2011-01-01

    Proprotein convertase 1 (PC1) is a member of the family of proprotein convertases (PCs), which are the processing enzymes of neuropeptides. Previous studies have addressed PC1 effects with regard to the neuroendocrine system. In this study, the developing changes of PC1 mRNA and PC1 protein in rat cortices after transient focal cerebral ischemia were investigated by fluorescent double labeling (both in situ hybridization and immunocytochemistry) using a transient focal cerebral ischemia model in rats. The results were compared with those of sham-operated rat cortices. Both the mRNA and protein levels of PC1 in ischemic cortices decreased gradually at 4, 8, and 16 hours of reperfusion after 100 minutes of middle cerebral artery occlusion. After 24 hours of reperfusion, enhanced intensities of signals for PC1 protein were observed, while signals for PC1 mRNA remained low. These results suggest that transient focal cerebral ischemia influences PC1 mRNA and protein expression in cortices of ischemic rats. Thus, PC1 is regulated by ischemic stress.

  3. DDR2 polymorphisms and mRNA expression in lung cancers of Japanese patients.

    Science.gov (United States)

    Sasaki, Hidefumi; Shitara, Masayuki; Yokota, Keisuke; Okuda, Katsuhiro; Hikosaka, Yu; Moriyama, Satoru; Yano, Motoki; Fujii, Yoshitaka

    2012-07-01

    Discoidin domain receptor 2, DDR2, is a tyrosine kinase receptor for fibrillar collagen that is involved in postnatal development, tissue repair and primary and metastatic cancer progression. Recently, mutations in the DDR2 kinase gene were identified in squamous cell lung cancer from large-scale Sanger sequencing. The present study investigated the DDR2 gene mutations and mRNA expression in surgically treated non-small cell lung cancer (NSCLC) of squamous histology cases. The presence or absence of DDR2 mutations at the kinase and discoidin domain was analyzed by direct sequencing. In this cohort, DDR2 mutations were not observed in the 166 patients with lung cancer, although DDR2 polymorphisms were observed (H136H, n=14) at the discoidin domain. mRNA levels of DDR2 in lung tumor samples and the adjacent normal lung samples were simultaneously analyzed. DDR2 mRNA levels were significantly decreased in tumor samples compared with normal lung samples. However, the DDR2 mRNA levels were elevated in the DDR2 polymorphism cases.

  4. Expression of growth hormone receptor and its mRNA in hepatic cirrhosis

    Institute of Scientific and Technical Information of China (English)

    Hong-Tao Wang; Shuang Chen; Jie Wang; Qing-Jia Ou; Chao Liu; Shu-Sen Zheng; Mei-Hai Deng; Xiao-Ping Liu

    2003-01-01

    AIM: To investigate the expression of growth hormone receptor (GHR) and mRNA of GHR in cirrhotic livers of rats with the intension to find the basis for application of recombinant human growth hormone (rhGH) to patients with liver cirrhosis.METHODS: Hepatic cirrhosis was induced in SpragueDawley rats by administration of thioacetamide intraperitoneally for 9-12 weeks. Collagenase Ⅳ was perfused in situ for isolation of hepatocytes. The expression of GHR and its mRNA in cirrhotic livers was studied with radio-ligand binding assay, RT-PCR and digital image analysis.RESULTS: One class of specific growth hormone-binding site, GHR, was detected in hepatocytes and hepatic tissue of cirrhotic livers. The binding capacity of GHR (RT, fmol/mg protein) in rat cirrhotic liver tissue (30.8±1.9) was significantly lower than that in normal control (74.9±3.9) at the time point of the ninth week after initiation of induction of cirrhosis (n=10, P<0.05), and it decreased gradually along with the accumulation of collagen in the process of formation and development of liver cirrhosis (P<0.05). The number of binding sites (×10 4/cell) of GHR on rat cirrhotic hepatocytes (0.86±0.16) was significantly lower than that (1.28±0.24)in control (n= 10, P<0.05). The binding affinity of GHR among liver tissue, hepatocytes of various groups had no significant difference (P>0.05). The expression of GHR mRNA (riOD,pixel) in rat cirrhotic hepatic tissues (23.3±3.1) was also significantly lower than that (29.3±3.4) in normal control (n=10, P<0.05).CONCLUSION: The growth hormone receptor was expressed in a reduced level in liver tissue of cirrhotic rats,and lesser expression of growth hormone receptors was found in a later stage of cirrhosis. The reduced expression of growth hormone receptor was partly due to its decreased expression on cirrhotic hepatocytes and the reduced expression of its mRNA in cirrhotic liver tissue.

  5. Keratinocyte growth factor mRNA expression in periodontal ligament fibroblasts

    DEFF Research Database (Denmark)

    Dabelsteen, S; Wandall, H H; Grøn, B

    1997-01-01

    Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF mRNA is expres......Keratinocyte growth factor (KGF) is a fibroblast growth factor which mediates epithelial growth and differentiation. KGF is expressed in subepithelial fibroblasts, but generally not in fibroblasts of deep connective tissue, such as fascia and ligaments. Here we demonstrate that KGF m...

  6. Modulation of IGFBP2 mRNA expression in white adipose tissue upon aging and obesity.

    Science.gov (United States)

    Li, Z; Picard, F

    2010-10-01

    The insulin/IGF-1 signaling pathway is a determinant of aging and age-related diseases. IGF-binding protein 2 (IGFBP2) is secreted by white adipocytes and contributes to the prevention of diet-induced obesity and age-related insulin resistance in mice. However, the expression levels of IGFBP2 in insulin resistance disorders have not been evaluated. The present study was aimed at determining IGFBP2 mRNA levels in adipose tissue in conditions of insulin resistance such as aging and obesity. In visceral white adipose tissue (WAT), but not in subcutaneous WAT, IGFBP2 mRNA levels were significantly lower in obese OB/OB, DB/DB and high fat-fed mice compared with those of their respective lean and chow-fed littermates. IGFBP2 mRNA levels were also decreased in visceral WAT of 12 and 24 months old mice compared with those of their 4 months old counterparts. Visceral WAT IGFBP2 expression was significantly associated with IGFBP2 circulating levels in mice, suggesting an important contribution from this tissue. The negative effect of aging on IGFBP2 mRNA levels in visceral WAT was confirmed in obese men. These findings demonstrate that the transcription of the IGFBP2 gene is modulated in a depot-specific fashion in obesity and aging in mice and men. Because IGFBP2 is an adipokine, an altered production from visceral WAT depots could impact on IGF-1 signaling and its downstream targets. This supports the need for further molecular and clinical studies to determine the factors regulating IGFBP2 expression and its relevance to metabolic diseases.

  7. Optimization of mRNA design for protein expression in the crustacean Daphnia magna.

    Science.gov (United States)

    Törner, Kerstin; Nakanishi, Takashi; Matsuura, Tomoaki; Kato, Yasuhiko; Watanabe, Hajime

    2014-08-01

    The water flea Daphnia is a new model organism for ecological, evolutionary, and toxicological genomics. Detailed functional analysis of genes newly discovered through genomic approaches often requires overexpression of the identified protein. In the present study, we report the microinjection of in vitro-synthesized RNAs into the eggs as a method for overexpressing ubiquitous proteins in Daphnia magna. We injected a 1.3-kb mRNA that coded for the red fluorescent protein (DsRed2) flanked by UTRs from the ubiquitously expressed elongation factor 1α-1 (EF1α-1) into D. magna embryos. DsRed2 fluorescence in the embryos was measured 24 h after microinjection. Unexpectedly, the reporter RNA containing the 522-bp full-length EF1α-1 3' UTR failed to induce fluorescence. To assess reporter expression, the length of the 3' UTR that potentially contained negative regulatory elements of protein expression, including AU-rich regions and Musashi binding elements, was serially reduced from the 3' end. Assessing all injected RNA alternatives, mRNA containing the first 60 bp of the 3' UTR gave rise to the highest fluorescence, 14 times the Daphnia auto-fluorescence. In contrast, mRNA lacking the entire 3' UTR hardly induced any change in fluorescence intensity. This is the first evaluation of UTRs of mRNAs delivered into Daphnia embryos by microinjection for overexpressing proteins. The mRNA with truncated 3' UTRs of Daphnia EF1α-1 will be useful not only for gain-of-function analyses but also for labeling proteins and organelles with fluorescent proteins in Daphnia.

  8. Expression of connective tissue growth factor in tumor tissues is an independent predictor of poor prognosis in patients with gastric cancer

    Institute of Scientific and Technical Information of China (English)

    Lu-Ying Liu; Yan-Chun Han; Shu-Hua Wu; Zeng-Hua Lv

    2008-01-01

    AIM: To examine the expression of connective tissue growth factor (CTGF), also known as CCN2, in gastric carcinoma (GC), and the correlation between the expression of CTGF, dinicopathologic features and clinical outcomes of patients with GC.METHODS: One hundred and twenty-two GC patients were included in the present study. All patients were followed up for at least 5 years. Proteins of CTGF were detected using the Powervision two-step immunostaining method.RESULTS: Of the specimens from 122 GC patients analyzed for CTGF expression, 58 (58/122, 47.5%) had a high CTGF expression in cytoplasm of gastric carcinoma cells and 64 (64/122, 52.5%) had a low CTGF expression. Patients with a high CTGF expression showed a higher incidence of lymph node metastasis than those with a low CTGF expression {P = 0.032). Patients with a high CTGF expression had significantly lower 5-year survival rate than those with a low CTGF expression (27.6% vs 46.9%, P = 0.0178), especially those staging I + H + m (35.7% vs 65.2%, P = 0.0027).CONCLUSION: GC patients with an elevated CTGF expression have more lymph node metastases and a shorter survival time. CTGF seems to be an independent prognostic factor for the successful differentiation of high-risk GC patients staging I + n + m. Over-expression of CTGF in human GC cells results in an increased aggressive ability.

  9. Expression of brain-derived neurotrophic factor mRNA in rat hippocampus after treatment with antipsychotic drugs.

    Science.gov (United States)

    Bai, Ou; Chlan-Fourney, Jennifer; Bowen, Rudy; Keegan, David; Li, Xin-Min

    2003-01-01

    Typical and atypical antipsychotic drugs, though both effective, act on different neurotransmitter receptors and are dissimilar in some clinical effects and side effects. The typical antipsychotic drug haloperidol has been shown to cause a decrease in the expression of brain-derived neurotrophic factor (BDNF), which plays an important role in neuronal cell survival, differentiation, and neuronal connectivity. However, it is still unknown whether atypical antipsychotic drugs similarly regulate BDNF expression. We examined the effects of chronic (28 days) administration of typical and atypical antipsychotic drugs on BDNF mRNA expression in the rat hippocampus using in situ hybridization. Quantitative analysis revealed that the typical antipsychotic drug haloperidol (1 mg/kg) down-regulated BDNF mRNA expression in both CA1 (P BDNF mRNA expression in CA1, CA3, and dentate gyrus regions of the rat hippocampus compared with their respective controls (P BDNF mRNA expression in rat hippocampus.

  10. Hormone therapy modulates ET(A) mRNA expression in the aorta of ovariectomised New Zealand White rabbits

    DEFF Research Database (Denmark)

    Pedersen, Susan Helene; Nielsen, Lars Bo; Pedersen, Nina Gros

    2009-01-01

    ) + NETA, CEE + MPA or placebo. The thoracic aorta and the epicardial coronary artery were used for mRNA expression and myograph analyses, respectively. RESULTS: E(2) and CEE alone significantly reduced ET-1 receptor subtype A (ET(A)) mRNA expression compared with placebo treatment. The E(2)-induced...... reduction in ET(A) mRNA expression persisted with the co-administration of NETA, but the CEE induced reduction in ET(A) mRNA expression was not maintained with the co-administration of MPA. Treatment with CEE alone significantly increased endotelin-1 converting enzyme (ECE) mRNA expression and CEE combined...... with MPA reduced prepro-endothelin-1 (ppET-1) mRNA expression when compared with placebo. ET-1 receptor subtype B mRNA expression and ET-1 induced vasocontraction was unaffected by treatment. CONCLUSIONS: E(2) and CEE treatment exert potentially beneficial vascular effects through regulation of the ET(A...

  11. mRNA expression profiling reveals a role of Helicobacter pylorivacuolating toxin in escaping host defense

    Institute of Scientific and Technical Information of China (English)

    Jian-Ping Yuan; Tao Li; Zhen-Hong Li; Gui-Zhen Yang; Bao-Yu Hu; Xiao-Dong Shi; Tie-Liu Shi; Shan-Qing Tong; Xiao-Kui Guo

    2004-01-01

    AIM: To study the immune response of host to Helicobacter pylori VacA.METHODS: The monocyte/macrophage-like U937 cells were infected with Helicobacter pylori vacA-positive strain NCTC 11638 or isogenic vacA-negative mutant. Differentially expressed genes were identified at 2, 6, 10, and 24 h postinfection by cDNA microarray. Differential expressions of some genes were confirmed by Northern blot.RESULTS: More than 100 genes altered their mRNA expression at different time points respectively, many of which were identified to be related to immune evasion.CONCLUSION: VacA is a crucial element for H pylorito escape from host immune defense by means of differentially regulating the expression of some related genes. These genes, previously known or unknown to be involved in the mechanism of immune evasion, deserve further investigation to unearth much more information complicated in the immune response.

  12. Effect of running training on uncoupling protein mRNA expression in rat brown adipose tissue

    Science.gov (United States)

    Yamashita, Hitoshi; Yamamoto, Mikio; Sato, Yuzo; Izawa, Tetsuya; Komabayashi, Takao; Saito, Daizo; Ohno, Hideki

    1993-03-01

    The effect was investigated of endurance training on the expression of uncoupling protein (UCP) mRNA in brown adipose tissue (BAT) of rats. The exercised rats were trained on a rodent treadmill for 5 days per week and a total of 9 weeks. After the training programme, a marked decrease in BAT mass was found in terms of weight or weight per unit body weight; there was a corresponding decrease in DNA content and a downward trend in RNA and glycogen levels. The UCP mRNA was present at a markedly decreased level in BAT of trained animals. In consideration of the reduced levels of mRNAs for hormone-sensitive lipase and acylCoA synthetase, the brown adipose tissue investigated appeared to be in a relatively atrophied and thermogenically quiescent state.

  13. Induction of Ski protein expression upon luteinization in rat granulosa cells without a change in its mRNA expression.

    Science.gov (United States)

    Kim, Hyun; Yamanouchi, Keitaro; Matsuwaki, Takashi; Nishihara, Masugi

    2012-01-01

    The Ski protein is implicated in the proliferation/differentiation of a variety of cells. We previously reported that the Ski protein is present in granulosa cells of atretic follicles, but not in preovulatory follicles, suggesting that Ski has a role in apoptosis of granulosa cells. However, granulosa cells cannot only undergo apoptosis but can alternatively differentiate into luteal cells. It is unknown whether Ski is expressed and has a role in granulosa cells undergoing luteinization. Thus, the aim of the present study was to determine the localization of the Ski protein in the rat ovary during luteinization to examine if Ski might play a role in this process. In order to examine the Ski protein expression during the progression of luteinization, follicular growth was induced in immature female rats by administration of equine chorionic gonadotropin, and luteinization was induced by human chorionic gonadotropin treatment to mimic the luteinizing hormone (LH) surge. While no Ski-positive granulosa cells were present in the preovulatory follicle, Ski protein expression was induced in response to the LH surge and was maintained after formation of the corpus luteum (CL). Although the Ski protein is absent from the granulosa cells of the preovulatory follicle, its mRNA (c-ski) was expressed, and the level of c-ski mRNA was unchanged even after the LH surge. The combined results demonstrated that Ski protein expression is induced in granulosa cells upon luteinization, and suggested that its expression is regulated posttranscriptionally.

  14. Changes of mRNA expression of enkephalin and prodynorphin in hippocampus of rats with chronic immobilization stress

    Institute of Scientific and Technical Information of China (English)

    Jia-Xu Chen; Wei Li; Xin Zhao; Jian-Xin Yang; Hong-Yan Xu; Zhu-Feng Wang; Guang-Xin Yue

    2004-01-01

    AIM: To observe the changes of enkephalin mRNA and prodynorphin mRNA in hippocampus of rats induced by chronic immobilization stress.METHODS: Thirty rats were randomly divided into three groups of 10 each: the normal control group (group A),the group induced by chronic immobilization stress for 7 d (group B) and the group induced by chronic immobilization stress for 21 d (group C). The changes of the enkephalin mRNA and prodynorphin mRNA in the rat hippocampus were detected by reverse transcription-polymerase chain reaction (RT-PCR).RESULTS: Expression levels of enkephalin mRNA and prodynorphin mRNA in rat hippocampus were significantly increased under chronic immobilization stress, and the expression of prodynorphin mRNA in the rat hippocampus in group C was remarkably higher than that in group B (0.624±0.026; n = 5; P<0.01).CONCLUSION: The increased enkephalin mRNA and prodynorphin mRNA gene expressions in rat hippocampus were involved in chronic stress.

  15. Downregulation of TIM-3 mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus

    Energy Technology Data Exchange (ETDEWEB)

    Cai, X.Z. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Department of Immunology, College of Basic Medical Sciences, China Medical University, Shenyang (China); Huang, W.Y.; Qiao, Y.; Chen, Y.; Du, S.Y.; Chen, D.; Yu, S. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Liu, N. [Department of Nephrology, First Affiliated Hospital, China Medical University, Shenyang (China); Dou, L.Y. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Jiang, Y. [Central Laboratory, First Affiliated Hospital, China Medical University, Shenyang (China); Department of Immunology, College of Basic Medical Sciences, China Medical University, Shenyang (China); Department of Dermatology, First Affiliated Hospital, China Medical University, Shenyang (China)

    2014-10-17

    The T-cell immunoglobulin and mucin domain (TIM) family is associated with autoimmune diseases, but its expression level in the immune cells of systemic lupus erythematosus (SLE) patients is not known. The aim of this study was to investigate whether the expression of TIM-3 mRNA is associated with pathogenesis of SLE. Quantitative real-time reverse transcription-polymerase chain reaction analysis (qRT-PCR) was used to determine TIM-1, TIM-3, and TIM-4 mRNA expression in peripheral blood mononuclear cells (PBMCs) from 132 patients with SLE and 62 healthy controls. The PBMC surface protein expression of TIMs in PBMCs from 20 SLE patients and 15 healthy controls was assayed by flow cytometry. Only TIM-3 mRNA expression decreased significantly in SLE patients compared with healthy controls (P<0.001). No significant differences in TIM family protein expression were observed in leukocytes from SLE patients and healthy controls (P>0.05). SLE patients with lupus nephritis (LN) had a significantly lower expression of TIM-3 mRNA than those without LN (P=0.001). There was no significant difference in the expression of TIM-3 mRNA within different classes of LN (P>0.05). Correlation of TIM-3 mRNA expression with serum IgA was highly significant (r=0.425, P=0.004), but was weakly correlated with total serum protein (r{sub s}=0.283, P=0.049) and serum albumin (r{sub s}=0.297, P=0.047). TIM-3 mRNA expression was weakly correlated with the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI; r{sub s}=-0.272, P=0.032). Our results suggest that below-normal expression of TIM-3 mRNA in PBMC may be involved in the pathogenesis of SLE.

  16. mRNA expression of dopamine receptors in peripheral blood lymphocytes of computer game addicts.

    Science.gov (United States)

    Vousooghi, Nasim; Zarei, Seyed Zeinolabedin; Sadat-Shirazi, Mitra-Sadat; Eghbali, Fatemeh; Zarrindast, Mohammad Reza

    2015-10-01

    Excessive playing of computer games like some other behaviors could lead to addiction. Addictive behaviors may induce their reinforcing effects through stimulation of the brain dopaminergic mesolimbic pathway. The status of dopamine receptors in the brain may be parallel to their homologous receptors in peripheral blood lymphocytes (PBLs). Here, we have investigated the mRNA expression of dopamine D3, D4 and D5 receptors in PBLs of computer game addicts (n = 20) in comparison to normal subjects (n = 20), using a real-time PCR method. The results showed that the expression level of D3 and D4 dopamine receptors in computer game addicts were not statistically different from the control group. However, the expression of the mRNA of D5 dopamine receptor was significantly down-regulated in PBLs of computer game addicts and reached 0.42 the amount of the control group. It is concluded that unlike with drug addiction, the expression levels of the D3 and D4 dopamine receptors in computer game addicts are not altered compared to the control group. However, reduced level of the D5 dopamine receptor in computer game addicts may serve as a peripheral marker in studies where the confounding effects of abused drugs are unwanted.

  17. Carboxylesterase 1 gene duplication and mRNA expression in adipose tissue are linked to obesity and metabolic function

    DEFF Research Database (Denmark)

    Friedrichsen, Martin; Poulsen, Pernille; Wojtaszewski, Jørgen;

    2013-01-01

    involved in the control of mRNA expression. Here, we investigated mRNA expression level in adipose tissue and its association with measures of adiposity and metabolic function in a population of elderly twins. Furthermore, the heritability of mRNA expression level in adipose tissue and the effect of gene......CONTEXT AND AIMS: Carboxylesterase 1 (CES1) appears to play an important role in the control of the metabolism of triglycerides and cholesterol in adipocytes and other cell types including hepatocytes. Therefore, it is relevant to gain insights into the genetic versus non-genetic mechanisms...

  18. Different structure and mRNA expression of Entamoeba invadens chitinases in the encystation and excystation.

    Science.gov (United States)

    Makioka, Asao; Kumagai, Masahiro; Hiranuka, Kazushi; Kobayashi, Seiki; Takeuchi, Tsutomu

    2011-08-01

    Entamoeba histolytica forms chitin-walled cysts during encystation process, where formation of the cyst wall needs not only chitin synthase but also chitinase. During excystation, quadruplet amoebae emerge from the chitin-walled cysts by dissolving the wall, so that chitinase may be necessary for excystation process as well. There is, however, no report on chitinase expression during excystation. In this study, we used Entamoeba invadens, a reptilian amoeba, as a model for encystation and excystation of E. histolytica, and studied chitinase mRNA expression in those processes. Although expression of three E. invadens chitinases designated EiChit1, EiChit2, and EiChit3 during encystation has been reported, we identified another enzyme named as EiChit4 in the E. invadens genome database. Therefore, we investigated the primary structure and mRNA expression of these four chitinases of Ei in the excystation as well as the encystation by real-time reverse transcription polymerase chain reaction (RT-PCR). Like EiChit1, EiChit4 had an 8 × Cys chitin-binding domain (CBD) and a hydrophilic spacer between the CBD and catalytic domain, and was also closer to EiChit1 than EiChit2 and EiChit3 in the phylogenetic tree. During encystation, the expression of all four chitinases increased in the early phase; the increase in EiChit1 and EiChit4 was much higher than in EiChit2 and EiChit3. Then, the expression of all four chitinases sharply decreased in the later phase. In cysts, EiChit1 was most abundantly expressed and EiChit4 was at a lower level, while the expressions of EiChit2 and EiChit3 were virtually absent. Following the induction of excystation, mRNA levels of EiChit1 and EiChit4 in cysts 5 h after induction were significantly lower than those in cysts before induction, while those of EiChit2 and EiChit3 were remarkably higher than before induction. The mRNAs of only EiChit2 and EiChit3 remarkably increased when the excystation was induced in the presence of cytochalasin D

  19. Contact call-driven zenk mRNA expression in the brain of the budgerigar (Melopsittacus undulatus).

    Science.gov (United States)

    Brauth, Steven E; Tang, Ye-Zhong; Liang, Wenru; Roberts, Todd F

    2003-09-10

    Contact call-driven zenk (zif268, egr1, NGF1A, Krox 24) mRNA expression was mapped with in situ hybridization histochemistry in a vocal learning parrot, the budgerigar (M. undulatus). Relative to controls, call stimulation induced high zenk mRNA expression in all auditory areas including those closely associated with the vocal system within the anterior forebrain (Brauth et al. (2001) J. Comp. Neurol. 432, 481; (2002) Learn. Memory 9, 76). Thus there is a high correspondence between the distributions of neurons exhibiting contact call-driven zenk protein and mRNA expression in budgerigars. Field L2a, an area reported previously to express only perinucleolar zenk protein localization (Brauth et al. (2002) Learn. Memory 9, 76) also showed zenk mRNA expression.

  20. CTGF is a central mediator of tissue remodeling and fibrosis and its inhibition can reverse the process of fibrosis.

    Science.gov (United States)

    Lipson, Kenneth E; Wong, Carol; Teng, Yuchin; Spong, Suzanne

    2012-01-01

    CTGF is a secreted matricellular protein with very complex biology. It has been shown to modulate many signaling pathways leading to cell adhesion and migration, angiogenesis, myofibroblast activation, and extracellular matrix deposition and remodeling, which together lead to tissue remodeling and fibrosis. It has been reported in the literature that inhibition of CTGF expression by siRNA prevents CCl4-induced liver fibrosis and can reverse fibrosis when administered after significant collagen deposition is observed. A monoclonal antibody to CTGF that is currently in clinical development (FG-3019) has demonstrated the ability to reverse vascular stiffening and improve cardiac function in a rat model of diabetic complications. FG-3019 has also exhibited activity in a murine radiation-induced pulmonary fibrosis model. When FG-3019 was administered to mice after a significant radiation-induced increase in lung density could be observed by CT imaging, the density of the lungs was observed to decrease over the period during which the antibody was administered and to remain stable after therapy had ceased. When considered together, these data indicate that inhibition of CTGF can prevent and reverse the process of fibrosis.

  1. LipL21 mRNA expression in lungs of hamsters infected with pathogenic Leptospira

    Institute of Scientific and Technical Information of China (English)

    Chintana Chirathaworn; Namo Suksomyos; Somchai Utivamek; Somboon Keelawat; Duangjai Suwancharoen; Duangporn Phulsuksombati; Yong Poovorawan

    2009-01-01

    Objective:Pulmonary haemorrhage is an increasing cause of death in leptospirosis patients.However,molecu-lar mechanism underlying pathologies in this organ is not clearly understood.It has been shown that sodium transport was disturbed following Leptospira infection.LipL21 is the second abundant outer membrane protein found only in pathogenic Leptospira.Its expression in vivo has been shown which suggests that this protein may be involved in survival in hosts or pathogenesis.However,the expression of this protein in host organs and its role in lung pathology has not been demonstrated.In this study we demonstrated the expression of LipL21 in lungs of hamsters infected with pathogenic Leptospira.Methods:Lung tissues were collected from Golden Syri-an hamsters injected with Leptospira interrogans serovar Pyrogenes at days 3,5 and 7 post-infection.Four ham-sters were used for each time point.Lungs from non-infected hamsters were collected as a control group.Li-pL21 mRNA expression in lung tissues was investigated by reverse transcription and nested PCR.Results:Li-pL21 mRNA expression was detected in all lung tissues from hamsters infected with pathogenic Leptospira.No PCR product was detected when tissues from non-infected hamsters were investigated.Conclusion:Our data demonstrated that LipL21 is expressed in lungs of hamsters infected with pathogenic Leptospira.Additional ex-periments such as quantitation and localization of LipL21 expression in lungs will provide further information whether this protein is involved in pathogenesis.

  2. Effects of glutamine supplementation on splenocyte cytokine mRNA expression in rats with septic peritonitis

    Institute of Scientific and Technical Information of China (English)

    Sung-Ling Yeh; Yu-Ni Lai; Huey-Fang Shang; Ming-Tsan Lin; Wan-Chun Chiu; Wei-Jao Chen

    2005-01-01

    AIM: To investigate the effects of glutamine (GLN)-enriched diets before and GLN-containing total parenteral nutrition (TPN) after sepsis or both on the secretion of cytokines and their mRNA expression levels in splenocytes of rats with septic peritonitis.METHODS: Rats were assigned to a control group and 4experimental groups. The control group and experimental groups 1 and 2 were fed a semipurified diet, while experimental groups 3 and 4 had part of the casein replaced by GLN which provided 25% of the total nitrogen.After rats were fed with these diets for 10 d, sepsis was induced by cecal ligation and puncture (CLP), whereas the control group underwent a sham operation, at the same time, an internal jugular vein was cannulated. All rats were maintained on TPN for 3 d. The control group and experimental groups 1 and 3 were infused with conventional TPN, while the TPN in experimental groups 2 and 4 was supplemented with GLN, providing 25% of the total nitrogen in the TPN solution. All rats were kiued 3 d after sham operation or CLP to examine their splenocyte subpopulation distribution and cytokine expression levels.RESULTS: Most cytokines could not be detected in plasma except for IL-10. No difference in plasma IL-10 was observed among the 5 groups. The IL-2, IL-4, IL-10, and TNF-α mRNA expression levels in splenocytes were significantly higher in experimental groups 2 and 4 than in the control group and group 1. The mRNA expression of IFN-γ was significantly higher in the GLN-supplemented groups than in the control group and experimental group 1. The proportion of CD45Ra+ was increased, while those of CD3+ and CD4+ were decreased in experimental group 1 after CLP was performed. There were no differences in spleen CD3+ lymphocyte distributions between the control and GLN-supplemented groups.CONCLUSION:GLN supplementation can maintain Tlymphocyte populations in the spleen and significantly enhance the mRNA expression levels of Th1 and Th2cytokines and TNF-α in

  3. Anti-CTGF single-chain variable fragment dimers inhibit human airway smooth muscle (ASM) cell proliferation by down-regulating p-Akt and p-mTOR levels.

    Science.gov (United States)

    Gao, Wei; Cai, Liting; Xu, Xudong; Fan, Juxiang; Xue, Xiulei; Yan, Xuejiao; Qu, Qinrong; Wang, Xihua; Zhang, Chen; Wu, Guoqiu

    2014-01-01

    Connective tissue growth factor (CTGF) contributes to airway smooth muscle (ASM) cell hyperplasia in asthma. Humanized single-chain variable fragment antibody (scFv) was well characterized as a CTGF antagonist in the differentiation of fibroblast into myofibroblast and pulmonary fibrosis in our previous studies. To further improve the bioactivity of scFv, we constructed a plasmid to express scFv-linker-matrilin-6×His fusion proteins that could self-assemble into the scFv dimers by disulfide bonds in matrilin under non-reducing conditions. An immunoreactivity assay demonstrated that the scFv dimer could highly bind to CTGF in a concentration-dependent manner. The MTT and EdU assay results revealed that CTGF (≥10 ng/mL) promoted the proliferation of ASM cells, and this effect was inhibited when the cells were treated with anti-CTGF scFv dimer. The western blot analysis results showed that increased phosphorylation of Akt and mTOR induced by CTGF could be suppressed by this scFv dimer. Based on these findings, anti-CTGF scFv dimer may be a potential agent for the prevention of airway remodeling in asthma.

  4. Exercise training does not increase muscle FNDC5 protein or mRNA expression in pigs.

    Science.gov (United States)

    Fain, John N; Company, Joseph M; Booth, Frank W; Laughlin, M Harold; Padilla, Jaume; Jenkins, Nathan T; Bahouth, Suleiman W; Sacks, Harold S

    2013-10-01

    Exercise training elevates circulating irisin and induces the expression of the FNDC5 gene in skeletal muscles of mice. Our objective was to determine whether exercise training also increases FNDC5 protein or mRNA expression in the skeletal muscles of pigs as well as plasma irisin. Castrated male pigs of the Rapacz familial hypercholesterolemic (FHM) strain and normal (Yucatan miniature) pigs were sacrificed after 16-20 weeks of exercise training. Samples of cardiac muscle, deltoid and triceps brachii muscle, subcutaneous and epicardial fat were obtained and FNDC5 mRNA, along with that of 6 other genes, was measured in all tissues of FHM pigs by reverse transcription polymerase chain reaction. FNDC protein in deltoid and triceps brachii was determined by Western blotting in both FHM and normal pigs. Citrate synthase activity was measured in the muscle samples of all pigs as an index of exercise training. Irisin was measured by an ELISA assay. There was no statistically significant effect of exercise training on FNDC5 gene expression in epicardial or subcutaneous fat, deltoid muscle, triceps brachii muscle or heart muscle. Exercise-training elevated circulating levels of irisin in the FHM pigs and citrate synthase activity in deltoid and triceps brachii muscle. A similar increase in citrate synthase activity was seen in muscle extracts of exercise-trained normal pigs but there was no alteration in circulating irisin. Exercise training in pigs does not increase FNDC5 mRNA or protein in the deltoid or triceps brachii of FHM or normal pigs while increasing circulating irisin only in the FHM pigs. These data indicate that the response to exercise training in normal pigs is not comparable to that seen in mice. Copyright © 2013 Elsevier Inc. All rights reserved.

  5. BCL6 mRNA Expression Level in Invasive Duct Carcinoma not otherwise Specified

    Science.gov (United States)

    Badr, Eman; Masoud, Eman; Eldien, Marwa Serag

    2016-01-01

    Introduction B-Cell Lymphoma 6 (BCL6) has an oncogenic role in tumourigenesis of various malignancies. It represses genes involved in terminal differentiation and plays complementary role with Signal Transducer and Activator of Transcription 3 (STAT3) in triple-negative breast cancer cellular function. Aim To evaluate the expression of BCL6 in cancer breast and determine its correlation with the clinico-pathological features including the molecular subtype of breast carcinoma. Materials and Methods This prospective case control study was carried out on 150 patients, divided into 100 cases of invasive duct carcinoma not otherwise specified and 50 benign breast lesions including fibroadenoma and fibrocystic disease. Fresh tissues were excised, which were then subjected to RNA extraction. The BCL6 mRNA level was assessed using real-time reverse transcription Polymerase Chain Reaction (PCR). Results There was a significant higher levels of BCL6 mRNA in malignant cases compared to benign ones (p<0.001). The level of BCL6 mRNA was higher in cases showing advanced tumor stage (p<0.04), triple negative subtype and associated in situ component (p<0.001) compared to cases with an early stage, luminal or Her 2-neu positive subtypes and those lacking in situ component. Conclusion BCL6 is up-regulated in breast cancer and is associated with poor prognostic features such as advanced stage and triple negative molecular subtype. BCL6 inhibitors might be considered as targeted therapy for breast cancer. PMID:28208987

  6. Changes of Survivin mRNA and Protein Expression during Paclitaxel Treatment in Breast Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    XIONG Huihua; YU Shiying; ZHUANG Liang; XIONG Hua

    2007-01-01

    In order to investigate the role of antiapoptosis gene, survivin in the resistance to palcitaxel, the expression of survivin mRNA and protein in the process of paclitaxel treatment in breast cancer cell line MCF-7 was detected. MCF-7 cells were incubated with paclitaxel at different concentrations. The growth inhibition rate of MCF-7 was investigated by tetrazolium bromide (MTT) colorimetry. The change of apoptosis was detected by Annexin-V/PI methods. The changes in the expression of survivin mRNA and protein were studied by reverse transcription polymerase chain reaction (RT-PCR) and Western-blot assay respectively. The growth inhibition rate of MCF-7 was increased in a concentration- and time-dependent manner. Paclitaxel of higher concentration could effectively induce apoptosis in MCF-7 cells after 48 h, while the expression of survivin was increased at early time (within 6 h) and decreased after 24 h regardless of treatment concentrations of paclitaxel. It suggested that tumor cells might evade the paclitaxel-induced cell cycle arrest and apoptosis by increasing the level of survivin at early treatment time.

  7. Biologically relevant effects of mRNA amplification on gene expression profiles

    Directory of Open Access Journals (Sweden)

    Smits Jos FM

    2006-04-01

    Full Text Available Abstract Background Gene expression microarray technology permits the analysis of global gene expression profiles. The amount of sample needed limits the use of small excision biopsies and/or needle biopsies from human or animal tissues. Linear amplification techniques have been developed to increase the amount of sample derived cDNA. These amplified samples can be hybridised on microarrays. However, little information is available whether microarrays based on amplified and unamplified material yield comparable results. In the present study we compared microarray data obtained from amplified mRNA derived from biopsies of rat cardiac left ventricle and non-amplified mRNA derived from the same organ. Biopsies were linearly amplified to acquire enough material for a microarray experiment. Both amplified and unamplified samples were hybridized to the Rat Expression Set 230 Array of Affymetrix. Results Analysis of the microarray data showed that unamplified material of two different left ventricles had 99.6% identical gene expression. Gene expression patterns of two biopsies obtained from the same parental organ were 96.3% identical. Similarly, gene expression pattern of two biopsies from dissimilar organs were 92.8% identical to each other. Twenty-one percent of reporters called present in parental left ventricular tissue disappeared after amplification in the biopsies. Those reporters were predominantly seen in the low intensity range. Sequence analysis showed that reporters that disappeared after amplification had a GC-content of 53.7+/-4.0%, while reporters called present in biopsy- and whole LV-samples had an average GC content of 47.8+/-5.5% (P Conclusion This study establishes that the gene expression profile obtained after amplification of mRNA of left ventricular biopsies is representative for the whole left ventricle of the rat heart. However, specific gene transcripts present in parental tissues were undetectable in the minute left

  8. Exercise Does Not Influence Myostatin and Follistatin mRNA expression in Young Women

    OpenAIRE

    Jensky, Nicole E.; Sims, Jennifer K.; Dieli-Conwright, Christina M.; Sattler, Fred R.; Rice, Judd C.; Schroeder, E. Todd

    2010-01-01

    We evaluated changes in myostatin, follistatin and MyoD mRNA gene expression using eccentric exercise (EE) and concentric exercise (CE) as probes to better understand the mechanisms of muscle hypertrophy in young women. Twelve women performed single-leg maximal eccentric (n=6, 25±1yr, 59±7kg) or concentric (n=6, 24±1 yr, 65±7kg) isokinetic knee extension exercise for 7 sessions. Muscle biopsies were taken from the vastus lateralis at baseline, 8hrs after the first exercise session, and 8hrs a...

  9. Alterations in Lipoxygenase and Cyclooxygenase-2 Catalytic Activity and mRNA Expression in Prostate Carcinoma

    Directory of Open Access Journals (Sweden)

    Scott B. Shappell

    2001-01-01

    Full Text Available Recent studies in prostate tissues and especially cell lines have suggested roles for arachidonic acid (AA metabolizing enzymes in prostate adenocarcinoma (Pca development or progression. The goal of this study was to more fully characterize lipoxygenase (LOX and cyclooxygenase-2 (COX-2 gene expression and AA metabolism in benign and malignant prostate using snap-frozen tissues obtained intraoperatively and mRNA analyses and enzyme assays. Formation of 15-hydroxyeicosatetraenoic acid (15-HETE was detected in 23/29 benign samples and 15-LOX-2 mRNA was detected in 21/25 benign samples. In pairs of pure benign and Pca from the same patients, 15-HETE production and 15-LOX-2 mRNA were reduced in Pca versus benign in 9/14 (P=.04 and 14/17 (P=.002, respectively. Under the same conditions, neither 5HETE nor 12-HETE formation was detectable in 29 benign and 24 tumor samples; with a more sensitive assay, traces were detected in some samples, but there was no clear association with tumor tissue. COX-2 mRNA was detected by nuclease protection assay in 7/16 benign samples and 5/16 tumors. In benign and tumor pairs from 10 patients, COX-2 was higher in tumor versus benign in only 2, with similar results by in situ hybridization. Paraffin immunoperoxidase for COX2 was performed in whole mount sections from 87 additional radical prostatectomy specimens, with strong expression in ejaculatory duct as a positive control and corroboration with in situ hybridization. No immunostaining was detected in benign prostate or tumor in 45% of cases. Greater immunostaining in tumor versus benign was present in only 17% of cases, and correlated with high tumor grade (Gleason score 8 and 9 vs. 5 to 7. In conclusion, reduced 15-LOX-2 expression and 15-HETE formation is the most characteristic alteration of AA metabolism in Pca. Increased 12-HETE and 5-HETE formation in Pca were not discernible. Increased COX-2 expression is not a typical abnormality in Pca in general, but

  10. Alterations in lipoxygenase and cyclooxygenase-2 catalytic activity and mRNA expression in prostate carcinoma.

    Science.gov (United States)

    Shappell, S B; Manning, S; Boeglin, W E; Guan, Y F; Roberts, R L; Davis, L; Olson, S J; Jack, G S; Coffey, C S; Wheeler, T M; Breyer, M D; Brash, A R

    2001-01-01

    Recent studies in prostate tissues and especially cell lines have suggested roles for arachidonic acid (AA) metabolizing enzymes in prostate adenocarcinoma (Pca) development or progression. The goal of this study was to more fully characterize lipoxygenase (LOX) and cyclooxygenase-2 (COX-2) gene expression and AA metabolism in benign and malignant prostate using snap-frozen tissues obtained intraoperatively and mRNA analyses and enzyme assays. Formation of 15-hydroxyeicosatetraenoic acid (15-HETE) was detected in 23/29 benign samples and 15-LOX-2 mRNA was detected in 21/25 benign samples. In pairs of pure benign and Pca from the same patients, 15-HETE production and 15-LOX-2 mRNA were reduced in Pca versus benign in 9/14 (P=.04) and 14/17 (P=.002), respectively. Under the same conditions, neither 5-HETE nor 12-HETE formation was detectable in 29 benign and 24 tumor samples; with a more sensitive assay, traces were detected in some samples, but there was no clear association with tumor tissue. COX-2 mRNA was detected by nuclease protection assay in 7/16 benign samples and 5/16 tumors. In benign and tumor pairs from 10 patients, COX-2 was higher in tumor versus benign in only 2, with similar results by in situ hybridization. Paraffin immunoperoxidase for COX-2 was performed in whole mount sections from 87 additional radical prostatectomy specimens, with strong expression in ejaculatory duct as a positive control and corroboration with in situ hybridization. No immunostaining was detected in benign prostate or tumor in 45% of cases. Greater immunostaining in tumor versus benign was present in only 17% of cases, and correlated with high tumor grade (Gleason score 8 and 9 vs. 5 to 7). In conclusion, reduced 15-LOX-2 expression and 15-HETE formation is the most characteristic alteration of AA metabolism in Pca. Increased 12-HETE and 5-HETE formation in Pca were not discernible. Increased COX-2 expression is not a typical abnormality in Pca in general, but occurs in high

  11. Tumor-associated antigens identified by mRNA expression profiling as tumor rejection epitopes

    DEFF Research Database (Denmark)

    Andersen, Marie Louise; Ruhwald, Morten; Thorn, Mette;

    2003-01-01

    immunization, but only two of these peptides (RAD23-31 and RAD24-31) were capable of generating a weak vaccination-induced protection against adoptive tumor growth. SM7 inoculated mice treated with a blocking antibody against the inhibitory T cell signal transducing molecule CTLA4 appeared to delay tumor take...... derived from potentially overexpressed tumor proteins, as identified by mRNA expression profiling of p53-/- thymoma cells, at best results in a weak tumor protection thus questioning this way of detection of new tumor rejection epitopes....

  12. Ustilago maydis natural antisense transcript expression alters mRNA stability and pathogenesis.

    Science.gov (United States)

    Donaldson, Michael E; Saville, Barry J

    2013-07-01

    Ustilago maydis infection of Zea mays leads to the production of thick-walled diploid teliospores that are the dispersal agent for this pathogen. Transcriptome analyses of this model biotrophic basidiomycete fungus identified natural antisense transcripts (NATs) complementary to 247 open reading frames. The U. maydis NAT cDNAs were fully sequenced and annotated. Strand-specific RT-PCR screens confirmed expression and identified NATs preferentially expressed in the teliospore. Targeted screens revealed four U. maydis NATs that are conserved in a related fungus. Expression of NATs in haploid cells, where they are not naturally occurring, resulted in increased steady-state levels of some complementary mRNAs. The expression of one NAT, as-um02151, in haploid cells resulted in a twofold increase in complementary mRNA levels, the formation of sense-antisense double-stranded RNAs, and unchanged Um02151 protein levels. This led to a model for NAT function in the maintenance and expression of stored teliospore mRNAs. In testing this model by deletion of the regulatory region, it was determined that alteration in NAT expression resulted in decreased pathogenesis in both cob and seedling infections. This annotation and functional analysis supports multiple roles for U. maydis NATs in controlling gene expression and influencing pathogenesis.

  13. BMP-15 m-RNA expression of mouse oocytes in vitro maturation in different droplet medium volume

    Institute of Scientific and Technical Information of China (English)

    Sri Rahayu; Nashi Widodo; Yumi Hoshino; Eimei Sato

    2015-01-01

    Objective:To investigate droplet medium volume effect on the BMP-15 mRNA expression. Methods:Oocytes are collected from mice ovaries by puncturing with a sterile 26-G needle. The droplet medium volumes are using 50 µL, 100 µL and 200 µL. The BMP-15 mRNA expression is determined in each group.Results:The results indicated that BMP-15 mRNA expression did not significantly differ when oocyte were cultured in 50 and 100 µL/droplet medium volume, but significant difference (P < 0.05) was found when oocytes were cultured in 200 µL/droplet medium volume.Conclusions:The highest BMP-15 m-RNA expression occur when oocytes are cultured in 200 µL/droplet medium volume.

  14. Time-course of 5-HT(6) receptor mRNA expression during memory consolidation and amnesia.

    Science.gov (United States)

    Huerta-Rivas, A; Pérez-García, G; González-Espinosa, C; Meneses, A

    2010-01-01

    Growing evidence indicates that antagonists of the 5-hydroxytryptamine (serotonin) receptor(6) (5-HT(6)) improve memory and reverse amnesia although the mechanisms involved are poorly understood. Hence, in this paper RT-PCR was used to evaluate changes in mRNA expression of 5-HT(6) receptor in trained and untrained rats treated with the 5-HT(6) receptor antagonist SB-399885 and amnesic drugs scopolamine or dizocilpine. Changes in mRNA expression of 5-HT(6) receptor were investigated at different times in prefrontal cortex, hippocampus and striatum. Data indicated that memory in the Pavlovian/instrumental autoshaping task was a progressive process associated to reduced mRNA expression of 5-HT(6) receptor in the three structures examined. SB-399885 improved long-term memory at 48h, while the muscarinic receptor antagonist scopolamine or the non-competitive NMDA receptor antagonist dizocilpine impaired it at 24h. Autoshaping training and treatment with SB-399885 increased 5-HT(6) receptor mRNA expression in (maximum increase) prefrontal cortex and striatum, 24 or 48h. The scopolamine-induced amnesia suppressed 5-HT(6) receptor mRNA expression while the dizocilpine-induced amnesia did not modify 5-HT(6) receptor mRNA expression. SB-399885 and scopolamine or dizocilpine were able to reestablish memory and 5-HT(6) receptor mRNA expression. These data confirmed previous memory evidence and of more interest is the observation that training, SB-399885 and amnesic drugs modulated 5-HT(6) receptor mRNA expression in prefrontal cortex, hippocampus and striatum. Further investigation in different memory tasks, times and amnesia models together with more complex control groups might provide further clues.

  15. Mycobacterium tuberculosis ESAT6 and CPF10 Induce Adenosine Deaminase 2 mRNA Expression in Monocyte-Derived Macrophages

    Science.gov (United States)

    Bae, Mi Jung; Ryu, Suyeon; Kim, Ha-Jeong; Cha, Seung Ick

    2017-01-01

    Background Delayed hypersensitivity plays a large role in the pathogenesis of tuberculous pleural effusion (TPE). Macrophages infected with live Mycobacterium tuberculosis (MTB) increase the levels of adenosine deaminase2 (ADA2) in the pleural fluid of TPE patients. However, it is as yet unclear whether ADA2 can be produced by macrophages when challenged with MTB antigens alone. This study therefore evaluated the levels of ADA2 mRNA expression, using monocyte-derived macrophages (MDMs) stimulated with MTB antigens. Methods Purified monocytes from the peripheral blood mononuclear cells of healthy volunteers were differentiated into macrophages using granulocyte-macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF). The MDMs were stimulated with early secretory antigenic target protein 6 (ESAT6) and culture filtrate protein 10 (CFP10). The mRNA expression levels for the cat eye syndrome chromosome region, candidate 1 (CECR1) gene encoding ADA2 were then measured. Results CECR1 mRNA expression levels were significantly higher in MDMs stimulated with ESAT6 and CFP10, than in the unstimulated MDMs. When stimulated with ESAT6, M-CSF-treated MDMs showed more pronounced CECR1 mRNA expression than GM-CSF-treated MDMs. Interferon-γ decreased the ESAT6- and CFP10-induced CECR1 mRNA expression in MDMs. CECR1 mRNA expression levels were positively correlated with mRNA expression of tumor necrosis factor α and interleukin 10, respectively. Conclusion ADA2 mRNA expression increased when MDMs were stimulated with MTB antigens alone. This partly indicates that pleural fluid ADA levels could increase in patients with culture-negative TPE. Our results may be helpful in improving the understanding of TPE pathogenesis.

  16. Early and late stimulation of ob mRNA expression in meal-fed and overfed rats.

    OpenAIRE

    Harris, R B; Ramsay, T G; Smith, S. R.; Bruch, R C

    1996-01-01

    ob protein is hypothesized to be a circulating feedback signal in the regulation of energy balance. Obese, overfed rats have high levels of ob mRNA expression and suppressed voluntary food intake, indicating the presence of a potent satiety factor. The objectives of this experiment were to determine whether feeding rats their normal daily intake in three meals, compared with ad libitum feeding, increased ob mRNA expression and to determine the degree of obesity required to stimulate expressio...

  17. Myometrial prostaglandin E2 synthetic enzyme mRNA expression: spatial and temporal variations with pregnancy and labour.

    Science.gov (United States)

    Sooranna, S R; Grigsby, P L; Engineer, N; Liang, Z; Sun, K; Myatt, L; Johnson, M R

    2006-10-01

    We have investigated the hypothesis that the expression of the enzymes involved in PGE(2) synthesis in the human uterus is co-ordinated. We have studied (i) the mRNA expression of the enzymes involved in PGE(2) synthesis [phospholipases (cPLA(2) and sPLA(2)), prostaglandin H synthase (PGHS)-2 and PG E synthases (PGES-1 and -2)] and their relationship to the expression of inflammatory cytokines in samples of myometrium obtained from pregnant women undergoing caesarean section (LSCS) either before or after the onset of labour at or before term; and (ii) the effect of IL-1beta, IL-6, TNF-alpha, PGE(2) and stretch on PGE(2) enzyme mRNA expression. We found that cPLA(2), sPLA(2) and PGHS-2 mRNA expression were greater in labour samples; cPLA(2), sPLA(2), PGHS-2, PGES-1 and -2 mRNA expression were greater in lower- than upper-segment samples; and there was no effect of gestational age. PGHS-2 mRNA levels correlated with those of PGES-1, cPLA(2), IL-1beta and IL-8; PGES-1 mRNA levels correlated with those of IL-1beta, IL-8 and cPLA(2). In primary cultures of uterine myocytes, cPLA(2) mRNA expression was increased by IL-1beta and IL-6; PGHS-2 mRNA expression was increased by IL-1beta, PGE(2) and stretch; and PGES-1 mRNA expression was increased by IL-1beta only. These data show that labour is associated with increased expression of the enzymes involved in PGE(2) synthesis and their expression is greater in the lower uterine segment. The presence of associations between the levels of PGE(2) enzyme mRNA expression and the effects of IL-1beta suggest that their expression is co-ordinated and that IL-1beta is the responsible factor.

  18. Expression of myosin heavy-chain mRNA in cultured myoblasts induced by centrifugal force.

    Science.gov (United States)

    Kurokawa, Katsuhide; Sakiyama, Koji; Abe, Shinichi; Hiroki, Emi; Naito, Kaoru; Nakajima, Kazunori; Takeda, Tomotaka; Inoue, Takashi; Ide, Yoshinobu; Ishigami, Keiichi

    2008-11-01

    Ballistic muscle training leads to hypertrophy of fast type fibers and training for endurance induces that of slow type fibers. Numerous studies have been conducted on electrical, extending and magnetic stimulation of cells, but the effect of centrifugal force on cells remains to be investigated. In this study, we investigated the effect of stimulating cultured myoblasts with centrifugal force at different speeds on cell proliferation and myosin heavy-chain (MyHC) mRNA expression in muscle fiber. Stimulation of myoblasts was carried out at 2 different speeds for 20 min using the Himac CT6D, a desk centrifuge, and cells were observed at 1, 3 and 5 days later. Number of cells 1 and 5 days after centrifugal stimulation was significantly larger in the 62.5 x g and 4,170 x g stimulation groups than in the control group. Expression of MyHC-2b mRNA 1 day after centrifugal stimulation was significantly higher in the 2 stimulation groups than in the control group. Almost no expression of MyHC-2a was observed in any group at 1 and 3 days after centrifugal stimulation. However, 5 days after stimulation, MyHC-2a was strongly expressed in the 2 stimulation groups in comparison to the control group. Three days after centrifugal stimulation, expression of MyHC-1 was significantly higher in the 2 stimulation groups than in the control group. The results of this study clarified the effect of different centrifugal stimulation speeds on muscle fiber characteristics, and suggest that centrifugal stimulation of myoblasts enhances cell proliferation.

  19. Distribution and mRNA Expression of BAMBI in Non-small-cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Shen MIAO

    2009-03-01

    Full Text Available Background and objective BAMBI structure is similar with that of the receptor Ⅰof TGF-β, it broadly participates in the control of TGF-β signaling. The aim of this study is to investigate the expression and its significance of BAMBI in non-small cell lung cancer (NSCLC and explore the relation between BAMBI and clinical and pathological factors of NSCLC. Methods Sixty-three cases with NSCLC and adjacent normal tissue specimens were used for immunohistochemical assay. Thirty-one fresh lung cancer tissue specimens and surrounding normal lung tissue specimens was preserved for RT-PCR in -70 ℃ after quick-frozen in liquid nitrogen immediately. Results The level of BAMBI mRNA in cancer tissues was higher than that in the corresponding adjacent tissues (0.358±0.135 vs 0.249±0.129, with the difference being statistically significant (P =0.003. BAMBI protein expressed mainly in the membrane and the cytoplasm close to the membrane, its expression in the cancer tissue was higher than that in the adjacent tissues, the difference was significant (P <0.01. Expression of BAMBI in the cancer tissue was higher than that in the adjacent tissues, and the expression of BAMBI in adenocarcinoma of lung is higher than that in squamous carcinoma. Conclusion The expressions of BAMBI significantly increase in NSCLC. It might be a common affair in carcinogenesis of NSCLC.

  20. The Expression of Antibiotic Resistance Methyltransferase Correlates with mRNA Stability Independently of Ribosome Stalling.

    Science.gov (United States)

    Dzyubak, Ekaterina; Yap, M N

    2016-12-01

    Members of the Erm methyltransferase family modify 23S rRNA of the bacterial ribosome and render cross-resistance to macrolides and multiple distantly related antibiotics. Previous studies have shown that the expression of erm is activated when a macrolide-bound ribosome stalls the translation of the leader peptide preceding the cotranscribed erm Ribosome stalling is thought to destabilize the inhibitory stem-loop mRNA structure and exposes the erm Shine-Dalgarno (SD) sequence for translational initiation. Paradoxically, mutations that abolish ribosome stalling are routinely found in hyper-resistant clinical isolates; however, the significance of the stalling-dead leader sequence is largely unknown. Here, we show that nonsense mutations in the Staphylococcus aureus ErmB leader peptide (ErmBL) lead to high basal and induced expression of downstream ErmB in the absence or presence of macrolide concomitantly with elevated ribosome methylation and resistance. The overexpression of ErmB is associated with the reduced turnover of the ermBL-ermB transcript, and the macrolide appears to mitigate mRNA cleavage at a site immediately downstream of the ermBL SD sequence. The stabilizing effect of antibiotics on mRNA is not limited to ermBL-ermB; cationic antibiotics representing a ribosome-stalling inducer and a noninducer increase the half-life of specific transcripts. These data unveil a new layer of ermB regulation and imply that ErmBL translation or ribosome stalling serves as a "tuner" to suppress aberrant production of ErmB because methylated ribosome may impose a fitness cost on the bacterium as a result of misregulated translation.

  1. Bcl-2/Bax protein and mRNA expression in yak (Bos grunniens) placentomes.

    Science.gov (United States)

    Fan, JiangFeng; Yu, SiJiu; Cui, Yan; Xu, Gengquan; Wang, Libin; Pan, Yangyang; He, Honghong

    2017-07-29

    Placental function is complex and influenced by various factors; furthermore, it depends on a delicate balance between cell proliferation, cell differentiation, and cell death. Bcl-2 and Bax proteins are key apoptosis regulators and are considered to play an important role in the maintenance of both dynamic balance and integrity of many tissues. Changes in Bcl-2 and Bax expressions have been described during different developmental stages in normal human placentas. Studies furthermore indicated several pathological placental changes to be related to abnormal Bcl-2 and Bax expressions. In the present study, we investigated both expression and distribution of Bcl-2 and Bax in yak placentas. For this, we collected placentas of 35 yaks at different stages of pregnancy as well as cotyledonary villi of four postpartum yaks. Protein and mRNA expressions of both Bcl-2 and Bax were investigated via immunohistochemistry, Western blot, and real-time PCR. Immunoreactive Bcl-2 protein was mainly localized near the fetal villous trophoblast at various gestational stages and post-partum. The Positive Index (PI) of Bcl-2 protein expression significantly decreased with increasing gestational age. Early during pregnancy (≤2 months), the Bax protein was widely distributed in the fetal villous trophoblast layer, the maternal caruncular crypt epithelium, and the stroma. Subsequently, the Bax protein distribution gradually concentrated in the fetal villous trophoblast layer. The staining intensity of Bax increased from the 3rd month to the prepartum of gestation. The PI reached a minimum of 9.4 ± 2.2 in fetal chorionic villi (FCV) and 1.3 ± 0.8 in maternal caruncular crypts (MCC) of the three months group. Both Bcl-2 and Bax had maximum immunoreactivity in the fetal villous trophoblast layer of placentas collected form postpartum yaks (with PIs of 36.6 ± 5.7 and 38.2 ± 4.8, respectively). Protein and mRNA expression of Bcl-2 and Bax investigated via Western blot and real

  2. Bacteria and Toll-like receptor and cytokine mRNA expression profiles associated with canine arthritis.

    Science.gov (United States)

    Riggio, Marcello P; Lappin, David F; Bennett, David

    2014-08-15

    The major forms of inflammatory canine arthritis are immune-mediated arthritis (IMA) and septic arthritis (SA), although some cases of cruciate disease (CD) are associated with significant levels of synovitis. In this study, the bacteria associated with canine arthritis were identified and mRNA expression levels of Toll-like receptors (TLRs) and pro-inflammatory cytokines determined. Of the 40 synovial fluid samples analysed, bacteria were isolated from 12 samples by culture (2 CD, 10 SA) and detected in 4 samples (3 CD, 1 SA) using culture-independent methods. Statistically significant increases in TLR2, tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and IL-12 mRNA expression were seen in all disease groups compared to normal controls. All disease groups had decreased mRNA expression of other TLRs compared to normal controls, but this did not reach statistical significance. Synovial fluid cell counts revealed that the highest number and proportion of mononuclear cells and neutrophils were found in the IMA and SA samples, respectively. Age had an effect on the TLR and cytokine mRNA expression profiles: TNF-α (p=0.043) and IL-12 (p=0.025) mRNA expression was increased and TLR4 mRNA expression was reduced (p=0.033) in dogs up to 4 years of age compared to older animals. In the 10 SA samples from which bacteria were isolated, statistically significant increases in TLR2, TLR7, TNF-α and IL-6 mRNA expression were observed. It is concluded that canine arthritis is associated with increased mRNA levels of pro-inflammatory cytokines, which could in some cases be mediated by bacteria through activation of TLR2.

  3. A comprehensive examination of Smad4, Smad6 and Smad7 mRNA expression in pancreatic ductal adenocarcinoma

    Directory of Open Access Journals (Sweden)

    P Singh

    2011-01-01

    Full Text Available Background: Smad4, Smad6 and Smad7 are important molecules in TGF-beta pathway, which plays an important role in pancreatic ductal adenocarcinoma (PDAC biology. Aims : This study examined the expression profiles of Smad4, Smad6 and Smad7 mRNA in patient samples of PDAC and their relationship to Smad protein expression, SMAD4 gene mutations, clinicopathological parameters and patient survival. Settings and Design: Surgically resected, paired normal and tumor tissues of 25 patients of PDAC were studied. Materials and Methods: Protein and mRNA levels were assessed by immunohistochemistry and RT-PCR, respectively. Statistical Methods: Statistical analysis was done using Student′s t-test, Pearson′s chi-square test, Spearman′s Rank Correlation, Pearson′s Correlation test and Kaplan-Meier Logrank test. Results: While there was a highly significant difference in the protein levels of all three Smads in tumor as compared to normal samples, mRNA levels were significantly different only for Smad4. Protein levels did not correlate significantly with mRNA levels for any of the three Smads. The mRNA levels of Smad4 and Smad6, Smad4 and Smad7, and Smad6 and Smad7 in tumor samples showed a significant positive correlation. The relationship of Smad4 mRNA expression to SMAD4 gene status and Smad4 protein expression was discordant and there was no significant correlation between mRNA expression and clinicopathological parameters and patient survival. Conclusion : The absence of concordance between SMAD4 gene status, mRNA expression and Smad4 protein expression suggests the presence of other regulatory mechanisms in Smad4 transcription and translation in PDAC.

  4. Molecular cloning and mRNA expression of duck invariant chain.

    Science.gov (United States)

    Zhong, Dalian; Yu, Weiyi; Bao, Min; Xu, Zhiben; Li, Lin; Liu, Jing

    2006-04-15

    In the present study we identified a duck invariant chain (Ii) cDNA, named duck Ii-1, by RT-PCR and RACE. It was 1190 bp in length and contained a 669 bp open reading frame. An alternative transcript encoding a thyroglobulin (Tg)-containing form of Ii, named duck Ii-2, was also found in duck. The putative amino acid sequence of duck Ii-1 showed an 82% similarity to chicken Ii-1 and about 60% similarity to its mammalian homologues. The similarity of the Tg domain between duck and chicken Ii-2 was 96%, and about 70% between duck and mammalian Ii. The result of RT-PCR showed that Ii mRNA was extensively expressed in various tissues. High levels of both Ii-1 and Ii-2 mRNA were observed in the spleen and bursa of Fabricius. The predicted three-dimensional (3D) structures of duck Ii trimerization and Tg domain are similar to the corresponding regions of human Ii analyzed by comparative protein modeling. These findings indicate that the two isoforms of duck Ii, which strongly expressed in the major immune organs, share structural identity with human Ii.

  5. RAT GDNF GENE TRANSFECTION AND EXPRESSION OF ITS mRNA AND PROTEIN IN SCHWANN CELLS

    Institute of Scientific and Technical Information of China (English)

    平萍; 范志宏; 李青峰; 张涤生

    2004-01-01

    Objective To investigate the possibility of the transfection of glial-cell line derived neurotrophic factor (GDNF) gene into Schwann cells(SCs). Methods SCs cultures from sciatic nerves of neonatal rats were established. A recombinant retrovirus vector containing GDNF gene was constructed and transferred into SCs.Expression levels of GDNF mRNA and protein were respectively identified with reverse transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemistry. Determination of GDNF synthesis rates from Retro. pLNCX2-GDNF-transduced SCs (GDNF-SCs) in vitro by enzyme-linked immunoassay sensitive assay ( ELISA ). Biololgical activity of conditioned medium from GENF-SCs was analysed by co-culture with rat motoneurons. Results Transfection of GDNF gene into SCs lead to significantly enhanced expression of GDNF mRNA and protein. The rate of GDNF secreted by GDNF-SCs was also enhanced(5. 1-fold), and more motoneurons survived co-cultured with conditioned medium of GNDF-SCs than with that of normal SCs. Conclusion GNDF gene transfection may be a better way to graft SCs promoting regeneration and repairing demyelination in PNS and CNS.

  6. Streamlining gene expression analysis: integration of co-culture and mRNA purification.

    Science.gov (United States)

    Berry, Scott M; Singh, Chandresh; Lang, Jessica D; Strotman, Lindsay N; Alarid, Elaine T; Beebe, David J

    2014-02-01

    Co-culture of multiple cell types within a single device enables the study of paracrine signaling events. However, extracting gene expression endpoints from co-culture experiments is laborious, due in part to pre-PCR processing of the sample (i.e., post-culture cell sorting and nucleic acid purification). Also, a significant loss of nucleic acid may occur during these steps, especially with microfluidic cell culture where lysate volumes are small and difficult to access. Here, we describe an integrated platform for performing microfluidic cell culture and extraction of mRNA for gene expression analysis. This platform was able to recover 30-fold more mRNA than a similar, non-integrated system. Additionally, using a breast cancer/bone marrow stroma co-culture, we recapitulated stromal-dependent, estrogen-independent growth of the breast cancer cells, coincident with transcriptional changes. We anticipate that this platform will be used for streamlined analysis of paracrine signaling events as well as for screening potential drugs and/or patient samples.

  7. Effect of GFR on plasma N-terminal connective tissue growth factor (CTGF) concentrations.

    NARCIS (Netherlands)

    Gerritsen, K.G.; Abrahams, A.C.; Peters, H.P.E.; Nguyen, T.Q.; Koeners, M.P.; Hoedt, C.H. den; Dendooven, A.; Dorpel, M.A. van den; Blankestijn, P.J.; Wetzels, J.F.M.; Joles, J.A.; Goldschmeding, R.; Kok, R.J.

    2012-01-01

    BACKGROUND: Connective tissue growth factor (CTGF) has a key role in the pathogenesis of renal and cardiac fibrosis. Its amino-terminal fragment (N-CTGF), the predominant form of CTGF detected in plasma, has a molecular weight in the middle molecular range (18 kDa). However, it is unknown whether N-

  8. FOXA2 mRNA expression is associated with relapse in patients with Triple-Negative/Basal-like breast carcinoma.

    Science.gov (United States)

    Perez-Balaguer, Ariadna; Ortiz-Martínez, Fernando; García-Martínez, Araceli; Pomares-Navarro, Critina; Lerma, Enrique; Peiró, Gloria

    2015-09-01

    The FOXA family of transcription factors regulates chromatin structure and gene expression especially during embryonic development. In normal breast tissue FOXA1 acts throughout mammary development; whereas in breast carcinoma its expression promotes luminal phenotype and correlates with good prognosis. However, the role of FOXA2 has not been previously studied in breast cancer. Our purpose was to analyze the expression of FOXA2 in breast cancer cells, to explore its role in breast cancer stem cells, and to correlate its mRNA expression with clinicopathological features and outcome in a series of patients diagnosed with breast carcinoma. We analyzed FOXA2 mRNA expression in a retrospective cohort of 230 breast cancer patients and in cell lines. We also knocked down FOXA2 mRNA expression by siRNA to determine the impact on cell proliferation and mammospheres formation using a cancer stem cells culture assay. In vitro studies demonstrated higher FOXA2 mRNA expression in Triple-Negative/Basal-like cells. Further, when it was knocked down, cells decreased proliferation and its capability of forming mammospheres. Similarly, FOXA2 mRNA expression was detected in 10% (23/230) of the tumors, especially in Triple-Negative/Basal-like phenotype (p Triple-Negative/Basal-like tumors, and is associated with increase relapses.

  9. Correlation of Baseline BCL-2 mRNA Expression and Clinical Response to Neoadjuvant Chemotherapy in Breast Cancer

    Directory of Open Access Journals (Sweden)

    Prihantono Prihantono

    2017-01-01

    Full Text Available Impairment of apoptosis is a hallmark of cancer. Tumor resistance to apoptosis usually caused by deregulation of the expression of BCL-2 family protein or mutation of the tumor suppressor gene p53. Over expression of Bcl-2 is commonly found in various types of cancer including breast cancer. Studies mentioned that analysis of Bcl-2 might predict response to selected endocrine and chemotherapies. This study is conducted to evaluate the correlation of BCL-2 mRNA expression and clinical response to neoadjuvant chemotherapy in breast cancer patients. Longitudinal study is used in this research, 30 subjects of breast cancer tissue samples prechemotherapy using cyclophosphamide-adriamycin-5FU regiment. Detection of mRNA expression of BCL-2 using qRT-PCR techniques. Evaluation of clinical response to chemotherapy is using RECIST criteria. Mean value of BCL-2 mRNA expression in breast cancer patients was 9.917± 2.568. Mean value of BCL-2 mRNA expression of responsive group was 9.887± 2.731. Mean value of BCL-2 mRNA expression of nonresponsive group was 10.017±2.122. Mean value of responsive group were lower than nonresponsive group, but there was no significant correlation between baseline mRNA expression of BCL-2 with clinical response to chemotherapy, value of r=0.378, p=0.223 (p>0.05. this study shows that there was no significant correlation between baseline expression of mRNA BCL-2 with clinical response to chemotherapy.

  10. MicroRNA-143-3p inhibits hyperplastic scar formation by targeting connective tissue growth factor CTGF/CCN2 via the Akt/mTOR pathway.

    Science.gov (United States)

    Mu, Shengzhi; Kang, Bei; Zeng, Weihui; Sun, Yaowen; Yang, Fan

    2016-05-01

    Post-traumatic hypertrophic scar (HS) is a fibrotic disease with excessive extracellular matrix (ECM) production, which is a response to tissue injury by fibroblasts. Although emerging evidence has indicated that miRNA contributes to hypertrophic scarring, the role of miRNA in HS formation remains unclear. In this study, we found that miR-143-3p was markedly downregulated in HS tissues and fibroblasts (HSFs) using qRT-PCR. The expression of connective tissue growth factor (CTGF/CCN2) was upregulated both in HS tissues and HSFs, which is proposed to play a key role in ECM deposition in HS. The protein expression of collagen I (Col I), collagen III (Col III), and α-smooth muscle actin (α-SMA) was obviously inhibited after treatment with miR-143-3p in HSFs. The CCK-8 assay showed that miR-143-3p transfection reduced the proliferation ability of HSFs, and flow cytometry showed that either early or late apoptosis of HSFs was upregulated by miR-143-3p. In addition, the activity of caspase 3 and caspase 9 was increased after miR-143-3p transfection. On the contrary, the miR-143-3p inhibitor was demonstrated to increase cell proliferation and inhibit apoptosis of HSFs. Moreover, miR-143-3p targeted the 3'-UTR of CTGF and caused a significant decrease of CTGF. Western blot demonstrated that Akt/mTOR phosphorylation and the expression of CTGF, Col I, Col III, and α-SMA were inhibited by miR-143-3p, but increased by CTGF overexpression. In conclusion, we found that miR-143-3p inhibits hypertrophic scarring by regulating the proliferation and apoptosis of human HSFs, inhibiting ECM production-associated protein expression by targeting CTGF, and restraining the Akt/mTOR pathway.

  11. Expression of CC Chemokine Ligand 20 and CC Chemokine Receptor 6 mRNA in Patients with Psoriasis Vulgaris

    Institute of Scientific and Technical Information of China (English)

    吴艳; 李家文

    2004-01-01

    Summary: In order to explore the possible role of CC chemokine ligand 20 (CCL20) and its receptor CC chemokine receptor 6 (CCR6) in the pathogenesis of psoriasis, the expression levels of mRNA of them in psoriatic lesions were investigated. The skin biopsies were collected from skin lesions in 35 cases of psoriasis vulgaris and 18 normal controls. RT-PCR was used to semi-quantitatively analyze the mRNA expression of CCL20 and CCR6 in the psoriatic lesions and the normal skin tissues.The results showed that the mRNA of CCL20 and CCR6 was present in every specimen. The expression levels of CCL20 mRNA in skin lesions were 1. 1397±0. 0521, which were greatly higher than those in normal controls (0.8681±0.0308) (P<0. 001). The expression levels of CCR6 mRNA in skin lesions were 1.1103±0.0538, significantly higher than in the controls (0.9131±0.0433, P<0. 001). These findings indicate that up-regulated expression of CCL20 and CCR6 mRNA might be related to the pathogenesis of psoriasis.

  12. Expression of TRAF6 and ubiquitin mRNA in skeletal muscle of gastric cancer patients

    Directory of Open Access Journals (Sweden)

    Sun Yuan-Shui

    2012-09-01

    Full Text Available Abstract Objective To investigate the prognostic significance of tumor necrosis factor receptor (TNFR,-associated factor 6 (TRAF6,-and ubiquitin in gastric cancer patients. Methods Biopsies of the rectus abdominis muscle were obtained intra operatively from 102 gastric cancer patients and 29 subjects undergoing surgery for benign abdominal diseases, and muscle TRAF6 and ubiquitin mRNA expression and proteasome proteolytic activities were assessed. Results TRAF6 was significantly upregulated in muscle of gastric cancer compared with the control muscles. TRAF6 was upregulated in 67.65% (69/102 muscle of gastric cancer. Over expression of TRAF6 in muscles of gastric cancer were associated with TNM stage, level of serum albumin and percent of weight loss. Ubiquitin was significantly upregulated in muscle of gastric cancer compared with the control muscles. Ubiquitin was upregulated in 58.82% (60/102 muscles of gastric cancer. Over expression of ubiquitin in muscles of gastric cancer were associated with TNM (Tumor-Node-Metastasis stage and weight loss. There was significant relation between TRAF6 and ubiquitin expression. Conclusions We found a positive correlation between TRAF6 and ubiquitin expression, suggesting that TRAF6 may up regulates ubiquitin activity in cancer cachexia. While more investigations are required to understand its mechanisms of TRAF6 and ubiquitin in skeletal muscle. Correct the catabolic-anabolic imbalance is essential for the effective treatment of cancer cachexia.

  13. Expression of CD52 mRNA in the rat embryo.

    Science.gov (United States)

    Ermini, Leonardo; Aldi, Silvia; Rosati, Floriana

    2008-09-01

    CD52 is a leukocyte differentiation antigen first discovered in humans as expressed on the surface of lymphocytes, monocytes and eosinophils. The human CD52 is found on chromosome 1, and two alleles are both known to be reasonably common. A closely homologous gene has been identified in the cynomologous monkey and related genes have been found in mouse, rat and dog. The role of CD52 in lymphocyte is still unclear but the anti-CD52 antibodies named CAMPATH-1 antibodies are largely used for therapy where depletion of lymphocytes is required. In the past expression of the antigen on progenitors of leukocytes in bone marrow had been excluded, but recent work indicates CD52 is highly expressed on cells with colony-forming and NOD/SCID (non-obese diabetic-severe combined immunodeficiency)-engrafting capacities, both at the mRNA and membrane protein level. We have investigated CD52 expression during development in rat embryos by in situ hybridization. We report here that the antigen is highly expressed in the liver that is the major organ where multipotent hematopietic stem cells differentiate but also in the splancnopleuric mesoderm, at early stages of embryo differentiation, where hematopietic stem cells are suggested to arise. CD52+ cells were found in areas active in vasculogenesis at early embryo stages and in the walls of the vessels in the liver at mid gestation. CD52+ cells were also found to emerge among c-Kit positive cells.

  14. Imipramine induces brain-derived neurotrophic factor mRNA expression in cultured astrocytes.

    Science.gov (United States)

    Takano, Katsura; Yamasaki, Hiroshi; Kawabe, Kenji; Moriyama, Mitsuaki; Nakamura, Yoichi

    2012-01-01

    Depression is one of the most prevalent and livelihood-threatening forms of mental illnesses and the neural circuitry underlying depression remains incompletely understood. Recent studies suggest that the neuronal plasticity involved with brain-derived neurotrophic factor (BDNF) plays an important role in the recovery from depression. Some antidepressants are reported to induce BDNF expression in vivo; however, the mechanisms have been considered solely in neurons and not fully elucidated. In the present study, we evaluated the effects of imipramine, a classic tricyclic antidepressant drug, on BDNF expression in cultured rat brain astrocytes. Imipramine dose-dependently increased BDNF mRNA expression in astrocytes. The imipramine-induced BDNF increase was suppressed with inhibitors for protein kinase A (PKA) or MEK/ERK. Moreover, imipramine exposure activated transcription factor cAMP response element binding protein (CREB) in a dose-dependent manner. These results suggested that imipramine induced BDNF expression through CREB activation via PKA and/or ERK pathways. Imipramine treatment in depression might exert antidepressant action through BDNF production from astrocytes, and glial BDNF expression might be a target of developing novel antidepressants.

  15. Oxidative stress induced Interleukin-32 mRNA expression in human bronchial epithelial cells

    Directory of Open Access Journals (Sweden)

    Kudo Megumi

    2012-03-01

    Full Text Available Abstract Background Chronic obstructive pulmonary disease (COPD is characterized by airflow obstruction and persistent inflammation in the airways and lung parenchyma. Oxidative stress contributes to the pathogenesis of COPD. Interleukin (IL-32 expression has been reported to increase in the lung tissue of patients with COPD. Here, we show that IFNγ upregulated IL-32 expression and that oxidative stress augmented IFNγ-induced-IL-32 expression in airway epithelial cells. We further investigated transcriptional regulation responsible for IFNγ induced IL-32 expression in human airway epithelial cells. Methods Human bronchial epithelial (HBE cells were stimulated with H2O2 and IFNγ, and IL-32 expression was evaluated. The cell viability was confirmed by MTT assay. The intracellular signaling pathways regulating IL-32 expression were investigated by examining the regulatory effects of MAPK inhibitors and JAK inhibitor after treatment with H2O2 and IFNγ, and by using a ChIP assay to identify transcription factors (i.e. c-Jun, CREB binding to the IL-32 promoter. Promoter activity assays were conducted after mutations were introduced into binding sites of c-Jun and CREB in the IL-32 promoter. IL-32 expression was also examined in HBE cells in which the expression of either c-Jun or CREB was knocked out by siRNA of indicated transcription factors. Results There were no significant differences of cell viability among groups. After stimulation with H2O2 or IFNγ for 48 hours, IL-32 expression in HBE cells was increased by IFNγ and synergistically upregulated by the addition of H2O2. The H2O2 augmented IFNγ induced IL-32 mRNA expression was suppressed by a JNK inhibitor, but not by MEK inhibitor, p38 inhibitor, and JAK inhibitor I. Significant binding of c-Jun and CREB to the IL-32 promoter was observed in the IFNγ + H2O2 stimulated HBE cells. Introducing mutations into the c-Jun/CREB binding sites in the IL-32 promoter prominently suppressed its

  16. 苦参碱对房颤犬心房肌胶原合成及TNF-α、TGF-β1和CTGF的影响%Effects of matrine on collagen proliferation and TNF-α, TGF-β1 and CTGF in atrial tissues of dogs with ;persistent atrial fibrillation

    Institute of Scientific and Technical Information of China (English)

    戴友平; 李七一; 张曙光; 吴佳佳; 王惠敏; 朱萱萱

    2013-01-01

    Objective To study the effects of matrine (mat) on collagen synthesis and expression of tumor necrosis factor alpha (TNF-α), transforming growth factor-β1 (TGF-β1) and connective tissue growth factor (CTGF) in atrial tissues of dogs with persistent atrial fibrillation (AF). Methods Ten healthy beagle dogs were randomly divided into two groups: AF group (n=5) and AF/Mat group (n=5), using right ventricular pacing to establish AF model. The collagen volume fraction (CVF) in atrial tissue were detected by sirius red staining to determine the level of fabrication. The level of TNF-α, TGF-β1 and CTGF were detected by immunohisto-chemistry. The mRNA expression level of TNF-α, TGF-β1 and CTGF were detected by reverse transcription-polymerase chain reaction (RT-PCR). Results Compared with the AF group, the fibration level of AF/Mat was decreased obviously (P<0.05), the expression levels of TNF-α, TGF-β1 and CTGF were decreased, and the mRNA expression level were decreased significantly in atrial tissues (P<0.05 and P<0.01). Conclusion Matrine may inhibit fibrosis in atrial tissues through inhibition collagen proliferation and expression of TNF-α, TGF-β1 and CTGF.%目的:探讨苦参碱对犬心房颤动(房颤)心房肌组织中胶原合成以及肿瘤坏死因子(TNF-α)、转化生长因子(TGF-β1)和结缔组织生长因子(CTGF)表达的影响。方法健康比格犬10只采用快速右心室起搏造房颤模型,随机分成房颤组和房颤+苦参碱组各5只。采用天狼星红染色,计算胶原容积分数(CVF)以测定纤维化程度;采用免疫组织化学法检测右心房TNF-α、TGF-β1和CTGF的蛋白表达情况;用逆转录-聚合酶链反应(RT-PCR)技术检测TNF-α、TGF-β1和CTGF的mRNA水平表达情况。结果与房颤组相比,房颤+苦参碱组纤维化程度降低,CVF 明显下降(P<0.05),TNF-α、TGF-β1和 CTGF 蛋白表达水平下降,且TNF-α和TGF-β1的mRNA

  17. Expression of survivin mRNA in peritoneal lavage fluid from patients with gastric carcinoma

    Institute of Scientific and Technical Information of China (English)

    王振宁; 徐惠绵; 姜莉; 周欣; 鲁翀; 张学

    2004-01-01

    Background Peritoneal dissemination is the most common pattern of metastasis in advanced gastric carcinoma with serosal invasion. In the present study, we reported the clinical relevance of a new diagnostic method involving RT-PCR, using survivin as the target gene, for the detection of free cancer cells in peritoneal washes.Methods Intraoperative peritoneal washes were obtained from 48 patients who underwent surgery for gastric cancer. RT-PCR analysis with primers specific for survivin and conventional cytological examinations were both performed.Results Survivin mRNA was not detected in any peritoneal wash samples from patients with benign disease, but was detected in 28 of 48 samples taken from patients with gastric cancer and in all metastastic nodules. Survivin expression in the peritoneal cavity significantly correlated with depth of cancer invasion, lymph node metastasis, and TNM stage. There were 92% of clinically evident peritoneal metastasis cases showed detectable survivin expression. The combination of survivin RT-PCR and cytological examination yielded positive results in 66.7% (32/48) of patients with gastric cancer, much higher than the results produced by cytological method alone. Conclusions Survivin mRNA detected in peritoneal lavage fluid might indicate the presence of free cancer cells in the peritoneal cavity. The high sensitivity of the RT-PCR-based survivin assay suggests that survivin serves as a molecular marker for detecting peritoneal micrometastasis. Its ubiquitous expression in peritoneal cancer cells and metastatic nodules also suggests a promising future therapeutic strategy based on survivin inhibition for cases of gastric cancer involving peritoneal metastasis.

  18. Gene expression in distinct regions of rat tendons in response to jump training combined with anabolic androgenic steroid administration

    DEFF Research Database (Denmark)

    Marqueti, Rita Cássia; Marqueti, Rita de Cássia; Heinemeier, Katja Maria;

    2012-01-01

    RNA levels of COL1A1, COL3A1, TIMP-1, TIMP-2, MMP-2, IGF-IEa, GAPDH, CTGF and TGF-ß-1 were evaluated by quantitative PCR. Our main results indicated that mRNA levels alter in different regions in each tendon of sedentary animals. The training did not alter the expression of COL1A1, COL3A, IGF-IEa and MMP-2...

  19. Oncogenic kinase NPM/ALK induces expression of HIF1α mRNA.

    Science.gov (United States)

    Marzec, M; Liu, X; Wong, W; Yang, Y; Pasha, T; Kantekure, K; Zhang, P; Woetmann, A; Cheng, M; Odum, N; Wasik, M A

    2011-03-17

    The mechanisms of malignant cell transformation mediated by the oncogenic anaplastic lymphoma kinase (ALK) tyrosine kinase remain only partially understood. In this study, we report that T-cell lymphoma (TCL) cells carrying the nucleophosmin (NPM)/ALK fusion protein (ALK+ TCL) strongly express hypoxia-induced factor 1α (HIF1α) mRNA, even under normoxic conditions, and markedly upregulate HIF1α protein expression under hypoxia. HIF1α expression is strictly dependent on the expression and enzymatic activity of NPM/ALK, as shown in BaF3 cells transfected with wild-type NPM/ALK and kinase-inactive NPM/ALK K210R mutant and by the inhibition of the NPM/ALK function in ALK+ TCL cells by a small-molecule ALK inhibitor. NPM/ALK induces HIF1α expression by upregulating its gene transcription through its key signal transmitter signal transducer and activator of transcription 3 (STAT3), which binds to the HIF1α gene promoter as shown by the chromatin immunoprecipitation assay and is required for HIF1α gene expression as demonstrated by its small interfering RNA-mediated depletion. In turn, depletion of HIF1α increases mammalian target of rapamycin complex 1 activation, cell growth and proliferation and decreases vascular endothelial growth factor synthesis. These results identify a novel cell-transforming property of NPM/ALK, namely its ability to induce the expression of HIF1α, a protein with an important role in carcinogenesis. These results also provide another rationale to therapeutically target NPM/ALK and STAT3 in ALK+ TCL.

  20. Successful TB treatment induces B-cells expressing FASL and IL5RA mRNA.

    Science.gov (United States)

    van Rensburg, Ilana C; Wagman, Chandre; Stanley, Kim; Beltran, Caroline; Ronacher, Katharina; Walzl, Gerhard; Loxton, Andre G

    2017-01-10

    Activated B-cells increase T-cell behaviour during autoimmune disease and other infections by means of cytokine production and antigen-presentation. Functional studies in experimental autoimmune encephalomyelitis (EAE) indicate that B-cell deficiencies, and a lack of IL10 and IL35 leads to a poor prognosis. We hypothesised that B-cells play a role during tuberculosis. We evaluated B-cell mRNA expression using real-time PCR from healthy community controls, individuals with other lung diseases and newly diagnosed untreated pulmonary TB patients at three different time points (diagnosis, month 2 and 6 of treatment).We show that FASLG, IL5RA, CD38 and IL4 expression was lower in B-cells from TB cases compared to healthy controls. The changes in expression levels of CD38 may be due to a reduced activation of B-cells from TB cases at diagnosis. By month 2 of treatment, there was a significant increase in the expression of APRIL and IL5RA in TB cases. Furthermore, after 6 months of treatment, APRIL, FASLG, IL5RA and CD19 were upregulated in B-cells from TB cases. The increase in the expression of APRIL and CD19 suggests that there may be restored activation of B-cells following anti-TB treatment. The upregulation of FASLG and IL5RA indicates that B-cells expressing regulatory genes may play an important role in the protective immunity against M.tb infection. Our results show that increased activation of B-cells is present following successful TB treatment, and that the expression of FASLG and IL5RA could potentially be utilised as a signature to monitor treatment response.

  1. Changes of bcl-xL and bax mRNA expression following traumatic brain injury in rats

    Institute of Scientific and Technical Information of China (English)

    骆纯; 卢亦成; 江基尧; 朱诚

    2002-01-01

    Objective: To investigate the changes of bcl-2 gene family and the molecular mechanism of neuronal apoptosis following traumatic brain injury (TBI) in rats.Methods: Male Sprague-Dawley (SD) rats were subjected to lateral fluid percussion brain injury (FPBI) of moderate severity. The bcl-xL and bax mRNA expression was detected by reverse transcription polymerase chain reaction (RT-PCR). In addition to morphological evidence of apoptosis, terminal deoxynucleotide transferase-mediated dUTP-biotin nick-end labeling (TUNEL) histochemistry was used to identify the DNA fragmentation in situ at both light and electron microscope levels, whereas characteristic internucleosomal DNA fragmentation of apoptosis was demonstrated by DNA gel electrophoresis.Results: The apoptotic response to trauma was regionally distinct and may be involved in both acute and delayed cell death. The bcl-xL mRNA expression of the impact site was significantly lower (67.42%±7.54%) than that of the ipsilateral hemisphere at 6 hours after injury (P<0.01). The decrease of bcl-xL mRNA expression preceded apoptosis at 24 hours after injury. The bax mRNA expression rose slowly, doubled at 3 days after injury and returned to the sham level slowly.Conclusions: Decreased expression of bcl-xL mRNA and increased expression of bax mRNA coincides with apoptosis following brain injury. The bcl-2 gene family is involved in neuronal apoptosis after TBI, and the changes of mRNA expression of the family members lead the neuronal cells to apoptosis.

  2. Increase in uncoupling protein-2 mRNA expression by BRL49653 and bromopalmitate in human adipocytes.

    Science.gov (United States)

    Viguerie-Bascands, N; Saulnier-Blache, J S; Dandine, M; Dauzats, M; Daviaud, D; Langin, D

    1999-03-05

    Uncoupling protein-2 (UCP2) is a novel mitochondrial protein that may be involved in the control of energy expenditure. We have previously reported an upregulation of adipose tissue UCP2 mRNA expression during fasting in humans. Analysis of changes in metabolic parameters suggested that fatty acids may be associated with the increased UCP2 mRNA level. Culture of human adipose tissue explants was used to study in vitro regulation of adipocyte UCP2 gene expression. A 48-h treatment with BRL49653 and bromopalmitate, two potent activators of PPARgamma, resulted in a dose-dependent increase in UCP2 mRNA levels. The induction by BRL49653 was rapid (from 6 h) and maintained up to 5 days. TNFalpha provoked a 2-fold decrease in UCP2 mRNA levels. Human recombinant leptin did not affect UCP2 mRNA expression. The data support the hypothesis that fatty acids are involved in the control of adipocyte UCP2 mRNA expression in humans. Copyright 1999 Academic Press.

  3. Effect of Radix Isatidis on the Expression of Moesin mRNA Induced by LPS in the Tissues of Mice

    Institute of Scientific and Technical Information of China (English)

    LI Jing; LIU Yunhai; FANG Jianguo; CHEN Xin; XIE Wei

    2007-01-01

    To investigate the effect of the anti-endotoxic part of Radix lsatidis on the expression of moesin mRNA in murine tissues induced by lipopolysaccharide (LPS), the sample solution of F022 part from Radix lsatidis was intraperitoneally administered to experimental mice, and the lipopolysaccharide (LPS) were injected into the tail vein, and then the tissues of liver, kidney and spleen were colleted and cut into slices. The mRNA was detected by moesin mRNA hybridization in situ. The staining results were observed under microscope. It was found that moesin mRNA expression was increased in the tissues of liver, kidndy and spleen in mice treated with LPS, while in the mice pre-treated with F022 part from Radix Isatidis, the LPS-induced moesin mRNA expressions in these tissues were inhibited in a dose-dependant manner. Our study showed that F022 part from Radix Isatidis can inhibit the LPS-induced expression of moesin mRNA in the tissues of liver, kidney and spleen in mice.

  4. Increased mRNA expression of cytochrome oxidase in dorsal raphe nucleus of depressive suicide victims

    Directory of Open Access Journals (Sweden)

    A Sanchez-Bahillo

    2008-04-01

    Full Text Available A Sanchez-Bahillo1, V Bautista-Hernandez1, Carlos Barcia Gonzalez1, R Bañon2, A Luna2, EC Hirsch3, Maria-Trinidad Herrero11Clinical and Experimental Neuroscience, Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED; 2Department of Legal Medicine, Department of Human Anatomy, School of Medicine, University of Murcia, Campus de Espinardo, Murcia 30100, Spain; 3INSERM U679 Hôpital de la Salpêtrière, Boulevard de l’Hôpital, Paris, FranceAbstract: Suicidal behavior is a problem with important social repercussions. Some groups of the population show a higher risk of suicide; for example, depression, alcoholism, psychosis or drug abuse frequently precedes suicidal behavior. However, the relationship between metabolic alterations in the brain and premorbid clinical symptoms of suicide remains uncertain. The serotonergic and noradrenergic systems have frequently been, implicated in suicidal behavior and the amount of serotonin in the brain and CSF of suicide victims has been found to be low compared with normal subjects. However, there are contradictory results regarding the role of noradrenergic neurons in the mediation of suicide attempts, possibly reflecting the heterogeneity of conditions that lead to a common outcome. In the present work we focus on the subgroup of suicide victims that share a common diagnosis of major depression. Based on post-mortem studies analyzing mRNA expression by in situ hybridization, serotonergic neurons from the dorsal raphe nucleus (DRN from depressive suicide victims are seen to over-express cytochrome oxidase mRNA. However, no corresponding changes were found in the expression of tyrosine hydroxylase (TH mRNA in the noradrenergic neurons of the Locus Coeruleus (LC. These results suggest that, despite of the low levels of serotonin described in suicide victims, the activity of DRN neurons could increase in the suicidally depressed, probably due to the over activation of

  5. Differential between Protein and mRNA Expression of CCR7 and SSTR5 Receptors in Crohn's Disease Patients

    Directory of Open Access Journals (Sweden)

    Nathalie Taquet

    2009-01-01

    Full Text Available Crohn's disease (CD is a multifactorial chronic inflammatory bowel disease of unknown cause. The aim of the present study was to explore if mRNA over-expression of SSTR5 and CCR7 found in CD patients could be correlated to respective protein expression. When compared to healthy donors, SSTR5 was over-expressed 417 ± 71 times in CD peripheral blood mononuclear cells (PBMCs. Flow cytometry experiments showed no correlation between mRNA and protein expression for SSTR5 in PBMCs. In an attempt to find a reason of such a high mRNA expression, SSTR5 present on CD PBMCs were tested and found as biologically active as on healthy cells. In biopsies of CD intestinal tissue, SSTR5 was not over-expressed but CCR7, unchanged in PBMCs, was over-expressed by 10 ± 3 times in the lamina propria. Confocal microscopy showed a good correlation of CCR7 mRNA and protein expression in CD intestinal biopsies. Our data emphasize flow and image cytometry as impossible to circumvent in complement to molecular biology so to avoid false interpretation on receptor expressions. Once confirmed by further large-scale studies, our preliminary results suggest a role for SSTR5 and CCR7 in CD pathogenesis.

  6. [Expression of c-fos mRNA following moderate lateral fluid percussion brain injury in rats].

    Science.gov (United States)

    Zhang, Y; Chen, G; Sun, G; Liu, M; Liao, Z; Wu, J; Wu, M

    2000-09-01

    This experiment was designed to study the expression of c-fos mRNA in brain following moderate lateral fluid percussion brain injury in rats and to observe the temporal pattern of its expressions following percussion. Male Sprague-Dawley rats were divided into normal control, sham operation control and injury groups. The rats of the injury group were subjected to moderate lateral fluid percussion injury (0.2 MPa). The injury group was then subdivided into 5 min, 15 min, 30 min, 1 h, 2 h groups according to the time elapsed after injury. The expression of c-fos mRNA was studied with reverse transcription polymerase chain reaction(RT-PCR) semi-quantitatively. c-fos mRNA in cortex and brain stem was expressed weakly in control groups. After 5 min of percussion, the expression of c-fos mRNA increased progressively and remained elevated up to 2 h after brain injury. This result suggested that the induction and expression of the c-fos mRNA in cortex and brain stem after fluid percussion brain injury were increased rapidly. The temporal pattern of induction in cortex was similar to that in brain stem.

  7. The mRNA expression of SATB1 and SATB2 in human breast cancer

    Directory of Open Access Journals (Sweden)

    Mansel Robert

    2009-07-01

    Full Text Available Abstract Background SATB1 is a nuclear protein that has been recently reported to be a 'genome organizer' which delineates specific epigenetic modifications at target gene loci, directly up-regulating metastasis-associated genes while down-regulating tumor-suppressor genes. In this study, the level of mRNA expression of SATB1 and SATB2 were assessed in normal and malignant breast tissue in a cohort of women with breast cancer and correlated to conventional clinico-pathological parameters. Materials and methods Breast cancer tissues (n = 115 and normal background tissues (n = 31 were collected immediately after excision during surgery. Following RNA extraction, reverse transcription was carried out and transcript levels were determined using real-time quantitative PCR and normalized against β-actin expression. Transcript levels within the breast cancer specimens were compared to the normal background tissues and analyzed against TNM stage, nodal involvement, tumour grade and clinical outcome over a 10 year follow-up period. Results The levels of SATB1 were higher in malignant compared with normal breast tissue (p = 0.0167. SATB1 expression increased with increasing TNM stage (TNM1 vs. TNM2 p = 0.0264, increasing tumour grade (grade1 vs. grade 3 p = 0.017; grade 2 vs. grade 3 p = 0.0437; grade 1 vs. grade 2&3 p = 0.021 and Nottingham Prognostic Index (NPI (NPI-1 vs. NPI-3 p = 0.0614; NPI-2 vs. NPI-3 p = 0.0495. Transcript levels were associated with oestrogen receptor (ER positivity (ER(- vs. ER(+ p = 0.046. SABT1 expression was also significantly correlated with downstream regulated genes IL-4 and MAF-1 (Pearson's correlation coefficient r = 0.21 and r = 0.162 and SATB2 (r = 0.506. After a median follow up of 10 years, there was a trend for higher SATB1 expression to be associated with shorter overall survival (OS. Higher levels of SATB2 were also found in malignant compared to background tissue (p = 0.049. SATB2 expression increased with

  8. EFFECTS OF MUTATION AND EXPRESSION OF PTEN GENE mRNA ON TUMORIGENESIS AND PROGRESSION OF EPITHELIAL OVARIAN CANCER

    Institute of Scientific and Technical Information of China (English)

    陈颖; 郑华川; 杨雪飞; 孙丽梅; 辛彦

    2004-01-01

    Objective To investigate the mutation and expression of tumor suppressor gene-PTEN mRNA and explore their roles in tumorigenesis and progression of ovarian cancer. Methods Mutated exon 5 of PTEN gene was examined in normal ovary (n = 5), ovarian cyst (n =5), ovarian borderline tumor (n=9), epithelial ovarian cancer (n=60), and ovarian cancer cell line (n= 1)by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). mRNA expression of PTEN gene was evaluated in corresponding tissues and cell line by reverse transcription polymerase chain reaction(RT-PCR). The mutation and mRNA expression of PTEN gene were compared with clinicopathological features of ovarian cancer. Results Mutated exon 5 of PTEN gene was detected only in 5 (7.1%) cases of epithelial ovarian cancer. mRNA expression level of PTEN gene in ovarian borderline tumor or ovarian cancer was lower than that in normal ovary or ovarian cyst (P < 0.05). The level of PTEN gene mRNA expression was negatively correlated with clinicopathological staging of ovarian cancer, whereas positively correlated with histological differentiation (P < 0.05). mRNA expression level of PTEN gene in ovarian endometrioid cancer was significantly lower than that in ovarian serous or mucinous cancer (P < 0.05). Conclusions Mutation of PTEN gene occurs in ovarian cancer. Down-regulated expression of PTEN is probably an important molecular event in tumorigenesis of ovarian cancer. Abnormal expression of PTEN gene is involved in progression of ovarian cancer. Reduced expression of PTEN gene is closely associated with tumorigenesis and pathobiological behaviors of ovarian endometrioid cancer.

  9. Rift Valley fever virus NS{sub S} gene expression correlates with a defect in nuclear mRNA export

    Energy Technology Data Exchange (ETDEWEB)

    Copeland, Anna Maria; Van Deusen, Nicole M.; Schmaljohn, Connie S., E-mail: Connie.s.schmaljohn.civ@mail.mil

    2015-12-15

    We investigated the localization of host mRNA during Rift Valley fever virus (RVFV) infection. Fluorescence in situ hybridization revealed that infection with RVFV altered the localization of host mRNA. mRNA accumulated in the nuclei of RVFV-infected but not mock-infected cells. Further, overexpression of the NS{sub S} gene, but not the N, G{sub N} or NS{sub M} genes correlated with mRNA nuclear accumulation. Nuclear accumulation of host mRNA was not observed in cells infected with a strain of RVFV lacking the gene encoding NS{sub S}, confirming that expression of NS{sub S} is likely responsible for this phenomenon. - Highlights: • Rift Valley fever virus (RVFV) infection alters the localization of host mRNA. • mRNA accumulates in the nuclei of RVFV-infected but not mock-infected cells. • NS{sub S} is likely responsible for mRNA relocalization to the nucleus.

  10. The mRNA expression profile of metabolic genes relative to MHC isoform pattern in human skeletal muscles

    DEFF Research Database (Denmark)

    Plomgaard, Peter; Penkowa, Milena; Leick, Lotte;

    2006-01-01

    was more than twofold higher in soleus and vastus than in triceps. Contrary, phosphofructokinase and total lactate dehydrogenase (LDH) activity was approximately three- and twofold higher in triceps than in both soleus and vastus. Expression of metabolic genes was assessed by determining the mRNA content...... of a broad range of metabolic genes. The triceps muscle had two- to fivefold higher MHC IIa, phosphofructokinase, and LDH A mRNA content and two- to fourfold lower MHC I, lipoprotein lipase, CD36, hormone-sensitive lipase, and LDH B and hexokinase II mRNA than vastus lateralis or soleus. Interestingly...

  11. Sensitive and precise quantification of insulin-like mRNA expression in Caenorhabditis elegans.

    Science.gov (United States)

    Baugh, L Ryan; Kurhanewicz, Nicole; Sternberg, Paul W

    2011-03-22

    Insulin-like signaling regulates developmental arrest, stress resistance and lifespan in the nematode Caenorhabditis elegans. However, the genome encodes 40 insulin-like peptides, and the regulation and function of individual peptides is largely uncharacterized. We used the nCounter platform to measure mRNA expression of all 40 insulin-like peptides as well as the insulin-like receptor daf-2, its transcriptional effector daf-16, and the daf-16 target gene sod-3. We validated the platform using 53 RNA samples previously characterized by high density oligonucleotide microarray analysis. For this set of genes and the standard nCounter protocol, sensitivity and precision were comparable between the two platforms. We optimized conditions of the nCounter assay by varying the mass of total RNA used for hybridization, thereby increasing sensitivity up to 50-fold and reducing the median coefficient of variation as much as 4-fold. We used deletion mutants to demonstrate specificity of the assay, and we used optimized conditions to assay insulin-like gene expression throughout the C. elegans life cycle. We detected expression for nearly all insulin-like genes and find that they are expressed in a variety of distinct patterns suggesting complexity of regulation and specificity of function. We identified insulin-like genes that are specifically expressed during developmental arrest, larval development, adulthood and embryogenesis. These results demonstrate that the nCounter platform provides a powerful approach to analyzing insulin-like gene expression dynamics, and they suggest hypotheses about the function of individual insulin-like genes.

  12. Sensitive and precise quantification of insulin-like mRNA expression in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    L Ryan Baugh

    Full Text Available Insulin-like signaling regulates developmental arrest, stress resistance and lifespan in the nematode Caenorhabditis elegans. However, the genome encodes 40 insulin-like peptides, and the regulation and function of individual peptides is largely uncharacterized. We used the nCounter platform to measure mRNA expression of all 40 insulin-like peptides as well as the insulin-like receptor daf-2, its transcriptional effector daf-16, and the daf-16 target gene sod-3. We validated the platform using 53 RNA samples previously characterized by high density oligonucleotide microarray analysis. For this set of genes and the standard nCounter protocol, sensitivity and precision were comparable between the two platforms. We optimized conditions of the nCounter assay by varying the mass of total RNA used for hybridization, thereby increasing sensitivity up to 50-fold and reducing the median coefficient of variation as much as 4-fold. We used deletion mutants to demonstrate specificity of the assay, and we used optimized conditions to assay insulin-like gene expression throughout the C. elegans life cycle. We detected expression for nearly all insulin-like genes and find that they are expressed in a variety of distinct patterns suggesting complexity of regulation and specificity of function. We identified insulin-like genes that are specifically expressed during developmental arrest, larval development, adulthood and embryogenesis. These results demonstrate that the nCounter platform provides a powerful approach to analyzing insulin-like gene expression dynamics, and they suggest hypotheses about the function of individual insulin-like genes.

  13. Changes of Expression of Stretch-activated Potassium Channel TREK-1 mRNA and Protein in Hypertrophic Myocardium

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The expression of stretch-activated potassium channel TREK-1 mRNA and protein of hypertrophic myocardium was measured. Using a model of hypertrophy induced by coarctation of abdominal aorta in male Wistar rats, the expression of TREK-1 mRNA and protein was detected by using semi quantitative RT PCR and Western blot respectively. At 4th and 8th week after constriction of the abdominal aorta, rats developed significant left ventricular hypertrophy. As compared to sham operated group, stretch activated potassium channel TREK-1 mRNA was strongly expressed and protein was up regulated in operation groups (P<0.05). It was concluded that the expression of TREK 1 was up regulated in hypertrophic myocardium induced by chronic pressure overload in Wistar rats.

  14. [Effect of airborne fungi on interleukin-4 mRNA expression in the peripheral blood mononuclear cells of rats].

    Science.gov (United States)

    Kan, H; Song, W; Jiang, R

    2001-09-01

    The effect of indoor airborne fungi on interleukin-4 (IL-4) mRNA expression in the peripheral blood mononuclear cells (PBMCs) of rats was studied by inoculating via trachea with fungi suspension. A simplified competitive RT-PCR method was used to measure the IL-4 mRNA expression in plasma. The results showed that interleukin-4 expression was much higher in the affected groups than that in the control one. It suggested that IL-4 might play an important role in the sensitization progress of fungi.

  15. Role of CTGF in White Matter Development in Tuberous Sclerosis

    Science.gov (United States)

    2015-02-01

    both in vitro and in vivo. KEYWORDS Tuberous Sclerosis Complex, myelination , CTGF ACCOMPLISHMENTS: We will review the accomplishments to date in...reporting burden for the collection of information is estimated to average 1 hour per response, including the time for reviewing instructions...searching existing data sources, gathering and maintaining the data needed, and completing and reviewing the collection of information. Send comments

  16. The vitamin D receptor localization and mRNA expression in ram testis and epididymis.

    Science.gov (United States)

    Jin, Hui; Huang, Yang; Jin, Guang; Xue, Yanrong; Qin, Xiaowei; Yao, Xiaolei; Yue, Wenbing

    2015-02-01

    The objectives of present study were to investigate the presence of vitamin D receptor (VDR) in testis and epididymis of ram by polymerase chain reaction (PCR), to locate VDR in testis and epididymis by immunohistochemistry and to compare difference of VDR expression between testis and epididymis before and after sexual maturation by Real time-PCR and Western blot. The results showed that VDR exists in the testis and epididymis of ram while VDR protein in testis and epididymis was localized in Leydig cells, spermatogonial stem cells, spermatocytes, Sertoli cells and principal cells. For the adult ram, the amounts of VDR mRNA and VDR protein were less (p ram, the result showed the same trend (p 0.05) between adult and prepubertal. In conclusion, VDR exists in testis and epididymis of ram, suggesting 1α,25-(OH)(2)VD(3) may play a role in ram reproduction.

  17. Multiple correlation analyses revealed complex relationship between DNA methylation and mRNA expression in human peripheral blood mononuclear cells.

    Science.gov (United States)

    Xie, Fang-Fei; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Hong; Wu, Jian; Guo, Yu-Fan; Zeng, Ke-Qin; Wang, Ming-Jun; Zhu, Xiao-Wei; Xia, Wei; Wang, Lan; He, Pei; Bing, Peng-Fei; Lu, Xin; Zhang, Yong-Hong; Lei, Shu-Feng

    2017-07-22

    DNA methylation is an important regulator on the mRNA expression. However, a genome-wide correlation pattern between DNA methylation and mRNA expression in human peripheral blood mononuclear cells (PBMCs) is largely unknown. The comprehensive relationship between mRNA and DNA methylation was explored by using four types of correlation analyses and a genome-wide methylation-mRNA expression quantitative trait locus (eQTL) analysis in PBMCs in 46 unrelated female subjects. An enrichment analysis was performed to detect biological function for the detected genes. Single pair correlation coefficient (r T1) between methylation level and mRNA is moderate (-0.63-0.62) in intensity, and the negative and positive correlations are nearly equal in quantity. Correlation analysis on each gene (T4) found 60.1% genes showed correlations between mRNA and gene-based methylation at P correlation (R T4 > 0.8). Methylation sites have regulation effects on mRNA expression in eQTL analysis, with more often observations in region of transcription start site (TSS). The genes under significant methylation regulation both in correlation analysis and eQTL analysis tend to cluster to the categories (e.g., transcription, translation, regulation of transcription) that are essential for maintaining the basic life activities of cells. Our findings indicated that DNA methylation has predictive regulation effect on mRNA with a very complex pattern in PBMCs. The results increased our understanding on correlation of methylation and mRNA and also provided useful clues for future epigenetic studies in exploring biological and disease-related regulatory mechanisms in PBMC.

  18. OIL FLY ASH AND VANADIUM DIMINISH NRAMP-2MRNA AND PROTEIN EXPRESSION IN HUMAN BRONCHIAL EPITHELIAL CELLS

    Science.gov (United States)

    The capacity of Nramp2 to transport iron and its ubiquitous expression make it a likely candidate for transferrin-independent uptake of iron in peripheral tissues. Airway epithelial cells increase both mRNA and expression of that isoform of Nramp-2 without an iron response ele...

  19. Duodenal calcium transporter mRNA expression in stressed male rats treated with diazepam, fluoxetine, reboxetine, or venlafaxine.

    Science.gov (United States)

    Charoenphandhu, Narattaphol; Teerapornpuntakit, Jarinthorn; Lapmanee, Sarawut; Krishnamra, Nateetip; Charoenphandhu, Jantarima

    2012-10-01

    Chronic stress has been reported to decrease bone density and intestinal calcium absorption, but its underlying mechanism remains elusive. Since long-term exposure to glucocorticoids, major stress hormones from adrenal gland, is known to downregulate the mRNA expression of intestinal calcium transporter TRPV6, the present study aimed to demonstrate whether decreases in mRNA expressions of duodenal calcium transporter genes were observed in male rats subjected to restraint stress for 4 weeks. The results from quantitative real-time PCR showed that restraint stress significantly downregulated the mRNA expressions of apical calcium channels (TRPV6 and Ca(v)1.3), cytoplasmic calcium-binding protein (calbindin-D(9k)), and basolateral calcium pump (PMCA(1b)), but not the expression of TRPV5 or NCX1. The mRNA expressions of paracellular genes, ZO-1, occludin, and claudin-3, were not altered by restraint stress. Since several antidepressant or anxiolytic drugs effectively alleviate stress-induced depressive and anxiety symptoms, we further hypothesized that these drugs may also enhance calcium transporter gene expression in stressed rats. As expected, 4-week daily administration of 10 mg/kg fluoxetine, 10 mg/kg reboxetine, or 10 mg/kg venlafaxine differentially increased calcium transporter mRNA expression in stressed rats, whereas 2 mg/kg diazepam had no such effect. It could, therefore, be concluded that 4-week restraint stress downregulated some important calcium transporter mRNA expression in the duodenal epithelial cells of male rats, which could be prevented by oral administration of fluoxetine, reboxetine, and venlafaxine. The present findings may be applied to help alleviate the stress-induced bone loss and osteoporosis by restoring intestinal calcium absorption to provide calcium for bone formation.

  20. Effects of corticosteroid on the expressions of neuropeptide and cytokine mRNA and on tenocyte viability in lateral epicondylitis

    Directory of Open Access Journals (Sweden)

    Han Soo

    2012-10-01

    Full Text Available Abstract Background The purpose of this study was to determine the reaction mechanism of corticosteroid by analyzing the expression patterns of neuropeptides (substance P (SP, calcitonin gene related peptide (CGRP and of cytokines (interleukin (IL-1α, tumor growth factor (TGF-β after corticosteroid treatment in lateral epicondylitis. In addition, we also investigated whether corticosteroid influenced tenocyte viability. Methods The corticosteroid triamcinolone acetonide (TAA was applied to cultured tenocytes of lateral epicondylitis, and the changes in the mRNA expressions of neuropeptides and cytokines and tenocyte viabilities were analyzed at seven time points. Quantitative real-time polymerase chain reaction and an MTT assay were used. Results The expression of SP mRNA was maximally inhibited by TAA at 24 hours but recovered at 72 hours, and the expressions of CGRP mRNA and IL-1α mRNA were inhibited at 24 and 3 hours, respectively. The expression of TGF-β mRNA was not significant. Tenocyte viability was significantly reduced by TAA at 24 hours. Conclusions We postulate that the reaction mechanism predominantly responsible for symptomatic relief after a corticosteroid injection involves the inhibitions of neuropeptides and cytokines, such as, CGRP and IL-1α. However the tenocyte viability was compromised by a corticosteroid.

  1. Calprotectin mRNA (MRP8/MRP14 expression in neutrophils of periodontitis patients with type 2 diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Ahmad Syaify

    2009-09-01

    Full Text Available Background: Calprotectin, a major cytosolic protein of leukocytes, is detected in neutrophils and monocytes/machrophages. This protein is known to be a marker for several inflammatory diseases including periodontitis. In type 2 diabetes mellitus patients, the severity of periodontitis was strongly thought to be caused by decreasing of leukocytes function such as neutrophils. Previous research found that the calprotectin level in serum of periodontitis patients with type 2 DM is higher than periodontits patients non DM. Purpose: The aim of this study was to determine calprotectin mRNA (MRP8/MRP14 expression in human neutrophils of periodontitis patients with type 2 diabetes mellitus. Methods: Neutrophils were isolated from the peripheral blood of periodontitis patients with uncontrolled type 2 DM, controlled type 2 DM, and non DM. The expression of calprotectin mRNA (MRP8 and MRP14 were detected by RTPCR. Result: The result showed that the value of mRNA calprotectin expression in DM patients were higher than non DM, and the highest expression was on the uncontrolled type 2 DM. Conclusion: The basal level of calprotectin mRNA MRP8/MRP14 expression increased in neutrophil of periodontitis patient with type 2 DM compared non diabetic subjects. It was suggested that high basal level of calprotectin mRNA has a role in the regulation of periodontitis severity with diabetes mellitus patients.

  2. Expression of heparanase mRNA in anti-sense oligonucleotide-transfected human esophageal cancer EC9706 cells

    Institute of Scientific and Technical Information of China (English)

    Kui-Sheng Chen; Lan Zhang; Lin Tang; Yun-Han Zhang; Dong-Ling Gao; Liang Yan; Lei Zhang

    2005-01-01

    AIM: To investigate the effects of anti-sense oligonucleotides (ASODNs) on mRNA expression of heparanase in human esophageal cancer EC9706 cells.METHODS: One non-sense oligonucleotide (N-ODN) and five ASODNs against different heparanase mRNA sites were transfected into EC9706 cells, then the expression of heparanase mRNA in EC9706 cells was studied byin situ hybridization.RESULTS: The expression of heparanase mRNA could be inhibited by ASODNs.There was no significant difference among five ASODNs (P>0.05), but there was a significant difference between ASODNs and N-ODN or non-transfected group (ASODN1: 2.25±0.25, ASODN2: 2.21±0.23, ASODN3:2.23±0.23, ASODN4:2.25±0.24 vs N-ODN: 3.47±2.80 or non- transfected group: 3.51±2.93 respectively, P<0.05).CONCLUSION: The expression of heparanase mRNA in EC9706 cells can be inhibited by ASODNs in vivo, and heparanase ASODNs can inhibit metastasis of esophageal squamous cell carcinoma or other tumors by inhibiting the expression of heparanase.

  3. Expression of MCMV M83 mRNA in Infected Testes and Its Relationship with Testicular Lesions

    Institute of Scientific and Technical Information of China (English)

    Jin-wen XIONG; Cheng-liang XIONG; Yuan ZHU; Yong-hong TIAN

    2005-01-01

    Objective To investigate the expression of murine cytomegalovirus (MCMV) M83mRNA in mouse testes at the different periods of MCMV infection and its relationshipwith the severity of testicular lesionsMethods Used digoxin-labeled MCMV mRNA oligonucleotide probe, the mousetesticular tissues at the different periods following MCMV inoculation were detectedby in situ hybridization(ISH). The average absorbency (A) of ISH positive signalsrepresented the resticular MCMV load. The expression of MCMV M83 mRNA in theinfected testes and its relation with the severity of testicular lesions were analyzed.Results At different periods after MCMV inoculation, the expression of MCMV M83mRNA was detectable in the testicular tissues. The ISH positive signals were mainlylocated in cytoplasm of the testicular interstitial cells. The average A values at d 9after MCMV infection were the highest (comparison between the experimental groups,P<0. 001). There were pathological inflammatory changes to varying degrees inseminiferous epithelia and interstitial cells at the different periods of infection, especiallyduring d 6 to d 9.Conclusion In testicular MCMV infection, the levels of MCMV M83 mRNA wererelated with the severity of testicular lesions, suggesting the dynamic detection of MCMVM83 mRNA could evaluate the prognosis of MCMV infection.

  4. Sequencing and expression analysis of hepcidin mRNA in donkey (Equus asinus liver

    Directory of Open Access Journals (Sweden)

    José P. Oliveira-Filho

    2012-10-01

    Full Text Available The hypoferremia that is observed during systemic inflammatory processes is mediated by hepcidin, which is a peptide that is mainly synthesized in the livers of several mammalian species. Hepcidin plays a key role in iron metabolism and in the innate immune system. It's up-regulation is particularly useful during acute inflammation, and it restricts the iron availability that is necessary for the growth of pathogenic microorganisms. In this study, the hepcidin mRNA of Equus asinus has been characterized, and the expression of donkey hepcidin in the liver has been determined. The donkey hepcidin sequence has an open reading frame (ORF of 261 nucleotides, and the deduced corresponding protein sequence has 86 amino acids. The amino acid sequence of donkey hepcidin was most homologous to Equus caballus (98%. The mature donkey hepcidin sequence (25 amino acids was 100% homologous to the equine mature hepcidin and has eight conserved cysteine residues that are found in all of the investigated hepcidin sequences. The expression profile of donkey hepcidin in the liver was high and was similar to the reference gene expression. The donkey hepcidin sequence was deposited in GenBankTM (HQ902884 and may be useful for additional studies on iron metabolism and the inflammatory process in this species.

  5. Expression of neuropeptide receptor mRNA during osteoblastic differentiation of mouse iPS cells.

    Science.gov (United States)

    Nagao, Satomi; Goto, Tetsuya; Kataoka, Shinji; Toyono, Takashi; Joujima, Takaaki; Egusa, Hiroshi; Yatani, Hirofumi; Kobayashi, Shigeru; Maki, Kenshi

    2014-12-01

    Various studies have shown a relationship between nerves and bones. Recent evidence suggests that both sensory and sympathetic nerves affect bone metabolism; however, little is known about how neuropeptides are involved in the differentiation of pluripotent stem cells into osteoblastic (OB) cells. To evaluate the putative effects of neuropeptides during the differentiation of mouse induced pluripotent stem (iPS) cells into calcified tissue-forming OB cells, we investigated the expression patterns of neuropeptide receptors at each differentiation stage. Mouse iPS cells were seeded onto feeder cells and then transferred to low-attachment culture dishes to form embryoid bodies (EBs). EBs were cultured for 4 weeks in osteoblastic differentiation medium. The expression of α1-adrenergic receptor (AR), α2-AR, β2-AR, neuropeptide Y1 receptor (NPY1-R), neuropeptide Y2 receptor (NPY2-R), calcitonin gene-related protein receptor (CGRP-R), and neurokinin 1-R (NK1-R) was assessed by reverse transcription-polymerase chain reaction (RT-PCR) and real-time PCR. Among these neuropeptide receptors, CGRP-R and β2-AR were expressed at all stages of cell differentiation, including the iPS cell stage, with peak expression occurring at the early osteoblastic differentiation stage. Another sensory nervous system receptor, NK1-R, was expressed mainly in the late osteoblastic differentiation stage. Furthermore, CGRP-R mRNA showed an additional small peak corresponding to EBs cultured for 3 days, suggesting that EBs may be affected by serum CGRP. These data suggest that the sensory nervous system receptor CGRP-R and the sympathetic nervous system receptor β2-AR may be involved in the differentiation of iPS cells into the osteoblastic lineage. It follows from these findings that CGRP and β2-AR may regulate cell differentiation in the iPS and EB stages, and that each neuropeptide has an optimal period of influence during the differentiation process.

  6. Rev-erb beta regulates the Srebp-1c promoter and mRNA expression in skeletal muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Ramakrishnan, Sathiya N.; Lau, Patrick; Crowther, Lisa M. [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia); Cleasby, Mark E. [Diabetes and Obesity Research Program, Garvan Institute of Medical Research, St. Vincent' s Hospital, 384 Victoria Street, Darlinghurst, Sydney, NSW 2010 (Australia); Millard, Susan; Leong, Gary M. [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia); Cooney, Gregory J. [Diabetes and Obesity Research Program, Garvan Institute of Medical Research, St. Vincent' s Hospital, 384 Victoria Street, Darlinghurst, Sydney, NSW 2010 (Australia); Muscat, George E.O., E-mail: g.muscat@imb.uq.edu.au [The University of Queensland, Institute for Molecular Bioscience, St. Lucia, Qld 4072 (Australia)

    2009-10-30

    The nuclear hormone receptor, Rev-erb beta operates as a transcriptional silencer. We previously demonstrated that exogenous expression of Rev-erb{beta}{Delta}E in skeletal muscle cells increased Srebp-1c mRNA expression. We validated these in vitro observations by injection of an expression vector driving Rev-erb{beta}{Delta}E expression into mouse tibialis muscle that resulted in increased Srebp-1c mRNA expression. Paradoxically, Rev-erb{beta} siRNA expression in skeletal muscle cells repressed Srebp-1c expression, and indicated that Rev-erb{beta} expression was necessary for Srebp-1c expression. ChIP analysis demonstrated that Rev-erb{beta} was recruited to the Srebp-1c promoter. Moreover, Rev-erb{beta} trans-activated the Srebp-1c promoter, in contrast, Rev-erb{beta} efficiently repressed the Rev-erb{alpha} promoter, a previously characterized target gene. Finally, treatment with the Rev-erb agonist (hemin) (i) increased the trans-activation of the Srebp-1c promoter by Rev-erb{beta}; and (ii) increased Rev-erb{beta} and Srebp-1c mRNA expression. These data suggest that Rev-erb{beta} has the potential to activate gene expression, and is a positive regulator of Srebp-1c, a regulator of lipogenesis.

  7. mRNA expression of genes regulating lipid metabolism in ringed seals (Pusa hispida) from differently polluted areas

    Energy Technology Data Exchange (ETDEWEB)

    Castelli, Martina Galatea [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway); University of Bergen, Department of Biology, 5020 Bergen (Norway); Rusten, Marte; Goksøyr, Anders [University of Bergen, Department of Biology, 5020 Bergen (Norway); Routti, Heli, E-mail: heli.routti@npolar.no [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway)

    2014-01-15

    Highlights: •Genes regulating lipid metabolism were studied in ringed seals. •We compared highly contaminated Baltic seals and less contaminated Svalbard seals. •mRNA expression of hepatic PPARγ was higher in the Baltic seals. •mRNA expression of adipose PPARγ target genes was higher in the Baltic seals. •Contaminant exposure may affect lipid metabolism in the Baltic ringed seals. -- Abstract: There is a growing concern about the ability of persistent organic pollutants (POPs) to influence lipid metabolism. Although POPs are found at high concentrations in some populations of marine mammals, for example in the ringed seal (Pusa hispida) from the Baltic Sea, little is known about the effects of POPs on their lipid metabolism. An optimal regulation of lipid metabolism is crucial for ringed seals during the fasting/molting season. This is a physiologically stressful period, during which they rely on the energy stored in their fat reserves. The mRNA expression levels for seven genes involved in lipid metabolism were analyzed in liver and/or blubber tissue from molting ringed seals from the polluted Baltic Sea and a less polluted reference location, Svalbard (Norway). mRNA expression of genes encoding peroxisome proliferator-activated receptors (PPAR) α and γ and their target genes acyl-coenzyme A oxidase 1 (ACOX1) and cluster of differentiation 36 (CD36) were analyzed in liver. mRNA expression level of genes encoding PPARβ, PPARγ and their target genes encoding fatty acid binding protein 4 (FABP4) and adiponectin (ADIPOQ) were measured in inner and middle blubber layers. In addition, we evaluated the influence of molting status on hepatic mRNA expression of genes encoding PPARs and their target genes in ringed seals from Svalbard. Our results show higher mRNA expression of genes encoding hepatic PPARγ and adipose PPARβ, FABP4, and ADIPOQ in the Baltic seals compared to the Svalbard seals. A positive relationship between mRNA expressions of genes

  8. cDNA cloning and mRNA expression of neuropeptide Y in orange spotted grouper, Epinephelus coioides.

    Science.gov (United States)

    Chen, Rong; Li, Wensheng; Lin, Haoran

    2005-09-01

    A full-length cDNA encoding the neuropeptide Y (NPY) was cloned from the hypothalamus of orange spotted grouper (Epinephelus coioides) by rapid amplification of cDNA ends approaches. The NPY cDNA sequence is 688 bp long and has an open reading frame of 300 bp encoding prepro-NPY with 99 amino acids. The deduced amino acid sequences contain a 28-amino-acids signal peptide followed by a 36-amino-acids mature NPY peptide. mRNA expression of NPY was determined using semi-quantitative RT-PCR followed by Southern blot analysis. NPY mRNA was expressed in olfactory bulb, telencephalon, pituitary, hypothalamus, optic tectum-thalamus, medulla oblongata, cerebellum and spinal cord. Low levels of NPY mRNA expression were found in retina, ovary and stomach, while much lower levels of expression were detected in liver, heart, gill, skin, anterior intestine, thymus and blood. No NPY mRNA expression was observed in unfertilized eggs, newly fertilized eggs, 16-cells stage and morula stage of the embryo and lower levels of expression were detected in the blastula, gastrula and neurula stages. It was highly expressed from lens formation stage to 52-day-old larval stage. NPY might be involved in the late embryonic and larval development of the orange spotted grouper.

  9. Cloning of human brevican cDNA and expression of its mRNA in human glioma

    Institute of Scientific and Technical Information of China (English)

    韩唏; 董艳; 由振东; 何成; 卢亦成

    2003-01-01

    Objective:To clone the cDNA of human brevican secreting isoform and to investigate its mRNA expression in human glioma.Methods:The full-length cDNA of human brevican secreted isoform was cloned from a human ahaplastic astrocytoma by RT-PCR,and the expression of human brevican mRNA in 22 cases of human glioma and 13 cases of non-glial brain tumors were investigated by in situ hybridization.Results:The cDNA which including the whole open reading frame of human brevican secreted isoform was obtained.In situ hybridization showed that brevican positive cells were present in all of the 22 cases of gliomas(100%),whereas none were found in the 13 cases of non-glial and metastasis brain tumors examined.Conclusion:The results suggest that brevican mRNA is highly and specifically expressed in human glioma.

  10. Expression of pulmonary mRNA encoding ICAM-1, VCAM-1, and P-selectin following thoracic irradiation in mice

    Energy Technology Data Exchange (ETDEWEB)

    Tsujino, Kayoko; Kodama, Akihisa; Nanaoka, Noriyoshi; Maruta, Tsutomu; Kono, Michio [Kobe Univ. (Japan). School of Medicine

    1999-08-01

    Recent studies have revealed that ionizing radiation induces the expression of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and P-selectin in vitro. The purpose of this study was to investigate the expression of these adhesion molecules in mouse lung following whole thoracic irradiation. C57BL/6J mice were irradiated with a single dose of 12 Gy to the thoraces and sacrificed at 4, 12, 24, and 48 hours and 1, 2, 4, and 8 weeks after irradiation. Expression of total lung mRNA for ICAM-1, VCAM-1, and P-selectin was quantified by the Northern blot method and normalized to {beta}-actin. There were increases in mRNA for ICAM-1 of 42% at 4 hours (p<0.05), 76% at 24 hours (p<0.01), and 51% at 48 hours (p<0.05) compared with the controls. There returned to the control level at 1 week. The expression of VCAM-1 mRNA was also increased by 49% (p<0.01) at 12 hours and was still increased by 25% at 1 week. P-selectin mRNA was transiently increased by 59% at 12 hours. These early inductions of mRNA for ICAM-1, VCAM-1, and P-selectin in mouse lung following thoracic irradiation were transient but significant, and are one of the most immediate changes reported in vivo. (author)

  11. Distinct regulation of vasoactive intestinal peptide (VIP) expression at mRNA and peptide levels in human neuroblastoma cells.

    Science.gov (United States)

    Agoston, D V; Colburn, S; Krajniak, K G; Waschek, J A

    1992-05-25

    Neuronal differentiation was induced in cultures of the human neuroblastoma cell line subclone SH-SY5Y by 14-day treatment with dibutyryl cAMP (dBcAMP), retinoic acid, and phorbol 12-myristate 13-acetate (PMA). An approximate 4-fold increase in vasoactive intestinal peptide (VIP) mRNA concentration was observed after differentiation with retinoic acid, whereas no change in VIP mRNA concentration was observed after differentiation with dBcAMP or PMA. A short-term treatment of cells with PMA did however result in a 5-fold transient increase in VIP mRNA; prior differentiation with retinoic acid or dBcAMP diminished this effect. Observed increases in VIP mRNA were in all cases accompanied by increases in VIP immunoreactivity. Remarkably, however, long-term treatment of cells with dBcAMP, which caused no change in mRNA levels, resulted in a six-fold increase in VIP immunoreactivity. Acute (36-h) treatment with carbachol also caused an increase in VIP immunoreactivity (about 2-fold, and blocked by atropine) without an increase in VIP mRNA level. Thus, a quantitative change in gene transcription or mRNA stability appears not to be a prerequisite for increased VIP expression, indicating that regulation can occur at translational or post-translational steps.

  12. Akt pathway activation and increased neuropeptide Y mRNA expression in the rat hippocampus: implications for seizure blockade.

    Science.gov (United States)

    Goto, Eduardo M; Silva, Marcelo de Paula; Perosa, Sandra R; Argañaraz, Gustavo A; Pesquero, João B; Cavalheiro, Esper A; Naffah-Mazzacoratti, Maria G; Teixeira, Vicente P C; Silva, José A

    2010-04-01

    The aim of this study was to analyze the expression of survival-related molecules such Akt and integrin-linked kinase (ILK) to evaluate Akt pathway activation in epileptogenesis process. Furthermore, was also investigated the mRNA expression of neuropeptide Y, a considered antiepileptic neuropeptide, in the pilocarpine-induced epilepsy. Male Wistar rats were submitted to the pilocarpine model of epilepsy. Hippocampi were removed 6h (acute phase), 12h (late acute), 5d (silent) and 60d (chronic) after status epilepticus (SE) onset, and from animals that received pilocarpine but did not develop SE (partial group). Hippocampi collected were used to specify mRNA expression using Real-Time PCR. Immunohistochemistry assay was employed to place ILK distribution in the hippocampus and Western blot technique was used to determine Akt activation level. A decrease in ILK mRNA content was found during acute (0.39+/-0.03) and chronic (0.48+/-0.06) periods when compared to control group (0.87+/-0.10). Protein levels of ILK were also diminished during both periods. Partial group showed increased ILK mRNA expression (0.80+/-0.06) when compared with animals in the acute stage. Silent group had ILK mRNA and immunoreactivity similar to control group. Western blot assay showed an augmentation in Akt activation in silent period (0.52+/-0.03) in comparison with control group (0.44+/-0.01). Neuropeptide Y mRNA expression increased in the partial group (1.67+/-0.22) and in the silent phase (1.45+/-0.29) when compared to control group (0.36+/-0.12). Results suggest that neuropeptide Y (as anticonvulsant) might act in protective mechanisms occurred during epileptic phenomena. Together with ILK expression and Akt activation, these molecules could be involved in hippocampal neuroprotection in epilepsy. Copyright 2009 Elsevier Ltd. All rights reserved.

  13. Expression of DNA Repair Enzyme hMTH1 mRNA and Protein in Hepatocellular Carcinoma

    Institute of Scientific and Technical Information of China (English)

    ZHOU Hejun; CHENG Bin; LIN Jusheng

    2005-01-01

    To study the expression of DNA repair enzyme hMTH1 mRNA and protein in hepatocellular carcinoma (HCC) tissues, tissues adjacent to the cancers, normal liver cells and hepatoma cell lines, and to investigate their function in the progress of HCC, semi-quantitative reverse-transcription polymerase chain reaction (RT-PCR) was employed to examine the expression of hMTH1 mRNA in matched HCC tissues (HT)/surrounding tissues (HST) of HCC, normal liver cell L02 and hepatoma cell lines SMMC7721, HepG2. hMTH1 protein was detected in corresponding HT as well as their HST by immunohistochemistry. Our results showed that the expression level of hMTH1 mRNA in HT was higher than that in HST (t=2.424, P <0.05). The expression level of hMTH1 mRNA in two hepatoma cell lines was higher than that in normal liver cell line (F=6.810, P <0.01). The expression of hMTH1 mRNA in SMMC7721 was similar to that in HepG2. hMTH1 protein was 88.2 % (15 of 17) positive in HT and 82.4 % (14 of 17) in HST. The protein level of hMTH1 in HT was correspondingly higher than in their HST (t=2.618,P<0.05). It is concluded that hMTH1 mRNA and protein were over-expressed in HCC and hepatoma cell lines. It may be one of the key events during the carcinogenesis,progression of HCC and may promote the malignant growth. These results suggest that hMTH1 plays a role in HCC and may be a candidate marker for the diagnosis of HCC.

  14. AT1a Receptor Has Interacted with Angiotensin-converting Enzymes 2 mRNA Expression in Mouse Brainstem

    Institute of Scientific and Technical Information of China (English)

    Zhanyi Lin; Shuguang Lin

    2008-01-01

    Objectives To examine in vivo interactions between angiotensin Ⅱ(Ang Ⅱ) AT1a receptor (AT1aR),angiotensin-converting enzymes (ACE) and ACE2 using small hairpin RNA (shRNA) gene-silencing methods in mice brainstem nucleus ttactus solitarius (NTS).Methods C57BL mice (n=8) were used as animal model.Method of microinjection in the nucleus of NTS was adopted.After ten days,mice were killed and their brain tissue were fixed and sectioned.The expression levels of AT1 aR,ACE and ACE2 mRNA at both sides of NTS were examined by in situ hybridization.Based on compared t-test,the changing for mRNA expression was examined.Results After the expression of AT1aR mRNA was significantly inhibited (61.6%±6.8% ) by AT1aR-shRNA,it was associated with decreases in ACE2 mRNA expression from (1.05±0.12) μCi/mg to (0.74±0.09) μCi/mg (29.0%±14.5%,P<0.01) on the same side of the brainstem.ACE mRNA expression was consistent at both sides (0.50 μCi/mg±0.09 μCi/mg and 0.53 μCi/mg±0.08 μCi/mg),with insignificant difference (P>0.05).Condusions The gene silencing result showed that there were interactions between brainstem AT1aR and ACE2.ACE mRNA expression was not altered by RNA interference treatment at AT1aR.

  15. Quantitative detection of metastasis-associated1mRNA and protein expression in breast cancer%Quantitative detection of metastasis-associated 1 mRNA and protein expression in breast cancer

    Institute of Scientific and Technical Information of China (English)

    Hiroko Yamashita; Tatsuya Toyama; Hiroshi Sugiura; Mei ZHANG; Shunzo Kobayashi; Yoshitaka Fujii; Hirotaka Iwase; Zhenhuan ZHANG

    2013-01-01

    Objective Understanding the mechanism of breast cancer metastasis will benefit those patients in need of aggressive treatment and avoid side effects caused by chemotherapy over treatment.Recently,a potential metastasis-associated gene and its product,the metastasis-associated 1 (MTA1),were identified; this gene has been found to be overexpressed in a variety of cancers.Methods In the present study,therefore,the level of expression of MTA1 mRNA has been assessed by LightCycler quantitative real-time RT-PCR in 160 cases of invasive carcinoma of the breast.MTA1 protein expression level was also detected by immunohistochemistry from available paired tissues of 154 cases.Associations between MTA1 mRNA and protein expression and clinicopathological factors were analyzed.Results It was found that MTA1 mRNA was expressed at significantly higher levels in patients with negative lymph node status,with ER and PgR positive and HER2 negative tumor.No difference was found between patient age,tumor size and histological grade groups.Patients with high levels of expression of MTA1 mRNA had a better prognosis than those with low expression.However,no difference was found between the protein level and clinicopathological factors.Univariate and multivariate prognostic analysis did not demonstrate that MTA1 mRNA was an independent prognostic factor for breast cancer.Conclusion In breast cancer,inconsistent with other tumor types,MTA1 gene expression is correlated with non-invasive clinicopathological factors and longer survival,which might suggest MTA1 gene is a tumor type specific metastasis associated gene.

  16. TP53 Promoter Methylation in Primary Glioblastoma: Relationship with TP53 mRNA and Protein Expression and Mutation Status

    OpenAIRE

    JESIONEK-KUPNICKA, DOROTA; Szybka, Malgorzata; Malachowska, Beata; Fendler, Wojciech; Potemski, Piotr; Piaskowski, Sylwester; Jaskolski, Dariusz; Papierz, Wielislaw; Skowronski, Wieslaw; Och, Waldemar; Kordek, Radzislaw; ZAWLIK, IZABELA

    2014-01-01

    Reduced expression of TP53 by promoter methylation has been reported in several neoplasms. It remains unclear whether TP53 promoter methylation is associated with reduced transcriptional and protein expression in glioblastoma (GB). The aim of our work was to study the impact of TP53 methylation and mutations on TP53 mRNA level and protein expression in 42 molecularly characterized primary GB tumors. We also evaluate the impact of all molecular alterations on the overall patient survival. The ...

  17. Negative correlation between serum uric acid and kidney URAT1 mRNA expression caused by resveratrol in rats.

    Science.gov (United States)

    Lee, Cheng-Tse; Chang, Li-Ching; Liu, Ching-Wen; Wu, Pei-Fung

    2017-10-01

    This study established a hyperuricemic rat model to elucidate the effect of resveratrol on the transport of UA in the kidney. Hyperuricemia was induced in rats through daily oral gavage of a potassium oxonate and UA mixture over 3 weeks. Our results revealed that resveratrol significantly reduced the serum UA levels but not creatinine, c-creative protein, alanine aminotransferase, or aspartate aminotransferase levels in these rats. Furthermore, renal URAT1 and OAT1 mRNA expression were significantly higher in the rats treated with allopurinol than in those with no treatment. Therefore, allopurinol not only inhibited UA production but also mediated renal URAT1 and OAT1 expression. The correlation analysis revealed that UA levels correlated negatively with renal IL-6 mRNA expression in rats treated with allopurinol. Moreover, URAT1 showed strong immunoreactivity in the distal convoluted tubule of rats treated with allopurinol or resveratrol and in hyperuricemic treated with allopurinol. Finally, in the rats treated with resveratrol, UA levels correlated negatively with renal URAT1 mRNA expression; thus, resveratrol reduced URAT1 mRNA expression under high UA levels, thereby reducing UA reabsorption in renal cells. Resveratrol contributes to URAT1 expression, which is potentially useful in therapeutic strategies aimed at treating hyperuricemia. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Microarray-based mRNA expression profiling of leukemia cells treated with the flavonoid, casticin.

    Science.gov (United States)

    Righeschi, Chiara; Eichhorn, Tolga; Karioti, Anastasia; Bilia, Anna Rita; Efferth, Thomas

    2012-01-01

    Natural polyphenols play an important role in tumor inhibition. We used a doxorubicin-sensitive acute T-lymphoblastic leukemia cell line (CCRF-CEM) and its multidrug-resistant subline (CEM/ADR5000) to evaluate the activity of 15 plant polyphenols isolated in our laboratory (hypericin and pseudohypericin, verbascoside, ellagic acid, casticin, kaempferol-3-O-(2'',6''-di-E-p-coumaroyl)-glucopyranoside, kaempferol-3-O-(3,4-diacetyl-2,6-di-E-p-coumaroyl) -glucopyranoside, tiliroside, salvianolic acid B, oleuropein, rosmarinic acid, bergenin) or of others from commercial sources (curcumin, epigallocatechin-3-gallate, silymarin). Casticin was the most potent compound (IC50 values of 0.28 ± 0.02 μM in CCRF-CEM and 0.44 ± 0.17 μM in CEM/ADR5000 cells. The IC50 values of the other compounds tested ranged from 1.52 μM to 164.1 μM. A microarray-based mRNA expression profiling of CCRF-CEM cells treated with casticin was performed in order to identify genes with altered expression following casticin treatment. Networks related to NF-κB, p38MAPK, histones H3 and H4, and follicle stimulating hormone were identified.

  19. Dysferlin expression in monocytes: a source of mRNA for mutation analysis.

    Science.gov (United States)

    De Luna, N; Freixas, A; Gallano, P; Caselles, L; Rojas-García, R; Paradas, C; Nogales, G; Dominguez-Perles, R; Gonzalez-Quereda, L; Vílchez, J J; Márquez, C; Bautista, J; Guerrero, A; Salazar, J A; Pou, A; Illa, I; Gallardo, E

    2007-01-01

    Dysferlin protein is expressed in peripheral blood monocytes. The genomic analysis of the DYSF gene has proved to be time consuming because it has 55 exons. We designed a mutational screening strategy based on cDNA from monocytes to find out whether the mutational analysis could be performed in mRNA from a source less invasive than the muscle biopsy. We studied 34 patients from 23 families diagnosed with dysferlinopathy. The diagnosis was based on clinical findings and on the absence of protein expression using either immunohistochemistry or Western blot of skeletal muscle and/or monocytes. We identified 28 different mutations, 13 of which were novel. The DYSF mutations in both alleles were found in 30 patients and only in one allele in four. The results were confirmed using genomic DNA in 26/34 patients. This is the first report to furnish evidence of reliable mutational analysis using monocytes cDNA and constitutes a good alternative to genomic DNA analysis.

  20. Comparative mRNA Expression Profiles of Riboflavin Biosynthesis Genes in Lactobacilli Isolated from Human Feces and Fermented Bamboo Shoots

    Science.gov (United States)

    Thakur, Kiran; Tomar, Sudhir K.; Wei, Zhao-Jun

    2017-01-01

    With the aim to bioprospect potent riboflavin producing lactobacilli, the present study was carried out to evaluate the relative mRNA expression of riboflavin biosynthesis genes namely Rib 1, Rib 2, Rib 3, and Rib 4 from potent riboflavin producers obtained from our previous studies. All the four genes were successfully cloned and sequenced for further analysis by in silico procedures. As studied by non-denaturing Polyacrylamide gel electrophoresis, no difference in size of all the four genes among those of various lactobacilli was observed. The relative fold increase in mRNA expression in Rib 1, Rib 2, Rib 3, and Rib 4 genes has been observed to be 10-, 1-, 0.7-, and 8.5-fold, respectively. Due to increase in relative mRNA expression for all the Rib genes as well as phenotypic production attribute, KTLF1 strain was used further for expression studies in milk and whey. The fold increase in mRNA expression for all the four Rib genes was higher at 12 and 18 h in milk and whey respectively. After exposure to roseoflavin, resistant variant of KTLF1 showed considerable increase in expression of all the targets genes. This is the first ever study to compare the mRNA expression of riboflavin biosynthesis pathway genes in lactobacilli and it also under lines the effect of media and harvesting time which significantly affect the expression of rib genes. The use of roseoflavin-resistant strains capable of synthesizing riboflavin in milk and whey paves a way for an exciting and economically viable biotechnological approach to develop novel riboflavin bio-enriched functional foods. PMID:28367143

  1. Expression of angiotensin-converting enzyme mRNA gene in the kidneys of patients with glomerulonephrites

    Directory of Open Access Journals (Sweden)

    Alsayed Ahmed Alnahal

    2012-01-01

    Full Text Available A little is known about the behavior of the renin-angiotensin system (RAS in glomerulo-nephritis (GN, although it is activated in other models of injury. To study renal angiotensin-converting enzyme (ACE messenger ribonucleic acid (mRNA gene expression in patients with GN to determine its role in the disease process and other factors that may influence the course of the disease and the prognosis, e.g. treatment with ACE inhibitor (ACEI drugs, we studied 20 patients with GN allocated to two groups: ten patients received an ACEI drug and ten patients did not receive ACEI in addition to a control group of ten healthy subjects. Routine and special laboratory investigation, histopathological studies and quantitative polymerase chain reaction analysis for renal ACE mRNA were done for both the study and the control groups. There was a statistically significant increase in ACE mRNA gene expression in the GN groups than in control group, but no statistically significant difference in ACE mRNA gene expression between the patients group that received and the group that did not receive ACEI. A significant correlation was found between the ACE mRNA gene expression and the mean blood pressure, serum creatinine, blood urea nitrogen and 24-h urinary protein. In conclusion, a higher level of ACE mRNA gene expression in patients suffering from GN may suggest a role of the RAS in the process of GN, perhaps contributing to glomerular hypertrophy and matrix overproduction. The use of ACEI drugs possibly slows the rate of progression of renal failure and plays a role in controlling the pathophysiology.

  2. Expression of angiotensin-converting enzyme mRNA gene in the kidneys of patients with glomerulonephrites.

    Science.gov (United States)

    Alnahal, Alsayed Ahmed; Khalil, Usama Ahmed; Diab, Magada Alsayed; Zanaty, Ali Fahmy

    2012-09-01

    A little is known about the behavior of the renin-angiotensin system (RAS) in glomerulo-nephritis (GN), although it is activated in other models of injury. To study renal angiotensin-converting enzyme (ACE) messenger ribonucleic acid (mRNA) gene expression in patients with GN to determine its role in the disease process and other factors that may influence the course of the disease and the prognosis, e.g. treatment with ACE inhibitor (ACEI) drugs, we studied 20 patients with GN allocated to two groups: ten patients received an ACEI drug and ten patients did not receive ACEI in addition to a control group of ten healthy subjects. Routine and special laboratory investigation, histopathological studies and quantitative polymerase chain reaction analysis for renal ACE mRNA were done for both the study and the control groups. There was a statistically significant increase in ACE mRNA gene expression in the GN groups than in control group, but no statistically significant difference in ACE mRNA gene expression between the patients group that received and the group that did not receive ACEI. A significant correlation was found between the ACE mRNA gene expression and the mean blood pressure, serum creatinine, blood urea nitrogen and 24-h urinary protein. In conclusion, a higher level of ACE mRNA gene expression in patients suffering from GN may suggest a role of the RAS in the process of GN, perhaps contributing to glomerular hypertrophy and matrix overproduction. The use of ACEI drugs possibly slows the rate of progression of renal failure and plays a role in controlling the pathophysiology.

  3. Triptolide inhibits COX-2 expression by regulating mRNA stability in TNF-{alpha}-treated A549 cells

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Lixin; Zhang, Shuang; Jiang, Zhenzhou; Huang, Xin; Wang, Tao; Huang, Xiao; Li, Han [Jiangsu Center for Drug Screening, China Pharmaceutical University, Nanjing 210009 (China); Zhang, Luyong, E-mail: lyzhang@cpu.edu.cn [Jiangsu Center for Drug Screening, China Pharmaceutical University, Nanjing 210009 (China)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Triptolide inhibited COX-2 expression and the half-life of COX-2 mRNA is decreased. Black-Right-Pointing-Pointer The HuR protein shuttling from nucleus to cytoplasm is inhibited by triptolide. Black-Right-Pointing-Pointer Triptolide inhibited 3 Prime -UTR fluorescence reporter gene activity. Black-Right-Pointing-Pointer COX-2 mRNA binding to HuR is decreased by triptolide in pull-down experiments. -- Abstract: Cyclooxygenase-2 (COX-2) over-expression is frequently associated with human non-small-cell lung cancer (NSCLC) and involved in tumor proliferation, invasion, angiogenesis and resistance to apoptosis. In the present study, the effects of triptolide on COX-2 expression in A549 cells were investigated and triptolide was found to inhibit TNF-{alpha}-induced COX-2 expression. In our further studies, it was found that triptolide decreased the half-life of COX-2 mRNA dramatically and that it inhibited 3 Prime -untranslated region (3 Prime -UTR) fluorescence reporter gene activity. Meanwhile, triptolide inhibited the HuR shuttling from nucleus to cytoplasm. After triptolide treatment, decreased COX-2 mRNA in pull-down experiments with anti-HuR antibodies was observed, indicating that the decreased cytoplasmic HuR is responsible for the decreased COX-2 mRNA. Taken together, our results provided evidence for the first time that triptolide inhibited COX-2 expression by COX-2 mRNA stability modulation and post-transcriptional regulation. These results provide a novel mechanism of action for triptolide which may be important in the treatment of lung cancer.

  4. Selenium Deficiency Influences the mRNA Expression of Selenoproteins and Cytokines in Chicken Erythrocytes.

    Science.gov (United States)

    Luan, Yilin; Zhao, Jinxin; Yao, Haidong; Zhao, Xia; Fan, Ruifeng; Zhao, Wenchao; Zhang, Ziwei; Xu, Shiwen

    2016-06-01

    Selenium (Se) deficiency induces hemolysis in chickens, but the molecular mechanism for this effect remains unclear. Se primarily elicits its function through the activity of selenoproteins, which contain the unique amino acid selenocysteine (Sec). In this study, we aimed to investigate the effect of Se deficiency on the expression of 24 selenoproteins and 10 cytokines. One hundred eighty chickens were randomly divided into 2 groups (90 chickens per group). During the entire experimental period, chickens were allowed ad libitum consumption of feed and water. The chickens were fed either a Se-deficient diet (0.008 mg Se/kg; produced in the Se-deficient area of Heilongjiang, China) or a Se-supplemented diet (as sodium selenite) at 0.2 mg/kg for 35 days. At the 35th day, the messenger RNA (mRNA) levels of 24 selenoproteins and 10 cytokines were examined in erythrocytes of 5 chickens per group, and the correlation was analyzed. The results showed that the expression of 24 selenoproteins and 7 cytokines (IL-2, IL-4, IL-8, IL-10, IL-12β, TGF-β4, and IFN-γ) decreased (P cytokines (IL-1γ, IL-6 and IL-7) was higher in the Se-deficient group. In both groups, glutathione peroxidase (GPX), thioredoxin 1 (Txnrd1), selenoprotein P1 (SELP), and selenoprotein synthetase (SPS2) were highly expressed compared to the other selenoproteins in chicken erythrocytes (P cytokines (IL-1γ, IL-6, and IL-7) suggested that the immune system of chickens was damaged by the Se deficiency. Correlation analysis suggested that although the expression of 24 selenoproteins and 7 cytokines decreased and that of 3 cytokines increased, there was a close correlation between their expression levels and a Se diet. These results suggested that Se deficiency influenced the expressions of 24 selenoproteins and 10 cytokines in chicken erythrocytes, revealing a relationship between Se and the chicken immune system. This study offers information regarding the mechanism of Se deficiency-induced hemolysis.

  5. Assessment of potential biomarkers, metallothionein and vitellogenin mRNA expressions in various chemically exposed benthic Chironomus riparius larvae

    Science.gov (United States)

    Park, Kiyun; Kwak, Inn-Sil

    2012-12-01

    The objective of this study was conducted to identify the possibility of using Chironomus metallothionein (MT) and vitellogenin (VTG) as biomarkers of stress caused by endocrinedisrupting chemicals (EDCs), heavy metals, herbicides and veterinary antibiotics. We characterized the MT and VTG cDNA in Chironomus riparius and evaluated their mRNA expression profiles following exposure to different environmental pollutants. The gene expression analysis showed that the MT mRNA levels increased significantly after long-term exposure to cadmium (Cd), copper (Cu), Lead (Pb), di(2-ethylhexyl) phthalate (DEHP), and 2,4-dichlorophenoxyacetic acid (2,4-D). Moreover, the VTG mRNA expression increased significantly in C. riparius larvae exposed to BPA, NP, DEHP, Cd, 2,4-D and fenbendazole. Evaluation of the long-term effects of environmental pollutants revealed up regulation of Chironomus MT mRNA in response to DEHP exposure among EDCs, and the level of the VTG mRNA was increased significantly following treatment with Cd and herbicide 2,4-D at all concentrations in a dose-dependent manner. These results indicate that VTG could be used as a potential biomarker of herbicide and Cd as well as EDCs, while MT was a potential biomarker of heavy metals such as Cd, Cu, and Pb in aquatic environments.

  6. Expression of RANTES mRNA in skin lesions of feline eosinophilic plaque.

    Science.gov (United States)

    Kimura, Tomoe; Kano, Rui; Maeda, Sadatoshi; Tsujimoto, Hajime; Nagata, Masahiko; Hasegawa, Atsuhiko

    2003-10-01

    One of the mechanisms of eosinophil infiltration is its induction by chemoattractants such as regulated upon activation, normal T-expressed and secreted (RANTES) which is a cysteine-cysteine chemokine that mediates chemotaxis and activation of eosinophils in humans and mice. Skin lesions of feline eosinophilic plaque are characterized by a predominant infiltration of eosinophils. The mechanism(s) of eosinophilic infiltration in the skin and/or mucosa of cats is unknown. It is possible that RANTES is involved. To investigate the presence of RANTES in the skin of cats with eosinophilic plaques and nonaffected skin, we cloned and sequenced the full-length feline RANTES cDNA gene, in order to determine whether it is present in the skin of cats with eosinophilic plaques and/or if it is present in normal adjacent skin. We were able to document the the expression of RANTES mRNAs in skin with feline eosinophilic plaque as well as in normal cat skin. The full-length cDNA sequence of the RANTES gene (742 bp) contained a single open reading frame of 276 bp encoding a protein of 92 amino acids. The amino acid sequence of feline RANTES shared 67 and 74% sequence identity with that of bovine and mouse RANTES genes, respectively. RT-PCR analysis on RANTES mRNA in the skin of cats with eosinophilic plaque revealed that its expression was higher in the eosinophilic plaque skin lesions than in the normal skin. The result suggested that RANTES might play a role to induce eosinophil infiltration in feline eosinophilic plaque lesions.

  7. Radioinduced intestinal fibrosis: from molecular mechanisms to therapy applications. Contribution of the TGF--{beta}1, of the CTGF and of the transduction pathway of the Rho/ROCK signal; La fibrose intestinale radio-induite: des mecanismes moleculaires aux applications therapeutiques. Roles du TGF-{beta}1, du CTGF et de la voie de transduction du signal Rho/ROCK

    Energy Technology Data Exchange (ETDEWEB)

    Haydont, V

    2006-12-15

    Delayed radiation enteritis is an intestinal fibrosis induced by accidental or therapeutic radiation for pelvic and abdominal cancer treatments. Studies of molecular mechanisms involved in the development and maintenance of fibrosis have showed the respective contribution of CTGF, low TGF-{beta}1 concentrations and Rho/ROCK pathway. Thus, based on the relationship between CTGF, TGF-{beta}1 and Rho pathway, 2 therapeutics strategies have been develop. First, a pravastatin curative gift leads to a fibro-lysis involving an inhibition of Rho and in cascade a reduction of CTGF expression and extracellular matrix deposition. The data suggest that reversal of established radiation fibrosis in the gut is possible. Second, a pravastatin prophylactic gift prevents the installation of a chronic fibrosis but does not protect the tumor. On the base of these results, the radiation therapy department of the Institut Gustave Roussy will soon initiate 2 clinical trials. (author)

  8. Changes in the expression of estrogen receptor mRNA in the utero-vaginal junction containing sperm storage tubules in laying hens after repeated artificial insemination.

    Science.gov (United States)

    Das, Shubash Chandra; Nagasaka, Naohiro; Yoshimura, Yukinori

    2006-03-01

    The objective was to determine whether expression of estrogen receptor (ER) mRNA in the utero-vaginal junction (UVJ) of laying hens was altered after repeated artificial insemination (AI). Semi-quantitative RT-PCR was used to determine the expression of mRNA of the two types of receptor, ERalpha and ERbeta. Only ERalpha mRNA was expressed in all segments of the oviducts of both virgin and artificially inseminated birds, whereas ERbeta mRNA was expressed in ovarian follicles but not in the oviduct. The expression of ERalpha mRNA in the UVJ was significantly decreased after repeated AI, whereas that in the uterus was not significantly different between virgin and inseminated birds. Since estrogen may be involved in maintaining the sperm storage function of sperm storage tubules, the decreased expression of ERalpha mRNA in the UVJ after repeated AI may contribute to reduced fertility in these birds.

  9. Over-expression of corticotropin-releasing factor mRNA in inferior olivary neurons of rolling mouse Nagoya.

    Science.gov (United States)

    Sawada, Kazuhiko; Kawano, Michihiro; Tsuji, Hiroshi; Sakata-Haga, Hiromi; Hisano, Setsuji; Fukui, Yoshihiro

    2003-10-01

    Expression of corticotropin-releasing factor (CRF) mRNA was examined in the inferior olivary nucleus (ION) of an ataxic mutant, rolling mouse Nagoya (RMN) by semi-quantitative in situ hybridization. The most marked difference in the level of CRF mRNA signals between RMN and non-ataxic littermates (control mice) was observed in the beta-subnucleus and ventrolateral protrusion of the ION. The level of signals in these subnuclei was about twofold higher in RMN than in the controls. Signal levels in the dorsal nucleus, principal nucleus and subnucleus A were slightly but significantly higher in RMN than in the controls. In the other subnuclei, there were no differences in signal level between RMN and controls. These results suggest a region-related over-expression of CRF mRNA in the ION of RMN. This may be responsible for the increased sensitivity of some Purkinje cells to glutamate, resulting in ataxic symptoms of RMN.

  10. Expression of nerve growth factor mRNA in splenic lymphocytes of bronchial asthma rats and its influencing actors

    Institute of Scientific and Technical Information of China (English)

    Jihong Dai; Yonghong Wang; Haixia He

    2008-01-01

    BACKGROUND: Previous research has proved that nerve growth factor (NGF) participates in the onset of asthma by the induction of neurogenic inflammation.OBJECTIVE: To investigate the effect of interleukin-13 (IL-13) and interferon-γ(IFN-γ) on the expression of NGF mRNA in the splenic lymphocytes of bronchial asthma rats.DESIGN, TIME AND SETTING: The experiment, a completely randomized study based on cellular immunology, was performed in the Laboratory of Neurology in Chongqing Medical University and the Department of Clinical Pharmacy in College of Clinical Medicine, Chongqing Medical University (Chongqing, China) from January 2006 to April 2007.MATERIALS: Four adult male Wistar rats were used in this study. Rat IL-13, IFN-γprobe and the total RNA extraction kit were produced by Shanghai Sangon Biological Technology & Services Co., Ltd (China). The NGF ELISA kit was a product of Wuhan Boster Bioengineering Co., Ltd (China). A Du-70 automatic UV spectrophotometer was produced by Beckman Company (USA).METHODS: Rats were subjected to 1-mL intraperitoneal injections each containing 100 mg of ovalbumin, and were sensitized by using antigen solution, which was sensitized with 5×109 Bacillus pertussis and 100 mg aluminum hydroxide powder. Four rats were challenged with 1% ovalbumin using an ultrasonic nebulizer for 60 minutes to establish an asthmatic model. After rats were anesthetized, splenic lymphocytes were isolated and cultured in medium, which was supplemented with IL-13 or IFN-γ, for 0, 12, 24 or 48 hours. A parallel study was conducted with cultured splenic lymphocytes, which were divided into a control group, an IL-13 group and an IFN-γ group. Culture medium was added with different concentrations of IL-13 (10, 50, 100 μg/L) and IFN-γ (1, 10, 50 μg/L); 24 hours later, all samples were harvested.MAIN OUTCOME MEASURES: The expression levels of NGF mRNA were detected by reverse transcription-polymerase chain reaction.RESULTS: In the control group, the

  11. Effect of zinc on gene expression in osteoblastic MC3T3-E1 cells: enhancement of Runx2, OPG, and regucalcin mRNA expressions.

    Science.gov (United States)

    Yamaguchi, Masayoshi; Goto, Maya; Uchiyama, Satoshi; Nakagawa, Taeko

    2008-05-01

    The effect of zinc sulfate on the mRNA expressions in Runx2, osteocalcin, alpha1(I) collagen, insulin-like growth factor-I (IGF-I), transforming growth factor-beta1 (TGF-beta1), osteoprotegerin (OPG), regucalcin, zinc transporter 1 (ZIP1), or glyceroaldehyde-3-phosphate dehydrogenase (G3PDH) in osteoblastic MC3T3-E1 cells in vitro was investigated. Cells with subconfluency were cultured for 48 h in a medium containing either vehicle or zinc sulfate (10(-6)-10(-4) M) without fetal bovine serum. Culture with zinc sulfate (10(-5) M) caused a significant increase in Runx2, OPG, or regucalcin mRNA expressions in the cells, while it did not have a significant effect on osteocalcin, alpha1(I) collagen, IGF-I, TGF-beta1, ZIP1, or G3PDH mRNA expressions. The effect of zinc sulfate (10(-4) M) in increasing Runx2 mRNA expression was seen at 24-72 h after culture. A significant increase in OPG mRNA expression was observed at 24 or 48 h after culture. Regucalcin mRNA expression was significantly increased at 48 or 72 h after culture with zinc sulfate (10(-4) M). The stimulatory effects of zinc sulfate on Runx2, OPG, or regucalcin mRNAs were significantly prevented in the presence of cycloheximide (10(-7) M), an inhibitor of protein synthesis, or 5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole (10(-6) M), an inhibitor of transcription activity. Culture with beta-alanyl-L-histidinato zinc (10(-5) M) caused a significant increase in Runx2 or regucalcin mRNA expressions, while zinc acexamate (10(-5) M) did not have a significant effect on Runx2, OPG, ZIP1, or regucalcin mRNA expressions. This study demonstrates that zinc sulfate has a role in the enhancement of Runx2, OPG, or regucalcin mRNA expression in osteoblastic cells in vitro, suggesting its role in the regulation of gene expression in the cells.

  12. Diurnal expression of Dnmt3b mRNA in mouse liver is regulated by feeding and hepatic clockwork.

    Science.gov (United States)

    Maekawa, Fumihiko; Shimba, Shigeki; Takumi, Shota; Sano, Tomoharu; Suzuki, Takehiro; Bao, Jinhua; Ohwada, Mika; Ehara, Tatsuya; Ogawa, Yoshihiro; Nohara, Keiko

    2012-09-01

    DNA methyltransferase 3B (DNMT3B) is critically involved in de novo DNA methylation and genomic stability, while the regulatory mechanism in liver is largely unknown. We previously reported that diurnal variation occurs in the mRNA expression of Dnmt3b in adult mouse liver. The aim of this study was to determine the mechanism underlying the diurnal expression pattern. The highest level and the lowest level of Dnmt3b mRNA expression were confirmed to occur at dawn and in the afternoon, respectively, and the expression pattern of Dnmt3b closely coincided with that of Bmal1. Since the diurnal pattern of Dnmt3b mRNA expression developed at weaning and scheduled feeding to separate the feeding cycle from the light/dark cycle led to a phase-shift in the expression, it could be assumed that feeding plays a critical role as an entrainment signal. In liver-specific Bmal1 knockout (L-Bmal1 KO) mice, L-Bmal1 deficiency resulted in significantly higher levels of Dnmt3b at all measured time points, and the time when the expression was the lowest in wild-type mice was shifted to earlier. Investigation of global DNA methylation revealed a temporal decrease of 5-methyl-cytosine percentage in the genome of wild-type mice in late afternoon. By contrast, no such decrease in 5-methyl-cytosine percentage was detected in L-Bmal1 KO mice, suggesting that altered Dnmt3b expression affects the DNA methylation state. Taken together, the results suggest that the feeding and hepatic clockwork generated by the clock genes, including Bmal1, regulate the diurnal variation in Dnmt3b mRNA expression and the consequent dynamic changes in global DNA methylation.

  13. A cis-acting element known to block 3' mRNA degradation enhances expression of polyA-minus mRNA in wild-type yeast cells and phenocopies a ski mutant.

    Science.gov (United States)

    Brown, J T; Johnson, A W

    2001-11-01

    mRNA lacking a 3' polyA tail is not translated efficiently in wild-type eukaryotic cells, but is translated efficiently in yeast ski mutants. This enhanced expression could be due to altered translational specificity. However, as the SKI genes are required for 3' mRNA degradation, it could be a consequence of inhibition of 3' mRNA decay. Therefore, we asked if inhibition of 3' decay of a polyA-minus mRNA in cis would allow its efficient expression in wild-type cells. Capped in vitro reporter transcripts were prepared with or without a 3' cis-acting element known to inhibit 3' degradation (oligoG) and electroporated into yeast cells. The addition of oligoG to a polyA-minus mRNA enhanced expression 30-fold in wild-type cells. This level of expression was the same as that for an oligoG-minus, polyA-minus transcript in a ski mutant. The addition of oligoG did not significantly enhance the expression of polyA-minus mRNA in a ski mutant. The oligoG-dependent increase in expression was due to an increase in initial rate of translation and an increase in the functional half-life of the mRNA, similar to the effects observed in a ski mutant. The enhanced expression of the oligoG-containing RNA did not require Pab1p. We conclude that the enhanced translation of polyA-minus RNA in a ski mutant is due to inhibition of 3' mRNA degradation. Furthermore, a polyA-minus mRNA is expressed in wild-type cells when terminated in an element known to inhibit 3' decay in cis.

  14. Sorting live stem cells based on Sox2 mRNA expression.

    Directory of Open Access Journals (Sweden)

    Hans M Larsson

    Full Text Available While cell sorting usually relies on cell-surface protein markers, molecular beacons (MBs offer the potential to sort cells based on the presence of any expressed mRNA and in principle could be extremely useful to sort rare cell populations from primary isolates. We show here how stem cells can be purified from mixed cell populations by sorting based on MBs. Specifically, we designed molecular beacons targeting Sox2, a well-known stem cell marker for murine embryonic (mES and neural stem cells (NSC. One of our designed molecular beacons displayed an increase in fluorescence compared to a nonspecific molecular beacon both in vitro and in vivo when tested in mES and NSCs. We sorted Sox2-MB(+SSEA1(+ cells from a mixed population of 4-day retinoic acid-treated mES cells and effectively isolated live undifferentiated stem cells. Additionally, Sox2-MB(+ cells isolated from primary mouse brains were sorted and generated neurospheres with higher efficiency than Sox2-MB(- cells. These results demonstrate the utility of MBs for stem cell sorting in an mRNA-specific manner.

  15. Molecular cloning and mRNA expression analysis of sheep MYL3 and MYL4 genes.

    Science.gov (United States)

    Zhang, Chunlan; Wang, Jianmin; Wang, Guizhi; Ji, Zhibin; Hou, Lei; Liu, Zhaohua; Chao, Tianle

    2016-02-15

    Using longissimus dorsi muscles of Dorper sheep as the experimental materials, the complete cDNAs of ovine MYL3 (Myosin light chain 3) and MYL4 (Myosin light chain 4) genes were cloned using RT-PCR, 5' RACE and 3' RACE. We obtained 925-bp and 869-bp full-length cDNAs and submitted their sequences to GenBank as accession numbers of KJ710703 and KJ768855, respectively. The cDNAs contained 600-bp and 582-bp open reading frames (ORFs) and encoded proteins comprising 199 and 193 amino acid residues, respectively. Neither protein was predicted to have a signal peptide, but both were predicted to have several N-glycosylation, O-glycosylation, and phosphorylation sites. The secondary structures of MYL3 and MYL4 were predicted to be 40.70% and 48.70% α- helical, respectively. Sequence alignment showed that the MYL3 and MYL4 proteins of Ovis aries both shared more than 91% amino acid sequence similarity with those of Mus musculus, Homo sapiens, Rattus norvegicus, Bos taurus, and Sus scrofa. The levels of MYL3 and MYL4 mRNA in various sheep tissues were determined using qRT-PCR. The results showed that both mRNAs were highly expressed in the heart. This study has established a foundation for further investigation of the ovine MYL3 and MYL4 genes.

  16. mRNA differential display on gene expression in settlement metamorphosis process of Ruditapes philippinarum larvae

    Institute of Scientific and Technical Information of China (English)

    Lu Sumin; Bao Zhenmin; Hu Jingjie; Hu Xiaoli; Mu Chunhua; Fang Jianguang

    2008-01-01

    The mRNA differential display (DDRT-PCR) technique was adopted to find out the genes related to settlement metamorphosis development process of Ruditapes philippinarum larvae.In this study, we have obtained three hundred and forty-six amplification bands in total from pedivehger larvae, veliger larvae,eye spot larvae and post-larvae.Sixty-five out of three hundred and forty-six bands are distinctly differential display from band pattern, which can be put into four groups, standing for different expression characters.Sixteen differential display bands were cloned, sequenced and analyzed and nine different sequences are obtained in the study.Three sequences have higher similarity to the eDNAs deposited in database and three are very similar to the rDNA of other species, considered as the rDNA of Ruditapes philippinarum.The rest three sequences are found to be novel sequences after analyzed.Their accession numbers are AY916799, AY916798, and AY916797 respectively.We thought the novel sequences are possibly relevant to the early embryo development of Ruditapes philippinarum larvae and can provide some fundamental understandings that are helpful for the improvement of scallop seed raising industry.

  17. Changes in mRNA expression of arcuate nucleus appetite-regulating peptides during lactation in rats.

    Science.gov (United States)

    Suzuki, Yoshihiro; Nakahara, Keiko; Maruyama, Keisuke; Okame, Rieko; Ensho, Takuya; Inoue, Yoshiyuki; Murakami, Noboru

    2014-04-01

    The contribution of hypothalamic appetite-regulating peptides to further hyperphagia accompanying the course of lactation in rats was investigated by using PCR array and real-time PCR. Furthermore, changes in the mRNA expression for appetite-regulating peptides in the hypothalamic arcuate nucleus (ARC) were analyzed at all stages of pregnancy and lactation, and also after weaning. Food intake was significantly higher during pregnancy, lactation, and after weaning than during non-lactation periods. During lactation, ARC expression of mRNAs for agouti-related protein (AgRP) and peptide YY was increased, whereas that of mRNAs for proopiomelanocortin (POMC) and cholecystokinin (CCK) was decreased, in comparison with non-lactation periods. The increase in AgRP mRNA expression during lactation was especially marked. The plasma level of leptin was significantly decreased during the course of lactation, whereas that of acyl-ghrelin was unchanged. In addition, food intake was negatively correlated with the plasma leptin level during lactation. This study has clarified synchronous changes in the expression of many appetite-regulating peptides in ARC of rats during lactation. Our results suggest that hyperphagia during lactation in rats is caused by decreases in POMC and CCK expression and increases in AgRP expression in ARC, the latter being most notable. Together with the decrease in the blood leptin level, such changes in mRNA expression may explain the further hyperphagia accompanying the course of lactation.

  18. EXPRESSION OF PEDF mRNA AND PROTEIN IN NORMAL MOUSE RETINA AND EXPERIMENTAL CHOROIDAL NEO-VASCULARIZATION TISSUES

    Institute of Scientific and Technical Information of China (English)

    张雷; 王康孙; 王玲; 张士胜; 石海云

    2004-01-01

    Objective To study the expression of pigment epithelium derived factor ( PEDF) in normal mouse retina and experimental choroidal neovascularization (CNV) tissues. Methods CNV mouse models were induced by diode laser. The expression of PEDF mRNA and protein in normal mouse retina and CNV tissues were detected by in situ hybridization and immunohistochemical study. Results In normal mouse retina,PEDF mRNA was observed in the ganglion cell layer, inner nuclear layer and RPE cell layer, and PEDF protein was observed mainly in the nerve fiber layer, ganglion cell layer, photoreceptor cell layer and RPE cell layer, and lower level expression of PEDF protein was also observed in the inner plexiform layer and outer plexiform layer. In CNV tissues,the expression of PEDF mRNA and protein was also observed. 3d and 1 week after photocoagulation, the expression level of PEDF was relatively lower, and increased following the development of CNV. The level was the highest 2 weeks after photocoagulation, then decreased at 3 weeks. Conclusion PEDF was expressed in different layers of retina and was obviously expressed in the CNV tissues induced by laser photocoagulation. These findings suggest that PEDF may participate and modulate the development of CNV.

  19. Integrated Analysis of DNA Methylation and mRNA Expression Profiles to Identify Key Genes in Severe Oligozoospermia

    Directory of Open Access Journals (Sweden)

    Zhiming Li

    2017-05-01

    Full Text Available Severe oligozoospermia (SO is a complex disorder, whose etiology is the combined effect of genetic factors and epigenetic conditions. In this study, we examined DNA methylation and mRNA expression status in a set of testicular tissues of SO patients (n = 3, and compared methylated data with those derived from obstructive azoospermia (OA patients (n = 3 with normal spermatogenesis phenotype. We identified 1,960 differentially methylated CpG sites showing significant alterations in SO vs. OA using the Illumina Infinium HumanMethylation450 bead array. By integrating above DNA methylation data and mRNA expression results, we totally identified 72 methylated CpG sites located in 65 genes with anti-correlation between DNA methylation and mRNA expression. Integrated pathways analysis indicates that these genes are involved in response to hormone stimulus, activation of protein kinase activity, and apoptotic process, among others. We also observed some genes with inversely correlated difference is novel in male infertility field, including PTPRN2, EPHX1, SERPINB9, SLIT3, etc. Our results lay a groundwork for further biological study of SO. Moreover, we generated a workflow for integrated analysis of DNA methylation and mRNA expression, which is expandable to other study types.

  20. Endurance exercise induces mRNA expression of oxidative enzymes in human skeletal muscle late in recovery

    DEFF Research Database (Denmark)

    Leick, Lotte; Plomgaard, Peter S.; Grønløkke, L.

    2010-01-01

    exercise. To test the hypothesis that mRNA expression of many oxidative enzymes is up-regulated late in recovery (10-24 h) after exercise, male subjects (n=8) performed a 90-min cycling exercise (70% VO(2-max)), with muscle biopsies obtained before exercise (pre), and after 10, 18 and 24 h of recovery...

  1. Lipopolysaccharide-induced expression of IP-10 mRNA in rat brain and in cultured rat astrocytes and microglia

    NARCIS (Netherlands)

    Ren, LQ; Gourmala, N; Boddeke, HWGM; Gebicke-Haerter, PJ

    1998-01-01

    Using mRNA differential display technique, we have found a differentially expressed band in rat brain, designated HAP(2)G1, which was the strongest one induced in response to peripheral administration of lipopolysaccharide (LPS). Sequence analysis showed that HAP(2)G1 cDNA is the rat homologue of th

  2. PAX5О± and PAX5ОІ mRNA expression in breast Cancer: Relation ...

    African Journals Online (AJOL)

    Manal Basyouni Ahmed

    Background: Many studies evaluated the role of paired box gene 5 (PAX5) in breast cancer. ... mRNA expression of PAX5a and PAX5b in breast cancer and assessing ... the molecular alterations that contribute to disease initiation and ... are spliced to common coding sequences of exons 2–10 [12]. .... basis of breast cancer.

  3. Intragraft interleukin 2 mRNA expression during acute cellular rejection and left ventricular total wall thickness after heart transplantation

    NARCIS (Netherlands)

    de Groot-Kruseman, H A; Baan, C C; Hagman, E M; Mol, W M; Niesters, H G; Maat, A P; Zondervan, P E; Weimar, W; Balk, A H

    OBJECTIVE: To assess whether diastolic graft function is influenced by intragraft interleukin 2 (IL-2) messenger RNA (mRNA) expression in rejecting cardiac allografts. DESIGN: 16 recipients of cardiac allografts were monitored during the first three months after transplantation. The presence of IL-2

  4. FUCOIDIN INHIBITS OXIDIZED LOW DENSITY LIPOPROTEIN FROM INDUCING HUMAN PERIPHERAL BLOOD MONOCYTE EXPRESSION OF PROINFLAMMATORY CYTOKINES mRNA

    Institute of Scientific and Technical Information of China (English)

    雷新军; 马爱群; 任冰稳; 耿涛; 张葳; 白玲

    2003-01-01

    Objective To study the significance of scavenger receptor class A(SR-A)in mediating human peripheral blood monocyte to uptake oxidized low density lipoprotein(OxLDL) and promoting the atherosclerotic immuno-pathological lesion in the local blood vessel. Methods With the Digoxenin-labeled Oligonucleotide-probes In situ Hybridization, this research investigated the effects of OxLDL on the mRNA expression of proinflammatory cytokines including MCP-1, bFGF, PDGF and IL-10 in the human peripheral blood monocyte and whether fucoidin, a peculiarly inhibitory ligand for SR-A, would influence this process. Results Monocyte was significantly increased the mRNA expression of MCP-1, bFGF, PDGF and IL-10 in a dose-dependent manner after incubating with OxLDL (10,15,20,25,30·mg·L-1, respectively)for 24 hours(P<0.001). Fucoidin(50,100,150,200,250·mg·mL-1, respectively)completely inhibited OxLDL(20·mg·L-1)from inducing monocyte the mRNA expression of above proinflammatory cytokines(P<0.001). Conclusion OxLDL can stimulate human peripheral blood monocyte to give expression to proinflammatory cytokines mRNA in a dose-dependent manner, while a peculiarly inhibitory ligand for SR-A-fucoidin has an obviously opposed role.

  5. Expression of c-fos mRNA following moderate lateral fluid percussion brain injury in rats

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    AIM: To study the expression of c-fos mRNA in brain following moderate lateral fluid percussion brain injury in rats, and to observe the temporal patterns of its expression following percussion. METHODS: Male Sprague-Dawley rats were divided into normal control, sham operation control and injury group. The rats of injury group subjected to moderate lateral fluid percussion injury (0.2 mPa). The injury groups were then subdivided into 5 min, 15 min, 30 min, 1h, 2h groups according to the time elapsed after injury. The expression of c-fos mRNA was studied with reverse transcription polymerase chain reaction (RT-PCR) semi-quantitatively.RESULTS: At 5 min after percussion, the induction of c-fos mRNA was increased, and remained elevated up to 2 h after brain injury.CONCLUSION: The induction and expression of the c-fos mRNA in cortex and brain stem after fluid percussion brain injury were increased rapidly.

  6. REAL-TIME DETECTION OF SURVIVIN mRNA EXPRESSION IN CERVICAL CANCER CELL LINES USING MOLECULAR BEACON IMAGING

    Institute of Scientific and Technical Information of China (English)

    An Ruifang; He Dalin; Xue Yan; Wang Shu; Xie Li; Zhao Jun; Wang Xinyang; Yang Lili

    2006-01-01

    Objective To detect the expression of survivin mRNA in cervical cancer cell lines using molecular beacon imaging technology. Methods Human cervical cancer cells (HeLa and SiHa) and human fetal lung fibroblast HFL-I were cultured in vitro. After adding 100 nmol/L survivin mRNA molecular beacon, the fluorescent signals were observed under fluorescent microscope. The expressions of survivin in cervical cancer cells and HFL-I cell were examined by immunocytochemical streptravidin-biothin peroxidase (SP) assay at the same time. Results Two kinds of survivin mRNA molecular beacon, with different color fluorescence, had strong fluorescent signal in cervical cancer cell lines, and the signal in SiHa cell line was stronger, but these signals were not found in HFL-I ; Immunocytochemical staining of positive survivin was located in the cytoplasm of cervical cancer cell lines HeLa and SiHa, whereas, no expression of survivin was detected in HFL-I cell line. Conclusion The technology of molecular beacon imaging can be used to detect the expression of survivin mRNA in viable cells successfully, and may provide a new approach to the diagnosis of early stage cervical cancer and the following-up in the clinic.

  7. Expression and autoregulation of transforming growth factor beta receptor mRNA in small-cell lung cancer cell lines

    DEFF Research Database (Denmark)

    Nørgaard, P; Spang-Thomsen, M; Poulsen, H S

    1996-01-01

    In small-cell lung cancer cell lines resistance to growth inhibition by transforming growth factor (TGF)-beta 1, was previously shown to correlate with lack of TGF-beta receptor I (RI) and II (RII) proteins. To further investigate the role of these receptors, the expression of mRNA for RI, RII...

  8. Expression of mRNA coding voltage - gated sodium channel α-subunit in spontaneously epileptic rat

    Institute of Scientific and Technical Information of China (English)

    DUWa; CAIJi-Qun

    2004-01-01

    OBJECTIVE Subtypes Ⅰ,Ⅱ and Ⅲ of sodium channel α- subunit mRNA were analyzed in adult rat brain of spontaneously epileptic rats, and investigated the relationship between sodium channel expression and epilepsy. METHODS Tissue samples were microdissected from occipital neocortex, CA1 and CA3 hippocampus areas and dentate gyms, observe

  9. Serum leptin concentrations, leptin mRNA expression, and food intake during the estrous cycle in rats

    DEFF Research Database (Denmark)

    Fungfuang, Wirasak; Nakada, Tomoaki; Nakao, Nobuhiro

    2013-01-01

    in leptin mRNA expression in adipose tissue during the proestrous period compared with the diestrous period. These findings suggest that increased leptin mRNA expression and serum leptin levels, which are induced by estrogen during the proestrous stage, may play a role in regulating appetitive behavior.......The aim of this study was to investigate food intake, serum leptin levels, and leptin mRNA expression during the sexual cycle in rats. Female Wistar-Imamichi rats aged 8-10 weeks were used in this experiment. Food intake was measured during the light and dark phases (light on at 07:00 and off at 19......:00) of the 4-day estrous cycle in female rats. Serum leptin levels were measured by ELISA, and leptin mRNA expression levels were analyzed using real-time PCR on diestrous- and proestrous-stage rats. Our results revealed that during the sexual cycle, food intake was significantly higher in the dark phase...

  10. Aberrant Expression of TNF-α and TGF-β1 mRNA in Spontaneous Abortion

    Institute of Scientific and Technical Information of China (English)

    Ji-fen HU; Hong-chu BAO; Feng-chuan ZHU; Cai-ling YOU

    2004-01-01

    Objective To investigate the aberrant expressions of TNF-α and TGF-β1 in peripheral blood mononuclear cells (PBMCs) and placental tissues in patients with early spontaneous abortionMethods Using the technique of semi-quantitative reverse transcript-polymerase chain reaction (RT-PCR), TNF-α mRNA and TGF-β1 mRNA in PBMCs were measured in spontaneous abortion group (30 cases), normal pregnancy group (25 cases) and nonpregnant group (25 cases). The expressive intension of TNF-α protein and TGF-β1 protein in placental tissues was also identified by immunohistochemistry.Results Both levels of TNF-α mRNA and TGF-β1 mRNA expressed in PBMCs were significantly different between the three groups respectively (P<0. 05). Levels of TNF-α in syncytiotrophoblastic and cytotrophoblastic cells of the two aborted groups were substantially higher than those of the non-pregnant group (P<0. 01), but the levels of TGF-β1 in syncytiotrophoblastic cells of the two aborted groups were markedly lower than those of the non-pregnant group (P<0. 01).Conclusion There is potential relation between TGF-β1 at the fetomaternal interface and spontaneous abortion. TGF-β1 may contribute to the maintenance of pregnancy,and low-level expression of TGF-β1 may be associated with pregnancy failure.

  11. CDKN2A (p16) mRNA decreased expression is a marker of poor prognosis in malignant high-grade glioma.

    Science.gov (United States)

    Sibin, M K; Bhat, Dhananjaya I; Narasingarao, K V L; Lavanya, Ch; Chetan, G K

    2015-09-01

    Human high-grade glioma is heterogeneous in nature based on pathological and genetic profiling. Various tumour suppressor gene alterations are considered as prognostic markers in high-grade glioma. Gene expression of CDKN2A (p16) is used in various cancers as a prognostic biomarker along with methylation and deletion status of this gene. Expression levels of p16 mRNA were not studied as a biomarker in gliomas before. In this study, we have performed mRNA quantification analysis on 48 high-grade glioma tissues and checked for a possible prognostic role. The decreased expression of p16 mRNA in majority of the tumour tissues (57.1 %) was observed when compared to control tissues (P = 0.02). mRNA expression level was correlated with clinical variables also. p16 deletion status and BMI1 mRNA expression were also considered for comparison. p16 mRNA was negatively correlated with the BMI1 mRNA (P = p16 deletion. p16 mRNA expression, midline shift in MRI and tumour type were able to predict patient survival in overall survival (OS) and progression-free survival (PFS). p16 mRNA could independently predict prognosis of OS (P = 0.0146) and PFS (P = 0.0305) in multivariate analysis. We have shown that p16 mRNA expression can act as an independent prognostic biomarker in high-grade glioma.

  12. Role of CTGF in White Matter Development in Tuberous Sclerosis

    Science.gov (United States)

    2016-04-01

    specimens from TSC patients taken at the time of epilepsy surgery . We stained paraffin sections of tubers from TSC patients and detected that CTGF is...Ebrahimi-Fakhari D. Tuberous Sclerosis Complex. Pediatr Clin North Am. 2015 Jun;62(3):633-648 Davis PE, Peters JM, Krueger DA and Sahin M. Tuberous...Neural Circuitry of Autism / Seminar Hope Center for Neurological Disorders Washington University School of Medicine St. Louis, MO 2015 Targeted

  13. Influence of mRNA decay rates on the computational prediction of transcription rate profiles from gene expression profiles

    Indian Academy of Sciences (India)

    Chi-Fang Chin; Arthur Chun-Chieh Shih; Kuo-Chin Fan

    2007-12-01

    The abundance of an mRNA species depends not only on the transcription rate at which it is produced, but also on its decay rate, which determines how quickly it is degraded. Both transcription rate and decay rate are important factors in regulating gene expression. With the advance of the age of genomics, there are a considerable number of gene expression datasets, in which the expression profiles of tens of thousands of genes are often non-uniformly sampled. Recently, numerous studies have proposed to infer the regulatory networks from expression profiles. Nevertheless, how mRNA decay rates affect the computational prediction of transcription rate profiles from expression profiles has not been well studied. To understand the influences, we present a systematic method based on a gene dynamic regulation model by taking mRNA decay rates, expression profiles and transcription profiles into account. Generally speaking, an expression profile can be regarded as a representation of a biological condition. The rationale behind the concept is that the biological condition is reflected in the changing of gene expression profile. Basically, the biological condition is either associated to the cell cycle or associated to the environmental stresses. The expression profiles of genes that belong to the former, so-called cell cycle data, are characterized by periodicity, whereas the expression profiles of genes that belong to the latter, so-called condition-specific data, are characterized by a steep change after a specific time without periodicity. In this paper, we examine the systematic method on the simulated expression data as well as the real expression data including yeast cell cycle data and condition-specific data (glucose-limitation data). The results indicate that mRNA decay rates do not significantly influence the computational prediction of transcription-rate profiles for cell cycle data. On the contrary, the magnitudes and shapes of transcription-rate profiles for

  14. Effect of GLP-1 on the expression of NADPH oxidase subunits in the kidney of type 1 diabetic rats

    Directory of Open Access Journals (Sweden)

    Jin-jin LIU

    2013-09-01

    Full Text Available Objective To observe the effect of exenatide, a glucagon-like peptide-1 (GLP-1 receptor agonist, on the expression of NADPH oxidase subunits NOX4 and p22phox and connective tissue growth factor (CTGF in the kidney of streptozotocin (STZ-induced type 1 diabetic rats, and explore the protective effects and mechanisms of exenatide on the kidney of diabetic rats. Methods Thirty male Sprague-Dawley (SD rats were divided into control group (group A, n=7 and diabetic model group (n=23. Type 1 diabetic model was reproduced by intraperitoneal injection of streptozotocin. It was successful in 19 rats. Diabetic rats were randomly divided into diabetic control group (group B, n=10 and diabetic with treatment of exenatide group (group C, n=9. Rats in group C were injected subcutaneously with exenatide in dose of 5μg/kg twice daily. Rats in group A and B were given equivalent volume of normal saline by subcutaneous injection. All rats were sacrificed after eight weeks. The mRNA expression of renal p22phox and NOX4 were detected by real-time fluorescence quantitative PCR. The protein expression of CTGF was detected by immunohistochemical staining. Results The levels of blood glucose, lipids, creatinine, and urea nitrogen, the albumin excretion rate, kidney index, the mRNA expressions of renal NOX4 and p22phox, and the protein expression of renal CTGF were significantly increased in group B compared with that in group A (P0.05. Conclusion Exenatide can decrease the expressions of renal NOX4, p22phox and CTGF, decline the index of urinary protein, and alleviate the kidney hypertrophy in type 1 diabetic rats, implying that exenatide exerted a protective effect on the kidney.

  15. Effect of Selenium Supplementation on Activity and Mrna Expression of Type 1 Deiodinase in Mice With Excessive lodine Intake

    Institute of Scientific and Technical Information of China (English)

    XUE-FENG YANG; XIAO-HUI HOU; JIAN XU; HUAI-LAN GUO; CHEN-JIANG YING; XIAO-YI CHEN; XIU-FA SUN

    2006-01-01

    To investigate the effect of selenium supplementation on the selenium status and selenoenzyme, especially the activity and mRNA expression of type 1 deiodinase (D1) in mice with excessive iodine (EI) intake and to explore the mechanism of selenium intervention on iodine-induced abnormities. Methods Weanling female BALB/c mice were given tap water or 3 mg/L of iodine or supplemented with 0.5 mg/L or 1.0 mg/L of selenium in the presence of excessive iodine for 5months. Selenium status, thyroid hormone level, hepatic and renal D 1 activity and mRNA expression were examined. Results Excessive iodine intake significantly decreased the selenium concentration in urine and liver, and the activity of glutathione peroxidase (GSH-Px) in liver. Meanwhile, serum total T4 (TT4) increased while serum total T3 (TT3) decreased. Hepatic D1enzyme activity and mRNA expression were reduced by 33% and 86%, respectively. Renal D1 enzyme activity and mRNA were reduced by 30% and 55%, respectively. Selenium supplementation obviously increased selenium concentration, activity of GSH-Px and D1 as well as mRNA expression of D1. However, increasing the supplementation of Se from 0.5 to 1.0 mg/L did not further increase selenoenzyme activity and expression. Conclusion Relative selenium deficiency caused by excessive iodine plays an essential role in the mechanism of iodine-induced abnormalities. An appropriate dose of selenium supplementation exercises a beneficial intervention.

  16. Type 1 iodothyronine deiodinase activity and mRNA expression in rat thyroid tissue with different iodine intakes

    Institute of Scientific and Technical Information of China (English)

    WANG Kun; SUN Yi-na; LIU Jia-yu; YAN Yu-qin; CHEN Zu-pei

    2006-01-01

    Background Type 1 deiodinase (D1) plays an important role in the metabolism of thyroid hormone and has close relationship with thyroid function. In this study we explore the effects of iodine intake on D1 activity and its mRNA expression and its possible mechanism.Methods Forty-eight Wistar rats were randomly divided into six groups with 8 in each: low iodine (LI), normal iodine (NI), five-fold iodine (HI5), ten-fold iodine (HI10), fifty-fold iodine (HI50), one hundred-fold iodine (HI100)group. Three months, six months and twelve months after admistration of potassium iodate, they were sacrificed and thyroids were excised. The expression of D1 mRNA in the thyroid tissue was determined by RT-PCR and D1 activity was analyzed by 125I-rT3 as substrate. The thyroid hormone was measured with radioimmunoassay method.Results Compared with NI group, D1 mRNA expression in LI groups slightly decreased, and D1 activity greatly increased. Both T3 and T4 in thyroid tissue significantly decreased, but the T3/T4 ratio increased. D1 mRNA expression decreased in all HI groups, and D1 activity was significantly lower in HI groups. There was a tendency of decrease in D 1 activity with increased doses of iodine intakes. There was no significant difference in T4 in thyroid tissue between HI groups and NI group, but a tendency of decrease in T3 level was found in all HI groups.Conclusions In the case of iodine deficiency, D1 activity increased greatly in order to convert more T4 to T3.Excess iodine can inhibit both D1 mRNA expression and its activity to protect organism from being injured by excessive T3.

  17. Cis-regulatory variants affect CHRNA5 mRNA expression in populations of African and European ancestry.

    Directory of Open Access Journals (Sweden)

    Jen-Chyong Wang

    Full Text Available Variants within the gene cluster encoding α3, α5, and β4 nicotinic receptor subunits are major risk factors for substance dependence. The strongest impact on risk is associated with variation in the CHRNA5 gene, where at least two mechanisms are at work: amino acid variation and altered mRNA expression levels. The risk allele of the non-synonymous variant (rs16969968; D398N primarily occurs on the haplotype containing the low mRNA expression allele. In populations of European ancestry, there are approximately 50 highly correlated variants in the CHRNA5-CHRNA3-CHRNB4 gene cluster and the adjacent PSMA4 gene region that are associated with CHRNA5 mRNA levels. It is not clear which of these variants contribute to the changes in CHRNA5 transcript level. Because populations of African ancestry have reduced linkage disequilibrium among variants spanning this gene cluster, eQTL mapping in subjects of African ancestry could potentially aid in defining the functional variants that affect CHRNA5 mRNA levels. We performed quantitative allele specific gene expression using frontal cortices derived from 49 subjects of African ancestry and 111 subjects of European ancestry. This method measures allele-specific transcript levels in the same individual, which eliminates other biological variation that occurs when comparing expression levels between different samples. This analysis confirmed that substance dependence associated variants have a direct cis-regulatory effect on CHRNA5 transcript levels in human frontal cortices of African and European ancestry and identified 10 highly correlated variants, located in a 9 kb region, that are potential functional variants modifying CHRNA5 mRNA expression levels.

  18. Negative and positive mRNA splicing elements act competitively to regulate human immunodeficiency virus type 1 vif gene expression.

    Science.gov (United States)

    Exline, C M; Feng, Z; Stoltzfus, C M

    2008-04-01

    Over 40 different human immunodeficiency virus type 1 (HIV-1) mRNAs are produced by alternative splicing of the primary HIV-1 RNA transcripts. In addition, approximately half of the viral RNA remains unspliced and is used as genomic RNA and as mRNA for the Gag and Pol gene products. Regulation of splicing at the HIV-1 3' splice sites (3'ss) requires suboptimal polypyrimidine tracts, and positive or negative regulation occurs through the binding of cellular factors to cis-acting splicing regulatory elements. We have previously shown that splicing at HIV-1 3'ss A1, which produces single-spliced vif mRNA and promotes the inclusion of HIV exon 2 into both completely and incompletely spliced viral mRNAs, is increased by optimizing the 5' splice site (5'ss) downstream of exon 2 (5'ss D2). Here we show that the mutations within 5'ss D2 that are predicted to lower or increase the affinity of the 5'ss for U1 snRNP result in reduced or increased Vif expression, respectively. Splicing at 5'ss D2 was not necessary for the effect of 5'ss D2 on Vif expression. In addition, we have found that mutations of the GGGG motif proximal to the 5'ss D2 increase exon 2 inclusion and Vif expression. Finally, we report the presence of a novel exonic splicing enhancer (ESE) element within the 5'-proximal region of exon 2 that facilitates both exon inclusion and Vif expression. This ESE binds specifically to the cellular SR protein SRp75. Our results suggest that the 5'ss D2, the proximal GGGG silencer, and the ESE act competitively to determine the level of vif mRNA splicing and Vif expression. We propose that these positive and negative splicing elements act together to allow the accumulation of vif mRNA and unspliced HIV-1 mRNA, compatible with optimal virus replication.

  19. The mRNA expression profile of metabolic genes relative to MHC isoform pattern in human skeletal muscles.

    Science.gov (United States)

    Plomgaard, Peter; Penkowa, Milena; Leick, Lotte; Pedersen, Bente K; Saltin, Bengt; Pilegaard, Henriette

    2006-09-01

    The metabolic profile of rodent muscle is generally reflected in the myosin heavy chain (MHC) fiber-type composition. The present study was conducted to test the hypothesis that metabolic gene expression is not tightly coupled with MHC fiber-type composition for all genes in human skeletal muscle. Triceps brachii, vastus lateralis quadriceps, and soleus muscle biopsies were obtained from normally physically active, healthy, young male volunteers, because these muscles are characterized by different fiber-type compositions. As expected, citrate synthase and 3-hydroxyacyl dehydrogenase activity was more than twofold higher in soleus and vastus than in triceps. Contrary, phosphofructokinase and total lactate dehydrogenase (LDH) activity was approximately three- and twofold higher in triceps than in both soleus and vastus. Expression of metabolic genes was assessed by determining the mRNA content of a broad range of metabolic genes. The triceps muscle had two- to fivefold higher MHC IIa, phosphofructokinase, and LDH A mRNA content and two- to fourfold lower MHC I, lipoprotein lipase, CD36, hormone-sensitive lipase, and LDH B and hexokinase II mRNA than vastus lateralis or soleus. Interestingly, such mRNA differences were not evident for any of the genes encoding mitochondrial oxidative proteins, 3-hydroxyacyl dehydrogenase, carnitine palmitoyl transferase I, citrate synthase, alpha-ketogluterate dehydrogenase, and cytochrome c, nor for the transcriptional regulators peroxisome proliferator activator receptor gamma coactivator-1alpha, forkhead box O1, or peroxisome proliferator activator receptor-alpha. Thus the mRNA expression of genes encoding mitochondrial proteins and transcriptional regulators does not seem to be fiber type specific as the genes encoding glycolytic and lipid metabolism genes, which suggests that basal mRNA regulation of genes encoding mitochondrial proteins does not match the wide differences in mitochondrial content of these muscles.

  20. Myocardial connective tissue growth factor (CCN2/CTGF attenuates left ventricular remodeling after myocardial infarction.

    Directory of Open Access Journals (Sweden)

    Jørgen Gravning

    Full Text Available AIMS: Myocardial CCN2/CTGF is induced in heart failure of various etiologies. However, its role in the pathophysiology of left ventricular (LV remodeling after myocardial infarction (MI remains unresolved. The current study explores the role of CTGF in infarct healing and LV remodeling in an animal model and in patients admitted for acute ST-elevation MI. METHODS AND RESULTS: Transgenic mice with cardiac-restricted overexpression of CTGF (Tg-CTGF and non-transgenic littermate controls (NLC were subjected to permanent ligation of the left anterior descending coronary artery. Despite similar infarct size (area of infarction relative to area at risk 24 hours after ligation of the coronary artery in Tg-CTGF and NLC mice, Tg-CTGF mice disclosed smaller area of scar tissue, smaller increase of cardiac hypertrophy, and less LV dilatation and deterioration of LV function 4 weeks after MI. Tg-CTGF mice also revealed substantially reduced mortality after MI. Remote/peri-infarct tissue of Tg-CTGF mice contained reduced numbers of leucocytes, macrophages, and cells undergoing apoptosis as compared with NLC mice. In a cohort of patients with acute ST-elevation MI (n = 42 admitted to hospital for percutaneous coronary intervention (PCI serum-CTGF levels (s-CTGF were monitored and related to infarct size and LV function assessed by cardiac MRI. Increase in s-CTGF levels after MI was associated with reduced infarct size and improved LV ejection fraction one year after MI, as well as attenuated levels of CRP and GDF-15. CONCLUSION: Increased myocardial CTGF activities after MI are associated with attenuation of LV remodeling and improved LV function mediated by attenuation of inflammatory responses and inhibition of apoptosis.

  1. A case of cervical cancer expressed three mRNA variant of Hyaluronan-mediated motility receptor

    Science.gov (United States)

    Villegas-Ruíz, Vanessa; Salcedo, Mauricio; Zentella-Dehesa, Alejandro; de Oca, Edén V Montes; Román-Basaure, Edgar; Mantilla-Morales, Alejandra; Dávila-Borja, Víctor M; Juárez-Méndez, Sergio

    2014-01-01

    Cervical cancer is the second malignancy in Mexico, little is known about the prognostic factors associated with this disease. Several cellular components are important in their transformation and progression. Alternative mRNA splice is an important mechanism for generating protein diversity, nevertheless, in cancer unknown mRNA diversity is expressed. Hyaluronan-mediated motility receptor (HMMR, RHAMM, CD168) is a family member of proteins, hyaluronan acid dependent, and has been associated with different malignant processes such as: angiogenesis, cell invasiveness, proliferation, metastasis and poor outcome in some tumors. In the present study we identified expression of HMMR in cervical cancer by means of RT-PCR and sequencing. Our results indicate co-expression of two HMMR variants in all samples, and one case expressed three alternative HMMR splice transcripts. These results showed the heterogeneity of mRNA transcripts of HMMR that could express in cancer and the expression of HMMR could be marker of malignancy in CC. PMID:24966934

  2. Studies on SSTR2 mRNA expression and its correlation to steroid receptors in human benign and malignant breast lesions

    Institute of Scientific and Technical Information of China (English)

    ZENG Xizhi(曾希志); YAO Zhenxiang(姚榛祥)

    2002-01-01

    Objective:This sudy was designed to observe somatostatin receptor subtype 2 (SSTR2) Mrna expression, and investigate the correlations between SSTR2 Mrna expression and steroid receptors in benign and malignant lesions of the breast. Methods: A total of 23 breast carcinomas,16 mammary hyperplasia and 9 mammary adenofibroma samples were analysed. The SSTR2 Mrna expression was examined by in situ hybridization using multiphase oligoprobes.The ER and PR were detected by immunohistochemical staining. A computerized image analysis system was utilized to estimate the relative contents of SSTR2 Mrna. Results: The positive rates of expression (87.0%) and relative contents (0.47) of SSTR2 Mrna in breast cancer were higher than those in benign breast lesions(64%,0.26) respectively( P<0.05). SSTR2 Mrna expression was closely correlated with ER and PR in breast cancer( P<0. 05), A positive correlation between SSTR2 Mrna expression and ER was also found in benign breast lesions. Conclusions: SSTR2 Mrna expressed both in benign and in malignant breast lesions, but higher in malignant than in benign ones. There was a significant positive correlation of SSTR2 Mrna expression with ER or PR. The results suggest that conbined treatment with an antiestrogen and a somatostatin analogue for ER-positive breast cancer is feasible.

  3. Relationship Between the DPD and TS mRNA Expression and the Response to S-1-Based Chemotherapy and Prognosis in Patients with Advanced Gastric Cancer.

    Science.gov (United States)

    Shen, Xiao-Ming; Zhou, Chong; Lian, Lian; Li, Li-Qun; Li, Wei; Tao, Min

    2015-04-01

    The aim was to determine changes in dihydropyrimidine dehydrogenase (DPD) and thymidylate synthase (TS) mRNAs in the blood of advanced gastric cancer (AGC) patients to see whether these enzymes affected the patients' response to S-1-based chemotherapy and prognosis. For this purpose, pretreatment DPD/TS mRNA expressions were determined in 40 AGC patients using RT-PCR. The patients were then administered with S-1-based regimen (S-1 + cisplatin) and toxicities were recorded. The relationship between the DPD/TS mRNA expressions and the chemotherapy response, drug resistance, and prognosis was analyzed. The data show that DPD mRNA expression correlated significantly with Lauren type while TS mRNA expression correlated with distant metastasis. Patients with higher DPD and/or TS mRNA expression(s) showed poor response, while those with low DPD mRNA expression showed better response to the chemotherapy. Pooled analysis showed that the patients with low DPD/TS mRNA expressions had better therapeutic response. The incidence of bone marrow suppression, diarrhea, and oral mucositis was high in patients with low DPD mRNA expression. Median overall survival (OS) in 40 patients was 13.5 months. It was 17 months for low and 10 months for high DPD (P = 0.044) and TS mRNA expression (P = 0.047). Pooled analysis showed that the patients with both low DPD/TS mRNA expressions had longer OS (P = 0.001). In conclusion, the detection of DPD and/or TS mRNA expression can be used to predict the response to S-1-based chemotherapy, drug resistance, and prognosis in AGC patients as well as to help guide the individualized treatment of gastric cancer.

  4. Expression of CD134 and CXCR4 mRNA in term placentas from FIV-infected and control cats.

    Science.gov (United States)

    Scott, Veronica L; Burgess, Shane C; Shack, Leslie A; Lockett, Nikki N; Coats, Karen S

    2008-05-15

    Feline immunodeficiency virus (FIV) causes a natural infection of domestic cats that resembles HIV-1 in pathogenesis and disease progression. Feline AIDS is characterized by depression of the CD4+ T cell population and fatal opportunistic infections. Maternal-fetal transmission of FIV readily occurs under experimental conditions, resulting in infected viable kittens and resorbed or arrested fetal tissues. Although both FIV and HIV use the chemokine receptor CXCR4 as a co-receptor, FIV does not utilize CD4 as the primary receptor. Rather, CD134 (OX40), a T cell activation antigen and co-stimulatory molecule, is the primary receptor for FIV. We hypothesized that placental expression of CD134 and CXCR4 may render the placenta vulnerable to FIV infection, possibly facilitating efficient vertical transmission of FIV, and impact pregnancy outcome. The purpose of this project was to quantify the relative expression of CD134 and CXCR4 mRNA from the term placentas of three groups of cats: uninfected queens producing viable offspring, experimentally-infected queens producing only viable offspring, and experimentally-infected queens producing viable offspring among mostly non-viable fetuses. Total RNA was extracted from term placental tissues from all groups of cats. Real-time one-step reverse transcriptase-PCR was used to measure gene expression. The FIV receptors CD134 and CXCR4 were expressed in all late term feline placental tissues. Placentas from FIV-infected queens producing litters of only viable offspring expressed more CD134 and CXCR4 mRNA than those from uninfected queens, suggesting that infection may cause upregulation of the receptors. On the other hand, placentas from FIV-infected cats with non-successful pregnancies expressed similar levels of CD134 mRNA and slightly less CXCR4 mRNA than those from uninfected queens. Thus, it appears that cells expressing these receptors may play a role in pregnancy maintenance.

  5. Identification of HAVCR1 gene haplotypes associated with mRNA expression levels and susceptibility to autoimmune diseases.

    Science.gov (United States)

    García-Lozano, José Raúl; Abad, Cristina; Escalera, Ana; Torres, Belén; Fernández, Olga; García, Alicia; Sánchez-Román, Julio; Sabio, José-Mario; Ortego-Centeno, Norberto; Raya-Alvarez, Enrique; Núñez-Roldán, Antonio; Martín, Javier; González-Escribano, María Francisca

    2010-08-01

    Human HAVCR1 gene maps on 5q33.2, a region linked with susceptibility to allergic and autoimmune diseases. The aims of the present study were to define the haplotypes of HAVCR1 gene taking into account both HapMap Project SNP haplotypes and exon 4 variants, to investigate a possible relationship between these haplotypes and mRNA expression levels, and to assess whether HAVCR1 gene is involved in susceptibility to rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Genotyping of three ins/del variants in the exon 4 was performed by fragment length analysis. Five tag SNPs genotypes and mRNA levels were determined using TaqMan assays. We defined four major haplotypes in our population: the two major haplotypes (named haplotypes A and B) bear both the 5383_5397del variant and the two most common SNP sets found in the CEU population. Quantification analysis revealed that genotype B/B had the highest median of mRNA expression levels (vs. BX + XX, p < 0.0001). Additionally, frequency of the genotype BB was significantly higher in RA patients than in controls (12.3 vs. 5.9% in controls, p = 0.0046, p (c) = 0.014, OR = 2.23, 95% CI 1.23-4.10). Our results support a relationship between HAVCR1 haplotypes and mRNA expression levels, and suggest an association of this gene with autoimmune diseases.

  6. Role of ghrelin on testosterone secretion and the mRNA expression of androgen receptors in adult rat testis.

    Science.gov (United States)

    Wang, Lin; Fang, Fugui; Li, Yunsheng; Zhang, Yunhai; Pu, Yong; Zhang, XiaoYong

    2011-06-01

    The present study was designed to determine the effects of ghrelin on in vivo and in vitro secretion of testosterone (T) and the expression of androgen receptor (AR) mRNA in the adult rat testis. The distribution of growth hormone secretagogue receptors (GHS-R(1a)) in the testis was also investigated. GHS-R(1a) immunoreactivity presented mainly in Sertoli and Leydig cells, primary spermatocytes, and secondary spermatocytes. Adult rats that were intracerebroventricularly (i.c.v.) administrated different dosages (1 nmol and 3 nmol) of ghrelin could significantly inhibit the secretion of T. The experession of AR mRNA in the testis was also notably reduced with 3 nmol ghrelin. Additionaly, in vitro exposure of the Leydig cells to increasing concentrations of ghrelin resulted in no obvious changes of T secretion in the culture media and AR mRNA expression of Leydig cells. Overall, our data demonstrate that the i.c.v. injection of ghrelin plays a physiological role in T secretion and AR mRNA expression in the testis, further confirming the reproductive role of ghrelin.

  7. [CDS cloning and relationship between intramuscular fat content and mRNA expression of PID1 gene in pig].

    Science.gov (United States)

    Qian, Yuan; Zeng, Yong-Qing; Du, Jin-Fang; Cui, Jing-Xiang; Li, Hua; Chen, Qi-Mei; Song, Yi-Ping; Chen, Wei

    2010-11-01

    To explore the relationship between the expression of PID1 gene and fat deposition, we cloned CDS of PID1 from porcine fat and muscle tissues by RT-PCR using degenerate primers, and investigated expression of this gene in various tissues (i.e., liver, backfat, and muscle tissues) of different breeds (i.e., Yorkshire, Laiwu, and Lulai Black) by real-time fluorescence quantitative PCR. The results showed that 654 bp CDS of porcine PID1 was obtained by sequencing and was 93.88%, 66.94% and 88.07% identical to those of the human, rat, and Bos taurus, respectively. The expression of PID1 mRNA in various tissues and breeds, on the whole, tended to be liver > fat > muscle and Laiwu > Lulai Black > Yorkshire, respectively. For different breeds, PID1 mRNA abundance in liver had significant difference (P 0.05). For the three groups of Laiwu pigs with high (LWH), intermediate (LWI), and low IMF content (LWL), PID1 mRNA level was higher in liver tissue of LWH than that of LWL significantly (P PID1 mRNA abundance was not correlated with IMF in these three tissues of Laiwu breed, but it was positively correlated with IMF in the tissues of these three breeds (P PID1 may be related to fat deposition.

  8. Studies on Androgen Receptor mRNA expression in Pancreas, Hypothalamus and Ovary of Androgen Sterilized Rats

    Institute of Scientific and Technical Information of China (English)

    Li WANG; Jing-wen HOU; Li-min LU; Jin YU; Sui-qi GUI

    2004-01-01

    Objective To investigate the androgen receptor (AR) mRNA expression in pancreas,hypothalamus and ovary of androgen sterilized rats (ASR)Methods ASR model was established by subcutaneous injection of testosterone propionate to SD female rats at the age of 9 days. Around the age of 106 days (proestrus),all rats were killed, serum △ 4-andronestedione (△ 4-A), total testosterone (TT), free testosterone (FT), insulin (Ins) and C-peptide (C-P)were measured by radioimmunoassay (RIA). Total RNA in pancreas, hypothalamus and ovary were extracted and the amount of AR mRNA was quantitatedly analyzed by RT-PCR with single base mutant template as inner standard. Results Serum concentrations of△ 4-A, TT, FT, Ins and C-P in ASR model rats were significantly higher than those in control group (P<0. 05, P<0. 01). The expression of AR mRNA in pancreas, hypothalamus and ovary increased significantly (P<0. 05,P<0. 01) of model rats as compared with control group. Conclusion The elevated serum androgen levels in ASR model could enhance the expression of AR mRNA levels in pancreas, hypothalamus and ovary, which further induce hyperinsulinemia and anovulation.

  9. A structural approach to the role of CCN (CYR61/CTGF/NOV proteins in tumourigenesis

    Directory of Open Access Journals (Sweden)

    Perbal Bernard

    2003-08-01

    Full Text Available Abstract The CCN (CYR61 [Cystein-rich61]/CTGF [connective tissue growth factor]/NOV [Nephroblastoma overexpressed] proteins constitute a family of regulatory factors involved in many aspects of cell proliferation and differentiation. An increasing body of evidence indicates that abnormal expression of the CCN proteins is associated to tumourgenesis. The multimodular architecture of the CCN proteins, and the production of truncated isoforms in tumours, raise interesting questions regarding the participation of each individual module to the various biological properties of these proteins. In this article, we review the current data regarding the involvement of CCN proteins in tumourigenesis. We also attempt to provide structural basis for the stimulatory and inhibitory functions of the full length and truncated CCN proteins that are expressed in various tumour tissues.

  10. Expression of fragile X mental retardation protein and Fmr1 mRNA during folliculogenesis in the rat.

    Science.gov (United States)

    Ferder, Ianina; Parborell, Fernanda; Sundblad, Victoria; Chiauzzi, Violeta; Gómez, Karina; Charreau, Eduardo H; Tesone, Marta; Dain, Liliana

    2013-04-01

    Fragile X mental retardation protein (FMRP) belongs to a small family of RNA-binding proteins. Its absence or inactivity is responsible for fragile X syndrome, the most common cause of inherited mental retardation. Despite its ubiquitous expression, FMRP function and expression remain almost understudied in non-neuronal tissues, though previous studies on germline development during oogenesis may suggest a special function of this protein also in ovarian tissue. In addition, the well-documented association of FMR1 premutation state with fragile X-related premature ovarian insufficiency adds interest to the role of FMRP in ovarian physiology. The aim of the present work was to investigate the expression of Fmr1 mRNA and its protein, FMRP, at different stages of rat follicular development. By immunohistochemical studies we demonstrated FMRP expression in granulosa, theca and germ cells in all stages of follicular development. In addition, changes in Fmr1 expression, both at the protein and mRNA levels, were observed. FMRP levels increased upon follicular development while preantral and early antral follicles presented similar levels of Fmr1 transcripts with decreased expression in preovulatory follicles. These observations suggest that Fmr1 expression in the ovary is regulated at different and perhaps independent levels. In addition, our results show expression of at least four different isoforms of FMRP during all stages of follicular growth with expression patterns that differ from those observed in brain and testis. Our study shows a regulated expression of Fmr1, both at mRNA and protein levels, during rat follicular development.

  11. Inducible expression of Pisum sativum xyloglucan fucosyltransferase in the pea root cap meristem, and effects of antisense mRNA expression on root cap cell wall structural integrity.

    Science.gov (United States)

    Wen, Fushi; Celoy, Rhodesia M; Nguyen, Trang; Zeng, Weiqing; Keegstra, Kenneth; Immerzeel, Peter; Pauly, Markus; Hawes, Martha C

    2008-07-01

    Mitosis and cell wall synthesis in the legume root cap meristem can be induced and synchronized by the nondestructive removal of border cells from the cap periphery. Newly synthesized cells can be examined microscopically as they differentiate progressively during cap development, and ultimately detach as a new population of border cells. This system was used to demonstrate that Pisum sativum L. fucosyl transferase (PsFut1) mRNA expression is strongly expressed in root meristematic tissues, and is induced >2-fold during a 5-h period when mitosis in the root cap meristem is increased. Expression of PsFut1 antisense mRNA in pea hairy roots under the control of the CaMV35S promoter, which exhibits meristem localized expression in pea root caps, resulted in a 50-60% reduction in meristem localized endogenous PsFut1 mRNA expression measured using whole mount in situ hybridization. Changes in gross levels of cell wall fucosylated xyloglucan were not detected, but altered surface localization patterns were detected using whole mount immunolocalization with CCRC-M1, an antibody that recognizes fucosylated xyloglucan. Emerging hairy roots expressing antisense PsFut1 mRNA appeared normal macroscopically but scanning electron microscopy of tissues with altered CCRC-M1 localization patterns revealed wrinkled, collapsed cell surfaces. As individual border cells separated from the cap periphery, cell death occurred in correlation with extrusion of cellular contents through breaks in the wall.

  12. Alterations of organ histopathology and metallolhionein mRNA expression in silver barb, Puntius gonionotus during subchronic cadmium exposure

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Common silver barb, Puntius gonionotus exposed to the nominal concentration of 0.06 mg/L Cd for 60 d, were assessed for histopathological alterations (gills, liver and kidney), metal accumulation, and metallothionein (MT) mRNA expression. Fish exhibited pathological symptoms such as hypertrophy and hyperplasia of primary and secondary gill lamellae, vacuolization in hepatocytes, and prominent tubular and glomerular damage in the kidney. In addition, kidney accumulated the highest content of cadmium, more than gills and liver. Expression of MT mRNA was increased in both liver and kidney of treated fish. Hepatic MT levels remained high after fish were removed to Cd-free water. In contrast, MT expression in kidney was peaked after 28 d of treatment and drastically dropped when fish were removed to Cd-free water. The high concentrations of Cd in hepatic tissues indicated an accumulation site or permanent damage on this tissue.

  13. Effect of acute resistance exercise and sex on human patellar tendon structural and regulatory mRNA expression

    DEFF Research Database (Denmark)

    Sullivan, B.E.; Carroll, C.C.; Jemiolo, B.;

    2009-01-01

    (6 men and 6 women). Collagen type I, collagen type III, and MMP-2 were downregulated (P 0.05) 24 h after RE. All other genes remained unchanged (P > 0.05) after RE. Women had higher resting mRNA expression (P ... = 0.08) toward lower resting expression of MMP-3 than men. All other genes were not influenced (P > 0.05) by sex. Acute RE appears to stimulate a change in collagen type I, collagen type III, and MMP-2 gene regulation in the human patellar tendon. Sex influences the structural and regulatory m...... and mechanical properties, it is uncertain what structural and regulatory components contribute to these responses. We measured the mRNA expression of tendon's main fibrillar collagens (type I and type III) and the main proteoglycans (decorin, biglycan, fibromodulin, and versican) and the regulatory enzymes MMP...

  14. Expression of LL-37, Human beta Defensin-2, and CCR6 mRNA in Patients with Psoriasis Vulgaris

    Institute of Scientific and Technical Information of China (English)

    李东升; 李家文; 段逸群; 周小勇

    2004-01-01

    To investigate whether LL-37 and human beta defensin-2 (hBD-2) is related to the patients with psoriasis seldom having skin infections and explore the role of the two peptides and CCR6 (the receptor of hBD-2) in the pathogenesis of psoriasis, the expression levels of mRNA of LL-37, hBD-2, and CCR6 in skin lesions of patients with psoriasis vulgaris were detected by using RT-PCR. The results showed that the mRNA expression levels of the two peptides and CCR6 in psoriatic lesions all increased compared with the normal skin (P<0. 001). It was suggested that upregulated expression of LL-37 and hBD-2 might be the main reason that result in the the skin of patients with psoriasis being seldom infected, and the two peptides and CCR6 might play crucial roles in the pathogenesis of psoriasis.

  15. Expression of PIK3CA, PTEN mRNA and PIK3CA mutations in primary breast cancer

    DEFF Research Database (Denmark)

    Palimaru, Irina; Brügmann, Anja; Wium-Andersen, Marie Kim;

    2013-01-01

    tissue samples of breast carcinoma and normal breast tissue were obtained from 175 breast cancer patients at the time of primary surgery, of these 105 patients were lymph node positive. Expression of PIK3CA and PTEN mRNA was quantified with Quantitative Real Time PCR. Somatic mutations in exon 9 and exon......PURPOSE: High activity of the intracellular phosphatidylinositol-3 kinase (PI3K) pathway is common in breast cancer. Here, we explore differences in expression of important PI3K pathway regulators: the activator, phosphatidylinositol-3-kinase catalytic subunit alpha (PIK3CA), and the tumour...... suppressor, phosphatase and tensin homolog (PTEN), in breast carcinoma tissue and normal breast tissue. Furthermore, we examine whether expression of PIK3CA and PTEN mRNA and occurrence of PIK3CA mutations are associated with lymph node metastases in patients with primary breast cancer. METHODS: Paired...

  16. Myogenic, matrix and growth factor mRNA expression in human skeletal muscle: effect of contraction intensity and feeding

    DEFF Research Database (Denmark)

    Agergaard, Jakob; Reitelseder, Søren; Pedersen, T.G.

    2013-01-01

    exercise. No major differences were seen in atrophy-related genes between HL and LL resistance exercise. No changes were seen over 12-week training for any of the targets. CONCLUSIONS: Resistance exercise at LL and HL elevated the expression of genes involved in skeletal muscle hypertrophy, although......INTRODUCTION: We examined short-term (3-hour) and long-term (12-week) training effects after heavy load [HL; 70% 1RM] and light load (LL; 16% 1RM) exercise. METHODS: mRNA expression of genes involved in skeletal muscle remodeling were analyzed and muscle activity (EMG measurements) was measured....... RESULTS: Relative muscle activity differed between HL and LL resistance exercise, whereas median power frequency was even, suggesting an equal muscle-fiber-type recruitment distribution. mRNA expression of Myf6, myogenin, and p21 was mostly increased, and myostatin was mostly depressed by HL resistance...

  17. The potential lipolysis function of musclin and its mRNA expression ...

    African Journals Online (AJOL)

    PRECIOUS

    2009-11-16

    Nov 16, 2009 ... Musclin mRNA levels in adipose, muscle tissues and primary adipocytes were examined by quantitative PCR. The ... reduced fat mass in mice. Here, we ..... increased, suggesting that musclin might be involved in adipocyte ...

  18. Effects of arginine supplementation on splenocyte cytokine mRNA expression in rats with gut-derived sepsis

    Institute of Scientific and Technical Information of China (English)

    Huey-Fang Shang; Chun-Sen Hsu; Chiu-Li Yeh; Man-Hui Pai; Sung-Ling Yeh

    2005-01-01

    AIM: To investigate the effects of arginine (Arg)-enriched diets before sepsis and/or Arg-containing total parenteral nutrition (TPN) after sepsis or both on cytokine mRNA expression levels in splenocytes of rats with gut-derived sepsis.METHODS: Rats were assigned to four experimental groups. Groups 1 and 2 were fed with a semipurified diet, while groups 3 and 4 had part of the casein replaced by Arg which provided 2% of the total calories.After the rats were fed with these diets for 10 d, sepsis was induced by cecal ligation and puncture (CLP), at the same time an internal jugular vein was cannulated. All rats were maintained on TPN for 3 d. Groups 1 and 3were infused with conventional TPN, while groups 2 and 4 were supplemented with Arg which provided 2% of the total calories in the TPN solution. All rats were killed 3 d after CLP to examine their splenocyte subpopulation distribution and cytokine expression levels.RESULTS: Plasma interleukin (IL)-2, IL-4, tumor necrosis factor-α (TNF-α) and interferon (IFN-γ) were not detectable 3 d after CLP. There were no differences in the distributions of CD45Ra+, CD3+, CD4+, and CD8+cells in whole blood and splenocytes among the four groups. The splenocyte IL-2 mRNA expression in the Arg-supplemented groups was significantly higher than that in group 1. IL-4 mRNA expression in groups 3 and 4was significantly higher than that in groups 1 and 2. The mRNA expression of IL-10 and IFN-γ was significantly higher in group 4 than in the other three groups. There was no difference in TNF-α mRNA expression among the four groups.CONCLUSION: The influence of Arg on the whole blood and splenic lymphocyte subpopulation distribution is not obvious. However, Arg administration, especially before and after CLP, significantly enhances the mRNA expression levels of Th1 and Th2 cytokines in the spleen of rats with gut-derived sepsis.

  19. Differential patterns of synaptotagmin7 mRNA expression in rats with kainate- and pilocarpine-induced seizures.

    Directory of Open Access Journals (Sweden)

    Gordana Glavan

    Full Text Available Previous studies in rat models of neurodegenerative disorders have shown disregulation of striatal synaptotagmin7 mRNA. Here we explored the expression of synaptotagmin7 mRNA in the brains of rats with seizures triggered by the glutamatergic agonist kainate (10 mg/kg or by the muscarinic agonist pilocarpine (30 mg/kg in LiCl (3 mEq/kg pre-treated (24 h rats, in a time-course experiment (30 min-1 day. After kainate-induced seizures, synaptotagmin7 mRNA levels were transiently and uniformly increased throughout the dorsal and ventral striatum (accumbens at 8 and 12 h, but not at 24 h, followed at 24 h by somewhat variable upregulation within different parts of the cerebral cortex, amigdala and thalamic nuclei, the hippocampus and the lateral septum. By contrast, after LiCl/pilocarpine-induced seizures, there was a more prolonged increase of striatal Synaptotagmin7 mRNA levels (at 8, 12 and 24 h, but only in the ventromedial striatum, while in some other of the aforementioned brain regions there was a decline to below the basal levels. After systemic post-treatment with muscarinic antagonist scopolamine in a dose of 2 mg/kg the seizures were either extinguished or attenuated. In scopolamine post-treated animals with extinguished seizures the striatal synaptotagmin7 mRNA levels (at 12 h after the onset of seizures were not different from the levels in control animals without seizures, while in rats with attenuated seizures, the upregulation closely resembled kainate seizures-like pattern of striatal upregulation. In the dose of 1 mg/kg, scopolamine did not significantly affect the progression of pilocarpine-induced seizures or pilocarpine seizures-like pattern of striatal upregulation of synaptotagmin7 mRNA. In control experiments, equivalent doses of scopolamine per se did not affect the expression of synaptotagmin7 mRNA. We conclude that here described differential time course and pattern of synaptotagmin7 mRNA expression imply regional

  20. Fas mRNA expression and calcium influx change in H2O2-induced apoptotic hepatocytes in vitro

    Institute of Scientific and Technical Information of China (English)

    Qi-Ping Lu; Lei Tian

    2005-01-01

    AIM: To investigate the relationship between Fas gene expression and calcium influx change in peroxide-induced apoptotic hepatocytes and the possible molecular mechanism of Rxa in protecting hepatocytes.METHODS: Single-cell Fas mRNA expression in H2O2-exposed L02 hepatocytes with or without treatment of Rxa,an extract from an anti-peroxidant, Radix Salviae Miltiorrhizae,was determined by all-cell patch clamp and single-cell reverse transcriptase polymerase chain reaction (RT-PCR). Transient calcium influx change ([Ca2+]i) in the cells was evaluated with all-cell patch clamp micro-fluorescence single-cytosolic free Ca2+ concentration technique. Fas protein expression, early apoptotic index (annexin-V+) and cell membrane change inthe cells were evaluated by immunohistochemistry, flow cytometry (FCM) and scan electron microscopy respectively.RESULTS: In cells exposed to H2O2 for 2 h, the specific lane for Fas mRNA was vivid on electrophoresis, with increased Fas protein expression, [Ca2+]i (from 143.66±34.21 to 1115.28±227.16), annexin-V+ index (from 4.00±0.79 to 16.18±0.72) and membrane vesicle formation. However, in cells exposed to H2O2 but pre-treated with Rxa, there was no increase in Fas mRNA or protein expression and [Ca2+]i (103.56±28.92). Annexin-V+ index (8.92±1.44) was lower than the controls (P<0.01), and the cell membrane was intact.CONCLUSION: H2O2 induces apoptosis of L02 cells by increasing cytosolic [Ca2+]i, and inducing Fas mRNA and protein expression. Rxa protects the L02 cells from apoptosis through anti-peroxidation, inhibition of calcium overloading and prevention of the activation of cytosolic Fas signal pathway.

  1. Change in mRNA expression of sirtuin 1 and sirtuin 3 in cats fed on high fat diet.

    Science.gov (United States)

    Ishikawa, Shingo; Li, Gebin; Takemitsu, Hiroshi; Fujiwara, Megumi; Mori, Nobuko; Yamamoto, Ichiro; Arai, Toshiro

    2013-09-27

    Mammalian sirtuins are homologs to the yeast silent information regulator 2 (Sir2), which is an NAD-dependent deacetylase. Sirtuins are comprised of 7 proteins, and each has different target proteins. Sirtuin 1 (SIRT1) plays important roles in maintaining metabolic functions and immune responses, and SIRT3 protects cells from oxidative stress-induced cell death. Both SIRT1 and SIRT3 are regulated by metabolic status and aging. Hence, SIRT1 and SIRT3 have been researched in metabolic diseases, such as type 2 diabetes mellitus (DM), fatty liver, and heart diseases. Although these diseases have been increasing, there is little information about relation between the diseases and SIRT1 and SIRT3 in cats. Therefore we cloned SIRT1 and SIRT3 cDNA, examined mRNA expression in cat tissues, and investigated the changes in SIRT1 and SIRT3 mRNA expression in peripheral blood leukocyte of cats fed on HFD for 6 weeks. Cat SIRT1 and SIRT3 contained a catalytic core region and showed high sequence homology with other vertebrate SIRT1 (>61.3%) and SIRT3 (>65.9%) amino acids. Real-time polymerase chain reaction analyses revealed that high expression levels were observed in the liver and skeletal muscle for SIRT1 and in the heart for SIRT3 in cats. In addition, both cat SIRT1 and SIRT3 expression levels in the pancreas were different between individuals. Cat SIRT1 mRNA expression in peripheral blood leukocytes was significantly elevated in obese cats fed on HFD (P Cat SIRT1 and SIRT3 genes are highly conserved among vertebrates, and HFD feeding may be related to SIRT1 mRNA expression mechanisms in cat peripheral blood leukocytes.

  2. BDNF and trkB mRNA expression in the rat hippocampus following entorhinal cortex lesions.

    Science.gov (United States)

    Lapchak, P A; Araujo, D M; Hefti, F

    1993-02-01

    Quantitative in situ hybridization was used to determine whether the prevalence or topographical distribution of brain-derived neurotrophic factor (BDNF) or tyrosine receptor kinase (trk) B mRNA is altered in the hippocampal formation following lesions of excitatory afferents from the entorhinal cortex which provides an external source of innervation for the hippocampal formation. BDNF mRNA levels were not altered in the hippocampal formation up to 10 days following entorhinal cortex lesions (ECLs). The levels of mRNA coding for all known forms of trkB receptors also remained unchanged. The prevalence of the synaptic plasticity marker SNAP-25 mRNA was increased in the CA2 and CA3 pyramidal cell layers and the dentate gyrus by 6 days following ECLs and remained elevated at 10 days following ECLs. Our findings indicate that hippocampal neuron sprouting which occurs in response to ECLs is not the result of changes in the expression of the BDNF or trkB mRNA.

  3. Functional characterization of recombinant rat macrophage inflammatory protein-1 alpha and mRNA expression in pulmonary inflammation.

    Science.gov (United States)

    Shi, M M; Chong, I W; Long, N C; Love, J A; Godleski, J J; Paulauskis, J D

    1998-02-01

    Chemokines are important inflammatory mediators that function by activating and recruiting leukocytes to an inflamed tissue. We have recently cDNA cloned the rat chemokine macrophage inflammatory protein-1 alpha (MIP-1 alpha) (1). In the present study, we characterize the biological function of recombinant MIP-1 alpha protein and describe expression of its mRNA both in vitro and in a rat model of lung inflammation. In vitro rat rMIP-1 alpha protein was chemotactic for both polymorphonuclear leukocytes (PMNs) and macrophages with maximal activity at 50 nM for both cell types. In in vivo studies, we found that intratracheal instillation of 1 and 5 micrograms of rMIP-1 alpha resulted in a significant (P < 0.05) influx of cells, primarily monocytes/macrophages, into the airspace of the lungs after 6 h. Mean numbers of lavagable PMNs were not elevated significantly (P < 0.05) for either dose of MIP-1 alpha. As a model of inflammation, rats were intratracheally instilled with 0.1 mg/kg bacterial lipopolysaccharide (LPS). Bronchoalveolar lavage (BAL) was performed 3 h later. Instillation of LPS resulted in an acute neutrophilia, but no significant change in lavagable macrophages. BAL cells from control animals (saline instilled) displayed no basal mRNA expression of either MIP-1 alpha or MIP-2 (positive control). In contrast, both MIP-1 alpha and MIP-2 mRNA levels increased markedly in BAL cells from rats instilled with LPS. The rat alveolar macrophage cell line (NR8383) also showed increased MIP-1 alpha mRNA levels in response to LPS (10 micrograms/ml) with a maximal increase after 6-8 h. The induction of MIP-1 alpha mRNA expression by LPS in NR8383 cells was attenuated by cotreatment with the antioxidants N-acetylcysteine and dimethylsulfoxide, suggesting that the induction of MIP-1 alpha mRNA by LPS is mediated via the generation of reactive oxygen species. We conclude that MIP-1 alpha is a potent chemoattractant for macrophages in vivo, and its mRNA expression in

  4. Expression of connective tissue growth factor in male breast cancer: clinicopathologic correlations and prognostic value.

    Directory of Open Access Journals (Sweden)

    Miangela M Lacle

    Full Text Available Connective tissue growth factor (CTGF/CCN2 is a member of the CCN family of secreted proteins that are believed to play an important role in the development of neoplasia. In particular, CTGF has been reported to play an important role in mammary tumorigenesis and to have prognostic value in female breast cancer (FBC. The aim of the present study was to investigate clinicopathologic correlations and prognostic value of CTGF in male breast cancer (MBC and to compare these findings with FBC. For this, we studied CTGF protein expression by immunohistochemistry in 109 MBC cases and 75 FBC cases. In MBC, stromal CTGF expression was seen in the majority of the cases 78% (85/109 with high expression in 31/109 cases (28.4%, but expression in tumor cells was only seen in 9.2% (10/109 of cases. High stromal CTGF expression correlated with high grade and high proliferation index (>15% assessed by MIB-1 immunohistochemical staining. CTGF expression in tumor epithelial cells did not correlate with any of the clinicopathologic features. In FBC, stromal CTGF expression positively correlated with mitotic count and tumor CTGF expression was associated with triple negative status of the tumor (p = 0.002. Neither stromal nor tumor epithelial cell CTGF expression had prognostic value in MBC and FBC. In conclusion, stromal CTGF expression was seen in a high percentage of MBC and was correlated with high grade and high proliferation index. In view of the important role of the microenvironment in cancer progression, this might suggest that stromal CTGF could be an interesting target for novel therapies and molecular imaging. However, the lack of association with prognosis warrants caution. The potential role of CTGF as a therapeutic target for triple negative FBC deserves to be further studied.

  5. Influence of Olmesartan on Sirtuin 1 mRNA Expression in 5/6 Nephrectomized Spontaneously Hypertensive Rats.

    Science.gov (United States)

    Takata, Tomoaki; Munemura, Chishio; Fukui, Takeaki; Fukuda, Satoko; Murawaki, Yoshikazu

    2015-06-01

    Recent studies revealed that sirtuin 1 (SIRT1) has a relation to the mechanism of transforming growth factor-beta (TGF-beta) mediated apoptosis in glomerular mesangial cells and plays an important role in blood pressure regulation. It has been suggested that SIRT1 contributes to the renoprotective effect of angiotensin receptor blocker (ARB), but this has not yet become clearly recognized. In this study, we examined the relationship between SIRT1 and the therapeutic effect of olmesartan on renal injury in nephrectomized spontaneously hypertensive rats (SHRs). Male Wistar rats and 5/6 nephrectomized (5/6Nx) SHRs were assigned to 5 groups as follows: group A, Wistar rats; group B, Wistar rats administered high-dose olmesartan (15 mg/kg/day); group C, 5/6Nx SHRs; group D, 5/6Nx SHRs administered low-dose (3 mg/kg/day) olmesartan; and group E, 5/6Nx SHRs administered high-dose olmesartan. The drugs were administered for 12 weeks. Blood pressure and urinary protein excretion were measured every 4 weeks. Serum creatinine, glomerular sclerosis, SIRT1 mRNA level, TGF-beta mRNA level and klotho mRNA level were measured at the end of the examination. Systolic blood pressure, urinary protein excretion, serum creatinine and glomerular sclerosis in Wistar rats were significantly lower than that of 5/6Nx SHRs. Among 5/6Nx SHRs, high doses of olmesartan significantly decreased urinary protein excretion, serum creatinine and glomerular sclerosis compared to the non-treated and low-dose olmesartan groups. Expression of SIRT1 and klotho mRNA were significantly downregulated in 5/6Nx SHRs; however, olmesartan did not attribute to any change in gene expression. Expression of TGF-beta mRNA was significantly increased in 5/6Nx SHRs, and olmesartan did not affect the level of TGF-beta mRNA expression. Expression of SIRT1 is decreased in 5/6Nx SHRs compared to Wistar rats. Olmesartan suppressed glomerular sclerosis, but did not increase the expression of SIRT1, suggesting that the

  6. Small Nuclear RNAs U11 and U12 Modulate Expression of TNR-CFTR mRNA in Mammalian Kidneys

    Science.gov (United States)

    Souza-Menezes, Jackson; Tukaye, Deepali N.; Novaira, Horacio Javier; Guggino, William B.; Morales, Marcelo M.

    2008-01-01

    TNR-CFTR, discovered as a splice variant of CFTR (Cystic Fibrosis Transmembrane conductance Regulator), is distributed in different tissues such as human and rat kidney, trachea, lungs etc and is a functional chloride channel. In Kidneys, our findings show TNR-CFTR to have an unique distribution pattern with low levels of expression in renal cortex and high levels of expression in renal medulla. As shown by us previously, TNR-CFTR mRNA lacks 145 bp corresponding to segments of exons 13 and 14. This deletion causes a frame shift mutation leading to reading of a premature termination codon in exon 14. Premature termination of translation produces a functional half molecule of CFTR; TNR-CFTR. Our analysis of TNR mRNA has shown that the putative alternatively spliced intron has in its 5′ and 3′ conserved element CT and AC, respectively, that can be recognized by snRNAs U11 and U12. With these findings, we hypothesize that TNR-CFTR mRNA alternative splicing is probably mediate by splicing pathways utilizing U11 and U12 snRNAs. In this study, we have determined sequences of snRNAs U11 and U12 derived from rat kidney, which show significant homology to human U11 and U12 snRNAs. We show that there is significantly lower expression of U11 and U12 snRNAs in renal cortex compared to renal medulla in both humans and rats. This renal pattern of distribution of U11 and U12 snRNAs in both humans and rats closely follows distribution pattern of renal TNR-CFTR. Further, we have shown that blocking U11 and/or U12 mRNAs, by using antisense probes transfected in Immortalized Rat Proximal Tubule Cell line (IRPTC), decreases TNR-CFTR mRNA expression but not wild-type CFTR mRNA expression. Our results suggest that expression of U11 and/or U12 snRNAs is important for non-conventional alternative splicing process that gives rise to mRNA transcript coding for TNR-CFTR. PMID:18769035

  7. Luteotropic and luteolytic factors regulate mRNA and protein expression of progesterone receptor isoforms A and B in the bovine endometrium.

    Science.gov (United States)

    Rekawiecki, Robert; Kowalik, Magdalena Karolina; Kotwica, Jan

    2014-12-17

    The aim of the present study was to examine the effects of luteotropic and luteolytic factors on the mRNA and protein levels of progesterone receptor isoforms A (PGRA) and B (PGRB) in the bovine endometrium. Endometrial slices from Days 6-10 and 17-20 of the oestrous cycle were treated with LH (100ngmL-1), oestradiol (E2; 1×10-8M), prostaglandin (PG) E2 (1×10-6M) and PGF2? (1×10-6M) and the nitric oxide donor NONOate (1×10-4M); these treatments lasted for 6h for mRNA expression analysis and 24h for protein expression analysis. On Days 6-10 of the oestrous cycle PGRAB (PGRAB; the entire PGRA mRNA sequence is common to the PGRB mRNA sequence) mRNA expression in endometrial slices was enhanced by E2 treatment (PPGRB mRNA expression was increased by LH (PPPPGRAB mRNA expression increased after E2 (P2 (PPGRB mRNA expression was increased by PGE2 (P2? (PPPPPP2? (P2 (P2? (P<0.001). These data suggest that luteotropic and luteolytic factors affect PGRA and PGRB mRNA and protein levels, and this may regulate the effects of progesterone on endometrial cells.

  8. Effect of gamma radiation on the expression of mRNA growth factors in glycerol cryopreserved human amniotic membrane.

    Science.gov (United States)

    Yatim, Rusidah Mat; Kannan, Thirumulu Ponnuraj; Ab Hamid, Suzina Sheikh

    2016-12-01

    Human amniotic membrane (HAM) due to its high biocompatibility, low immunogenicity, anti-microbial, anti-viral properties as well as the presence of growth factors has been used in various clinical applications. The growth factors play an important role in wound healing. The current study aimed to explore the effect of 15 kGy gamma radiation dose on selected growth factors and receptors mRNA present in HAM. Eight growth factors, namely, EGF, HGF, KGF, TGF-α, TGF-β1, TGF-β2, TGF-β3 and bFGF and two growth factor receptors, HGFR and KGFR were evaluated in this study. The total RNA was extracted and converted to complimentary DNA using commercial kits. Subsequently, the mRNA expressions of these growth factors were evaluated using real-time PCR and the results were statistically analyzed using REST-MCS software. This study confirmed the presence of these mRNA growth factors and receptors in fresh, glycerol cryopreserved and irradiated glycerol cryopreserved HAM. In glycerol cryopreserved HAM, the results showed up-regulation of HGF and bFGF and down-regulation of EGF, HGFR, KGF, KGFR, TGF-α, TGF-β1, TGF-β2 and TGF-β3 relative to the fresh HAM which acted as the control, whereas in irradiated glycerol cryopreserved HAM, the results showed up-regulation of EGF, HGF, KGF, KGFR, TGF-β1, TGF-β2 and TGF-β3 and down-regulation of HGFR, TGF-α and bFGF relative to the glycerol cryopreserved HAM which acted as the control. However, these mRNA expressions did not show any statistical significant difference compared to the control groups. This study concluded that a dose of 15 kGy of gamma radiation did not affect the mRNA expression for the growth factors' and receptors' in the glycerol cryopreserved HAM.

  9. Using DNA sequencing electrophoresis compression artifacts as reporters of stable mRNA structures affecting gene expression.

    Science.gov (United States)

    Kapoor, Divya; Chandrayan, Sanjeev Kumar; Ahmed, Shubbir; Guptasarma, Purnananda

    2007-11-01

    The formation of secondary structure in oligonucleotide DNA is known to lead to "compression" artifacts in electropherograms produced through DNA sequencing. Separately, the formation of secondary structure in mRNA is known to suppress translation; in particular, when such structures form in a region covered by the ribosome either during, or shortly after, initiation of translation. Here, we demonstrate how a DNA sequencing compression artifact provides important clues to the location(s) of translation-suppressing secondary structural elements in mRNA. Our study involves an engineered version of a gene sourced from Rhodothermus marinus encoding an enzyme called Cel12A. We introduced this gene into Escherichia coli with the intention of overexpressing it, but found that it expressed extremely poorly. Intriguingly, the gene displayed a remarkable compression artifact during DNA sequencing electrophoresis. Selected "designer" silent mutations destroyed the artifact. They also simultaneously greatly enhanced the expression of the cel12A gene, presumably by destroying stable mRNA structures that otherwise suppress translation. We propose that this method of finding problem mRNA sequences is superior to software-based analyses, especially if combined with low-temperature CE.

  10. Lithium decreases VEGF mRNA expression in leukocytes of healthy subjects and patients with bipolar disorder.

    Science.gov (United States)

    Kikuchi, Kumiko; Iga, Jun-ichi; Tayoshi, Sumiko; Nakataki, Masahito; Watanabe, Shinya; Numata, Shusuke; Ohmori, Tetsuro

    2011-01-01

    Vascular endothelial growth factor (VEGF) is thought to be involved in the pathophysiology of mood disorders and the target of antidepressants. The aim of this study was to elucidate molecular effects of lithium on VEGF expression by using leukocytes of healthy subjects and patients with bipolar disorder. Eight healthy male subjects participated in the first study. Lithium was prescribed for 2 weeks, enough to reach therapeutic serum concentration. Leukocyte counts and serum lithium concentrations were determined at baseline, at 1- and 2-week medication, and at 2 weeks after stopping medication. VEGF mRNA levels were also examined in nine lithium-treated bipolar patients and healthy controls in the second study. In the first study, leukocyte counts were significantly increased at 2 weeks compared with those at baseline and were normalized after 2 weeks. VEGF mRNA levels were significantly decreased at 2 weeks and after 2 weeks compared with those at baseline. Consistent with the first study, VEGF mRNA levels were significantly decreased in the lithium-treated bipolar patients compared with healthy controls. Our investigation suggests that VEGF mRNA expression may be useful as a peripheral marker of the effects of lithium. Copyright © 2011 John Wiley & Sons, Ltd.

  11. NMDAR1 mRNA expression and glutamate receptor stimulated increase in cytosolic calcium concentration in rat and mouse cerebellar granule cells

    DEFF Research Database (Denmark)

    Mogensen, H S; Jørgensen, Ole Steen

    1996-01-01

    RNA and was normalized to that of the constitutively expressed H3.3 histone mRNA. The glutamate and NMDA stimulated increase in cytosolic Ca(2+)-ion concentration was measured using the fluorescent Ca(2+)-chelator Fluo3. In contrast to the hypothesis, we found NMDAR1 mRNA expression to be lower in mouse than in rat...

  12. Plasma cytokines do not reflect expression of pro- and anti-inflammatory cytokine mRNA at organ level after cardiopulmonary bypass in neonatal pigs

    DEFF Research Database (Denmark)

    Brix-Christensen, V.; Vestergaard, C.; Chew, M.

    2003-01-01

    Background: Plasma concentrations of inflammatory markers are increased in response to the trauma of cardiac surgery and cardiopulmonary bypass (CPB). It is, however, unknown whether the plasma cytokine levels and cytokine mRNA expression at organ level reflect each other. Methods: Twenty-six pig...... poorly reflected mRNA expression of the pro- and anti-inflammatory cytokines....

  13. Enhanced alpha 1(I) mRNA expression in frozen shoulder and dupuytren tissue.

    Science.gov (United States)

    Kilian, Olaf; Pfeil, U; Wenisch, S; Heiss, C; Kraus, R; Schnettler, R

    2007-12-14

    The purpose of this study has been to investigate collagen I and III synthesis during the fibrosing stage of frozen shoulder and Dupuytren samples in comparison to normal capsule tissue. - By using the quantitative PCR significantly increased levels of alpha 1(I) mRNA transcription in samples of frozen shoulder (p = 0.016) and Duypuytren (p = 0.041) could be demonstrated, whereas alpha 2(I) and alpha 1(III) chains have shown the same mRNA levels as in normal capsule tissue. - Despite an enhancement of alpha 1(I) mRNA transcription in frozen shoulder and Dupuytren samples the intracellular precursor procollagen I and extracellular mature collagen I was detected immunohistochemically in reduced levels. - The structural alteration of collagen I assembly might be caused by disturbed post-translation from the polypeptide chains into the triple helices procollagen I though alpha 1(I) mRNA transcription was significantly increased and alpha 2(I) mRNA transcription was in normal range. Fibroblasts might release high quantities of free alpha 1(I) polypeptide chains or (alpha 1(I)) 3 homotrimer into the extracellular space during the fibrosing stage of frozen shoulder and Dupuytren disease. - In all samples neither differences of alpha 1(III) mRNA transcription nor differences of immunohistochemical staining intensity of collagen III could be seen. This might result from apoptosis of myofibroblasts in the final phase of the fibrosing processes. - The stimulating effect of insulin-like growth factor type I (IGF-I) to induce fibrosis in connective tissue such as scarlet is known. In all patients suffering from frozen shoulder and Dupuytren disease the serum IGF-I level was in a normal range and the IGF-I receptor - (IGFR-I) mRNA transcription in the samples was also in the same level compared with normal capsule tissue.

  14. Effects of AFP gene silencing on Survivin mRNA expression inhibition in HepG2 cells.

    Science.gov (United States)

    Fang, Z L; Fang, N; Han, X N; Huang, G; Fu, X J; Xie, G S; Wang, N R; Xiong, J P

    2015-04-10

    We investigated the effects of alpha-fetoprotein (AFP) gene silencing on Survivin expression in HepG2 cells. Small interfering RNA technology was used to downregulate AFP expression in HepG2 cells. An enzyme-linked immunosorbent assay was used to measure AFP concentration in the supernatant before and after transfection. An MTT assay was used to detect cell proliferation activity before and after transfection. We performed flow cytometric analysis to detect the cell apoptosis rate, and reverse transcription-polymerase chain reaction to detect Survivin mRNA levels before and after transfection. Forty-eight hours after transfection, AFP concentration in the supernatant of the experimental group significantly decreased, hepatocellular carcinoma cell growth was inhibited by 43.1%, and the apoptosis rate increased by 24.3%. Survivin mRNA expression was reduced by 78.0% in HepG2 cells. These indicators in the control group and in the blank group did not change significantly. Silencing of AFP expression in HepG2 cells can effectively inhibit the growth of hepatoma cells and promote apoptosis, which may be useful for reducing intracellular Survivin mRNA levels.

  15. Freund's adjuvant-induced inflammation: clinical findings and its effect on hepcidin mRNA expression in horses

    Directory of Open Access Journals (Sweden)

    José P. Oliveira-Filho

    2014-01-01

    Full Text Available Hypoferremia observed during systemic inflammatory disorders is regulated by hepcidin. Hepcidin up-regulation is particularly important during acute inflammation, as it restricts the availability of iron, which is necessary for pathogenic microorganism growth before adaptive immunity occurs. The aim of this study was to evaluate the clinical findings and hepatic hepcidin mRNA expression in horses using a Freund's complete adjuvant (FCA model of inflammation. The expression of hepcidin mRNA in the liver was determined in healthy horses following two intramuscular injections of FCA at 0 h and 12 h. Plasma iron and fibrinogen concentrations were measured at multiple time points between 0 h and 240 h post-FCA injection (PI. Hepcidin mRNA expression was determined by RT-qPCR using liver biopsy samples performed at 0 h (control, 6 h and 18 h PI. The mean plasma fibrinogen level was significantly different from the control values only between 120 and 216 h PI. The mean plasma iron level was significantly lower than the control between 16 and 72 h PI, reaching the lowest levels at 30 h PI (33 % of the initial value, and returned to the reference value from 96 h PI to the end of the experiment. Hepcidin mRNA expression increased at 6 h PI and remained high at 18 h PI. The iron plasma concentration was an earlier indicator of inflammatory processes in horses when compared with fibrinogen and might be useful for the early detection of inflammation in the horse. FCA administration caused the rapid onset of hypoferremia, and this effect was likely the result of up-regulated hepatic hepcidin gene expression. This study emphasizes the importance of hepcidin and iron metabolism during inflammation in horses.

  16. SOCS3 and SOCS5 mRNA expressions may predict initial steroid response in nephrotic syndrome children

    Directory of Open Access Journals (Sweden)

    Witold Szaflarski

    2012-01-01

    Full Text Available Suppressors of Cytokine Signaling (SOCS inhibit Signal Transducers and Activators of Transcription (STATs phosphorylation by binding and inhibiting Janus Kinases (JaKs. The aim of the present study was to evaluate the influence of glucocorticosteroids on the JaK/STAT signaling pathway in the leukocytes of nephrotic syndrome (NS patients. The study group was composed of 34 steroid sensitive NS (SSNS children and 20 steroid resistant NS (SRNS subjects. Gene expression was assessed by real-time PCR using pre-designed human JaK/STAT PCR array. Protein expression was evaluated using ELISA assay (plasma concentration and immunofluorescence (in situ protein expression. In SSNS children, the initial increased expression of JaK1, JaK2, JaK3, STAT1, STAT2, STAT6, TYK2, SOCS1, SOCS2, SOCS3, SOCS4 and SOCS5 was reduced back to the control limits. Similarly, in SRNS patients the increased levels of almost all mRNA expressions for the abovementioned genes were decreased, with the exceptions of SOCS3 and SOCS5 expressions. These mRNA expressions were still significantly increased and correlated with early unfavorable course of nephrotic syndrome in children. Plasma levels of SOCS3, SOCS5, IL-6 and IL-20 were significantly increased in SRNS subjects after six weeks of steroids medication compared to SSNS and control participants. We conclude that SOCS3 and SOCS5 increased mRNA expressions might predict initial resistance to steroids in NS patients. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 4, pp. 719–728

  17. Endoplasmic reticulum-directed recombinant mRNA displays subcellular localization equal to endogenous mRNA during transient expression in CHO cells

    DEFF Research Database (Denmark)

    Beuchert Kallehauge, Thomas; Kol, Stefan; Andersen, Mikael Rørdam

    2016-01-01

    for this is the direction of the mRNA encoding the recombinant protein to the surface of the endoplasmic reticulum (ER) for subsequent protein translocation into the secretory pathway. To evaluate the efficiency of this process in Chinese hamster ovary (CHO) cells, the subcellular localization of recombinant mRNA encoding...... the therapeutic proteins, erythropoietin (EPO) and Rituximab, was determined. The results show that ER-directed recombinant mRNAs exhibited an efficient recruitment to the ER when compared to an endogenous ER-directed mRNA, with no cytoplasmic translation of ER-directed recombinant proteins observed....... These observations indicate that the recombinant mRNA, encoding ER-directed proteins, follows the same distribution pattern as endogenous mRNA directed towards the ER. Furthermore, the previous established fractionation method proves to be an efficient tool to study not only recombinant mRNA localization, but also...

  18. LDOC1 mRNA is differentially expressed in chronic lymphocytic leukemia and predicts overall survival in untreated patients

    Science.gov (United States)

    Duzkale, Hatice; Schweighofer, Carmen D.; Coombes, Kevin R.; Barron, Lynn L.; Ferrajoli, Alessandra; O'Brien, Susan; Wierda, William G.; Pfeifer, John; Majewski, Tadeusz; Czerniak, Bogdan A.; Jorgensen, Jeffrey L.; Medeiros, L. Jeffrey; Freireich, Emil J; Keating, Michael J.

    2011-01-01

    We previously identified LDOC1 as one of the most significantly differentially expressed genes in untreated chronic lymphocytic leukemia (CLL) patients with respect to the somatic mutation status of the immunoglobulin heavy-chain variable region genes. However, little is known about the normal function of LDOC1, its contribution to the pathophysiology of CLL, or its prognostic significance. In this study, we have investigated LDOC1 mRNA expression in a large cohort of untreated CLL patients, as well as in normal peripheral blood B-cell (NBC) subsets and primary B-cell lymphoma samples. We have confirmed that LDOC1 is dramatically down-regulated in mutated CLL cases compared with unmutated cases, and have identified a new splice variant, LDOC1S. We show that LDOC1 is expressed in NBC subsets (naive > memory), suggesting that it may play a role in normal B-cell development. It is also expressed in primary B-cell lymphoma samples, in which its expression is associated with somatic mutation status. In CLL, we show that high levels of LDOC1 correlate with biomarkers of poor prognosis, including cytogenetic markers, unmutated somatic mutation status, and ZAP70 expression. Finally, we demonstrate that LDOC1 mRNA expression is an excellent predictor of overall survival in untreated CLL patients. PMID:21310924

  19. The quantification of COMT mRNA in post mortem cerebellum tissue: diagnosis, genotype, methylation and expression

    Directory of Open Access Journals (Sweden)

    Craig Ian W

    2006-02-01

    Full Text Available Abstract Background The COMT gene is located on chromosome 22q11, a region strongly implicated in the aetiology of several psychiatric disorders, in particular schizophrenia. Previous research has suggested that activity and expression of COMT is altered in schizophrenia, and is mediated by one or more polymorphisms within the gene, including the functional Val158Met polymorphism. Method In this study we examined the expression levels of COMT mRNA using quantitative RT-PCR in 60 post mortem cerebellum samples derived from individuals with schizophrenia, bipolar disorder, depression, and no history of psychopathology. Furthermore, we have examined the methylation status of two CpG sites in the promoter region of the gene. Results We found no evidence of altered COMT expression or methylation in any of the psychiatric diagnoses examined. We did, however, find evidence to suggest that genotype is related to COMT gene expression, replicating the findings of two previous studies. Specifically, val158met (rs165688; Val allele rs737865 (G allele and rs165599 (G allele all showed reduced expression (P COMT expression, with females exhibiting significantly greater levels of COMT mRNA. Conclusion The expression of COMT does not appear to be altered in the cerebellum of individuals suffering from schizophrenia, bipolar disorder or depression, but does appear to be influenced by single nucleotide polymorphisms within the gene.

  20. Electroacupuncture-regulated neurotrophic factor mRNA expression in the substantia nigra of Parkinson’s disease rats

    Institute of Scientific and Technical Information of China (English)

    Shuju Wang; Jianqiao Fang; Jun Ma; Yanchun Wang; Shaorong Liang; Dan Zhou; Guojie Sun

    2013-01-01

    Acupuncture for the treatment of Parkinson's disease has a precise clinical outcome. This study investigated the effect of electroacupuncture at Fengfu (GV16) and Taichong (LR3) acupoints in rat models of Parkinson's disease induced by subcutaneous injection of rotenone into rat neck and back. Reverse transcription-PCR demonstrated that brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor mRNA expression was significantly increased in the substantia nigra of rat models of Parkinson's disease, and that abnormal behavior of rats was significantly improved following electroacupuncture treatment. These results indicated that electroacupuncture treatment upregulated brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor mRNA expression in the substantia nigra of rat models of Parkinson's disease. Thus, electroacupuncture may be useful in the treatment of Parkinson's disease.

  1. Electroacupuncture-regulated neurotrophic factor mRNA expression in the substantia nigra of Parkinson's disease rats.

    Science.gov (United States)

    Wang, Shuju; Fang, Jianqiao; Ma, Jun; Wang, Yanchun; Liang, Shaorong; Zhou, Dan; Sun, Guojie

    2013-02-25

    Acupuncture for the treatment of Parkinson's disease has a precise clinical outcome. This study investigated the effect of electroacupuncture at Fengfu (GV16) and Taichong (LR3) acupoints in rat models of Parkinson's disease induced by subcutaneous injection of rotenone into rat neck and back. Reverse transcription-PCR demonstrated that brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor mRNA expression was significantly increased in the substantia nigra of rat models of Parkinson's disease, and that abnormal behavior of rats was significantly improved following electroacupuncture treatment. These results indicated that electroacupuncture treatment upregulated brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor mRNA expression in the substantia nigra of rat models of Parkinson's disease. Thus, electroacupuncture may be useful in the treatment of Parkinson's disease.

  2. Estrogen and exercise interact to regulate brain-derived neurotrophic factor mRNA and protein expression in the hippocampus.

    Science.gov (United States)

    Berchtold, N C; Kesslak, J P; Pike, C J; Adlard, P A; Cotman, C W

    2001-12-01

    We investigated the possibility that estrogen and exercise interact in the hippocampus and regulate brain-derived neurotrophic factor (BDNF), a molecule increasingly recognized for its role in plasticity and neuron function. An important aspect of this study is to examine the effect of different time intervals between estrogen loss and estrogen replacement intervention. We demonstrate that in the intact female rat, physical activity increases hippocampal BDNF mRNA and protein levels. However, the exercise effect on BDNF up-regulation is reduced in the absence of estrogen, in a time-dependent manner. In addition, voluntary activity itself is stimulated by the presence of estrogen. In exercising animals, estrogen deprivation reduced voluntary activity levels, while estrogen replacement restored activity to normal levels. In sedentary animals, estrogen deprivation (ovariectomy) decreased baseline BDNF mRNA and protein, which were restored by estrogen replacement. Despite reduced activity levels in the ovariectomized condition, exercise increased BDNF mRNA levels in the hippocampus after short-term (3 weeks) estrogen deprivation. However, long-term estrogen-deprivation blunted the exercise effect. After 7 weeks of estrogen deprivation, exercise alone no longer affected either BDNF mRNA or protein levels. However, exercise in combination with long-term estrogen replacement increased BDNF protein above the effects of estrogen replacement alone. Interestingly, protein levels across all conditions correlated most closely with mRNA levels in the dentate gyrus, suggesting that expression of mRNA in this hippocampal region may be the major contributor to the hippocampal BDNF protein pool. The interaction of estrogen, physical activity and hippocampal BDNF is likely to be an important issue for maintenance of brain health, plasticity and general well-being, particularly in women.

  3. HCV core protein promotes liver fibrogenesis via up-regulation of CTGF with TGF-beta1.

    Science.gov (United States)

    Shin, Ju Yeop; Hur, Wonhee; Wang, Jin Sang; Jang, Jeong Won; Kim, Chang Wook; Bae, Si Hyun; Jang, Sung Key; Yang, Se-Hwan; Sung, Young Chul; Kwon, Oh-Joo; Yoon, Seung Kew

    2005-04-30

    Liver cirrhosis is one of the major complications of hepatitis C virus (HCV) infection, but the mechanisms underlying HCV-related fibrogenesis are still not clear. Although the roles of HCV core protein remain poorly understood, it is supposed to play an important role in the regulation of cellular growth and hepatocarcinogenesis. The aim of this study was to examine the role of HCV core protein on the hepatic fibrogenesis. We established an in vitro co-culture system with primary hepatic stellate cell (HSC) isolated from rats, and a stable HepG2-HCV core cell line which had been transfected with HCV core gene. The expressions of fibrosis-related molecules transforming growth factor beta1 (TGF-beta1), transforming growth factor beta receptor II (TGFbetaRII), alpha-smooth muscle actin (alpha-SMA) and connective tissue growth factor (CTGF) were analyzed via histological or molecular methods. In addition, the expression levels of matrix metaloprotinase-2 (MMP-2) and collagen type I (Col I) from the co-cultured media were measured by zymogram and ELISA, respectively. The expressions of alpha-SMA, TGF-beta1, Col I, TGFbetaRII and MMP-2 were significantly increased in the co-culture of stable HepG2-HCV core with HSC. Moreover, the significant increases of CTGF and TGF-beta1 in the HCV core-expressing cells were observed by either Northern or Western blot analysis. These results suggest that HCV core protein may contribute to the hepatic fibrogenesis via up-regulation of CTGF and TGF-beta1.

  4. Changes of bcl—XL and bax mRNA expression following traumatic brain injury in rats

    Institute of Scientific and Technical Information of China (English)

    骆纯; 卢亦成; 等

    2002-01-01

    Objective:To investigate the changes of bcl-2 gene family and the molecular mechanism of neuromal apoptosis following traumatic brain injury(TBI)in rats.Methods:Male Sprague-Dawley(SD)rats were subjected to lateral fluid percussion brain injury(FPBI)of moderate severity.Thebcl-XLand baxmRNA expression was detected by reverse transcription polymerase chain reaction(RT-PCR)expression of the impact site sas significantly lower(67.42%±7.54)than that of the ipsilateral hemisphere at 6hours after injury(P<0.01).The decrease of bcl-XLmRNA expression preceded apoptosis at 24 hours after injury.The bax mRNA expression rose slowly,doubled at 3days after injury and returned to the sham level slowly.Conclusions:Decreased expression of bcl-XLmRNA and increased expression of bax mRNA coincides tith apoptosis followwin brain injury.The bcl-2gene family is involved in neuronal apoptosis after TBI,and the changes of mRNA expression of the family members lead the neuronal cells to apoptosis.

  5. Promoter Methylation and Relative mRNA Expression of the p16 Gene in Cervical Cancer in North Indians.

    Science.gov (United States)

    Gupta, Amita; Ahmad, Mohammad Kaleem; Mahndi, Abbas Ali; Singh, Renu; Pradeep, Yashodhara

    2016-01-01

    Cervical carcinoma is one of the main causes of mortality in women worldwide as well as in India. It occurs as a result of various molecular events that develop from the combined influences of an individual's genetic predisposition and external agents such as smoking and menstrual hygiene, for example. However, infection with human papillomavirus (HPV) is the established major risk factor. The aim of the current study was to investigate p16 CpG island methylation and establish any correlation with mRNA expression in a north Indian population. We analyzed 196 woman volunteers out of which 98 were cases and 98 healthy controls. For the analysis of methylation pattern, DNA extracted from blood samples was modified with a bisulfate kit and used as template for methylation specific PCR (MSP). Quantitative real-time PCR (QRT-PCR) was performed to check mRNA expression. Correlation between methylation status of p16 gene and poor menstrual hygiene was significant (p=0.006), high parity cases showed methylation of p16 gene (p=0.031) with increased risk up to 1.86 times for cervical cancer and smoking was a strong risk factor associated with cervical cancer. We analyzed methylation pattern and found 60.3% methylation in cases with low mRNA expression level (0.014) as compared to controls (1.24). It was also observed that promoter methylation of p16 gene was significantly greater in FIGO stage III. We conclude that p16 methylation plays an important role in cervical cancer in the north Indian population and its methylation decreases mRNA expression. It can be used as an important and consistent blood biomarker in cervical cancer patients.

  6. ESTRADIOL IN FEMALES MAY NEGATE SKELETAL MUSCLE MYOSTATIN MRNA EXPRESSION AND SERUM MYOSTATIN PROPEPTIDE LEVELS AFTER ECCENTRIC MUSCLE CONTRACTIONS

    Directory of Open Access Journals (Sweden)

    Darryn S. Willoughby

    2006-12-01

    Full Text Available Eccentric contractions produce a significant degree of inflammation and muscle injury that may increase the expression of myostatin. Due to its anti- oxidant and anti-flammatory effects, circulating 17-β estradiol (E2 may attenuate myostatin expression. Eight males and eight females performed 7 sets of 10 reps of eccentric contractions of the knee extensors at 150% 1-RM. Each female performed the eccentric exercise bout on a day that fell within her mid-luteal phase (d 21-23 of her 28-d cycle. Blood and muscle samples were obtained before and 6 and 24 h after exercise, while additional blood samples were obtained at 48 and 72 h after exercise. Serum E2 and myostatin LAP/propeptide (LAP/pro levels were determined with ELISA, and myostatin mRNA expression determined using RT-PCR. Data were analyzed with two-way ANOVA and bivariate correlations (p 0.05. Compared to pre-exercise, males had significant increases (p < 0.05 in LAP/propetide and mRNA of 78% and 28%, respectively, at 24 h post-exercise, whereas females underwent respective decreases of 10% and 21%. E2 and LAP/propeptide were correlated at 6 h (r = -0.804, p = 0.016 and 24 h post- exercise (r = -0.841, p = 0.009 in males, whereas in females E2 levels were correlated to myostatin mRNA at 6 h (r =0.739, p = 0.036 and 24 h (r = 0.813, p = 0.014 post-exercise and LAP/propeptide at 6 h (r = 0.713, p = 0.047 and 24 h (r = 0.735, p = 0.038. In females, myostatin mRNA expression and serum LAP/propeptide levels do not appear to be significantly up-regulated following eccentric exercise, and may be due to higher levels of circulating E2

  7. Effect of Heat Stress on the Expression of GABA Receptor mRNA in the HPG Axis of Wenchang Chickens

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    LJ Xie

    Full Text Available ABSTRACT We investigated the effect of heat stress (HS on the expression of the GABA receptor in the hypothalamic-pituitary-gonadal (HPG axis of Wenchang chickens. Real-time quantitative RT-PCR (qRT-PCR was used to quantify the GABA receptor mRNA levels along the HPG axis of chickens under HS (40±0.5 °C for 1-6 weeks. Our results showed that the expression of GABAA and GABAB receptor at the mRNAs levels in the tissues of HPG axis exhibited fluctuation and variability. After HS, the mRNA level of GABAA receptor was significantly reduced in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but significantly increased in the pituitary of 1-, 4-, and 5-week-old chickens. The GABAB receptor mRNA level significantly declined in the hypothalamus of 1-week-old and in the pituitary of 3-week-old chickens, but was significantly upregulated in the pituitary and testis of 1- and 2-week-old chickens. At other time points, the expressions of GABAA receptor and GABAB receptor showed no significant differences compared with control group. These results indicated that the levels of GABAA receptor and GABAB receptor mRNAs varied in different tissues of the HPG axis in chickens of different ages, displaying temporal and spatial variations. GABA receptor behaved as a positively-regulated gene by HS, i.e., its mRNA was increased by HS; similarly, it was a negatively-regulated gene by HS, when its expression was reduced by HS.

  8. Changes in mRNA expression of arcuate nucleus appetite-regulating peptides during lactation in rats

    OpenAIRE

    Suzuki, Yoshihiro; Nakahara, Keiko; MARUYAMA, Keisuke; OKAME, Rieko; Ensho, Takuya; Inoue, Yoshiyuki; Murakami, Noboru

    2014-01-01

    The contribution of hypothalamic appetite-regulating peptides to further hyperphagia accompanying the course of lactation in rats was investigated by using PCR array and real-time PCR. Furthermore, changes in the mRNA expression for appetite-regulating peptides in the hypothalamic arcuate nucleus (ARC) were analyzed at all stages of pregnancy and lactation, and also after weaning. Food intake was significantly higher during pregnancy, lactation, and after weaning than during non-lactation per...

  9. Analysis of p130 protein and mRNA expression in ten patients with uterine papillary serous carcinoma

    Directory of Open Access Journals (Sweden)

    Shao-ting XU

    2011-11-01

    Full Text Available Objective To examine p130 protein and mRNA expression in uterine papillary serous carcinoma(UPSC and their clinical and pathologic significance.Methods A total of 10 UPSC patients(Stage I were included,with 10 cases of high-level endometrial carcinoma of the same stage taken as the control group and 10 cases of normal proliferative stage endometrium(EM taken as the disease control group.The level of p130 protein expression was determined by hematoxylin and eosin staining,microscopic observation,and immunohistochemistry,whereas the p130 mRNA levels were examined through real-time quantitative reverse transcriptase polymerase chain reaction.The clinicopathologic analysis was carried out in combination with clinical data.Results The p130 protein and p130 mRNA expression levels in the UPSC group(0.46±0.01 and 0.56±0.06,respectively were apparently less than that of the normal proliferative stage endometrium group(0.91±0.04 and 2.81±0.40,respectively;P < 0.01 and also less than those in high-level endometrial carcinoma(P < 0.05.Clinicopathologic analysis shows that all patients are post-menopausal women with symptoms of irregular vaginal bleeding and the average tumor size was 7.5cm(range: 1.2-14.8cm.The pathologic features are same as that of high-level ovarian papillary serous carcinoma.Conclusion Reduced p130 protein and p130 mRNA expression in UPSC might correlate with poor prognosis in UPSC patients.

  10. Induction of mRNA expression of osteogenesis-related genes by guaiacol in human dental pulp cells.

    Science.gov (United States)

    Kato, Takashi; Shirayama, Kumiko; Tsutsui, Takeo W; Tsutsui, Takeki

    2010-07-01

    To investigate the stimulating effect of endodontic medications on the mRNA expression of some osteogenesis-related genes associated with reparative dentinogenesis and hard-tissue formation, human dental pulp cells (D824 cells) were treated with calcium hydroxide (Ca (OH)(2)), formocresol, or guaiacol. The effect on growth was determined by growth curves of D824 cells treated for 1-3 days with 0.03-0.3 mM Ca (OH)(2), 0.0007%-0.0014% formocresol, or 0.24-2.43 mM guaiacol. The mitotic activity of individual cells and the mRNA expression of the osteogenesis-related genes for alkaline phosphatase (ALP), type I collagen (COL-1), and bone sialoprotein (BSP) in the cells treated for 24 h with the same concentrations of the medications as described above were determined by colony-forming efficiency and by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis, respectively. Cellular growth and mitotic activity were scarcely affected by Ca (OH)(2), but were significantly reduced by formocresol or guaiacol. The mRNA expression of the osteogenesis-related genes was little affected by Ca (OH)(2) or formocresol, but was significantly enhanced by guaiacol. The results indicate that guaiacol may stimulate the mRNA expression of genes associated with reparative dentinogenesis and hard-tissue formation in human dental pulp cells, suggesting that the novel property of guaiacol provides new insights into the utilization of guaiacol in endodontic therapy.

  11. Ivermectin induces P-glycoprotein expression and function through mRNA stabilization in murine hepatocyte cell line.

    Science.gov (United States)

    Ménez, Cécile; Mselli-Lakhal, Laïla; Foucaud-Vignault, Magali; Balaguer, Patrick; Alvinerie, Michel; Lespine, Anne

    2012-01-15

    Ivermectin is widely used in human and veterinary medicine for the control of helminth infections. Ivermectin is known to interact with P-glycoprotein (P-gp/MDR1), being a good substrate and a potent inhibitor, however, the influence of ivermectin on the expression of the transporter has not been investigated. Expression of P-glycoprotein was investigated in cultured mouse hepatocytes acutely exposed to ivermectin. The two P-glycoprotein murine isoforms, Mdr1a and Mdr1b, mRNA levels were assessed by real-time RT-PCR. Ivermectin induced a clear time- and concentration-dependent up-regulation of Mdr1a and Mdr1b mRNA levels (as early as a 12-h exposure and up to 2.5-fold at 10μM). Moreover, ivermectin-treated cells displayed enhanced cellular efflux of the P-glycoprotein substrate calcein that was inhibited by the P-glycoprotein blocker valspodar, providing evidence that the ivermectin-induced P-glycoprotein was functional. The mechanisms underlying these effects were investigated. Ivermectin-mediated Mdr1 mRNA induction was independent of the two nuclear receptors CAR and PXR, which are known to be involved in drug transporters regulation. Moreover, by using reporter cell lines that detects specific ligand-activated transcription factors, we showed that ivermectin did not displayed CAR, PXR or AhR ligand activities. However, studies with actinomycin D revealed that the half-life of Mdr1a and Mdr1b mRNA were significantly prolonged by two-fold in ivermectin-treated cells suggesting a post-transcriptional mode of ivermectin regulation. This study demonstrates for the first time that ivermectin induces P-glycoprotein overexpression through post-transcriptional mRNA stabilization, thus offering insight into the mechanism of reduced therapeutic efficacy and development of ivermectin-resistant parasites.

  12. Effects of simulated microgravity on microRNA and mRNA expression profile of rat soleus

    Science.gov (United States)

    Xu, Hongjie; Wu, Feng; Cao, Hongqing; Kan, Guanghan; Zhang, Hongyu; Yeung, Ella W.; Shang, Peng; Dai, Zhongquan; Li, Yinghui

    2015-02-01

    Spaceflight induces muscle atrophy but mechanism is not well understood. Here, we quantified microRNAs (miRNAs) and mRNA shifts of rat soleus in response to microgravity. MiRNAs and mRNA microarray of soleus after tail suspension (TS) for 7 and 14 days were performed followed by target gene and function annotation analysis and qRT-PCR. Relative muscle mass lost by 37.0% in TS-7 but less than 10% in the following three weeks. TS altered 23 miRNAs and 1313 mRNAs with at least 2-fold. QRT-PCR confirmed some of these changes. MiR-214, miR-486-5p and miR-221 continuously decreased. MiR-674 and Let-7e decreased only in TS-7, while miR-320b and miR-187 decreased only in TS-14. But there was no alteration of miR-320 and miR-206 in both time point. For mRNA detection, actn3 (5.1-fold and 13.8-fold) and myh4 (38-fold and 51.6-fold) increased abundantly and a3galt2 decreased. Predicted targeted genes (whyz, ywhaz and SFRP2) of altered miRNAs decreased. GO terms and cellular pathway of these alteration showed enrichment in regulation of muscle metabolism. Integration analysis of the miRNA and mRNA expression profiles confirmed that eleven genes were differently regulated by four miRNAs. This is the first study that showed expression pattern and synergistical regulation of miRNA and mRNA in rat soleus of TS for up to 14 days.

  13. Mixture effects of levonorgestrel and ethinylestradiol: estrogenic biomarkers and hormone receptor mRNA expression during sexual programming.

    Science.gov (United States)

    Säfholm, Moa; Jansson, Erika; Fick, Jerker; Berg, Cecilia

    2015-04-01

    Synthetic progesterone (progestins) and estrogens are widely used pharmaceuticals. Given that their simultaneous unintentional exposure occurs in wildlife and also in human infants, data on mixture effects of combined exposures to these hormones during development is needed. Using the Xenopus (Silurana) tropicalis test system we investigated mixture effects of levonorgestrel (LNG) and ethinylestradiol (EE2) on hormone sensitive endpoints. After larval exposure to LNG (0.1nM), or EE2 (0.1nM) singly, or in combination with LNG (0.01, 0.1, 1.0nM), the gonadal sex ratio was determined histologically and hepatic mRNA levels of genes encoding vitellogenin (vtg beta1) and the estrogen (esr1, esr2), progesterone (ipgr) and androgen (ar) receptors were quantified using quantitative PCR. All EE2-exposed groups showed female-biased sex ratios and increased vtg beta1 mRNA levels compared with the controls. Compared with the EE2-alone group (positive control) there were no significant alterations in vtg beta1 levels or in sex ratios in the co-exposure groups. Exposure to LNG-alone caused an increase in ar mRNA levels in females, but not in males, compared to the controls and the co-exposed groups, indicating that co-exposure to EE2 counteracted the LNG-induced ar levels. No treatment related impacts on the mRNA expression of esr1, esr2, and ipgr in female tadpoles were found, suggesting that these endpoints are insensitive to long-term exposure to estrogen or progestin. Due to the EE2-induced female-biased sex ratios, the mRNA expression data for the low number of males in the EE2-exposed groups were not statistically analyzed. In conclusion, our results suggest that induced vtg expression is a robust biomarker for estrogenic activity in exposure scenarios involving both estrogens and progestins. Developmental exposure to LNG caused an induction of hepatic ar mRNA expression that was antagonized by combined exposure to EE2 and LNG. To our knowledge this is the first study to

  14. Expression of GLUT4 mRNA of peripheral tissues and insulin resistance in rats with severe traumatic brain injury

    Institute of Scientific and Technical Information of China (English)

    CHEN Da-qing; ZHU Lie-lie; LI Yong-ling

    2007-01-01

    Objective: To evaluate the expression of glucose transporter-4 (GLUT4) mRNA in skeletal muscle and subcutaneous adipose tissues and investigate the mechanism of posttraumatic insulin resistance.Methods: Sixteen adult male Wistar rats were randomly divided into 2 group (n=8 in each group), i.e., severe traumatic brain injury (TBI) group due to falls from a height and normal control group. Blood glucose and serum insulin were measured at 0.5 h before trauma and 3 h, 24 h, 72 h, 7 d after trauma, respectively. And insulin sensitivity was calculated by insulin activity index (IAI) formula. Skeletal muscle and subcutaneous adipose tissue samples were collected at the same time when blood was sampled. The changes of expression of GLUT4 mRNA were observed using reverse transcription-polymerase chain reaction (RT-PCR).Results: Accompanied by the decrease of insulin sensitivity, the expression of GLUT4 mRNA was significantly decreased in adipose tissues at 24 h and 72 h after trauma (P<0.01), however, such phenomena did not appear in skeletal muscle samples.Conclusions: To some extent, the development of posttraumatic insulin resistance is related to the abnormality of transcription activity of GLUT4 gene. Adipose tissues show some difference in the transcriptional level of GLUT4 gene after trauma as compared with skeletal muscle tissues.

  15. Expression of trkB mRNA is altered in rat hippocampus after experimental brain trauma.

    Science.gov (United States)

    Hicks, R R; Zhang, L; Dhillon, H S; Prasad, M R; Seroogy, K B

    1998-08-31

    Recent investigations have shown that expression of mRNAs for the neurotrophins brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) is differentially altered in the hippocampus following traumatic brain injury. In the present study, modulation of neurotrophin receptor expression was examined in the hippocampus in a rat model of traumatic brain injury using in situ hybridization. Messenger RNA for trkB, the high-affinity receptor for BDNF and neurotrophin-4 (NT-4), was increased between 3 and 6 h bilaterally in the dentate gyrus following a lateral fluid-percussion brain injury of moderate severity (2.0-2.1 atm). No time-dependent alterations were observed for trkB mRNA in hippocampal subfields CA1 and CA3. Levels of mRNA for trkC, the high-affinity receptor for NT-3, did not change in any region of the hippocampus. These data demonstrate that lateral fluid-percussion injury modulates expression of trkB mRNA in the hippocampus and support a role for BDNF/trkB signalling mechanisms in secondary events associated with traumatic brain injury.

  16. FLT3-ITD and MLL-PTD influence the expression of MDR-1, MRP-1, and BCRP mRNA but not LRP mRNA assessed with RQ-PCR method in adult acute myeloid leukemia.

    Science.gov (United States)

    Nasilowska-Adamska, Barbara; Solarska, Iwona; Paluszewska, Monika; Malinowska, Iwona; Jedrzejczak, Wieslaw W; Warzocha, Krzysztof

    2014-04-01

    Fms-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) and mixed-lineage leukemia gene-partial tandem duplication (MLL-PTD) are aberrations associated with leukemia which indicate unsatisfactory prognosis. Downstream regulatory targets of FLT3-ITD and MLL-PTD are not well defined. We have analyzed the expression of MDR-1, multidrug resistant protein-1 (MRP-1), breast cancer resistance protein (BCRP), and lung resistance protein (LRP) messenger RNA (mRNA) in relation to the mutational status of FLT3-ITD and MLL-PTD in 185 acute myeloid leukemia (AML) adult patients. The real-time quantitative polymerase chain reaction method was performed to assess the expression of the MDR-1, MRP-1, BCRP, and LRP mRNA, and the results were presented as coefficients calculated using an intermediate method according to Pfaffl's rule. Significantly higher expressions of MDR-1 mRNA were found in patients who did not harbor FLT3-ITD (0.20 vs. 0.05; p = 0.0001) and MRP-1 mRNA in patients with this mutation (0.96 vs. 0.70; p = 0.002) and of BCRP mRNA in patients with MLL-PTD (0.61 vs. 0.38; p = 0.03). In univariate analysis, the high expression of MDR-1 mRNA (≥0.1317) negatively influenced the outcome of induction therapy (p = 0.05), whereas the high expression of BCRP mRNA (≥1.1487) was associated with a high relapse rate (RR) (p = 0.013). We found that the high expression of MDR-1 (≥0.1317), MRP-1 (≥0.8409), and BCRP mRNA (≥1.1487) significantly influenced disease-free survival (DFS; p = 0.059, 0.032, and 0.009, respectively) and overall survival (0.048, 0.014, and 0.059, respectively). Moreover, a high expression of BCRP mRNA (≥1.1487) proved to be an independent prognostic factor for RR (p = 0.01) and DFS (p = 0.002) in multivariate analysis. The significant correlation between the expression of MDR-1, MRP-1, and BCRP mRNA and FLT3-ITD or MLL-PTD in AML patients requires further investigation.

  17. A circadian neuropeptide PDF in the honeybee, Apis mellifera: cDNA cloning and expression of mRNA.

    Science.gov (United States)

    Sumiyoshi, Miho; Sato, Seiji; Takeda, Yukimasa; Sumida, Kazunori; Koga, Keita; Itoh, Tsunao; Nakagawa, Hiroyuki; Shimohigashi, Yasuyuki; Shimohigashi, Miki

    2011-12-01

    Pigment-dispersing factor (PDF) is a pacemaker hormone regulating the locomotor rhythm in insects. In the present study, we cloned the cDNAs encoding the Apis PDF precursor protein, and found that there are at least seven different pdf mRNAs yielded by an alternative splicing site and five alternative polyadenylation sites in the 5'UTR and 3'UTR regions. The amino acid sequence of Apis PDF peptide has a characteristic novel amino acid residue, aspargine (Asn), at position 17. Quantitative real-time PCR of total and 5'UTR insertion-type pdf mRNAs revealed, for the first time, that the expression levels change in a circadian manner with a distinct trough at the beginning of night in LD conditions, and at the subjective night under DD conditions. In contrast, the expression level of 5'UTR deletion-type pdf mRNAs was about half of that of the insertion type, and the expression profile failed to show a circadian rhythm. As the expression profile of the total pdf mRNA exhibited a circadian rhythm, transcription regulated at the promoter region was supposed to be controlled by some of the clock components. Whole mount in situ hybridization revealed that 14 lateral neurons at the frontal margin of the optic lobe express these mRNA isoforms. PDF expressing cells examined with a newly produced antibody raised against Apis PDF were also found to have a dense supply of axon terminals in the optic lobes and the central brain.

  18. Constitutive and heat-shock induced expression of Hsp70 mRNA during chicken testicular development and regression.

    Science.gov (United States)

    Mezquita, B; Mezquita, J; Durfort, M; Mezquita, C

    2001-01-01

    The constitutive and heat shock induced expression of Hsp70 mRNA was investigated in normal adult chicken testis and in adult testis after testicular regression induced by diethylstilbestrol (DES) treatment. In addition to the canonical form of Hsp70 mRNA, we have detected transcripts with an extended 5'UTR and transcripts containing, in the 5'UTR, sequences of the 18S ribosomal RNA. Hsp70 was expressed in unstressed male gonads in adult and regressed testis, being the expression much lower in regressed testis. Upon heat shock at 44 degrees C or 46 degrees C, Hsp70 was highly induced in both tissues. However, when testicular seminiferous tubules were incubated at the chicken internal temperature of 39 degrees C, no induction of Hsp70 was observed in mature testis, while the expression markedly increased in regressed testis. Induction at 39 degrees C was completely inhibited in the presence of 6 mM aspirin. Aspirin in the range 3-10 mM decreases the expression of Hsp70 in unstressed and stressed testicular cells, in striking contrast with the effect observed in other tissues as liver. These data suggest that the expression of Hsp70 is regulated in a specific manner in chicken testis and particularly in the male gonad undergoing regression. Copyright 2001 Wiley-Liss, Inc.

  19. Characteristics of mRNA dynamic expression related to spinal cord ischemia/reperfusion injury:a transcriptomics study

    Institute of Scientific and Technical Information of China (English)

    Zhi-ping Qi; Peng Xia; Ting-ting Hou; Ding-yang Li; Chang-jun Zheng; Xiao-yu Yang

    2016-01-01

    Following spinal cord ischemia/reperfusion injury, an endogenous damage system is immediately activated and participates in a cascade reac-tion. It is dififcult to interpret dynamic changes in these pathways, but the examination of the transcriptome may provide some information. The transcriptome relfects highly dynamic genomic and genetic information and can be seen as a precursor for the proteome. We used DNA microarrays to measure the expression levels of dynamic evolution-related mRNA after spinal cord ischemia/reperfusion injury in rats. The abdominal aorta was blocked with a vascular clamp for 90 minutes and underwent reperfusion for 24 and 48 hours. The simple ischemia group and sham group served as controls. After rats had regained consciousness, hindlimbs showed varying degrees of functional impairment, and gradually improved with prolonged reperfusion in spinal cord ischemia/reperfusion injury groups. Hematoxylin-eosin staining demon-strated that neuronal injury and tissue edema were most severe in the 24-hour reperfusion group, and mitigated in the 48-hour reperfusion group. There were 8,242 differentially expressed mRNAs obtained by Multi-Class Dif in the simple ischemia group, 24-hour and 48-hour reperfusion groups. Sixteen mRNA dynamic expression patterns were obtained by Serial Test Cluster. Of them, ifve patterns were signiifcant. In the No. 28 pattern, all differential genes were detected in the 24-hour reperfusion group, and their expressions showed a trend in up-regu-lation. No. 11 pattern showed a decreasing trend in mRNA whereas No. 40 pattern showed an increasing trend in mRNA from ischemia to 48 hours of reperfusion, and peaked at 48 hours. In the No. 25 and No. 27 patterns, differential expression appeared only in the 24-hour and 48-hour reperfusion groups. Among the ifve mRNA dynamic expression patterns, No. 11 and No. 40 patterns could distinguish normal spinal cord from pathological tissue. No. 25 and No. 27 patterns could distinguish

  20. Characteristics of mRNA dynamic expression related to spinal cord ischemia/reperfusion injury: a transcriptomics study

    Directory of Open Access Journals (Sweden)

    Zhi-ping Qi

    2016-01-01

    Full Text Available Following spinal cord ischemia/reperfusion injury, an endogenous damage system is immediately activated and participates in a cascade reaction. It is difficult to interpret dynamic changes in these pathways, but the examination of the transcriptome may provide some information. The transcriptome reflects highly dynamic genomic and genetic information and can be seen as a precursor for the proteome. We used DNA microarrays to measure the expression levels of dynamic evolution-related mRNA after spinal cord ischemia/reperfusion injury in rats. The abdominal aorta was blocked with a vascular clamp for 90 minutes and underwent reperfusion for 24 and 48 hours. The simple ischemia group and sham group served as controls. After rats had regained consciousness, hindlimbs showed varying degrees of functional impairment, and gradually improved with prolonged reperfusion in spinal cord ischemia/reperfusion injury groups. Hematoxylin-eosin staining demonstrated that neuronal injury and tissue edema were most severe in the 24-hour reperfusion group, and mitigated in the 48-hour reperfusion group. There were 8,242 differentially expressed mRNAs obtained by Multi-Class Dif in the simple ischemia group, 24-hour and 48-hour reperfusion groups. Sixteen mRNA dynamic expression patterns were obtained by Serial Test Cluster. Of them, five patterns were significant. In the No. 28 pattern, all differential genes were detected in the 24-hour reperfusion group, and their expressions showed a trend in up-regulation. No. 11 pattern showed a decreasing trend in mRNA whereas No. 40 pattern showed an increasing trend in mRNA from ischemia to 48 hours of reperfusion, and peaked at 48 hours. In the No. 25 and No. 27 patterns, differential expression appeared only in the 24-hour and 48-hour reperfusion groups. Among the five mRNA dynamic expression patterns, No. 11 and No. 40 patterns could distinguish normal spinal cord from pathological tissue. No. 25 and No. 27 patterns

  1. Arabidopsis mRNA polyadenylation machinery: comprehensive analysis of protein-protein interactions and gene expression profiling

    Directory of Open Access Journals (Sweden)

    Mo Min

    2008-05-01

    Full Text Available Abstract Background The polyadenylation of mRNA is one of the critical processing steps during expression of almost all eukaryotic genes. It is tightly integrated with transcription, particularly its termination, as well as other RNA processing events, i.e. capping and splicing. The poly(A tail protects the mRNA from unregulated degradation, and it is required for nuclear export and translation initiation. In recent years, it has been demonstrated that the polyadenylation process is also involved in the regulation of gene expression. The polyadenylation process requires two components, the cis-elements on the mRNA and a group of protein factors that recognize the cis-elements and produce the poly(A tail. Here we report a comprehensive pairwise protein-protein interaction mapping and gene expression profiling of the mRNA polyadenylation protein machinery in Arabidopsis. Results By protein sequence homology search using human and yeast polyadenylation factors, we identified 28 proteins that may be components of Arabidopsis polyadenylation machinery. To elucidate the protein network and their functions, we first tested their protein-protein interaction profiles. Out of 320 pair-wise protein-protein interaction assays done using the yeast two-hybrid system, 56 (~17% showed positive interactions. 15 of these interactions were further tested, and all were confirmed by co-immunoprecipitation and/or in vitro co-purification. These interactions organize into three distinct hubs involving the Arabidopsis polyadenylation factors. These hubs are centered around AtCPSF100, AtCLPS, and AtFIPS. The first two are similar to complexes seen in mammals, while the third one stands out as unique to plants. When comparing the gene expression profiles extracted from publicly available microarray datasets, some of the polyadenylation related genes showed tissue-specific expression, suggestive of potential different polyadenylation complex configurations. Conclusion An

  2. Reduced mRNA expression of PTGDS in peripheral blood mononuclear cells of rapid-cycling bipolar disorder patients compared with healthy control subjects

    DEFF Research Database (Denmark)

    Munkholm, Klaus; Peijs, Lone; Kessing, Lars Vedel;

    2015-01-01

    that mRNA expression of PTGDS and AKR1C3 is deregulated in rapid-cycling disorder patients in a euthymic or current affective state compared with healthy control subjects, and that expression alters with affective states. METHODS: PTGDS and AKR1C3 mRNA expression in peripheral blood mononuclear cells...... was measured in 37 rapid-cycling bipolar disorder patients and 40 age- and gender-matched healthy control subjects using reverse transcription quantitative real-time polymerase chain reaction. Repeated measurements of PTGDS and AKR1C3 mRNA expression were obtained in various affective states during 6-12 months...

  3. Interleukin-6 modifies mRNA expression in mouse skeletal muscle

    DEFF Research Database (Denmark)

    Hassing, Helle Adser; Wojtaszewski, Jørgen; Jakobsen, Anne Hviid;

    2011-01-01

    Aim: The aim of the present study was to test the hypothesis that interleukin-6 plays a role in exercise-induced PGC-1a and TNFa mRNA responses in skeletal muscle and to examine the potential IL-6 mediated AMPK regulation in these responses. Methods: Whole body IL-6 knockout and wildtype (WT) male...

  4. Gravitational loading of a simulated launch alters mRNA expression in osteoblasts

    Science.gov (United States)

    Fitzgerald, J.; Hughes-Fulford, M.

    1996-01-01

    Serum-deprived mouse osteoblastic cells (MC3T3-E1a) were centrifuged under a regime designed to simulate a space shuttle launch (maximum of 3g). Messenger RNA levels for eight genes involved in bone growth and maintenance were determined using RT-PCR. Following 30 min of centrifugation, mRNA level for early response gene c-fos was significantly increased 89% (P gene osteocalcin was significantly decreased to 44% of control level (P basal mRNA level for TGFbeta3 was detected. In addition, no change in the steady-state synthesis of prostaglandin E2 was detected, possibly due to lack of lipid substrates in serum-deprived cells, suggesting that the increase in c-fos mRNA in response to gravitational loading is a result of mechanical stimulation. These results indicate that a small magnitude mechanical loading, such as that experienced during a shuttle launch, can alter mRNA levels in quiescent osteoblastic cells.

  5. Relative Expression of HIF-1α mRNA in Rat Heart, Brain and Blood During Induced Systemic Hypoxia

    Directory of Open Access Journals (Sweden)

    Syarifah Dewi

    2009-11-01

    Full Text Available Hypoxia is a pathological condition in which the body as a whole or region of the body (tissue or cell deprived of adequate oxygen supply. The transcriptional regulator hypoxia inducible factor-1 (HIF-1 is an essential mediator of O2 homeostasis. Unlike the β sub unit (HIF-1β, the activity of HIF-1α is controlled in an oxygen-dependent manner. It has been reported that the stability and expression of HIF-1α during hypoxia is remarkably higher than those under normoxic conditions.The aim of this study was to analyze the adaptive tissue responses during induced systemic hypoxia by comparation of relative expression of mRNA HIF-1α in rat heart, brain and blood. Twenty-five male Sprague Dawley rats were subjected to systemic hypoxia by placing them in the hypoxic chamber supplied by 8-10% of O2 for 0, 1, 7, 14 and 21 days, respectively. The relative expression level of HIF-1α mRNA in brain, heart and leucocyte cells were analyzed using quantitative RT-PCR assay (Real Time PCR based on Pfaff's formula. This study demonstrates that the increased of relative expression of HIF-1α mRNA during induced systemic hypoxia reached its maximum level at day 7 (in heart or at day 14 (in brain, whereas in leucocyte cells the stimulation of HIF-1α expression was intensively maintained up to 21 days although the expression has reached the remarkably high level. We could conclude that HIF-1α as an oxygen sensing during systemic hypoxia has different capacity and sensitivity in brain, heart and blood tissues, due to the importance of oxygen homeostasis in each tissue.

  6. Leptin and cholecystokinin in Schizothorax prenanti: molecular cloning, tissue expression, and mRNA expression responses to periprandial changes and fasting.

    Science.gov (United States)

    Yuan, Dengyue; Wang, Tao; Zhou, Chaowei; Lin, Fangjun; Chen, Hu; Wu, Hongwei; Wei, Rongbin; Xin, Zhiming; Li, Zhiqiong

    2014-08-01

    In the present study, full-length cDNA sequences of leptin and cholecystokinin (CCK) were cloned from Schizothorax prenanti (S. prenanti), and applied real-time quantitative PCR to characterize the tissue distribution, and appetite regulatory effects of leptin and CCK in S. prenanti. The S. prenanti leptin and CCK full-length cDNA sequences were 1121 bp and 776 bp in length, encoding the peptide of 171 and 123 amino acid residues, respectively. Tissue distribution analysis showed that leptin mRNA was mainly expressed in the liver of S. prenanti. CCK was widely expressed, with the highest levels of expression in the hypothalamus, myelencephalon, telencephalon and foregut of S. prenanti. The CCK mRNA expression was highly elevated after feeding, whereas the leptin mRNA expression was not affected by single meal. These results suggested that CCK is a postprandial satiety signal in S. prenanti, but leptin might not be. In present study, leptin and CCK gene expression were both decreased after fasting and increased after refeeding, which suggested leptin and CCK might be involved in regulation of appetite in S. prenanti. This study provides an essential groundwork to further elucidate the appetite regulatory systems of leptin and CCK in S. prenanti as well as in other teleosts.

  7. A Candida albicans gene expressed in Saccharomyces cerevisiae results in a distinct pattern of mRNA processing.

    Science.gov (United States)

    Iborra, A; Sentandreu, R; Gozalbo, D

    1996-09-01

    Two plasmids (derived from YCplac22 and YEplac112) carrying a Candida albicans gene (including the 5' non-coding promoter sequences) coding for a 30 kDa membrane-bound protein, were used to transform Saccharomyces cerevisiae cells. A 30 kDa protein was immunodetected by Western blot in the membrane fraction of transformants. Northern analysis showed the presence of three mRNA species (of about 1.1, 0.7 and 0.5 kb) hybridizing with the C. albicans gene as a probe. The same result was obtained using the 5' and 3' regions of the gene as probes, whereas only a 1.1 kb mRNA was found in C. albicans and none was detected in S. cerevisiae control transformants. Thus, heterologous expression of this gene in S. cerevisiae results in a distinct pattern of mRNA processing, either due to the location on plasmid vectors and/or to differences in the mRNA processing systems in the two microorganisms.

  8. Expression of RFamide-Related Peptide-3 (RFRP-3) mRNA in Dorsomedial Hypothalamic Nucleus and KiSS-1 mRNA in Arcuate Nucleus of Rat during Pregnancy

    Science.gov (United States)

    Sabet Sarvestani, Fatemeh; Tamadon, Amin; Koohi-Hosseinabadi, Omid; Mohammadi Nezhad, Saeed; Rahmanifar, Farhad; Jafarzadeh Shirazi, Mohammad Reza; Tanideh, Nader; Moghadam, Ali; Niazi, Ali

    2014-01-01

    Background RFamide-related peptide-3 (RFRP-3) and kisspeptin (KiSS-1) are known to respectively inhibit and stimulate gonadotropin releasing hormone (GnRH) and lute- inizing hormone (LH) secretion in rat. The aim of the present study was to evaluate the relative mRNA expression of RFRP-3 and KiSS-1 in the hypothalamus of pregnant rats. Materials and Methods In a randomized controlled experimental study, the exact preg- nancy day of 18 Sprague-Dawley rats were confirmed using the vaginal smear method and were equally assigned to three groups of days 7, 14 and 21 of pregnancy. Four non- pregnant female rats were ovariectomized and assigned as the control group. All rats were decapitated, and the dorsomedial hypothalamic nucleus (DMH) and the arcuate nucleus (ARC) for detection of KiSS-1 mRNA were separated from their hypothalamus to detect RFRP-3 and KiSS-1 mRNA respectively. Then, their relative expressions were compared between control and pregnant groups using real-time polymerase chain reac- tion (PCR). Results The relative expression of RFRP-3 mRNA in DMH did not change significantly during pregnancy (p>0.01). However, the relative expression of KiSS-1 mRNA in ARC was at its highest in day 7 of pregnancy and decreased until day 21 of pregnancy (p<0.01). Conclusion Decrease in GnRH and LH secretion during the pregnancy of rat may be controlled by constant expression of RFRP-3 mRNA and reduced expression of KiSS-1 mRNA in hypothalamus. PMID:25379163

  9. Expression of RFamide-Related Peptide-3 (RFRP-3) mRNA in Dorsomedial Hypothalamic Nucleus and KiSS-1 mRNA in Arcuate Nucleus of Rat during Pregnancy.

    Science.gov (United States)

    Sabet Sarvestani, Fatemeh; Tamadon, Amin; Koohi-Hosseinabadi, Omid; Mohammadi Nezhad, Saeed; Rahmanifar, Farhad; Jafarzadeh Shirazi, Mohammad Reza; Tanideh, Nader; Moghadam, Ali; Niazi, Ali

    2014-10-01

    RFamide-related peptide-3 (RFRP-3) and kisspeptin (KiSS-1) are known to respectively inhibit and stimulate gonadotropin releasing hormone (GnRH) and lute- inizing hormone (LH) secretion in rat. The aim of the present study was to evaluate the relative mRNA expression of RFRP-3 and KiSS-1 in the hypothalamus of pregnant rats. In a randomized controlled experimental study, the exact preg- nancy day of 18 Sprague-Dawley rats were confirmed using the vaginal smear method and were equally assigned to three groups of days 7, 14 and 21 of pregnancy. Four non- pregnant female rats were ovariectomized and assigned as the control group. All rats were decapitated, and the dorsomedial hypothalamic nucleus (DMH) and the arcuate nucleus (ARC) for detection of KiSS-1 mRNA were separated from their hypothalamus to detect RFRP-3 and KiSS-1 mRNA respectively. Then, their relative expressions were compared between control and pregnant groups using real-time polymerase chain reac- tion (PCR). The relative expression of RFRP-3 mRNA in DMH did not change significantly during pregnancy (p>0.01). However, the relative expression of KiSS-1 mRNA in ARC was at its highest in day 7 of pregnancy and decreased until day 21 of pregnancy (p<0.01). Decrease in GnRH and LH secretion during the pregnancy of rat may be controlled by constant expression of RFRP-3 mRNA and reduced expression of KiSS-1 mRNA in hypothalamus.

  10. Guipi decoction effects on brain somatostatin levels and receptor mRNA expression in rats with spleen deficiency

    Institute of Scientific and Technical Information of China (English)

    Huinan Qian; Le Wang; Libo Shen; Xueqin Hu

    2008-01-01

    BACKGROUND:Somatostatin is abundant in the hypothalamus,cerebral cortex,limbic system,and mesencephalon.Somatostatin mRNA expression in the brain of rats with spleen deficiency is noticeably reduced,as well as attenuation of cognitive function. OBJECTIVE:To observe the interventional effect of Guipi decoction on somatostatin level and somatostatin receptor 1(SSTRI)mRNA expression in different encephalic regions of rats with spleen deficiency,and to compare the interventional effects of Guipi decoction,Chaihu Shugan powder,and Tianwang Buxin pellet. DESIGN:A randomized controlled observation. SETTING:Basic Medical College,Beijing University of Traditional Chinese Medicine.MATERIALS:Fifty adult Wistar male rats,of clean grade,weighing(160 ± 10)g,were provided by Beijing Weitong Lihua Laboratory Animal Technology Co.,Ltd.The protocol was performed in accordance with ethical guidelines for the use and care of animals.Somatostatin 1 polyclonal anti-rabbit antibody and SSTR1 in situ hybridization kit were provided by Department of Neuroanatomy,Shanghai Second Military Medical University of Chinese PLA.The drug for developing rat models of spleen deficiency was composed of Dahuang,Houpu and Zhishi,and prepared at 2:1:1.Guipi decoction,Chaihu Shugan powder,and Tianwang Buxin pellet recipes were made according to previous studies.METHODS:This study was performed at the Basic Medical College,Beijing University of Traditional Chinese Medicine from March 2002 to March 2005.The rats were randomly divided into 5 groups,with 10 rats in each group:normal,model,Guipi decoction,Chaihu Shugan powder,and Tianwang Buxin pelletgroups.Rat models of the latter 4 groups were developed by methods of purgation with bitter and cold nature drugs,improper diet,and overstrain.The rats received 7.5 g/kg of the drugs each morning and were fasted every other day,but were allowed free access to water at all times,The rats were forced to swim in 25℃ water until fatigued.Rats in the normal group

  11. REAL-TIME DETECTION OF SURVIVIN mRNA EXPRESSION IN CERVICAL CANCER CELL LINES USING MOLECULAR BEACON IMAGING

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The initiated growth of human cancer cells of-ten mostly come fromthe abnor mal expression ofgenes.Survivinis anapotosis inhibitor of IAPfami-ly,cloned by Ambrosini in1997usingthe cDNAofeffector cell protease receptor-1(EPR-1),and is thekey gene for the development and advancement oftumor.Inthe present study,the feasibility of detec-ting the expression of survivin mRNA was exam-inedincervical cancer cell lines using molecular bea-coni maging technology.MATERIALS AND METHODS1Cervical cancer cell lines and ce...

  12. Expression of Endogenous Beta Retroviruses and Hyal-2 mRNA in Immune Organs of Fetuses and Lambs

    Institute of Scientific and Technical Information of China (English)

    Jing-wei Qi; Xiao-li Wu; Shu-ying Liu; Gui-fang Cao

    2012-01-01

    Endogenous beta retroviruses (enJSRV) are highly homologous with Jaagsiekte sheep retrovirus (exJSRV),this exogenous retrovirus is the aetiological agent of ovine pulmonary adenocarcinoma (OPA).The aim of this study was to clarify the function of enJSRV and the immunological mechanisms of its corresponding antibody,that is undetectable in JSRV-infected ovine serum.The expression of enJSRV envelope protein and Hyal-2 mRNA in immune organs and lungs of ovine fetuses and lambs were analyzed by Real-Time reverse transcription PCR and In Situ Hybridization using specific probes.In Situ Hybridization results indicated that the enJSRV envelope protein and Hyal-2 mRNA were expressed in thymus,spleen,mesenteric lymph nodes and lungs at different times,while no positive signals were detected in the negative controls.On the other hand,results from Real-Time reverse transcription PCR analysis showed that in 130d fetuses and 3d newborn lambs the enJSRV mRNA levels were much higher in organs associated with the immune system than that in lungs,especially in the thymus and spleen,but levels of Hyal-2 mRNA expression was not significantly different in all collected tissue.These results provided evidence from an immunology point of view to understand why the circulating antibodies against exJSRV are undetectable in JSRV-infected ovine,and will help to unravel the pathogenesis of JSRV-infected ovine.

  13. Comparative Analysis of mRNA Isoform Expression in Cardiac Hypertrophy and Development Reveals Multiple Post-Transcriptional Regulatory Modules

    Science.gov (United States)

    Park, Ji Yeon; Li, Wencheng; Zheng, Dinghai; Zhai, Peiyong; Zhao, Yun; Matsuda, Takahisa; Vatner, Stephen F.; Sadoshima, Junichi; Tian, Bin

    2011-01-01

    Cardiac hypertrophy is enlargement of the heart in response to physiological or pathological stimuli, chiefly involving growth of myocytes in size rather than in number. Previous studies have shown that the expression pattern of a group of genes in hypertrophied heart induced by pressure overload resembles that at the embryonic stage of heart development, a phenomenon known as activation of the “fetal gene program”. Here, using a genome-wide approach we systematically defined genes and pathways regulated in short- and long-term cardiac hypertrophy conditions using mice with transverse aortic constriction (TAC), and compared them with those regulated at different stages of embryonic and postnatal development. In addition, exon-level analysis revealed widespread mRNA isoform changes during cardiac hypertrophy resulting from alternative usage of terminal or internal exons, some of which are also developmentally regulated and may be attributable to decreased expression of Fox-1 protein in cardiac hypertrophy. Genes with functions in certain pathways, such as cell adhesion and cell morphology, are more likely to be regulated by alternative splicing. Moreover, we found 3′UTRs of mRNAs were generally shortened through alternative cleavage and polyadenylation in hypertrophy, and microRNA target genes were generally de-repressed, suggesting coordinated mechanisms to increase mRNA stability and protein production during hypertrophy. Taken together, our results comprehensively delineated gene and mRNA isoform regulation events in cardiac hypertrophy and revealed their relations to those in development, and suggested that modulation of mRNA isoform expression plays an importance role in heart remodeling under pressure overload. PMID:21799842

  14. The effect of hypothermia on the expression of neurotrophin mRNA in the hippocampus following transient cerebral ischemia in the rat.

    Science.gov (United States)

    Boris-Möller, F; Kamme, F; Wieloch, T

    1998-12-10

    The expression of the mRNAs of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT3) and the neurotrophin receptor, TrkB, was studied in the rat hippocampus by in situ hybridization following normothermic (37 degreesC) and protective hypothermic (33 degreesC) transient cerebral ischemia of 15 min duration. In the resistant dentate gyrus, normothermic ischemia transiently induced NGF mRNA at around 8 h of recovery, while the NT3 mRNA levels were depressed over at least a 24-h recovery period. The levels of BDNF and TrkB were transiently and markedly elevated with a maximal expression at 24 h of recovery. Intraischemic hypothermia reduced the induction of NGF mRNA, while the increase of BDNF mRNA expression occurred earlier during recovery, and the post-ischemic NT3 mRNA depression was not affected. Also, the expression of TrkB mRNA was enhanced, and occurred concomitantly with the elevation of BDNF mRNA. In contrast, there were no changes in neurotrophin and TrkB mRNA in the CA3 and CA1 regions. The expression of BDNF mRNA at 24 h after normothermic ischemia, was attenuated by intraischemic hypothermia. We conclude that, the expressions of NGF, BDNF, NT3 or TrkB mRNA in ischemia-sensitive hippocampal subregions are not increased by protective hypothermia. In contrast, hypothermia induces neurotrophin mRNA alterations in the ischemia-resistant dentate gyrus that may convey protection to sensitive regions.

  15. Gene expression analysis in sections and tissue microarrays of archival tissues by mRNA in situ hybridization.

    Science.gov (United States)

    Henke, R T; Maitra, A; Paik, S; Wellstein, A

    2005-01-01

    Altered expression of genes in diseased tissues can prognosticate a distinct natural progression of the disease as well as predict sensitivity or resistance to particular therapies. Archival tissues from patients with a known medical history and treatments are an invaluable resource to validate the utility of candidate genes for prognosis and prediction of therapy outcomes. However, stored tissues with associated long-term follow-up information typically are formalin-fixed, paraffin-embedded specimen and this can severely restrict the methods applicable for gene expression analysis. We report here on the utility of tissue microarrays (TMAs) that use valuable tissues sparingly and provide a platform for simultaneous analysis of gene expression in several hundred samples. In particular, we describe a stable method applicable to mRNA expression screening in such archival tissues. TMAs are constructed from sections of small drill cores, taken from tissue blocks of archival tissues and multiple samples can thus be arranged on a single microscope slide. We used mRNA in situ hybridization (ISH) on >500 full sections and >100 TMAs for >10 different cDNAs that yielded >10,000 data points. We provide detailed experimental protocols that can be implemented without major hurdles in a molecular pathology laboratory and discuss quantitative analysis and the advantages and limitations of ISH. We conclude that gene expression analysis in archival tissues by ISH is reliable and particularly useful when no protein detection methods are available for a candidate gene.

  16. Sex Differences in mRNA Expression of Reduced Folate Carrier-1, Folypolyformyl Glutamate Synthase, and γ-Glutamyl Hydrolase in a Healthy Japanese Population.

    Science.gov (United States)

    Hashiguchi, Masayuki; Tanaka, Takanori; Shimizu, Mikiko; Tsuru, Tomomi; Chiyoda, Takeshi; Miyawaki, Kumika; Irie, Shin; Takeuchi, Osamu; Hakamata, Jun; Mochizuki, Mayumi

    2016-12-01

    Sex differences in the prevalence of autoimmune diseases such as rheumatoid arthritis (RA) are well known, but little is known about those differences in relation to therapeutic response. Reduced folate carrier-1 (RFC-1), folypolyformyl glutamate synthase (FPGS), and γ-glutamyl hydrolase (GGH) are important transporters and enzymes that convert methotrexate (MTX) in the body. This study investigated the sex differences in mRNA expression of RFC-1, FPGS, and GGH in 190 unrelated healthy Japanese people. The genotypes and mRNA expression were determined using the real-time PCR method. Significant differences between men and women were observed in RFC-1, FPGS, and GGH mRNA expression. The mRNA expression of FPGS and GGH was greater in women than that in men, but the expression of RFC-1 was less in the former than the latter. In stratified analysis by genotype, significant differences in sex-specific mRNA expression were observed in G/G of FPGS, C/C of GGH 452, and C/C of GGH -401. All showed greater mRNA expression in women than in men. In the 5 single-nucleotide polymorphisms RFC-1 80G>A, RFC-1 -43T>C, FPGS 1994G>A, GGH 452C>T, and GGH -401C>T examined, the FPGS 1994 G/G (1.46-fold), GGH 452 C/C (2.16-fold), and GGH -401 C/C (2.68-fold) genotypes showed significantly higher mRNA expression in women than in men. Healthy Japanese adults in this study showed sex-specific differences in mRNA expression that differed among RFC-1, FPGS, and GGH. Therefore, the relationship between genetic polymorphisms and mRNA expression including sex differences might contribute to the variation in the efficacy/toxicity of MTX in patients with RA.

  17. The role of tumor cell-derived connective tissue growth factor (CTGF/CCN2) in pancreatic tumor growth

    DEFF Research Database (Denmark)

    Bennewith, Kevin L; Huang, Xin; Ham, Christine M

    2009-01-01

    Pancreatic cancer is highly aggressive and refractory to existing therapies. Connective tissue growth factor (CTGF/CCN2) is a fibrosis-related gene that is thought to play a role in pancreatic tumor progression. However, CCN2 can be expressed in a variety of cell types, and the contribution of CCN2...... adenocarcinomas. Furthermore, we found increased CCN2 staining in clinical pancreatic tumor tissue relative to stromal cells surrounding the tumor, supporting our assertion that tumor cell-derived CCN2 is important for pancreatic tumor growth. Taken together, these data improve our understanding of the mechanisms...

  18. Influence of clonidine and ketamine on m-RNA expression in a model of opioid-induced hyperalgesia in mice.

    Directory of Open Access Journals (Sweden)

    Henning Ohnesorge

    Full Text Available BACKGROUND: We investigated the influence of morphine and ketamine or clonidine in mice on the expression of genes that may mediate pronociceptive opioid effects. MATERIAL AND METHODS: C57BL/6 mice received morphine injections thrice daily using increasing doses (5-20 mg∙kg(-1 for 3 days (sub-acute, n=6 or 14 days (chronic, n=6 and additionally either s-ketamine (5 mg∙kg(-1, n=6 or clonidine (0.1 mg∙kg(-1, n=6. Tail flick test and the assessment of the mechanical withdrawal threshold of the hindpaw was performed during and 4 days after cessation of opioid treatment. Upon completion of the behavioural testing the mRNA-concentration of the NMDA receptor (NMDAR1 and β-arrestin 2 (Arrb2 were measured by PCR. RESULTS: Chronic opioid treatment resulted in a delay of the tail flick latency with a rapid on- and offset. Simultaneously the mice developed a static mechanical hyperalgesia with a delayed onset that that outlasted the morphine treatment. Sub-acute morphine administration resulted in a decrease of NMDAR1 and Arrb2 whereas during longer opioid treatment the expression NMDAR1 and Arrb2 mRNA increased again to baseline values. Coadministration of s-ketamine or clonidine resulted in a reversal of the mechanical hyperalgesia and inhibited the normalization of NMDAR1 mRNA expression but had no effect on the expression of Arrb2 mRNA. CONCLUSION: In the model of chronic morphine therapy the antinociceptive effects of morphine are represented by the thermal analgesia while the proniceptive effects are represented by the mechanical hyperalgesia. The results indicate that the regulation of the expression of NMDAR1 and Arrb2 may be associated to the development of OIH in mice. PERSPECTIVE: The results indicate that co-administration of clonidine or ketamine may influence the underlying mechanisms of OIH.

  19. ALDH1A1 mRNA expression in association with prognosis of triple-negative breast cancer.

    Science.gov (United States)

    Liu, Yan; Baglia, Michelle; Zheng, Ying; Blot, William; Bao, Ping-Ping; Cai, Hui; Nechuta, Sarah; Zheng, Wei; Cai, Qiuyin; Shu, Xiao Ou

    2015-12-01

    ALDH1 is a crucial element in the retinoic acid signaling pathway regulating the self-renewal and differentiation of normal stem cells, and may play an important role in cancer progression. However, research on ALDH1 gene expression and breast cancer prognosis has yielded conflicting results. We evaluated the association between tumor tissue ALDH1A1/ALDH1A3 mRNA expression and triple-negative breast cancer (TNBC) prognosis in the Shanghai Breast Cancer Survival Study (SBCSS, N=463), Nashville Breast Health Study (NBHS, N=86), and Southern Community Cohort Study (SCCS, N=47). Gene expression was measured in RNA isolated from breast cancer tissues. In the SBCSS, higher ALDH1A1 mRNA level was associated with improved disease-free (HR=0.87, 95% CI: 0.80-0.95, per log unit change) and overall survival (HR=0.85, 95% CI: 0.78-0.93 per log unit change) independent of age at diagnosis, TNM stage and treatment. We replicated the findings for overall survival in the NBHS and SCCS (HR = 0.27, 95% CI: 0.10-0.73) and for disease-free survival by a meta-analysis of four publicly-available gene expression datasets (HR = 0.86, 95% CI: 0.76-0.97). No significant association was found for ALDH1A3.Our study suggests high expression of ALDH1A1 mRNA in tumor tissues may be an independent predictor of a favorable TNBC outcome.

  20. Effect of long real space flight on the whole genome mRNA expression properties in medaka Oryzias latipes

    Science.gov (United States)

    Kozlova, Olga; Gusev, Oleg; Levinskikh, Margarita; Sychev, Vladimir; Poddubko, Svetlana

    The current study is addressed to the complex analysis of whole genome mRNA expression profile and properties of splicing variants formation in different organs of medaka fish exposed to prolonged space flight in the frame of joint Russia-Japan research program “Aquarium-AQH”. The fish were kept in the AQH joint-aquariums system in October-December 2013, followed by fixation in RNA-preserving buffers and freezing during the space flight. The samples we returned to the Earth frozen in March 2013 and mRNAs from four fish were sequenced in organ-specific manner using HiSeq Illumina sequencing platform. The ground group fish treated in the same way was used as a control. The comparison between the groups revealed space group-specific specific mRNA expression pattern. More than 50 genes (including several types of myosins) were down-regulated in the space group. Moreover, we found an evidence for formation of space group-specific splicing variants of mRNA. Taking together, the data suggest that in spite of aquatic environment, space flight-associated factors have a strong effect on the activity of fish genome. This work was supported in part by subsidy of the Russian Government to support the Program of competitive growth of Kazan Federal University among world class academic centres and universities.

  1. The Csr system regulates genome-wide mRNA stability and transcription and thus gene expression in Escherichia coli.

    Science.gov (United States)

    Esquerré, Thomas; Bouvier, Marie; Turlan, Catherine; Carpousis, Agamemnon J; Girbal, Laurence; Cocaign-Bousquet, Muriel

    2016-04-26

    Bacterial adaptation requires large-scale regulation of gene expression. We have performed a genome-wide analysis of the Csr system, which regulates many important cellular functions. The Csr system is involved in post-transcriptional regulation, but a role in transcriptional regulation has also been suggested. Two proteins, an RNA-binding protein CsrA and an atypical signaling protein CsrD, participate in the Csr system. Genome-wide transcript stabilities and levels were compared in wildtype E. coli (MG1655) and isogenic mutant strains deficient in CsrA or CsrD activity demonstrating for the first time that CsrA and CsrD are global negative and positive regulators of transcription, respectively. The role of CsrA in transcription regulation may be indirect due to the 4.6-fold increase in csrD mRNA concentration in the CsrA deficient strain. Transcriptional action of CsrA and CsrD on a few genes was validated by transcriptional fusions. In addition to an effect on transcription, CsrA stabilizes thousands of mRNAs. This is the first demonstration that CsrA is a global positive regulator of mRNA stability. For one hundred genes, we predict that direct control of mRNA stability by CsrA might contribute to metabolic adaptation by regulating expression of genes involved in carbon metabolism and transport independently of transcriptional regulation.

  2. Monitoring p21 mRNA expression in living cell based on molecular beacon fluorescence increasing rate

    Institute of Scientific and Technical Information of China (English)

    TANG HongXing; YANG XiaoHai; WANG KeMin; TAN WeiHong; LIU Bin; HE LiFang; WANG Wei

    2008-01-01

    Studying the expression level of mRNA in living cells will offer tremendous opportunities for ad-vancement in cell biology research, disease diagnostics, and drug discovery. In this paper, a molecular beacon (MB) specific for the important tumor suppressor gene p21 has been designed and synthesized. The fluorescence signal was detected in real-time after the MB entered the cytoplasm of nasopharyn-geal carcinoma cells. After injecting the p21MB into nasopharyngeal carcinoma cell and p33-trans-fected nasopharyngeal carcinoma cell, the consistent increase of fluorescent signal intensity was de-tected in both cell lines, and maximum fluorescence intensity achieved in about 15 min. In about 4 min following microinjection, the fluorescence increasing rate was significantly different between these two cell lines, which indicate the different p21 mRNA expression levels. The results obtained in the real-time detection were also validated by RT-PCR. Analysis of the initial fluorescence increasing rate can effi-ciently reduce the side effect of enzyme and improve the accuracy in living cell mRNA detection.

  3. Occupational Toluene Exposure Induces Cytochrome P450 2E1 mRNA Expression in Peripheral Lymphocytes

    Science.gov (United States)

    Mendoza-Cantú, Ania; Castorena-Torres, Fabiola; de León, Mario Bermúdez; Cisneros, Bulmaro; López-Carrillo, Lizbeth; Rojas-García, Aurora E.; Aguilar-Salinas, Alberto; Manno, Maurizio; Albores, Arnulfo

    2006-01-01

    Print workers are exposed to organic solvents, of which the systemic toxicant toluene is a main component. Toluene induces expression of cytochrome P450 2E1 (CYP2E1), an enzyme involved in its own metabolism and that of other protoxicants, including some procarcinogens. Therefore, we investigated the association between toluene exposure and the CYP2E1 response, as assessed by mRNA content in peripheral lymphocytes or the 6-hydroxychlorzoxazone (6OH-CHZ)/chlorzoxazone (CHZ) quotient (known as CHZ metabolic ratio) in plasma, and the role of genotype (5′-flanking region RsaI/PstI polymorphic sites) in 97 male print workers. The geometric mean (GM) of toluene concentration in the air was 52.80 ppm (10–760 ppm); 54% of the study participants were exposed to toluene concentrations that exceeded the maximum permissible exposure level (MPEL). The GM of urinary hippuric acid at the end of a work shift (0.041 g/g creatinine) was elevated relative to that before the shift (0.027 g/g creatinine; p < 0.05). The GM of the CHZ metabolic ratio was 0.33 (0–9.3), with 40% of the subjects having ratios below the GM. However, the average CYP2E1 mRNA level in peripheral lymphocytes was 1.07 (0.30–3.08), and CYP2E1 mRNA levels within subjects correlated with the toluene exposure ratio (environmental toluene concentration:urinary hippuric acid concentration) (p = 0.014). Genotype did not alter the association between the toluene exposure ratio and mRNA content. In summary, with further validation, CYP2E1 mRNA content in peripheral lymphocytes could be a sensitive and noninvasive biomarker for the continuous monitoring of toluene effects in exposed persons. PMID:16581535

  4. Occupational toluene exposure induces cytochrome P450 2E1 mRNA expression in peripheral lymphocytes.

    Science.gov (United States)

    Mendoza-Cantú, Ania; Castorena-Torres, Fabiola; Bermúdez de León, Mario; Cisneros, Bulmaro; López-Carrillo, Lizbeth; Rojas-García, Aurora E; Aguilar-Salinas, Alberto; Manno, Maurizio; Albores, Arnulfo

    2006-04-01

    Print workers are exposed to organic solvents, of which the systemic toxicant toluene is a main component. Toluene induces expression of cytochrome P450 2E1 (CYP2E1), an enzyme involved in its own metabolism and that of other protoxicants, including some procarcinogens. Therefore, we investigated the association between toluene exposure and the CYP2E1 response, as assessed by mRNA content in peripheral lymphocytes or the 6-hydroxychlorzoxazone (6OH-CHZ)/chlorzoxazone (CHZ) quotient (known as CHZ metabolic ratio) in plasma, and the role of genotype (5 -flanking region RsaI/PstI polymorphic sites) in 97 male print workers. The geometric mean (GM) of toluene concentration in the air was 52.80 ppm (10-760 ppm); 54% of the study participants were exposed to toluene concentrations that exceeded the maximum permissible exposure level (MPEL). The GM of urinary hippuric acid at the end of a work shift (0.041 g/g creatinine) was elevated relative to that before the shift (0.027 g/g creatinine; p < 0.05). The GM of the CHZ metabolic ratio was 0.33 (0-9.3), with 40% of the subjects having ratios below the GM. However, the average CYP2E1 mRNA level in peripheral lymphocytes was 1.07 (0.30-3.08), and CYP2E1 mRNA levels within subjects correlated with the toluene exposure ratio (environmental toluene concentration:urinary hippuric acid concentration) (p = 0.014). Genotype did not alter the association between the toluene exposure ratio and mRNA content. In summary, with further validation, CYP2E1 mRNA content in peripheral lymphocytes could be a sensitive and noninvasive biomarker for the continuous monitoring of toluene effects in exposed persons.

  5. Amitriptyline induces brain-derived neurotrophic factor (BDNF) mRNA expression through ERK-dependent modulation of multiple BDNF mRNA variants in primary cultured rat cortical astrocytes and microglia.

    Science.gov (United States)

    Hisaoka-Nakashima, Kazue; Kajitani, Naoto; Kaneko, Masahiro; Shigetou, Takahiro; Kasai, Miho; Matsumoto, Chie; Yokoe, Toshiki; Azuma, Honami; Takebayashi, Minoru; Morioka, Norimitsu; Nakata, Yoshihiro

    2016-03-01

    A significant role of brain-derived neurotrophic factor (BDNF) has been previously implicated in the therapeutic effect of antidepressants. To ascertain the contribution of specific cell types in the brain that produce BDNF following antidepressant treatment, the effects of the tricyclic antidepressant amitriptyline on rat primary neuronal, astrocytic and microglial cortical cultures were examined. Amitriptyline increased the expression of BDNF mRNA in astrocytic and microglial cultures but not neuronal cultures. Antidepressants with distinct mechanisms of action, such as clomipramine, duloxetine and fluvoxamine, also increased BDNF mRNA expression in astrocytic and microglial cultures. There are multiple BDNF mRNA variants (exon I, IIA, IV and VI) expressed in astrocytes and microglia and the variant induced by antidepressants has yet to be elaborated. Treatment with antidepressants increased the expression of exon I, IV and VI in astrocyte and microglia. Clomipramine alone significantly upregulated expression of exon IIA. The amitriptyline-induced expression of both total and individual BDNF mRNA variants (exon I, IV and VI) were blocked by MEK inhibitor U0126, indicating MEK/ERK signaling is required in the expression of BDNF. These findings indicate that non-neural cells are a significant target of antidepressants and further support the contention that glial production of BDNF is crucial role in the therapeutic effect of antidepressants. The current data suggest that targeting of glial function could lead to the development of antidepressants with a truly novel mechanism of action.

  6. Associations among SPARC mRNA expression in adipose tissue, serum SPARC concentration and metabolic parameters in Korean women.

    Science.gov (United States)

    Lee, S H; Lee, J A; Park, H S; Song, Y S; Jang, Y J; Kim, J H; Lee, Y J; Heo, Y

    2013-11-01

    Secreted protein acidic and rich in cysteine (SPARC) is expressed in most tissues and is also secreted by adipocytes. The associations of SPARC mRNA expression in visceral adipose tissue (VAT), subcutaneous abdominal adipose tissue (SAT), serum SPARC concentration, and metabolic parameters in Korean women are investigated. This is a cross-sectional study. Fifty-eight women were recruited, of whom 15 women who underwent bariatric surgery for morbid obesity (BMI mean ± SD: 40.2±5.7 kg/m(2) ), 16 who underwent metabolic surgery for type 2 diabetes (BMI: 28.9±4.5 kg/m(2) ), and, as a control group, 27 who underwent gynecological surgery (BMI: 22.7±2.4 kg/m(2) ). Anthropometric variables, metabolic parameters, SPARC mRNA expression in adipose tissue, and serum SPARC concentration were measured. In all subjects, SPARC mRNA expression was significantly higher in SAT than in VAT. Serum SPARC concentrations (mean ± SE) in morbidly obese subjects, subjects with type 2 diabetes, and normal weight subjects were 267.3±40.2 ng/mL, 130.4±33.0 ng/mL, and 53.1±2.8 ng/mL, respectively. SPARC mRNA in SAT was significantly correlated with BMI, whereas SPARC mRNA in VAT was significantly correlated with BMI and VAT area. Serum SPARC concentration was significantly correlated with BMI, waist circumference, total adipose tissue area, and SAT area. After BMI adjustment, serum SPARC concentration was significantly correlated with fasting insulin concentration and HOMA-IR score. Multivariate regression analysis showed that BMI and HOMA-IR were independently associated with serum SPARC concentration. Serum SPARC concentration is significantly correlated with obesity indices and might be influenced by insulin resistance. These findings suggest that SPARC may contribute to the metabolic dysregulation associated with obesity in humans. Copyright © 2012 The Obesity Society.

  7. Expression of intestinal mucosa tight junctions claudin proteins and mRNA in patients with irritable bowel syndrome

    Institute of Scientific and Technical Information of China (English)

    KONG Wu-ming; GONG Jun; DONG Lei; LU Xiao-lan; XU Jun-rong

    2007-01-01

    Objective:To investigate the changes of intestinal mucosa tight junctions(TJs)claudin-1,-3,-4 proteins and mRNA changes in patients with irritable bowel syndrome(IBS)and to elucidate their possible roles in the changes of bowel evacuation habit and formation.Methods:Claudin-1,-3,-4 proteins and mRNA were evaluated in intestinal mucosa in control group,D-IBS(diarrhea IBS)group and C-IBS (constipation IBS)group with immunohistochemical assay and Realtime-PCR.Results:It was observed that claudin-1,-3,-4 proteins were localized in the membranes of epithelial cells along the entire length of the plasma membrane including the apical end of the epithelial cells.The claudins were concentrated at the site of TJs only.Claudin-1,-3,-4 mRNA and claudin-1 protein in small intestinal mucosa and colonal mucous in D-IBS group were significantly downregulated(P<0.05).Claudin-1,-3,-4 mRNA and proteins in small intestinal mucosa and co1onal mucous in C-IBS group were significantly upregulated(P<0.05).There was no significant difference in the expression of claudin-3 protein in both small intestinal mucosa and colonal mucous between D-IBS group and control group(P>0.05).Similarly,no significantly different expression of claudin-4 protein in colonal mucous in D-IBS group was found compared with control group(P>0.05).Otherwise,the expression of claudin-4 protein in small intestinal mucosa decreased in D-IBS group(P<0.05).Conclusion:Claudin-1,-3,-4 may play a potential important role in the changes of bowel evacuation habit and formation in patients with IBS.It is not due to the localization changes of claudin proteins in TJ,but may be caused by the quantitative changes of the expression of TJ proteins and mRNA.

  8. Cytokine mRNA Expression in Lesions in Cats with Chronic Gingivostomatitis

    OpenAIRE

    Harley, R.; Helps, C. R.; Harbour, D. A.; Gruffydd-Jones, T.J.; Day, M J

    1999-01-01

    Semiquantitative reverse transcription-PCR assays were developed to measure feline interleukin-2 (IL-2), IL-4, IL-5, IL-6, IL-10, and IL-12 (p35 & p40); gamma interferon (IFN-γ); and glyceraldehyde-3-phosphate dehydrogenase mRNA concentrations in biopsies of feline oral mucosa. Biopsies were collected from 30 cats with chronic gingivostomatitis (diseased) prior to each cat receiving one of four treatments. In 23 cases replicate biopsies were collected 3 months after tr...

  9. Heterologous Expression of Membrane and Soluble Proteins Derepresses GCN4 mRNA Translation in the Yeast Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Steffensen, L.; Pedersen, P. A.

    2006-01-01

    of the membrane-bound 1ß1 Na,K-ATPase from pig kidney, the rat pituitary adenylate cyclase seven-transmembrane-domain receptor, or a 401-residue soluble part of the Na,K-ATPase 1 subunit derepressed GCN4 mRNA translation up to 70-fold. GCN4 translation was very sensitive to the presence of heterologous protein......This paper describes the first physiological response at the translational level towards heterologous protein production in Saccharomyces cerevisiae. In yeast, the phosphorylation of eukaryotic initiation factor 2 (eIF-2 ) by Gcn2p protein kinase mediates derepression of GCN4 mRNA translation. Gcn4......, as a density of 1 of heterologous membrane protein derepressed translation maximally. Translational derepression of GCN4 was not triggered by misfolding in the endoplasmic reticulum, as expression of the wild type or temperature-sensitive folding mutants of the Na,K-ATPase increased GCN4 translation...

  10. Upregulation of aggrecan and type II collagen mRNA expression in bovine chondrocytes by the application of hydrostatic pressure.

    Science.gov (United States)

    Toyoda, Takashi; Seedhom, Bahaa B; Kirkham, Jennifer; Bonass, William A

    2003-01-01

    The aim of this study was to investigate the effect of hydrostatic pressure on the expression of messenger ribonucleic acid (mRNA) for specific extracellular matrix proteins in chondrocytes. Chondrocytes obtained from bovine metatarsophalangeal joints were embedded in cylindrical 2% agarose gels. A novel experimental system was used to apply 5 MPa of static hydrostatic pressure to these chondrocytes for 4 hours. The application of hydrostatic pressure caused a significant increase in the level of aggrecan mRNA by almost four fold (papplication of hydrostatic pressure, in the absence of cell deformation, can bring about changes in the matrix components which may play an important role in the homeostasis and mechanical properties of articular cartilage.

  11. Effect of siRNAs on HSV-1 Plaque Formation and Relative Expression Levels of RR mRNA

    Institute of Scientific and Technical Information of China (English)

    Zhe Ren; Pei-zhuo Zhang; Shen Li; Qiao-li Wang; Yang-fei Xiang; Yun-xia Cui; Yi-fei Wang; Ren-bin Qi; Da-xiang Lu; Shu-min Zhang

    2011-01-01

    RNA interference(RNAi)is a process by which introduced small interfering RNA(siRNA)can cause the specific degradation of mRNA with identical sequences. The human herpes simplex virus type 1(HSV-1)RR is composed of two distinct homodimeric subunits encoded by UL39 and UL40, respectively. In this study, we applied siRNAs targeting the UL39 and UL40 genes of HSV-1. We showed that synthetic siRNA silenced effectively and specifically UL39 and UL40 mRNA expression and inhibited HSV-1 replication. Our work offers new possibilities for RNAi as a genetic tool for inhibition of HSV-1 replication.

  12. Complex control of GABA(A receptor subunit mRNA expression: variation, covariation, and genetic regulation.

    Directory of Open Access Journals (Sweden)

    Megan K Mulligan

    Full Text Available GABA type-A receptors are essential for fast inhibitory neurotransmission and are critical in brain function. Surprisingly, expression of receptor subunits is highly variable among individuals, but the cause and impact of this fluctuation remains unknown. We have studied sources of variation for all 19 receptor subunits using massive expression data sets collected across multiple brain regions and platforms in mice and humans. Expression of Gabra1, Gabra2, Gabrb2, Gabrb3, and Gabrg2 is highly variable and heritable among the large cohort of BXD strains derived from crosses of fully sequenced parents--C57BL/6J and DBA/2J. Genetic control of these subunits is complex and highly dependent on tissue and mRNA region. Remarkably, this high variation is generally not linked to phenotypic differences. The single exception is Gabrb3, a locus that is linked to anxiety. We identified upstream genetic loci that influence subunit expression, including three unlinked regions of chromosome 5 that modulate the expression of nine subunits in hippocampus, and that are also associated with multiple phenotypes. Candidate genes within these loci include, Naaa, Nos1, and Zkscan1. We confirmed a high level of coexpression for subunits comprising the major channel--Gabra1, Gabrb2, and Gabrg2--and identified conserved members of this expression network in mice and humans. Gucy1a3, Gucy1b3, and Lis1 are novel and conserved associates of multiple subunits that are involved in inhibitory signaling. Finally, proximal and distal regions of the 3' UTRs of single subunits have remarkably independent expression patterns in both species. However, corresponding regions of different subunits often show congruent genetic control and coexpression (proximal-to-proximal or distal-to-distal, even in the absence of sequence homology. Our findings identify novel sources of variation that modulate subunit expression and highlight the extraordinary capacity of biological networks to buffer

  13. Integrated Analysis of Dysregulated ncRNA and mRNA Expression Profiles in Humans Exposed to Carbon Nanotubes.

    Directory of Open Access Journals (Sweden)

    Anna A Shvedova

    Full Text Available As the application of carbon nanotubes (CNT in consumer products continues to rise, studies have expanded to determine the associated risks of exposure on human and environmental health. In particular, several lines of evidence indicate that exposure to multi-walled carbon nanotubes (MWCNT could pose a carcinogenic risk similar to asbestos fibers. However, to date the potential markers of MWCNT exposure are not yet explored in humans.In the present study, global mRNA and ncRNA expression profiles in the blood of exposed workers, having direct contact with MWCNT aerosol for at least 6 months (n = 8, were compared with expression profiles of non-exposed (n = 7 workers (e.g., professional and/or technical staff from the same manufacturing facility.Significant changes in the ncRNA and mRNA expression profiles were observed between exposed and non-exposed worker groups. An integrative analysis of ncRNA-mRNA correlations was performed to identify target genes, functional relationships, and regulatory networks in MWCNT-exposed workers. The coordinated changes in ncRNA and mRNA expression profiles revealed a set of miRNAs and their target genes with roles in cell cycle regulation/progression/control, apoptosis and proliferation. Further, the identified pathways and signaling networks also revealed MWCNT potential to trigger pulmonary and cardiovascular effects as well as carcinogenic outcomes in humans, similar to those previously described in rodents exposed to MWCNTs.This study is the first to investigate aberrant changes in mRNA and ncRNA expression profiles in the blood of humans exposed to MWCNT. The significant changes in several miRNAs and mRNAs expression as well as their regulatory networks are important for getting molecular insights into the MWCNT-induced toxicity and pathogenesis in humans. Further large-scale prospective studies are necessary to validate the potential applicability of such changes in mRNAs and miRNAs as prognostic markers

  14. Promoter methylation and mRNA expression of HLA-G in relation to HLA-G protein expression in colorectal cancer.

    Science.gov (United States)

    Swets, Marloes; Seneby, Lina; Boot, Arnoud; van Wezel, Tom; Gelderblom, Hans; van de Velde, Cornelis J H; van den Elsen, Peter J; Kuppen, Peter J K

    2016-09-01

    Expression of human leukocyte antigen-G (HLA-G) is a suggested mechanism used by tumor cells to escape from host immune recognition and destruction. Advances in the field have made it evident that HLA-G is expressed in different types of malignancies including colorectal cancer (CRC). We analyzed HLA-G expression in 21 low passage CRC cell lines. The level of DNA methylation of the HLA-G gene and the presence of mRNA encoding HLA-G was measured. Moreover, HLA-G protein expression was determined by flow cytometry and immunohistochemistry (IHC). IHC was performed with three different monoclonal antibodies (mAbs) (4H84, MEM-G/1 and MEM-G/2). In addition, HLA-G protein expression was measured in matching primary tumor tissues. RNA analysis using RT-PCR followed by sequencing in 6 samples indicated strong homology of the PCR product with HLA-G3 in 5 samples. In accordance, in none of the cell lines, HLA-G1 expression was detected by flow-cytometry. Furthermore, no association between HLA-G DNA methylation patterns and HLA-G mRNA expression was observed. In addition, different immunohistochemical staining profiles among various anti-HLA-G mAbs were observed. In conclusion, the results of this study show that the HLA-G3 isoform was expressed in some of the CRC cell lines irrespective of the level of DNA methylation of HLA-G.

  15. Expression of TLR2 mRNA and TLR4 mRNA in psoriatic lesions%TLR2 mRNA和TLR4 mRNA在银屑病皮损中的表达

    Institute of Scientific and Technical Information of China (English)

    吴娜; 郑焱; 郗彦萍

    2011-01-01

    Objective To detect the expression of toll-like receptor 2 (TLR2) mRNA and toll-like receptor 4 (TLR4) mRNA in psoriatic skin lesions. Methods By using in situ hybridization, TLR2 mRNA and TLR4 mRNA expression levels were detected in psoriatic lesional and normal skin tissues. Results TLR2 mRNA and TLR4 mRNA were highly expressed in the suprabasal layer of psoriatic lesions, while in normal epidermis TLR2 mRNA and TLR4 mRNA were expressed in the basal layer of epidermis. There was a significant difference between psoriatic lesion and the normal epiderm (P<0.05), and TLR2 mRNA and TLR4 mRNA expressions were higher in psoriatic lesional epidermis than in the normal epidermis. Conclusion TLR2 and TLR4 may play a crucial role in the pathogenesis and development of psoriasis.%目的 探讨Toll样受体2(TLR2)mRNA和Toll样受体4(TLR4)mRNA在银屑病皮损中表达的变化.方法 采用原位杂交技术检测银屑病皮损处和正常皮肤中TLR2 mRNA和TLR4m RNA的表达情况.结果 寻常型银屑病皮损中TLR2 mRNA和TLR4 mRNA主要表达于表皮基底上层;正常皮肤则表达于表皮的基底层.统计学分析表明,TLR2 mRNA和TLR4 mRNA在正常皮肤和银屑病皮损处基底层和基底上层的表达差异均有统计学意义(P<0.05),且TLR2 mRNA和TLR4 mRNA在银屑病皮损中的表达均高于正常皮肤组织(P<0.05).结论 TLR2和TLR4可能在银屑病的发生与发展过程中发挥重要的作用.

  16. mRNA expression levels in failing human hearts predict cellular electrophysiological remodeling: a population-based simulation study.

    Directory of Open Access Journals (Sweden)

    John Walmsley

    Full Text Available Differences in mRNA expression levels have been observed in failing versus non-failing human hearts for several membrane channel proteins and accessory subunits. These differences may play a causal role in electrophysiological changes observed in human heart failure and atrial fibrillation, such as action potential (AP prolongation, increased AP triangulation, decreased intracellular calcium transient (CaT magnitude and decreased CaT triangulation. Our goal is to investigate whether the information contained in mRNA measurements can be used to predict cardiac electrophysiological remodeling in heart failure using computational modeling. Using mRNA data recently obtained from failing and non-failing human hearts, we construct failing and non-failing cell populations incorporating natural variability and up/down regulation of channel conductivities. Six biomarkers are calculated for each cell in each population, at cycle lengths between 1500 ms and 300 ms. Regression analysis is performed to determine which ion channels drive biomarker variability in failing versus non-failing cardiomyocytes. Our models suggest that reported mRNA expression changes are consistent with AP prolongation, increased AP triangulation, increased CaT duration, decreased CaT triangulation and amplitude, and increased delay between AP and CaT upstrokes in the failing population. Regression analysis reveals that changes in AP biomarkers are driven primarily by reduction in I[Formula: see text], and changes in CaT biomarkers are driven predominantly by reduction in I(Kr and SERCA. In particular, the role of I(CaL is pacing rate dependent. Additionally, alternans developed at fast pacing rates for both failing and non-failing cardiomyocytes, but the underlying mechanisms are different in control and heart failure.

  17. Synaptic adaptations by alcohol and drugs of abuse: changes in microRNA expression and mRNA regulation

    Directory of Open Access Journals (Sweden)

    Dana eMost

    2014-12-01

    Full Text Available Local translation of mRNAs is a mechanism by which cells can rapidly remodel synaptic structure and function. There is ample evidence for a role of synaptic translation in the neuroadaptations resulting from chronic drug use and abuse. Persistent and coordinated changes of many mRNAs, globally and locally, may have a causal role in complex disorders such as addiction. In this review we examine the evidence that translational regulation by microRNAs drives synaptic remodeling and mRNA expression, which may regulate the transition from recreational to compulsive drug use.MicroRNAs are small, non-coding RNAs that control the translation of mRNAs in the cell and within spatially restricted sites such as the synapse. MicroRNAs typically repress the translation of mRNAs into protein by binding to the 3’UTR of their targets. As ‘master regulators’ of many mRNAs, changes in microRNAs could account for the systemic alterations in mRNA and protein expression observed with drug abuse and dependence. Recent studies indicate that manipulation of microRNAs affects addiction-related behaviors such as the rewarding properties of cocaine, cocaine-seeking behavior and self-administration rates of alcohol. There is limited evidence, however, regarding how synaptic microRNAs control local mRNA translation during chronic drug exposure and how this contributes to the development of dependence.Here, we discuss research supporting microRNA regulation of local mRNA translation and how drugs of abuse may target this process. The ability of synaptic microRNAs to rapidly regulate mRNAs provides a discrete, localized system that could potentially be used as diagnostic and treatment tools for alcohol and other addiction disorders.

  18. Increased IL-17 and 22 mRNA expression in pediatric patients with otitis media with effusion.

    Science.gov (United States)

    Kwon, Oh Eun; Park, Sang Hyun; Kim, Sung Su; Shim, Haeng Seon; Kim, Min Gyeong; Kim, Young Il; Kim, Sang Hoon; Yeo, Seung Geun

    2016-11-01

    Middle ear effusion has been reported to be associated with immune responses in patients with otitis media with effusion (OME). Although various cytokines are involved in immunologic responses in patients with OME, no study to date has assessed the involvement of the pro-inflammatory cytokines interleukin (IL)-17 and IL-22. This study analyzed the levels of expression of IL-17 and IL-22 in the middle ear effusion of patients with OME. Patients aged <11 years who were diagnosed with chronic OME and underwent ventilation tube insertion from May 2013 to August 2015 were enrolled. Effusion fluid samples were obtained during surgery and levels of IL-17 and IL-22 mRNAs assessed by real-time PCR. IL-17 and IL-22 mRNA levels were compared in patients with effusion fluid positive and negative for bacteria; in patients with and without accompanying diseases, recurrent disease, and re-operation; and relative to fluid characteristics. The study cohort included 70 pediatric patients, 46 boys and 24 girls, of mean age 4.31 ± 2.11 years. The levels of IL-17 and IL-22 mRNA were higher in patients with than without sinusitis, but only IL-22 mRNA levels differed significantly (p < 0.05). The level of IL-17 mRNA was significantly higher in patients who did than did not undergo T&A (p < 0.05). The level of IL-22 expression was significantly higher in mucoid and purulent middle ear fluid samples than in serous fluid samples (p < 0.05). IL-17 and IL-22 mRNAs are involved in the pathophysiology of OME and are significantly higher in subjects with than without accompanying diseases. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  19. Enhancing effect of taurine on CYP7A1 mRNA expression in Hep G2 cells.

    Science.gov (United States)

    Lam, N V; Chen, W; Suruga, K; Nishimura, N; Goda, T; Yokogoshi, H

    2006-02-01

    Taurine has been reported to enhance cholesterol 7alpha-hydroxylase (CYP7A1) mRNA expression in animal models. However, no in vitro studies of this effect have been reported. The Hep G2 human hepatoma cell line has been recognized as a good model for studying the regulation of human CYP7A1. This work characterizes the effects of taurine on CYP7A1 mRNA levels of Hep G2 cells in a dose- and time-dependent manner. In the dose-dependent experiment, Hep G2 cells were treated with 0, 2, 10 or 20 mM taurine in the presence or absence of cholesterol 0.2 mM for 48 h. In the time-dependent experiment, Hep G2 cells were treated with 0 or 20 mM taurine for 4, 24 and 48 h with and without cholesterol 0.2 mM. Our data revealed that taurine showed time- and dose-response effects on CYP7A1 mRNA levels in Hep G2 cells. However, glycine - a structural analogue of taurine - did not have an effect on CYP7A1 gene expression. These results show that, in agreement to previous studies on animal models, taurine induces the mRNA levels of CYP7A1 in Hep G2 cells, which could enhance cholesterol conversion into bile acids. Also, Hep G2 cell line may be an appropriate model to study the effects of taurine on human cholesterol metabolism.

  20. Biological analysis of chronic lymphocytic leukemia: integration of mRNA and microRNA expression profiles.

    Science.gov (United States)

    Dong, L; Bi, K H; Huang, N; Chen, C Y

    2016-01-08

    Chronic lymphocytic leukemia (CLL) is a disease that involves progressive accumulation of nonfunctioning lymphocytes and has a low cure rate. There is an urgent requirement to determine the molecular mechanism underlying this disease in order to improve the early diagnosis and treatment of CLL. In this study, genes differentially expressed between CLL samples and age-matched controls were identified using microRNA (miRNA) and mRNA expression profiles. Differentially expressed (DE) miRNA targets were predicted by combining five algorithms. Common genes were obtained on overlapping the DE mRNA and DE miRNA targets. Then, network and module analyses were performed. A total of 239 miRNA targets were predicted and 357 DE mRNAs were obtained. On intersecting miRNA targets and DE mRNAs, 33 common genes were obtained. The protein-protein interaction network and module analysis identified several crucial genes and modules that might be associated with the development of CLL. These DE mRNAs were significantly enriched in the hematopoietic cell lineage (P = 2.58E-4), mitogen-activated protein kinase signaling pathway (P = 0.0025), and leukocyte transendothelial migration pathway (P = 0.0026). Thus, we conducted biological analysis on integration of DE mRNAs and DE miRNAs in CLL, determined gene expression patterns, and screened out several important genes that might be related to CLL.

  1. The effect of anastrozole on mRNA expression of oestrogen related gene in MCF-7 breast cancer cells

    Institute of Scientific and Technical Information of China (English)

    SONG Zhang-jun; WU Yi; MA Qing-yong

    2006-01-01

    Objective: To look for additional markers of molecular biology response to anastrozole, a new aromatase inhibitor, on the growth and mRNA expression level of MCF-7 cell. Methods: We investigated the effect of anastrzole on growth and gene expression in the human breast cancer cell line MCF-7and compared with the most widely used antiestrogen tamoxifen. We chose 4 genes to examine regulation of gene expression of estrogen regulated genes: PR A, PR B, ErbB-2 and cyclin D1. Results: Compared with the tamoxifen, a statistically significant growth inhibition was observed with anastrozole. The PRA,PR B and cyclin D1 mRNA level in anastrozole treated cells was sigificantly below the level in tamoxifen treated cells (P<0. 05). They had agonistic effect on ErbB gene (P>0.05). Conclusion: The third generation of aromatase inhibitors anastrozole exert more inhibit function in some expression of estrogen regulated genes than tomoxifen in MCF-7 cell line.

  2. Transforming Growth Factor β1 (TGF-β1) Activates Hepcidin mRNA Expression in Hepatocytes.

    Science.gov (United States)

    Chen, Simeng; Feng, Teng; Vujić Spasić, Maja; Altamura, Sandro; Breitkopf-Heinlein, Katja; Altenöder, Jutta; Weiss, Thomas S; Dooley, Steven; Muckenthaler, Martina U

    2016-06-17

    The hepatic hormone hepcidin is the master regulator of systemic iron homeostasis. Its expression level is adjusted to alterations in iron levels, inflammatory cues, and iron requirements for erythropoiesis. Bone morphogenetic protein 6 (BMP6) contributes to the iron-dependent control of hepcidin. In addition, TGF-β1 may stimulate hepcidin mRNA expression in murine hepatocytes and human leukocytes. However, receptors and downstream signaling proteins involved in TGF-β1-induced hepcidin expression are still unclear. Here we show that TGF-β1 treatment of mouse and human hepatocytes, as well as ectopic expression of TGF-β1 in mice, increases hepcidin mRNA levels. The hepcidin response to TGF-β1 depends on functional TGF-β1 type I receptor (ALK5) and TGF-β1 type II receptor (TβRII) and is mediated by a noncanonical mechanism that involves Smad1/5/8 phosphorylation. Interestingly, increasing availability of canonical Smad2/3 decreases TGF-β1-induced hepcidin regulation, whereas the BMP6-hepcidin signal was enhanced, indicating a signaling component stoichiometry-dependent cross-talk between the two pathways. Although ALK2/3-dependent hepcidin activation by BMP6 can be modulated by each of the three hemochromatosis-associated proteins: HJV (hemojuvelin), HFE (hemochromatosis protein), and TfR2 (transferrin receptor 2), these proteins do not control the ALK5-mediated hepcidin response to TGF-β1. TGF-β1 mRNA levels are increased in mouse models of iron overload, indicating that TGF-β1 may contribute to hepcidin synthesis under these conditions. In conclusion, these data demonstrate that a complex regulatory network involving TGF-β1 and BMP6 may control the sensing of systemic and/or hepatic iron levels.

  3. Characterizing dose-responses of catalase to nitrofurazone exposure in model ciliated protozoan Euplotes vannus for ecotoxicity assessment: enzyme activity and mRNA expression.

    Science.gov (United States)

    Li, Jiqiu; Zhou, Liang; Lin, Xiaofeng; Yi, Zhenzhen; Al-Rasheid, Khaled A S

    2014-02-01

    In environmental studies, some biological responses, known as biomarkers, have been used as a powerful bioassay tool for more than four decades. Disparity between enzyme activity and mRNA abundance leads to correlation equivocality, which makes the application of biomarkers for environmental risk assessment more complicated. This study investigates this disparity in the case of catalase when used as a biomarker for detecting ecotoxicity induced by antibiotics in aquatic ecosystems. In particular, dose-responses for catalase activity and mRNA expression abundance were investigated in Euplotes vannus which were exposed to graded doses of nitrofurazone for several discrete durations, and dose-response models were developed to characterize the dose-response dynamics. Significant differences were found in both catalase activity and mRNA expression abundance among the E. vannus treated with nitrofurazone. Catalase activity showed a hormetic-like effect in terms of dose-response, characterized by a biphasic relationship which was more clearly evident after a longer exposure period, while mRNA expression abundance increased linearly with the exposure duration. Additionally, the correlation between catalase activity and mRNA expression abundance reversed along with the duration of exposure to nitrofurazone. Taken together, our results demonstrate that catalase mRNA expression offers a more straightforward dose-response model than enzyme activity. Our findings suggest that both catalase enzyme activity and mRNA expression abundance can be used jointly as bioassay tools for detecting ecotoxicity induced by nitrofurazone in aquatic ecosystems.

  4. Selective cognitive deficits and reduced hippocampal brain-derived neurotrophic factor mRNA expression in small-conductance calcium-activated K+ channel deficient mice

    DEFF Research Database (Denmark)

    Jacobsen, J P R; Redrobe, J P; Hansen, H H

    2009-01-01

    (BDNF) mRNA expression in the dentate gyrus and CA3 of the hippocampus. BDNF mRNA levels in the frontal cortex were not affected. BDNF has been crucially implicated in many cognitive processes. Hence, the biological substrate for the cognitive impairments in T/T mice could conceivably entail reduced...

  5. Posttranscriptional regulation of GAP-43 gene expression in PC12 cells through protein kinase C-dependent stabilization of the mRNA.

    Science.gov (United States)

    Perrone-Bizzozero, N I; Cansino, V V; Kohn, D T

    1993-03-01

    We have previously shown that nerve growth factor (NGF) selectively stabilizes the GAP-43 mRNA in PC12 cells. To study the cellular mechanisms for this post-transcriptional control and to determine the contribution of mRNA stability to GAP-43 gene expression, we examined the effects of several agents that affect PC12 cell differentiation on the level of induction and rate of degradation of the GAP-43 mRNA. The NGF-mediated increase in GAP-43 mRNA levels and neurite outgrowth was mimicked by the phorbol ester TPA, but not by dibutyryl cAMP or the calcium ionophore A12783. Downregulation of protein kinase C (PKC) by high doses of phorbol esters or selective PKC inhibitors prevented the induction of this mRNA by NGF, suggesting that NGF and TPA act through a common PKC-dependent pathway. In mRNA decay studies, phorbol esters caused a selective 6-fold increase in the half-life of the GAP-43 mRNA, which accounts for most of the induction of this mRNA by TPA. The phorbol ester-induced stabilization of GAP-43 mRNA was blocked by the protein kinase inhibitor polymyxin B and was partially inhibited by dexamethasone, an agent that blocks GAP-43 expression and neuronal differentiation in PC12 cells. In contrast, the rates of degradation and the levels of the GAP-43 mRNA in control and TPA-treated cells were not affected by cycloheximide treatment. Thus, changes in GAP-43 mRNA turnover do not appear to require continuous protein synthesis. In conclusion, these data suggest that PKC activity regulates the levels of the GAP-43 mRNA in PC12 cells through a novel, translation-independent mRNA stabilization mechanism.

  6. Benzo(a)pyrene induces hepatic AKR1A1 mRNA expression in tilapia fish (Oreochromis niloticus).

    Science.gov (United States)

    Osorio-Yáñez, Citlalli; García-Tavera, José Luis; Pérez-Núñez, Maria Teresa; Poblete-Naredo, Irais; Muñoz, Balam; Barron-Vivanco, Briscia S; Rothenberg, Stephen J; Zapata-Pérez, Omar; Albores, Arnulfo

    2012-07-01

    AKR1A1 or aldehyde reductase is a member of the aldo-keto reductases superfamily that is evolutionarily conserved among species. AKR1A1 is one of the five AKRs (AKR1A1 and 1C1-1C4) implicated in the metabolic benzo(a)pyrene (BaP) activation to reactive BaP 7,8-dione. BaP is a polycyclic aromatic hydrocarbon (PAH) widely distributed in aquatic ecosystems and its metabolic activation is necessary to produce its toxic effects. Although the presence of AKR1A1 in fish has been reported, its tissue distribution in tilapia (Oreochromis niloticus) and AKR1A1 inducibility by BaP are not known yet. Moreover, cytochrome P4501A (CYP1A) mRNA expression in fish has been used as a PAH biomarker of effect. Therefore, BaP effects on AKR1A1 and CYP1A gene expressions in tilapia, a species of commercial interest, were investigated by real-time RT-PCR. A partial AKR1A1 cDNA was identified, sequenced and compared with AKR1A1 reported sequences in the GenBank DNA database. Constitutive AKR1A1 mRNA expression was detected mainly in liver, similarly to that of CYP1A. BaP exposure resulted in statistically significant AKR1A1 and CYP1A mRNA induction in liver (20- and 120-fold, respectively) at 24 h. On the other hand, ethoxyquin (EQ) was used as control inducer for AKR1A1 mRNA. Interestingly, EQ also induced CYP1A mRNA levels in tilapia liver. Our results suggest that teleost AKR1A1, in addition to CYP1A, are inducible by BaP. The mechanism of AKR1A1 induction by BaP and its role in fish susceptibility to BaP toxic effects remains to be elucidated.

  7. Effect of experimental treatment on GAPDH mRNA expression as a housekeeping gene in human diploid fibroblasts

    Directory of Open Access Journals (Sweden)

    Zainuddin Azalina

    2010-08-01

    Full Text Available Abstract Background Several genes have been used as housekeeping genes and choosing an appropriate reference gene is important for accurate quantitative RNA expression in real time RT-PCR technique. The expression levels of reference genes should remain constant between the cells of different tissues and under different experimental conditions. The purpose of this study was to determine the effect of different experimental treatments on the expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH mRNA so that the reliability of GAPDH as reference gene for quantitative real time RT-PCR in human diploid fibroblasts (HDFs can be validated. HDFs in 4 different treatment groups viz; young (passage 4, senescent (passage 30, H2O2-induced oxidative stress and γ-tocotrienol (GTT-treated groups were harvested for total RNA extraction. Total RNA concentration and purity were determined prior to GAPDH mRNA quantification. Standard curve of GAPDH expression in serial diluted total RNA, melting curve analysis and agarose gel electrophoresis were used to determine the reliability of GAPDH as reference gene. Results HDFs with different experimental treatments exhibited diverse cell morphology with different expression of senescence-associated β-galactosidase (SA β-gal activity. However the expression level of GAPDH was consistent in all treatment groups. Conclusion The study demonstrated that GAPDH is reliable as reference gene for quantitative gene expression analysis in HDFs. Therefore it can be used as housekeeping gene for quantitative real time RT-PCR technique in human diploid fibroblasts particularly in studying cellular senescence.

  8. Effect of acidic ribosomal phosphoprotein mRNA 5'-untranslated region on gene expression and protein accumulation.

    Science.gov (United States)

    Bermejo, B; Remacha, M; Ortiz-Reyes, B; Santos, C; Ballesta, J P

    1994-02-11

    Constructions were made from genes encoding ribosomal acidic phosphoproteins YP1 beta (L44') and YP2 beta (L45) from Saccharomyces cerevisiae in which different parts of the 5'-untranslated regions were included. The constructs were inserted into centromeric plasmids under the control of the GAL1 promoter and expressed in yeast strains in which the genes coding for each acidic protein family, P1 and P2, had been disrupted. Deletions in the 5' region of the two genes have been found to oppositely affect their expression. Deletion of most of this region strongly stimulates the expression of YP2 beta (L45), increasing the translation efficiency of the mRNA, and generating a 6-fold excess of protein in the cell. A similar deletion in the rpYP1 beta gene represses the expression of the protein, reducing drastically the amount of the mRNA in the cell. The overexpression of rpYP2 beta affects the cell growth by inhibiting protein synthesis at the level of initiation. Reduction of the YP2 beta(L45) overproduction by growing in controlled concentrations of glucose abolishes the inhibitory effect. The excess protein, probably as a high molecular weight complex, apparently interferes with the joining of the 60 S subunit to the initiation complex generating the accumulation of polysome half-mers. In addition, the results indicate the existence of a regulatory mechanism by which each one of the two acidic proteins controls the expression of the other polypeptide. YP1 beta(L44') represses the expression of YP2 beta(L45), while this protein stimulates the expression of YP1 beta(L44').

  9. Regulation of FAT/CD36 mRNA gene expression by long chain fatty acids in the differentiated 3T3-L1 cells.

    Science.gov (United States)

    Yang, Yingkui; Chen, Min; Loux, Tara J; Harmon, Carroll M

    2007-07-01

    Defects in fatty acid translocase (FAT/CD36) have been identified as a major factor in insulin resistance and defective fatty acid and glucose metabolism. Therefore, understanding of the regulation of FAT/CD36 expression and function is important for a potential therapeutic target for type II diabetes. We differentiated 3T3-L1 preadipocytes into matured adipocytes and examined the roles of insulin and long chain fatty acids on FAT/CD36 expression and function. Our results indicate that FAT/CD36 mRNA expression was not detected at preadipocyte but was significantly increased at matured adipocyte. In fully differentiated 3T3-L1 adipocytes, insulin significantly increased FAT/CD36 mRNA and protein expression in a dose dependent manner. The free fatty acid stearic acid reduced FAT/CD36 mRNA expression while the non-metabolizable free fatty acid alpha-bromopalmitate (2-BP) significantly increased FAT/CD36 mRNA and protein expression. Isoproterenol, in contrast, dose-dependently reduced FAT/CD36 mRNA expression and increased free fatty acid release. Mechanism analysis indicated that the effect of insulin and 2-BP on the FAT/CD36 mRNA gene expression may be mediated through activation of PPAR-gamma, suggesting that FAT/CD36 may have important implications in the pathophysiology of defective fatty acid metabolism.

  10. The effects of pilates exercise on lipid metabolism and inflammatory cytokines mRNA expression in female undergraduates.

    Science.gov (United States)

    Kim, Hyo-Jin; Kim, Jiyeon; Kim, Chang-Sun

    2014-09-01

    The purpose of the study was to verify the effects of Pilates exercise by observing the impact of 8 weeks of Pilates exercise on lipid metabolism and inflammatory cytokine mRNA expression in female undergraduates in their 20s who had no prior experience in Pilates exercise and had not exercised in the previous 6 months. There were 18 subjects with no prior experience in Pilates exercise. The subjects were separated into the Pilates exercise group (n = 9) and the non-exercise control group (n = 9). The former performed Pilates exercise for 60-70 minutes over 8 weeks with a gradual strength increase of 9-16 in the Rating of Perceived Exercise (RPE). The body composition, creatine kinase in the bloodstream and lipid metabolism (TC, LDL-C, HDL-C, TG) were measured before and after the experiment and Real-Time PCR was used to investigate the mRNA expression of the inflammatory cytokines IL-6 and TNF-⍺. The creatine kinase (CK) in the blood had significant diffe