Koreksi gigitan terbalik posterior dan anterior dengan alat cekat rapid maxillary expansion dan elastik intermaksila

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Retno Dewati

2014-06-01

Full Text Available Background: Children with anterior and posterior crossbite usually have a complaint in aesthetic and masticatory function. It could caused by bad habits and hereditary factors which made worse condition. Purpose: The purpose of this case report was to report the use of orthodontic appliance rapid maxillary expansion (RPE and intermaxillary elastic to correct posterior and anterior crossbite in teenage patient. Case: A fourteen years-old teenage female patient came to Dental Hospital Dentistry Universitas Airlangga with case of anterior posterior cross bite and unerupted permanent teeth. Case management: The case was treated using orthodontic fixed appliance rapid maxillary expansion (RPE and followed by intermaxillary elastics. The posterior cross bite treatment took 4 weeks used of orthodontic fixed appliance RPE, while, treatment of anterior cross bite which used intermaxillary elactic was done within three month to achieved normal occlusion. Conclusion: This case report showed that the orthodontic appliance rapid maxillary expansion (RPE and intermaxillary elastic could be used to correct posterior and anterior crossbite.Latar belakang: Anak dengan gigitan terbalik anterior dan posterior pada umumnya mempunyai keluhan dalam hal estetik dan fungsi pengunyahan. Kondisi gigitan terbalik biasanya disebabkan oleh adanya kebiasaan buruk dan faktor keturunan yang semakin memperparah keadaan tersebut. Tujuan: Laporan kasus ini melaporkan pemakaian alat cekat rapid maxillary expansion (RPE dan elastik intermaksila untuk mengkoreksi gigitan terbalik posterior dan anterior pada anak remaja. Kasus: Pasien remaja perempuan berusia 14 tahun datang ke Rumah Sakit Gigi dan Mulut Fakultas Kedokteran Gigi Universitas Airlangga Surabaya dengan kasus gigitan terbalik anterior posterior dan terdapat gigi permanen yang tidak tumbuh. Tatalaksana kasus: Perawatan yang dilakukan adalah koreksi gigitan terbalik dengan menggunakan alat ortodonsia cekat rapid maxillary

Cell Expansion-Dependent Inflammatory and Metabolic Profile of Human Bone Marrow Mesenchymal Stem Cells.

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Prieto, Patricia; Fernández-Velasco, María; Fernández-Santos, María E; Sánchez, Pedro L; Terrón, Verónica; Martín-Sanz, Paloma; Fernández-Avilés, Francisco; Boscá, Lisardo

2016-01-01

Stem cell therapy has emerged as a promising new area in regenerative medicine allowing the recovery of viable tissues. Among the many sources of adult stem cells, bone marrow-derived are easy to expand in culture via plastic adherence and their multipotentiality for differentiation make them ideal for clinical applications. Interestingly, several studies have indicated that MSCs expansion in vitro may be limited mainly due to "cell aging" related to the number of cell divisions in culture. We have determined that MSCs exhibit a progressive decline across successive passages in the expression of stem cell markers, in plasticity and in the inflammatory response, presenting low immunogenicity. We have exposed human MSCs after several passages to TLRs ligands and analyzed their inflammatory response. These cells responded to pro-inflammatory stimuli (i.e., NOS-2 expression) and to anti-inflammatory cytokines (i.e., HO1 and Arg1) until two expansions, rapidly declining upon subculture. Moreover, in the first passages, MSCs were capable to release IL1β, IL6, and IL8, as well as to produce active MMPs allowing them to migrate. Interestingly enough, after two passages, anaerobic glycolysis was enhanced releasing high levels of lactate to the extracellular medium. All these results may have important implications for the safety and efficacy of MSCs-based cell therapies.

EDAG promotes the expansion and survival of human CD34+ cells.

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Ke Zhao

Full Text Available EDAG is multifunctional transcriptional regulator primarily expressed in the linloc-kit+Sca-1+ hematopoietic stem cells (HSC and CD34+ progenitor cells. Previous studies indicate that EDAG is required for maintaining hematopoietic lineage commitment balance. Here using ex vivo culture and HSC transplantation models, we report that EDAG enhances the proliferative potential of human cord blood CD34+ cells, increases survival, prevents cell apoptosis and promotes their repopulating capacity. Moreover, EDAG overexpression induces rapid entry of CD34+ cells into the cell cycle. Gene expression profile analysis indicate that EDAG knockdown leads to down-regulation of various positive cell cycle regulators including cyclin A, B, D, and E. Together these data provides novel insights into EDAG in regulation of expansion and survival of human hematopoietic stem/progenitor cells.

Modified hyrax splint for rapid maxillary expansion in esthetically concerned patients

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Sarabjeet Singh Sandhu

2015-01-01

Full Text Available The orthodontic treatment of Class III malocclusion with a maxillary deficiency is often treated with maxillary protraction either with or without maxillary expansion. The routine procedure for rapid maxillary expansion includes banding on first premolars/first deciduous molars and the permanent first molars. However in some patients who are esthetically very conscious, banding of the first premolar would not be a good esthetic option. So for such circumstances we have designed a modified hyrax splint, which does not need the first premolars to be banded.

Ex-vivo expansion of nonhuman primate CD34+ cells by stem cell factor Sall4B

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Bin Shen

2016-10-01

Full Text Available Abstract Background Hematopoietic CD34+ stem cells are widely used in the clinical therapy of complicated blood diseases. Stem cell factor Sall4B is a zinc finger transcription factor that plays a vital role in hematopoietic stem cell expansion. The purpose of our current study is to further evaluate how Sall4B might affect the expansion of CD34+ cells derived from nonhuman primates. Methods Sall4B was overexpressed in nonhuman primate bone marrow-derived CD34+ cells via a lentiviral transduction system. The granulocyte–erythrocyte–macrophage–megakaryocyte colony-forming unit (CFU assay evaluated the differentiation potential of primate CD34+ cells that were expanded with Sall4B. Furthermore, an in-vivo murine system was employed to evaluate the hematopoietic potential of primate Sall4B-expanded CD34+ cells. Results Overexpression of Sall4B promoted ex-vivo nonhuman primate CD34+ cell expansion by 9.21 ± 1.94-fold on day 9, whereas lentiviral transduction without Sall4B expanded cells by only 2.95 ± 0.77-fold. Sall4B maintained a significant percentage of CD34+ cells as well. The CFU assay showed that the Sall4B-expanded CD34+ cells still possessed multilineage differentiation potential. A study using nonobese diabetic/severe combined immunodeficiency (NOD/SCID mice in vivo revealed that Sall4B led to an increase in the number of repopulating cells and the 9-day-old Sall4B-transduced CD34+ cells still possess self-renewal and multilineage differentiation capacity in vivo, which are similar stemness characteristics to those in freshly isolated primate bone marrow-derived CD34+ cells. Conclusions We investigated the expansion of nonhuman primate bone marrow-derived CD34+ cells using the Sall4B lentiviral overexpression approach; our findings provide a new perspective on mechanisms of rapid stem cell proliferation. The utilization of Sall4B to expand CD34+ cells on a large scale through use of suitable model systems would prove

In Vitro Efficient Expansion of Tumor Cells Deriving from Different Types of Human Tumor Samples

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Ilaria Turin

2014-03-01

Full Text Available Obtaining human tumor cell lines from fresh tumors is essential to advance our understanding of antitumor immune surveillance mechanisms and to develop new ex vivo strategies to generate an efficient anti-tumor response. The present study delineates a simple and rapid method for efficiently establishing primary cultures starting from tumor samples of different types, while maintaining the immuno-histochemical characteristics of the original tumor. We compared two different strategies to disaggregate tumor specimens. After short or long term in vitro expansion, cells analyzed for the presence of malignant cells demonstrated their neoplastic origin. Considering that tumor cells may be isolated in a closed system with high efficiency, we propose this methodology for the ex vivo expansion of tumor cells to be used to evaluate suitable new drugs or to generate tumor-specific cytotoxic T lymphocytes or vaccines.

Bleb Expansion in Migrating Cells Depends on Supply of Membrane from Cell Surface Invaginations.

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Goudarzi, Mohammad; Tarbashevich, Katsiaryna; Mildner, Karina; Begemann, Isabell; Garcia, Jamie; Paksa, Azadeh; Reichman-Fried, Michal; Mahabaleshwar, Harsha; Blaser, Heiko; Hartwig, Johannes; Zeuschner, Dagmar; Galic, Milos; Bagnat, Michel; Betz, Timo; Raz, Erez

2017-12-04

Cell migration is essential for morphogenesis, organ formation, and homeostasis, with relevance for clinical conditions. The migration of primordial germ cells (PGCs) is a useful model for studying this process in the context of the developing embryo. Zebrafish PGC migration depends on the formation of cellular protrusions in form of blebs, a type of protrusion found in various cell types. Here we report on the mechanisms allowing the inflation of the membrane during bleb formation. We show that the rapid expansion of the protrusion depends on membrane invaginations that are localized preferentially at the cell front. The formation of these invaginations requires the function of Cdc42, and their unfolding allows bleb inflation and dynamic cell-shape changes performed by migrating cells. Inhibiting the formation and release of the invaginations strongly interfered with bleb formation, cell motility, and the ability of the cells to reach their target. Copyright © 2017 Elsevier Inc. All rights reserved.

Truncal anaesthesia of the maxillary nerve for outpatient surgically assisted rapid maxillary expansion.

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Robiony, M; Demitri, V; Costa, F; Politi, M; Cugini, U

1998-10-01

We present our experience of transcutaneous truncal anaesthesia of the maxillary nerve in association with transmucosal anaesthesia of the sphenopalatine ganglion in surgically assisted rapid maxillary expansion. Twelve patients with a skeletal transverse discrepancy of the maxilla were treated in our department from 1994 to 1995. Maxillary transcutaneous nerve block was done with a Quincke 8 cm spinal needle together with transmucosal anaesthesia of the sphenopalatine ganglion. Mepivacaine without adrenaline and sodium bicarbonate 1/10 was used for truncal anaesthesia and lidocaine-prilocaine cream for transmucosal anaesthesia. A Le Fort I osteotomy, lateral nasal wall osteotomy, pterygomaxillary osteotomy, and a palatal osteotomy were done for all patients before the maxillary expansion. Total anaesthesia of the maxillary area facilitated the operations and appreciably reduced the amount of postoperative pain. The ease of achieving effective anaesthesia before and after operation and the absence of side-effects make this form of anaesthetic particularly useful in surgically assisted rapid maxillary expansion.

Rapid expansion method (REM) for time‐stepping in reverse time migration (RTM)

KAUST Repository

Pestana, Reynam C.; Stoffa, Paul L.

2009-01-01

an analytical approximation for the Bessel function where we assume that the time step is sufficiently small. From this derivation we find that if we consider only the first two Chebyshev polynomials terms in the rapid expansion method we can obtain the second

Immediate impact of rapid maxillary expansion on upper airway dimensions and on the quality of life of mouth breathers

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Edna Namiko Izuka

2015-06-01

Full Text Available OBJECTIVE: To assess short-term tomographic changes in the upper airway dimensions and quality of life of mouth breathers after rapid maxillary expansion (RME. METHODS: A total of 25 mouth breathers with maxillary atresia and a mean age of 10.5 years old were assessed by means of cone-beam computed tomography (CBCT and a standardized quality of life questionnaire answered by patients' parents/legal guardians before and immediately after rapid maxillary expansion. RESULTS: Rapid maxillary expansion resulted in similar and significant expansion in the width of anterior (2.8 mm, p < 0.001 and posterior nasal floor (2.8 mm, p < 0.001. Although nasopharynx and nasal cavities airway volumes significantly increased (+1646.1 mm3, p < 0.001, oropharynx volume increase was not statistically significant (+1450.6 mm3, p = 0.066. The results of the quality of life questionnaire indicated that soon after rapid maxillary expansion, patients' respiratory symptoms significantly decreased in relation to their initial respiratory conditions. CONCLUSIONS: It is suggested that RME produces significant dimensional increase in the nasal cavity and nasopharynx. Additionally, it also positively impacts the quality of life of mouth-breathing patients with maxillary atresia.

Regulatory activity of azabisphosphonate-capped dendrimers on human CD4+ T cell proliferation enhances ex-vivo expansion of NK cells from PBMCs for immunotherapy

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Caminade Anne-Marie

2009-09-01

Full Text Available Abstract Background Adoptive cell therapy with allogenic NK cells constitutes a promising approach for the treatment of certain malignancies. Such strategies are currently limited by the requirement of an efficient protocol for NK cell expansion. We have developed a method using synthetic nanosized phosphonate-capped dendrimers allowing such expansion. We are showing here that this is due to a specific inhibitory activity towards CD4+ T cell which could lead to further medical applications of this dendrimer. Methods Mononuclear cells from human peripheral blood were used to investigate the immunomodulatory effects of nanosized phosphonate-capped dendrimers on interleukin-2 driven CD4+T cell expansion. Proliferation status was investigated using flow cytometry analysis of CFSE dilution and PI incorporation experiments. Magnetic bead cell sorting was used to address activity towards individual or mixed cell sub-populations. We performed equilibrium binding assay to assess the interaction of fluorescent dendrimers with pure CD4+ T cells. Results Phosphonate-capped dendrimers are inhibiting the activation, and therefore the proliferation; of CD4+ T cells in IL-2 stimulated PBMCs, without affecting their viability. This allows a rapid enrichment of NK cells and further expansion. We found that dendrimer acts directly on T cells, as their regulatory property is maintained when stimulating purified CD4+ T cells with anti-CD3/CD28 microbeads. Performing equilibrium binding assays using a fluorescent analogue, we show that the phosphonate capped-dendrimers are specifically interacting with purified CD4+ T cells. Ultimately, we found that our protocol prevents the IL-2 related expansion of regulatory T cells that would be deleterious for the activity of infused NK cells. Conclusion High yield expansion of NK cells from human PBMCs by phosphonate-capped dendrimers and IL-2 occurs through the specific inhibition of the CD4+ lymphocyte compartment. Given the

Accelerated production of antigen-specific T-cells for pre-clinical and clinical applications using Gas-permeable Rapid Expansion cultureware (G-Rex)

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Vera, Juan F.; Brenner, Lara J.; Gerdemann, Ulrike; Ngo, Minhtran C.; Sili, Uluhan; Liu, Hao; Wilson, John; Dotti, Gianpietro; Heslop, Helen E.; Leen, Ann M.; Rooney, Cliona M.

2009-01-01

The clinical manufacture of antigen-specific cytotoxic T lymphocytes (CTL) for adoptive immunotherapy is limited by the complexity and time required to produce large numbers with the desired function and specificity. The culture conditions required are rigorous, and in some cases only achieved in 2cm2 wells in which cell growth is limited by gas exchange, nutrients and waste accumulation. Bioreactors developed to overcome these issues tend to be complex, expensive and not always conducive to CTL growth. We observed that antigen-specific CTL undergo seven to ten divisions post-stimulation. However the expected CTL numbers were achieved only in the first week of culture. By recreating the culture conditions present during this first week - low frequency of antigen-specific T-cells and high frequency of feeder cells - we were able to increase CTL expansion to expected levels which could be sustained for several weeks without affecting phenotype or function. However, the number of 24-well plates needed was excessive and cultures required frequent media changes, increasing complexity and manufacturing costs. Therefore, we evaluated novel gas-permeable culture devices (G-Rex) with a silicone membrane at the base allowing gas exchange to occur uninhibited by depth of medium above. This system effectively supports the expansion of CTL and actually increases output by up to 20-fold while decreasing required technician time. Importantly, this amplified cell expansion is not due to more cell divisions but to reduced cell death. This bioprocess optimization increased T-cell output while decreasing the complexity and cost of CTL manufacture, making cell therapy more accessible. PMID:20445351

An Arabidopsis kinase cascade influences auxin-responsive cell expansion.

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Enders, Tara A; Frick, Elizabeth M; Strader, Lucia C

2017-10-01

Mitogen-activated protein kinase (MPK) cascades are conserved mechanisms of signal transduction across eukaryotes. Despite the importance of MPK proteins in signaling events, specific roles for many Arabidopsis MPK proteins remain unknown. Multiple studies have suggested roles for MPK signaling in a variety of auxin-related processes. To identify MPK proteins with roles in auxin response, we screened mpk insertional alleles and identified mpk1-1 as a mutant that displays hypersensitivity in auxin-responsive cell expansion assays. Further, mutants defective in the upstream MAP kinase kinase MKK3 also display hypersensitivity in auxin-responsive cell expansion assays, suggesting that this MPK cascade affects auxin-influenced cell expansion. We found that MPK1 interacts with and phosphorylates ROP BINDING PROTEIN KINASE 1 (RBK1), a protein kinase that interacts with members of the Rho-like GTPases from Plants (ROP) small GTPase family. Similar to mpk1-1 and mkk3-1 mutants, rbk1 insertional mutants display auxin hypersensitivity, consistent with a possible role for RBK1 downstream of MPK1 in influencing auxin-responsive cell expansion. We found that RBK1 directly phosphorylates ROP4 and ROP6, supporting the possibility that RBK1 effects on auxin-responsive cell expansion are mediated through phosphorylation-dependent modulation of ROP activity. Our data suggest a MKK3 • MPK1 • RBK1 phosphorylation cascade that may provide a dynamic module for altering cell expansion. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

Treatment resistant adolescent depression with upper airway resistance syndrome treated with rapid palatal expansion: a case report

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Miller Paul

2012-12-01

Full Text Available Abstract Introduction To the best of our knowledge, this is the first report of a case of treatment-resistant depression in which the patient was evaluated for sleep disordered breathing as the cause and in which rapid palatal expansion to permanently treat the sleep disordered breathing produced a prolonged symptom-free period off medication. Case presentation An 18-year-old Caucasian man presented to our sleep disorders center with chronic severe depression that was no longer responsive to medication but that had recently responded to electroconvulsive therapy. Ancillary, persistent symptoms included mild insomnia, moderate to severe fatigue, mild sleepiness and severe anxiety treated with medication. Our patient had no history of snoring or witnessed apnea, but polysomnography was consistent with upper airway resistance syndrome. Although our patient did not have an orthodontic indication for rapid palatal expansion, rapid palatal expansion was performed as a treatment of his upper airway resistance syndrome. Following rapid palatal expansion, our patient experienced a marked improvement of his sleep quality, anxiety, fatigue and sleepiness. His improvement has been maintained off all psychotropic medication and his depression has remained in remission for approximately two years following his electroconvulsive therapy. Conclusions This case report introduces the possibility that unrecognized sleep disordered breathing may play a role in adolescent treatment-resistant depression. The symptoms of upper airway resistance syndrome are non-specific enough that every adolescent with depression, even those responding to medication, may have underlying sleep disordered breathing. In such patients, rapid palatal expansion, by widening the upper airway and improving airflow during sleep, may produce a prolonged improvement of symptoms and a tapering of medication. Psychiatrists treating adolescents may benefit from having another treatment option for

Fast recovery of platelet production in NOD/SCID mice after transplantation with ex vivo expansion of megakaryocyte from cord blood CD34+ cells

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Hailian Wang

2018-01-01

Conclusion: Platelets can recover rapidly in vivo by means of expanded CD34+ cells with various cytokines. In our system, a group of TPO, SCF, FL, and IL-6 represents the best cytokine combination for expansion of Mk progenitor cells from CB CD34+ cells.

Rapid expansion method (REM) for time‐stepping in reverse time migration (RTM)

KAUST Repository

Pestana, Reynam C.

2009-01-01

We show that the wave equation solution using a conventional finite‐difference scheme, derived commonly by the Taylor series approach, can be derived directly from the rapid expansion method (REM). After some mathematical manipulation we consider an analytical approximation for the Bessel function where we assume that the time step is sufficiently small. From this derivation we find that if we consider only the first two Chebyshev polynomials terms in the rapid expansion method we can obtain the second order time finite‐difference scheme that is frequently used in more conventional finite‐difference implementations. We then show that if we use more terms from the REM we can obtain a more accurate time integration of the wave field. Consequently, we have demonstrated that the REM is more accurate than the usual finite‐difference schemes and it provides a wave equation solution which allows us to march in large time steps without numerical dispersion and is numerically stable. We illustrate the method with post and pre stack migration results.

Coupling of solute transport and cell expansion in pea stems

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Schmalstig, J. G.; Cosgrove, D. J.

1990-01-01

As cells expand and are displaced through the elongation zone of the epicotyl of etiolated pea (Pisum sativum L. var Alaska) seedlings, there is little net dilution of the cell sap, implying a coordination between cell expansion and solute uptake from the phloem. Using [14C] sucrose as a phloem tracer (applied to the hypogeous cotyledons), the pattern of label accumulation along the stem closely matched the growth rate pattern: high accumulation in the growing zone, little accumulation in nongrowing regions. Several results suggest that a major portion of phloem contents enters elongating cells through the symplast. We propose that the coordination between phloem transport and cell expansion is accomplished via regulatory pathways affecting both plasmodesmata conductivity and cell expansion.

Cellulose synthesis inhibition, cell expansion, and patterns of cell wall deposition in Nitella internodes

International Nuclear Information System (INIS)

Richmond, P.A.; Metraux, J.P.

1984-01-01

The authors have investigated the pattern of wall deposition and maturation and correlated it with cell expansion and cellulose biosynthesis. The herbicide 2,6-dichlorobenzonitrile (DCB) was found to be a potent inhibitor of cellulose synthesis, but not of cell expansion in Nitella internodal cells. Although cellulose synthesis is inhibited during DCB treatment, matrix substances continue to be synthesized and deposited. The inhibition of cellulose microfibril deposition can be demonstrated by various techniques. These results demonstrate that matrix deposition is by apposition, not by intussusception, and that the previously deposited wall moves progressively outward while stretching and thinning as a result of cell expansion

Alveolar bone changes after asymmetric rapid maxillary expansion.

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Akin, Mehmet; Baka, Zeliha Muge; Ileri, Zehra; Basciftci, Faruk Ayhan

2015-09-01

To quantitatively evaluate the effects of asymmetric rapid maxillary expansion (ARME) on cortical bone thickness and buccal alveolar bone height (BABH), and to determine the formation of dehiscence and fenestration in the alveolar bone surrounding the posterior teeth, using cone-beam computed tomography (CBCT). The CBCT records of 23 patients with true unilateral posterior skeletal crossbite (10 boys, 14.06 ± 1.08 years old, and 13 girls, 13.64 ± 1.32 years old) who had undergone ARME were selected from our clinic archives. The bonded acrylic ARME appliance, including an occlusal stopper, was used on all patients. CBCT records had been taken before ARME (T1) and after the 3-month retention period (T2). Axial slices of the CBCT images at 3 vertical levels were used to evaluate the buccal and palatal aspects of the canines, first and second premolars, and first molars. Paired samples and independent sample t-tests were used for statistical comparison. The results suggest that buccal cortical bone thickness of the affected side was significantly more affected by the expansion than was the unaffected side (P ARME significantly reduced the BABH of the canines (P ARME also increased the incidence of dehiscence and fenestration on the affected side. ARME may quantitatively decrease buccal cortical bone thickness and height on the affected side.

Atkinesin-13A Modulates Cell-Wall Synthesis and Cell Expansion in Arabidopsis thaliana via the THESEUS1 Pathway

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Fujikura, Ushio; Elsaesser, Lore; Breuninger, Holger; Sánchez-Rodríguez, Clara; Ivakov, Alexander; Laux, Thomas; Findlay, Kim; Persson, Staffan; Lenhard, Michael

2014-01-01

Growth of plant organs relies on cell proliferation and expansion. While an increasingly detailed picture about the control of cell proliferation is emerging, our knowledge about the control of cell expansion remains more limited. We demonstrate here that the internal-motor kinesin AtKINESIN-13A (AtKIN13A) limits cell expansion and cell size in Arabidopsis thaliana, with loss-of-function atkin13a mutants forming larger petals with larger cells. The homolog, AtKINESIN-13B, also affects cell expansion and double mutants display growth, gametophytic and early embryonic defects, indicating a redundant role of the two genes. AtKIN13A is known to depolymerize microtubules and influence Golgi motility and distribution. Consistent with this function, AtKIN13A interacts genetically with ANGUSTIFOLIA, encoding a regulator of Golgi dynamics. Reduced AtKIN13A activity alters cell wall structure as assessed by Fourier-transformed infrared-spectroscopy and triggers signalling via the THESEUS1-dependent cell-wall integrity pathway, which in turn promotes the excess cell expansion in the atkin13a mutant. Thus, our results indicate that the intracellular activity of AtKIN13A regulates cell expansion and wall architecture via THESEUS1, providing a compelling case of interplay between cell wall integrity sensing and expansion. PMID:25232944

Anchorage onto deciduous teeth: effectiveness of early rapid maxillary expansion in increasing dental arch dimension and improving anterior crowding

OpenAIRE

Mutinelli, Sabrina; Manfredi, Mario; Guiducci, Antonio; Denotti, Gloria; Cozzani, Mauro

2015-01-01

Background Anchorage onto permanent dentition is a common procedure in rapid maxillary expansion. However, replacing first permanent molars with the second deciduous molars seems to be an option to reduce some negative side effects during orthodontic treatment. The purpose of this study was to evaluate the dental effect of rapid maxillary expansion with anchorage exclusively onto deciduous teeth performed in the first period of transition. Methods Twenty patients with a lateral cross-bite tre...

The endoplasmic reticulum exerts control over organelle streaming during cell expansion.

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Stefano, Giovanni; Renna, Luciana; Brandizzi, Federica

2014-03-01

Cytoplasmic streaming is crucial for cell homeostasis and expansion but the precise driving forces are largely unknown. In plants, partial loss of cytoplasmic streaming due to chemical and genetic ablation of myosins supports the existence of yet-unknown motors for organelle movement. Here we tested a role of the endoplasmic reticulum (ER) as propelling force for cytoplasmic streaming during cell expansion. Through quantitative live-cell analyses in wild-type Arabidopsis thaliana cells and mutants with compromised ER structure and streaming, we demonstrate that cytoplasmic streaming undergoes profound changes during cell expansion and that it depends on motor forces co-exerted by the ER and the cytoskeleton.

Impact of rapid maxillary expansion on nasomaxillary complex volume in mouth-breathers

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Mario Cappellette Jr.

Full Text Available ABSTRACT Objective: To assess the volumetric changes that occur in the nasomaxillary complex of mouth-breathing patients with transverse maxillary deficiency subjected to rapid maxillary expansion (RME. Methods: This was a controlled, prospective intervention study involving 38 mouth-breathing patients presenting with transverse maxillary deficiency, regardless of malocclusion type or race. Twenty-three of them comprised the experimental group, which was composed of 11 (47.8% boys, and 12 (52.2% girls, with a mean age of 9.6 years, ranging from 6.4 to 14.2 years and standard deviation of 2.3 years; and 15 of them comprised the control group, composed of 9 (60% boys and 6 (40% girls with an mean age of 10.5 years, ranging from 8.0 to 13.6 years, and standard deviation of 1.9 years. All patients were scanned (CT according to a standard protocol: Initial CT (T1, and CT three months thereafter (T2, and the patients in the experimental group were treated with RME using a Hyrax expander for the correction of maxillary deficiency during the T1-T2 interval. The CT scans were manipulated using Dolphin® Imaging version 11.7 software for total and partial volumetric assessment of the nasomaxillary complex. Results: The results revealed that in the experimental group there was a significant increase in the size of the structures of interest compared to the control group, both in general aspect and in specific regions. Conclusions: Rapid maxillary expansion (RME provided a significant expansion in all the structures of the nasomaxillary complex (nasal cavity, oropharynx, right and left maxillary sinuses.

A novel monoclonal antibody of human stem cell factor inhibits umbilical cord blood stem cell ex vivo expansion

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Fan Jie

2012-12-01

Full Text Available Abstract Stem cell factor (SCF activates hematopoietic stem cell (HSC self-renewal and is being used to stimulate the ex vivo expansion of HSCs. The mechanism by which SCF supports expansion of HSCs remains poorly understood. In cord blood ex vivo expansion assays, a newly produced anti-SCF monoclonal antibody (clone 23C8 was found to significantly inhibit the expansion of CD34+ cells. This antibody appears to bind directly to a part of SCF that is critical for biological activity toward expansion of CD34+ cells, which is located in the first 104 amino acids from the NH2-terminus.

Volumetric changes of the nose and nasal airway 2 years after tooth-borne and bone-borne surgically assisted rapid maxillary expansion

NARCIS (Netherlands)

Nada, R.M.; Loon, B. van; Schols, J.G.J.H.; Maal, T.J.J.; Koning, M.J.J. de; Mostafa, Y.A.; Kuijpers-Jagtman, A.M.

2013-01-01

This study aimed to assess the effects of bone-borne and tooth-borne surgically assisted rapid maxillary expansion on the volumes of the nose and nasal airway 2 yr after maxillary expansion. This prospective cohort study included 32 patients with transverse maxillary hypoplasia. Expansion was

Expansion of mycobacterium-reactive gamma delta T cells by a subset of memory helper T cells.

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Vila, L M; Haftel, H M; Park, H S; Lin, M S; Romzek, N C; Hanash, S M; Holoshitz, J

1995-04-01

Human gamma delta T cells expressing the V gamma 9/V delta 2 T-cell receptor have been previously found to proliferate in response to certain microorganisms and to expand throughout life, presumably because of extrathymic activation by foreign antigens. In vitro expansion of V gamma 9/V delta 2 cells by mycobacteria has been previously shown to be dependent on accessory cells. In order to gain an insight into the mechanisms involved in the expansion of these cells, we have undertaken to identify the peripheral blood subset of cells on which proliferation of V gamma 9/V delta 2 cells in response to mycobacteria is dependent. Contrary to their role in antigen presentation to alpha beta T cells, professional antigen-presenting cells, such as monocytes, B cells, and dendritic cells, were unable to provide the cellular support for the expansion of V gamma 9/V delta 2 cells. Selective depletion of T-cell subsets, as well as the use of highly purified T-cell populations, indicated that the only subset of peripheral blood cells that could expand V gamma 9/V delta 2 cells were CD4+ CD45RO+ CD7- alpha beta T cells. These cells underwent distinct intracellular signaling events after stimulation with the mycobacterial antigen. Expansion of V gamma 9/V delta 2 cells by alpha beta T cells was dependent on cell-cell contact. This is the first evidence that a small subset of the memory helper T-cell population is exclusively responsible for the peripheral expansion of V gamma 9/V delta 2 cells. These data illustrate a unique aspect of antigen recognition by gamma delta T cells and provide new means to study their immune defense role.

Three-dimensional prospective evaluation of tooth-borne and bone-borne surgically assisted rapid maxillary expansion

NARCIS (Netherlands)

Nada, R.M.; Fudalej, P.S.; Maal, T.J.J.; Berge, S.J.; Mostafa, Y.A.; Kuijpers-Jagtman, A.M.

2012-01-01

AIM: To three-dimensionally (3D) assess the long-term effects of tooth-borne and bone-borne surgically assisted rapid maxillary expansion (SARME). SUBJECTS AND METHODS: This prospective cohort study comprised 45 consecutive skeletally mature non-syndromic patients with transverse maxillary

Analyzing suitability for urban expansion under rapid coastal urbanization with remote sensing and GIS techniques: a case study of Lianyungang, China

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Zhao, Wenjun; Zhu, Xiaodong; Reenberg, Anette; Sun, Xiang

2010-10-01

Beginning in 2000, Lianyungang's urbanization entered a period of rapid growth, spatially as well as economically. Rapid and intensive expansion of "construction land" imposed increasing pressures on regional environment. With the support of remote sensing data and GIS tools, this paper reports a "present-capacity-potential" integrated suitability analysis framework, in order to characterize and evaluate the suitability of urban expansion in Lianyungang. We found that during the rapid coastal urbanization process from 2000 to 2008, the characteristics of physical expansion in the study area were characterized by a combination of high-density expansion and sprawling development. The land use conversion driven by urbanization and industrialization has not occurred only in city districts, but also the surrounding areas that were spatially absorbed by urban growth, while closely associated and greatly influenced by the explosive growth of industrial establishment. The over-consumption of land resources in the areas with low environmental carrying capacity, particularly in the eastern coastal area, should be strictly controlled. Compared to conventional land suitability analysis methods, the proposed integrated approach could better review the potential environmental impacts of urban expansion and provide guidance for decision makers.

Memory phenotype CD4 T cells undergoing rapid, nonburst-like, cytokine-driven proliferation can be distinguished from antigen-experienced memory cells.

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Souheil-Antoine Younes

2011-10-01

Full Text Available Memory phenotype (CD44(bright, CD25(negative CD4 spleen and lymph node T cells (MP cells proliferate rapidly in normal or germ-free donors, with BrdU uptake rates of 6% to 10% per day and Ki-67 positivity of 18% to 35%. The rapid proliferation of MP cells stands in contrast to the much slower proliferation of lymphocytic choriomeningitis virus (LCMV-specific memory cells that divide at rates ranging from <1% to 2% per day over the period from 15 to 60 days after LCMV infection. Anti-MHC class II antibodies fail to inhibit the in situ proliferation of MP cells, implying a non-T-cell receptor (TCR-driven proliferation. Such proliferation is partially inhibited by anti-IL-7Rα antibody. The sequence diversity of TCRβ CDR3 gene segments is comparable among the proliferating and quiescent MP cells from conventional and germ-free mice, implying that the majority of proliferating MP cells have not recently derived from a small cohort of cells that expand through multiple continuous rounds of cell division. We propose that MP cells constitute a diverse cell population, containing a subpopulation of slowly dividing authentic antigen-primed memory cells and a majority population of rapidly proliferating cells that did not arise from naïve cells through conventional antigen-driven clonal expansion.

Novel micronisation β-carotene using rapid expansion supercritical solution with co-solvent

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Kien, Le Anh

2017-09-01

Rapid expansion of supercritical solution (RESS) is the most common approach of pharmaceutical pacticle forming methods using supercritical fluids. The RESS method is a technology producing a small solid product with a very narrow particle size distribution, organic solvent-free particles. This process is also simple and easy to control the operating parameters in comparision with other ways based on supercritical techniques. In this study, β-carotene, a strongly colored red-orange pigment abundant in plants and fruits, has been forming by RESS. In addition, the size and morphology effect of four different RESS parameters including co-solvent, extraction temperature, and extraction pressure and expansion nozzle temperature has surveyed. The particle size distribution has been determined by using laser diffraction experiment. SEM has conducted to analyze the surface structure, DSC and FTIR for thermal and chemical structure analysis.

Effects of rapid maxillary expansion in cleft patients resulting from the use of two different expanders

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Daniel Santos Fonseca Figueiredo

Full Text Available ABSTRACT Introduction: The aim of this study was to evaluate the skeletal and dental effects of rapid maxillary expansion (RME in cleft patients using two types of expanders. Methods: Twenty unilateral cleft lip and palate patients were randomly divided into two groups, according to the type of expander used: (I modified Hyrax and (II inverted Mini-Hyrax. A pretreatment cone-beam computed tomographic image (T0 was taken as part of the initial orthodontic records and three months after RME as need for bone graft planning (T1. Results: In general, there was no significant difference among groups (p > 0.05. Both showed significant transverse maxillary expansion (p 0.05. There was greater crown than apical expansion. Maxillary posterior expansion tended to be larger than anterior opening (p 0.05. Conclusions: The appliances tested are effective for transverse expansion of the maxilla. However, these appliances should be better indicated to cleft cases also presenting posterior transverse discrepancy, since there was greater expansion in the posterior maxillary region than in the anterior one.

Cell fiber-based three-dimensional culture system for highly efficient expansion of human induced pluripotent stem cells.

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Ikeda, Kazuhiro; Nagata, Shogo; Okitsu, Teru; Takeuchi, Shoji

2017-06-06

Human pluripotent stem cells are a potentially powerful cellular resource for application in regenerative medicine. Because such applications require large numbers of human pluripotent stem cell-derived cells, a scalable culture system of human pluripotent stem cell needs to be developed. Several suspension culture systems for human pluripotent stem cell expansion exist; however, it is difficult to control the thickness of cell aggregations in these systems, leading to increased cell death likely caused by limited diffusion of gases and nutrients into the aggregations. Here, we describe a scalable culture system using the cell fiber technology for the expansion of human induced pluripotent stem (iPS) cells. The cells were encapsulated and cultured within the core region of core-shell hydrogel microfibers, resulting in the formation of rod-shaped or fiber-shaped cell aggregations with sustained thickness and high viability. By encapsulating the cells with type I collagen, we demonstrated a long-term culture of the cells by serial passaging at a high expansion rate (14-fold in four days) while retaining its pluripotency. Therefore, our culture system could be used for large-scale expansion of human pluripotent stem cells for use in regenerative medicine.

Crop domestication facilitated rapid geographic expansion of a specialist pollinator, the squash bee Peponapis pruinosa

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Over the past 10,000 years, humans have facilitated rapid range expansions of animal, plants and microorganisms, often accompanying agriculture’s spread. Three squash species were early domesticates in the New World. Their spreading cultivation out of the Southwest across much of today’s USA has bee...

Conditionally reprogrammed cells (CRC) methodology does not allow the in vitro expansion of patient-derived primary and metastatic lung cancer cells.

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Sette, Giovanni; Salvati, Valentina; Giordani, Ilenia; Pilozzi, Emanuela; Quacquarini, Denise; Duranti, Enrico; De Nicola, Francesca; Pallocca, Matteo; Fanciulli, Maurizio; Falchi, Mario; Pallini, Roberto; De Maria, Ruggero; Eramo, Adriana

2018-07-01

Availability of tumor and non-tumor patient-derived models would promote the development of more effective therapeutics for non-small cell lung cancer (NSCLC). Recently, conditionally reprogrammed cells (CRC) methodology demonstrated exceptional potential for the expansion of epithelial cells from patient tissues. However, the possibility to expand patient-derived lung cancer cells using CRC protocols is controversial. Here, we used CRC approach to expand cells from non-tumoral and tumor biopsies of patients with primary or metastatic NSCLC as well as pulmonary metastases of colorectal or breast cancers. CRC cultures were obtained from both tumor and non-malignant tissues with extraordinary high efficiency. Tumor cells were tracked in vitro through tumorigenicity assay, monitoring of tumor-specific genetic alterations and marker expression. Cultures were composed of EpCAM+ lung epithelial cells lacking tumorigenic potential. NSCLC biopsies-derived cultures rapidly lost patient-specific genetic mutations or tumor antigens. Similarly, pulmonary metastases of colon or breast cancer generated CRC cultures of lung epithelial cells. All CRC cultures examined displayed epithelial lung stem cell phenotype and function. In contrast, brain metastatic lung cancer biopsies failed to generate CRC cultures. In conclusion, patient-derived primary and metastatic lung cancer cells were negatively selected under CRC conditions, limiting the expansion to non-malignant lung epithelial stem cells from either tumor or non-tumor tissue sources. Thus, CRC approach cannot be applied for direct therapeutic testing of patient lung tumor cells, as the tumor-derived CRC cultures are composed of (non-tumoral) airway basal cells. © 2018 UICC.

The evidence of the rugoscopy effectiveness as a human identification method in patients submitted to rapid palatal expansion.

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Barbieri, Ana A; Scoralick, Raquel A; Naressi, Suely C M; Moraes, Mari E L; Daruge, Eduardo; Daruge, Eduardo

2013-01-01

The objective of this study was to demonstrate the effectiveness of rugoscopy as a human identification method, even when the patient is submitted to rapid palatal expansion, which in theory would introduce doubt. With this intent, the Rugoscopic Identity was obtained for each subject using the classification formula proposed by Santos based on the intra-oral casts made before and after treatment from patients who were subjected to palatal expansion. The casts were labeled with the patients' initials and randomly arranged for studying. The palatine rugae kept the same patterns in every case studied. The technical error of the intra-evaluator measurement provided a confidence interval of 95%, making rugoscopy a reliable identification method for patients who were submitted to rapid palatal expansion, because even in the presence of intra-oral changes owing to the use of palatal expanders, the palatine rugae retained the biological and technical requirements for the human identification process. © 2012 American Academy of Forensic Sciences.

[Piezosurgery for surgically assisted rapid maxillary expansion under local anesthesia].

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Sun, Hao; Li, Biao; Sun, Hao; Liu, Zhixu; Wang, Xudong

2014-08-01

This study evaluates piezosurgery for surgically assisted rapid maxillary expansion (SARME) under local anesthesia. SARME was performed on adults with maxillary transverse deficiency under local anesthesia with a piezosurgical device. Fourteen patients (six males and eight females) underwent lateral maxillary osteotomies, midpalatal osteotomies, and bilateral pterygomaxillary disjunction. The feelings of patients during the operation were determined through questionnaires. All patients underwent SARME in the out-patient operating room. The surgical procedures were completed under local anesthesia. All patients exhibited satisfactory tolerance. Ultrasonic bone-cutting surgery was recently introduced as a feasible alternative to the conventional tools of cranio-maxillofacial surgery for its technical characteristics of precision and safety. The device used was unique in that cutting action occurred when the tool was employed on mineralized tissues, but stoped on soft tissues. The results of the questionnaires showed that eight (57.14%) patients felt a mild sensation of ultrasonic vibration, tweleve (85.7 1%) felt mild tolerable pain and tooth soreness during surgery, and eleven (78.57%) felt little fear and hardly heard the ultrasonic sound. Preoperative and postoperative six months later measurements showed an evident effect of expansion. Piezosurgery enabled patients to undergo all the steps of SARME under local anesthesia, but more cases and longer follow-up are needed to verif ' the results.

Protecting tropical forests from the rapid expansion of rubber using carbon payments.

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Warren-Thomas, Eleanor M; Edwards, David P; Bebber, Daniel P; Chhang, Phourin; Diment, Alex N; Evans, Tom D; Lambrick, Frances H; Maxwell, James F; Nut, Menghor; O'Kelly, Hannah J; Theilade, Ida; Dolman, Paul M

2018-03-02

Expansion of Hevea brasiliensis rubber plantations is a resurgent driver of deforestation, carbon emissions, and biodiversity loss in Southeast Asia. Southeast Asian rubber extent is massive, equivalent to 67% of oil palm, with rapid further expansion predicted. Results-based carbon finance could dis-incentivise forest conversion to rubber, but efficacy will be limited unless payments match, or at least approach, the costs of avoided deforestation. These include opportunity costs (timber and rubber profits), plus carbon finance scheme setup (transaction) and implementation costs. Using comprehensive Cambodian forest data, exploring scenarios of selective logging and conversion, and assuming land-use choice is based on net present value, we find that carbon prices of $30-$51 per tCO 2 are needed to break even against costs, higher than those currently paid on carbon markets or through carbon funds. To defend forests from rubber, either carbon prices must be increased, or other strategies are needed, such as corporate zero-deforestation pledges, and governmental regulation and enforcement of forest protection.

Differential response of cell-cycle and cell-expansion regulators to heat stress in apple (Malus domestica) fruitlets.

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Flaishman, Moshe A; Peles, Yuval; Dahan, Yardena; Milo-Cochavi, Shira; Frieman, Aviad; Naor, Amos

2015-04-01

Temperature is one of the most significant factors affecting physiological and biochemical aspects of fruit development. Current and progressing global warming is expected to change climate in the traditional deciduous fruit tree cultivation regions. In this study, 'Golden Delicious' trees, grown in a controlled environment or commercial orchard, were exposed to different periods of heat treatment. Early fruitlet development was documented by evaluating cell number, cell size and fruit diameter for 5-70 days after full bloom. Normal activities of molecular developmental and growth processes in apple fruitlets were disrupted under daytime air temperatures of 29°C and higher as a result of significant temporary declines in cell-production and cell-expansion rates, respectively. Expression screening of selected cell cycle and cell expansion genes revealed the influence of high temperature on genetic regulation of apple fruitlet development. Several core cell-cycle and cell-expansion genes were differentially expressed under high temperatures. While expression levels of B-type cyclin-dependent kinases and A- and B-type cyclins declined moderately in response to elevated temperatures, expression of several cell-cycle inhibitors, such as Mdwee1, Mdrbr and Mdkrps was sharply enhanced as the temperature rose, blocking the cell-cycle cascade at the G1/S and G2/M transition points. Moreover, expression of several expansin genes was associated with high temperatures, making them potentially useful as molecular platforms to enhance cell-expansion processes under high-temperature regimes. Understanding the molecular mechanisms of heat tolerance associated with genes controlling cell cycle and cell expansion may lead to the development of novel strategies for improving apple fruit productivity under global warming. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Intra-hydrogel culture prevents transformation of mesenchymal stem cells induced by monolayer expansion.

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Jiang, Tongmeng; Liu, Junting; Ouyang, Yiqiang; Wu, Huayu; Zheng, Li; Zhao, Jinmin; Zhang, Xingdong

2018-05-01

In this study, we report that the intra-hydrogel culture system mitigates the transformation of mesenchymal stem cells (MSCs) induced by two-dimensional (2D) expansion. MSCs expanded in monolayer culture prior to encapsulation in collagen hydrogels (group eMSCs-CH) featured impaired stemness in chondrogenesis, comparing with the freshly isolated bone marrow mononuclear cells seeded directly in collagen hydrogels (group fMSCs-CH). The molecular mechanism of the in vitro expansion-triggered damage to MSCs was detected through genome-wide microarray analysis. Results indicated that pathways such as proteoglycans in cancer and pathways in cancer expansion were highly enriched in eMSCs-CH. And multiple up-regulated oncoma-associated genes were verified in eMSCs-CH compared with fMSCs-CH, indicating that expansion in vitro triggered cellular transformation was associated with signaling pathways related to tumorigenicity. Besides, focal adhesion (FA) and mitogen-activated protein kinase (MAPK) signaling pathways were also involved in in vitro expansion, indicating restructuring of the cell architecture. Thus, monolayer expansion in vitro may contribute to vulnerability of MSCs through the regulation of FA and MAPK. This study indicates that intra-hydrogel culture can mitigate the monolayer expansion induced transformation of MSCs and maintain the uniformity of the stem cells, which is a viable in vitro culture system for stem cell therapy.

Mesenchymal Stromal Cell Phenotype is not Influenced by Confluence during Culture Expansion

DEFF Research Database (Denmark)

Haack-Sørensen, Mandana; Hansen, Susanne Kofoed; Hansen, Louise

2013-01-01

BACKGROUND: Accumulating preclinical and clinical evidence indicates that human mesenchymal stromal cells (MSCs) are good candidates for cell therapy. For clinical applications of MSCs extensive in vitro expansion is required to obtain an adequate number of cells. It is evident that the pursuit...... differentiation. This phenotype persisted independent of increasing cell densities. DISCUSSION: These data demonstrate that MSC characteristics and plasticity can be maintained during culture expansion from bone marrow mononuclear cells to MSCs and that a homogeneous phenotype of undifferentiated MSCs which...... persists independent of cell density can be used for clinical therapies....

Suppression of IL-7-dependent Effector T-cell Expansion by Multipotent Adult Progenitor Cells and PGE2

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Reading, James L; Vaes, Bart; Hull, Caroline; Sabbah, Shereen; Hayday, Thomas; Wang, Nancy S; DiPiero, Anthony; Lehman, Nicholas A; Taggart, Jen M; Carty, Fiona; English, Karen; Pinxteren, Jef; Deans, Robert; Ting, Anthony E; Tree, Timothy I M

2015-01-01

T-cell depletion therapy is used to prevent acute allograft rejection, treat autoimmunity and create space for bone marrow or hematopoietic cell transplantation. The evolved response to T-cell loss is a transient increase in IL-7 that drives compensatory homeostatic proliferation (HP) of mature T cells. Paradoxically, the exaggerated form of this process that occurs following lymphodepletion expands effector T-cells, often causing loss of immunological tolerance that results in rapid graft rejection, autoimmunity, and exacerbated graft-versus-host disease (GVHD). While standard immune suppression is unable to treat these pathologies, growing evidence suggests that manipulating the incipient process of HP increases allograft survival, prevents autoimmunity, and markedly reduces GVHD. Multipotent adult progenitor cells (MAPC) are a clinical grade immunomodulatory cell therapy known to alter γ-chain cytokine responses in T-cells. Herein, we demonstrate that MAPC regulate HP of human T-cells, prevent the expansion of Th1, Th17, and Th22 effectors, and block the development of pathogenic allograft responses. This occurs via IL-1β-primed secretion of PGE2 and activates T-cell intrinsic regulatory mechanisms (SOCS2, GADD45A). These data provide proof-of-principle that HP of human T-cells can be targeted by cellular and molecular therapies and lays a basis for the development of novel strategies to prevent immunopathology in lymphodepleted patients. PMID:26216515

Cone-beam computed tomography evaluation of dentoskeletal changes after asymmetric rapid maxillary expansion.

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Baka, Zeliha Muge; Akin, Mehmet; Ucar, Faruk Izzet; Ileri, Zehra

2015-01-01

The aims of this study were to quantitatively evaluate the changes in arch widths and buccolingual inclinations of the posterior teeth after asymmetric rapid maxillary expansion (ARME) and to compare the measurements between the crossbite and the noncrossbite sides with cone-beam computed tomography (CBCT). From our clinic archives, we selected the CBCT records of 30 patients with unilateral skeletal crossbite (13 boys, 14.2 ± 1.3 years old; 17 girls, 13.8 ± 1.3 years old) who underwent ARME treatment. A modified acrylic bonded rapid maxillary expansion appliance including an occlusal locking mechanism was used in all patients. CBCT records had been taken before ARME treatment and after a 3-month retention period. Fourteen angular and 80 linear measurements were taken for the maxilla and the mandible. Frontally clipped CBCT images were used for the evaluation. Paired sample and independent sample t tests were used for statistical comparisons. Comparisons of the before-treatment and after-retention measurements showed that the arch widths and buccolingual inclinations of the posterior teeth increased significantly on the crossbite side of the maxilla and on the noncrossbite side of the mandible (P ARME treatment, the crossbite side of the maxilla and the noncrossbite side of the mandible were more affected than were the opposite sides. Copyright © 2015. Published by Elsevier Inc.

Low interleukin-2 concentration favors generation of early memory T cells over effector phenotypes during chimeric antigen receptor T-cell expansion.

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Kaartinen, Tanja; Luostarinen, Annu; Maliniemi, Pilvi; Keto, Joni; Arvas, Mikko; Belt, Heini; Koponen, Jonna; Loskog, Angelica; Mustjoki, Satu; Porkka, Kimmo; Ylä-Herttuala, Seppo; Korhonen, Matti

2017-06-01

Adoptive T-cell therapy offers new options for cancer treatment. Clinical results suggest that T-cell persistence, depending on T-cell memory, improves efficacy. The use of interleukin (IL)-2 for in vitro T-cell expansion is not straightforward because it drives effector T-cell differentiation but does not promote the formation of T-cell memory. We have developed a cost-effective expansion protocol for chimeric antigen receptor (CAR) T cells with an early memory phenotype. Lymphocytes were transduced with third-generation lentiviral vectors and expanded using CD3/CD28 microbeads. The effects of altering the IL-2 supplementation (0-300 IU/mL) and length of expansion (10-20 days) on the phenotype of the T-cell products were analyzed. High IL-2 levels led to a decrease in overall generation of early memory T cells by both decreasing central memory T cells and augmenting effectors. T memory stem cells (T SCM , CD95 + CD45RO - CD45RA + CD27 + ) were present variably during T-cell expansion. However, their presence was not IL-2 dependent but was linked to expansion kinetics. CD19-CAR T cells generated in these conditions displayed in vitro antileukemic activity. In summary, production of CAR T cells without any cytokine supplementation yielded the highest proportion of early memory T cells, provided a 10-fold cell expansion and the cells were functionally potent. The number of early memory T cells in a T-cell preparation can be increased by simply reducing the amount of IL-2 and limiting the length of T-cell expansion, providing cells with potentially higher in vivo performance. These findings are significant for robust and cost-effective T-cell manufacturing. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Culture expansion of adipose derived stromal cells. A closed automated Quantum Cell Expansion System compared with manual flask-based culture

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Mandana Haack-Sørensen

2016-11-01

Full Text Available Abstract Background Adipose derived stromal cells (ASCs are a rich and convenient source of cells for clinical regenerative therapeutic approaches. However, applications of ASCs often require cell expansion to reach the needed dose. In this study, cultivation of ASCs from stromal vascular fraction (SVF over two passages in the automated and functionally closed Quantum Cell Expansion System (Quantum system is compared with traditional manual cultivation. Methods Stromal vascular fraction was isolated from abdominal fat, suspended in α-MEM supplemented with 10% Fetal Bovine Serum and seeded into either T75 flasks or a Quantum system that had been coated with cryoprecipitate. The cultivation of ASCs from SVF was performed in 3 ways: flask to flask; flask to Quantum system; and Quantum system to Quantum system. In all cases, quality controls were conducted for sterility, mycoplasmas, and endotoxins, in addition to the assessment of cell counts, viability, immunophenotype, and differentiation potential. Results The viability of ASCs passage 0 (P0 and P1 was above 96%, regardless of cultivation in flasks or Quantum system. Expression of surface markers and differentiation potential was consistent with ISCT/IFATS standards for the ASC phenotype. Sterility, mycoplasma, and endotoxin tests were consistently negative. An average of 8.0 × 107 SVF cells loaded into a Quantum system yielded 8.96 × 107 ASCs P0, while 4.5 × 106 SVF cells seeded per T75 flask yielded an average of 2.37 × 106 ASCs—less than the number of SVF cells seeded. ASCs P1 expanded in the Quantum system demonstrated a population doubling (PD around 2.2 regardless of whether P0 was previously cultured in flasks or Quantum, while ASCs P1 in flasks only reached a PD of 1.0. Conclusion: Manufacturing of ASCs in a Quantum system enhances ASC expansion rate and yield significantly relative to manual processing in T-flasks, while maintaining the purity and quality essential to

Culture expansion of adipose derived stromal cells. A closed automated Quantum Cell Expansion System compared with manual flask-based culture.

Science.gov (United States)

Haack-Sørensen, Mandana; Follin, Bjarke; Juhl, Morten; Brorsen, Sonja K; Søndergaard, Rebekka H; Kastrup, Jens; Ekblond, Annette

2016-11-16

Adipose derived stromal cells (ASCs) are a rich and convenient source of cells for clinical regenerative therapeutic approaches. However, applications of ASCs often require cell expansion to reach the needed dose. In this study, cultivation of ASCs from stromal vascular fraction (SVF) over two passages in the automated and functionally closed Quantum Cell Expansion System (Quantum system) is compared with traditional manual cultivation. Stromal vascular fraction was isolated from abdominal fat, suspended in α-MEM supplemented with 10% Fetal Bovine Serum and seeded into either T75 flasks or a Quantum system that had been coated with cryoprecipitate. The cultivation of ASCs from SVF was performed in 3 ways: flask to flask; flask to Quantum system; and Quantum system to Quantum system. In all cases, quality controls were conducted for sterility, mycoplasmas, and endotoxins, in addition to the assessment of cell counts, viability, immunophenotype, and differentiation potential. The viability of ASCs passage 0 (P0) and P1 was above 96%, regardless of cultivation in flasks or Quantum system. Expression of surface markers and differentiation potential was consistent with ISCT/IFATS standards for the ASC phenotype. Sterility, mycoplasma, and endotoxin tests were consistently negative. An average of 8.0 × 10 7 SVF cells loaded into a Quantum system yielded 8.96 × 10 7 ASCs P0, while 4.5 × 10 6 SVF cells seeded per T75 flask yielded an average of 2.37 × 10 6 ASCs-less than the number of SVF cells seeded. ASCs P1 expanded in the Quantum system demonstrated a population doubling (PD) around 2.2 regardless of whether P0 was previously cultured in flasks or Quantum, while ASCs P1 in flasks only reached a PD of 1.0. Manufacturing of ASCs in a Quantum system enhances ASC expansion rate and yield significantly relative to manual processing in T-flasks, while maintaining the purity and quality essential to safe and robust cell production. Notably, the use of the Quantum

Bilateral cross-bite treated by repeated rapid maxillary expansions: a 17-year follow-up case.

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Cozzani, M; Mazzotta, L; Caprioglio, A

2014-07-01

The objective of this paper is to show the clinical results after the repeated application of a Haas expander for rapid maxillary expansion (RME) anchored onto deciduous teeth in a 7-year-old patient that presented bilateral cross-bite, superior crowding and no space for permanent lateral incisors eruption. A first Haas expander was applied to the patient. She was told to activate it once a day, each activation was equal to 0.20 mm. After the first RME, the bilateral cross-bite was solved but still there was not enough space for lateral incisor eruption. A second and then a third Haas expander were applied, with the same activation protocol as the first one, in order to gain space in the anterior region and to achieve proper eruption of the lateral incisors. The patient was then treated with fixed appliances. At debonding the patient presented well aligned arch-forms: space for lateral incisor eruption was gained and superior crowding was solved. Bilateral cross-bite was also corrected. She was seen again 10 years and 17 years after expansions: she showed no relapse and presented a good functional occlusion that had remained stable, and an aesthetically pleasant smile, however she exhibited gingival recessions. Repeated rapid maxillary expansion, anchored onto deciduous teeth, performed in early mixed dentition represents a safe and successful treatment to correct severe bilateral cross- bites and to create space for maxillary incisor eruption.

Microenvironmental cues enhance mesenchymal stem cell-mediated immunomodulation and regulatory T-cell expansion.

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Kadle, Rohini L; Abdou, Salma A; Villarreal-Ponce, Alvaro P; Soares, Marc A; Sultan, Darren L; David, Joshua A; Massie, Jonathan; Rifkin, William J; Rabbani, Piul; Ceradini, Daniel J

2018-01-01

Mesenchymal stem cells (MSCs) are known to both have powerful immunosuppressive properties and promote allograft tolerance. Determining the environmental oxygen tension and inflammatory conditions under which MSCs are optimally primed for this immunosuppressive function is essential to their utilization in promoting graft tolerance. Of particular interest is the mechanisms governing the interaction between MSCs and regulatory T cells (Tregs), which is relatively unknown. We performed our experiments utilizing rat bone marrow derived MSCs. We observed that priming MSCs in hypoxia promotes maintenance of stem-like characteristics, with greater expression of typical MSC cell-surface markers, increased proliferation, and maintenance of differentiation potential. Addition of autologous MSCs to CD4+/allogeneic endothelial cell (EC) co-culture increases regulatory T cell (Treg) proliferation, which is further enhanced when MSCs are primed in hypoxia. Furthermore, MSC-mediated Treg expansion does not require direct cell-cell contact. The expression of indolamine 2,3-dioxygenase, a mediator of MSC immunomodulation, increases when MSCs are primed in hypoxia, and inhibition of IDO significantly decreases the expansion of Tregs. Priming with inflammatory cytokines IFNγ and TNFα increases also expression of markers associated with MSC immunomodulatory function, but decreases MSC proliferation. The expression of IDO also increases when MSCs are primed with inflammatory cytokines. However, there is no increase in Treg expansion when MSCs are primed with IFNγ, suggesting an alternate mechanism for inflammatory-stimulated MSC immunomodulation. Overall, these results suggest that MSCs primed in hypoxia or inflammatory conditions are optimally primed for immunosuppressive function. These results provide a clearer picture of how to enhance MSC immunomodulation for clinical use.

Suitability of human mesenchymal stem cells for gene therapy depends on the expansion medium

International Nuclear Information System (INIS)

Apel, Anja; Groth, Ariane; Schlesinger, Sabine; Bruns, Helge; Schemmer, Peter; Buechler, Markus W.; Herr, Ingrid

2009-01-01

Great hope is set in the use of mesenchymal stem cells for gene therapy and regenerative medicine. Since the frequency of this subpopulation of stem cells in bone marrow is low, mesenchymal stem cells are expanded ex vivo and manipulated prior to experimental or clinical use. Different methods for isolation and expansion are available, but the particular effect on the stem cell character is unclear. While the isolation of mesenchymal stem cells by density centrifugation followed by selection of the plastic adherent fraction is frequently used, the composition of expansion media differs. Thus, in the present study we cultured mesenchymal stem cells isolated from five healthy young volunteers in three widely used expansion media and performed a detailed analysis of the effect on morphology, proliferation, clonogenicity, passaging, differentiation and senescence. By this way we clearly show that the type of expansion medium used determines the stem cell character and time of senescence which is critical for future gene therapeutic and regenerative approaches using mesenchymal stem cells

Pleural mesothelial cells promote expansion of IL-17-producing CD8+ T cells in tuberculous pleural effusion.

Science.gov (United States)

Li, X; Zhou, Q; Yang, W B; Xiong, X Z; Du, R H; Zhang, J C

2013-05-01

IL-17-producing CD8(+) T lymphocytes (Tc17 cells) have recently been detected in many cancers and autoimmune diseases. However, the possible implication of Tc17 cells in tuberculous pleural effusion remains unclarified. In this study, distribution and phenotypic features of Tc17 cells in both tuberculous pleural effusion (TPE) and peripheral blood from patients with tuberculosis were determined. The effects of proinflammatory cytokines and local accessory cells (pleural mesothelial cells) on Tc17 cell expansion were also explored. We found that TPE contained more Tc17 cells than the blood. Compared with IFN-γ-producing CD8(+) T cells, Tc17 cells displayed higher expression of chemokine receptors (CCRs) and lower expression of cytotoxic molecules. In particularly, Tc17 cells in TPE exhibited high expression levels of CCR6, which could migrate in response to CCL20. Furthermore, IL-1β, IL-6, IL-23, or their various combinations could promote Tc17 cell expansion from CD8(+) T cells, whereas the proliferative response of Tc17 cells to above cytokines was lower than that of Th17 cells. Pleural mesothelial cells (PMCs) were able to stimulate Tc17 cell expansion via cell contact in an IL-1β/IL-6/IL-23 independent fashion. Thus this study demonstrates that Tc17 cells marks a subset of non-cytotoxic, CCR6(+) CD8(+) T lymphocytes with low proliferative capacity. The overrepresentation of Tc17 cells in TPE may be due to Tc17 cell expansion stimulated by pleural proinflammatory cytokines and to recruitment of Tc17 cells from peripheral blood. Additionally, PMCs may promote the production of IL-17 by CD8(+) T cells at sites of TPE via cell-cell interactions.

Rapid evolution of phenology during range expansion with recent climate change.

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Lustenhouwer, Nicky; Wilschut, Rutger A; Williams, Jennifer L; van der Putten, Wim H; Levine, Jonathan M

2018-02-01

Although climate warming is expected to make habitat beyond species' current cold range edge suitable for future colonization, this new habitat may present an array of biotic or abiotic conditions not experienced within the current range. Species' ability to shift their range with climate change may therefore depend on how populations evolve in response to such novel environmental conditions. However, due to the recent nature of thus far observed range expansions, the role of rapid adaptation during climate change migration is only beginning to be understood. Here, we evaluated evolution during the recent native range expansion of the annual plant Dittrichia graveolens, which is spreading northward in Europe from the Mediterranean region. We examined genetically based differentiation between core and edge populations in their phenology, a trait that is likely under selection with shorter growing seasons and greater seasonality at northern latitudes. In parallel common garden experiments at range edges in Switzerland and the Netherlands, we grew plants from Dutch, Swiss, and central and southern French populations. Population genetic analysis following RAD-sequencing of these populations supported the hypothesized central France origins of the Swiss and Dutch range edge populations. We found that in both common gardens, northern plants flowered up to 4 weeks earlier than southern plants. This differentiation in phenology extended from the core of the range to the Netherlands, a region only reached from central France over approximately the last 50 years. Fitness decreased as plants flowered later, supporting the hypothesized benefits of earlier flowering at the range edge. Our results suggest that native range expanding populations can rapidly adapt to novel environmental conditions in the expanded range, potentially promoting their ability to spread. © 2017 John Wiley & Sons Ltd.

The effects of micro-implant assisted rapid palatal expansion (MARPE) on the nasomaxillary complex--a finite element method (FEM) analysis.

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MacGinnis, Matt; Chu, Howard; Youssef, George; Wu, Kimberley W; Machado, Andre Wilson; Moon, Won

2014-08-29

Orthodontic palatal expansion appliances have been widely used with satisfactory and, most often, predictable clinical results. Recently, clinicians have successfully utilized micro-implants with palatal expander designs to work as anchors to the palate to achieve more efficient skeletal expansion and to decrease undesired dental effects. The purpose of the study was to use finite element method (FEM) to determine the stress distribution and displacement within the craniofacial complex when simulated conventional and micro-implant-assisted rapid palatal expansion (MARPE) expansion forces are applied to the maxilla. The simulated stress distribution produced within the palate and maxillary buttresses in addition to the displacement and rotation of the maxilla could then be analyzed to determine if micro-implants aid in skeletal expansion. A three-dimensional (3D) mesh model of the cranium with associated maxillary sutures was developed using computed tomography (CT) images and Mimics modeling software. To compare transverse expansion stresses in rapid palatal expansion (RPE) and MARPE, expansion forces were distributed to differing points on the maxilla and evaluated with ANSYS simulation software. The stresses distributed from forces applied to the maxillary teeth are distributed mainly along the trajectories of the three maxillary buttresses. In comparison, the MARPE showed tension and compression directed to the palate, while showing less rotation, and tipping of the maxillary complex. In addition, the conventional hyrax displayed a rotation of the maxilla around the teeth as opposed to the midpalatal suture of the MARPE. This data suggests that the MARPE causes the maxilla to bend laterally, while preventing unwanted rotation of the complex. In conclusion, the MARPE may be beneficial for hyperdivergent patients, or those that have already experienced closure of the midpalatal suture, who require palatal expansion and would worsen from buccal tipping of the teeth

Mechanical feedback coordinates cell wall expansion and assembly in yeast mating morphogenesis

Science.gov (United States)

2018-01-01

The shaping of individual cells requires a tight coordination of cell mechanics and growth. However, it is unclear how information about the mechanical state of the wall is relayed to the molecular processes building it, thereby enabling the coordination of cell wall expansion and assembly during morphogenesis. Combining theoretical and experimental approaches, we show that a mechanical feedback coordinating cell wall assembly and expansion is essential to sustain mating projection growth in budding yeast (Saccharomyces cerevisiae). Our theoretical results indicate that the mechanical feedback provided by the Cell Wall Integrity pathway, with cell wall stress sensors Wsc1 and Mid2 increasingly activating membrane-localized cell wall synthases Fks1/2 upon faster cell wall expansion, stabilizes mating projection growth without affecting cell shape. Experimental perturbation of the osmotic pressure and cell wall mechanics, as well as compromising the mechanical feedback through genetic deletion of the stress sensors, leads to cellular phenotypes that support the theoretical predictions. Our results indicate that while the existence of mechanical feedback is essential to stabilize mating projection growth, the shape and size of the cell are insensitive to the feedback. PMID:29346368

Measurement of Rapid Protein Diffusion in the Cytoplasm by Photo-Converted Intensity Profile Expansion

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Rotem Gura Sadovsky

2017-03-01

Full Text Available The fluorescence microscopy methods presently used to characterize protein motion in cells infer protein motion from indirect observables, rather than measuring protein motion directly. Operationalizing these methods requires expertise that can constitute a barrier to their broad utilization. Here, we have developed PIPE (photo-converted intensity profile expansion to directly measure the motion of tagged proteins and quantify it using an effective diffusion coefficient. PIPE works by pulsing photo-convertible fluorescent proteins, generating a peaked fluorescence signal at the pulsed region, and analyzing the spatial expansion of the signal. We demonstrate PIPE’s success in measuring accurate diffusion coefficients in silico and in vitro and compare effective diffusion coefficients of native cellular proteins and free fluorophores in vivo. We apply PIPE to measure diffusion anomality in the cell and use it to distinguish free fluorophores from native cellular proteins. PIPE’s direct measurement and ease of use make it appealing for cell biologists.

Asymmetric rapid maxillary expansion in true unilateral crossbite malocclusion: a prospective controlled clinical study.

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Ileri, Zehra; Basciftci, Faruk Ayhan

2015-03-01

To investigate the short-term effects of the asymmetric rapid maxillary (ARME) appliance on the vertical, sagittal, and transverse planes in patients with true unilateral posterior crossbite. Subjects were divided into two groups. The treatment group was comprised of 21 patients with unilateral posterior crossbite (mean age  =  13.3 ± 2.1 years). Members of this group were treated with the ARME appliance. The control group was comprised of 17 patients with Angle Class I who were kept under observation (mean age  =  12.3 ± 0.8 years). Lateral and frontal cephalograms were taken before the expansion (T1), immediately after expansion (T2), and at postexpansion retention (T3) in the treatment group and at preobservation (T1) and postobservation (T2) in the control group. A total of 34 measurements were assessed on cephalograms. For statistical analysis, the Wilcoxon test and analysis of covariance were used. The ARME appliance produced significant increases in nasal, maxillary base, upper arch, and lower arch dimensions (P ARME appliance created asymmetric increments in the transversal dimensions of the nose, maxilla, and upper arch in the short term. Asymmetric expansion therapy for subjects with unilateral maxillary deficiency may provide satisfactory outcomes in adolescents, with the exception of mandibular arch expansion. The triangular pattern of expansion caused clockwise rotation of the mandible and the occlusal plane and produced significant alterations in the vertical facial dimensions, whereas it created no displacement in maxilla in the sagittal plane.

Cytomegalovirus-Driven Adaptive-Like Natural Killer Cell Expansions Are Unaffected by Concurrent Chronic Hepatitis Virus Infections

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David F. G. Malone

2017-05-01

Full Text Available Adaptive-like expansions of natural killer (NK cell subsets are known to occur in response to human cytomegalovirus (CMV infection. These expansions are typically made up of NKG2C+ NK cells with particular killer-cell immunoglobulin-like receptor (KIR expression patterns. Such NK cell expansion patterns are also seen in patients with viral hepatitis infection. Yet, it is not known if the viral hepatitis infection promotes the appearance of such expansions or if effects are solely attributed to underlying CMV infection. In sizeable cohorts of CMV seropositive hepatitis B virus (HBV, hepatitis C virus (HCV, and hepatitis delta virus (HDV infected patients, we analyzed NK cells for expression of NKG2A, NKG2C, CD57, and inhibitory KIRs to assess the appearance of NK cell expansions characteristic of what has been seen in CMV seropositive healthy individuals. Adaptive-like NK cell expansions observed in viral hepatitis patients were strongly associated with CMV seropositivity. The number of subjects with these expansions did not differ between CMV seropositive viral hepatitis patients and corresponding healthy controls. Hence, we conclude that adaptive-like NK cell expansions observed in HBV, HCV, and/or HDV infected individuals are not caused by the chronic hepatitis infections per se, but rather are a consequence of underlying CMV infection.

Allogeneic cell therapy bioprocess economics and optimization: single-use cell expansion technologies.

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Simaria, Ana S; Hassan, Sally; Varadaraju, Hemanthram; Rowley, Jon; Warren, Kim; Vanek, Philip; Farid, Suzanne S

2014-01-01

For allogeneic cell therapies to reach their therapeutic potential, challenges related to achieving scalable and robust manufacturing processes will need to be addressed. A particular challenge is producing lot-sizes capable of meeting commercial demands of up to 10(9) cells/dose for large patient numbers due to the current limitations of expansion technologies. This article describes the application of a decisional tool to identify the most cost-effective expansion technologies for different scales of production as well as current gaps in the technology capabilities for allogeneic cell therapy manufacture. The tool integrates bioprocess economics with optimization to assess the economic competitiveness of planar and microcarrier-based cell expansion technologies. Visualization methods were used to identify the production scales where planar technologies will cease to be cost-effective and where microcarrier-based bioreactors become the only option. The tool outputs also predict that for the industry to be sustainable for high demand scenarios, significant increases will likely be needed in the performance capabilities of microcarrier-based systems. These data are presented using a technology S-curve as well as windows of operation to identify the combination of cell productivities and scale of single-use bioreactors required to meet future lot sizes. The modeling insights can be used to identify where future R&D investment should be focused to improve the performance of the most promising technologies so that they become a robust and scalable option that enables the cell therapy industry reach commercially relevant lot sizes. The tool outputs can facilitate decision-making very early on in development and be used to predict, and better manage, the risk of process changes needed as products proceed through the development pathway. © 2013 Wiley Periodicals, Inc.

Analyzing suitability for urban expansion under rapid coastal urbanization with remote sensing and GIS techniques: a case study of Linanyungang, China

DEFF Research Database (Denmark)

Zhao, Wenjun; Zhu, Xiaodong; Reenberg, Anette

2010-01-01

Beginning in 2000, Lianyungang's urbanization entered a period of rapid growth, spatially as well as economically. Rapid and intensive expansion of "construction land" imposed increasing pressures on regional environment. With the support of remote sensing data and GIS tools, this paper reports a...

With respect to coefficient of linear thermal expansion, bacterial vegetative cells and spores resemble plastics and metals, respectively.

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Nakanishi, Koichi; Kogure, Akinori; Fujii, Takenao; Kokawa, Ryohei; Deuchi, Keiji; Kuwana, Ritsuko; Takamatsu, Hiromu

2013-10-09

If a fixed stress is applied to the three-dimensional z-axis of a solid material, followed by heating, the amount of thermal expansion increases according to a fixed coefficient of thermal expansion. When expansion is plotted against temperature, the transition temperature at which the physical properties of the material change is at the apex of the curve. The composition of a microbial cell depends on the species and condition of the cell; consequently, the rate of thermal expansion and the transition temperature also depend on the species and condition of the cell. We have developed a method for measuring the coefficient of thermal expansion and the transition temperature of cells using a nano thermal analysis system in order to study the physical nature of the cells. The tendency was seen that among vegetative cells, the Gram-negative Escherichia coli and Pseudomonas aeruginosa have higher coefficients of linear expansion and lower transition temperatures than the Gram-positive Staphylococcus aureus and Bacillus subtilis. On the other hand, spores, which have low water content, overall showed lower coefficients of linear expansion and higher transition temperatures than vegetative cells. Comparing these trends to non-microbial materials, vegetative cells showed phenomenon similar to plastics and spores showed behaviour similar to metals with regards to the coefficient of liner thermal expansion. We show that vegetative cells occur phenomenon of similar to plastics and spores to metals with regard to the coefficient of liner thermal expansion. Cells may be characterized by the coefficient of linear expansion as a physical index; the coefficient of linear expansion may also characterize cells structurally since it relates to volumetric changes, surface area changes, the degree of expansion of water contained within the cell, and the intensity of the internal stress on the cellular membrane. The coefficient of linear expansion holds promise as a new index for

Effects of strontium ranelate on bone formation in the mid-palatal suture after rapid maxillary expansion

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Zhao SY

2015-05-01

Full Text Available Shuya Zhao,1,* Xuxia Wang,2,* Na Li,3 Yun Chen,1 Yuran Su,1 Jun Zhang1 1Department of Orthodontics, 2Department of Oral and Maxillofacial Surgery, Faculty of Stomatology, Shandong University; 3Department of Orthodontics, Shandong Provincial Qianfoshan Hospital, Jinan, People’s Republic of China *These authors contributed equally to this work Background: The aim of this experimental study was to investigate the effects of strontium ranelate on bone regeneration in the mid-palatal suture in response to rapid maxillary expansion (RME.Methods: Thirty-six male 6-week-old Wistar rats were randomly divided into three groups, ie, an expansion only (EO group, an expansion plus strontium ranelate (SE group, and a control group. An orthodontic appliance was set between the right and left upper molars of rats with an initial expansive force of 0.98 N. Rats in the SE group were administered strontium ranelate (600 mg/kg body weight and then euthanized in batches on days 4, 7, and 10. Morphological changes in the mid-palatal suture were investigated using micro-computed tomography and hematoxylin and eosin staining after RME. Bone morphogenetic protein-2 expression in the suture was also examined to evaluate bone formation in the mid-palatal suture. Image-Pro Plus software was then used to determine the mean optical density of the immunohistochemical images. Analysis of variance was used for statistical evaluation at the P<0.05 level.Results: With expansive force, the mid-palatal suture was expanded, but there was no statistically significant difference (P>0.05 between the SE and EO groups. The bone volume of the suture decreased after RME, but was higher in the SE group than in the EO group on days 7 and 10. Further, expression of bone morphogenetic protein-2 in the SE group was higher than in the other two groups (P<0.05.Conclusion: Strontium ranelate may hasten new bone formation in the expanded mid-palatal suture, which may be therapeutically

Xeno-Free Strategies for Safe Human Mesenchymal Stem/Stromal Cell Expansion: Supplements and Coatings

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M. Cimino

2017-01-01

Full Text Available Human mesenchymal stem/stromal cells (hMSCs have generated great interest in regenerative medicine mainly due to their multidifferentiation potential and immunomodulatory role. Although hMSC can be obtained from different tissues, the number of available cells is always low for clinical applications, thus requiring in vitro expansion. Most of the current protocols for hMSC expansion make use of fetal bovine serum (FBS as a nutrient-rich supplement. However, regulatory guidelines encourage novel xeno-free alternatives to define safer and standardized protocols for hMSC expansion that preserve their intrinsic therapeutic potential. Since hMSCs are adherent cells, the attachment surface and cell-adhesive components also play a crucial role on their successful expansion. This review focuses on the advantages/disadvantages of FBS-free media and surfaces/coatings that avoid the use of animal serum, overcoming ethical issues and improving the expansion of hMSC for clinical applications in a safe and reproducible way.

Large-Scale Culture and Genetic Modification of Human Natural Killer Cells for Cellular Therapy.

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Lapteva, Natalia; Parihar, Robin; Rollins, Lisa A; Gee, Adrian P; Rooney, Cliona M

2016-01-01

Recent advances in methods for the ex vivo expansion of human natural killer (NK) cells have facilitated the use of these powerful immune cells in clinical protocols. Further, the ability to genetically modify primary human NK cells following rapid expansion allows targeting and enhancement of their immune function. We have successfully adapted an expansion method for primary NK cells from peripheral blood mononuclear cells or from apheresis products in gas permeable rapid expansion devices (G-Rexes). Here, we describe an optimized protocol for rapid and robust NK cell expansion as well as a method for highly efficient retroviral transduction of these ex vivo expanded cells. These methodologies are good manufacturing practice (GMP) compliant and could be used for clinical-grade product manufacturing.

The C. elegans engrailed homolog ceh-16 regulates the self-renewal expansion division of stem cell-like seam cells.

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Huang, Xinxin; Tian, E; Xu, Yanhua; Zhang, Hong

2009-09-15

Stem cells undergo symmetric and asymmetric division to maintain the dynamic equilibrium of the stem cell pool and also to generate a variety of differentiated cells. The homeostatic mechanism controlling the choice between self-renewal and differentiation of stem cells is poorly understood. We show here that ceh-16, encoding the C. elegans ortholog of the transcription factor Engrailed, controls symmetric and asymmetric division of stem cell-like seam cells. Loss of function of ceh-16 causes certain seam cells, which normally undergo symmetric self-renewal expansion division with both daughters adopting the seam cell fate, to divide asymmetrically with only one daughter retaining the seam cell fate. The human engrailed homolog En2 functionally substitutes the role of ceh-16 in promoting self-renewal expansion division of seam cells. Loss of function of apr-1, encoding the C. elegans homolog of the Wnt signaling component APC, results in transformation of self-renewal maintenance seam cell division to self-renewal expansion division, leading to seam cell hyperplasia. The apr-1 mutation suppresses the seam cell division defect in ceh-16 mutants. Our study reveals that ceh-16 interacts with the Wnt signaling pathway to control the choice between self-renewal expansion and maintenance division and also demonstrates an evolutionarily conserved function of engrailed in promoting cell proliferation.

A review of decellularized stem cell matrix: a novel cell expansion system for cartilage tissue engineering

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M Pei

2011-11-01

Full Text Available Cell-based therapy is a promising biological approach for the treatment of cartilage defects. Due to the small size of autologous cartilage samples available for cell transplantation in patients, cells need to be expanded to yield a sufficient cell number for cartilage repair. However, chondrocytes and adult stem cells tend to become replicatively senescent once they are expanded on conventional plastic flasks. Many studies demonstrate that the loss of cell properties is concomitant with the decreased cell proliferation capacity. This is a significant challenge for cartilage tissue engineering and regeneration. Despite much progress having been made in cell expansion, there are still concerns over expanded cell size and quality for cell transplantation applications. Recently, in vivo investigations in stem cell niches have suggested the importance of developing an in vitro stem cell microenvironment for cell expansion and tissue-specific differentiation. Our and other investigators’ work indicates that a decellularized stem cell matrix (DSCM may provide such an expansion system to yield large-quantity and high-quality cells for cartilage tissue engineering and regeneration. This review briefly introduces key parameters in an in vivo stem cell niche and focuses on our recent work on DSCM for its rejuvenating or reprograming effect on various adult stem cells and chondrocytes. Since research in DSCM is still in its infancy, we are only able to discuss some potential mechanisms of DSCM on cell proliferation and chondrogenic potential. Further investigations of the underlying mechanism and in vivo regeneration capacity will allow this approach to be used in clinics.

Endothelial Cells Promote Expansion of Long‐Term Engrafting Marrow Hematopoietic Stem and Progenitor Cells in Primates

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Gori, Jennifer L.; Butler, Jason M.; Kunar, Balvir; Poulos, Michael G.; Ginsberg, Michael; Nolan, Daniel J.; Norgaard, Zachary K.; Adair, Jennifer E.; Rafii, Shahin

2016-01-01

Abstract Successful expansion of bone marrow (BM) hematopoietic stem and progenitor cells (HSPCs) would benefit many HSPC transplantation and gene therapy/editing applications. However, current expansion technologies have been limited by a loss of multipotency and self‐renewal properties ex vivo. We hypothesized that an ex vivo vascular niche would provide prohematopoietic signals to expand HSPCs while maintaining multipotency and self‐renewal. To test this hypothesis, BM autologous CD34+ cells were expanded in endothelial cell (EC) coculture and transplanted in nonhuman primates. CD34+C38− HSPCs cocultured with ECs expanded up to 17‐fold, with a significant increase in hematopoietic colony‐forming activity compared with cells cultured with cytokines alone (colony‐forming unit‐granulocyte‐erythroid‐macrophage‐monocyte; p < .005). BM CD34+ cells that were transduced with green fluorescent protein lentivirus vector and expanded on ECs engrafted long term with multilineage polyclonal reconstitution. Gene marking was observed in granulocytes, lymphocytes, platelets, and erythrocytes. Whole transcriptome analysis indicated that EC coculture altered the expression profile of 75 genes in the BM CD34+ cells without impeding the long‐term engraftment potential. These findings show that an ex vivo vascular niche is an effective platform for expansion of adult BM HSPCs. Stem Cells Translational Medicine 2017;6:864–876 PMID:28297579

Wall relaxation and the driving forces for cell expansive growth

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Cosgrove, D. J.

1987-01-01

When water uptake by growing cells is prevented, the turgor pressure and the tensile stress in the cell wall are reduced by continued wall loosening. This process, termed in vivo stress relaxation, provides a new way to study the dynamics of wall loosening and to measure the wall yield threshold and the physiological wall extensibility. Stress relaxation experiments indicate that wall stress supplies the mechanical driving force for wall yielding. Cell expansion also requires water absorption. The driving force for water uptake during growth is created by wall relaxation, which lowers the water potential of the expanding cells. New techniques for measuring this driving force show that it is smaller than believed previously; in elongating stems it is only 0.3 to 0.5 bar. This means that the hydraulic resistance of the water transport pathway is small and that rate of cell expansion is controlled primarily by wall loosening and yielding.

Intrinsic Plasma Cell Differentiation Defects in B Cell Expansion with NF-κB and T Cell Anergy Patient B Cells

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Swadhinya Arjunaraja

2017-08-01

Full Text Available B cell Expansion with NF-κB and T cell Anergy (BENTA disease is a novel B cell lymphoproliferative disorder caused by germline, gain-of-function mutations in the lymphocyte scaffolding protein CARD11, which drives constitutive NF-κB signaling. Despite dramatic polyclonal expansion of naive and immature B cells, BENTA patients also present with signs of primary immunodeficiency, including markedly reduced percentages of class-switched/memory B cells and poor humoral responses to certain vaccines. Using purified naive B cells from our BENTA patient cohort, here we show that BENTA B cells exhibit intrinsic defects in B cell differentiation. Despite a profound in vitro survival advantage relative to normal donor B cells, BENTA patient B cells were severely impaired in their ability to differentiate into short-lived IgDloCD38hi plasmablasts or CD138+ long-lived plasma cells in response to various stimuli. These defects corresponded with diminished IgG antibody production and correlated with poor induction of specific genes required for plasma cell commitment. These findings provide important mechanistic clues that help explain both B cell lymphocytosis and humoral immunodeficiency in BENTA disease.

Coordinate expansion of murine hematopoietic and mesenchymal stem cell compartments by SHIPi.

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Brooks, R; Iyer, S; Akada, H; Neelam, S; Russo, C M; Chisholm, J D; Kerr, W G

2015-03-01

Promoting the expansion of adult stem cell populations offers the potential to ameliorate radiation or chemotherapy-induced bone marrow failure and allows for expedited recovery for patients undergoing these therapies. Previous genetic studies suggested a pivotal role for SH2 domain-containing inositol-5-phosphatase 1 (SHIP1) in limiting the size of the hematopoietic stem cell (HSC) compartment. The aim of this study was to determine whether our recent development of small molecule SHIP1 inhibitors offers the potential for pharmacological expansion of the HSC compartment in vivo. We show here that treatment of mice with aminosteroid inhibitors of SHIP1 (SHIPi) more than doubles the size of the adult mesenchymal stem cell (MSC) compartment while simultaneously expanding the HSC pool sixfold. Consistent with its ability to target SHIP1 function in vivo, SHIPi also significantly increases plasma granulocyte colony-stimulating factor (G-CSF) levels, a growth factor that supports proliferation of HSC. Here, we show that SHIPi-induced G-CSF production mediates HSC and MSC expansion, as in vivo neutralization of G-CSF abrogates the SHIPi-induced expansion of both the HSC and MSC compartments. Due to its expansionary effect on adult stem cell compartments, SHIPi represents a potential novel strategy to improve declining stem cell function in both therapy induced and genetically derived bone marrow failure syndromes. © 2014 AlphaMed Press.

Endothelial Cells Promote Expansion of Long-Term Engrafting Marrow Hematopoietic Stem and Progenitor Cells in Primates.

Science.gov (United States)

Gori, Jennifer L; Butler, Jason M; Kunar, Balvir; Poulos, Michael G; Ginsberg, Michael; Nolan, Daniel J; Norgaard, Zachary K; Adair, Jennifer E; Rafii, Shahin; Kiem, Hans-Peter

2017-03-01

Successful expansion of bone marrow (BM) hematopoietic stem and progenitor cells (HSPCs) would benefit many HSPC transplantation and gene therapy/editing applications. However, current expansion technologies have been limited by a loss of multipotency and self-renewal properties ex vivo. We hypothesized that an ex vivo vascular niche would provide prohematopoietic signals to expand HSPCs while maintaining multipotency and self-renewal. To test this hypothesis, BM autologous CD34 + cells were expanded in endothelial cell (EC) coculture and transplanted in nonhuman primates. CD34 + C38 - HSPCs cocultured with ECs expanded up to 17-fold, with a significant increase in hematopoietic colony-forming activity compared with cells cultured with cytokines alone (colony-forming unit-granulocyte-erythroid-macrophage-monocyte; p < .005). BM CD34 + cells that were transduced with green fluorescent protein lentivirus vector and expanded on ECs engrafted long term with multilineage polyclonal reconstitution. Gene marking was observed in granulocytes, lymphocytes, platelets, and erythrocytes. Whole transcriptome analysis indicated that EC coculture altered the expression profile of 75 genes in the BM CD34 + cells without impeding the long-term engraftment potential. These findings show that an ex vivo vascular niche is an effective platform for expansion of adult BM HSPCs. Stem Cells Translational Medicine 2017;6:864-876. © 2016 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Colony Expansion of Socially Motile Myxococcus xanthus Cells Is Driven by Growth, Motility, and Exopolysaccharide Production.

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Pintu Patra

2016-06-01

Full Text Available Myxococcus xanthus, a model organism for studies of multicellular behavior in bacteria, moves exclusively on solid surfaces using two distinct but coordinated motility mechanisms. One of these, social (S motility is powered by the extension and retraction of type IV pili and requires the presence of exopolysaccharides (EPS produced by neighboring cells. As a result, S motility requires close cell-to-cell proximity and isolated cells do not translocate. Previous studies measuring S motility by observing the colony expansion of cells deposited on agar have shown that the expansion rate increases with initial cell density, but the biophysical mechanisms involved remain largely unknown. To understand the dynamics of S motility-driven colony expansion, we developed a reaction-diffusion model describing the effects of cell density, EPS deposition and nutrient exposure on the expansion rate. Our results show that at steady state the population expands as a traveling wave with a speed determined by the interplay of cell motility and growth, a well-known characteristic of Fisher's equation. The model explains the density-dependence of the colony expansion by demonstrating the presence of a lag phase-a transient period of very slow expansion with a duration dependent on the initial cell density. We propose that at a low initial density, more time is required for the cells to accumulate enough EPS to activate S-motility resulting in a longer lag period. Furthermore, our model makes the novel prediction that following the lag phase the population expands at a constant rate independent of the cell density. These predictions were confirmed by S motility experiments capturing long-term expansion dynamics.

Colony Expansion of Socially Motile Myxococcus xanthus Cells Is Driven by Growth, Motility, and Exopolysaccharide Production.

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Patra, Pintu; Kissoon, Kimberley; Cornejo, Isabel; Kaplan, Heidi B; Igoshin, Oleg A

2016-06-01

Myxococcus xanthus, a model organism for studies of multicellular behavior in bacteria, moves exclusively on solid surfaces using two distinct but coordinated motility mechanisms. One of these, social (S) motility is powered by the extension and retraction of type IV pili and requires the presence of exopolysaccharides (EPS) produced by neighboring cells. As a result, S motility requires close cell-to-cell proximity and isolated cells do not translocate. Previous studies measuring S motility by observing the colony expansion of cells deposited on agar have shown that the expansion rate increases with initial cell density, but the biophysical mechanisms involved remain largely unknown. To understand the dynamics of S motility-driven colony expansion, we developed a reaction-diffusion model describing the effects of cell density, EPS deposition and nutrient exposure on the expansion rate. Our results show that at steady state the population expands as a traveling wave with a speed determined by the interplay of cell motility and growth, a well-known characteristic of Fisher's equation. The model explains the density-dependence of the colony expansion by demonstrating the presence of a lag phase-a transient period of very slow expansion with a duration dependent on the initial cell density. We propose that at a low initial density, more time is required for the cells to accumulate enough EPS to activate S-motility resulting in a longer lag period. Furthermore, our model makes the novel prediction that following the lag phase the population expands at a constant rate independent of the cell density. These predictions were confirmed by S motility experiments capturing long-term expansion dynamics.

Dynamics of gas cell coalescence during baking expansion of leavened dough.

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Miś, Antoni; Nawrocka, Agnieszka; Lamorski, Krzysztof; Dziki, Dariusz

2018-01-01

The investigation of the dynamics of gas cell coalescence, i.e. a phenomenon that deteriorates the homogeneity of the cellular structure of bread crumb, was carried out performing simultaneously measurements of the dough volume, pressure, and viscosity. It was demonstrated that, during the baking expansion of chemically leavened wheat flour dough, the maximum growth rate of the gas cell radius determined from the ratio of pressure exerted by the expanded dough to its viscosity was on average four-fold lower than that calculated from volume changes in the gas phase of the dough. Such a high discrepancy was interpreted as a result of the course of coalescence, and a formula for determination of its rate was developed. The coalescence rate in the initial baking expansion phase had negative values, indicating nucleation of newly formed gas cells, which increased the number of gas cells even by 8%. In the next baking expansion phase, the coalescence rate started to exhibit positive values, reflecting dominance of the coalescence phenomenon over nucleation. The maximum coalescence rates indicate that, during the period of the most intensive dough expansion, the number of gas cells decreased by 2-3% within one second. At the end of the formation of bread crumb, the number of the gas cells declined by 55-67% in comparison with the initial value. The correctness of the results was positively verified using X-ray micro-computed tomography. The developed method can be a useful tool for more profound exploration of the coalescence phenomenon at various stages of evolution of the cellular structure and its determinants, which may contribute to future development of more effective methods for improving the texture and sensory quality of bread crumb. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rapid replacement of bridge deck expansion joints study - phase I.

Science.gov (United States)

2014-12-01

Bridge deck expansion joints are used to allow for movement of the bridge deck due to thermal expansion, dynamics loading, and : other factors. More recently, expansion joints have also been utilized to prevent the passage of winter de-icing chemical...

Evaluation of the rapid and slow maxillary expansion using cone-beam computed tomography: a randomized clinical trial

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Juliana da S. Pereira

Full Text Available ABSTRACT OBJECTIVE: The aim of this randomized clinical trial was to evaluate the dental, dentoalveolar, and skeletal changes occurring right after the rapid maxillary expansion (RME and slow maxillary expansion (SME treatment using Haas-type expander. METHODS: All subjects performed cone-beam computed tomography (CBCT before installation of expanders (T1 and right after screw stabilization (T2. Patients who did not follow the research parameters were excluded. The final sample resulted in 21 patients in RME group (mean age of 8.43 years and 16 patients in SME group (mean age of 8.70 years. Based on the skewness and kurtosis statistics, the variables were judged to be normally distributed and paired t-test and student t-test were performed at significance level of 5%. RESULTS: Intermolar angle changed significantly due to treatment and RME showed greater buccal tipping than SME. RME showed significant changes in other four measurements due to treatment: maxilla moved forward and mandible showed backward rotation and, at transversal level both skeletal and dentoalveolar showed significant changes due to maxillary expansion. SME showed significant dentoalveolar changes due to maxillary expansion. CONCLUSIONS: Only intermolar angle showed significant difference between the two modalities of maxillary expansion with greater buccal tipping for RME. Also, RME produced skeletal maxillary expansion and SME did not. Both maxillary expansion modalities were efficient to promote transversal gain at dentoalveolar level. Sagittal and vertical measurements did not show differences between groups, but RME promoted a forward movement of the maxilla and backward rotation of the mandible.

Xenografts in zebrafish embryos as a rapid functional assay for breast cancer stem-like cell identification.

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Eguiara, Arrate; Holgado, Olaia; Beloqui, Izaskun; Abalde, Leire; Sanchez, Yolanda; Callol, Carles; Martin, Angel G

2011-11-01

The cancer stem cell is defined by its capacity to self-renew, the potential to differentiate into all cells of the tumor and the ability to proliferate and drive the expansion of the tumor. Thus, targeting these cells may provide novel anti-cancer treatment strategies. Breast cancer stem cells have been isolated according to surface marker expression, ability to efflux fluorescent dyes, increased activity of aldehyde dehydrogenase or the capacity to form spheres in non-adherent culture conditions. In order to test novel drugs directed towards modulating self-renewal of cancer stem cells, rapid, easy and inexpensive assays must be developed. Using 2 days-post-fertilization (dpf) zebrafish embryos as transplant recipients, we show that cells grown in mammospheres from breast carcinoma cell lines migrate to the tail of the embryo and form masses with a significantly higher frequency than parental monolayer populations. When stem-like self-renewal was targeted in the parental population by the use of the dietary supplement curcumin, cell migration and mass formation were reduced, indicating that these effects were associated with stem-like cell content. This is a proof of principle report that proposes a rapid and inexpensive assay to target in vivo cancer stem-like cells, which may be used to unravel basic cancer stem cell biology and for drug screening.

Umbilical cord bloods hematopoietic stem cells ex vivo expansion (the literature review

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T. V. Shamanskaya

2012-01-01

Full Text Available Umbilical cord blood (CB is now an attractive source of hematopoietic stem cells (HSCs for transplantation in pediatric and adult patients with various malignant and non-malignant diseases. However, its clinical application is limited by low cells numbers in graft, which correlates with delayed engraftment, an extension of time to platelets and neutrophils recovery and increasing risk of infectious complications. Several strategies have been suggested to overcome this limitation, one of which is obtaining a sufficient number of hematopoietic progenitor cells by ex vivo expansion. Literature review about CB HSCs expansion in given article is presented.

Integrated processes for expansion and differentiation of human pluripotent stem cells in suspended microcarriers cultures

International Nuclear Information System (INIS)

Lam, Alan Tin-Lun; Chen, Allen Kuan-Liang; Ting, Sherwin Qi-Peng; Reuveny, Shaul; Oh, Steve Kah-Weng

2016-01-01

Current methods for human pluripotent stem cells (hPSC) expansion and differentiation can be limited in scalability and costly (due to their labor intensive nature). This can limit their use in cell therapy, drug screening and toxicity assays. One of the approaches that can overcome these limitations is microcarrier (MC) based cultures in which cells are expanded as cell/MC aggregates and then directly differentiated as embryoid bodies (EBs) in the same agitated reactor. This integrated process can be scaled up and eliminate the need for some culture manipulation used in common monolayer and EBs cultures. This review describes the principles of such microcarriers based integrated hPSC expansion and differentiation process, and parameters that can affect its efficiency (such as MC type and extracellular matrix proteins coatings, cell/MC aggregates size, and agitation). Finally examples of integrated process for generation cardiomyocytes (CM) and neural progenitor cells (NPC) as well as challenges to be solved are described. - Highlights: • Expansion of hPSC on microcarriers. • Differentiation of hPSC on microcarriers. • Parameters that can affect the expansion and differentiation of hPSC on microcarriers. • Integration of expansion and differentiation of hPSC on microcarriers in one unit operation.

Integrated processes for expansion and differentiation of human pluripotent stem cells in suspended microcarriers cultures

Energy Technology Data Exchange (ETDEWEB)

Lam, Alan Tin-Lun, E-mail: alan_lam@bti.a-star.edu.sg; Chen, Allen Kuan-Liang; Ting, Sherwin Qi-Peng; Reuveny, Shaul; Oh, Steve Kah-Weng, E-mail: steve_oh@bti.a-star.edu.sg

2016-05-06

Current methods for human pluripotent stem cells (hPSC) expansion and differentiation can be limited in scalability and costly (due to their labor intensive nature). This can limit their use in cell therapy, drug screening and toxicity assays. One of the approaches that can overcome these limitations is microcarrier (MC) based cultures in which cells are expanded as cell/MC aggregates and then directly differentiated as embryoid bodies (EBs) in the same agitated reactor. This integrated process can be scaled up and eliminate the need for some culture manipulation used in common monolayer and EBs cultures. This review describes the principles of such microcarriers based integrated hPSC expansion and differentiation process, and parameters that can affect its efficiency (such as MC type and extracellular matrix proteins coatings, cell/MC aggregates size, and agitation). Finally examples of integrated process for generation cardiomyocytes (CM) and neural progenitor cells (NPC) as well as challenges to be solved are described. - Highlights: • Expansion of hPSC on microcarriers. • Differentiation of hPSC on microcarriers. • Parameters that can affect the expansion and differentiation of hPSC on microcarriers. • Integration of expansion and differentiation of hPSC on microcarriers in one unit operation.

Residues 39-56 of Stem Cell Factor Protein Sequence Are Capable of Stimulating the Expansion of Cord Blood CD34+ Cells.

Science.gov (United States)

Shen, Bin; Jiang, Wenhong; Fan, Jie; Dai, Wei; Ding, Xinxin; Jiang, Yongping

2015-01-01

Stem cell factor (SCF) can stimulate hematopoietic stem cell (HSC) expansion; however, the specific structural region(s) of SCF protein that are critical for this function are still unknown. A novel monoclonal antibody (named 23C8) against recombinant human SCF (rhSCF) was previously found to inhibit the ability of rhSCF to enhance HSC expansion, making it possible to identify the relevant active region to HSC. Eleven polypeptides were synthesized, which were designed to cover the full-length of rhSCF, with overlaps that are at least 3 amino acids long. ELISA was used to identify the polypeptide(s) that specifically react with the anti-SCF. The effects of the synthetic polypeptides on human HSC expansion, or on the ability of the full-length rhSCF to stimulate cell proliferation, were evaluated ex vivo. Total cell number was monitored using hemocytometer whereas CD34+ cell number was calculated based on the proportion determined via flow cytometry on day 6 of culture. Of all polypeptides analyzed, only one, named P0, corresponding to the SCF protein sequence at residues 39-56, was recognized by 23C8 mAb during ELISA. P0 stimulated the expansion of CD34+ cells derived from human umbilical cord blood (UCB). Addition of P0 increased the numbers of total mononucleated cells and CD34+ cells, by ~2 fold on day 6. P0 also showed partial competition against full-length rhSCF in the ex vivo cell expansion assay. Residues 39-56 of rhSCF comprise a critical functional region for its ability to enhance expansion of human UCB CD34+ cells. The peptide P0 is a potential candidate for further development as a synthetic substitute for rhSCF in laboratory and clinical applications.

Good manufacturing practice-compliant expansion of marrow-derived stem and progenitor cells for cell therapy.

Science.gov (United States)

Gastens, Martin H; Goltry, Kristin; Prohaska, Wolfgang; Tschöpe, Diethelm; Stratmann, Bernd; Lammers, Dirk; Kirana, Stanley; Götting, Christian; Kleesiek, Knut

2007-01-01

Ex vivo expansion is being used to increase the number of stem and progenitor cells for autologous cell therapy. Initiation of pivotal clinical trials testing the efficacy of these cells for tissue repair has been hampered by the challenge of assuring safe and high-quality cell production. A strategy is described here for clinical-scale expansion of bone marrow (BM)-derived stem cells within a mixed cell population in a completely closed process from cell collection through postculture processing using sterile connectable devices. Human BM mononuclear cells (BMMNC) were isolated, cultured for 12 days, and washed postharvest using either standard open procedures in laminar flow hoods or using automated closed systems. Conditions for these studies were similar to long-term BM cultures in which hematopoietic and stromal components are cultured together. Expansion of marrow-derived stem and progenitor cells was then assessed. Cell yield, number of colony forming units (CFU), phenotype, stability, and multilineage differentiation capacity were compared from the single pass perfusion bioreactor and standard flask cultures. Purification of BMMNC using a closed Ficoll gradient process led to depletion of 98% erythrocytes and 87% granulocytes, compared to 100% and 70%, respectively, for manual processing. After closed system culture, mesenchymal progenitors, measured as CD105+CD166+CD14-CD45- and fibroblastic CFU, expanded 317- and 364-fold, respectively, while CD34+ hematopoietic progenitors were depleted 10-fold compared to starting BMMNC. Cultured cells exhibited multilineage differentiation by displaying adipogenic, osteogenic, and endothelial characteristics in vitro. No significant difference was observed between manual and bioreactor cultures. Automated culture and washing of the cell product resulted in 181 x 10(6) total cells that were viable and contained fibroblastic CFU for at least 24 h of storage. A combination of closed, automated technologies enabled

Linear thermal expansion coefficient measurement technology in hot cell

International Nuclear Information System (INIS)

Park, Dae Gyu; Choo, Yong Sun; Ahn, Sang Bok; Hong, Kwon Pyo; Lee, K. S.

1998-06-01

To establish linear thermal expansion coefficient measurement technology in hot cell, we reviewed and evaluated various measuring technology by paper and these were compared with the data produced with pre-installed dilatometer in hot cell. Detailed contents are as follows; - The theory of test. - Review of characteristics for various measurement technology and compatibility with hot cell. - Review of standard testing regulations(ASTM). - System calibration of pre-installed dilatometer. - Performance test of pre-installed dilatometer. (author). 12 refs., 15 tabs., 8 figs

The Role of Auxin in Cell Wall Expansion.

Science.gov (United States)

Majda, Mateusz; Robert, Stéphanie

2018-03-22

Plant cells are surrounded by cell walls, which are dynamic structures displaying a strictly regulated balance between rigidity and flexibility. Walls are fairly rigid to provide support and protection, but also extensible, to allow cell growth, which is triggered by a high intracellular turgor pressure. Wall properties regulate the differential growth of the cell, resulting in a diversity of cell sizes and shapes. The plant hormone auxin is well known to stimulate cell elongation via increasing wall extensibility. Auxin participates in the regulation of cell wall properties by inducing wall loosening. Here, we review what is known on cell wall property regulation by auxin. We focus particularly on the auxin role during cell expansion linked directly to cell wall modifications. We also analyze downstream targets of transcriptional auxin signaling, which are related to the cell wall and could be linked to acid growth and the action of wall-loosening proteins. All together, this update elucidates the connection between hormonal signaling and cell wall synthesis and deposition.

Resistin enhances the expansion of regulatory T cells through modulation of dendritic cells

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Han Seung

2010-06-01

Full Text Available Abstract Background Resistin, a member of adipokine family, is known to be involved in the modulation of immune responses including inflammatory activity. Interestingly, resistin is secreted by adipocytes in mice and rats whereas it is secreted by leukocytes in humans. However, the mechanism behind the effect of resistin on the expansion of regulatory T cells (Tregs remains poorly understood. Therefore, we examined regulatory effect of resistin on the induction and cellular modification of Tregs. Results Both protein and mRNA expression of FoxP3, a representative marker of Tregs, increased in a dose-dependent manner when peripheral blood mononuclear cells were treated with resistin. At the same time, resistin had no direct effect on the induction of FoxP3 in CD4+ T cells, suggesting an indirect role through other cells type(s. Since DCs are an important player in the differentiation of T cells, we focused on the role of DCs in the modulation of Tregs by resistin. Resistin suppressed the expression of interferon regulatory factor (IRF-1 and its target cytokines, IL-6, IL-23p19 and IL-12p40, in DCs. Furthermore, FoxP3 expression is increased in CD4+ T cells when co-cultured with DCs and concomitantly treated with resistin. Conclusion Our results suggest that resistin induces expansion of functional Tregs only when co-cultured with DCs.

A Rapid Cell Expansion Process for Production of Engineered Autologous CAR-T Cell Therapies.

Science.gov (United States)

Lu, Tangying Lily; Pugach, Omar; Somerville, Robert; Rosenberg, Steven A; Kochenderfer, James N; Better, Marc; Feldman, Steven A

2016-12-01

The treatment of B-cell malignancies by adoptive cell transfer (ACT) of anti-CD19 chimeric antigen receptor T cells (CD19 CAR-T) has proven to be a highly successful therapeutic modality in several clinical trials. 1-6 The anti-CD19 CAR-T cell production method used to support initial trials relied on numerous manual, open process steps, human serum, and 10 days of cell culture to achieve a clinical dose. 7 This approach limited the ability to support large multicenter clinical trials, as well as scale up for commercial cell production. Therefore, studies were completed to streamline and optimize the original National Cancer Institute production process by removing human serum from the process in order to minimize the risk of viral contamination, moving process steps from an open system to functionally closed system operations in order to minimize the risk of microbial contamination, and standardizing additional process steps in order to maximize process consistency. This study reports a procedure for generating CD19 CAR-T cells in 6 days, using a functionally closed manufacturing process and defined, serum-free medium. This method is able to produce CD19 CAR-T cells that are phenotypically and functionally indistinguishable from cells produced for clinical trials by the previously described production process.

Efficient expansion of mesenchymal stromal cells in a disposable fixed bed culture system.

Science.gov (United States)

Mizukami, Amanda; Orellana, Maristela D; Caruso, Sâmia R; de Lima Prata, Karen; Covas, Dimas T; Swiech, Kamilla

2013-01-01

The need for efficient and reliable technologies for clinical-scale expansion of mesenchymal stromal cells (MSC) has led to the use of disposable bioreactors and culture systems. Here, we evaluate the expansion of cord blood-derived MSC in a disposable fixed bed culture system. Starting from an initial cell density of 6.0 × 10(7) cells, after 7 days of culture, it was possible to produce of 4.2(±0.8) × 10(8) cells, which represents a fold increase of 7.0 (±1.4). After enzymatic retrieval from Fibra-Cell disks, the cells were able to maintain their potential for differentiation into adipocytes and osteocytes and were positive for many markers common to MSC (CD73, CD90, and CD105). The results obtained in this study demonstrate that MSC can be efficiently expanded in the culture system. This novel approach presents several advantages over the current expansion systems, based on culture flasks or microcarrier-based spinner flasks and represents a key element for MSC cellular therapy according to GMP compliant clinical-scale production system. Copyright © 2013 American Institute of Chemical Engineers.

Development of maternal seed tissue in barley is mediated by regulated cell expansion and cell disintegration and coordinated with endosperm growth.

Science.gov (United States)

Radchuk, Volodymyr; Weier, Diana; Radchuk, Ruslana; Weschke, Winfriede; Weber, Hans

2011-01-01

After fertilization, filial grain organs are surrounded by the maternal nucellus embedded within the integuments and pericarp. Rapid early endosperm growth must be coordinated with maternal tissue development. Parameters of maternal tissue growth and development were analysed during early endosperm formation. In the pericarp, cell proliferation is accomplished around the time of fertilization, followed by cell elongation predominantly in longitudinal directions. The rapid cell expansion coincides with endosperm cellularization. Distribution of TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling)-positive nuclei reveals distinct patterns starting in the nucellus at anthesis and followed later by the inner cell rows of the pericarp, then spreading to the whole pericarp. The pattern suggests timely and spatially regulated programmed cell death (PCD) processes in maternal seed tissues. When the endosperm is coenocytic, PCD events are only observed within the nucellus. Thereby, remobilization of nucellar storage compounds by PCD could nourish the early developing endosperm when functional interconnections are absent between maternal and filial seed organs. Specific proteases promote PCD events. Characterization of the barley vacuolar processing enzyme (VPE) gene family identified seven gene members specifically expressed in the developing grain. HvVPE2a (known as nucellain) together with closely similar HvVPE2b and HvVPE2d might be involved in nucellar PCD. HvVPE4 is strongly cell specific for pericarp parenchyma. Correlative evidence suggests that HvVPE4 plays a role in PCD events in the pericarp. Possible functions of PCD in the maternal tissues imply a potential nutritive role or the relief of a physical restraint for endosperm growth. PCD could also activate post-phloem transport functions.

Model of oronasal rehabilitation in children with obstructive sleep apnea syndrome undergoing rapid maxillary expansion: Research review

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Luca Levrini

2014-12-01

Full Text Available Rapid maxillary expansion (RME is a widely used practice in orthodontics. Scientific evidence shows that RME can be helpful in modifying the breathing pattern in mouth-breathing patients. In order to promote the restoration of physiological breathing we have developed a rehabilitation program associated with RME in children. The aim of the study was a literature review and a model of orofacial rehabilitation in children with obstructive sleep apnea undergoing treatment with rapid maxillary expansion. Muscular training (local exercises and general ones is the key factor of the program. It also includes hygienic and behavior instructions as well as other therapeutic procedures such as rhinosinusal washes, a postural re-education (Alexander technique and, if necessary, a pharmacological treatment aimed to improve nasal obstruction. The program should be customized for each patient. If RME is supported by an adequate functional rehabilitation, the possibility to change the breathing pattern is considerably amplified. Awareness, motivation and collaboration of the child and their parents, as well as the cooperation among specialists, such as orthodontist, speech therapist, pediatrician and otolaryngologist, are necessary conditions to achieve the goal.

Orthodontic Treatment with Rapid Maxillary Expansion for Treating a Boy with Severe Obstructive Sleep Apnea

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Myungrip Kim

2014-06-01

Full Text Available This case report shows that orthodontic treatment with rapid maxillary expansion (RME is an effective treatment option for managing pediatric obstructive sleep apnea (OSA. An 11-year-old boy with severe pediatric OSA received comprehensive orthodontic treatment with RME. Four sleep studies were done: before orthodontic treatment, after RME, just after comprehensive orthodontic treatment and at the 2-year and 5-month follow-up. Polysomnographic findings showed that the orthodontic treatment with RME was successful for managing severe OSA in the patient.

Clonal expansion under the microscope: studying lymphocyte activation and differentiation using live-cell imaging.

Science.gov (United States)

Polonsky, Michal; Chain, Benjamin; Friedman, Nir

2016-03-01

Clonal expansion of lymphocytes is a hallmark of vertebrate adaptive immunity. A small number of precursor cells that recognize a specific antigen proliferate into expanded clones, differentiate and acquire various effector and memory phenotypes, which promote effective immune responses. Recent studies establish a large degree of heterogeneity in the level of expansion and in cell state between and within expanding clones. Studying these processes in vivo, while providing insightful information on the level of heterogeneity, is challenging due to the complex microenvironment and the inability to continuously track individual cells over extended periods of time. Live cell imaging of ex vivo cultures within micro fabricated arrays provides an attractive methodology for studying clonal expansion. These experiments facilitate continuous acquisition of a large number of parameters on cell number, proliferation, death and differentiation state, with single-cell resolution on thousands of expanding clones that grow within controlled environments. Such data can reveal stochastic and instructive mechanisms that contribute to observed heterogeneity and elucidate the sequential order of differentiation events. Intercellular interactions can also be studied within these arrays by following responses of a controlled number of interacting cells, all trapped within the same microwell. Here we describe implementations of live-cell imaging within microwell arrays for studies of lymphocyte clonal expansion, portray insights already gained from these experiments and outline directions for future research. These tools, together with in vivo experiments tracking single-cell responses, will expand our understanding of adaptive immunity and the ways by which it can be manipulated.

Treatment of skeletal class III malocclusion using face mask therapy with alternate rapid maxillary expansion and constriction (Alt-RAMEC protocol

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Anand Ramchandra Rathi

2015-01-01

Full Text Available Class III malocclusion is very common malocclusion and can be due to maxillary retrusion, mandibular prognathism, or combination. Ellis and McNamara found a combination of maxillary retrusion and mandibular protrusion to be the most common skeletal relationship (30%. The treatment should be carried out as early as possible for permitting normal growth of the skeletal bases. Reverse pull head gear combined with maxillary expansion can effectively correct skeletal Class III malocclusion due to maxillary deficiency in growing patient. An eight-year-old female patient with chief complaint of prognathic mandible and anterior crossbite was successfully treated in duration of 5 months with facemask and expansion therapy based on Alternate Rapid Maxillary Expansion and Constriction (Alt-RAMEC protocol.

Efficient 3D frequency response modeling with spectral accuracy by the rapid expansion method

KAUST Repository

Chu, Chunlei

2012-07-01

Frequency responses of seismic wave propagation can be obtained either by directly solving the frequency domain wave equations or by transforming the time domain wavefields using the Fourier transform. The former approach requires solving systems of linear equations, which becomes progressively difficult to tackle for larger scale models and for higher frequency components. On the contrary, the latter approach can be efficiently implemented using explicit time integration methods in conjunction with running summations as the computation progresses. Commonly used explicit time integration methods correspond to the truncated Taylor series approximations that can cause significant errors for large time steps. The rapid expansion method (REM) uses the Chebyshev expansion and offers an optimal solution to the second-order-in-time wave equations. When applying the Fourier transform to the time domain wavefield solution computed by the REM, we can derive a frequency response modeling formula that has the same form as the original time domain REM equation but with different summation coefficients. In particular, the summation coefficients for the frequency response modeling formula corresponds to the Fourier transform of those for the time domain modeling equation. As a result, we can directly compute frequency responses from the Chebyshev expansion polynomials rather than the time domain wavefield snapshots as do other time domain frequency response modeling methods. When combined with the pseudospectral method in space, this new frequency response modeling method can produce spectrally accurate results with high efficiency. © 2012 Society of Exploration Geophysicists.

A Case of Re-Expansion Pulmonary Edema after Rapid Pleural Evacuation

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SH Shahbazi

2007-07-01

Full Text Available Introduction & Objective: Pulmonary edema after chest tube insertion is a rare complication and is associated with high mortality. The cause of this phenomenon is not clear, although causes such as decrease in surfactant and inflammatory process have been defined. Early diagnosis and treatment decrease the mortality. This study introduces a case of re-expansion pulmonary edema after rapid pleural evacuation. Case: The case is a 4.5 y/o boy, a case of Tetralogy of Fallot, who developed respiratory distress after surgery (Total Correction in ICU of Namazi Hospital in 1385. Chest X ray showed pneumothorax of left lung. For the patient, chest tube was inserted and the symptoms improved. After few hours the patient developed tachypnea, tachycardia, and CXR showed pulmonary edema of left lung. Appropriate treatment was done for the patient and his condition improved. Conclusion: Pulmonary edema after sudden evacuation of pleura is a rare phenomenon and early diagnosis decreases the mortality.

Comparative analysis in continuous expansion of bovine and human primary nucleus pulposus cells for tissue repair applications

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DH Rosenzweig

2017-03-01

Full Text Available Autologous NP cell implantation is a potential therapeutic avenue for intervertebral disc (IVD degeneration. However, monolayer expansion of cells isolated from surgical samples may negatively impact matrix production by way of dedifferentiation. Previously, we have used a continuous expansion culture system to successfully preserve a chondrocyte phenotype. In this work, we hypothesised that continuous expansion culture could also preserve nucleus pulposus (NP phenotype. We confirmed that serial passaging drove NP dedifferentiation by significantly decreasing collagen type II, aggrecan and chondroadherin (CHAD gene expression, compared to freshly isolated cells. Proliferation, gene expression profile and matrix production in both culture conditions were compared using primary bovine NP cells. Both standard culture and continuous culture produced clinically relevant cell populations. However, continuous culture cells maintained significantly higher collagen type II, aggrecan and CHAD transcript expression levels. Also, continuous expansion cells generated greater amounts of proteoglycan, collagen type II and aggrecan protein deposition in pellet cultures. To our surprise, continuous expansion of human intervertebral disc cells – isolated from acute herniation tissue – produced less collagen type II, aggrecan and CHAD genes and proteins, compared to standard culture. Also, continuous culture of cells isolated from young non-degenerate tissue did not preserve gene and protein expression, compared to standard culture. These data indicated that primary bovine and human NP cells responded differently to continuous culture, where the positive effects observed for bovine cells did not translate to human cells. Therefore, caution must be exercised when choosing animal models and cell sources for pre-clinical studies.

Anti-IL-2 treatment impairs the expansion of T(reg cell population during acute malaria and enhances the Th1 cell response at the chronic disease.

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Cláudia A Zago

Full Text Available Plasmodium chabaudi infection induces a rapid and intense splenic CD4(+ T cell response that contributes to both disease pathogenesis and the control of acute parasitemia. The subsequent development of clinical immunity to disease occurs concomitantly with the persistence of low levels of chronic parasitemia. The suppressive activity of regulatory T (T(reg cells has been implicated in both development of clinical immunity and parasite persistence. To evaluate whether IL-2 is required to induce and to sustain the suppressive activity of T(reg cells in malaria, we examined in detail the effects of anti-IL-2 treatment with JES6-1 monoclonal antibody (mAb on the splenic CD4(+ T cell response during acute and chronic P. chabaudi AS infection in C57BL/6 mice. JES6-1 treatment on days 0, 2 and 4 of infection partially inhibits the expansion of the CD4(+CD25(+Foxp3(+ cell population during acute malaria. Despite the concomitant secretion of IL-2 and expression of high affinity IL-2 receptor by large CD4(+ T cells, JES6-1 treatment does not impair effector CD4(+ T cell activation and IFN-γ production. However, at the chronic phase of the disease, an enhancement of cellular and humoral responses occurs in JES6-1-treated mice, with increased production of TNF-α and parasite-specific IgG2a antibodies. Furthermore, JES6-1 mAb completely blocked the in vitro proliferation of CD4(+ T cells from non-treated chronic mice, while it further increased the response of CD4(+ T cells from JES6-1-treated chronic mice. We conclude that JES6-1 treatment impairs the expansion of T(reg cell population during early P. chabaudi malaria and enhances the Th1 cell response in the late phase of the disease.

Model-Based Analysis of Arabidopsis Leaf Epidermal Cells Reveals Distinct Division and Expansion Patterns for Pavement and Guard Cells1[W][OA

Science.gov (United States)

Asl, Leila Kheibarshekan; Dhondt, Stijn; Boudolf, Véronique; Beemster, Gerrit T.S.; Beeckman, Tom; Inzé, Dirk; Govaerts, Willy; De Veylder, Lieven

2011-01-01

To efficiently capture sunlight for photosynthesis, leaves typically develop into a flat and thin structure. This development is driven by cell division and expansion, but the individual contribution of these processes is currently unknown, mainly because of the experimental difficulties to disentangle them in a developing organ, due to their tight interconnection. To circumvent this problem, we built a mathematic model that describes the possible division patterns and expansion rates for individual epidermal cells. This model was used to fit experimental data on cell numbers and sizes obtained over time intervals of 1 d throughout the development of the first leaf pair of Arabidopsis (Arabidopsis thaliana). The parameters were obtained by a derivative-free optimization method that minimizes the differences between the predicted and experimentally observed cell size distributions. The model allowed us to calculate probabilities for a cell to divide into guard or pavement cells, the maximum size at which it can divide, and its average cell division and expansion rates at each point during the leaf developmental process. Surprisingly, average cell cycle duration remained constant throughout leaf development, whereas no evidence for a maximum cell size threshold for cell division of pavement cells was found. Furthermore, the model predicted that neighboring cells of different sizes within the epidermis expand at distinctly different relative rates, which could be verified by direct observations. We conclude that cell division seems to occur independently from the status of cell expansion, whereas the cell cycle might act as a timer rather than as a size-regulated machinery. PMID:21693673

A fully defined and scalable 3D culture system for human pluripotent stem cell expansion and differentiation

Science.gov (United States)

Lei, Yuguo; Schaffer, David V.

2013-12-01

Human pluripotent stem cells (hPSCs), including human embryonic stem cells and induced pluripotent stem cells, are promising for numerous biomedical applications, such as cell replacement therapies, tissue and whole-organ engineering, and high-throughput pharmacology and toxicology screening. Each of these applications requires large numbers of cells of high quality; however, the scalable expansion and differentiation of hPSCs, especially for clinical utilization, remains a challenge. We report a simple, defined, efficient, scalable, and good manufacturing practice-compatible 3D culture system for hPSC expansion and differentiation. It employs a thermoresponsive hydrogel that combines easy manipulation and completely defined conditions, free of any human- or animal-derived factors, and entailing only recombinant protein factors. Under an optimized protocol, the 3D system enables long-term, serial expansion of multiple hPSCs lines with a high expansion rate (∼20-fold per 5-d passage, for a 1072-fold expansion over 280 d), yield (∼2.0 × 107 cells per mL of hydrogel), and purity (∼95% Oct4+), even with single-cell inoculation, all of which offer considerable advantages relative to current approaches. Moreover, the system enabled 3D directed differentiation of hPSCs into multiple lineages, including dopaminergic neuron progenitors with a yield of ∼8 × 107 dopaminergic progenitors per mL of hydrogel and ∼80-fold expansion by the end of a 15-d derivation. This versatile system may be useful at numerous scales, from basic biological investigation to clinical development.

Human platelet lysate in mesenchymal stromal cell expansion according to a GMP grade protocol: a cell factory experience.

Science.gov (United States)

Becherucci, Valentina; Piccini, Luisa; Casamassima, Serena; Bisin, Silvia; Gori, Valentina; Gentile, Francesca; Ceccantini, Riccardo; De Rienzo, Elena; Bindi, Barbara; Pavan, Paola; Cunial, Vanessa; Allegro, Elisa; Ermini, Stefano; Brugnolo, Francesca; Astori, Giuseppe; Bambi, Franco

2018-05-02

The use of platelet lysate (PL) for the ex-vivo expansion of mesenchymal stromal/stem cells (MSCs) was initially proposed by Doucet et al. in 2005, as an alternative to animal serum. Moreover, regulatory authorities discourage the use of fetal bovine serum (FBS) or other animal derivatives, to avoid risk of zoonoses and xenogeneic immune reactions. Even if many studies investigated PL composition, there still are some open issues related to its use in ex-vivo MSC expansion, especially according to good manufacturing practice (GMP) grade protocols. As an authorized cell factory, we report our experience using standardized PL produced by Azienda Ospedaliero Universitaria Meyer Transfusion Service for MSC expansion according to a GMP grade clinical protocol. As suggested by other authors, we performed an in-vitro test on MSCs versus MSCs cultured with FBS that still represents the best way to test PL batches. We compared 12 MSC batches cultured with DMEM 5% PL with similar batches cultured with DMEM 10% FBS, focusing on the MSC proliferation rate, MSC surface marker expression, MSC immunomodulatory and differentiation potential, and finally MSC relative telomere length. Results confirmed the literature data as PL increases cell proliferation without affecting the MSC immunophenotype, immunomodulatory potential, differentiation potential and relative telomere length. PL can be considered a safe alternative to FBS for ex-vivo expansion of MSC according to a GMP grade protocol. Our experience confirms the literature data: a large number of MSCs for clinical applications can be obtained by expansion with PL, without affecting the MSC main features. Our experience underlines the benefits of a close collaboration between the PL producers (transfusion service) and the end users (cell factory) in a synergy of skills and experiences that can lead to standardized PL production.

Ex vivo expansion of human umbilical cord blood hematopoietic stem and progenitor cells

NARCIS (Netherlands)

N. Kusadasi (Nuray)

2002-01-01

textabstractThe goal of this thesis research is to establish ex vivo expansion conditions for HSCs derived from UCB. To realize the expansion of HSCs, CD34+ or ACJ33+ UCB cells were cultured in the absence or presence of various cocktails of early acting cytokines including Flt3-L, Tpo, SCF or IL6

Incorporation of a hinge domain improves the expansion of chimeric antigen receptor T cells

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Le Qin

2017-03-01

Full Text Available Abstract Background Multiple iterations of chimeric antigen receptors (CARs have been developed, mainly focusing on intracellular signaling modules. However, the effect of non-signaling extracellular modules on the expansion and therapeutic efficacy of CARs remains largely undefined. Methods We generated two versions of CAR vectors, with or without a hinge domain, targeting CD19, mesothelin, PSCA, MUC1, and HER2, respectively. Then, we systematically compared the effect of the hinge domains on the growth kinetics, cytokine production, and cytotoxicity of CAR T cells in vitro and in vivo. Results During in vitro culture period, the percentages and absolute numbers of T cells expressing the CARs containing a hinge domain continuously increased, mainly through the promotion of CD4+ CAR T cell expansion, regardless of the single-chain variable fragment (scFv. In vitro migration assay showed that the hinges enhanced CAR T cells migratory capacity. The T cells expressing anti-CD19 CARs with or without a hinge had similar antitumor capacities in vivo, whereas the T cells expressing anti-mesothelin CARs containing a hinge domain showed enhanced antitumor activities. Conclusions Hence, our results demonstrate that a hinge contributes to CAR T cell expansion and is capable of increasing the antitumor efficacy of some specific CAR T cells. Our results suggest potential novel strategies in CAR vector design.

Expansion of Adult Human Pancreatic Tissue Yields Organoids Harboring Progenitor Cells with Endocrine Differentiation Potential

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Cindy J.M. Loomans

2018-03-01

Full Text Available Summary: Generating an unlimited source of human insulin-producing cells is a prerequisite to advance β cell replacement therapy for diabetes. Here, we describe a 3D culture system that supports the expansion of adult human pancreatic tissue and the generation of a cell subpopulation with progenitor characteristics. These cells display high aldehyde dehydrogenase activity (ALDHhi, express pancreatic progenitors markers (PDX1, PTF1A, CPA1, and MYC, and can form new organoids in contrast to ALDHlo cells. Interestingly, gene expression profiling revealed that ALDHhi cells are closer to human fetal pancreatic tissue compared with adult pancreatic tissue. Endocrine lineage markers were detected upon in vitro differentiation. Engrafted organoids differentiated toward insulin-positive (INS+ cells, and circulating human C-peptide was detected upon glucose challenge 1 month after transplantation. Engrafted ALDHhi cells formed INS+ cells. We conclude that adult human pancreatic tissue has potential for expansion into 3D structures harboring progenitor cells with endocrine differentiation potential. : In the context of β cell replacement therapy for diabetes, de Koning and colleagues describe a 3D culture platform that supports ex vivo expansion of human pancreatic tissue as organoids. These organoids harbor a subpopulation of ALDHhi cells that display proliferative capacity and can differentiate to an endocrine fate. Keywords: pancreas, organoid, human, ALDH, endocrine differentiation, beta cells, insulin, progenitor, fetal, diabetes

Ex Vivo Expansion of Functional Human UCB-HSCs/HPCs by Coculture with AFT024-hkirre Cells

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Muti ur Rehman Khan

2014-01-01

Full Text Available Kiaa1867 (human Kirre, hKirre has a critical role in brain development and/or maintenance of the glomerular slit diaphragm in kidneys. Murine homolog of this gene, mKirre expressed in OP9 and AFT024 cells could support hematopoietic stem cells/hematopoietic progenitor cells (HSC/HPC expansion in vitro. HKirre is also expressed in human FBMOB-hTERT cell line and fetal liver fibroblast-like cells but its function has remained unclear. In this paper, we cloned a hKirre gene from human fetal liver fibroblast-like cells and established a stably overexpressing hKirre-AFT024 cell line. Resultant cells could promote self-renewal and ex vivo expansion of HSCs/HPCs significantly higher than AFT024-control cells transformed with mock plasmid. The Expanded human umbilical cord blood (hUCB CD34+ cells retained the capacity of multipotent differentiation as long as 8 weeks and successfully repopulated the bone marrow of sublethally irradiated NOD/SCID mice, which demonstrated the expansion of long-term primitive transplantable HSCs/HPCs. Importantly, hkirre could upregulate the expressions of Wnt-5A, BMP4, and SDF-1 and downregulate TGF-β with other hematopoietic growth factors. By SDS-PAGE and Western Blot analysis, a ~89 kDa protein in total lysate of AFT024-hKirre was identified. Supernatants from AFT024-hkirre could also support CD34+CD38− cells expansion. These results demonstrated that the AFT024-hKirre cells have the ability to efficiently expand HSCs/HPCs.

Co-Expansion of Cytokine-Induced Killer Cells and Vγ9Vδ2 T Cells for CAR T-Cell Therapy.

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Shou-Hui Du

Full Text Available Gamma delta (γδ T cells and cytokine-induced killer (CIK cells, which are a heterogeneous population of T lymphocytes and natural killer T (NKT cells, have been separately expanded ex vivo and shown to be capable of targeting and mediating cytotoxicity against various tumor cells in a major histocompatibility complex-unrestricted manner. However, the co-expansion and co-administration of these immune cells have not been explored. In this study we describe an efficient method to expand simultaneously both CIK and Vγ9Vδ2 T cells, termed as CIKZ cells, from human peripheral blood mononuclear cells (PBMCs using Zometa, interferon-gamma (IFN-γ, interleukin 2 (IL-2, anti-CD3 antibody and engineered K562 feeder cells expressing CD64, CD137L and CD86. A 21-day culture of PBMCs with this method yielded nearly 20,000-fold expansion of CIKZ cells with γδ T cells making up over 20% of the expanded population. The expanded CIKZ cells exhibited antitumor cytotoxicity and could be modified to express anti-CD19 chimeric antigen receptor (CAR, anti-CEA CAR, and anti-HER2 CAR to enhance their specificity and cytotoxicity against CD19-, CEA-, or HER2-positive tumor cells. The tumor inhibitory activity of anti-CD19 CAR-modified CIKZ cells was further demonstrated in vivo in a Raji tumor mouse model. The findings herein substantiate the feasibility of co-expanding CIK and γδ cells for adoptive cellular immunotherapy applications such as CAR T-cell therapy against cancer.

Chromosomal aberrations and deoxyribonucleic acid single-strand breaks in adipose-derived stem cells during long-term expansion in vitro.

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Froelich, Katrin; Mickler, Johannes; Steusloff, Gudrun; Technau, Antje; Ramos Tirado, Mario; Scherzed, Agmal; Hackenberg, Stephan; Radeloff, Andreas; Hagen, Rudolf; Kleinsasser, Norbert

2013-07-01

Adipose-derived stem cells (ASCs) are a promising mesenchymal cell source for tissue engineering approaches. To obtain an adequate cell amount, in vitro expansion of the cells may be required in some cases. To monitor potential contraindications for therapeutic applications in humans, DNA strand breaks and chromosomal aberrations in ASCs during in vitro expansion were examined. After isolation of ASC from human lipoaspirates of seven patients, in vitro expansion over 10 passages was performed. Cells from passages 1, 2, 3, 5 and 10 were used for the alkaline single-cell microgel electrophoresis (comet) assay to detect DNA single-strand breaks and alkali labile as well as incomplete excision repair sites. Chromosomal changes were examined by means of the chromosomal aberration test. During in vitro expansion, ASC showed no DNA single-strand breaks in the comet assay. With the chromosomal aberration test, however, a significant increase in chromosomal aberrations were detected. The study showed that although no DNA fragmentation could be determined, the safety of ASC cannot be ensured with respect to chromosome stability during in vitro expansion. Thus, reliable analyses for detecting ASC populations, which accumulate chromosomal aberrations or even undergo malignant transformation during extensive in vitro expansion, must be implemented as part of the safety evaluation of these cells for stem cell-based therapy. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Thin-plate spline analysis of the short- and long-term effects of rapid maxillary expansion.

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Franchi, Lorenzo; Baccetti, Tiziano; Cameron, Christopher G; Kutcipal, Elizabeth A; McNamara, James A

2002-04-01

The aim of this study was to investigate the short- and long-term effects induced by rapid maxillary expansion (RME) on the shape of the maxillary and circummaxillary structures by means of thin-plate spline (TPS) analysis. The sample consisted of 42 patients who were compared with a control sample of 20 subjects. The treated subjects underwent Haas-type RME, followed by fixed appliance therapy. Postero-anterior (PA) cephalograms were analysed for each treated subject at T1 (pre-treatment), T2 (immediate post-expansion), and T3 (long-term observation), and were available at T1 and T3 for the control group (CG). The mean age at T1 was 11 years and 10 months for both groups. The mean chronological ages at T3 were 20 years, 6 months for the treated group (TG) and 17 years, 8 months for the control group. The study focused on shape changes in the maxillary, nasal, zygomatic, and orbital regions. TPS analysis revealed significant shape changes in the TG. They consisted of an upward and lateral displacement of the two halves of the naso-maxillary complex as a result of active expansion in the short-term, and normalization of maxillary shape in the transverse dimension in the long-term (the initial transverse deficiency of the maxilla in the treated group was eliminated by RME therapy both in the short- and long-term). At the end of the observation period, the nasal cavities were larger when compared with both their pre-expansion configuration and the final configuration in the controls. RME with the Haas appliance appears to be an efficient therapeutic means to induce permanent favourable changes in the shape of the naso-maxillary complex.

Miniscrew-assisted rapid palatal expansion for managing arch perimeter in an adult patient

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Amanda Carneiro da Cunha

Full Text Available ABSTRACT Introduction: Etiology of dental crowding may be related to arch constriction in diverse dimensions, and an appropriate manipulation of arch perimeter by intervening in basal bone discrepancies cases, may be a key for crowding relief, especially when incisors movement is limited due to underlying pathology, periodontal issues or restrictions related to soft tissue profile. Objectives: This case report illustrates a 24-year old woman, with maxillary transverse deficiency, upper and lower arches crowding, Class II, division 1, subdivision right relationship, previous upper incisors traumatic episode and straight profile. A non-surgical and non-extraction treatment approach was feasible due to the miniscrew-assisted rapid palatal expansion technique (MARPE. Methods: The MARPE appliance consisted of a conventional Hyrax expander supported by four orthodontic miniscrews. A slow expansion protocol was adopted, with an overall of 40 days of activation and a 3-month retention period. Intrusive traction miniscrew-anchored mechanics were used for correcting the Class II subdivision relationship, managing lower arch perimeter and midline deviation before including the upper central incisors. Results: Post-treatment records show an intermolar width increase of 5 mm, bilateral Class I molar and canine relationships, upper and lower crowding resolution, coincident dental midlines and proper intercuspation. Conclusions: The MARPE is an effective treatment approach for managing arch-perimeter deficiencies related to maxillary transverse discrepancies in adult patients.

Evaluation of umbilical cord blood CD34+ hematopoietic stem cells expansion with inhibition of TGF-β receptorII in co-culture with bone marrow mesenchymal stromal cells.

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Sohrabi Akhkand, Saman; Amirizadeh, Naser; Nikougoftar, Mahin; Alizadeh, Javad; Zaker, Farhad; Sarveazad, Arash; Joghataei, Mohammad Taghi; Faramarzi, Mahmood

2016-08-01

Umbilical cord blood (UCB) is an important source of hematopoietic stem cells (HSCs). However, low number of HSCs in UCB has been an obstacle for adult hematopoietic stem cell transplantation. The expansion of HSCs in culture is one approach to overcome this problem. In this study, we investigated the expansion of UCB-HSCs by using human bone marrow mesenchymal stromal cells (MSCs) as feeder layer as well as inhibiting the TGF-β signaling pathway through reduction of TGF-βRII expression. CD34(+) cells were isolated from UCB and transfected by SiRNA targeting TGF-βRII mRNA. CD34(+) cells were expanded in four culture media with different conditions, including 1) expansion of CD34(+) cells in serum free medium containing growth factors, 2) expansion of cells transfected with SiRNA targeting TGF-βRII in medium containing growth factors, 3) expansion of cells in presence of growth factors and MSCs, 4) expansion of cells transfected with SiRNA targeting TGF-βRII on MSCs feeder layer in medium containing growth factors. These culture conditions were evaluated for the number of total nucleated cells (TNCs), CD34 surface marker as well as using CFU assay on 8th day after culture. The fold increase in CD34(+) cells, TNCs, and colony numbers (71.8±6.9, 93.2±10.2 and 128±10, respectively) was observed to be highest in fourth culture medium compared to other culture conditions. The difference between number of cells in four culture media in 8th day compared to unexpanded cells (0day) before expansion was statistically significant (P<0.05). The results showed that transfection of CD34(+) cells with SiRNA targeting TGF-βRII and their co-culture with MSCs could considerably increase the number of progenitors. Therefore, this method could be useful for UCB-HSCs expansion. Copyright © 2016 Elsevier Ltd. All rights reserved.

Quantifying the mechanical micro-environment during three-dimensional cell expansion on microbeads by means of individual cell-based modelling.

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Smeets, Bart; Odenthal, Tim; Tijskens, Engelbert; Ramon, Herman; Van Oosterwyck, Hans

2013-10-01

Controlled in vitro three-dimensional cell expansion requires culture conditions that optimise the biophysical micro-environment of the cells during proliferation. In this study, we propose an individual cell-based modelling platform for simulating the mechanics of cell expansion on microcarriers. The lattice-free, particle-based method considers cells as individual interacting particles that deform and move over time. The model quantifies how the mechanical micro-environment of individual cells changes during the time of confluency. A sensitivity analysis is performed, which shows that changes in the cell-specific properties of cell-cell adhesion and cell stiffness cause the strongest change in the mechanical micro-environment of the cells. Furthermore, the influence of the mechanical properties of cells and microbead is characterised. The mechanical micro-environment is strongly influenced by the adhesive properties and the size of the microbead. Simulations show that even in the absence of strong biological heterogeneity, a large heterogeneity in mechanical stresses can be expected purely due to geometric properties of the culture system.

Spatial Linkage and Urban Expansion: AN Urban Agglomeration View

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Jiao, L. M.; Tang, X.; Liu, X. P.

2017-09-01

Urban expansion displays different characteristics in each period. From the perspective of the urban agglomeration, studying the spatial and temporal characteristics of urban expansion plays an important role in understanding the complex relationship between urban expansion and network structure of urban agglomeration. We analyze urban expansion in the Yangtze River Delta Urban Agglomeration (YRD) through accessibility to and spatial interaction intensity from core cities as well as accessibility of road network. Results show that: (1) Correlation between urban expansion intensity and spatial indicators such as location and space syntax variables is remarkable and positive, while it decreases after rapid expansion. (2) Urban expansion velocity displays a positive correlation with spatial indicators mentioned above in the first (1980-1990) and second (1990-2000) period. However, it exhibits a negative relationship in the third period (2000-2010), i.e., cities located in the periphery of urban agglomeration developing more quickly. Consequently, the hypothesis of convergence of urban expansion in rapid expansion stage is put forward. (3) Results of Zipf's law and Gibrat's law show urban expansion in YRD displays a convergent trend in rapid expansion stage, small and medium-sized cities growing faster. This study shows that spatial linkage plays an important but evolving role in urban expansion within the urban agglomeration. In addition, it serves as a reference to the planning of Yangtze River Delta Urban Agglomeration and regulation of urban expansion of other urban agglomerations.

Expansion and conversion of human pancreatic ductal cells into insulin-secreting endocrine cells.

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Lee, Jonghyeob; Sugiyama, Takuya; Liu, Yinghua; Wang, Jing; Gu, Xueying; Lei, Ji; Markmann, James F; Miyazaki, Satsuki; Miyazaki, Jun-Ichi; Szot, Gregory L; Bottino, Rita; Kim, Seung K

2013-11-19

Pancreatic islet β-cell insufficiency underlies pathogenesis of diabetes mellitus; thus, functional β-cell replacement from renewable sources is the focus of intensive worldwide effort. However, in vitro production of progeny that secrete insulin in response to physiological cues from primary human cells has proven elusive. Here we describe fractionation, expansion and conversion of primary adult human pancreatic ductal cells into progeny resembling native β-cells. FACS-sorted adult human ductal cells clonally expanded as spheres in culture, while retaining ductal characteristics. Expression of the cardinal islet developmental regulators Neurog3, MafA, Pdx1 and Pax6 converted exocrine duct cells into endocrine progeny with hallmark β-cell properties, including the ability to synthesize, process and store insulin, and secrete it in response to glucose or other depolarizing stimuli. These studies provide evidence that genetic reprogramming of expandable human pancreatic cells with defined factors may serve as a general strategy for islet replacement in diabetes. DOI: http://dx.doi.org/10.7554/eLife.00940.001.

IBO Case Report: Management of Skeletal Class III Malocclusion with Combined Rapid Maxillary Expansion: Facemask Therapy and 5-Year Follow-up

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Ashok Surana

2012-01-01

Full Text Available This case report describes the management of skeletal Class III malocclusion with maxillary deficiency in an adolescent girl, using combined rapid maxillary expansion-facemask approach, followed by comprehensive fixed appliance mechanotherapy. Excellent long-term stability is demonstrated up to 5 years post-treatment.

A chemically defined substrate for the expansion and neuronal differentiation of human pluripotent stem cell-derived neural progenitor cells.

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Tsai, Yihuan; Cutts, Josh; Kimura, Azuma; Varun, Divya; Brafman, David A

2015-07-01

Due to the limitation of current pharmacological therapeutic strategies, stem cell therapies have emerged as a viable option for treating many incurable neurological disorders. Specifically, human pluripotent stem cell (hPSC)-derived neural progenitor cells (hNPCs), a multipotent cell population that is capable of near indefinite expansion and subsequent differentiation into the various cell types that comprise the central nervous system (CNS), could provide an unlimited source of cells for such cell-based therapies. However the clinical application of these cells will require (i) defined, xeno-free conditions for their expansion and neuronal differentiation and (ii) scalable culture systems that enable their expansion and neuronal differentiation in numbers sufficient for regenerative medicine and drug screening purposes. Current extracellular matrix protein (ECMP)-based substrates for the culture of hNPCs are expensive, difficult to isolate, subject to batch-to-batch variations, and, therefore, unsuitable for clinical application of hNPCs. Using a high-throughput array-based screening approach, we identified a synthetic polymer, poly(4-vinyl phenol) (P4VP), that supported the long-term proliferation and self-renewal of hNPCs. The hNPCs cultured on P4VP maintained their characteristic morphology, expressed high levels of markers of multipotency, and retained their ability to differentiate into neurons. Such chemically defined substrates will eliminate critical roadblocks for the utilization of hNPCs for human neural regenerative repair, disease modeling, and drug discovery. Copyright © 2015. Published by Elsevier B.V.

A chemically defined substrate for the expansion and neuronal differentiation of human pluripotent stem cell-derived neural progenitor cells

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Yihuan Tsai

2015-07-01

Full Text Available Due to the limitation of current pharmacological therapeutic strategies, stem cell therapies have emerged as a viable option for treating many incurable neurological disorders. Specifically, human pluripotent stem cell (hPSC-derived neural progenitor cells (hNPCs, a multipotent cell population that is capable of near indefinite expansion and subsequent differentiation into the various cell types that comprise the central nervous system (CNS, could provide an unlimited source of cells for such cell-based therapies. However the clinical application of these cells will require (i defined, xeno-free conditions for their expansion and neuronal differentiation and (ii scalable culture systems that enable their expansion and neuronal differentiation in numbers sufficient for regenerative medicine and drug screening purposes. Current extracellular matrix protein (ECMP-based substrates for the culture of hNPCs are expensive, difficult to isolate, subject to batch-to-batch variations, and, therefore, unsuitable for clinical application of hNPCs. Using a high-throughput array-based screening approach, we identified a synthetic polymer, poly(4-vinyl phenol (P4VP, that supported the long-term proliferation and self-renewal of hNPCs. The hNPCs cultured on P4VP maintained their characteristic morphology, expressed high levels of markers of multipotency, and retained their ability to differentiate into neurons. Such chemically defined substrates will eliminate critical roadblocks for the utilization of hNPCs for human neural regenerative repair, disease modeling, and drug discovery.

Three dimensional evaluation of alveolar bone changes in response to different rapid palatal expansion activation rates

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Brian LaBlonde

Full Text Available ABSTRACT Introduction: The aim of this multi-center retrospective study was to quantify the changes in alveolar bone height and thickness after using two different rapid palatal expansion (RPE activation protocols, and to determine whether a more rapid rate of expansion is likely to cause more adverse effects, such as alveolar tipping, dental tipping, fenestration and dehiscence of anchorage teeth. Methods: The sample consisted of pre- and post-expansion records from 40 subjects (age 8-15 years who underwent RPE using a 4-banded Hyrax appliance as part of their orthodontic treatment to correct posterior buccal crossbites. Subjects were divided into two groups according to their RPE activation rates (0.5 mm/day and 0.8 mm/day; n = 20 each group. Three-dimensional images for all included subjects were evaluated using Dolphin Imaging Software 11.7 Premium. Maxillary base width, buccal and palatal cortical bone thickness, alveolar bone height, and root angulation and length were measured. Significance of the changes in the measurements was evaluated using Wilcoxon signed-rank test and comparisons between groups were done using ANOVA. Significance was defined at p ≤ 0.05. Results: RPE activation rates of 0.5 mm per day (Group 1 and 0.8 mm per day (Group 2 caused significant increase in arch width following treatment; however, Group 2 showed greater increases compared to Group 1 (p < 0.01. Buccal alveolar height and width decreased significantly in both groups. Both treatment protocols resulted in significant increases in buccal-lingual angulation of teeth; however, Group 2 showed greater increases compared to Group 1 (p < 0.01. Conclusion: Both activation rates are associated with significant increase in intra-arch widths. However, 0.8 mm/day resulted in greater increases. The 0.8 mm/day activation rate also resulted in more increased dental tipping and decreased buccal alveolar bone thickness over 0.5 mm/day.

Three-dimensional evaluation of soft tissue changes in the orofacial region after tooth-borne and bone-borne surgically assisted rapid maxillary expansion

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Nada, R.M.; Loon, B. van; Maal, T.J.J.; Berge, S.J.; Mostafa, Y.A.; Kuijpers-Jagtman, A.M.; Schols, J.G.J.H.

2013-01-01

OBJECTIVES: This study seeks to three-dimensionally assess soft tissue changes in the orofacial region following tooth-borne and bone-borne surgically assisted rapid maxillary expansion (SARME). MATERIALS AND METHODS: This prospective cohort study included 40 skeletally mature patients with

The Important Role of FLT3-L in Ex Vivo Expansion of Hematopoietic Stem Cells following Co-Culture with Mesenchymal Stem Cells.

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Oubari, Farhad; Amirizade, Naser; Mohammadpour, Hemn; Nakhlestani, Mozhdeh; Zarif, Mahin Nikougoftar

2015-01-01

Hematopoietic stem cells (HSCs) transplantation using umbilical cord blood (UCB) has improved during the last decade. Because of cell limitations, several studies focused on the ex vivo expansion of HSCs. Numerous investigations were performed to introduce the best cytokine cocktails for HSC expansion The majority used the Fms-related tyrosine kinase 3 ligand (FLT3-L) as a critical component. According to FLT3-L biology, in this study we have investigated the hypothesis that FLT3-L only effectively induces HSCs expansion in the presence of a mesenchymal stem cell (MSC) feeder. In this experimental study, HSCs and MSCs were isolated from UCB and placenta, respectively. HSCs were cultured in different culture conditions in the presence and absence of MSC feeder and cytokines. After ten days of culture, total nucleated cell count (TNC), cluster of differentiation 34+(CD34(+)) cell count, colony forming unit assay (CFU), long-term culture initiating cell (LTC-IC), homeobox protein B4 (HoxB4) mRNA and surface CD49d expression were evaluated. The fold increase for some culture conditions was compared by the t test. HSCs expanded in the presence of cytokines and MSCs feeder. The rate of expansion in the co-culture condition was two-fold more than culture with cytokines (Pculture condition at a level of 20-fold equal to the presence of stem cell factor (SCF), thrombopoietin (TPO) and FLT3-L without feeder cells. The number of extracted colonies from LTC-IC and CD49d expression compared with a cytokine cocktail condition meaningfully increased (Pculture with MSCs can induce high yield expansion of HSCs and be a substitute for the universal cocktail of SCF, TPO and FLT3-L in feeder-free culture.

Pathogen-free, plasma-poor platelet lysate and expansion of human mesenchymal stem cells.

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Iudicone, Paola; Fioravanti, Daniela; Bonanno, Giuseppina; Miceli, Michelina; Lavorino, Claudio; Totta, Pierangela; Frati, Luigi; Nuti, Marianna; Pierelli, Luca

2014-01-27

Supplements to support clinical-grade cultures of mesenchymal stem cells (MSC) are required to promote growth and expansion of these cells. Platelet lysate (PL) is a human blood component which may replace animal serum in MSC cultures being rich in various growth factors. Here, we describe a plasma poor pathogen-free platelet lysate obtained by pooling 12 platelet (PLT) units, to produce a standardized and safe supplement for clinical-grade expansion of MSC. PL lots were obtained by combining 2 6-unit PLT pools in additive solution (AS) following a transfusional-based procedure including pathogen inactivation (PI) by Intercept technology and 3 cycles of freezing/thawing, followed by membrane removal. Three PI-PL and 3 control PL lots were produced to compare their ability to sustain bone marrow derived MSC selection and expansion. Moreover, two further PL, subjected to PI or not, were also produced starting from the same initial PLT pools to evaluate the impact of PI on growth factor concentration and capacity to sustain cell growth. Additional PI-PL lots were used for comparison with fetal bovine serum (FBS) on MSC expansion. Immunoregulatory properties of PI-PL-generated MSC were documented in vitro by mixed lymphocyte culture (MLC) and peripheral blood mononuclear cells (PBMC) mitogen induced proliferation. PI-PL and PL control lots had similar concentrations of 4 well-described growth factors endowed with MSC stimulating ability. Initial growth and MSC expansion by PI-PL and PL controls were comparable either using different MSC populations or in head to head experiments. Moreover, PI-PL and PL control sustained similar MSC growth of frozen/thawed MSC. Multilineage differentiation of PI-derived and PI-PL-derived MSC were maintained in any MSC cultures as well as their immunoregulatory properties. Finally, no direct impact of PI on growth factor concentration and MSC growth support was observed, whereas the capacity of FBS to sustain MSC expansion in basic

Expansion and differentiation of germline-derived pluripotent stem cells on biomaterials.

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Hoss, Mareike; Šarić, Tomo; Denecke, Bernd; Peinkofer, Gabriel; Bovi, Manfred; Groll, Jürgen; Ko, Kinarm; Salber, Jochen; Halbach, Marcel; Schöler, Hans R; Zenke, Martin; Neuss, Sabine

2013-05-01

Stem cells with broad differentiation potential, such as the recently described germline-derived pluripotent stem cells (gPS cells), are an appealing source for tissue engineering strategies. Biomaterials can inhibit, support, or induce proliferation and differentiation of stem cells. Here we identified (1) polymers that maintain self-renewal and differentiation potential of gPS cells for feeder-free expansion and (2) polymers supporting the cardiomyogenic fate of gPS cells by analyzing a panel of polymers of an established biomaterial bank previously used to assess growth of diverse stem cell types. Identification of cytocompatible gPS cell/biomaterial combinations required analysis of several parameters, including morphology, viability, cytotoxicity, apoptosis, proliferation, and differentiation potential. Pluripotency of gPS cells was visualized by the endogenous Oct4-promoter-driven GFP and by Sox2 and Nanog immunofluorescence. Viability assay, proliferation assay, and flow cytometry showed that gPS cells efficiently adhere and are viable on synthetic polymers, such as Resomer(®) LR704 (poly(L-lactic-D,L-lactic acid), poly(tetrafluor ethylene) (PTFE), poly(vinylidene fluoride) (PVDF), and on gelatine-coated tissue culture polystyrene. Expansion experiments showed that Resomer LR704 is an alternative substrate for feeder-free gPS cell maintenance. Resomer LR704, PTFE, and PVDF were found to be suitable for gPS cell differentiation. Spontaneous beating in embryoid bodies cultured on Resomer LR704 occurred already on day 8 of differentiation, much earlier compared to the other surfaces. This indicates that Resomer LR704 supports spontaneous cardiomyogenic differentiation of gPS cells, which was also confirmed on molecular, protein and functional level.

Microencapsulation Technology: A Powerful Tool for Integrating Expansion and Cryopreservation of Human Embryonic Stem Cells

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Malpique, Rita; Brito, Catarina; Jensen, Janne; Bjorquist, Petter; Carrondo, Manuel J. T.; Alves, Paula M.

2011-01-01

The successful implementation of human embryonic stem cells (hESCs)-based technologies requires the production of relevant numbers of well-characterized cells and their efficient long-term storage. In this study, cells were microencapsulated in alginate to develop an integrated bioprocess for expansion and cryopreservation of pluripotent hESCs. Different three-dimensional (3D) culture strategies were evaluated and compared, specifically, microencapsulation of hESCs as: i) single cells, ii) aggregates and iii) immobilized on microcarriers. In order to establish a scalable bioprocess, hESC-microcapsules were cultured in stirred tank bioreactors. The combination of microencapsulation and microcarrier technology resulted in a highly efficient protocol for the production and storage of pluripotent hESCs. This strategy ensured high expansion ratios (an approximately twenty-fold increase in cell concentration) and high cell recovery yields (>70%) after cryopreservation. When compared with non-encapsulated cells, cell survival post-thawing demonstrated a three-fold improvement without compromising hESC characteristics. Microencapsulation also improved the culture of hESC aggregates by protecting cells from hydrodynamic shear stress, controlling aggregate size and maintaining cell pluripotency for two weeks. This work establishes that microencapsulation technology may prove a powerful tool for integrating the expansion and cryopreservation of pluripotent hESCs. The 3D culture strategy developed herein represents a significant breakthrough towards the implementation of hESCs in clinical and industrial applications. PMID:21850261

Expansion of donor-reactive host T cells in primary graft failure after allogeneic hematopoietic SCT following reduced-intensity conditioning.

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Koyama, M; Hashimoto, D; Nagafuji, K; Eto, T; Ohno, Y; Aoyama, K; Iwasaki, H; Miyamoto, T; Hill, G R; Akashi, K; Teshima, T

2014-01-01

Graft rejection remains a major obstacle in allogeneic hematopoietic SCT following reduced-intensity conditioning (RIC-SCT), particularly after cord blood transplantation (CBT). In a murine MHC-mismatched model of RIC-SCT, primary graft rejection was associated with activation and expansion of donor-reactive host T cells in peripheral blood and BM early after SCT. Donor-derived dendritic cells are at least partly involved in host T-cell activation. We then evaluated if such an expansion of host T cells could be associated with graft rejection after RIC-CBT. Expansion of residual host lymphocytes was observed in 4/7 patients with graft rejection at 3 weeks after CBT, but in none of the 17 patients who achieved engraftment. These results suggest the crucial role of residual host T cells after RIC-SCT in graft rejection and expansion of host T cells could be a marker of graft rejection. Development of more efficient T cell-suppressive conditioning regimens may be necessary in the context of RIC-SCT.

Expression of a constitutively activated plasma membrane H+-ATPase in Nicotiana tabacum BY-2 cells results in cell expansion.

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Niczyj, Marta; Champagne, Antoine; Alam, Iftekhar; Nader, Joseph; Boutry, Marc

2016-11-01

Increased acidification of the external medium by an activated H + -ATPase results in cell expansion, in the absence of upstream activating signaling. The plasma membrane H + -ATPase couples ATP hydrolysis with proton transport outside the cell, and thus creates an electrochemical gradient, which energizes secondary transporters. According to the acid growth theory, this enzyme is also proposed to play a major role in cell expansion, by acidifying the external medium and so activating enzymes that are involved in cell wall-loosening. However, this theory is still debated. To challenge it, we made use of a plasma membrane H + -ATPase isoform from Nicotiana plumbaginifolia truncated from its C-terminal auto-inhibitory domain (ΔCPMA4), and thus constitutively activated. This protein was expressed in Nicotiana tabacum BY-2 suspension cells using a heat shock inducible promoter. The characterization of several independent transgenic lines showed that the expression of activated ΔCPMA4 resulted in a reduced external pH by 0.3-1.2 units, as well as in an increased H + -ATPase activity by 77-155 % (ATP hydrolysis), or 70-306 % (proton pumping) of isolated plasma membranes. In addition, ΔCPMA4-expressing cells were 17-57 % larger than the wild-type cells and displayed abnormal shapes. A proteomic comparison of plasma membranes isolated from ΔCPMA4-expressing and wild-type cells revealed the altered abundance of several proteins involved in cell wall synthesis, transport, and signal transduction. In conclusion, the data obtained in this work showed that H + -ATPase activation is sufficient to induce cell expansion and identified possible actors which intervene in this process.

ERK Regulates Renal Cell Proliferation and Renal Cyst Expansion in inv Mutant Mice

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Okumura, Yasuko; Sugiyama, Noriyuki; Tanimura, Susumu; Nishida, Masashi; Hamaoka, Kenji; Kohno, Michiaki; Yokoyama, Takahiko

2009-01-01

Nephronophthisis (NPHP) is the most frequent genetic cause of end-stage kidney disease in children and young adults. Inv mice are a model for human nephronophthisis type 2 (NPHP2) and characterized by multiple renal cysts and situs inversus. Renal epithelial cells in inv cystic kidneys show increased cell proliferation. We studied the ERK pathway to understand the mechanisms that induce cell proliferation and renal cyst progression in inv kidneys. We studied the effects of ERK suppression by administering PD184352, an oral mitogen-activated protein kinase kinase (MEK) inhibitor on renal cyst expansion, extracellular signal-regulated protein kinase (ERK) activity, bromo-deoxyuridine (BrdU) incorporation and expression of cell-cycle regulators in invΔC kidneys. Phosphorylated ERK (p-ERK) level increased along with renal cyst enlargement. Cell-cycle regulators showed a high level of expression in invΔC kidneys. PD184352 successfully decreased p-ERK level and inhibited renal cyst enlargement. The inhibitor also decreased expression of cell-cycle regulators and BrdU incorporation in renal epithelial cells. The present results showed that ERK regulated renal cell proliferation and cyst expansion in inv mutants

Periodontal and dental effects of surgically assisted rapid maxillary expansion, assessed by using digital study models

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Siqueira, Danilo Furquim; Cardoso, Mauricio de Almeida; Capelozza, Leopoldino; Goldenberg, Dov Charles; Fernandes, Mariana dos Santos

2015-01-01

OBJECTIVE: The present study assessed the maxillary dental arch changes produced by surgically assisted rapid maxillary expansion (SARME). METHODS: Dental casts from 18 patients (mean age of 23.3 years) were obtained at treatment onset (T1), three months after SARME (T2) and 6 months after expansion (T3). The casts were scanned in a 3D scanner (D-250, 3Shape, Copenhagen, Denmark). Maxillary dental arch width, dental crown tipping and height were measured and assessed by ANOVA and Tukey's test. RESULTS: Increased transversal widths from T1 and T2 and the maintenance of these values from T2 and T3 were observed. Buccal teeth tipping also showed statistically significant differences, with an increase in all teeth from T1 to T2 and a decrease from T2 to T3. No statistically significant difference was found for dental crown height, except for left first and second molars, although clinically irrelevant. CONCLUSION: SARME proved to be an effective and stable procedure, with minimum periodontal hazards. PMID:26154457

Dental arch changes associated with rapid maxillary expansion: A retrospective model analysis study

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Ivor M D′Souza

2015-01-01

Full Text Available Introduction: Transverse deficiency of the maxilla is a common clinical problem in orthodontics and dentofacial orthopedics. Transverse maxillary deficiency, isolated or associated with other dentofacial deformities, results in esthetic and functional impairment giving rise to several clinical manifestations such as asymmetrical facial growth, positional and functional mandibular deviations, altered dentofacial esthetics, adverse periodontal responses, unstable dental tipping, and other functional problems. Orthopedic maxillary expansion is the preferred treatment approach to increase the maxillary transverse dimension in young patients by splitting of the mid palatal suture. This orthopedic procedure has lately been subject of renewed interest in orthodontic treatment mechanics because of its potential for increasing arch perimeter to alleviate crowding in the maxillary arch without adversely affecting facial profile. Hence, the present investigation was conducted to establish a correlation between transverse expansion and changes in the arch perimeter, arch width and arch length. Methods: For this purpose, 10 subjects (five males, five females were selected who had been treated by rapid maxillary expansion (RME using hyrax rapid palatal expander followed by fixed mechanotherapy (PEA. Pretreatment (T1, postexpansion (T2, and posttreatment (T3 dental models were compared for dental changes brought about by RME treatment and its stability at the end of fixed mechanotherapy. After model measurements were made, the changes between T1-T2, T2-T3 and T1-T3 were determined for each patient. The mean difference between T1-T2, T2-T3 and T1-T3 were compared to assess the effects of RME on dental arch measurements. Results are expressed as mean ± standard deviation and are compared by repeated measures analysis of variance followed by a post-hoc test. Arch perimeter changes are correlated with changes in arch widths at the canine, premolar and molar

Progenitor cells in liver regeneration: molecular responses controlling their activation and expansion

DEFF Research Database (Denmark)

Santoni-Rugiu, Eric; Jelnes, Peter; Thorgeirsson, Snorri S

2005-01-01

created in the liver by a certain insult. This review will focus on molecular responses controlling activation and expansion of the hepatic progenitor cell niche, emphasizing similarities and differences in the microenvironments orchestrating regeneration by recruitment of progenitor cell populations...... cells, and recruited inflammatory cells as well as the variety of growth-modulating molecules produced and/or harboured by these elements. The cellular and molecular responses to different regenerative stimuli seem to depend on the injury inflicted and consequently on the molecular microenvironment...

Whole transcriptome analysis for T cell receptor-affinity and IRF4-regulated clonal expansion of T cells

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Wei Shi

2014-12-01

Full Text Available Clonal population expansion of T cells during an immune response is dependent on the affinity of the T cell receptor (TCR for its antigen [1]. However, there is little understanding of how this process is controlled transcriptionally. We found that the transcription factor IRF4 was induced in a manner dependent on TCR-affinity and was critical for the clonal expansion and maintenance of effector function of antigen-specific CD8+ T cells. We performed a genome-wide expression profiling experiment using RNA sequencing technology (RNA-seq to interrogate global expression changes when IRF4 was deleted in CD8+ T cells activated with either a low or high affinity peptide ligand. This allowed us not only to determine IRF4-dependent transcriptional changes but also to identify transcripts dependent on TCR-affinity [2]. Here we describe in detail the analyses of the RNA-seq data, including quality control, read mapping, quantification, normalization and assessment of differential gene expression. The RNA-seq data can be accessed from Gene Expression Omnibus database (accession number GSE49929.

LED phototherapy on midpalatal suture after rapid maxilla expansion: a Raman spectroscopic study

Science.gov (United States)

Rosa, Cristiane B.; Habib, Fernando Antonio L.; de Araújo, Telma M.; dos Santos, Jean N.; Cangussu, Maria Cristina T.; Barbosa, Artur Felipe S.; de Castro, Isabele Cardoso V.; Soares, Luiz Guilherme P.; Pinheiro, Antonio L. B.

2015-03-01

A quick bone formation after maxillary expansion would reduce treatment timeand the biomodulating effects of LED light could contribute for it. The aim of this study was to analyze the effect of LED phototherapy on the acceleration of bone formation at the midpalatal suture after maxilla expansion. Thirty rats divided into 6 groups were used on the study at 2 time points - 7 days: Control; Expansion; and Expansion + LED; and 14 days: Expansion; Expansion + LED in the first week; Expansion and LED in the first and second weeks. LED irradiation occurred at every 48 h during 2 weeks. Expansion was accomplished using a spatula and maintained with a triple helicoid of 0.020" stainless steel orthodontic wire. A LED light (λ850 ± 10nm, 150mW ± 10mW, spot of 0.5cm2, t=120 sec, SAEF of 18J/cm2) was applied in one point in the midpalatal suture immediately behind the upper incisors. Near infrared Raman spectroscopic analysis of the suture region was carried and data submitted to statistical analyzes (p≤0.05). Raman spectrum analysis demonstrated that irradiation increased hydroxyapatite in the midpalatal suture after expansion. The results of this indicate that LED irradiation; have a positive biomodulation contributing to the acceleration of bone formation in the midpalatal suture after expansion procedure.

Decellularized extracellular matrices produced from immortal cell lines derived from different parts of the placenta support primary mesenchymal stem cell expansion.

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Gina D Kusuma

Full Text Available Mesenchymal stem/stromal cells (MSCs exhibit undesired phenotypic changes during ex vivo expansion, limiting production of the large quantities of high quality primary MSCs needed for both basic research and cell therapies. Primary MSCs retain many desired MSC properties including proliferative capacity and differentiation potential when expanded on decellularized extracellular matrix (dECM prepared from primary MSCs. However, the need to use low passage number primary MSCs (passage 3 or lower to produce the dECM drastically limits the utility and impact of this technology. Here, we report that primary MSCs expanded on dECM prepared from high passage number (passage 25 human telomerase reverse transcriptase (hTERT transduced immortal MSC cell lines also exhibit increased proliferation and osteogenic differentiation. Two hTERT-transduced placenta-derived MSC cell lines, CMSC29 and DMSC23 [derived from placental chorionic villi (CMSCs and decidua basalis (DMSCs, respectively], were used to prepare dECM-coated substrates. These dECM substrates showed structural and biochemical differences. Primary DMSCs cultured on dECM-DMSC23 showed a three-fold increase in cell number after 14 days expansion in culture and increased osteogenic differentiation compared with controls. Primary CMSCs cultured on the dECM-DMSC23 exhibited a two-fold increase in cell number and increased osteogenic differentiation. We conclude that immortal MSC cell lines derived from different parts of the placenta produce dECM with varying abilities for supporting increased primary MSC expansion while maintaining important primary MSC properties. Additionally, this is the first demonstration of using high passage number cells to produce dECM that can promote primary MSC expansion, and this advancement greatly increases the feasibility and applicability of dECM-based technologies.

Expansion of CD16-Negative Natural Killer Cells in the Peripheral Blood of Patients with Metastatic Melanoma

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Shernan G. Holtan

2011-01-01

Full Text Available Altered natural killer (NK cell function is a component of the global immune dysregulation that occurs in advanced malignancies. Another condition associated with altered NK homeostasis is normal pregnancy, where robust infiltration with CD16− CD9+ NK cells can be identified in decidual tissues, along with a concomitant expansion of CD16− NK cells in the maternal peripheral blood. In metastatic melanoma, we identified a similar expansion of peripheral blood CD16− NK cells (median 7.4% in 41 patients with melanoma compared with 3.0% in 29 controls, P<.001. A subset of NK cells in melanoma patients also expresses CD9, which is characteristically expressed only on NK cells within the female reproductive tract. Expansion of CD16− NK cells was associated with elevated plasma transforming growth factor-beta (TGF-β levels (median 20 ng/ml, Spearman's ρ=0.81,P=.015. These findings suggest the possibility of exploring anti-TGF-β therapy to restore NK function in melanoma.

Nutrient regulation by continuous feeding removes limitations on cell yield in the large-scale expansion of Mammalian cell spheroids.

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Bradley P Weegman

Full Text Available Cellular therapies are emerging as a standard approach for the treatment of several diseases. However, realizing the promise of cellular therapies across the full range of treatable disorders will require large-scale, controlled, reproducible culture methods. Bioreactor systems offer the scale-up and monitoring needed, but standard stirred bioreactor cultures do not allow for the real-time regulation of key nutrients in the medium. In this study, β-TC6 insulinoma cells were aggregated and cultured for 3 weeks as a model of manufacturing a mammalian cell product. Cell expansion rates and medium nutrient levels were compared in static, stirred suspension bioreactors (SSB, and continuously fed (CF SSB. While SSB cultures facilitated increased culture volumes, no increase in cell yields were observed, partly due to limitations in key nutrients, which were consumed by the cultures between feedings, such as glucose. Even when glucose levels were increased to prevent depletion between feedings, dramatic fluctuations in glucose levels were observed. Continuous feeding eliminated fluctuations and improved cell expansion when compared with both static and SSB culture methods. Further improvements in growth rates were observed after adjusting the feed rate based on calculated nutrient depletion, which maintained physiological glucose levels for the duration of the expansion. Adjusting the feed rate in a continuous medium replacement system can maintain the consistent nutrient levels required for the large-scale application of many cell products. Continuously fed bioreactor systems combined with nutrient regulation can be used to improve the yield and reproducibility of mammalian cells for biological products and cellular therapies and will facilitate the translation of cell culture from the research lab to clinical applications.

Nutrient Regulation by Continuous Feeding Removes Limitations on Cell Yield in the Large-Scale Expansion of Mammalian Cell Spheroids

Science.gov (United States)

Weegman, Bradley P.; Nash, Peter; Carlson, Alexandra L.; Voltzke, Kristin J.; Geng, Zhaohui; Jahani, Marjan; Becker, Benjamin B.; Papas, Klearchos K.; Firpo, Meri T.

2013-01-01

Cellular therapies are emerging as a standard approach for the treatment of several diseases. However, realizing the promise of cellular therapies across the full range of treatable disorders will require large-scale, controlled, reproducible culture methods. Bioreactor systems offer the scale-up and monitoring needed, but standard stirred bioreactor cultures do not allow for the real-time regulation of key nutrients in the medium. In this study, β-TC6 insulinoma cells were aggregated and cultured for 3 weeks as a model of manufacturing a mammalian cell product. Cell expansion rates and medium nutrient levels were compared in static, stirred suspension bioreactors (SSB), and continuously fed (CF) SSB. While SSB cultures facilitated increased culture volumes, no increase in cell yields were observed, partly due to limitations in key nutrients, which were consumed by the cultures between feedings, such as glucose. Even when glucose levels were increased to prevent depletion between feedings, dramatic fluctuations in glucose levels were observed. Continuous feeding eliminated fluctuations and improved cell expansion when compared with both static and SSB culture methods. Further improvements in growth rates were observed after adjusting the feed rate based on calculated nutrient depletion, which maintained physiological glucose levels for the duration of the expansion. Adjusting the feed rate in a continuous medium replacement system can maintain the consistent nutrient levels required for the large-scale application of many cell products. Continuously fed bioreactor systems combined with nutrient regulation can be used to improve the yield and reproducibility of mammalian cells for biological products and cellular therapies and will facilitate the translation of cell culture from the research lab to clinical applications. PMID:24204645

Rapid maxillary expansion effects: An alternative assessment method by means of cone-beam tomography

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Camilo Aquino Melgaço

2014-10-01

Full Text Available INTRODUCTION: This study aims to develop a method to assess the changes in palatal and lingual cross-sectional areas in patients submitted to rapid maxillary expansion (RME. METHODS: The sample comprised 31 Class I malocclusion individuals submitted to RME and divided into two groups treated with Haas (17 patients and Hyrax (14 patients expanders. Cone-beam computed tomography scans were acquired at T0 (before expansion and T1 (six months after screw stabilization. Maxillary and mandibular cross-sectional areas were assessed at first permanent molars and first premolars regions and compared at T0 and T1. Mandibular occlusal area was also analyzed. RESULTS: Maxillary cross-sectional areas increased in 56.18 mm2 and 44.32 mm2 for the posterior and anterior regions. These values were smaller for the mandible, representing augmentation of 40.32 mm2 and 39.91 mm2 for posterior and anterior sections. No differences were found when comparing both expanders. Mandibular occlusal area increased 43.99mm2 and mandibular incisors proclined. Increments of 1.74 mm and 1.7 mm occurred in mandibular intermolar and interpremolar distances. These same distances presented increments of 5.5 mm and 5.57 mm for the maxillary arch. CONCLUSION: Occlusal and cross-sectional areas increased significantly after RME. The method described seems to be reliable and precise to assess intraoral area changes.

Cumulus Cell Expansion, Its Role in Oocyte Biology and Perspectives of Measurement: A Review

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Nevoral J.

2015-01-01

Full Text Available Cumulus expansion of the cumulus-oocyte complex is necessary for meiotic maturation and acquiring developmental competence. Cumulus expansion is based on extracellular matrix synthesis by cumulus cells. Hyaluronic acid is the most abundant component of this extracellular matrix. Cumulus expansion takes place during meiotic oocyte maturation under in vivo and in vitro conditions. Quantification and measurement of cumulus expansion intensity is one possible method of determining oocyte quality and optimizing conditions for in vitro cultivation. Currently, subjective methods of expanded area and more exact cumulus expansion measurement by hyaluronic acid assessment are available. Among the methods of hyaluronic acid measurement is the use of radioactively labelled synthesis precursors. Alternatively, immunological and analytical methods, including enzyme-linked immunosorbent assay (ELISA, spectrophotometry, and high-performance liquid chromatography (HPLC in UV light, could be utilized. The high sensitivity of these methods could provide a precise analysis of cumulus expansion without the use of radioisotopes. Therefore, the aim of this review is to summarize and compare available approaches of cumulus expansion measurement, respecting special biological features of expanded cumuli, and to suggest possible solutions for exact cumulus expansion analysis.

POLYGALACTURONASE INVOLVED IN EXPANSION1 functions in cell elongation and flower development in Arabidopsis.

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Xiao, Chaowen; Somerville, Chris; Anderson, Charles T

2014-03-01

Pectins are acidic carbohydrates that comprise a significant fraction of the primary walls of eudicotyledonous plant cells. They influence wall porosity and extensibility, thus controlling cell and organ growth during plant development. The regulated degradation of pectins is required for many cell separation events in plants, but the role of pectin degradation in cell expansion is poorly defined. Using an activation tag screen designed to isolate genes involved in wall expansion, we identified a gene encoding a putative polygalacturonase that, when overexpressed, resulted in enhanced hypocotyl elongation in etiolated Arabidopsis thaliana seedlings. We named this gene POLYGALACTURONASE INVOLVED IN EXPANSION1 (PGX1). Plants lacking PGX1 display reduced hypocotyl elongation that is complemented by transgenic PGX1 expression. PGX1 is expressed in expanding tissues throughout development, including seedlings, roots, leaves, and flowers. PGX1-GFP (green fluorescent protein) localizes to the apoplast, and heterologously expressed PGX1 displays in vitro polygalacturonase activity, supporting a function for this protein in apoplastic pectin degradation. Plants either overexpressing or lacking PGX1 display alterations in total polygalacturonase activity, pectin molecular mass, and wall composition and also display higher proportions of flowers with extra petals, suggesting PGX1's involvement in floral organ patterning. These results reveal new roles for polygalacturonases in plant development.

DOES INTRAVENOUS TRANEXAMIC ACID REDUCE BLOOD LOSS DURING SURGICALLY ASSISTED RAPID PALATAL EXPANSION?

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Emine AKBAŞ

2017-10-01

Full Text Available Purpose: The purpose of this study was to evaluate the efficacy of tranexamic acid (TXA in reducing blood loss during surgically assisted rapid palatal expansion (SARPE procedure. Subjects and Methods: A total of 34 patients (12 male, 22 female who had been treated surgically under general anesthesia with SARPE including pterygoid disjunction for transverse maxillary deficiency (TMD were included in this study. The study group (n=17 received intravenous (IV TXA 10 mg/kg as a preoperative bolus; the control group (n=17 received normal saline solution. Preoperative and postoperative haemoglobin and haematocrit values, intraoperative blood loss, and any blood product transfusion were recorded. Results: Blood loss during SARPE was statistically significantly less in the study group than the control group (p=0.0001. Conclusion: Preoperative IV administration of TXA can effectively control blood loss during when SARPE with pterygoid disjunction is performed.

Characterization and Application of a Disposable Rotating Bed Bioreactor for Mesenchymal Stem Cell Expansion.

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Neumann, Anne; Lavrentieva, Antonina; Heilkenbrinker, Alexandra; Loenne, Maren; Kasper, Cornelia

2014-11-27

Recruitment of mesenchymal stromal cells (MSC) into the field of tissue engineering is a promising development since these cells can be expanded vivo to clinically relevant numbers and, after expansion, retain their ability to differentiate into various cell lineages. Safety requirements and the necessity to obtain high cell numbers without frequent subcultivation of cells raised the question of the possibility of expanding MSC in one-way (single-use) disposable bioreactors. In this study, umbilical cord-derived MSC (UC-MSC) were expanded in a disposable Z 2000 H bioreactor under dynamic conditions. Z was characterized regarding residence time and mixing in order to evaluate the optimal bioreactor settings, enabling optimal mass transfer in the absence of shear stress, allowing an reproducible expansion of MSC, while maintaining their stemness properties. Culture of the UC-MSC in disposable Z 2000 H bioreactor resulted in a reproducible 8-fold increase of cell numbers after 5 days. Cells were shown to maintain specific MSC surface marker expression as well as trilineage differentiation potential and lack stress-induced premature senescence.

Characterization and Application of a Disposable Rotating Bed Bioreactor for Mesenchymal Stem Cell Expansion

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Anne Neumann

2014-11-01

Full Text Available Recruitment of mesenchymal stromal cells (MSC into the field of tissue engineering is a promising development since these cells can be expanded vivo to clinically relevant numbers and, after expansion, retain their ability to differentiate into various cell lineages. Safety requirements and the necessity to obtain high cell numbers without frequent subcultivation of cells raised the question of the possibility of expanding MSC in one-way (single-use disposable bioreactors. In this study, umbilical cord-derived MSC (UC-MSC were expanded in a disposable Z 2000 H bioreactor under dynamic conditions. Z was characterized regarding residence time and mixing in order to evaluate the optimal bioreactor settings, enabling optimal mass transfer in the absence of shear stress, allowing an reproducible expansion of MSC, while maintaining their stemness properties. Culture of the UC-MSC in disposable Z 2000 H bioreactor resulted in a reproducible 8-fold increase of cell numbers after 5 days. Cells were shown to maintain specific MSC surface marker expression as well as trilineage differentiation potential and lack stress-induced premature senescence.

Optimising cell aggregate expansion in a perfused hollow fibre bioreactor via mathematical modelling.

KAUST Repository

Chapman, Lloyd A C

2014-08-26

The need for efficient and controlled expansion of cell populations is paramount in tissue engineering. Hollow fibre bioreactors (HFBs) have the potential to meet this need, but only with improved understanding of how operating conditions and cell seeding strategy affect cell proliferation in the bioreactor. This study is designed to assess the effects of two key operating parameters (the flow rate of culture medium into the fibre lumen and the fluid pressure imposed at the lumen outlet), together with the cell seeding distribution, on cell population growth in a single-fibre HFB. This is achieved using mathematical modelling and numerical methods to simulate the growth of cell aggregates along the outer surface of the fibre in response to the local oxygen concentration and fluid shear stress. The oxygen delivery to the cell aggregates and the fluid shear stress increase as the flow rate and pressure imposed at the lumen outlet are increased. Although the increased oxygen delivery promotes growth, the higher fluid shear stress can lead to cell death. For a given cell type and initial aggregate distribution, the operating parameters that give the most rapid overall growth can be identified from simulations. For example, when aggregates of rat cardiomyocytes that can tolerate shear stresses of up to 0:05 Pa are evenly distributed along the fibre, the inlet flow rate and outlet pressure that maximise the overall growth rate are predicted to be in the ranges 2.75 x 10(-5) m(2) s(-1) to 3 x 10(-5) m(2) s(-1) (equivalent to 2.07 ml min(-1) to 2.26 ml min(-1)) and 1.077 x 10(5) Pa to 1.083 x 10(5) Pa (or 15.6 psi to 15.7 psi) respectively. The combined effects of the seeding distribution and flow on the growth are also investigated and the optimal conditions for growth found to depend on the shear tolerance and oxygen demands of the cells.

Cryo-electron microscopy and single molecule fluorescent microscopy detect CD4 receptor induced HIV size expansion prior to cell entry

International Nuclear Information System (INIS)

Pham, Son; Tabarin, Thibault; Garvey, Megan; Pade, Corinna; Rossy, Jérémie; Monaghan, Paul; Hyatt, Alex; Böcking, Till; Leis, Andrew; Gaus, Katharina; Mak, Johnson

2015-01-01

Viruses are often thought to have static structure, and they only remodel after the viruses have entered target cells. Here, we detected a size expansion of virus particles prior to viral entry using cryo-electron microscopy (cryo-EM) and single molecule fluorescence imaging. HIV expanded both under cell-free conditions with soluble receptor CD4 (sCD4) targeting the CD4 binding site on the HIV-1 envelope protein (Env) and when HIV binds to receptor on cellular membrane. We have shown that the HIV Env is needed to facilitate receptor induced virus size expansions, showing that the ‘lynchpin’ for size expansion is highly specific. We demonstrate that the size expansion required maturation of HIV and an internal capsid core with wild type stability, suggesting that different HIV compartments are linked and are involved in remodelling. Our work reveals a previously unknown event in HIV entry, and we propose that this pre-entry priming process enables HIV particles to facilitate the subsequent steps in infection. - Highlights: • Cell free viruses are able to receive external trigger that leads to apparent size expansion. • Virus envelope and CD4 receptor engagement is the lynchpin of virus size expansion. • Internal capsid organisation can influence receptor mediated virus size expansion. • Pre-existing virus-associated lipid membrane in cell free virus can accommodate the receptor mediated virus size expansion.

Cryo-electron microscopy and single molecule fluorescent microscopy detect CD4 receptor induced HIV size expansion prior to cell entry

Energy Technology Data Exchange (ETDEWEB)

Pham, Son [Deakin University, Victoria 3216 (Australia); CSIRO Australian Animal Health Laboratory, Victoria 3220 (Australia); Tabarin, Thibault [ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, New South Wales 3220 (Australia); Garvey, Megan; Pade, Corinna [Deakin University, Victoria 3216 (Australia); CSIRO Australian Animal Health Laboratory, Victoria 3220 (Australia); Rossy, Jérémie [ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, New South Wales 3220 (Australia); Monaghan, Paul; Hyatt, Alex [CSIRO Australian Animal Health Laboratory, Victoria 3220 (Australia); Böcking, Till [ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, New South Wales 3220 (Australia); Leis, Andrew [CSIRO Australian Animal Health Laboratory, Victoria 3220 (Australia); Gaus, Katharina, E-mail: k.gaus@unsw.edu.au [ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, New South Wales 3220 (Australia); Mak, Johnson, E-mail: j.mak@deakin.edu.au [Deakin University, Victoria 3216 (Australia); CSIRO Australian Animal Health Laboratory, Victoria 3220 (Australia)

2015-12-15

Viruses are often thought to have static structure, and they only remodel after the viruses have entered target cells. Here, we detected a size expansion of virus particles prior to viral entry using cryo-electron microscopy (cryo-EM) and single molecule fluorescence imaging. HIV expanded both under cell-free conditions with soluble receptor CD4 (sCD4) targeting the CD4 binding site on the HIV-1 envelope protein (Env) and when HIV binds to receptor on cellular membrane. We have shown that the HIV Env is needed to facilitate receptor induced virus size expansions, showing that the ‘lynchpin’ for size expansion is highly specific. We demonstrate that the size expansion required maturation of HIV and an internal capsid core with wild type stability, suggesting that different HIV compartments are linked and are involved in remodelling. Our work reveals a previously unknown event in HIV entry, and we propose that this pre-entry priming process enables HIV particles to facilitate the subsequent steps in infection. - Highlights: • Cell free viruses are able to receive external trigger that leads to apparent size expansion. • Virus envelope and CD4 receptor engagement is the lynchpin of virus size expansion. • Internal capsid organisation can influence receptor mediated virus size expansion. • Pre-existing virus-associated lipid membrane in cell free virus can accommodate the receptor mediated virus size expansion.

Use of high throughput qPCR screening to rapidly clone low frequency tumour specific T-cells from peripheral blood for adoptive immunotherapy

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Serrano Oscar K

2008-10-01

Full Text Available Abstract Background The adoptive transfer of autologous tumor reactive lymphocytes can mediate significant tumor regression in some patients with refractory metastatic cancer. However, a significant obstacle for this promising therapy has been the availability of highly efficient methods to rapidly isolate and expand a variety of potentially rare tumor reactive lymphocytes from the natural repertoire of cancer patients. Methods We developed a novel in vitro T cell cloning methodology using high throughput quantitative RT-PCR (qPCR assay as a rapid functional screen to detect and facilitate the limiting dilution cloning of a variety of low frequency T cells from bulk PBMC. In preclinical studies, this strategy was applied to the isolation and expansion of gp100 specific CD8+ T cell clones from the peripheral blood of melanoma patients. Results In optimization studies, the qPCR assay could detect the reactivity of 1 antigen specific T cell in 100,000 background cells. When applied to short term sensitized PBMC microcultures, this assay could detect T cell reactivity against a variety of known melanoma tumor epitopes. This screening was combined with early limiting dilution cloning to rapidly isolate gp100154–162 reactive CD8+ T cell clones. These clones were highly avid against peptide pulsed targets and melanoma tumor lines. They had an effector memory phenotype and showed significant proliferative capacity to reach cell numbers appropriate for adoptive transfer trials (~1010 cells. Conclusion This report describes a novel high efficiency strategy to clone tumor reactive T cells from peripheral blood for use in adoptive immunotherapy.

Early peroxisome proliferator-activated receptor gamma regulated genes involved in expansion of pancreatic beta cell mass

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Vivas Yurena

2011-12-01

Full Text Available Abstract Background The progression towards type 2 diabetes depends on the allostatic response of pancreatic beta cells to synthesise and secrete enough insulin to compensate for insulin resistance. The endocrine pancreas is a plastic tissue able to expand or regress in response to the requirements imposed by physiological and pathophysiological states associated to insulin resistance such as pregnancy, obesity or ageing, but the mechanisms mediating beta cell mass expansion in these scenarios are not well defined. We have recently shown that ob/ob mice with genetic ablation of PPARγ2, a mouse model known as the POKO mouse failed to expand its beta cell mass. This phenotype contrasted with the appropriate expansion of the beta cell mass observed in their obese littermate ob/ob mice. Thus, comparison of these models islets particularly at early ages could provide some new insights on early PPARγ dependent transcriptional responses involved in the process of beta cell mass expansion Results Here we have investigated PPARγ dependent transcriptional responses occurring during the early stages of beta cell adaptation to insulin resistance in wild type, ob/ob, PPARγ2 KO and POKO mice. We have identified genes known to regulate both the rate of proliferation and the survival signals of beta cells. Moreover we have also identified new pathways induced in ob/ob islets that remained unchanged in POKO islets, suggesting an important role for PPARγ in maintenance/activation of mechanisms essential for the continued function of the beta cell. Conclusions Our data suggest that the expansion of beta cell mass observed in ob/ob islets is associated with the activation of an immune response that fails to occur in POKO islets. We have also indentified other PPARγ dependent differentially regulated pathways including cholesterol biosynthesis, apoptosis through TGF-β signaling and decreased oxidative phosphorylation.

Culture expansion of adipose derived stromal cells. A closed automated Quantum Cell Expansion System compared with manual flask-based culture

DEFF Research Database (Denmark)

Haack-Sørensen, Mandana; Follin, Bjarke; Juhl, Morten

2016-01-01

) over two passages in the automated and functionally closed Quantum Cell Expansion System (Quantum system) is compared with traditional manual cultivation. Methods: Stromal vascular fraction was isolated from abdominal fat, suspended in α-MEM supplemented with 10% Fetal Bovine Serum and seeded...... into either T75 flasks or a Quantum system that had been coated with cryoprecipitate. The cultivation of ASCs from SVF was performed in 3 ways: flask to flask; flask to Quantum system; and Quantum system to Quantum system. In all cases, quality controls were conducted for sterility, mycoplasmas......, and endotoxins, in addition to the assessment of cell counts, viability, immunophenotype, and differentiation potential. Results: The viability of ASCs passage 0 (P0) and P1 was above 96%, regardless of cultivation in flasks or Quantum system. Expression of surface markers and differentiation potential...

Neonatal colonisation expands a specific intestinal antigen-presenting cell subset prior to CD4 T-cell expansion, without altering T-cell repertoire.

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Charlotte F Inman

Full Text Available Interactions between the early-life colonising intestinal microbiota and the developing immune system are critical in determining the nature of immune responses in later life. Studies in neonatal animals in which this interaction can be examined are central to understanding the mechanisms by which the microbiota impacts on immune development and to developing therapies based on manipulation of the microbiome. The inbred piglet model represents a system that is comparable to human neonates and allows for control of the impact of maternal factors. Here we show that colonisation with a defined microbiota produces expansion of mucosal plasma cells and of T-lymphocytes without altering the repertoire of alpha beta T-cells in the intestine. Importantly, this is preceded by microbially-induced expansion of a signal regulatory protein α-positive (SIRPα(+ antigen-presenting cell subset, whilst SIRPα(-CD11R1(+ antigen-presenting cells (APCs are unaffected by colonisation. The central role of intestinal APCs in the induction and maintenance of mucosal immunity implicates SIRPα(+ antigen-presenting cells as orchestrators of early-life mucosal immune development.

[The effect of bFGF and sucralfate on cell proliferation during continuous tissue expansion].

Science.gov (United States)

Hu, Ya-lan; Guo, Shu-zhong; Lu, Kai-hua

2003-05-01

To investigate the effect of local application of bFGF combined with sucralfate on the cell proliferation during continuous tissue expansion (CTE). Nine white pigs were selected to undergo the continuous tissue expansion in this study and treated with bFGF and sucralfate, respectively as the following groups: group 1 with both bFGF and sucralfate, group 2 only with bFGF, group 3 with only sucralfate, and group 4 with saline as control. Fifteen samples were taken in each pig for immunohistochemistry analysis 1-14 days and 6 weeks after the operation. In the group with both bFGF and sucralfate, the epidermic basal cells proliferated significantly after the operation and reached top level in 3 days, which was statistical higher than the control group, but the multiplication of basal cell was the lowest 14 days after the operation, still more than the control group. In dermal layer, proliferation of fibroblasts, vessel endothelial cells, hair follicles epidermic cells and sweat gland epicytes was also significant higher in the group with both bFGF and sucralfate than that the control group and reached top level 7 day after the operation, but the proliferation of cells decreased obviously 14 days after the operation, still higher than the control group. The mitotic activity of cells returned to the basal level in 42 days. There were no significant differences among the group 2, group 3 and group 4. Local application of both bFGF and sucralfate could be more effect to induce cells multiplication during early skin expansion to facilitate the growth of neoformed skin soft tissue.

Self-renewal of single mouse hematopoietic stem cells is reduced by JAK2V617F without compromising progenitor cell expansion.

Science.gov (United States)

Kent, David G; Li, Juan; Tanna, Hinal; Fink, Juergen; Kirschner, Kristina; Pask, Dean C; Silber, Yvonne; Hamilton, Tina L; Sneade, Rachel; Simons, Benjamin D; Green, Anthony R

2013-01-01

Recent descriptions of significant heterogeneity in normal stem cells and cancers have altered our understanding of tumorigenesis, emphasizing the need to understand how single stem cells are subverted to cause tumors. Human myeloproliferative neoplasms (MPNs) are thought to reflect transformation of a hematopoietic stem cell (HSC) and the majority harbor an acquired V617F mutation in the JAK2 tyrosine kinase, making them a paradigm for studying the early stages of tumor establishment and progression. The consequences of activating tyrosine kinase mutations for stem and progenitor cell behavior are unclear. In this article, we identify a distinct cellular mechanism operative in stem cells. By using conditional knock-in mice, we show that the HSC defect resulting from expression of heterozygous human JAK2V617F is both quantitative (reduced HSC numbers) and qualitative (lineage biases and reduced self-renewal per HSC). The defect is intrinsic to individual HSCs and their progeny are skewed toward proliferation and differentiation as evidenced by single cell and transplantation assays. Aged JAK2V617F show a more pronounced defect as assessed by transplantation, but mice that transform reacquire competitive self-renewal ability. Quantitative analysis of HSC-derived clones was used to model the fate choices of normal and JAK2-mutant HSCs and indicates that JAK2V617F reduces self-renewal of individual HSCs but leaves progenitor expansion intact. This conclusion is supported by paired daughter cell analyses, which indicate that JAK2-mutant HSCs more often give rise to two differentiated daughter cells. Together these data suggest that acquisition of JAK2V617F alone is insufficient for clonal expansion and disease progression and causes eventual HSC exhaustion. Moreover, our results show that clonal expansion of progenitor cells provides a window in which collaborating mutations can accumulate to drive disease progression. Characterizing the mechanism(s) of JAK2V617F

Cone-beam computed tomography evaluation of dental, skeletal, and alveolar bone changes associated with bonded rapid maxillary expansion

Directory of Open Access Journals (Sweden)

Namrata Dogra

2016-01-01

Full Text Available Aims and Objectives: To evaluate skeletal changes in maxilla and its surrounding structures, changes in the maxillary dentition and maxillary alveolar bone changes produced by bonded rapid maxillary expansion (RME using cone-beam computed tomography (CBCT. Materials and Methods: The sample consisted of 10 patients (6 males and 4 females with age range 12 to 15 years treated with bonded RME. CBCT scans were performed at T1 (pretreatment and at T2 (immediately after expansion to evaluate the dental, skeletal, and alveolar bone changes. Results: RME treatment increased the overall skeletal parameters such as interorbital, zygomatic, nasal, and maxillary widths. Significant increases in buccal maxillary width was observed at first premolar, second premolar, and first molar level. There was a significant increase in arch width both on the palatal side and on the buccal side. Significant tipping of right and left maxillary first molars was seen. There were significant reductions in buccal bone plate thickness and increase in palatal bone plate thickness. Conclusions: Total expansion achieved with RME was a combination of dental, skeletal and alveolar bone changes. At the first molar level, 28.45% orthopedic, 16.03% alveolar bone bending, and 55.5% orthodontic changes were observed.

Ultraviolet B irradiation induces expansion of intraepithelial tumor cells in a tissue model of early cancer progression.

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Mudgil, Adarsh V; Segal, Nadav; Andriani, Frank; Wang, Youai; Fusenig, Norbert E; Garlick, Jonathan A

2003-07-01

Ultraviolet B irradiation is thought to enable skin cancer progression as clones of genetically damaged keratinocytes escape apoptosis and expand at the expense of adjacent normal cells. Mechanisms through which potentially malignant cells in human skin undergo clonal expansion, however, are not well understood. The goal of this study was to characterize the role of ultraviolet B irradiation on the intraepithelial expansion of early stage human tumor cells in organotypic skin cultures. To accomplish this, we have studied the effect of ultraviolet B irradiation on organotypic cultures that were fabricated by mixing normal human keratinocytes with beta-galactosidase-marked, intraepithelial tumor cells (HaCaT-ras, clone II-4), which bear mutations in both p53 alleles and harbor an activated H-ras oncogene. We found that when organotypic mixtures were exposed to an ultraviolet B dose of 50 mJ per cm2, intraepithelial tumor cells underwent a significant degree of proliferative expansion compared to nonirradiated cultures. To understand this response, organotypic cultures of nor-mal keratinocytes were exposed to ultraviolet B and showed a dose-dependent increase in numbers of sunburn cells and TUNEL-positive cells although their proliferation was suppressed. In contrast, neither the apoptotic nor the proliferative response of II-4 cells was altered by ultraviolet B in organotypic cultures. The differential response of these cell types suggested that II-4 cells were resistant to ultraviolet-B-induced alterations, which allowed these intraepithelial tumor cells to gain a selective growth and survival advantage relative to neighboring normal cells. These findings demonstrate that ultraviolet B exposure can induce the intraepithelial expansion of apoptosis-resistant, p53-mutant, and ras-activated keratinocytes, suggesting that this agent can act to promote the early stages of epithelial carcinogenesis.

Optimization of Human NK Cell Manufacturing: Fully Automated Separation, Improved Ex Vivo Expansion Using IL-21 with Autologous Feeder Cells, and Generation of Anti-CD123-CAR-Expressing Effector Cells.

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Klöß, Stephan; Oberschmidt, Olaf; Morgan, Michael; Dahlke, Julia; Arseniev, Lubomir; Huppert, Volker; Granzin, Markus; Gardlowski, Tanja; Matthies, Nadine; Soltenborn, Stephanie; Schambach, Axel; Koehl, Ulrike

2017-10-01

The administration of ex vivo expanded natural killer (NK) cells as potential antitumor effector cells appears to be suitable for effector cell-based immunotherapies in high-risk cancer patients. However, good manufacturing practice (GMP)-compliant manufacturing of clinical-grade NK cells at sufficiently high numbers represents a great challenge. Therefore, previous expansion protocols for those effector cells were improved and optimized by using newly developed culture medium, interleukin (IL)-21, and autologous feeder cells (FCs). Separation of primary human NK cells (CD56 + CD3 - ) was carried out with the CliniMACS Prodigy ® in a single process, starting with approximately 1.2 × 10 9 leukocytes collected by small-scale lymphapheresis or from buffy coats. Enriched NK cells were adjusted to starting cell concentrations within approximately 1 × 10 6 effector cells/mL and cultured in comparative expansion experiments for 14 days with IL-2 (1,000 IU/mL) in different GMP-compliant media (X-VIVO ™ 10, CellGro ® , TexMACS ™ , and NK MACS ® ). After medium optimization, beneficial effects for functionality and phenotype were investigated at the beginning of cell expansion with irradiated (25 Gy) autologous FCs at a ratio of 20:1 (feeder: NK) in the presence or absence of IL-21 (100 ng/mL). Additionally, expanded NK cells were gene modified to express chimeric antigen receptors (CARs) against CD123, a common marker for acute myeloid leukemia (AML). Cytotoxicity, degranulation, and cytokine release of transduced NK cells were determined against KG1a cells in flow cytometric analysis and fluorescent imaging. The Prodigy manufacturing process revealed high target cell viabilities (median 95.4%), adequate NK cell recovery (median 60.4%), and purity of 95.4% in regard to CD56 + CD3 - target cells. The process in its early phase of development led to a median T-cell depletion of log 3.5 after CD3 depletion and log 3.6 after the whole process, including CD3

Ex Vivo Expansion of Hematopoietic Stem Cells to Improve Engraftment in Stem Cell Transplantation.

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Ko, Kap-Hyoun; Nordon, Robert; O'Brien, Tracey A; Symonds, Geoff; Dolnikov, Alla

2017-01-01

The efficient use of hematopoietic stem cells (HSC) for transplantation is often limited by the relatively low numbers of HSC collected. The ex vivo expansion of HSC for clinical use is a potentially valuable and safe approach to increase HSC numbers thereby increasing engraftment and reducing the risk of morbidity from infection. Here, we describe a protocol for the robust ex vivo expansion of human CD34(+) HSC isolated from umbilical cord blood. The protocol described can efficiently generate large numbers of HSC. We also describe a flow cytometry-based method using high-resolution division tracking to characterize the kinetics of HSC growth and differentiation. Utilizing the guidelines discussed, it is possible for investigators to use this protocol as presented or to modify it for their specific needs.

Cell expansion of human articular chondrocytes on macroporous gelatine scaffolds-impact of microcarrier selection on cell proliferation

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Pettersson, Sofia; Kratz, Gunnar [Laboratory for Reconstructive Plastic Surgery, Department of Clinical and Experimental Medicine, Linkoeping University, SE-581 85 Linkoeping (Sweden); Wetteroe, Jonas [Rheumatology/AIR, Department of Clinical and Experimental Medicine, Linkoeping University, SE-581 85 Linkoeping (Sweden); Tengvall, Pentti, E-mail: sofia.pettersson@liu.se [Institute of Clinical Sciences, Department of Biomaterials, The Sahlgrenska Academy at University of Gothenburg, SE-405 30 Gothenburg (Sweden)

2011-12-15

This study investigates human chondrocyte expansion on four macroporous gelatine microcarriers (CultiSpher) differing with respect to two manufacturing processes-the amount of emulsifier used during initial preparation and the gelatine cross-linking medium. Monolayer-expanded articular chondrocytes from three donors were seeded onto the microcarriers and cultured in spinner flask systems for a total of 15 days. Samples were extracted every other day to monitor cell viability and establish cell counts, which were analysed using analysis of variance and piecewise linear regression. Chondrocyte densities increased according to a linear pattern for all microcarriers, indicating an ongoing, though limited, cell proliferation. A strong chondrocyte donor effect was seen during the initial expansion phase. The final cell yield differed significantly between the microcarriers and our results indicate that manufacturing differences affected chondrocyte densities at this point. Remaining cells stained positive for chondrogenic markers SOX-9 and S-100 but extracellular matrix formation was modest to undetectable. In conclusion, the four gelatine microcarriers supported chondrocyte adhesion and proliferation over a two week period. The best yield was observed for microcarriers produced with low emulsifier content and cross-linked in water and acetone. These results add to the identification of optimal biomaterial parameters for specific cellular processes and populations.

Environmentally benign formation of polymeric microspheres by rapid expansion of supercritical carbon dioxide solution with a nonsolvent.

Science.gov (United States)

Matsuyama, K; Mishima, K; Umemoto, H; Yamaguchi, S

2001-10-15

A novel method is reported for forming polymer microparticles, which reduce atmospheric emissions of environmentally harmful volatile organic compounds such as toluene and xylene used as paint solvent in paint industry. The polymer microparticles have formed through rapid expansion from supercritical solution with a nonsolvent (RESS-N). Solubilization of poly(styrene)-b-(poly(methyl methacrylate)-co-poly (glycidyl methacrylate)) copolymer(PS-b-(PMMA-co-PGMA), MW = 5000, PS/PMMA/PGMA = 2/5/3), poly(ethylene glycol) (PEG, M. W = 4000), bisphenol A type epoxy resin (EP, MW = 3000), poly(methyl methacrylate) (PMMA; MW = 15000, 75000, 120000), and poly(oxyalkylene) alkylphenyl ether (MW = 4000) in carbon dioxide (CO2) was achieved with the use of small alcohols as cosolvents. The solubility of the PS-b-(PMMA-co-PGMA) is extremely low in either CO2 or ethanol but becomes 20 wt % in a mixture of the two. Because ethanol is a nonsolvent for the polymer, it can be used as a cosolvent in rapid expansion from supercritical solution to produce 1-3 microm particles that do not agglomerate. Obtained polymer particles by RESS-N were applied as powder coatings. The resulting coatings have a smooth and coherent film. The particle size distribution of microspheres was controlled by changing the polymer concentration, preexpansion pressure, temperature, and injection distance. The feed compositions were more effective than the other factors in controlling the particle size. The polymeric microparticles formed by RESS-N method can be utilized to make the thin coating film without anytoxic organic solvents and/or surfactants.

Skin cell isolation and expansion for cell transplantation is limited in patients using tobacco, alcohol, or are exhibiting diabetes mellitus.

Science.gov (United States)

Johnen, Christa; Hartmann, Bernd; Steffen, Ingo; Bräutigam, Kirsten; Witascheck, Tom; Toman, Nidal; Küntscher, Markus V; Gerlach, Jörg C

2006-03-01

The aim of this exploratory study was to investigate the isolation and expansion of keratinocytes and fibroblasts from donors with certain medical histories. Biopsies were taken from donors (N=32) falling into one or more of the following categories: a history of heavy smoking and/or alcohol abuse, drug abuse, diabetes mellitus or steroid treatment. Cells from donors who did not fall into any of the above-mentioned categories were used as controls. Proliferation and growth behaviour of cells were analyzed by measurement of passage duration, absorbance (MTT-assay) and light microscopy. Donors with a specific medical history required larger biopsy areas than the control group for isolating a sufficient number of fibroblasts and keratinocytes. Times to confluence were significantly prolonged and absorbances (MTT) were significantly reduced in several donor groups when compared to control cultures. Biopsies from donors with steroid treatment, drug abuse and combined nicotine and alcohol abuse could not be established beyond passage 0 degrees or 1 degree, respectively. We conclude that isolation and expansion of skin cells from donors with certain medical histories may require larger biopsies, prolonged expansion times or may even result in failure. These findings may therefore be of clinical importance in the field of autologous skin cell transplantation.

Rapid, global demographic expansions after the origins of agriculture.

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Gignoux, Christopher R; Henn, Brenna M; Mountain, Joanna L

2011-04-12

The invention of agriculture is widely assumed to have driven recent human population growth. However, direct genetic evidence for population growth after independent agricultural origins has been elusive. We estimated population sizes through time from a set of globally distributed whole mitochondrial genomes, after separating lineages associated with agricultural populations from those associated with hunter-gatherers. The coalescent-based analysis revealed strong evidence for distinct demographic expansions in Europe, southeastern Asia, and sub-Saharan Africa within the past 10,000 y. Estimates of the timing of population growth based on genetic data correspond neatly to dates for the initial origins of agriculture derived from archaeological evidence. Comparisons of rates of population growth through time reveal that the invention of agriculture facilitated a fivefold increase in population growth relative to more ancient expansions of hunter-gatherers.

Evaluation of surgically assisted rapid maxillary expansion with piezosurgery versus oscillating saw and chisel osteotomy - a randomized prospective trial.

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Rana, Majeed; Gellrich, Nils-Claudius; Rana, Madiha; Piffkó, Jozsef; Kater, Wolfgang

2013-02-17

Ultrasonic bone-cutting surgery has been introduced as a feasible alternative to the conventional sharp instruments used in craniomaxillofacial surgery because of its precision and safety. The piezosurgery medical device allows the efficient cutting of mineralized tissues with minimal trauma to soft tissues. Piezoelectric osteotome has found its role in surgically assisted rapid maxillary expansion (SARME), a procedure well established to correct transverse maxillary discrepancies. The advantages include minimal risk to critical anatomic structures. The purpose of this clinical comparative study (CIS 2007-237-M) was to present the advantages of the piezoelectric cut as a minimally invasive device in surgically assisted, rapid maxillary expansion by protecting the maxillary sinus mucosal lining. Thirty patients (18 females and 12 males) at the age of 18 to 54 underwent a surgically assisted palatal expansion of the maxilla with a combined orthodontic and surgical approach. The patients were randomly divided into two separate treatment groups. While Group 1 received conventional surgery using an oscillating saw, Group 2 was treated with piezosurgery. The following parameters were examined: blood pressure, blood values, required medication, bleeding level in the maxillary sinus, duration of inpatient stay, duration of surgery and height of body temperature. The results displayed no statistically significant differences between the two groups regarding laboratory blood values and inpatient stay. The duration of surgery revealed a significant discrepancy. Deploying piezosurgery took the surgeon an average of 10 minutes longer than working with a conventional-saw technique. However, the observation of the bleeding level in the paranasal sinus presented a major and statistically significant advantage of piezosurgery: on average the bleeding level was one category above the one of the remaining patients. This method of piezoelectric surgery with all its advantages is going

Anchorage onto deciduous teeth: effectiveness of early rapid maxillary expansion in increasing dental arch dimension and improving anterior crowding.

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Mutinelli, Sabrina; Manfredi, Mario; Guiducci, Antonio; Denotti, Gloria; Cozzani, Mauro

2015-01-01

Anchorage onto permanent dentition is a common procedure in rapid maxillary expansion. However, replacing first permanent molars with the second deciduous molars seems to be an option to reduce some negative side effects during orthodontic treatment. The purpose of this study was to evaluate the dental effect of rapid maxillary expansion with anchorage exclusively onto deciduous teeth performed in the first period of transition. Twenty patients with a lateral cross-bite treated exclusively by a Haas expander in early mixed dentition were retrospectively analyzed before treatment, at appliance removal, and at 21 months out of retention. The sagittal and transverse dimensions, together with the inter-canine arch and irregularity index, were digitally measured on scanned images of dental casts. The patients were compared with three balanced control groups (in total, 60 individuals) matched for gender. Two control groups had the same canine dental class as the treated group at T1, were in the inter-transitional period, and either had or lacked a lateral cross-bite. The last control group was comprised of adolescents in permanent dentition with a dental class I. The statistical analysis was performed by means of repeated-measures ANOVA for paired data and one-way ANOVA, the Kruskal-Wallis test, and the Mann-Whitney test for independent measures (α-level p dentition), the dental arch dimensions of treated patients were similar to those of adolescents with a dental class I and significantly wider than those of patients with a lateral cross-bite. Also, the anterior irregularity index was lower among patients who had undergone expansion treatments than in all untreated study participants. The Haas expander anchored to the deciduous teeth is effective in increasing the dental arch width in patients with a lateral cross-bite. The dimensions of the dental arch were modified earlier toward the values of the permanent dentition.

Influence of patient related factors on number of mesenchymal stromal cells reached after in vitro culture expansion for clinical treatment

DEFF Research Database (Denmark)

Qayyum, Abbas Ali; Kaur, Kamal Preet; Mathiasen, Anders Bruun

2017-01-01

of autologous stromal cells reached after in vitro culture expansion for clinical therapy. METHODS: Culture expansion data from 111 patients with IHD treated with autologous stromal cells in three clinical trials were used. We correlated the final cell count after two passages of cultivation with different...... correlation between left ventricular ejection fraction and number of MSCs was found (r = -0.287, p = .017). CONCLUSIONS: Patient related factors such as BMI, hypertension and gender may influence the number of MSCs reached after in vitro culture expansion....... patient factors. RESULTS: There was a significant relation between body mass index (BMI) and the number of adipose derived stromal cells (ASCs) reached after culture expansion and for all patients included into the three studies (r = 0.375, p = .019 and r = 0.200, p = .036, respectively). Moreover...

Characterization and Application of a Disposable Rotating BedBioreactor for Mesenchymal Stem Cell Expansion

OpenAIRE

Neumann, Anne; Lavrentieva, Antonina; Heilkenbrinker, Alexandra; Loenne, Maren; Kasper, Cornelia

2014-01-01

Recruitment of mesenchymal stromal cells (MSC) into the field of tissue engineering is a promising development since these cells can be expanded vivo to clinically relevant numbers and, after expansion, retain their ability to differentiate into various cell lineages. Safety requirements and the necessity to obtain high cell numbers without frequent subcultivation of cells raised the question of the possibility of expanding MSC in one-way (single-use) disposable bioreactors. In this study, u...

Interceptive treatment of palatal impaction of maxillary canines with rapid maxillary expansion: a randomized clinical trial.

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Baccetti, Tiziano; Mucedero, Manuela; Leonardi, Maria; Cozza, Paola

2009-11-01

Our aim in this prospective randomized clinical study was to assess the prevalence rate of eruption of palatally displaced canines, diagnosed at an early developmental stage with posteroanterior radiographs and consequently treated by rapid maxillary expansion. A sample of 60 subjects in the early mixed dentition with palatally displaced canines diagnosed on posteroanterior radiographs was enrolled in the trial. Their age range at the first observation (T1) was 7.6 to 9.6 years, with a prepubertal stage of skeletal maturity (CS1 or CS2). The 60 subjects were randomly allocated to the treatment group (TG, 35 subjects) or the no-treatment group (NTG, 25 subjects). The TG was treated with a banded rapid maxillary expander; after expansion, all patients were retained with the expander in place for 6 months. Thereafter, the expander was removed, and the patients wore a retention plate at night for a year. The NTG received no treatment. All subjects were reevaluated in the early permanent dentition (T2) (postpubertal CS4). The number of dropouts was recorded. The main outcome recorded at T2 was successful or unsuccessful eruption of the maxillary permanent canines. The starting forms at T1 for measurements on posteroanterior and panoramic films were compared in the 2 groups with the Mann-Whitney U test (P <0.05). The prevalence rates of successful and unsuccessful treatments in the TG were compared with those in NTG with chi-square tests (P <0.05). From T1 to T2, there were 3 dropouts in each group. The final sample comprised 32 subjects in the TG and 22 subjects in the NTG. No statistically significant differences were found for any variable at T1. The prevalence rates of successful eruption of the maxillary canines were 65.7% (21 subjects) in the TG and 13.6% (3 subjects) in the NTG. The comparison was statistically significant (chi-square = 12.4; P <0.001). Subjects with palatally displaced canines in the early mixed dentition do not have transverse deficiency of the

Elastic hydrogel substrate supports robust expansion of murine myoblasts and enhances their engraftment

International Nuclear Information System (INIS)

Ding, Ke; Yang, Zhong; Xu, Jian-zhong; Liu, Wen-ying; Zeng, Qiang; Hou, Fang; Lin, Sen

2015-01-01

The application of satellite cell-derived myoblasts in regenerative medicine has been restricted by the rapid loss of stemness during in vitro cell expansion using traditional culture systems. However, studies published in the past decade have highlighted the influence of substrate elasticity on stem cell fate and revealed that culture on a soft hydrogel substrate can promote self-renewal and prolong the regenerative potential of muscle stem cells. Whether hydrogel substrates have similar effects after long-term robust expansion remains to be determined. Herein we prepared an elastic chitosan/beta-glycerophosphate/collagen hydrogel mimicking the soft microenvironment of muscle tissues for use as the substrate for satellite cell culture and investigated its influence on long-term cell expansion. After 20 passages in culture, satellite cell-derived myoblasts cultured on our hydrogel substrate exhibited significant improvements in proliferation capability, cell viability, colony forming frequency, and potential for myogenic differentiation compared to those cultured on a routine rigid culture surface. Immunochemical staining and western blot analysis both confirmed that myoblasts cultured on the hydrogel substrate expressed higher levels of several differentiation-related markers, including Pax7, Pax3, and SSEA-1, and a lower level of MyoD compared to myoblasts cultured on rigid culture plates (all p<0.05). After transplantation into the tibialis anterior of nude mice, myoblasts that had been cultured on the hydrogel substrate demonstrated a significantly greater engraftment efficacy than those cultured on the traditional surface. Collectively, these results indicate that the elastic hydrogel substrate supported robust expansion of murine myoblasts and enhanced their engraftment in vivo. - Highlights: • An elastic hydrogel was designed to mimic the pliable muscle tissue microenvironment. • Myoblasts retained their stemness in long-term culture on the elastic

Elastic hydrogel substrate supports robust expansion of murine myoblasts and enhances their engraftment

Energy Technology Data Exchange (ETDEWEB)

Ding, Ke, E-mail: dk1118@yeah.net [Department of Pediatric Surgery, Sichuan Academy of Medical Sciences & Sichuan Provincial People’s Hospital, Chengdu 610072 (China); Yang, Zhong [Department of Clinical Hematology, Southwest Hospital, Third Military Medical University, Chongqing 400038 (China); Xu, Jian-zhong, E-mail: xjzspine@163.com [Department of Orthopaedics, Southwest Hospital, Third Military Medical University, Chongqing 400038 (China); Liu, Wen-ying; Zeng, Qiang; Hou, Fang [Department of Pediatric Surgery, Sichuan Academy of Medical Sciences & Sichuan Provincial People’s Hospital, Chengdu 610072 (China); Lin, Sen [Department of Anatomy and Histology & Embryology, Chengdu Medical College, Chengdu 610500 (China)

2015-09-10

The application of satellite cell-derived myoblasts in regenerative medicine has been restricted by the rapid loss of stemness during in vitro cell expansion using traditional culture systems. However, studies published in the past decade have highlighted the influence of substrate elasticity on stem cell fate and revealed that culture on a soft hydrogel substrate can promote self-renewal and prolong the regenerative potential of muscle stem cells. Whether hydrogel substrates have similar effects after long-term robust expansion remains to be determined. Herein we prepared an elastic chitosan/beta-glycerophosphate/collagen hydrogel mimicking the soft microenvironment of muscle tissues for use as the substrate for satellite cell culture and investigated its influence on long-term cell expansion. After 20 passages in culture, satellite cell-derived myoblasts cultured on our hydrogel substrate exhibited significant improvements in proliferation capability, cell viability, colony forming frequency, and potential for myogenic differentiation compared to those cultured on a routine rigid culture surface. Immunochemical staining and western blot analysis both confirmed that myoblasts cultured on the hydrogel substrate expressed higher levels of several differentiation-related markers, including Pax7, Pax3, and SSEA-1, and a lower level of MyoD compared to myoblasts cultured on rigid culture plates (all p<0.05). After transplantation into the tibialis anterior of nude mice, myoblasts that had been cultured on the hydrogel substrate demonstrated a significantly greater engraftment efficacy than those cultured on the traditional surface. Collectively, these results indicate that the elastic hydrogel substrate supported robust expansion of murine myoblasts and enhanced their engraftment in vivo. - Highlights: • An elastic hydrogel was designed to mimic the pliable muscle tissue microenvironment. • Myoblasts retained their stemness in long-term culture on the elastic

Wnt3a nanodisks promote ex vivo expansion of hematopoietic stem and progenitor cells

DEFF Research Database (Denmark)

Lalefar, Nahal R.; Witkowski, Andrzej; Simonsen, Jens Bæk

2016-01-01

Background : Wnt proteins modulate development, stem cell fate and cancer through interactions with cell surface receptors. Wnts are cysteine-rich, glycosylated, lipid modified, two domain proteins that are prone to aggregation. The culprit responsible for this behavior is a covalently bound palm...... to Lin- Sca-1+ c-Kit+ cell expansion, an effect that was not mediated through β-catenin. Conclusions : The data indicate Wnt3a ND constitute a water-soluble transport vehicle capable of promoting ex vivo expansion of HSPC.......Background : Wnt proteins modulate development, stem cell fate and cancer through interactions with cell surface receptors. Wnts are cysteine-rich, glycosylated, lipid modified, two domain proteins that are prone to aggregation. The culprit responsible for this behavior is a covalently bound...... palmitoleoyl moiety in the N-terminal domain. Results : By combining murine Wnt3a with phospholipid and apolipoprotein A-I, ternary complexes termed nanodisks (ND) were generated. ND-associated Wnt3a is soluble in the absence of detergent micelles and gel filtration chromatography revealed that Wnt3a co...

* Tissue-Specific Extracellular Matrix Enhances Skeletal Muscle Precursor Cell Expansion and Differentiation for Potential Application in Cell Therapy.

Science.gov (United States)

Zhang, Deying; Zhang, Yong; Zhang, Yuanyuan; Yi, Hualin; Wang, Zhan; Wu, Rongpei; He, Dawei; Wei, Guanghui; Wei, Shicheng; Hu, Yun; Deng, Junhong; Criswell, Tracy; Yoo, James; Zhou, Yu; Atala, Anthony

2017-08-01

Skeletal muscle precursor cells (MPCs) are considered a key candidate for cell therapy in the treatment of skeletal muscle dysfunction due to injury, disease, or age. However, expansion of a sufficient number of functional skeletal muscle cells in vitro from a small tissue biopsy has been challenging due to changes in phenotypic expression of these cells under traditional culture conditions. Thus, the aim of the study was to develop a better culture system for the expansion and myo-differentiation of MPCs that could further be used for therapy. For this purpose, we developed an ideal method of tissue decellularization and compared the ability of different matrices to support MPC growth and differentiation. Porcine-derived skeletal muscle and liver and kidney extracellular matrix (ECM) were generated by decellularization methods consisting of distilled water, 0.2 mg/mL DNase, or 5% fetal bovine serum. Acellular matrices were further homogenized, dissolved, and combined with a hyaluronic acid-based hydrogel decorated with heparin (ECM-HA-HP). The cell proliferation and myogenic differentiation capacity of human MPCs were assessed when grown on gel alone, ECM, or each ECM-HA-HP substrate. Human MPC proliferation was significantly enhanced when cultured on the ECM-HA-HP substrates compared to the other substrates tested, with the greatest proliferation on the muscle ECM-HA-HP (mECM-HA-HP) substrate. The number of differentiated myotubes was significantly increased on the mECM-HA-HP substrate compared to the other gel-ECM substrates, as well as the numbers of MPCs expressing specific myogenic cell markers (i.e., myosin, desmin, myoD, and myf5). In conclusion, skeletal mECM-HA-HP as a culture substrate provided an optimal culture microenvironment potentially due to its similarity to the in vivo environment. These data suggest a potential use of skeletal muscle-derived ECM gel for the expansion and differentiation of human MPCs for cell-based therapy for skeletal muscle

Phosphoantigen/IL2 expansion and differentiation of Vγ2Vδ2 T cells increase resistance to tuberculosis in nonhuman primates.

Directory of Open Access Journals (Sweden)

Crystal Y Chen

2013-08-01

Full Text Available Dominant Vγ2Vδ2 T-cell subset exist only in primates, and recognize phosphoantigen from selected pathogens including M. tuberculosis(Mtb. In vivo function of Vγ2Vδ2 T cells in tuberculosis remains unknown. We conducted mechanistic studies to determine whether earlier expansion/differentiation of Vγ2Vδ2 T cells during Mtb infection could increase immune resistance to tuberculosis in macaques. Phosphoantigen/IL-2 administration specifically induced major expansion and pulmonary trafficking/accumulation of phosphoantigen-specific Vγ2Vδ2 T cells, significantly reduced Mtb burdens and attenuated tuberculosis lesions in lung tissues compared to saline/BSA or IL-2 controls. Expanded Vγ2Vδ2 T cells differentiated into multifunctional effector subpopulations capable of producing anti-TB cytokines IFNγ, perforin and granulysin, and co-producing perforin/granulysin in lung tissue. Mechanistically, perforin/granulysin-producing Vγ2Vδ2 T cells limited intracellular Mtb growth, and macaque granulysin had Mtb-bactericidal effect, and inhibited intracellular Mtb in presence of perforin. Furthermore, phosphoantigen/IL2-expanded Vγ2Vδ2 T effector cells produced IL-12, and their expansion/differentiation led to enhanced pulmonary responses of peptide-specific CD4+/CD8+ Th1-like cells. These results provide first in vivo evidence implicating that early expansion/differentiation of Vγ2Vδ2 T effector cells during Mtb infection increases resistance to tuberculosis. Thus, data support a rationale for conducting further studies of the γδ T-cell-targeted treatment of established TB, which might ultimately help explore single or adjunctive phosphoantigen expansion of Vγ2Vδ2 T-cell subset as intervention of MDR-tuberculosis or HIV-related tuberculosis.

Expansion of highly activated invariant natural killer T cells with altered phenotype in acute dengue infection

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Kamaladasa, A.; Wickramasinghe, N.; Adikari, T. N.; Gomes, L.; Shyamali, N. L. A.; Salio, M.; Cerundolo, V.; Ogg, G. S.

2016-01-01

Summary Invariant natural killer T (iNKT) cells are capable of rapid activation and production of cytokines upon recognition of antigenic lipids presented by CD1d molecules. They have been shown to play a significant role in many viral infections and were observed to be highly activated in patients with acute dengue infection. In order to characterize further their role in dengue infection, we investigated the proportion of iNKT cells and their phenotype in adult patients with acute dengue infection. The functionality of iNKT cells in patients was investigated by both interferon (IFN)‐γ and interleukin (IL)−4 ex‐vivo enzyme‐linked immunospot (ELISPOT) assays following stimulation with alpha‐galactosyl‐ceramide (αGalCer). We found that circulating iNKT cell proportions were significantly higher (P = 0·03) in patients with acute dengue when compared to healthy individuals and were predominantly of the CD4+ subset. iNKT cells of patients with acute dengue had reduced proportions expressing CD8α and CD161 when compared to healthy individuals. The iNKT cells of patients were highly activated and iNKT activation correlated significantly with dengue virus‐specific immunoglobulin (Ig)G antibody levels. iNKT cells expressing Bcl‐6 (P = 0·0003) and both Bcl‐6 and inducible T cell co‐stimulator (ICOS) (P = 0·006) were increased significantly in patients when compared to healthy individuals. Therefore, our data suggest that in acute dengue infection there is an expansion of highly activated CD4+ iNKT cells, with reduced expression of CD161 markers. PMID:26874822

Expansion on stromal cells preserves the undifferentiated state of human hematopoietic stem cells despite compromised reconstitution ability.

Science.gov (United States)

Magnusson, Mattias; Sierra, Maria I; Sasidharan, Rajkumar; Prashad, Sacha L; Romero, Melissa; Saarikoski, Pamela; Van Handel, Ben; Huang, Andy; Li, Xinmin; Mikkola, Hanna K A

2013-01-01

Lack of HLA-matched hematopoietic stem cells (HSC) limits the number of patients with life-threatening blood disorders that can be treated by HSC transplantation. So far, insufficient understanding of the regulatory mechanisms governing human HSC has precluded the development of effective protocols for culturing HSC for therapeutic use and molecular studies. We defined a culture system using OP9M2 mesenchymal stem cell (MSC) stroma that protects human hematopoietic stem/progenitor cells (HSPC) from differentiation and apoptosis. In addition, it facilitates a dramatic expansion of multipotent progenitors that retain the immunophenotype (CD34+CD38-CD90+) characteristic of human HSPC and proliferative potential over several weeks in culture. In contrast, transplantable HSC could be maintained, but not significantly expanded, during 2-week culture. Temporal analysis of the transcriptome of the ex vivo expanded CD34+CD38-CD90+ cells documented remarkable stability of most transcriptional regulators known to govern the undifferentiated HSC state. Nevertheless, it revealed dynamic fluctuations in transcriptional programs that associate with HSC behavior and may compromise HSC function, such as dysregulation of PBX1 regulated genetic networks. This culture system serves now as a platform for modeling human multilineage hematopoietic stem/progenitor cell hierarchy and studying the complex regulation of HSC identity and function required for successful ex vivo expansion of transplantable HSC.

Expansion on stromal cells preserves the undifferentiated state of human hematopoietic stem cells despite compromised reconstitution ability.

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Mattias Magnusson

Full Text Available Lack of HLA-matched hematopoietic stem cells (HSC limits the number of patients with life-threatening blood disorders that can be treated by HSC transplantation. So far, insufficient understanding of the regulatory mechanisms governing human HSC has precluded the development of effective protocols for culturing HSC for therapeutic use and molecular studies. We defined a culture system using OP9M2 mesenchymal stem cell (MSC stroma that protects human hematopoietic stem/progenitor cells (HSPC from differentiation and apoptosis. In addition, it facilitates a dramatic expansion of multipotent progenitors that retain the immunophenotype (CD34+CD38-CD90+ characteristic of human HSPC and proliferative potential over several weeks in culture. In contrast, transplantable HSC could be maintained, but not significantly expanded, during 2-week culture. Temporal analysis of the transcriptome of the ex vivo expanded CD34+CD38-CD90+ cells documented remarkable stability of most transcriptional regulators known to govern the undifferentiated HSC state. Nevertheless, it revealed dynamic fluctuations in transcriptional programs that associate with HSC behavior and may compromise HSC function, such as dysregulation of PBX1 regulated genetic networks. This culture system serves now as a platform for modeling human multilineage hematopoietic stem/progenitor cell hierarchy and studying the complex regulation of HSC identity and function required for successful ex vivo expansion of transplantable HSC.

The Glycine-Alanine Dipeptide Repeat from C9orf72 Hexanucleotide Expansions Forms Toxic Amyloids Possessing Cell-to-Cell Transmission Properties.

Science.gov (United States)

Chang, Yu-Jen; Jeng, U-Ser; Chiang, Ya-Ling; Hwang, Ing-Shouh; Chen, Yun-Ru

2016-03-04

Hexanucleotide expansions, GGGGCC, in the non-coding regions of the C9orf72 gene were found in major frontotemporal lobar dementia and amyotrophic lateral sclerosis patients (C9FTD/ALS). In addition to possible RNA toxicity, several dipeptide repeats (DPRs) are translated through repeat-associated non-ATG-initiated translation. The DPRs, including poly(GA), poly(GR), poly(GP), poly(PR), and poly(PA), were found in the brains and spinal cords of C9FTD/ALS patients. Among the DPRs, poly(GA) is highly susceptible to form cytoplasmic inclusions, which is a characteristic of C9FTD/ALS. To elucidate DPR aggregation, we used synthetic (GA)15 DPR as a model system to examine the aggregation and structural properties in vitro. We found that (GA)15 with 15 repeats fibrillates rapidly and ultimately forms flat, ribbon-type fibrils evidenced by transmission electron microscopy and atomic force microscopy. The fibrils are capable of amyloid dye binding and contain a characteristic cross-β sheet structure, as revealed by x-ray scattering. Furthermore, using neuroblastoma cells, we demonstrated the neurotoxicity and cell-to-cell transmission property of (GA)15 DPR. Overall, our results show the structural and toxicity properties of GA DPR to facilitate future DPR-related therapeutic development. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

COBRA, an Arabidopsis extracellular glycosyl-phosphatidyl inositol-anchored protein, specifically controls highly anisotropic expansion through its involvement in cellulose microfibril orientation.

Science.gov (United States)

Roudier, François; Fernandez, Anita G; Fujita, Miki; Himmelspach, Regina; Borner, Georg H H; Schindelman, Gary; Song, Shuang; Baskin, Tobias I; Dupree, Paul; Wasteneys, Geoffrey O; Benfey, Philip N

2005-06-01

The orientation of cell expansion is a process at the heart of plant morphogenesis. Cellulose microfibrils are the primary anisotropic material in the cell wall and thus are likely to be the main determinant of the orientation of cell expansion. COBRA (COB) has been identified previously as a potential regulator of cellulose biogenesis. In this study, characterization of a null allele, cob-4, establishes the key role of COB in controlling anisotropic expansion in most developing organs. Quantitative polarized-light and field-emission scanning electron microscopy reveal that loss of anisotropic expansion in cob mutants is accompanied by disorganization of the orientation of cellulose microfibrils and subsequent reduction of crystalline cellulose. Analyses of the conditional cob-1 allele suggested that COB is primarily implicated in microfibril deposition during rapid elongation. Immunodetection analysis in elongating root cells revealed that, in agreement with its substitution by a glycosylphosphatidylinositol anchor, COB was polarly targeted to both the plasma membrane and the longitudinal cell walls and was distributed in a banding pattern perpendicular to the longitudinal axis via a microtubule-dependent mechanism. Our observations suggest that COB, through its involvement in cellulose microfibril orientation, is an essential factor in highly anisotropic expansion during plant morphogenesis.

A Rapid Embryonic Stem Cell-Based Mouse Model for B-cell Lymphomas Driven by Epstein-Barr Virus Protein LMP1.

Science.gov (United States)

Ba, Zhaoqing; Meng, Fei-Long; Gostissa, Monica; Huang, Pei-Yi; Ke, Qiang; Wang, Zhe; Dao, Mai N; Fujiwara, Yuko; Rajewsky, Klaus; Zhang, Baochun; Alt, Frederick W

2015-06-01

The Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) contributes to oncogenic human B-cell transformation. Mouse B cells conditionally expressing LMP1 are not predisposed to B-cell malignancies, as LMP1-expressing B cells are eliminated by T cells. However, mice with conditional B-cell LMP1 expression and genetic elimination of α/β and γ/δ T cells ("CLT" mice) die early in association with B-cell lymphoproliferation and lymphomagenesis. Generation of CLT mice involves in-breeding multiple independently segregating alleles. Thus, although introduction of additional activating or knockout mutations into the CLT model is desirable for further B-cell expansion and immunosurveillance studies, doing such experiments by germline breeding is time-consuming, expensive, and sometimes unfeasible. To generate a more tractable model, we generated clonal CLT embryonic stem (ES) cells from CLT embryos and injected them into RAG2-deficient blastocysts to generate chimeric mice, which, like germline CLT mice, harbor splenic CLT B cells and lack T cells. CLT chimeric mice generated by this RAG2-deficient blastocyst complementation ("RDBC") approach die rapidly in association with B-cell lymphoproliferation and lymphoma. Because CLT lymphomas routinely express the activation-induced cytidine deaminase (AID) antibody diversifier, we tested potential AID roles by eliminating the AID gene in CLT ES cells and testing them via RDBC. We found that CLT and AID-deficient CLT ES chimeras had indistinguishable phenotypes, showing that AID is not essential for LMP1-induced lymphomagenesis. Beyond expanding accessibility and utility of CLT mice as a cancer immunotherapy model, our studies provide a new approach for facilitating generation of genetically complex mouse cancer models. ©2015 American Association for Cancer Research.

Angiopoietin-like protein 3 promotes preservation of stemness during ex vivo expansion of murine hematopoietic stem cells.

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Farahbakhshian, Elnaz; Verstegen, Monique M; Visser, Trudi P; Kheradmandkia, Sima; Geerts, Dirk; Arshad, Shazia; Riaz, Noveen; Grosveld, Frank; van Til, Niek P; Meijerink, Jules P P

2014-01-01

Allogeneic hematopoietic stem cell (HSC) transplantations from umbilical cord blood or autologous HSCs for gene therapy purposes are hampered by limited number of stem cells. To test the ability to expand HSCs in vitro prior to transplantation, two growth factor cocktails containing stem cell factor, thrombopoietin, fms-related tyrosine kinase-3 ligand (STF) or stem cell factor, thrombopoietin, insulin-like growth factor-2, fibroblast growth factor-1 (STIF) either with or without the addition of angiopoietin-like protein-3 (Angptl3) were used. Culturing HSCs in STF and STIF media for 7 days expanded long-term repopulating stem cells content in vivo by ∼6-fold and ∼10-fold compared to freshly isolated stem cells. Addition of Angptl3 resulted in increased expansion of these populations by ∼17-fold and ∼32-fold, respectively, and was further supported by enforced expression of Angptl3 in HSCs through lentiviral transduction that also promoted HSC expansion. As expansion of highly purified lineage-negative, Sca-1+, c-Kit+ HSCs was less efficient than less pure lineage-negative HSCs, Angptl3 may have a direct effect on HCS but also an indirect effect on accessory cells that support HSC expansion. No evidence for leukemia or toxicity was found during long-term follow up of mice transplanted with ex vivo expanded HSCs or manipulated HSC populations that expressed Angptl3. We conclude that the cytokine combinations used in this study to expand HSCs ex vivo enhances the engraftment in vivo. This has important implications for allogeneic umbilical cord-blood derived HSC transplantations and autologous HSC applications including gene therapy.

Megakaryocytes promote murine osteoblastic HSC niche expansion and stem cell engraftment after radioablative conditioning

OpenAIRE

Olson, Timothy S.; Caselli, Anna; Otsuru, Satoru; Hofmann, Ted J.; Williams, Richard; Paolucci, Paolo; Dominici, Massimo; Horwitz, Edwin M.

2013-01-01

After radioablative conditioning, host megakaryocytes promote endosteal HSC niche expansion and donor stem cell engraftment.Thrombopoietin administration before radiation and bone marrow transplant enhances megakaryocyte promotion of HSC engraftment.

LMP1-mediated glycolysis induces myeloid-derived suppressor cell expansion in nasopharyngeal carcinoma.

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Ting-Ting Cai

2017-07-01

Full Text Available Myeloid-derived suppressor cells (MDSCs are expanded in tumor microenvironments, including that of Epstein-Barr virus (EBV-associated nasopharyngeal carcinoma (NPC. The link between MDSC expansion and EBV infection in NPC is unclear. Here, we show that EBV latent membrane protein 1 (LMP1 promotes MDSC expansion in the tumor microenvironment by promoting extra-mitochondrial glycolysis in malignant cells, which is a scenario for immune escape initially suggested by the frequent, concomitant detection of abundant LMP1, glucose transporter 1 (GLUT1 and CD33+ MDSCs in tumor sections. The full process has been reconstituted in vitro. LMP1 promotes the expression of multiple glycolytic genes, including GLUT1. This metabolic reprogramming results in increased expression of the Nod-like receptor family protein 3 (NLRP3 inflammasome, COX-2 and P-p65 and, consequently, increased production of IL-1β, IL-6 and GM-CSF. Finally, these changes in the environment of malignant cells result in enhanced NPC-derived MDSC induction. One key step is the physical interaction of LMP1 with GLUT1 to stabilize the GLUT1 protein by blocking its K48-ubiquitination and p62-dependent autolysosomal degradation. This work indicates that LMP1-mediated glycolysis regulates IL-1β, IL-6 and GM-CSF production through the NLRP3 inflammasome, COX-2 and P-p65 signaling pathways to enhance tumor-associated MDSC expansion, which leads to tumor immunosuppression in NPC.

Gelatin–PMVE/MA composite scaffold promotes expansion of embryonic stem cells

International Nuclear Information System (INIS)

Chhabra, Hemlata; Gupta, Priyanka; Verma, Paul J.; Jadhav, Sameer; Bellare, Jayesh R.

2014-01-01

We introduce a new composite scaffold of gelatin and polymethyl vinyl ether-alt-maleic anhydride (PMVE/MA) for expansion of embryonic stem cells (ESCs) in an in vitro environment. To optimize the scaffold, we prepared a gelatin scaffold (G) and three composite scaffolds namely GP-1, GP-2, and GP-3 with varying PMVE/MA concentrations (0.2–1%) and characterized them by scanning electron microscopy (SEM), swelling study, compression testing and FTIR. SEM micrographs revealed interconnected porous structure in all the scaffolds. The permissible hemolysis ratio and activation of platelets by scaffolds confirmed the hemocompatibility of scaffolds. Initial biocompatibility assessment of scaffolds was conducted using hepatocarcinoma (Hep G2) cells and adhesion, proliferation and infiltration of Hep G2 cells in depth of scaffolds were observed, proving the scaffold's biocompatibility. Further Oct4B2 mouse embryonic stem cells (mESCs), which harbor a green fluorescence protein transgene under regulatory control of the Oct4 promotor, were examined for expansion on scaffolds with MTT assay. The GP-2 scaffold demonstrated the best cell proliferation and was further explored for ESC adherence and infiltration in depth (SEM and confocal), and pluripotent state of mESCs was assessed with the expression of Oct4-GFP and stage-specific embryonic antigen-1 (SSEA-1). This study reports the first demonstration of biocompatibility of gelatin–PMVE/MA composite scaffold and presents this scaffold as a promising candidate for embryonic stem cell based tissue engineering. - Highlights: • Composite scaffolds of gelatin and PMVE/MA were prepared by freeze-drying method. • SEM micrographs showed porous structure in all scaffolds of varying pore dimension. • GP-2 composite exhibited better cellular response in comparison to other scaffolds. • mESCs proliferated and expressed Oct-4 and SSEA-1, when cultured on GP-2 scaffold

Gelatin–PMVE/MA composite scaffold promotes expansion of embryonic stem cells

Energy Technology Data Exchange (ETDEWEB)

Chhabra, Hemlata [Department of Chemical Engineering, Indian Institute of Technology Bombay, Powai, Mumbai (India); Gupta, Priyanka [Department of Chemical Engineering, Indian Institute of Technology Bombay, Powai, Mumbai (India); IITB-Monash Research Academy, Mumbai (India); Department of Chemical Engineering, Monash University, Melbourne (Australia); Verma, Paul J. [Turretfield Research Centre, South Australian Research and Development Institute, Rosedale, South Australia (Australia); Jadhav, Sameer; Bellare, Jayesh R. [Department of Chemical Engineering, Indian Institute of Technology Bombay, Powai, Mumbai (India)

2014-04-01

We introduce a new composite scaffold of gelatin and polymethyl vinyl ether-alt-maleic anhydride (PMVE/MA) for expansion of embryonic stem cells (ESCs) in an in vitro environment. To optimize the scaffold, we prepared a gelatin scaffold (G) and three composite scaffolds namely GP-1, GP-2, and GP-3 with varying PMVE/MA concentrations (0.2–1%) and characterized them by scanning electron microscopy (SEM), swelling study, compression testing and FTIR. SEM micrographs revealed interconnected porous structure in all the scaffolds. The permissible hemolysis ratio and activation of platelets by scaffolds confirmed the hemocompatibility of scaffolds. Initial biocompatibility assessment of scaffolds was conducted using hepatocarcinoma (Hep G2) cells and adhesion, proliferation and infiltration of Hep G2 cells in depth of scaffolds were observed, proving the scaffold's biocompatibility. Further Oct4B2 mouse embryonic stem cells (mESCs), which harbor a green fluorescence protein transgene under regulatory control of the Oct4 promotor, were examined for expansion on scaffolds with MTT assay. The GP-2 scaffold demonstrated the best cell proliferation and was further explored for ESC adherence and infiltration in depth (SEM and confocal), and pluripotent state of mESCs was assessed with the expression of Oct4-GFP and stage-specific embryonic antigen-1 (SSEA-1). This study reports the first demonstration of biocompatibility of gelatin–PMVE/MA composite scaffold and presents this scaffold as a promising candidate for embryonic stem cell based tissue engineering. - Highlights: • Composite scaffolds of gelatin and PMVE/MA were prepared by freeze-drying method. • SEM micrographs showed porous structure in all scaffolds of varying pore dimension. • GP-2 composite exhibited better cellular response in comparison to other scaffolds. • mESCs proliferated and expressed Oct-4 and SSEA-1, when cultured on GP-2 scaffold.

Digital radiographic evaluation of the midpalatal suture in patients submitted to rapid maxillary expansion

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Maria de Fátima Batista de Melo

2013-01-01

Full Text Available Aims: To analyze the density of the midpalatal suture by means of digital radiographs three months after retention to evaluate if this period of retention is really sufficient for bone repair. Materials and Methods: This prospective study consisted of 31 patients (11 girls and 20 boys in the mixed or permanent dentition stage, treated using a tooth-tissue borne expanders (Haas. Occlusal digital radiographs were taken at three stages: prior to rapid maxillary expansion (Stage I; immediately after desired maxillary expansion (Stage II; and after three months of retention (Stage III. Radiographs were taken on a dental X-ray machine, set at 70 kVp and 7 mA with an exposure time of 0.04 s. A phosphor storage plate system, imaging plate size n. 2 (35 × 45 × 1.6 mm, was used. Three regions (A, B and C measured 0.02 mm² were selected for optical density analysis. The difference between the measurements was evaluated with the paired t-test. Results: The optical density was reduced at Stages II and III compared with Stage I. Between-stage comparison showed statistically significant changes for all variables (P < 0.05, with the highest mean optical density at Stage I and the lowest at Stage II, in all groups. Actually, there was an increase in optical density between Stages II and III, but they are reduced compared with Stage I. Conclusion: The results strongly suggest that bone formation did not occur as expected, and that a longer retention period for bone repair may be necessary.

An organ-specific role for ethylene in rose petal expansion during dehydration and rehydration

Science.gov (United States)

Liu, Daofeng; Liu, Xiaojing; Meng, Yonglu; Sun, Cuihui; Tang, Hongshu; Jiang, Yudong; Khan, Muhammad Ali; Xue, Jingqi; Ma, Nan; Gao, Junping

2013-01-01

Dehydration is a major factor resulting in huge loss from cut flowers during transportation. In the present study, dehydration inhibited petal cell expansion and resulted in irregular flowers in cut roses, mimicking ethylene-treated flowers. Among the five floral organs, dehydration substantially elevated ethylene production in the sepals, whilst rehydration caused rapid and elevated ethylene levels in the gynoecia and sepals. Among the five ethylene biosynthetic enzyme genes (RhACS1–5), expression of RhACS1 and RhACS2 was induced by dehydration and rehydration in the two floral organs. Silencing both RhACS1 and RhACS2 significantly suppressed dehydration- and rehydration-induced ethylene in the sepals and gynoecia. This weakened the inhibitory effect of dehydration on petal cell expansion. β-glucuronidase activity driven by both the RhACS1 and RhACS2 promoters was dramatically induced in the sepals, pistil, and stamens, but not in the petals of transgenic Arabidopsis. This further supports the organ-specific induction of these two genes. Among the five rose ethylene receptor genes (RhETR1–5), expression of RhETR3 was predominantly induced by dehydration and rehydration in the petals. RhETR3 silencing clearly aggravated the inhibitory effect of dehydration on petal cell expansion. However, no significant difference in the effect between RhETR3-silenced flowers and RhETR-genes-silenced flowers was observed. Furthermore, RhETR-genes silencing extensively altered the expression of 21 cell expansion-related downstream genes in response to ethylene. These results suggest that induction of ethylene biosynthesis by dehydration proceeds in an organ-specific manner, indicating that ethylene can function as a mediator in dehydration-caused inhibition of cell expansion in rose petals. PMID:23599274

Controlled auto-ignition characteristics of methane-air mixture in a rapid intake compression and expansion machine

Energy Technology Data Exchange (ETDEWEB)

Cho, Gyubaek; Jeong, Dongsoo [Engine Research Team, Eco-Machinery Research Division, Korea Institute of Machinery and Materials, 104 Sinseongno, Yuseong-gu, Daejeon 305-701 (Korea); Moon, Gunfeel [Department of Clean Environmental system, University of Science and Technology, 52 Eoeun-dong, Yuseong-gu, Daejeon (Korea); Bae, Choongsik [Engine Laboratory, Department of Mechanical Engineering, Korea Advanced Institute of Science and Technology, 373-1 GuSeong-Dong, Yuseong-Gu, Daejeon 305-701 (Korea)

2010-10-15

The characteristics of controlled auto-ignition (CAI) were investigated with a methane-air mixture and simulated residual gas, that represents internal exhaust gas recirculation (IEGR). Supply systems were additionally installed on the conventional rapid compression machine (RCM), and this modified machine - a rapid intake compression and expansion machine (RICEM) - was able to simulate an intake stroke for the evaluation of controlled auto-ignition with fuel-air mixture. The fuel-air mixture and the simulated residual gas were introduced separately into the combustion chamber through the spool valves. Various IEGR rates and temperatures of the IEGR gas were tested. The initial reaction and the development in controlled auto-ignition combustion were compared with spark-ignited combustion by visualization with a high-speed digital camera. Under the controlled auto-ignition operation, multi-point ignition and faster combustion were observed. With increasing the temperature of IEGR gas, the auto-ignition timing was advanced and burning duration was shortened. The higher rate of IEGR had the same effects on the combustion of the controlled auto-ignition. However, this trend was reversed with more than 47 per cent of IEGR. (author)

Evaluation of GMP-compliant culture media for in vitro expansion of human bone marrow mesenchymal stromal cells.

Science.gov (United States)

Wuchter, Patrick; Vetter, Marcel; Saffrich, Rainer; Diehlmann, Anke; Bieback, Karen; Ho, Anthony D; Horn, Patrick

2016-06-01

Mesenchymal stromal cells (MSCs) from human bone marrow serve as a resource for cell-based therapies in regenerative medicine. Clinical applications require standardized protocols according to good manufacturing practice (GMP) guidelines. Donor variability as well as the intrinsic heterogeneity of MSC populations must be taken into consideration. The composition of the culture medium is a key factor in successful MSC expansion. The aim of this study was to comparatively assess the efficiency of xeno-free human platelet lysate (HPL)-based cell expansion with two commercially available media-StemPro MSC SFM CTS (for human ex vivo tissue and cell culture processing applications) and MSCGM (non-GMP-compliant, for research only)-in an academic setting as the first optimization step toward GMP-compliant manufacturing. We report the feasibility of MSC expansion up to the yielded cell number with all three media. MSCs exhibited the typical fibroblastoid morphology, with distinct differences in cell size depending on the medium. The differentiation capacity and characteristic immunophenotype were confirmed for all MSC populations. Proliferation was highest using StemPro MSC SFM CTS, whereas HPL medium was more cost-effective and its composition could be adjusted individually according to the respective needs. In summary, we present a comprehensive evaluation of GMP-compatible culture media for MSC expansion. Both StemPro and HPL medium proved to be suitable for clinical application and allowed sufficient cell proliferation. Specific differences were observed and should be considered according to the intended use. This study provides a detailed cost analysis and tools that may be helpful for the establishment of GMP-compliant MSC expansion. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

Expansion of Human Tregs from Cryopreserved Umbilical Cord Blood for GMP-Compliant Autologous Adoptive Cell Transfer Therapy

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Howard R. Seay

2017-03-01

Full Text Available Umbilical cord blood is a traditional and convenient source of cells for hematopoietic stem cell transplantation. Thymic regulatory T cells (Tregs are also present in cord blood, and there is growing interest in the use of autologous Tregs to provide a low-risk, fully human leukocyte antigen (HLA-matched cell product for treating autoimmune diseases, such as type 1 diabetes. Here, we describe a good manufacturing practice (GMP-compatible Treg expansion protocol using fluorescence-activated cell sorting, resulting in a mean 2,092-fold expansion of Tregs over a 16-day culture for a median yield of 1.26 × 109 Tregs from single-donor cryopreserved units. The resulting Tregs passed prior clinical trial release criteria for Treg purity and sterility, including additional rigorous assessments of FOXP3 and Helios expression and epigenetic analysis of the FOXP3 Treg-specific demethylated region (TSDR. Compared with expanded adult peripheral blood Tregs, expanded cord blood Tregs remained more naive, as assessed by continued expression of CD45RA, produced reduced IFN-γ following activation, and effectively inhibited responder T cell proliferation. Immunosequencing of the T cell receptor revealed a remarkably diverse receptor repertoire within cord blood Tregs that was maintained following in vitro expansion. These data support the feasibility of generating GMP-compliant Tregs from cord blood for adoptive cell transfer therapies and highlight potential advantages in terms of safety, phenotypic stability, autoantigen specificity, and tissue distribution.

Plaque index and gingival index during rapid maxillary expansion of patients with unilateral cleft lip and palate

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Maria Olívia Rocha

Full Text Available ABSTRACT Objective: To assess, during rapid maxillary expansion, the plaque index (PI and the gingival index (GI of patients with unilateral cleft lip and palate(UCLP using Hyrax (HX or inverted mini-Hyrax (IMHX rapid maxillary expanders (RME considering patients’ sex and age. Methods: PI (Quigley Index modified by Turesky et al and GI (Löe and Silness of 28 UCLP (11 females; 17 males: aged 8 to 15 years submitted to daily RME activation were assessed before (T0 and 7 (T1, 28 (T2 and 90 (T3 days after activation. Log-linear models and Bonferroni correction were performed to analyze possible differences in PI and GI between RME, sexes or age groups over time. Results: Intra-group comparison revealed significant increases in PI of patients using HX (T0 < T2, IMHX (T0 < T3; T1< T3, males (T0 < T1; T0 < T2; T0 < T3 or aged 12-15 years (T0 < T1; T0 < T2; T0 < T3, and in GI of patients using IMHX (T0 < T3; T1 < T3, females (T1 < T3; T2 < T3 or aged 12-15 years (T0 < T3; T2 < T3. One inter-group difference in GI according to patients’ age (8-11 < 12-15; T1 was observed. Conclusions: Since a single difference between groups was encountered, the results of this study indicated that PI and GI during maxillary expansion were similar between HX and IMHX, sexes and the analyzed age groups. Therefore, orthodontists can use these RME in UCLP patients according to the patient’s necessity or their preferences.

Effect of the laser and light-emitting diode (LED) phototherapy on midpalatal suture bone formation after rapid maxilla expansion: a Raman spectroscopy analysis.

Science.gov (United States)

Rosa, Cristiane Becher; Habib, Fernando Antonio Lima; de Araújo, Telma Martins; Aragão, Juliana Silveira; Gomes, Rafael Soares; Barbosa, Artur Felipe Santos; Silveira, Landulfo; Pinheiro, Antonio L B

2014-05-01

The aim of this study was to analyze the effect of laser or light-emitting diode (LED) phototherapy on the bone formation at the midpalatal suture after rapid maxilla expansion. Twenty young adult male rats were divided into four groups with 8 days of experimental time: group 1, no treatment; group 2, expansion; group 3, expansion and laser irradiation; and group 4, expansion and LED irradiation. In groups 3 and 4, light irradiation was in the first, third, and fifth experimental days. In all groups, the expansion was accomplished with a helicoid 0.020" stainless steel orthodontic spring. A diode laser (λ780 nm, 70 mW, spot of 0.04 cm(2), t = 257 s, spatial average energy fluence (SAEF) of 18 J/cm(2)) or a LED (λ850 nm, 150 mW ± 10 mW, spot of 0.5 cm(2), t = 120 s, SAEF of 18 J/cm(2)) were used. The samples were analyzed by Raman spectroscopy carried out at midpalatal suture and at the cortical area close to the suture. Two Raman shifts were analyzed: ∼ 960 (phosphate hydroxyapatite) and ∼ 1,450 cm(-1) (lipids and protein). Data was submitted to statistical analysis. Significant statistical difference (p ≤ 0.05) was found in the hydroxyapatite (CHA) peaks among the expansion group and the expansion and laser or LED groups. The LED group presented higher mean peak values of CHA. No statistical differences were found between the treated groups as for collagen deposition, although LED also presented higher mean peak values. The results of this study using Raman spectral analysis indicate that laser and LED light irradiation improves deposition of CHA in the midpalatal suture after orthopedic expansion.

Phloem unloading and cell expansion in pea stems

International Nuclear Information System (INIS)

Schmalstig, J.G.; Cosgrove, D.J.

1989-01-01

Phloem unloading into elongating stems of dark-grown pea seedlings was greater in regions with higher relative growth rates. Phloem transport was monitored over 1 h by measuring accumulation of radiolabel from 14 C-sucrose added between the cotyledons. The apical hook and plumule and 8 mm of the growing region of an intact plant were sealed in a pressure chamber and the pressure was raised to stop elongation. Phloem unloading was inhibited in the pressurized zone of elongation and accelerated in the apical hook and plumule, with the result that the magnitude of phloem transport into the stem was unchanged. The results demonstrate a coupling between cell expansion and phloem unloading

Chemical graph-theoretic cluster expansions

International Nuclear Information System (INIS)

Klein, D.J.

1986-01-01

A general computationally amenable chemico-graph-theoretic cluster expansion method is suggested as a paradigm for incorporation of chemical structure concepts in a systematic manner. The cluster expansion approach is presented in a formalism general enough to cover a variety of empirical, semiempirical, and even ab initio applications. Formally such approaches for the utilization of chemical structure-related concepts may be viewed as discrete analogues of Taylor series expansions. The efficacy of the chemical structure concepts then is simply bound up in the rate of convergence of the cluster expansions. In many empirical applications, e.g., boiling points, chromatographic separation coefficients, and biological activities, this rate of convergence has been observed to be quite rapid. More note will be made here of quantum chemical applications. Relations to questions concerning size extensivity of energies and size consistency of wave functions are addressed

Rapid kinetics of lysis in human natural cell-mediated cytotoxicity: some implications

International Nuclear Information System (INIS)

Bloom, E.T.; Babbitt, J.T.

1983-01-01

The entire lytic process of natural cell-mediated cytotoxicity against sensitive target cells can occur rapidly, within minutes. This was demonstrated by 51 chromium release and in single-cell assays. At the cellular level, most of the target cell lysis occurred within 15-30 min after binding to effector cells. The enriched natural killer cell subpopulation of lymphocytes obtained by Percoll density gradient centrifugation (containing greater than 70% large granular lymphocytes (LGL)) was the most rapidly lytic population by 51 chromium release. However, in the single-cell assay, the rate of lysis of bound target cells was quite similar for the LGL-enriched effector subpopulation and the higher density subpopulation of effector cells recognized previously. Both the light and dense effector cells contained similar numbers of target binding cells. Therefore, that the light subpopulation effected lysis more rapidly and to a greater extent than the dense subpopulation suggested that the low-density effector cells probably recycled more rapidly than those of higher density. This was corroborated by the finding that when conjugates were formed at 29 degrees C for the single-cell assay, a significant number of dead unconjugated targets could be observed only on the slides made with the LGL-enriched effector cells but not on those made with dense effector cell. Lysis continued to increase in the chromium-release assay probably because of recycling, recruitment, and/or heterogeneity of the effector cells, and/or because of heterogeneity or delayed death of the target cells

Mandibular dental arch short and long-term spontaneous dentoalveolar changes after slow or rapid maxillary expansion: a systematic review

Directory of Open Access Journals (Sweden)

Arthur César de Medeiros Alves

Full Text Available ABSTRACT Objective: The aim of this systematic review was to analyze the short and long-term spontaneous dentoalveolar changes of the mandibular dental arch after slow (SME or rapid (RME maxillary expansion in the mixed and early permanent dentitions. Methods: An electronic search was performed in the following databases: PubMed/Medline, Cochrane Library, Scopus, Embase and Web of Science. Eligibility criteria for article selection included randomized controlled trials and prospective studies written in English, with no restriction of year of publication, involving patients who underwent SME or RME during the mixed or early permanent dentitions. A double-blind search of articles was performed by two reviewers. Initially, the title and the abstract of the studies were read, and their references were also hand-searched for possible missing studies. A methodological quality scoring scale was used to analyze the selected articles. Results: The search retrieved 373 articles, but only 6 were selected for review after application of the eligibility and exclusion criteria. Non-clinically significant spontaneous dentoalveolar changes of approximately 1mm were found in the mandibular dental arch in the short and long-term, after slow or rapid maxillary expansions. Furthermore, no significant differences were found between treated and control groups. Conclusions: There is enough evidence to conclude that negligible short and long-term spontaneous dentoalveolar changes tend to occur in the mandibular dental arch after SME or RME in the mixed and early permanent dentitions. More randomized studies with appropriate control group are required to better evaluate this issue.

An improved out-cell to in-cell rapid transfer system at the HFEF/South

International Nuclear Information System (INIS)

Bacca, J.P.; Sherman, E.K.

1991-01-01

This paper reports on Argonne National Laboratory's Fuel Cycle Facility (FCF) (formerly named Hot Fuel Examination Facility-South) (HFEF/South) which is currently being refurbished and upgraded in preparation for demonstrating remote, fast reactor metal-fuel reprocessing and refabrication, as part of the Integral Fast Reactor (IFR) Program. Among the FCF hot-cell system upgrades being provided is a newly fabricated, direct, out-of-cell to in-cell, small-item transfer system for the FCF argon cell. This system will enable the rapid transfer of selected small items from the hot cell exterior into the argon cell (argon-gas atmosphere) of the facility, without necessitating the use of formerly employed, very time-consuming, and quite laborious procedures. The new system will be especially valuable for the rapid insertion of IFR fuel processing makeup materials and small tools into the argon cell, and for use in argon cell and overall FCF radioactive contamination-control activities

An improved out-cell to in-cell rapid transfer system at the HFEF-south

International Nuclear Information System (INIS)

Bacca, J.P.; Sherman, E.K.

1990-01-01

The Argonne National Laboratory (ANL) Hot Fuel Examination Facility-South (HFEF-S), located at the ANL-West site of the Idaho National Engineering Laboratory, is currently undergoing extensive refurbishment and modifications in preparation for its use, beginning in 1991, in demonstrating remote recycling of fast reactor, metal-alloy fuel as part of the US Department of Energy liquid-metal reactor, Integral Fast Reactor (IFR) program. Included in these improvements to HFEF-S is a new, small-item, rapid transfer system (RTS). When installed, this system will enable the rapid transfer of small items from the hot-cell exterior into the argon cell (argon-gas atmosphere) of the facility without necessitating the use of time-consuming and laborious procedures. The new RTS will also provide another important function associated with HFEF-S hot-cell operation in the IFR Fuel Recycle Program; namely, the rapid insertion of clean, radioactive contamination-measuring smear paper specimens into the hot cells for area surveys, and the expedited removal of these contaminated (including alpha as well as beta/gamma contamination) smears from the argon cell for transfer to an adjacent health physics field laboratory in the facility for nuclear contamination/radiation counting

Nras Overexpression Results in Granulocytosis, T-Cell Expansion and Early Lethality in Mice

DEFF Research Database (Denmark)

Lassen, Louise Berkhoudt; Gonzalez, Borja Ballarin; Schmitz, Alexander

2012-01-01

NRAS is a proto-oncogene involved in numerous myeloid malignancies. Here, we report on a mouse line bearing a single retroviral long terminal repeat inserted into Nras. This genetic modification resulted in an increased level of wild type Nras mRNA giving the possibility of studying the function ...... the increment in immature myeloid cells detected in these mice. The short latency period indicates that Nras overexpression alone is sufficient to cause dose-dependent granulocytosis and T-cell expansion....

The Tec kinase ITK regulates thymic expansion, emigration, and maturation of γδ NKT cells.

Science.gov (United States)

Yin, Catherine C; Cho, Ok Hyun; Sylvia, Katelyn E; Narayan, Kavitha; Prince, Amanda L; Evans, John W; Kang, Joonsoo; Berg, Leslie J

2013-03-15

The Tec family tyrosine kinase, Itk, regulates signaling downstream of the TCR. The absence of Itk in CD4(+) T cells results in impaired Th2 responses along with defects in maturation, cytokine production, and survival of iNKT cells. Paradoxically, Itk(-/-) mice have spontaneously elevated serum IgE levels, resulting from an expansion of the Vγ1.1(+)Vδ6.3(+) subset of γδ T cells, known as γδ NKT cells. Comparisons between γδ NKT cells and αβ iNKT cells showed convergence in the pattern of cell surface marker expression, cytokine profiles, and gene expression, suggesting that these two subsets of NKT cells undergo similar differentiation programs. Hepatic γδ NKT cells have an invariant TCR and are derived predominantly from fetal progenitors that expand in the thymus during the first weeks of life. The adult thymus contains these invariant γδ NKT cells plus a heterogeneous population of Vγ1.1(+)Vδ6.3(+) T cells with diverse CDR3 sequences. This latter population, normally excluded from the liver, escapes the thymus and homes to the liver when Itk is absent. In addition, Itk(-/-) γδ NKT cells persistently express high levels of Zbtb16 (PLZF) and Il4, genes that are normally downregulated in the most mature subsets of NKT cells. These data indicate that Itk signaling is required to prevent the expansion of γδ NKT cells in the adult thymus, to block their emigration, and to promote terminal NKT cell maturation.

3D printing human induced pluripotent stem cells with novel hydroxypropyl chitin bioink: scalable expansion and uniform aggregation.

Science.gov (United States)

Li, Yang; Jiang, Xulin; Li, Ling; Chen, Zhi-Nan; Gao, Ge; Yao, Rui; Sun, Wei

2018-06-28

Human induced pluripotent stem cells (hiPSCs) are more likely to successfully avoid the immunological rejection and ethical problems that are often encountered by human embryonic stem cells in various stem cell studies and applications. To transfer hiPSCs from the laboratory to clinical applications, researchers must obtain sufficient cell numbers. In this study, 3D cell printing was used as a novel method for iPSC scalable expansion. Hydroxypropyl chitin (HPCH), utilized as a new type of bioink, and a set of optimized printing parameters were shown to achieve high cell survival (> 90%) after the printing process and high proliferation efficiency (~ 32.3 folds) during subsequent 10-day culture. After the culture, high levels of pluripotency maintenance were recognized by both qualitative and quantitative detections. Compared with static suspension (SS) culture, hiPSC aggregates formed in 3D printed constructs showed a higher uniformity in size. Using novel dual-fluorescent labelling method, hiPSC aggregates in the constructs were found more inclined to form by in situ proliferation rather than multicellular aggregation. This study revealed unique advantages of non-ionic crosslinking bioink material HPCH, including high gel strength and rapid temperature response in hiPSC printing, and achieved primed state hiPSC printing for the first time. Features achieved in this study, such as high cell yield, high pluripotency maintenance and uniform aggregation provide good foundations for further hiPSC studies on 3D micro-tissue differentiation and drug screening. © 2018 IOP Publishing Ltd.

Rapid-Cycling Bubble-Chamber, details

CERN Multimedia

CERN PhotoLab

1980-01-01

Parts of the hydraulic expansion system of the Rapid-Cycling Bubble-Chamber (RCBC). RCBC was the largest of 3 rapid-cycling bubble-chambers (the others were LEBC and HOLEBC), used as target- and vertex-detectors within the European Hybrid Spectrometer (EHS) in the SPS North Area (EHN1). RCBC contained 250 l of liquid hydrogen and was located inside a 3 T superconducting magnet. It was designed for 30 expansions/s (100 times faster than BEBC), the system shown here allowed 50 expansions/s. RCBC operated from 1981 to 1983 for experiments NA21, NA22 and NA23 at a rate of 15 expansions/s, clocking up a total of over 4 million. In the rear, at left, is bearded Lucien Veillet; Augustin Didona is at the right. See also 8001009. The installation of the piston assembly in the RCBC chamber body is shown in the Annual Report 1980, p.65.

Rapid cell separation with minimal manipulation for autologous cell therapies

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Smith, Alban J.; O'Rorke, Richard D.; Kale, Akshay; Rimsa, Roberts; Tomlinson, Matthew J.; Kirkham, Jennifer; Davies, A. Giles; Wälti, Christoph; Wood, Christopher D.

2017-02-01

The ability to isolate specific, viable cell populations from mixed ensembles with minimal manipulation and within intra-operative time would provide significant advantages for autologous, cell-based therapies in regenerative medicine. Current cell-enrichment technologies are either slow, lack specificity and/or require labelling. Thus a rapid, label-free separation technology that does not affect cell functionality, viability or phenotype is highly desirable. Here, we demonstrate separation of viable from non-viable human stromal cells using remote dielectrophoresis, in which an electric field is coupled into a microfluidic channel using shear-horizontal surface acoustic waves, producing an array of virtual electrodes within the channel. This allows high-throughput dielectrophoretic cell separation in high conductivity, physiological-like fluids, overcoming the limitations of conventional dielectrophoresis. We demonstrate viable/non-viable separation efficacy of >98% in pre-purified mesenchymal stromal cells, extracted from human dental pulp, with no adverse effects on cell viability, or on their subsequent osteogenic capabilities.

Recombinant TAT-BMI-1 fusion protein induces ex vivo expansion of human umbilical cord blood-derived hematopoietic stem cells.

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Codispoti, Bruna; Rinaldo, Nicola; Chiarella, Emanuela; Lupia, Michela; Spoleti, Cristina Barbara; Marafioti, Maria Grazia; Aloisio, Annamaria; Scicchitano, Stefania; Giordano, Marco; Nappo, Giovanna; Lucchino, Valeria; Moore, Malcolm A S; Zhou, Pengbo; Mesuraca, Maria; Bond, Heather Mandy; Morrone, Giovanni

2017-07-04

Transplantation of hematopoietic stem cells (HSCs) is a well-established therapeutic approach for numerous disorders. HSCs are typically derived from bone marrow or peripheral blood after cytokine-induced mobilization. Umbilical cord blood (CB) represents an appealing alternative HSC source, but the small amounts of the individual CB units have limited its applications. The availability of strategies for safe ex vivo expansion of CB-derived HSCs (CB-HSCs) may allow to extend the use of these cells in adult patients and to avoid the risk of insufficient engraftment or delayed hematopoietic recovery.Here we describe a system for the ex vivo expansion of CB-HSCs based on their transient exposure to a recombinant TAT-BMI-1 chimeric protein. BMI-1 belongs to the Polycomb family of epigenetic modifiers and is recognized as a central regulator of HSC self-renewal. Recombinant TAT-BMI-1 produced in bacteria was able to enter the target cells via the HIV TAT-derived protein transduction peptide covalently attached to BMI-1, and conserved its biological activity. Treatment of CB-CD34+ cells for 3 days with repeated addition of 10 nM purified TAT-BMI-1 significantly enhanced total cell expansion as well as that of primitive hematopoietic progenitors in culture. Importantly, TAT-BMI-1-treated CB-CD34+ cells displayed a consistently higher rate of multi-lineage long-term repopulating activity in primary and secondary xenotransplants in immunocompromised mice. Thus, recombinant TAT-BMI-1 may represent a novel, effective reagent for ex vivo expansion of CB-HSC for therapeutic purposes.

Downregulation of IL-17-producing T cells is associated with regulatory T cell expansion and disease progression in chronic lymphocytic leukemia.

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Jadidi-Niaragh, Farhad; Ghalamfarsa, Ghasem; Memarian, Ali; Asgarian-Omran, Hossein; Razavi, Seyed Mohsen; Sarrafnejad, Abdolfattah; Shokri, Fazel

2013-04-01

Little is known about the immunobiology of interleukin-17 (IL-17)-producing T cells and regulatory T cells (Treg) in chronic lymphocytic leukemia (CLL). In this study, the frequencies of Th17, Tc17, and CD39(+) Treg cells were enumerated in peripheral T cells isolated from 40 CLL patients and 15 normal subjects by flow cytometry. Our results showed a lower frequency of Th17 and Tc17 cells in progressive (0.99 ± 0.12 % of total CD3(+)CD4(+) cells; 0.44 ± 0.09 % of total CD8(+) cells) compared to indolent patients (1.57 ± 0.24 %, p = 0.042; 0.82 ± 0.2 %, p = 0.09) and normal subjects (1.78 ± 0.2 %, p = 0.003; 0.71 ± 0.09 %, p = 0.04). Decrease in IL-17-producing T cells was associated with CD39(+) Treg cells expansion. Variation of IL-17-producing cells and Treg cells in indolent and progressive patients was neither associated to the expression levels of Th1- and Th2-specific transcription factors T-bet and GATA-3 nor to the frequencies of IFN-γ and IL-4-producing CD4(+) T cells in a selected number of samples. Additionally, suppressive potential of CD4(+) Treg was similar in CLL patients and normal subjects. Our data indicate that progression of CLL is associated with downregulation of IL-17-producing T cells and expansion of Treg cells, implying contribution of these subsets of T cells in the progression of CLL.

Clinical grade purification and expansion of NK cell products for an optimized manufacturing protocol

Directory of Open Access Journals (Sweden)

Ulrike eKoehl

2013-05-01

Full Text Available Allogeneic Natural Killer (NK cells are used for adoptive immunotherapy after stem cell transplantation. In order to overcome technical limitations in NK cell purification and activation, the following study investigates the impact of different variables on NK cell recovery, cytotoxicity and T cell depletion during GMP-grade NK cell selection. 40 NK cell products were derived from 54 unstimulated donor leukaphereses using immunomagnetic CD3 T-cell depletion, followed by a CD56 cell enrichment step. For T cell depletion, either the depletion 2.1 program in single or double procedure (D2.1 1depl, n=18; D2.1 2depl, n=13 or the faster depletion 3.1 (D3.1, n=9 was used on the CliniMACS instrument. 17 purified NK cell products were activated in vitro by IL-2 for 12 days. The whole process resulted in a median number of 7.59x10e8 CD56+CD3- cells with both purity and viability of 94%, respectively. The T-cell depletion was significantly better using D2.1 1depl/2depl compared to D3.1 (log 4.6/log 4.9 vs. log 3.7; p<0.01 and double procedure in two stages led always to residual T cells below 0.1%. In contrast D3.1 was superior to D2.1 1depl/2depl with regard to recovery of CD56+CD3- NK cells (68% vs 41%/38%. Concomitant monocytes and especially IL-2 activation led to increased NK cell activity against malignant target cells compared to unstimulated NK cells, which correlated with both up-regulation of natural cytotoxicity receptors and intracellular signaling. Overall, wide variations in the NK cell expansion rate and the distribution of NK cell subpopulations were found. In conclusion, our results indicate that GMP-grade purification of NK cells might be improved by a sequential processing of T cell depletion program D2.1 and D3.1. In addition NK cell expansion protocols need to be further optimized.

An increase in immature β-cells lacking Glut2 precedes the expansion of β-cell mass in the pregnant mouse.

Directory of Open Access Journals (Sweden)

Christine A Beamish

Full Text Available A compensatory increase in β-cell mass occurs during pregnancy to counter the associated insulin resistance, and a failure in adaptation is thought to contribute to gestational diabetes. Insulin-expressing but glucose-transporter-2-low (Ins+Glut2LO progenitor cells are present in mouse and human pancreas, being predominantly located in extra-islet β-cell clusters, and contribute to the regeneration of the endocrine pancreas following induced ablation. We therefore sought to investigate the contribution of Ins+Glut2LO cells to β-cell mass expansion during pregnancy. Female C57Bl/6 mice were time mated and pancreata were collected at gestational days (GD 6, 9, 12, 15, and 18, and postpartum D7 (n = 4/time-point and compared to control (non-pregnant animals. Beta cell mass, location, proliferation (Ki67+, and proportion of Ins+Glut2LO cells were measured using immunohistochemistry and bright field or confocal microscopy. Beta cell mass tripled by GD18 and β-cell proliferation peaked at GD12 in islets (≥6 β-cells and small β-cell clusters (1-5 β-cells. The proportion and fraction of Ins+Glut2LO cells undergoing proliferation increased significantly at GD9 in both islets and clusters, preceding the increase in β-cell mass and proliferation, and their proliferation within clusters persisted until GD15. The overall number of clusters increased significantly at GD9. Quantitative PCR showed a significant increase in Pdx1 presence at GD9 vs. GD18 or control pancreas, and Pdx1 was visualized by immunohistochemistry within both Ins+Glut2LO and Ins+Glut2HI cells within clusters. These results indicate that Ins+Glut2LO cells are likely to contribute to β-cell mass expansion during pregnancy.

Evaporation variability of Nam Co Lake in the Tibetan Plateau and its role in recent rapid lake expansion

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Ma, Ning; Szilagyi, Jozsef; Niu, Guo-Yue; Zhang, Yinsheng; Zhang, Teng; Wang, Binbin; Wu, Yanhong

2016-06-01

Previous studies have shown that the majority of the lakes in the Tibetan Plateau (TP) started to expand rapidly since the late 1990s. However, the causes are still not well known. For Nam Co, being a closed lake with no outflow, evaporation (EL) over the lake surface is the only way water may leave the lake. Therefore, quantifying EL is key for investigating the mechanism of lake expansion in the TP. EL can be quantified by Penman- and/or bulk-transfer-type models, requiring only net radiation, temperature, humidity and wind speed for inputs. However, interpolation of wind speed data may be laden with great uncertainty due to extremely sparse ground meteorological observations, the highly heterogeneous landscape and lake-land breeze effects. Here, evaporation of Nam Co Lake was investigated within the 1979-2012 period at a monthly time-scale using the complementary relationship lake evaporation (CRLE) model which does not require wind speed data. Validations by in-situ observations of E601B pan evaporation rates at the shore of Nam Co Lake as well as measured EL over an adjacent small lake using eddy covariance technique suggest that CRLE is capable of simulating EL well since it implicitly considers wind effects on evaporation via its vapor transfer coefficient. The multi-year average of annual evaporation of Nam Co Lake is 635 mm. From 1979 to 2012, annual evaporation of Nam Co Lake expressed a very slight decreasing trend. However, a more significant decrease in EL occurred during 1998-2008 at a rate of -12 mm yr-1. Based on water-level readings, this significant decrease in lake evaporation was found to be responsible for approximately 4% of the reported rapid water level increase and areal expansion of Nam Co Lake during the same period.

Feasibility of mesenchymal stem cell culture expansion for a phase I clinical trial in multiple sclerosis.

Science.gov (United States)

Planchon, Sarah M; Lingas, Karen T; Reese Koç, Jane; Hooper, Brittney M; Maitra, Basabi; Fox, Robert M; Imrey, Peter B; Drake, Kylie M; Aldred, Micheala A; Lazarus, Hillard M; Cohen, Jeffrey A

2018-01-01

Multiple sclerosis is an inflammatory, neurodegenerative disease of the central nervous system for which therapeutic mesenchymal stem cell transplantation is under study. Published experience of culture-expanding multiple sclerosis patients' mesenchymal stem cells for clinical trials is limited. To determine the feasibility of culture-expanding multiple sclerosis patients' mesenchymal stem cells for clinical use. In a phase I trial, autologous, bone marrow-derived mesenchymal stem cells were isolated from 25 trial participants with multiple sclerosis and eight matched controls, and culture-expanded to a target single dose of 1-2 × 10 6 cells/kg. Viability, cell product identity and sterility were assessed prior to infusion. Cytogenetic stability was assessed by single nucleotide polymorphism analysis of mesenchymal stem cells from 18 multiple sclerosis patients and five controls. One patient failed screening. Mesenchymal stem cell culture expansion was successful for 24 of 25 multiple sclerosis patients and six of eight controls. The target dose was achieved in 16-62 days, requiring two to three cell passages. Growth rate and culture success did not correlate with demographic or multiple sclerosis disease characteristics. Cytogenetic studies identified changes on one chromosome of one control (4.3%) after extended time in culture. Culture expansion of mesenchymal stem cells from multiple sclerosis patients as donors is feasible. However, culture time should be minimized for cell products designated for therapeutic administration.

Forced expression of Sox2 or Nanog in human bone marrow derived mesenchymal stem cells maintains their expansion and differentiation capabilities

International Nuclear Information System (INIS)

Go, Masahiro J.; Takenaka, Chiemi; Ohgushi, Hajime

2008-01-01

Mesenchymal stem cells (MSCs) derived from human bone marrow have capability to differentiate into cells of mesenchymal lineage. The cells have already been applied in various clinical situations because of their expansion and differentiation capabilities. The cells lose their capabilities after several passages, however. With the aim of conferring higher capability on human bone marrow MSCs, we introduced the Sox2 or Nanog gene into the cells. Sox2 and Nanog are not only essential for pluripotency and self-renewal of embryonic stem cells, but also expressed in somatic stem cells that have superior expansion and differentiation potentials. We found that Sox2-expressing MSCs showed consistent proliferation and osteogenic capability in culture media containing basic fibroblast growth factor (bFGF) compared to control cells. Significantly, in the presence of bFGF in culture media, most of the Sox2-expressing cells were small, whereas the control cells were elongated in shape. We also found that Nanog-expressing cells even in the absence of bFGF had much higher capabilities for expansion and osteogenesis than control cells. These results demonstrate not only an effective way to maintain proliferation and differentiation potentials of MSCs but also an important implication about the function of bFGF for self-renewal of stem cells including MSCs

Co-stimulation through 4-1BB/CD137 improves the expansion and function of CD8(+ melanoma tumor-infiltrating lymphocytes for adoptive T-cell therapy.

Directory of Open Access Journals (Sweden)

Jessica Ann Chacon

Full Text Available Adoptive T-cell therapy (ACT using tumor-infiltrating lymphocytes (TIL can induce tumor regression in up to 50% or more of patients with unresectable metastatic melanoma. However, current methods to expand melanoma TIL, especially the "rapid expansion protocol" (REP were not designed to enhance the generation of optimal effector-memory CD8(+ T cells for infusion. One approach to this problem is to manipulate specific co-stimulatory signaling pathways to enhance CD8(+ effector-memory T-cell expansion. In this study, we determined the effects of activating the TNF-R family member 4-1BB/CD137, specifically induced in activated CD8(+ T cells, on the yield, phenotype, and functional activity of expanded CD8(+ T cells during the REP. We found that CD8(+ TIL up-regulate 4-1BB expression early during the REP after initial TCR stimulation, but neither the PBMC feeder cells in the REP or the activated TIL expressed 4-1BB ligand. However, addition of an exogenous agonistic anti-4-1BB IgG4 (BMS 663513 to the REP significantly enhanced the frequency and total yield of CD8(+ T cells as well as their maintenance of CD28 and increased their anti-tumor CTL activity. Gene expression analysis found an increase in bcl-2 and survivin expression induced by 4-1BB that was associated with an enhanced survival capability of CD8(+ post-REP TIL when re-cultured in the absence or presence of cytokines. Our findings suggest that adding an agonistic anti-4-1BB antibody during the time of TIL REP initiation produces a CD8(+ T cell population capable of improved effector function and survival. This may greatly improve TIL persistence and anti-tumor activity in vivo after adoptive transfer into patients.

β-catenin is selectively required for the expansion and regeneration of mature pancreatic acinar cells in mice

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Matthew D. Keefe

2012-07-01

The size of the pancreas is determined by intrinsic factors, such as the number of progenitor cells, and by extrinsic signals that control the fate and proliferation of those progenitors. Both the exocrine and endocrine compartments of the pancreas undergo dramatic expansion after birth and are capable of at least partial regeneration following injury. Whether the expansion of these lineages relies on similar mechanisms is unknown. Although we have shown that the Wnt signaling component β-catenin is selectively required in mouse embryos for the generation of exocrine acinar cells, this protein has been ascribed various functions in the postnatal pancreas, including proliferation and regeneration of islet as well as acinar cells. To address whether β-catenin remains important for the maintenance and expansion of mature acinar cells, we have established a system to follow the behavior and fate of β-catenin-deficient cells during postnatal growth and regeneration in mice. We find that β-catenin is continuously required for the establishment and maintenance of acinar cell mass, extending from embryonic specification through juvenile and adult self-renewal and regeneration. This requirement is not shared with islet cells, which proliferate and function normally in the absence of β-catenin. These results make distinct predictions for the relative role of Wnt–β-catenin signaling in the etiology of human endocrine and exocrine disease. We suggest that loss of Wnt–β-catenin activity is unlikely to drive islet dysfunction, as occurs in type 2 diabetes, but that β-catenin is likely to promote human acinar cell proliferation following injury, and might therefore contribute to the resolution of acute or chronic pancreatitis.

Ex vivo expansion of bovine corneal endothelial cells in xeno-free medium supplemented with platelet releasate.

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Ming-Li Chou

Full Text Available Clinical-grade ex vivo expansion of corneal endothelial cells can increase the availability of corneal tissues for transplantation and treatment of corneal blindness. However, these cells have very limited proliferative capacity. Successful propagation has required so far to use very complex growth media supplemented with fetal bovine serum and other xenocomponents. We hypothesized that human platelet releasates rich in multiple growth factors, and in particular neurotrophins, could potentially be a useful supplement for ex vivo expansion of corneal endothelium cells due to their neural crest origin. Platelet releasates were prepared by calcium salt activation of apheresis platelet concentrates, subjected or not to complement inactivation by heat treatment at 56°C for 30 minutes. Platelet releasates were characterized for their content in proteins and were found to contain high amount of growth factors including platelet-derived growth factor-AB (30.56 to 39.08 ng/ml and brain-derived neurotrophic factor (30.57 to 37.11 ng/ml neurotrophins. We compared the growth and viability of corneal endothelium cells in DMEM-F12 medium supplemented with different combinations of components, including 2.5%∼10% of the platelet releasates. Corneal endothelium cells expanded in platelet releasates exhibited good adhesion and a typical hexagonal morphology. Their growth and viability were enhanced when using the complement-inactivated platelet releasate at a concentration of 10%. Immunostaining and Western blots showed that CECs maintained the expressions of four important membrane markers: Na-K ATPase α1, zona occludens-1, phospho-connexin 43 and N-cadherin. In conclusion, our study provides the first proof-of-concept that human platelet releasates can be used for ex vivo expansion of corneal endothelium cells. These findings open a new paradigm for ex vivo propagation protocols of corneal endothelium cells in compliance with good tissue culture practices

Assessment of the dental and skeletal effects of fan-type rapid maxillary expansion screw and Hyrax screw on craniofacial structures

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Umarevathi Gopalakrishnan

2017-01-01

Full Text Available Aims and Objectives: The purpose of the study was to assess the skeletal and dental effects of fan-type rapid maxillary expansion (RME appliance and Hyrax RME appliance on the craniofacial structures. Materials and Methods: The sample of the study included 12 patients with constricted maxillary arches. Acrylic bonded type of attachment was used for both groups. Changes in sagittal, vertical, and transverse relationship were assessed with lateral and frontal cephalograms, respectively. Intercanine and intermolar widths were measured with stone models. Pre- and immediate post-treatment records were statistically analyzed with Wilcoxon signed-rank test. The differences between the groups were evaluated using Mann–Whitney U-test. Since the data pertaining to intercanine width and intermolar width were normally distributed, parametric test of signifi cance (unpaired t-test was used to compare them. Results: Results showed that Hyrax presented with signifi cantly greater increments for both nasal cavity width and maxillary width when compared to fan-type RME. Both groups had retroclination of incisors. The increase in the intercanine width was almost similar in both groups. Conclusion: Fan-type RME caused only minimal expansion of the intermolar width when compared to the Hyrax. The ratio between the intercanine and intermolar width expansion was nearly 4:1 in the fan-type RME and 0.75:1 in Hyrax.

Bioactive poly(ethylene glycol) hydrogels to recapitulate the HSC niche and facilitate HSC expansion in culture.

Science.gov (United States)

Cuchiara, Maude L; Coşkun, Süleyman; Banda, Omar A; Horter, Kelsey L; Hirschi, Karen K; West, Jennifer L

2016-04-01

Hematopoietic stem cells (HSCs) have been used therapeutically for decades, yet their widespread clinical use is hampered by the inability to expand HSCs successfully in vitro. In culture, HSCs rapidly differentiate and lose their ability to self-renew. We hypothesize that by mimicking aspects of the bone marrow microenvironment in vitro we can better control the expansion and differentiation of these cells. In this work, derivatives of poly(ethylene glycol) diacrylate hydrogels were used as a culture substrate for hematopoietic stem and progenitor cell (HSPC) populations. Key HSC cytokines, stem cell factor (SCF) and interferon-γ (IFNγ), as well as the cell adhesion ligands RGDS and connecting segment 1 were covalently immobilized onto the surface of the hydrogels. With the use of SCF and IFNγ, we observed significant expansion of HSPCs, ∼97 and ∼104 fold respectively, while maintaining c-kit(+) lin(-) and c-kit(+) Sca1(+) lin(-) (KSL) populations and the ability to form multilineage colonies after 14 days. HSPCs were also encapsulated within degradable poly(ethylene glycol) hydrogels for three-dimensional culture. After expansion in hydrogels, ∼60% of cells were c-kit(+), demonstrating no loss in the proportion of these cells over the 14 day culture period, and ∼50% of colonies formed were multilineage, indicating that the cells retained their differentiation potential. The ability to tailor and use this system to support HSC growth could have implications on the future use of HSCs and other blood cell types in a clinical setting. © 2015 Wiley Periodicals, Inc.

Did the Higgs boson drive the universe's expansion?

CERN Multimedia

2008-01-01

The Higgs boson has been moonlighting. Not content with its day job of giving other particles their mass, it may also have driven the expansion of the early universe, given a little tinkering, according to two separate studies. Soon after the big bang the early universe is believed to have undergone a period of rapid expansion, known as inflation.

In vitro expansion of Lin+ and Lin− mononuclear cells from human peripheral blood

International Nuclear Information System (INIS)

Norhaiza, H. Siti; Zarina, Z. A. Intan; Hisham, Z. A. Shahrul; Rohaya, M. A. W.

2013-01-01

Haematopoietic stem cells (HSCs) are used in the therapy of blood disorders due to the ability of these cells to reconstitute haematopoietic lineage cells when transplanted into myeloablative recipients. However, substantial number of cells is required in order for the reconstitution to take place. Since HSCs present in low frequency, larger number of donor is required to accommodate the demand of transplantable HSCs. Therefore, in vitro expansion of HSCs will have profound impact on clinical purposes. The aim of this study was to expand lineage negative (Lin − ) stem cells from human peripheral blood. Total peripheral blood mononuclear cells (PBMNCs) were fractionated from human blood by density gradient centrifugation. Subsequently, PBMNCs were subjected to magnetic assisted cell sorter (MACS) which depletes lineage positive (Lin + ) mononuclear cells expressing lineage positive markers such as CD2, CD3, CD11b, CD14, CD15, CD16, CD19, CD56, CD123, and CD235a to obtained Lin − cell population. The ability of Lin + and Lin − to survive in vitro was explored by culturing both cell populations in complete medium consisting of Alpha-Minimal Essential Medium (AMEM) +10% (v/v) Newborn Calf Serum (NBCS)+ 2% (v/v) pen/strep. In another experiment, Lin + and Lin − were cultured with complete medium supplemented with 10ng/mL of the following growth factors: stem cell factor (SCF), interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF), 2IU/mL of Erythropoietin (Epo) and 20ng/mL of IL-6. Three samples were monitored in static culture for 22 days. The expansion potential was assessed by the number of total viable cells, counted by trypan blue exclusion assay. It was found that Lin + mononuclear cells were not able to survive either in normal proliferation medium or proliferation medium supplemented with cytokines. Similarly, Lin − stem cells were not able to survive in proliferation medium however, addition of cytokines into the proliferation

In vitro expansion of Lin+ and Lin- mononuclear cells from human peripheral blood

Science.gov (United States)

Norhaiza, H. Siti; Rohaya, M. A. W.; Zarina, Z. A. Intan; Hisham, Z. A. Shahrul

2013-11-01

Haematopoietic stem cells (HSCs) are used in the therapy of blood disorders due to the ability of these cells to reconstitute haematopoietic lineage cells when transplanted into myeloablative recipients. However, substantial number of cells is required in order for the reconstitution to take place. Since HSCs present in low frequency, larger number of donor is required to accommodate the demand of transplantable HSCs. Therefore, in vitro expansion of HSCs will have profound impact on clinical purposes. The aim of this study was to expand lineage negative (Lin-) stem cells from human peripheral blood. Total peripheral blood mononuclear cells (PBMNCs) were fractionated from human blood by density gradient centrifugation. Subsequently, PBMNCs were subjected to magnetic assisted cell sorter (MACS) which depletes lineage positive (Lin+) mononuclear cells expressing lineage positive markers such as CD2, CD3, CD11b, CD14, CD15, CD16, CD19, CD56, CD123, and CD235a to obtained Lin- cell population. The ability of Lin+ and Lin- to survive in vitro was explored by culturing both cell populations in complete medium consisting of Alpha-Minimal Essential Medium (AMEM) +10% (v/v) Newborn Calf Serum (NBCS)+ 2% (v/v) pen/strep. In another experiment, Lin+ and Lin- were cultured with complete medium supplemented with 10ng/mL of the following growth factors: stem cell factor (SCF), interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF), 2IU/mL of Erythropoietin (Epo) and 20ng/mL of IL-6. Three samples were monitored in static culture for 22 days. The expansion potential was assessed by the number of total viable cells, counted by trypan blue exclusion assay. It was found that Lin+ mononuclear cells were not able to survive either in normal proliferation medium or proliferation medium supplemented with cytokines. Similarly, Lin- stem cells were not able to survive in proliferation medium however, addition of cytokines into the proliferation medium support Lin

Evaluating Clonal Expansion of HIV-Infected Cells: Optimization of PCR Strategies to Predict Clonality

Science.gov (United States)

Laskey, Sarah B.; Pohlmeyer, Christopher W.; Bruner, Katherine M.; Siliciano, Robert F.

2016-01-01

In HIV-infected individuals receiving suppressive antiretroviral therapy, the virus persists indefinitely in a reservoir of latently infected cells. The proliferation of these cells may contribute to the stability of the reservoir and thus to the lifelong persistence of HIV-1 in infected individuals. Because the HIV-1 replication process is highly error-prone, the detection of identical viral genomes in distinct host cells provides evidence for the clonal expansion of infected cells. We evaluated alignments of unique, near-full-length HIV-1 sequences to determine the relationship between clonality in a short region and clonality in the full genome. Although it is common to amplify and sequence short, subgenomic regions of the viral genome for phylogenetic analysis, we show that sequence identity of these amplicons does not guarantee clonality across the full viral genome. We show that although longer amplicons capture more diversity, no subgenomic region can recapitulate the diversity of full viral genomes. Consequently, some identical subgenomic amplicons should be expected even from the analysis of completely unique viral genomes, and the presence of identical amplicons alone is not proof of clonally expanded HIV-1. We present a method for evaluating evidence of clonal expansion in the context of these findings. PMID:27494508

Evaluating Clonal Expansion of HIV-Infected Cells: Optimization of PCR Strategies to Predict Clonality.

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Sarah B Laskey

2016-08-01

Full Text Available In HIV-infected individuals receiving suppressive antiretroviral therapy, the virus persists indefinitely in a reservoir of latently infected cells. The proliferation of these cells may contribute to the stability of the reservoir and thus to the lifelong persistence of HIV-1 in infected individuals. Because the HIV-1 replication process is highly error-prone, the detection of identical viral genomes in distinct host cells provides evidence for the clonal expansion of infected cells. We evaluated alignments of unique, near-full-length HIV-1 sequences to determine the relationship between clonality in a short region and clonality in the full genome. Although it is common to amplify and sequence short, subgenomic regions of the viral genome for phylogenetic analysis, we show that sequence identity of these amplicons does not guarantee clonality across the full viral genome. We show that although longer amplicons capture more diversity, no subgenomic region can recapitulate the diversity of full viral genomes. Consequently, some identical subgenomic amplicons should be expected even from the analysis of completely unique viral genomes, and the presence of identical amplicons alone is not proof of clonally expanded HIV-1. We present a method for evaluating evidence of clonal expansion in the context of these findings.

Synthetic surface for expansion of human mesenchymal stem cells in xeno-free, chemically defined culture conditions.

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Paula J Dolley-Sonneville

Full Text Available Human mesenchymal stem cells (HMSCS possess three properties of great interest for the development of cell therapies and tissue engineering: multilineage differentiation, immunomodulation, and production of trophic factors. Efficient ex vivo expansion of hMSCs is a challenging requirement for large scale production of clinical grade cells. Low-cost, robust, scalable culture methods using chemically defined materials need to be developed to address this need. This study describes the use of a xeno-free synthetic peptide acrylate surface, the Corning® Synthemax® Surface, for culture of hMSCs in serum-free, defined medium. Cell performance on the Corning Synthemax Surface was compared to cells cultured on biological extracellular matrix (ECM coatings in xeno-free defined medium and in traditional conditions on tissue culture treated (TCT plastic in fetal bovine serum (FBS supplemented medium. Our results show successful maintenance of hMSCs on Corning Synthemax Surface for eight passages, with cell expansion rate comparable to cells cultured on ECM and significantly higher than for cells in TCT/FBS condition. Importantly, on the Corning Synthemax Surface, cells maintained elongated, spindle-like morphology, typical hMSC marker profile and in vitro multilineage differentiation potential. We believe the Corning Synthemax Surface, in combination with defined media, provides a complete synthetic, xeno-free, cell culture system for scalable production of hMSCs.

Functional and Structural Characterization of a Receptor-Like Kinase Involved in Germination and Cell Expansion in Arabidopsis

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Wu, Zhen; Liang, Shan; Song, Wen; Lin, Guangzhong; Wang, Weiguang; Zhang, Heqiao; Han, Zhifu; Chai, Jijie

2017-01-01

Leucine-rich repeat receptor-like kinases (LRR-RLKs) are widespread in different plant species and play important roles in growth and development. Germination inhibition is vital for the completion of seed maturation and cell expansion is a fundamental cellular process driving plant growth. Here, we report genetic and structural characterizations of a functionally uncharacterized LRR-RLK, named GRACE (Germination Repression and Cell Expansion receptor-like kinase). Overexpression of GRACE in Arabidopsis exhibited delayed germination, enlarged cotyledons, rosette leaves and stubbier petioles. Conversely, these phenotypes were reversed in the T-DNA insertion knock-down mutant grace-1 plants. A crystal structure of the extracellular domain of GRACE (GRACE-LRR) determined at the resolution of 3.0 Å revealed that GRACE-LRR assumed a right-handed super-helical structure with an island domain (ID). Structural comparison showed that structure of the ID in GRACE-LRR is strikingly different from those observed in other LRR-RLKs. This structural observation implies that GRACE might perceive a new ligand for signaling. Collectively, our data support roles of GRACE in repressing seed germination and promoting cell expansion of Arabidopsis, presumably by perception of unknown ligand(s). PMID:29213277

Up-regulating the abscisic acid inactivation gene ZmABA8ox1b contributes to seed germination heterosis by promoting cell expansion.

Science.gov (United States)

Li, Yangyang; Wang, Cheng; Liu, Xinye; Song, Jian; Li, Hongjian; Sui, Zhipeng; Zhang, Ming; Fang, Shuang; Chu, Jinfang; Xin, Mingming; Xie, Chaojie; Zhang, Yirong; Sun, Qixin; Ni, Zhongfu

2016-04-01

Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Methods of ex vivo expansion of human cord blood cells: challenges, successes and clinical implications.

Science.gov (United States)

Baron, Frédéric; Ruggeri, Annalisa; Nagler, Arnon

2016-03-01

More than 40,000 unrelated cord blood transplantations (UCBT) have been performed worldwide as treatment for patients with malignant or non-malignant life threatening hematologic disorders. However, low absolute numbers of hematopoietic stem and progenitor cells (HSPCs) within a single cord blood unit has remained a limiting factor for this transplantation modality, particularly in adult recipients. Further, because UCB contains low numbers of mostly naïve T cells, immune recovery after UCBT is slow, predisposing patients to severe infections. Other causes of UCBT failure has included graft-versus-host disease (GVHD) and relapse of the underlying disease. In this article, we first review the current landscape of cord blood engineering aimed at improving engraftment. This includes approaches of UCB-HSPCs expansion and methods aimed at improving UCB-HSCPs homing. We then discuss recent approaches of cord blood engineering developed to prevent infection [generation of multivirus-specific cytotoxic T cells (VSTs) from UCB], relapse [transduction of UCB-T cells with tumor-specific chimeric receptor antigens (CARs)] and GVHD (expansion of regulatory T cells from UCB). Although many of these techniques of UCB engineering remain currently technically challenging and expensive, they are likely to revolutionize the field of UCBT in the next decades.

Modeling of thermal expansion coefficient of perovskite oxide for solid oxide fuel cell cathode

Science.gov (United States)

Heydari, F.; Maghsoudipour, A.; Alizadeh, M.; Khakpour, Z.; Javaheri, M.

2015-09-01

Artificial intelligence models have the capacity to eliminate the need for expensive experimental investigation in various areas of manufacturing processes, including the material science. This study investigates the applicability of adaptive neuro-fuzzy inference system (ANFIS) approach for modeling the performance parameters of thermal expansion coefficient (TEC) of perovskite oxide for solid oxide fuel cell cathode. Oxides (Ln = La, Nd, Sm and M = Fe, Ni, Mn) have been prepared and characterized to study the influence of the different cations on TEC. Experimental results have shown TEC decreases favorably with substitution of Nd3+ and Mn3+ ions in the lattice. Structural parameters of compounds have been determined by X-ray diffraction, and field emission scanning electron microscopy has been used for the morphological study. Comparison results indicated that the ANFIS technique could be employed successfully in modeling thermal expansion coefficient of perovskite oxide for solid oxide fuel cell cathode, and considerable savings in terms of cost and time could be obtained by using ANFIS technique.

Dominant role of antigen dose in CD4+Foxp3+ regulatory T cell induction and expansion1

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Turner, Michael S.; Kane, Lawrence P.; Morel, Penelope A.

2009-01-01

The definitions of tolerogenic vs. immunogenic dendritic cells (DC) remain controversial. Immature DC have been shown to induce T regulatory cells (Treg) specific for foreign and allo-antigens. However, we have previously reported that mature DC (G4DC) prevented the onset of autoimmune diabetes whereas immature DC (GMDC) were therapeutically ineffective. In this study, islet-specific CD4+ T cells from BDC2.5 TCR Tg mice were stimulated, in the absence of exogenous cytokine, with GMDC or G4DC pulsed with high- or low-affinity antigenic peptides and examined for Treg induction. Both GMDC and G4DC presenting low peptide doses induced weak TCR signaling via the Akt/mTOR pathway, resulting in significant expansion of Foxp3+ Treg. Furthermore, unpulsed G4DC, but not GMDC, also induced Treg. High peptide doses induced strong Akt/mTOR signaling and favored the expansion of Foxp3neg Th cells. The inverse correlation of Foxp3 and Akt/mTOR signaling was also observed in DO11.10 and OT-II TCR-Tg T cells and was recapitulated with anti-CD3/CD28 stimulation in the absence of DC. IL-6 production in these cultures correlated positively with antigen dose and inversely with Treg expansion. Studies with T cells or DC from IL-6−/− mice revealed that IL-6 production by T cells was more important in the inhibition of Treg induction at low antigen doses. These studies indicate that strength of Akt/mTOR signaling, a critical T cell intrinsic determinant for Treg vs Th induction, can be controlled by adjusting the dose of antigenic peptide. Furthermore, this operates in a dominant fashion over DC phenotype and cytokine production. PMID:19801514

IL-6 trans-Signaling-Dependent Rapid Development of Cytotoxic CD8+ T Cell Function

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Jan P. Böttcher

2014-09-01

Full Text Available Immune control of infections with viruses or intracellular bacteria relies on cytotoxic CD8+ T cells that use granzyme B (GzmB for elimination of infected cells. During inflammation, mature antigen-presenting dendritic cells instruct naive T cells within lymphoid organs to develop into effector T cells. Here, we report a mechanistically distinct and more rapid process of effector T cell development occurring within 18 hr. Such rapid acquisition of effector T cell function occurred through cross-presenting liver sinusoidal endothelial cells (LSECs in the absence of innate immune stimulation and known costimulatory signaling. Rather, interleukin-6 (IL-6 trans-signaling was required and sufficient for rapid induction of GzmB expression in CD8+ T cells. Such LSEC-stimulated GzmB-expressing CD8+ T cells further responded to inflammatory cytokines, eliciting increased and protracted effector functions. Our findings identify a role for IL-6 trans-signaling in rapid generation of effector function in CD8+ T cells that may be beneficial for vaccination strategies.

Effects of rapid maxillary expansion on head posture, postural stability, and fall risk

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Fatih Celebi

2017-01-01

Full Text Available Objective: The aim of this study was to investigate the effects of rapid maxillary expansion (RME on head posture, postural stability, and fall risk. Materials and Methods: A sample of 51 adolescent patients was randomly divided into two groups. In the first group, which consisted of 28 patients (15 females and 13 males, RME was performed as a part of routine orthodontic treatment. The remaining 23 individuals (12 females and 11 males served as the control group. Lateral cephalometric radiographs taken in natural head position, postural stability, and fall risk scores were obtained during the first visit. They were repeated on average 3.8 months and 3.5 months later for the study and control groups, respectively. The changes were analyzed using the Wilcoxon signed-rank test, paired samples t-test, Mann–Whitney U-test, and independent samples t-test. Results: As a result of RME, a statistically significant decrease was detected in the fall risk score (P < 0.05 in the study group, while the head position and postural stability remained unchanged. For the control group, no significant changes were observed in all measurements. Conclusions: The result of the present study suggests that RME has a capacity of improving fall risk.

A protein-based hydrogel for in vitro expansion of mesenchymal stem cells.

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Jingyu Wang

Full Text Available Hydrogels are widely used as scaffolds in tissue engineering because they can provide excellent environments for bioactive components including growth factors and cells. We reported in this study on a physical hydrogel formed by a specific protein-peptide interaction, which could be used for the three dimensional (3D cell culture of murine mesenchymal stem cells (mMSC. The mMSC kept dividing during the 7-day culture period and the metabolic-active cell number at day 7 was 359% more than that at day 1. This kind of physical hydrogel could be converted to a homogeneous solution by firstly adding an equal volume of culture medium and then pipeting for several times. Therefore, mMSC post culture could be easily separated from cell-gel constructs. We believed that the protein-based hydrogel system in this study could be developed into a promising scaffold for in vitro expansion of stem cells and cell therapy. This work would be in the general interests of researchers in the fields of biomaterials and supramolecular chemistry.

Atypical PKC-iota Controls Stem Cell Expansion via Regulation of the Notch Pathway

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In Kyoung Mah

2015-11-01

Full Text Available The number of stem/progenitor cells available can profoundly impact tissue homeostasis and the response to injury or disease. Here, we propose that an atypical PKC, Prkci, is a key player in regulating the switch from an expansion to a differentiation/maintenance phase via regulation of Notch, thus linking the polarity pathway with the control of stem cell self-renewal. Prkci is known to influence symmetric cell division in invertebrates; however a definitive role in mammals has not yet emerged. Using a genetic approach, we find that loss of Prkci results in a marked increase in the number of various stem/progenitor cells. The mechanism used likely involves inactivation and symmetric localization of NUMB, leading to the activation of NOTCH1 and its downstream effectors. Inhibition of atypical PKCs may be useful for boosting the production of pluripotent stem cells, multipotent stem cells, or possibly even primordial germ cells by promoting the stem cell/progenitor fate.

Ex vivo expansion of hematopoietic stem cell by fusion protein TAT-Zfx

International Nuclear Information System (INIS)

Xu Chong; Zhang Yanbing; Jiang Hua

2009-01-01

The relative inability of hemopoietic stem cells (HSCs) to reproduce themselves (self-renew) ex vivo imposes substantial limitations on the current use of HSC transplantation. Recently, the transcription factor Zfx has been demonstrated that played an important in controlling the self-renewal of hematopoietic stem cells. Here, we reported that Zfx could enable high-level expansion of HSCs in vitro, by combination of protein transduction domain, TAT. Furthermore, we also demonstrated that expanded HSCs population retains their normal in vivo potential of pluripotency. It is thus that TAT-Zfx has the potential to expand HSCs significantly in vitro, and will have enormous clinical potentials.

Tumor-associated mesenchymal stem cells inhibit naïve T cell expansion by blocking cysteine export from dendritic cells.

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Ghosh, Tithi; Barik, Subhasis; Bhuniya, Avishek; Dhar, Jesmita; Dasgupta, Shayani; Ghosh, Sarbari; Sarkar, Madhurima; Guha, Ipsita; Sarkar, Koustav; Chakrabarti, Pinak; Saha, Bhaskar; Storkus, Walter J; Baral, Rathindranath; Bose, Anamika

2016-11-01

Mesenchymal stem cells (MSCs) represent an important cellular constituent of the tumor microenvironment, which along with tumor cells themselves, serve to regulate protective immune responses in support of progressive disease. We report that tumor MSCs prevent the ability of dendritic cells (DC) to promote naïve CD4(+) and CD8(+) T cell expansion, interferon gamma secretion and cytotoxicity against tumor cells, which are critical to immune-mediated tumor eradication. Notably, tumor MSCs fail to prevent DC-mediated early T cell activation events or the ability of responder T cells to produce IL-2. The immunoregulatory activity of tumor MSCs is IL-10- and STAT3-dependent, with STAT3 repressing DC expression of cystathionase, a critical enzyme that converts methionine-to-cysteine. Under cysteine-deficient priming conditions, naïve T cells exhibit defective cellular metabolism and proliferation. Bioinformatics analyses as well as in vitro observations suggest that STAT3 may directly bind to a GAS-like motif within the cystathionase promoter (-269 to -261) leading to IL-10-STAT3 mediated repression of cystathionase gene transcription. Our collective results provide evidence for a novel mechanism of tumor MSC-mediated T cell inhibition within tumor microenvironment. © 2016 UICC.

Acquisition and Expansion of Adult Rat Bone Marrow Multipotent Mesenchymal Stromal Cells

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Šulla I.

2017-03-01

Full Text Available This study was initiated in order to test a mini-invasive method of mesenchymal stem/progenitor cells (MS/PCs isolation from a rat bone marrow (BM, and subsequently their expansion, differentiation, and evaluation of their immunophenotypic characteristics; and later their preservation as donor cells in an optimal condition for potential autotransplantation. The study group comprised of 6 adult male Sprague-Dawley (S-D rats, weighing 480—690 g. The rats were anaesthetised by isoflurane with room air in a Plexiglas box and maintained by inhalation of a mixture of isoflurane and O2. Their femurs were surgically exposed and their diaphyses double-trephined. Then BM cells were flushed out by saline with heparin and aspirated into a syringe with a solution of DMEM (Dulbecco’s modified eagle’s medium and heparin. The mononuclear cells from the BM were isolated by centrifugation and expanded in a standard culture medium supplemented with ES-FBS (es-cell-qualified foetal bovine serum, L-glutamine and rh LIF (recombinant human leukemia inhibitory factor. Following 14 days of passaging cultures, the cells were split into 2 equal parts. The first culture continued with the original medium. The second culture received additional supplementation with a human FGFβ (fibroblast growth factor beta and EGF (epidermal growth factor. The populations of these cells were analysed by light-microscopy, then the mean fluorescence intensities (MFIs of CD90 and Nestin were evaluated by a tricolour flow cytometry using monoclonal antibodies. The type of general anaesthesia used proved to be appropriate for the surgical phase of the experiments. All rats survived the harvesting of the BM without complications. The total number of mononuclear cells was 1.5—4.0 × 106 per sample and the proportion of CD90/Nestin expressing cells was < 1 %. Following 14 days of expansion, the cells became larger, adherent, with fibrillary morphology; the proportion of cells expressing

Ferns are less dependent on passive dilution by cell expansion to coordinate leaf vein and stomatal spacing than angiosperms.

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Madeline R Carins Murphy

Full Text Available Producing leaves with closely spaced veins is a key innovation linked to high rates of photosynthesis in angiosperms. A close geometric link between veins and stomata in angiosperms ensures that investment in enhanced venous water transport provides the strongest net carbon return to the plant. This link is underpinned by "passive dilution" via expansion of surrounding cells. However, it is not known whether this 'passive dilution' mechanism is present in plant lineages other than angiosperms and is another key feature of the angiosperms' evolutionary success. Consequently, we sought to determine whether the 'passive dilution' mechanism is; (i exclusive to the angiosperms, (ii a conserved mechanism that evolved in the common ancestor of ferns and angiosperms, or (iii has evolved continuously over time. To do this we first we assessed the plasticity of vein and stomatal density and epidermal cell size in ferns in response to light environment. We then compared the relationships between these traits found among ferns with modelled relationships that assume vein and stomatal density respond passively to epidermal cell expansion, and with those previously observed in angiosperms. Vein density, stomatal density and epidermal cell size were linked in ferns with remarkably similar relationships to those observed in angiosperms, except that fern leaves had fewer veins per stomata. However, plasticity was limited in ferns and stomatal spacing was dependent on active stomatal differentiation as well as passive cell expansion. Thus, ferns (like angiosperms appear to coordinate vein and stomatal density with epidermal cell expansion to some extent to maintain a constant ratio between veins and stomata in the leaf. The different general relationships between vein density and stomatal density in ferns and angiosperms suggests the groups have different optimum balances between the production of vein tissue dedicated to water supply and stomatal tissue for gas

Expansion and maintenance of human embryonic stem cell–derived endothelial cells by TGFβ inhibition is Id1 dependent

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James, Daylon; Nam, Hyung-song; Seandel, Marco; Nolan, Daniel; Janovitz, Tyler; Tomishima, Mark; Studer, Lorenz; Lee, Gabsang; Lyden, David; Benezra, Robert; Zaninovic, Nikica; Rosenwaks, Zev; Rabbany, Sina Y; Rafii, Shahin

2010-01-01

Previous efforts to differentiate human embryonic stem cells (hESCs) into endothelial cells have not achieved sustained expansion and stability of vascular cells. To define vasculogenic developmental pathways and enhance differentiation, we used an endothelial cell–specific VE-cadherin promoter driving green fluorescent protein (GFP) (hVPr-GFP) to screen for factors that promote vascular commitment. In phase 1 of our method, inhibition of transforming growth factor (TGF)β at day 7 of differentiation increases hVPr-GFP+ cells by tenfold. In phase 2, TGFβ inhibition maintains the proliferation and vascular identity of purified endothelial cells, resulting in a net 36-fold expansion of endothelial cells in homogenous monolayers, which exhibited a transcriptional profile of Id1highVEGFR2highVE-cadherin+ ephrinB2+. Using an Id1-YFP hESC reporter line, we showed that TGFβ inhibition sustains Id1 expression in hESC-derived endothelial cells and that Id1 is required for increased proliferation and preservation of endothelial cell commitment. Our approach provides a serum-free method for differentiation and long-term maintenance of hESC-derived endothelial cells at a scale relevant to clinical application. PMID:20081865

The immune receptor Trem1 cooperates with diminished DNA damage response to induce preleukemic stem cell expansion.

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Du, W; Amarachintha, S; Wilson, A; Pang, Q

2017-02-01

Fanconi anemia (FA) is an inherited bone marrow failure syndrome with extremely high risk of leukemic transformation. Here we investigate the relationship between DNA damage response (DDR) and leukemogenesis using the Fanca knockout mouse model. We found that chronic exposure of the Fanca -/- hematopoietic stem cells to DNA crosslinking agent mitomycin C in vivo leads to diminished DDR, and the emergence/expansion of pre-leukemia stem cells (pre-LSCs). Surprisingly, although genetic correction of Fanca deficiency in the pre-LSCs restores DDR and reduces genomic instability, but fails to prevent pre-LSC expansion or delay leukemia development in irradiated recipients. Furthermore, we identified transcription program underlying dysregulated DDR and cell migration, myeloid proliferation, and immune response in the Fanca -/- pre-LSCs. Forced expression of the downregulated DNA repair genes, Rad51c or Trp53i13, in the Fanca -/- pre-LSCs partially rescues DDR but has no effect on leukemia, whereas shRNA knockdown of the upregulated immune receptor genes Trem1 or Pilrb improves leukemia-related survival, but not DDR or genomic instability. Furthermore, Trem1 cooperates with diminished DDR in vivo to promote Fanca -/- pre-LSC expansion and leukemia development. Our study implicates diminishing DDR as a root cause of FA leukemogenesis, which subsequently collaborates with other signaling pathways for leukemogenic transformation.

Proteome analysis of human Wharton's jelly cells during in vitro expansion

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Sulpizio Marilisa

2010-03-01

Full Text Available Abstract Background The human umbilical cord contains mucoid connective tissue and fibroblast-like cells. These cells named Wharton's jelly cells, (WJCs display properties similar to mesenchymal stem cells therefore representing a rich source of primitive cells to be potentially used in regenerative medicine. Results To better understand their self-renewal and potential in vitro expansion capacity, a reference 2D map was constructed as a proteomic data set. 158 unique proteins were identified. More than 30% of these proteins belong to cytoskeleton compartment. We also found that several proteins including Shootin1, Adenylate kinase 5 isoenzyme and Plasminogen activator-inhibitor 2 are no longer expressed after the 2nd passage of in vitro replication. This indicates that the proliferative potency of these cells is reduced after the initial stage of in vitro growing. At the end of cellular culturing, new synthesized proteins, including, ERO1-like protein alpha, Aspartyl-tRNA synthetase and Prolyl-4-hydroxylase were identified. It is suggested that these new synthesized proteins are involved in the impairment of cellular surviving during replication and differentiation time. Conclusions Our work represents an essential step towards gaining knowledge of the molecular properties of WJCs so as to better understand their possible use in the field of cell therapy and regenerative medicine.

Defining chemical expansion: the choice of units for the stoichiometric expansion coefficient

DEFF Research Database (Denmark)

Marrocchelli, Dario; Chatzichristodoulou, Christodoulos; Bishop, Sean R.

2014-01-01

Chemical expansion refers to the spatial dilation of a material that occurs upon changes in its composition. When this dilation is caused by a gradual, iso-structural increase in the lattice parameter with composition, it is related to the composition change by the stoichiometric expansion coeffi...... are provided for changes in oxygen content in fluorite, perovskite, and Ruddlesden-Popper (K2NiF4) phase materials used in solid oxide fuel cells....

Umbilical cord blood regulatory T-cell expansion and functional effects of tumor necrosis factor receptor family members OX40 and 4-1BB expressed on artificial antigen-presenting cells

Science.gov (United States)

Harker-Murray, Paul; Porter, Stephen B.; Merkel, Sarah C.; Londer, Aryel; Taylor, Dawn K.; Bina, Megan; Panoskaltsis-Mortari, Angela; Rubinstein, Pablo; Van Rooijen, Nico; Golovina, Tatiana N.; Suhoski, Megan M.; Miller, Jeffrey S.; Wagner, John E.; June, Carl H.; Riley, James L.

2008-01-01

Previously, we showed that human umbilical cord blood (UCB) regulatory T cells (Tregs) could be expanded approximately 100-fold using anti-CD3/28 monoclonal antibody (mAb)–coated beads to provide T-cell receptor and costimulatory signals. Because Treg numbers from a single UCB unit are limited, we explored the use of cell-based artificial antigen-presenting cells (aAPCs) preloaded with anti-CD3/28 mAbs to achieve higher levels of Treg expansion. Compared with beads, aAPCs had similar expansion properties while significantly increasing transforming growth factor β (TGF-β) secretion and the potency of Treg suppressor function. aAPCs modified to coexpress OX40L or 4-1BBL expanded UCB Tregs to a significantly greater extent than bead- or nonmodified aAPC cultures, reaching mean expansion levels exceeding 1250-fold. Despite the high expansion and in contrast to studies using other Treg sources, neither OX40 nor 4-1BB signaling of UCB Tregs reduced in vitro suppression. UCB Tregs expanded with 4-1BBL expressing aAPCs had decreased levels of proapoptotic bim. UCB Tregs expanded with nonmodified or modified aAPCs versus beads resulted in higher survival associated with increased Treg persistence in a xeno-geneic graft-versus-host disease lethality model. These data offer a novel approach for UCB Treg expansion using aAPCs, including those coexpressing OX40L or 4-1BBL. PMID:18645038

Human skeletal muscle-derived stem cells retain stem cell properties after expansion in myosphere culture

International Nuclear Information System (INIS)

Wei, Yan; Li, Yuan; Chen, Chao; Stoelzel, Katharina; Kaufmann, Andreas M.; Albers, Andreas E.

2011-01-01

Human skeletal muscle contains an accessible adult stem-cell compartment in which differentiated myofibers are maintained and replaced by a self-renewing stem cell pool. Previously, studies using mouse models have established a critical role for resident stem cells in skeletal muscle, but little is known about this paradigm in human muscle. Here, we report the reproducible isolation of a population of cells from human skeletal muscle that is able to proliferate for extended periods of time as floating clusters of rounded cells, termed 'myospheres' or myosphere-derived progenitor cells (MDPCs). The phenotypic characteristics and functional properties of these cells were determined using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and immunocytochemistry. Our results showed that these cells are clonogenic, express skeletal progenitor cell markers Pax7, ALDH1, Myod, and Desmin and the stem cell markers Nanog, Sox2, and Oct3/4 significantly elevated over controls. They could be maintained proliferatively active in vitro for more than 20 weeks and passaged at least 18 times, despite an average donor-age of 63 years. Individual clones (4.2%) derived from single cells were successfully expanded showing clonogenic potential and sustained proliferation of a subpopulation in the myospheres. Myosphere-derived cells were capable of spontaneous differentiation into myotubes in differentiation media and into other mesodermal cell lineages in induction media. We demonstrate here that direct culture and expansion of stem cells from human skeletal muscle is straightforward and reproducible with the appropriate technique. These cells may provide a viable resource of adult stem cells for future therapies of disease affecting skeletal muscle or mesenchymal lineage derived cell types.

Isotropic Negative Thermal Expansion Metamaterials.

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Wu, Lingling; Li, Bo; Zhou, Ji

2016-07-13

Negative thermal expansion materials are important and desirable in science and engineering applications. However, natural materials with isotropic negative thermal expansion are rare and usually unsatisfied in performance. Here, we propose a novel method to achieve two- and three-dimensional negative thermal expansion metamaterials via antichiral structures. The two-dimensional metamaterial is constructed with unit cells that combine bimaterial strips and antichiral structures, while the three-dimensional metamaterial is fabricated by a multimaterial 3D printing process. Both experimental and simulation results display isotropic negative thermal expansion property of the samples. The effective coefficient of negative thermal expansion of the proposed models is demonstrated to be dependent on the difference between the thermal expansion coefficient of the component materials, as well as on the circular node radius and the ligament length in the antichiral structures. The measured value of the linear negative thermal expansion coefficient of the three-dimensional sample is among the largest achieved in experiments to date. Our findings provide an easy and practical approach to obtaining materials with tunable negative thermal expansion on any scale.

Reaction of the nasal septal cartilage to rapid expansion of the maxillary dental arch

International Nuclear Information System (INIS)

Horisaka, Takashi

1981-01-01

1) In the control group, in which maxillary expansion was not performed, the cell proliferation and uptake of [ 3 H] proline in the septal cartilage were found to be more remarkable at the area of the septoethmoidal junction than other areas of the cartilage. In the experimental group, however, higher activity of the cell proliferation and more intense uptake of [ 3 H] proline were found at the area adjacent to the septoethmoidal junction than the control, while no change was observed in other areas of the septal cartilage. 2) The cartilaginous cells at the septoethmoidal junction represented the columnar pattern in their alignment and the cells were classified into five zones according to their morphological characteristics, i.e. resting, proliferative, transitional, hypertrophic and erosion zones. 3) Distribution of the labeled nuclei with [methyl- 3 H] thymidine was confined almost to the proliferative and transitional zones. In these two zones, the cell proliferation indicated by labeling index increased about 50% in the experimental group as compared with the control. 4) Uptake of [ 3 H] proline was the highest in the transitional zone followed by the proliferative zone. Uptake of [ 3 H] proline indicated by grain counts/2000 μm 2 increased about 50% in the transitional zone and about 30% in the proliferative zone respectively in the experimental group as compared with control. 5) The septo-premaxillary ligament named by Latham (1970) was found also in rats to arise from the perichondrium of antero-inferior portion of the nasal septum, to course postero-inferiorly and to insert into the anterior portion of the premaxillary bones. Uptake of [ 3 H] proline indicated by grain counts/400 μm 2 increased about 30% in the area of insertion of the ligament in the experimental group as compared with the control. (J.P.N.)

Automated Expansion of Primary Human T Cells in Scalable and Cell-Friendly Hydrogel Microtubes for Adoptive Immunotherapy.

Science.gov (United States)

Lin, Haishuang; Li, Qiang; Wang, Ou; Rauch, Jack; Harm, Braden; Viljoen, Hendrik J; Zhang, Chi; Van Wyk, Erika; Zhang, Chi; Lei, Yuguo

2018-05-11

Adoptive immunotherapy is a highly effective strategy for treating many human cancers, such as melanoma, cervical cancer, lymphoma, and leukemia. Here, a novel cell culture technology is reported for expanding primary human T cells for adoptive immunotherapy. T cells are suspended and cultured in microscale alginate hydrogel tubes (AlgTubes) that are suspended in the cell culture medium in a culture vessel. The hydrogel tubes protect cells from hydrodynamic stresses and confine the cell mass less than 400 µm (in radial diameter) to ensure efficient mass transport, creating a cell-friendly microenvironment for growing T cells. This system is simple, scalable, highly efficient, defined, cost-effective, and compatible with current good manufacturing practices. Under optimized culture conditions, the AlgTubes enable culturing T cells with high cell viability, low DNA damage, high growth rate (≈320-fold expansion over 14 days), high purity (≈98% CD3+), and high yield (≈3.2 × 10 8 cells mL -1 hydrogel). All offer considerable advantages compared to current T cell culturing approaches. This new culture technology can significantly reduce the culture volume, time, and cost, while increasing the production. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Homogenous Population Genetic Structure of the Non-Native Raccoon Dog (Nyctereutes procyonoides) in Europe as a Result of Rapid Population Expansion

Science.gov (United States)

Drygala, Frank; Korablev, Nikolay; Ansorge, Hermann; Fickel, Joerns; Isomursu, Marja; Elmeros, Morten; Kowalczyk, Rafał; Baltrunaite, Laima; Balciauskas, Linas; Saarma, Urmas; Schulze, Christoph; Borkenhagen, Peter; Frantz, Alain C.

2016-01-01

The extent of gene flow during the range expansion of non-native species influences the amount of genetic diversity retained in expanding populations. Here, we analyse the population genetic structure of the raccoon dog (Nyctereutes procyonoides) in north-eastern and central Europe. This invasive species is of management concern because it is highly susceptible to fox rabies and an important secondary host of the virus. We hypothesized that the large number of introduced animals and the species’ dispersal capabilities led to high population connectivity and maintenance of genetic diversity throughout the invaded range. We genotyped 332 tissue samples from seven European countries using 16 microsatellite loci. Different algorithms identified three genetic clusters corresponding to Finland, Denmark and a large ‘central’ population that reached from introduction areas in western Russia to northern Germany. Cluster assignments provided evidence of long-distance dispersal. The results of an Approximate Bayesian Computation analysis supported a scenario of equal effective population sizes among different pre-defined populations in the large central cluster. Our results are in line with strong gene flow and secondary admixture between neighbouring demes leading to reduced genetic structuring, probably a result of its fairly rapid population expansion after introduction. The results presented here are remarkable in the sense that we identified a homogenous genetic cluster inhabiting an area stretching over more than 1500km. They are also relevant for disease management, as in the event of a significant rabies outbreak, there is a great risk of a rapid virus spread among raccoon dog populations. PMID:27064784

Limit-dilution assay and clonal expansion of all T cells capable of proliferation

Energy Technology Data Exchange (ETDEWEB)

Chen, W.F.; Wilson, A.; Scollay, R.; Shortman, K. (Walter and Eliza Hall Inst. of Medical Research, Parkville (Australia))

1982-08-13

A limit-dilution microculture system is presented in which almost all mature T cells, cultured at a level of about 1 cell/well, grow and expand to clones averaging 60,000 cells over an 8-9 day period. Cloning efficiency is 70-100%, so the set of expanded clones is representative of the starting T-cell population. T cells of all Lyt phenotypes form clones of progeny cells. The system involves culture in flat-bottom microtitre trays, in the presence of concanavalin A as the initiating stimulus, together with appropriately irradiated spleen filler cells and a supplementary source of soluble T cell growth factors. The resultant clones may be screened for cytolytic function, as described in the accompanying paper. The system may be used to assay the level of T cells capable of expansion or precursor function (PTL-p) by using (/sup 3/H)TdR uptake as a readout for the presence or absence of proliferating clones. Analysis of the frequency of positive cultures shows a good fit to the expected Poisson distribution, with no evidence of complicating suppressor or helper effects.

Limit-dilution assay and clonal expansion of all T cells capable of proliferation

International Nuclear Information System (INIS)

Chen, W.-F.; Wilson, A.; Scollay, R.; Shortman, K.

1982-01-01

A limit-dilution microculture system is presented in which almost all mature T cells, cultured at a level of about 1 cell/well, grow and expand to clones averaging 60,000 cells over an 8-9 day period. Cloning efficiency is 70-100%, so the set of expanded clones is representative of the starting T-cell population. T cells of all Lyt phenotypes form clones of progeny cells. The system involves culture in flat-bottom microtitre trays, in the presence of concanavalin A as the initiating stimulus, together with appropriately irradiated spleen filler cells and a supplementary source of soluble T cell growth factors. The resultant clones may be screened for cytolytic function, as described in the accompanying paper. The system may be used to assay the level of T cells capable of expansion or precursor function (PTL-p) by using [ 3 H]TdR uptake as a readout for the presence or absence of proliferating clones. Analysis of the frequency of positive cultures shows a good fit to the expected Poisson distribution, with no evidence of complicating suppressor or helper effects. (Auth.)

The influence of capillarization on satellite cell pool expansion and activation following exercise-induced muscle damage in healthy young men.

Science.gov (United States)

Nederveen, Joshua P; Joanisse, Sophie; Snijders, Tim; Thomas, Aaron C Q; Kumbhare, Dinesh; Parise, Gianni

2018-03-15

Skeletal muscle stem cells (satellite cells) play a crucial role in repair and remodelling of muscle in response to exercise. Satellite cells are in close spatial proximity to muscle capillaries and therefore may be influenced by them. In this study, we describe the activation and expansion of the satellite cell pool in response to eccentric contraction-induced muscle damage in individuals with significantly different levels of muscle capillarization. Individuals with greater capillarization and capacity for muscle perfusion demonstrated enhanced activation and/or expansion of the satellite cell pool allowing for an accelerated recovery of muscle function. These results provide insight into the critical relationship between muscle capillarization and satellite cells during skeletal muscle repair. Factors that determine the skeletal muscle satellite cell (SC) response remain incompletely understood. It is known, however, that SC activation status is closely related to the anatomical relationship between SCs and muscle capillaries. We investigated the impact of muscle fibre capillarization on the expansion and activation status of SCs following a muscle-damaging exercise protocol in healthy young men. Twenty-nine young men (21 ± 0.5 years) performed 300 unilateral eccentric contractions (180 deg s -1 ) of the knee extensors. Percutaneous muscle biopsies from the vastus lateralis and blood samples from the antecubital vein were taken prior to (Pre) exercise and at 6, 24, 72 and 96 h of post-exercise recovery. A comparison was made between subjects who had a relative low mixed muscle capillary-to-fibre perimeter exchange index (CFPE; Low group) and high mixed muscle CFPE index (High group) at baseline. Type I and type II muscle fibre size, myonuclear content, capillarization, and SC response were determined via immunohistochemistry. Overall, there was a significant correlation (r = 0.39; P < 0.05) between the expansion of SC content (change in total Pax7

Strategic camouflage treatment of skeletal Class III malocclusion (mandibular prognathism) using bone-borne rapid maxillary expansion and mandibular anterior subapical osteotomy.

Science.gov (United States)

Seo, Yu-Jin; Lin, Lu; Kim, Seong-Hun; Chung, Kyu-Rhim; Nelson, Gerald

2016-01-01

This case report presents the camouflage treatment that successfully improved the facial profile of a patient with a skeletal Class III malocclusion using bone-borne rapid maxillary expansion and mandibular anterior subapical osteotomy. The patient was an 18-year-old woman with chief complaints of crooked teeth and a protruded jaw. Camouflage treatment was chosen because she rejected orthognathic surgery under general anesthesia. A hybrid type of bone-borne rapid maxillary expander with palatal mini-implants was used to correct the transverse discrepancy, and a mandibular anterior subapical osteotomy was conducted to achieve proper overjet with normal incisal inclination and to improve her lip and chin profile. As a result, a Class I occlusion with a favorable inclination of the anterior teeth and a good esthetic profile was achieved with no adverse effects. Therefore, the hybrid type of bone-borne rapid maxillary expander and a mandibular anterior subapical osteotomy can be considered effective camouflage treatment of a skeletal Class III malocclusion, providing improved inclination of the dentition and lip profile. Copyright © 2016 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.

Theoretical calculation of rapid x-ray transients and radius expansion

International Nuclear Information System (INIS)

Starrfield, S.; Sparks, W.; Truran, J.; Kenyon, S.

1984-01-01

We present the results of a calculation of a thermonuclear runaway on a 10 km neutron star which produced a precursor, radius expansion, and after the envelope had begun to shrink, a seconds x-ray burst about 2500 second later. Although such an event has not yet been observed, decreasing the initial envelope mass should bring the calculations into better agreement with the observations

Brain and muscle Arnt-like 1 promotes skeletal muscle regeneration through satellite cell expansion

Energy Technology Data Exchange (ETDEWEB)

Chatterjee, Somik [Center for Diabetes Research, Department of Medicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Yin, Hongshan [Center for Diabetes Research, Department of Medicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Department of Cardiovascular Medicine, Third Affiliated Hospital, Hebei Medical University, Shijiazhuang 050051, Hebei (China); Nam, Deokhwa [Center for Diabetes Research, Department of Medicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Li, Yong [Department of Pediatric Surgery, Center for Stem Cell Research and Regenerative Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030 (United States); Ma, Ke, E-mail: kma@houstonmethodist.org [Center for Diabetes Research, Department of Medicine, Houston Methodist Research Institute, Houston, TX 77030 (United States)

2015-02-01

Circadian clock is an evolutionarily conserved timing mechanism governing diverse biological processes and the skeletal muscle possesses intrinsic functional clocks. Interestingly, although the essential clock transcription activator, Brain and muscle Arnt-like 1 (Bmal1), participates in maintenance of muscle mass, little is known regarding its role in muscle growth and repair. In this report, we investigate the in vivo function of Bmal1 in skeletal muscle regeneration using two muscle injury models. Bmal1 is highly up-regulated by cardiotoxin injury, and its genetic ablation significantly impairs regeneration with markedly suppressed new myofiber formation and attenuated myogenic induction. A similarly defective regenerative response is observed in Bmal1-null mice as compared to wild-type controls upon freeze injury. Lack of satellite cell expansion accounts for the regeneration defect, as Bmal1{sup −/−} mice display significantly lower satellite cell number with nearly abolished induction of the satellite cell marker, Pax7. Furthermore, satellite cell-derived primary myoblasts devoid of Bmal1 display reduced growth and proliferation ex vivo. Collectively, our results demonstrate, for the first time, that Bmal1 is an integral component of the pro-myogenic response that is required for muscle repair. This mechanism may underlie its role in preserving adult muscle mass and could be targeted therapeutically to prevent muscle-wasting diseases. - Highlights: • Bmal1 is highly inducible by muscle injury and myogenic stimuli. • Genetic ablation of Bmal1 significantly impairs muscle regeneration. • Bmal1 promotes satellite cell expansion during muscle regeneration. • Bmal1-deficient primary myoblasts display attenuated growth and proliferation.

Brain and muscle Arnt-like 1 promotes skeletal muscle regeneration through satellite cell expansion

International Nuclear Information System (INIS)

Chatterjee, Somik; Yin, Hongshan; Nam, Deokhwa; Li, Yong; Ma, Ke

2015-01-01

Circadian clock is an evolutionarily conserved timing mechanism governing diverse biological processes and the skeletal muscle possesses intrinsic functional clocks. Interestingly, although the essential clock transcription activator, Brain and muscle Arnt-like 1 (Bmal1), participates in maintenance of muscle mass, little is known regarding its role in muscle growth and repair. In this report, we investigate the in vivo function of Bmal1 in skeletal muscle regeneration using two muscle injury models. Bmal1 is highly up-regulated by cardiotoxin injury, and its genetic ablation significantly impairs regeneration with markedly suppressed new myofiber formation and attenuated myogenic induction. A similarly defective regenerative response is observed in Bmal1-null mice as compared to wild-type controls upon freeze injury. Lack of satellite cell expansion accounts for the regeneration defect, as Bmal1 −/− mice display significantly lower satellite cell number with nearly abolished induction of the satellite cell marker, Pax7. Furthermore, satellite cell-derived primary myoblasts devoid of Bmal1 display reduced growth and proliferation ex vivo. Collectively, our results demonstrate, for the first time, that Bmal1 is an integral component of the pro-myogenic response that is required for muscle repair. This mechanism may underlie its role in preserving adult muscle mass and could be targeted therapeutically to prevent muscle-wasting diseases. - Highlights: • Bmal1 is highly inducible by muscle injury and myogenic stimuli. • Genetic ablation of Bmal1 significantly impairs muscle regeneration. • Bmal1 promotes satellite cell expansion during muscle regeneration. • Bmal1-deficient primary myoblasts display attenuated growth and proliferation

The genetic structure of Turnip mosaic virus population reveals the rapid expansion of a new emergent lineage in China.

Science.gov (United States)

Li, Xiangdong; Zhu, Tiansheng; Yin, Xiao; Zhang, Chengling; Chen, Jia; Tian, Yanping; Liu, Jinliang

2017-08-29

Turnip mosaic virus (TuMV) is one of the most widespread and economically important virus infecting both crop and ornamental species of the family Brassicaceae. TuMV isolates can be classified to five phylogenetic lineages, basal-B, basal-BR, Asian-BR, world-B and Orchis. To understand the genetic structure of TuMV from radish in China, the 3'-terminal genome of 90 TuMV isolates were determined and analyzed with other available Chinese isolates. The results showed that the Chinese TuMV isolates from radish formed three groups: Asian-BR, basal-BR and world-B. More than half of these isolates (52.54%) were clustered to basal-BR group, and could be further divided into three sub-groups. The TuMV basal-BR isolates in the sub-groups I and II were genetically homologous with Japanese ones, while those in sub-group III formed a distinct lineage. Sub-populations of TuMV basal-BR II and III were new emergent and in a state of expansion. The Chinese TuMV radish populations were under negative selection. Gene flow between TuMV populations from Tai'an, Weifang and Changchun was frequent. The genetic structure of Turnip mosaic virus population reveals the rapid expansion of a new emergent lineage in China.

Optimising cell aggregate expansion in a perfused hollow fibre bioreactor via mathematical modelling.

KAUST Repository

Chapman, Lloyd A C; Shipley, Rebecca J; Whiteley, Jonathan P; Ellis, Marianne J; Byrne, Helen M; Waters, Sarah L

2014-01-01

, the higher fluid shear stress can lead to cell death. For a given cell type and initial aggregate distribution, the operating parameters that give the most rapid overall growth can be identified from simulations. For example, when aggregates of rat

World NGL markets continue rapid expansion

International Nuclear Information System (INIS)

Otto, K.; Gist, R.; Whitley, C.; Haun, R.

1998-01-01

The international LPG industry has expanded rapidly during the 1990s and undergone significant changes. LPG consumption has expanded at nearly twice the rate of world petroleum demand. In particular, LPG use in residential and commercial markets has more than doubled in many developing countries. Markets for LPG and other petroleum products have been opened in many countries, accelerating demand growth and creating investment opportunities in all downstream segments. This has led to an overall strengthening of global LPG pricing and the development of many new export gas-processing projects. The paper discusses world LPG demand in residential and commercial markets and in petrochemicals, world LPG supply, regional increases, international trade, the US situation in natural gas, NGL supply, and NGL demand

Closely related T-memory stem cells correlate with in vivo expansion of CAR.CD19-T cells and are preserved by IL-7 and IL-15

Science.gov (United States)

Xu, Yang; Zhang, Ming; Ramos, Carlos A.; Durett, April; Liu, Enli; Dakhova, Olga; Liu, Hao; Creighton, Chad J.; Gee, Adrian P.; Heslop, Helen E.; Rooney, Cliona M.; Savoldo, Barbara

2014-01-01

Adoptive transfer of T lymphocytes expressing a CD19-specific chimeric antigen receptor (CAR.CD19) induces complete tumor regression in patients with lymphoid malignancies. Although in vivo persistence of CAR-T cells correlates with clinical responses, it remains unknown whether specific cell subsets within the CAR–T-cell product correlate with their subsequent in vivo expansion and persistence. We analyzed 14 patients with B-cell malignancies infused with autologous CAR.CD19-redirected T cells expanded ex vivo using IL-2, and found that their in vivo expansion only correlated with the frequency within the infused product of a CD8+CD45RA+CCR7+ subset, whose phenotype is closest to “T-memory stem cells.” Preclinical models showed that increasing the frequency of CD8+CD45RA+CCR7+ CAR-T cells in the infused line by culturing the cells with IL-7 and IL-15 produced greater antitumor activity of CAR-T cells mediated by increased resistance to cell death, following repetitive encounters with the antigen, while preserving their migration to secondary lymphoid organs. This trial was registered at www.clinicaltrials.gov as #NCT00586391 and #NCT00709033. PMID:24782509

Diclofenac in Arabidopsis cells: Rapid formation of conjugates.

Science.gov (United States)

Fu, Qiuguo; Ye, Qingfu; Zhang, Jianbo; Richards, Jaben; Borchardt, Dan; Gan, Jay

2017-03-01

Pharmaceutical and personal care products (PPCPs) are continuously introduced into the soil-plant system, through practices such as agronomic use of reclaimed water and biosolids containing these trace contaminants. Plants may accumulate PPCPs from soil, serving as a conduit for human exposure. Metabolism likely controls the final accumulation of PPCPs in plants, but is in general poorly understood for emerging contaminants. In this study, we used diclofenac as a model compound, and employed 14 C tracing, and time-of-flight (TOF) and triple quadruple (QqQ) mass spectrometers to unravel its metabolism pathways in Arabidopsis thaliana cells. We further validated the primary metabolites in Arabidopsis seedlings. Diclofenac was quickly taken up into A. thaliana cells. Phase I metabolism involved hydroxylation and successive oxidation and cyclization reactions. However, Phase I metabolites did not accumulate appreciably; they were instead rapidly conjugated with sulfate, glucose, and glutamic acid through Phase II metabolism. In particular, diclofenac parent was directly conjugated with glutamic acid, with acyl-glutamatyl-diclofenac accounting for >70% of the extractable metabolites after 120-h incubation. In addition, at the end of incubation, >40% of the spiked diclofenac was in the non-extractable form, suggesting extensive sequestration into cell matter. The rapid formation of non-extractable residue and dominance of diclofenac-glutamate conjugate uncover previously unknown metabolism pathways for diclofenac. In particular, the rapid conjugation of parent highlights the need to consider conjugates of emerging contaminants in higher plants, and their biological activity and human health implications. Copyright © 2016 Elsevier Ltd. All rights reserved.

Rapid expansion and pseudo spectral implementation for reverse time migration in VTI media

KAUST Repository

Pestana, Reynam C

2012-04-24

In isotropic media, we use the scalar acoustic wave equation to perform reverse time migration (RTM) of the recorded pressure wavefield data. In anisotropic media, P- and SV-waves are coupled, and the elastic wave equation should be used for RTM. For computational efficiency, a pseudo-acoustic wave equation is often used. This may be solved using a coupled system of second-order partial differential equations. We solve these using a pseudo spectral method and the rapid expansion method (REM) for the explicit time marching. This method generates a degenerate SV-wave in addition to the P-wave arrivals of interest. To avoid this problem, the elastic wave equation for vertical transversely isotropic (VTI) media can be split into separate wave equations for P- and SV-waves. These separate wave equations are stable, and they can be effectively used to model and migrate seismic data in VTI media where |ε- δ| is small. The artifact for the SV-wave has also been removed. The independent pseudo-differential wave equations can be solved one for each mode using the pseudo spectral method for the spatial derivatives and the REM for the explicit time advance of the wavefield. We show numerically stable and high-resolution modeling and RTM results for the pure P-wave mode in VTI media. © 2012 Sinopec Geophysical Research Institute.

Rapid expansion and pseudo spectral implementation for reverse time migration in VTI media

KAUST Repository

Pestana, Reynam C; Ursin, Bjø rn; Stoffa, Paul L

2012-01-01

In isotropic media, we use the scalar acoustic wave equation to perform reverse time migration (RTM) of the recorded pressure wavefield data. In anisotropic media, P- and SV-waves are coupled, and the elastic wave equation should be used for RTM. For computational efficiency, a pseudo-acoustic wave equation is often used. This may be solved using a coupled system of second-order partial differential equations. We solve these using a pseudo spectral method and the rapid expansion method (REM) for the explicit time marching. This method generates a degenerate SV-wave in addition to the P-wave arrivals of interest. To avoid this problem, the elastic wave equation for vertical transversely isotropic (VTI) media can be split into separate wave equations for P- and SV-waves. These separate wave equations are stable, and they can be effectively used to model and migrate seismic data in VTI media where |ε- δ| is small. The artifact for the SV-wave has also been removed. The independent pseudo-differential wave equations can be solved one for each mode using the pseudo spectral method for the spatial derivatives and the REM for the explicit time advance of the wavefield. We show numerically stable and high-resolution modeling and RTM results for the pure P-wave mode in VTI media. © 2012 Sinopec Geophysical Research Institute.

A simulation environment for dry-expansion evaporators with application to the design of autotuning control algorithms for electronic expansion valves

Energy Technology Data Exchange (ETDEWEB)

Beghi, Alessandro [Dipartimento di Ingegneria dell' Informazione, Universita di Padova, via Gradenigo 6/B, I-35131 Padova (Italy); Cecchinato, Luca [Dipartimento di Fisica Tecnica, Universita di Padova, via Venezia 1, I-35131 Padova (Italy)

2009-11-15

In this paper some results of a research project aimed at deriving high-performance, adaptive control algorithms for electronic expansion valves (EEVs) to be used in finned-coiled, dry-expansion evaporators for refrigeration systems are reported. With the aim of developing a software environment that can be used for controller design, rapid prototyping, optimization of data collection and test design, virtual prototyping approach to design was adopted. The development of a distributed dynamic simulation model of the evaporator coupled with an electronic expansion valve, and its use for deriving autotuning PID control algorithms is described. Experimental results confirm the effectiveness of this kind of approach. (author)

Rapid lentiviral transduction preserves the engraftment potential of Fanca(-/-) hematopoietic stem cells.

Science.gov (United States)

Müller, Lars U W; Milsom, Michael D; Kim, Mi-Ok; Schambach, Axel; Schuesler, Todd; Williams, David A

2008-06-01

Fanconi anemia (FA) is a rare recessive syndrome, characterized by congenital anomalies, bone marrow failure, and predisposition to cancer. Two earlier clinical trials utilizing gamma-retroviral vectors for the transduction of autologous FA hematopoietic stem cells (HSCs) required extensive in vitro manipulation and failed to achieve detectable long-term engraftment of transduced HSCs. As a strategy for minimizing ex vivo manipulation, we investigated the use of a "rapid" lentiviral transduction protocol in a murine Fanca(-/-) model. Importantly, while this and most murine models of FA fail to completely mimic the human hematopoietic phenotype, we observed a high incidence of HSC transplant engraftment failure and low donor chimerism after conventional transduction (CT) of Fanca(-/-) donor cells. In contrast, rapid transduction (RT) of Fanca(-/-) HSCs preserved engraftment to the level achieved in wild-type cells, resulting in long-term multilineage engraftment of gene-modified cells. We also demonstrate the correction of the characteristic hypersensitivity of FA cells against the cross-linking agent mitomycin C (MMC), and provide evidence for the advantage of using pharmacoselection as a means of further increasing gene-modified cells after RT. Collectively, these data support the use of rapid lentiviral transduction for gene therapy in FA.

Avaliação cefalométrica das alterações verticais e anteroposteriores associadas ao uso do expansor maxilar com cobertura oclusal Cephalometric evaluation of vertical and anteroposterior changes associated with the use of bonded rapid maxillary expansion appliance

Directory of Open Access Journals (Sweden)

Moara De Rossi

2010-06-01

Full Text Available INTRODUÇÃO: os aparelhos expansores maxilares com cobertura oclusal têm sido sugeridos para controlar o aumento na dimensão vertical da face após a expansão rápida da maxila, porém ainda não há um consenso na literatura sobre seus reais efeitos. OBJETIVO: o objetivo deste trabalho foi avaliar as alterações cefalométricas verticais e anteroposteriores associadas à expansão da maxila realizada com o aparelho expansor com cobertura oclusal. MÉTODOS: a amostra foi composta por 25 crianças, de ambos os gêneros, com idades entre 6 e 10 anos, portadoras de mordida cruzada posterior esquelética. Após a expansão maxilar, o próprio aparelho expansor foi utilizado como contenção fixa. Foram analisadas telerradiografias em norma lateral tomadas antes do início do tratamento e após a remoção do aparelho expansor. CONCLUSÃO: com base nos resultados, pôde-se concluir que o uso do aparelho expansor com cobertura oclusal não alterou significativamente as medidas cefalométricas verticais e anteroposteriores das crianças.INTRODUCTION: Bonded rapid maxillary expansion appliances have been suggested to control increases in the vertical dimension of the face after rapid maxillary expansion but there is still no consensus in the literature concerning its actual effectiveness. OBJECTIVE: The purpose of this study was to evaluate the vertical and anteroposterior cephalometric changes associated with maxillary expansion performed using bonded rapid maxillary expansion appliances. METHODS: The sample consisted of 25 children of both genders, aged between 6 and 10 years old, with skeletal posterior crossbite. After maxillary expansion, the expansion appliance itself was used for fixed retention. Were analyzed lateral teleradiographs taken prior to treatment onset and after removal of the expansion appliance. CONCLUSION: Based on the results, it can be concluded that the use of bonded rapid maxillary expansion appliance did not significantly

Study of instantaneous unsteady heat transfer in a rapid compression-expansion machine using zero dimensional k- ε turbulence model

International Nuclear Information System (INIS)

Bakhshan, Y.; Karim, G. A.; Mansouri, S. H.

2003-01-01

In this investigation, the instantaneous unsteady heat transfer within a pneumatically driven rapid compression-expansion machine that offers simple, well-controlled and known boundary conditions was studied. Values of the instantaneous apparent overall heat flux from the cylinder gas to the wall surfaces were calculated using a thermodynamics analysis of the experimentally measured pressure and volume temporal development. Corresponding heat flux values were also calculated through the application of a zero-dimensional k- ε turbulence model the characteristic velocity is a contribution of turbulence kinetic energy, mean kinetic energy of charged air into cylinder and piston motion for the calculation of Reynolds, Nusselt and Prandtl numbers. Comparison of the zero-dimensional k- ε turbulence model prediction with experimental data shows good agreement for all compression ratios

Nephron progenitor cell death elicits a limited compensatory response associated with interstitial expansion in the neonatal kidney

Directory of Open Access Journals (Sweden)

Sree Deepthi Muthukrishnan

2018-01-01

Full Text Available The final nephron number in an adult kidney is regulated by nephron progenitor cell availability and collecting duct branching in the fetal period. Fetal environmental perturbations that cause reductions in cell numbers in these two compartments result in low nephron endowment. Previous work has shown that maternal dietary factors influence nephron progenitor cell availability, with both caloric restriction and protein deprivation leading to reduced cell numbers through apoptosis. In this study, we evaluate the consequences of inducing nephron progenitor cell death on progenitor niche dynamics and on nephron endowment. Depletion of approximately 40% of nephron progenitor cells by expression of diphtheria toxin A at embryonic day 15 in the mouse results in 10-20% nephron reduction in the neonatal period. Analysis of cell numbers within the progenitor cell pool following induction of apoptosis reveals a compensatory response in which surviving progenitor cells increase their proliferation and replenish the niche. The proliferative response is temporally associated with infiltration of macrophages into the nephrogenic zone. Colony stimulating factor 1 (CSF1 has a mitogenic effect on nephron progenitor cells, providing a potential explanation for the compensatory proliferation. However, CSF1 also promotes interstitial cell proliferation, and the compensatory response is associated with interstitial expansion in recovering kidneys which can be pharmacologically inhibited by treatment with clodronate liposomes. Our findings suggest that the fetal kidney employs a macrophage-dependent compensatory regenerative mechanism to respond to acute injury caused by death of nephron progenitor cells, but that this regenerative response is associated with neonatal interstitial expansion.

NOTCH-Mediated Maintenance and Expansion of Human Bone Marrow Stromal/Stem Cells: A Technology Designed for Orthopedic Regenerative Medicine.

Science.gov (United States)

Dong, Yufeng; Long, Teng; Wang, Cuicui; Mirando, Anthony J; Chen, Jianquan; O'Keefe, Regis J; Hilton, Matthew J

2014-12-01

Human bone marrow-derived stromal/stem cells (BMSCs) have great therapeutic potential for treating skeletal disease and facilitating skeletal repair, although maintaining their multipotency and expanding these cells ex vivo have proven difficult. Because most stem cell-based applications to skeletal regeneration and repair in the clinic would require large numbers of functional BMSCs, recent research has focused on methods for the appropriate selection, expansion, and maintenance of BMSC populations during long-term culture. We describe here a novel biological method that entails selection of human BMSCs based on NOTCH2 expression and activation of the NOTCH signaling pathway in cultured BMSCs via a tissue culture plate coated with recombinant human JAGGED1 (JAG1) ligand. We demonstrate that transient JAG1-mediated NOTCH signaling promotes human BMSC maintenance and expansion while increasing their skeletogenic differentiation capacity, both ex vivo and in vivo. This study is the first of its kind to describe a NOTCH-mediated methodology for the maintenance and expansion of human BMSCs and will serve as a platform for future clinical or translational studies aimed at skeletal regeneration and repair. ©AlphaMed Press.

Thermal expansion of doped lanthanum gallates

Indian Academy of Sciences (India)

Administrator

Since the components are in intimate mechanical contact, any stress generated due to their thermal expansion mis- match during thermal cycling could lead to catastrophic failure of the cell. The functional materials must have similar thermal expansions to avoid mechanical stresses. Hence it is useful to study the thermal ...

A bilinear elastic constitutive model applied for midpalatal suture behavior during rapid maxillary expansion

Directory of Open Access Journals (Sweden)

Larissa Carvalho Trojan Serpe

Full Text Available Introduction : This study aims to evaluate the influence of the biomechanical behavior of the midpalatal suture (MPS during the rapid maxillary expansion (RME when modeled by the Finite Element Method. Methods Four simulation alternatives are discussed and, for each analysis, the suture is considered as a functional unit with a different mechanical behavior: (i without MPS elements, (ii MPS with Young's modulus (E equal to 1 MPa, (ii MPS with E equal to 0.01 MPa and (iv MPS with bilinear elastic behavior. Results The stress analysis showed that, when MPS is not considered in the model, stress peaks are reduced in magnitude and their distribution is restricted to a smaller area when compared to the model with the inclusion of MPS (E=1 MPa. The increased suture stiffness also has a direct influence on MPS displacements after 30 expander activations. Conclusion The consideration of the MPS in RME computer models influences greatly the calculated displacements between the suture bone ends, even as the stress levels in maxillary structures. Furthermore, as proposed for the described model, the elastic bilinear behavior assigned to MPS allows coherent prediction of stresses and displacements results, being a good representation for this suture overall behavior.

Changes in occlusal relationships in mixed dentition patients treated with rapid maxillary expansion. A prospective clinical study.

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McNamara, James A; Sigler, Lauren M; Franchi, Lorenzo; Guest, Susan S; Baccetti, Tiziano

2010-03-01

To prospectively measure occlusal changes in mixed dentition patients who underwent a standardized early expansion protocol. The treatment sample consisted of 500 patients who were assigned to three groups according to molar relationship: Class I (n = 204), end-to-end (n = 166), and Class II (n = 130). All patients were treated with a bonded rapid maxillary expander (RME) followed by a removable maintenance plate and a transpalatal arch. Mean age at the start of treatment was 8.8 years (T(1)), with a pre-phase 2 treatment cephalogram (T(2)) taken 3.7 years later. The control sample consisted of the cephalometric records of 188 untreated subjects (Class 1, n = 79; end-to-end, n = 51; Class II, n = 58). The largest change in molar relationship was noted when the Class II treatment group (1.8 mm) was compared with the matched control group (0.3 mm). A positive change was seen in 81% of the Class II treatment group, with almost half of the group improving by > or = 2.0 mm. The end-to-end treatment group had a positive change of 1.4 mm, compared with a control value of 0.6 mm, and the Class I group of about 1 mm compared with controls, who remained unchanged (0.1 mm). Skeletal changes were not significant when any of the groups were compared with controls. The expansion protocol had a significantly favorable effect on the sagittal occlusal relationships of Class II, end-to-end, and Class I patients treated in the early mixed dentition.

In vivo Expansion of Naïve CD4+CD25high FOXP3+ Regulatory T Cells in Patients with Colorectal Carcinoma after IL-2 Administration

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Beyer, Marc; Schumak, Beatrix; Weihrauch, Martin R.; Andres, Bettina; Giese, Thomas; Endl, Elmar; Knolle, Percy A.; Classen, Sabine; Limmer, Andreas; Schultze, Joachim L.

2012-01-01

Regulatory T cells (Treg cells) are increased in context of malignancies and their expansion can be correlated with higher disease burden and decreased survival. Initially, interleukin 2 (IL-2) has been used as T-cell growth factor in clinical vaccination trials. In murine models, however, a role of IL-2 in development, differentiation, homeostasis, and function of Treg cells was established. In IL-2 treated cancer patients a further Treg-cell expansion was described, yet, the mechanism of expansion is still elusive. Here we report that functional Treg cells of a naïve phenotype - as determined by CCR7 and CD45RA expression - are significantly expanded in colorectal cancer patients. Treatment of 15 UICC stage IV colorectal cancer patients with IL-2 in a phase I/II peptide vaccination trial further enlarges the already increased naïve Treg-cell pool. Higher frequencies of T-cell receptor excision circles in naïve Treg cells indicate IL-2 dependent thymic generation of naïve Treg cells as a mechanism leading to increased frequencies of Treg cells post IL-2 treatment in cancer patients. This finding could be confirmed in naïve murine Treg cells after IL-2 administration. These results point to a more complex regulation of Treg cells in context of IL-2 administration. Future strategies therefore might aim at combining IL-2 therapy with novel strategies to circumvent expansion and differentiation of naïve Treg cells. PMID:22276195

Wnt3a protein reduces growth factor-driven expansion of human hematopoietic stem and progenitor cells in serum-free cultures

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L.E. Duinhouwer (Lucia); N. Tüysüz (Nesrin); E.J. Rombouts (Elwin); M.N.D. Ter Borg (Mariëtte N. D.); E. Mastrobattista; J. Spanholtz (Jan); J.J. Cornelissen (Jan); D. ten Berge (Derk); E. Braakman (Eric)

2015-01-01

textabstractAbstract Ex vivo expansion of hematopoietic stem and progenitor cells (HSPC) is a promising approach to improve insufficient engraftment after umbilical cord blood stem cell transplantation (UCB-SCT). Although culturing HSPC with hematopoietic cytokines results in

TET proteins regulate the lineage specification and TCR-mediated expansion of iNKT cells.

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Tsagaratou, Ageliki; González-Avalos, Edahí; Rautio, Sini; Scott-Browne, James P; Togher, Susan; Pastor, William A; Rothenberg, Ellen V; Chavez, Lukas; Lähdesmäki, Harri; Rao, Anjana

2017-01-01

TET proteins oxidize 5-methylcytosine in DNA to 5-hydroxymethylcytosine and other oxidation products. We found that simultaneous deletion of Tet2 and Tet3 in mouse CD4 + CD8 + double-positive thymocytes resulted in dysregulated development and proliferation of invariant natural killer T cells (iNKT cells). Tet2-Tet3 double-knockout (DKO) iNKT cells displayed pronounced skewing toward the NKT17 lineage, with increased DNA methylation and impaired expression of genes encoding the key lineage-specifying factors T-bet and ThPOK. Transfer of purified Tet2-Tet3 DKO iNKT cells into immunocompetent recipient mice resulted in an uncontrolled expansion that was dependent on the nonclassical major histocompatibility complex (MHC) protein CD1d, which presents lipid antigens to iNKT cells. Our data indicate that TET proteins regulate iNKT cell fate by ensuring their proper development and maturation and by suppressing aberrant proliferation mediated by the T cell antigen receptor (TCR).

Association of Monoclonal Expansion of Epstein-Barr Virus-Negative CD158a+ NK Cells Secreting Large Amounts of Gamma Interferon with Hemophagocytic Lymphohistiocytosis▿

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López-Álvarez, María R.; Martínez-Sánchez, María V.; Salgado-Cecilia, María G.; Campillo, José A.; Heine-Suñer, Damian; Villar-Permuy, Florentina; Fuster, José L.; Bas, Águeda; Gil-Herrera, Juana; Muro, Manuel; García-Alonso, Ana M.; Álvarez-López, María R.; Minguela, Alfredo

2009-01-01

We report the first case of hemophagocytic lymphohistiocytosis (HLH) induced by the monoclonal expansion of Epstein-Barr virus (EBV)-negative NK cells. Consanguinity of the patient's parents made it necessary to discard familial HLH in the patient and her sister with identical HLA markers and demonstrate that no cause other than the expansion of NK cells, which secrete high levels of gamma interferon, was inducing HLH in this patient. PMID:19020108

Modified SARME (Surgically Assisted Rapid Maxillary Expansion) in Conjunction with Orthodontic Treatment-A Case Report.

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Dahiya, Sagar; Chitra, Prasad; Rao, Sadam Srinivas; Bindra, Sukhvinder

2015-10-01

Transverse maxillary hypoplasia or maxillary constriction in conjunction with unilateral or bilateral posterior cross bites is a common finding in cleft palate patients. These situations are also commonly encountered in adults who have not had recourse to orthodontic treatment in childhood. In adults, after ossification of the mid palatal suture is complete, the accepted means of correcting transverse skeletal discrepancies is by Surgically Assisted Rapid Maxillary Expansion (SARME). The disadvantage of this technique in the Indian scenario is reduced patient acceptance and increased treatment costs. Le Fort-I down fracture and mid palatal suture sectioning requires hospitalization and increases morbidity. A case of a 21-year-old non-cleft male who presented with Class I malocclusion with transverse skeletal discrepancy and bilateral posterior cross bites is presented. A modified SAARME technique was performed without pterygomaxillary disjunction, as an outpatient procedure. The results obtained were satisfactory and the desired amount of transverse skeletal correction was achieved. The patient was discharged the same day. The technique can be used to successfully treat a large number of patients in India with maxillary skeletal transverse problems with increased predictability, reduced costs and morbidity and higher rates of acceptance.

Short-term, serum-free, static culture of cord blood-derived CD34+ cells: effects of FLT3-L and MIP-1alpha on in vitro expansion of hematopoietic progenitor cells.

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Capmany, G; Querol, S; Cancelas, J A; García, J

1999-08-01

The use of ex vivo expanded cells has been suggested as a possible means to accelerate the speed of engraftment in cord blood (CB) transplantation. The aim of this study was to fix the optimal condition for the generation of committed progenitors without affecting the stem cell compartment. Analysis of the effects of FLT3-L and MIP-1alpha when combined with SCF, IL-3 and IL-6, in short-term (6 days), serum-free expansion cultures of CB-selected CD34+ cells. An important expansion was obtained that ranged between 8-15 times for CFU-GM, 21-51 times for the BFU-E/CFU-Mix population and 11 to 30 times for CD34+ cells assessed by flow cytometry. From the combinations tested, those in which FLT3-L was present had a significant increase in the expansion of committed progenitors, while the presence of MIP-1alpha had a detrimental effect on the generation of more differentiated cells. However, stem cell candidates assessed by week 5 CAFC assay could be maintained in culture when both MIP-1a and FLT3-L were present (up to 91% recovery). This culture system was also able to expand megakaryocytic precursors as determined by the co-expression of CD34 and CD61 antigens (45-70 times), in spite of the use of cytokines non-specific for the megakaryocytic lineage. The results obtained point to the combination of SCF, IL-3, IL-6, FLT3-L and MIP-1alpha as the best suited for a pre-clinical short-term serum-free static ex vivo expansion protocol of CB CD34+ cells, since it can generate large numbers of committed progenitor cells as well as maintaining week 5 CAFC.

Convergence of hyperspherical adiabatic expansion for helium-like systems

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Abrashkevich, A.G.; Abrashkevich, D.G.; Pojda, V.Yu.; Vinitskij, S.I.; Kaschiev, M.S.; Puzynin, I.V.

1988-01-01

The convergence of hyperspherical adiabatic expansion has been studied numerically. The spectral problems arising after separation of variables are solved by the finite-difference and finite element methods. The energies of the ground and some doubly excited staes of a hydrogen ion are calculated in the six-channel approximation within the 10 -4 a.u. accuracy. Obtained results demonstrate a rapid convergence of the hyperspherical adiabatic expansion. 14 refs.; 5 tabs

Surgical retrieval, isolation and in vitro expansion of human anterior cruciate ligament-derived cells for tissue engineering applications.

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Gupta, Ashim; Sharif, Kevin; Walters, Megan; Woods, Mia D; Potty, Anish; Main, Benjamin J; El-Amin, Saadiq F

2014-04-30

Injury to the ACL is a commonly encountered problem in active individuals. Even partial tears of this intra-articular knee ligament lead to biomechanical deficiencies that impair function and stability. Current options for the treatment of partial ACL tears range from nonoperative, conservative management to multiple surgical options, such as: thermal modification, single-bundle repair, complete reconstruction, and reconstruction of the damaged portion of the native ligament. Few studies, if any, have demonstrated any single method for management to be consistently superior, and in many cases patients continue to demonstrate persistent instability and other comorbidities. The goal of this study is to identify a potential cell source for utilization in the development of a tissue engineered patch that could be implemented in the repair of a partially torn ACL. A novel protocol was developed for the expansion of cells derived from patients undergoing ACL reconstruction. To isolate the cells, minced hACL tissue obtained during ACL reconstruction was digested in a Collagenase solution. Expansion was performed using DMEM/F12 medium supplemented with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin (P/S). The cells were then stored at -80 ºC or in liquid nitrogen in a freezing medium consisting of DMSO, FBS and the expansion medium. After thawing, the hACL derived cells were then seeded onto a tissue engineered scaffold, PLAGA (Poly lactic-co-glycolic acid) and control Tissue culture polystyrene (TCPS). After 7 days, SEM was performed to compare cellular adhesion to the PLAGA versus the control TCPS. Cellular morphology was evaluated using immunofluorescence staining. SEM (Scanning Electron Microscope) micrographs demonstrated that cells grew and adhered on both PLAGA and TCPS surfaces and were confluent over the entire surfaces by day 7. Immunofluorescence staining showed normal, non-stressed morphological patterns on both surfaces. This technique is

Rapidly dissociated autologous meniscus tissue enhances meniscus healing: An in vitro study.

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Numpaisal, Piya-On; Rothrauff, Benjamin B; Gottardi, Riccardo; Chien, Chung-Liang; Tuan, Rocky S

Treatment of meniscus tears is a persistent challenge in orthopedics. Although cell therapies have shown promise in promoting fibrocartilage formation in in vitro and preclinical studies, clinical application has been limited by the paucity of autologous tissue and the need for ex vivo cell expansion. Rapid dissociation of the free edges of the anterior and posterior meniscus with subsequent implantation in a meniscus lesion may overcome these limitations. The purpose of this study was to explore the effect of rapidly dissociated meniscus tissue in enhancing neotissue formation in a radial meniscus tear, as simulated in an in vitro explant model. All experiments in this study, performed at minimum with biological triplicates, utilized meniscal tissues from hind limbs of young cows. The effect of varying collagenase concentration (0.1%, 0.2% and 0.5% w/v) and treatment duration (overnight and 30 minutes) on meniscus cell viability, organization of the extracellular matrix (ECM), and gene expression was assessed through a cell metabolism assay, microscopic examination, and quantitative real-time reverse transcription polymerase chain reaction analysis, respectively. Thereafter, an explant model of a radial meniscus tear was used to evaluate the effect of a fibrin gel seeded with one of the following: (1) fibrin alone, (2) isolated and passaged (P2) meniscus cells, (3) overnight digested tissue, and (4) rapidly dissociated tissue. The quality of in vitro healing was determined through histological analysis and derivation of an adhesion index. Rapid dissociation in 0.2% collagenase yielded cells with higher levels of metabolism than either 0.1% or 0.5% collagenase. When seeded in a three-dimensional fibrin hydrogel, both overnight digested and rapidly dissociated cells expressed greater levels of collagens type I and II than P2 meniscal cells at 1 week. At 4 and 8 weeks, collagen type II expression remained elevated only in the rapid dissociation group. Histological

Rapid shear alignment of sub-10 nm cylinder-forming block copolymer films based on thermal expansion mismatch

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Nicaise, Samuel M.; Gadelrab, Karim R.; G, Amir Tavakkoli K.; Ross, Caroline A.; Alexander-Katz, Alfredo; Berggren, Karl K.

2018-01-01

Directed self-assembly of block copolymers (BCPs) provided by shear-stress can produce aligned sub-10 nm structures over large areas for applications in integrated circuits, next-generation data storage, and plasmonic structures. In this work, we present a fast, versatile BCP shear-alignment process based on coefficient of thermal expansion mismatch of the BCP film, a rigid top coat and a substrate. Monolayer and bilayer cylindrical microdomains of poly(styrene-b-dimethylsiloxane) aligned preferentially in-plane and orthogonal to naturally-forming or engineered cracks in the top coat film, allowing for orientation control over 1 cm2 substrates. Annealing temperatures, up to 275 °C, provided low-defect alignment up to 2 mm away from cracks for rapid (<1 min) annealing times. Finite-element simulations of the stress as a function of annealing time, annealing temperature, and distance from cracks showed that shear stress during the cooling phase of the thermal annealing was critical for the observed microdomain alignment.

Rapid transcriptional pulsing dynamics of high expressing retroviral transgenes in embryonic stem cells.

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Mandy Y M Lo

Full Text Available Single cell imaging studies suggest that transcription is not continuous and occurs as discrete pulses of gene activity. To study mechanisms by which retroviral transgenes can transcribe to high levels, we used the MS2 system to visualize transcriptional dynamics of high expressing proviral integration sites in embryonic stem (ES cells. We established two ES cell lines each bearing a single copy, self-inactivating retroviral vector with a strong ubiquitous human EF1α gene promoter directing expression of mRFP fused to an MS2-stem-loop array. Transfection of MS2-EGFP generated EGFP focal dots bound to the mRFP-MS2 stem loop mRNA. These transcription foci colocalized with the transgene integration site detected by immunoFISH. Live tracking of single cells for 20 minutes detected EGFP focal dots that displayed frequent and rapid fluctuations in transcription over periods as short as 25 seconds. Similarly rapid fluctuations were detected from focal doublet signals that colocalized with replicated proviral integration sites by immunoFISH, consistent with transcriptional pulses from sister chromatids. We concluded that retroviral transgenes experience rapid transcriptional pulses in clonal ES cell lines that exhibit high level expression. These events are directed by a constitutive housekeeping gene promoter and may provide precedence for rapid transcriptional pulsing at endogenous genes in mammalian stem cells.

Rapid palatal expansion effects on mandibular transverse dimensions in unilateral posterior crossbite patients: a three-dimensional digital imaging study

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Alessandro Ugolini

2016-01-01

Full Text Available Abstract Background The purpose of this controlled study was to investigate indirect effects on mandibular arch dimensions, 1 year after rapid palatal expansion (RPE therapy. Methods Thirty-three patients in mixed dentition (mean age 8.8 years showing unilateral posterior crossbite and maxillary deficiency were treated with a RPE (Haas type cemented on the first permanent molars. Treatment protocol consisted of two turns per day until slight overcorrection of the molar transverse relationship occurred. The Haas expander was kept on the teeth as a passive retainer for an average of 6 months. Study models were taken prior (T1 and 15 months on average (T2 after expansion. A control group of 15 untreated subjects with maxillary deficiency (mean age 8.3 years was also recorded with a 12-month interval. Stone casts were digitized with a 3D scanner (3Shape, DK. Results In the treated group, both mandibular intermolar distance (+1.9 mm and mandibular molar angulation (+9° increased. Mandibular incisor angulation showed an increase of 1.9°. There was little effect on intercanine distance and canine angulation. Controls showed a reduction in transverse arch dimension and a decrease in molar and canine angulation values. Conclusions RPE protocol has indirect widening effects on the mandibular incisors and first molars.

Abnormal ion content, hydration and granule expansion of the secretory granules from cystic fibrosis airway glandular cells

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Baconnais, S.; Delavoie, F.; Zahm, J.M.; Milliot, M.; Terryn, C.; Castillon, N.; Banchet, V.; Michel, J.; Danos, O.; Merten, M.; Chinet, T.; Zierold, K.; Bonnet, N.; Puchelle, E.; Balossier, G.

2005-01-01

The absence or decreased expression of cystic fibrosis transmembrane conductance regulator (CFTR) induces increased Na + absorption and hyperabsorption of the airway surface liquid (ASL) resulting in a dehydrated and hyperviscous ASL. Although the implication of abnormal airway submucosal gland function has been suggested, the ion and water content in the Cystic Fibrosis (CF) glandular secretory granules, before exocytosis, is unknown. We analyzed, in non-CF and CF human airway glandular cell lines (MM-39 and KM4, respectively), the ion content in the secretory granules by electron probe X-ray microanalysis and the water content by quantitative dark field imaging on freeze-dried cryosections. We demonstrated that the ion content (Na + , Mg 2+ , P, S and Cl - ) is significantly higher and the water content significantly lower in secretory granules from the CF cell line compared to the non-CF cell line. Using videomicroscopy, we observed that the secretory granule expansion was deficient in CF glandular cells. Transfection of CF cells with CFTR cDNA or inhibition of non-CF cells with CFTR inh -172, respectively restored or decreased the water content and granule expansion, in parallel with changes in ion content. We hypothesize that the decreased water and increased ion content in glandular secretory granules may contribute to the dehydration and increased viscosity of the ASL in CF

Electrospun polystyrene scaffolds as a synthetic substrate for xeno-free expansion and differentiation of human induced pluripotent stem cells.

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Leong, Meng Fatt; Lu, Hong Fang; Lim, Tze Chiun; Du, Chan; Ma, Nina K L; Wan, Andrew C A

2016-12-01

The use of human induced pluripotent stem cells (hiPSCs) for clinical tissue engineering applications requires expansion and differentiation of the cells using defined, xeno-free substrates. The screening and selection of suitable synthetic substrates however, is tedious, as their performance relies on the inherent material properties. In the present work, we demonstrate an alternative concept for xeno-free expansion and differentiation of hiPSCs using synthetic substrates, which hinges on the structure-function relationship between electrospun polystyrene scaffolds (ESPS) and pluripotent stem cell growth. ESPS of differential porosity was obtained by fusing the fibers at different temperatures. The more porous, loosely fused scaffolds were found to efficiently trap the cells, leading to a large number of three-dimensional (3D) aggregates which were shown to be pluripotent colonies. Immunostaining, PCR analyses, in vitro differentiation and in vivo teratoma formation studies demonstrated that these hiPSC aggregates could be cultured for up to 10 consecutive passages (P10) with maintenance of pluripotency. Flow cytometry showed that more than 80% of the cell population stained positive for the pluripotent marker OCT4 at P1, P5 and P10. P10 cells could be differentiated to neuronal-like cells and cultured within the ESPS for up to 18months. Our results suggest the usefulness of a generic class of synthetic substrates, exemplified by ESPS, for 'trapped aggregate culture' of hiPSCs. To realize the potential of human induced pluripotent stem cells (hiPSCs) in clinical medicine, robust, xeno-free substrates for expansion and differentiation of iPSCs are required. In the existing literature, synthetic materials have been reported that meet the requirement for non-xenogeneic substrates. However, the self-renewal and differentiation characteristics of hiPSCs are affected differently by the biocompatibility and physico-chemical properties of individual substrates. Although

HLA-B27-Homodimer-Specific Antibody Modulates the Expansion of Pro-Inflammatory T-Cells in HLA-B27 Transgenic Rats.

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Osiris Marroquin Belaunzaran

Full Text Available HLA-B27 is a common genetic risk factor for the development of Spondyloarthritides (SpA. HLA-B27 can misfold to form cell-surface heavy chain homodimers (B272 and induce pro-inflammatory responses that may lead to SpA pathogenesis. The presence of B272 can be detected on leukocytes of HLA-B27+ Ankylosing spondylitis (AS patients and HLA-B27 transgenic rats. We characterized a novel B272-specific monoclonal antibody to study its therapeutic use in HLA-B27 associated disorders.The monoclonal HD5 antibody was selected from a phage library to target cell-surface B272 homodimers and characterized for affinity, specificity and ligand binding. The immune modulating effect of HD5 was tested in HLA-B27 transgenic rats. Onset and progression of disease profiles were monitored during therapy. Cell-surface B272 and expansion of pro-inflammatory cells from blood, spleen and draining lymph nodes were assessed by flow cytometry.HD5 bound B272 with high specificity and affinity (Kd = 0.32 nM. HD5 blocked cell-surface interaction of B272 with immune regulatory receptors KIR3DL2, LILRB2 and Pirb. In addition, HD5 modulated the production of TNF from CD4+ T-cells by limiting B272 interactions in vitro. In an HLA-B27 transgenic rat model repetitive dosing of HD5 reduced the expansion of pro-inflammatory CD4+ T-cells, and decreased the levels of soluble TNF and number of cell-surface B272 molecules.HD5 predominantly inhibits early TNF production and expansion of pro-inflammatory CD4+ T-cells in HLA-B27 transgenic rats. Monoclonal antibodies targeting cell-surface B272 propose a new concept for the modulation of inflammatory responses in HLA-B27 related disorders.

HLA-B27-Homodimer-Specific Antibody Modulates the Expansion of Pro-Inflammatory T-Cells in HLA-B27 Transgenic Rats

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Marroquin Belaunzaran, Osiris; Kleber, Sascha; Schauer, Stefan; Hausmann, Martin; Nicholls, Flora; Van den Broek, Maries; Payeli, Sravan; Ciurea, Adrian; Milling, Simon; Stenner, Frank; Shaw, Jackie; Kollnberger, Simon; Bowness, Paul; Petrausch, Ulf; Renner, Christoph

2015-01-01

Objectives HLA-B27 is a common genetic risk factor for the development of Spondyloarthritides (SpA). HLA-B27 can misfold to form cell-surface heavy chain homodimers (B272) and induce pro-inflammatory responses that may lead to SpA pathogenesis. The presence of B272 can be detected on leukocytes of HLA-B27+ Ankylosing spondylitis (AS) patients and HLA-B27 transgenic rats. We characterized a novel B272–specific monoclonal antibody to study its therapeutic use in HLA-B27 associated disorders. Methods The monoclonal HD5 antibody was selected from a phage library to target cell-surface B272 homodimers and characterized for affinity, specificity and ligand binding. The immune modulating effect of HD5 was tested in HLA-B27 transgenic rats. Onset and progression of disease profiles were monitored during therapy. Cell-surface B272 and expansion of pro-inflammatory cells from blood, spleen and draining lymph nodes were assessed by flow cytometry. Results HD5 bound B272 with high specificity and affinity (Kd = 0.32 nM). HD5 blocked cell-surface interaction of B272 with immune regulatory receptors KIR3DL2, LILRB2 and Pirb. In addition, HD5 modulated the production of TNF from CD4+ T-cells by limiting B272 interactions in vitro. In an HLA-B27 transgenic rat model repetitive dosing of HD5 reduced the expansion of pro-inflammatory CD4+ T-cells, and decreased the levels of soluble TNF and number of cell-surface B272 molecules. Conclusion HD5 predominantly inhibits early TNF production and expansion of pro-inflammatory CD4+ T-cells in HLA-B27 transgenic rats. Monoclonal antibodies targeting cell-surface B272 propose a new concept for the modulation of inflammatory responses in HLA-B27 related disorders. PMID:26125554

Wnt3a protein reduces growth factor-driven expansion of human hematopoietic stem and progenitor cells in serum-free cultures

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Duinhouwer, Lucia E.; Tüysüz, Nesrin; Rombouts, Elwin W J C; Ter Borg, Mariette N D; Mastrobattista, Enrico; Spanholtz, Jan; Cornelissen, Jan J.; Berge, Derk Ten; Braakman, Eric

2015-01-01

Ex vivo expansion of hematopoietic stem and progenitor cells (HSPC) is a promising approach to improve insufficient engraftment after umbilical cord blood stem cell transplantation (UCB-SCT). Although culturing HSPC with hematopoietic cytokines results in robust proliferation, it is accompanied with

Hyperkalemia caused by rapid red cell transfusion and the potassium absorption filter

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Yasuhiko Imashuku

2017-01-01

Full Text Available We report a case of transient hyperkalemia during hysterectomy after cesarean section, due to preoperatively undiagnosed placenta accreta that caused unforeseen massive hemorrhage and required rapid red cell transfusion. Hyperkalemia-induced by rapid red cell transfusion is a well-known severe complication of transfusion; however, in patients with sudden massive hemorrhage, rapid red cell transfusion is necessary to save their life. In such cases, it is extremely important to monitor serum potassium levels. For an emergency situation, a system should be developed to ensure sufficient preparation for immediate transfusion and laboratory tests. Furthermore, sufficient stock of preparations to treat hyperkalemia, such as calcium preparations, diuretics, glucose, and insulin is required. Moreover, a transfusion filter that absorbs potassium has been developed and is now available for clinical use in Japan. The filter is easy to use and beneficial, and should be prepared when it is available.

Hyperkalemia caused by rapid red cell transfusion and the potassium absorption filter

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Imashuku, Yasuhiko; Kitagawa, Hirotoshi; Mizuno, Takayoshi; Fukushima, Yutaka

2017-01-01

We report a case of transient hyperkalemia during hysterectomy after cesarean section, due to preoperatively undiagnosed placenta accreta that caused unforeseen massive hemorrhage and required rapid red cell transfusion. Hyperkalemia-induced by rapid red cell transfusion is a well-known severe complication of transfusion; however, in patients with sudden massive hemorrhage, rapid red cell transfusion is necessary to save their life. In such cases, it is extremely important to monitor serum potassium levels. For an emergency situation, a system should be developed to ensure sufficient preparation for immediate transfusion and laboratory tests. Furthermore, sufficient stock of preparations to treat hyperkalemia, such as calcium preparations, diuretics, glucose, and insulin is required. Moreover, a transfusion filter that absorbs potassium has been developed and is now available for clinical use in Japan. The filter is easy to use and beneficial, and should be prepared when it is available. PMID:28217070

Intraclonal Cell Expansion and Selection Driven by B Cell Receptor in Chronic Lymphocytic Leukemia

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Colombo, Monica; Cutrona, Giovanna; Reverberi, Daniele; Fabris, Sonia; Neri, Antonino; Fabbi, Marina; Quintana, Giovanni; Quarta, Giovanni; Ghiotto, Fabio; Fais, Franco; Ferrarini, Manlio

2011-01-01

The mutational status of the immunoglobulin heavy-chain variable region (IGHV) genes utilized by chronic lymphocytic leukemia (CLL) clones defines two disease subgroups. Patients with unmutated IGHV have a more aggressive disease and a worse outcome than patients with cells having somatic IGHV gene mutations. Moreover, up to 30% of the unmutated CLL clones exhibit very similar or identical B cell receptors (BcR), often encoded by the same IG genes. These “stereotyped” BcRs have been classified into defined subsets. The presence of an IGHV gene somatic mutation and the utilization of a skewed gene repertoire compared with normal B cells together with the expression of stereotyped receptors by unmutated CLL clones may indicate stimulation/selection by antigenic epitopes. This antigenic stimulation may occur prior to or during neoplastic transformation, but it is unknown whether this stimulation/selection continues after leukemogenesis has ceased. In this study, we focused on seven CLL cases with stereotyped BcR Subset #8 found among a cohort of 700 patients; in six, the cells expressed IgG and utilized IGHV4-39 and IGKV1-39/IGKV1D-39 genes, as reported for Subset #8 BcR. One case exhibited special features, including expression of IgM or IgG by different subclones consequent to an isotype switch, allelic inclusion at the IGH locus in the IgM-expressing cells and a particular pattern of cytogenetic lesions. Collectively, the data indicate a process of antigenic stimulation/selection of the fully transformed CLL cells leading to the expansion of the Subset #8 IgG-bearing subclone. PMID:21541442

In vivo expansion of naïve CD4+ CD25(high FOXP3+ regulatory T cells in patients with colorectal carcinoma after IL-2 administration.

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Marc Beyer

Full Text Available Regulatory T cells (T(reg cells are increased in context of malignancies and their expansion can be correlated with higher disease burden and decreased survival. Initially, interleukin 2 (IL-2 has been used as T-cell growth factor in clinical vaccination trials. In murine models, however, a role of IL-2 in development, differentiation, homeostasis, and function of T(reg cells was established. In IL-2 treated cancer patients a further T(reg-cell expansion was described, yet, the mechanism of expansion is still elusive. Here we report that functional T(reg cells of a naïve phenotype--as determined by CCR7 and CD45RA expression--are significantly expanded in colorectal cancer patients. Treatment of 15 UICC stage IV colorectal cancer patients with IL-2 in a phase I/II peptide vaccination trial further enlarges the already increased naïve T(reg-cell pool. Higher frequencies of T-cell receptor excision circles in naïve T(reg cells indicate IL-2 dependent thymic generation of naïve T(reg cells as a mechanism leading to increased frequencies of T(reg cells post IL-2 treatment in cancer patients. This finding could be confirmed in naïve murine T(reg cells after IL-2 administration. These results point to a more complex regulation of T(reg cells in context of IL-2 administration. Future strategies therefore might aim at combining IL-2 therapy with novel strategies to circumvent expansion and differentiation of naïve T(reg cells.

MicroRNA Cluster miR-17-92 Regulates Neural Stem Cell Expansion and Transition to Intermediate Progenitors in the Developing Mouse Neocortex

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Shan Bian

2013-05-01

Full Text Available During development of the embryonic neocortex, tightly regulated expansion of neural stem cells (NSCs and their transition to intermediate progenitors (IPs are critical for normal cortical formation and function. Molecular mechanisms that regulate NSC expansion and transition remain unclear. Here, we demonstrate that the microRNA (miRNA miR-17-92 cluster is required for maintaining proper populations of cortical radial glial cells (RGCs and IPs through repression of Pten and Tbr2 protein. Knockout of miR-17-92 and its paralogs specifically in the developing neocortex restricts NSC proliferation, suppresses RGC expansion, and promotes transition of RGCs to IPs. Moreover, Pten and Tbr2 protectors specifically block silencing activities of endogenous miR-17-92 and control proper numbers of RGCs and IPs in vivo. Our results demonstrate a critical role for miRNAs in promoting NSC proliferation and modulating the cell-fate decision of generating distinct neural progenitors in the developing neocortex.

Comparison of human platelet lysate alternatives using expired and freshly isolated platelet concentrates for adipose-derived stromal cell expansion.

Science.gov (United States)

Dessels, Carla; Durandt, Chrisna; Pepper, Michael S

2018-03-19

Pooled human platelet lysate (pHPL) has been used to expand adipose-derived stromal cells (ASCs) and can be formulated using fresh or expired buffy coats (BCs) which are then resuspended in either plasma or an additive solution. Not much is known about the effects that expired products and additive solutions have on ASC expansion, and the need for quality control and release criteria has been expressed. This pilot study compared proliferation, cell size, morphology and immunophenotype of ASCs expanded in the different pHPL alternatives versus foetal bovine serum (FBS). Quality control criteria were assessed prior to and during the manufacture of the pHPL alternatives. ASCs were then expanded in 1%, 2.5%, 5% or 10% of the different pHPL alternatives or in 10% FBS. Cell size, morphology, cell number and immunophenotype were measured using microscopy and flow cytometry. The majority of the pHPL alternatives were within the recommended ranges for the quality control criteria. ASCs expanded in the pHPL alternatives were smaller in size, displayed a tighter spindle-shaped morphology, increased cell growth and had a similar immunophenotype (with the exception of CD34 and CD36) when compared to ASCs expanded in FBS. Here we report on the effects that expired BC products and additive solutions have on ASC expansion. When taken together, our findings indicate that all of the pHPL alternatives can be considered to be suitable replacements for FBS for ASC expansion, and that expired BC products can be used as an alternative to fresh BC products.

Comparison of allogeneic platelet lysate and fetal bovine serum for in vitro expansion of equine bone marrow-derived mesenchymal stem cells.

Science.gov (United States)

Seo, Jong-pil; Tsuzuki, Nao; Haneda, Shingo; Yamada, Kazutaka; Furuoka, Hidefumi; Tabata, Yasuhiko; Sasaki, Naoki

2013-10-01

Mesenchymal stem cells (MSCs) are promising candidates for cell-based therapy and tissue engineering approaches. Fetal bovine serum (FBS) is commonly used for in vitro MSC expansion; however, the use of FBS may be associated with ethical, scientific, and safety issues. This study aimed to compare the ability of allogeneic platelet lysate (PL) and FBS to cause equine bone marrow-derived MSC expansion. MSCs were isolated from bone marrow aspirate in media supplemented with either PL or FBS, and cell proliferation properties and characteristics were examined. There were no significant differences in MSC yield, colony-forming unit-fibroblast (CFU-F) assay, and population doubling time between PL and FBS cultures. In addition, both PL-MSCs and FBS-MSCs showed similar results in term of ALP staining, osteogenic differentiation, and RT-PCR, although there were subtle differences in morphology, growth pattern, and adhesive properties. These results suggest that PL is a suitable alternative to FBS for use in equine MSC expansion, without the problems related to FBS use. Published by Elsevier India Pvt Ltd.

Thin-plate spline analysis of treatment effects of rapid maxillary expansion and face mask therapy in early Class III malocclusions.

Science.gov (United States)

Baccetti, T; Franchi, L; McNamara, J A

1999-06-01

An effective morphometric method (thin-plate spline analysis) was applied to evaluate shape changes in the craniofacial configuration of a sample of 23 children with Class III malocclusions in the early mixed dentition treated with rapid maxillary expansion and face mask therapy, and compared with a sample of 17 children with untreated Class III malocclusions. Significant treatment-induced changes involved both the maxilla and the mandible. Major deformations consisted of forward displacement of the maxillary complex from the pterygoid region and of anterior morphogenetic rotation of the mandible, due to a significant upward and forward direction of growth of the mandibular condyle. Significant differences in size changes due to reduced increments in mandibular dimensions were associated with significant shape changes in the treated group.

Chimeric antigen receptor T cells form nonclassical and potent immune synapses driving rapid cytotoxicity.

Science.gov (United States)

Davenport, A J; Cross, R S; Watson, K A; Liao, Y; Shi, W; Prince, H M; Beavis, P A; Trapani, J A; Kershaw, M H; Ritchie, D S; Darcy, P K; Neeson, P J; Jenkins, M R

2018-02-27

Chimeric antigen receptor T (CAR-T) cells are effective serial killers with a faster off-rate from dying tumor cells than CAR-T cells binding target cells through their T cell receptor (TCR). Here we explored the functional consequences of CAR-mediated signaling using a dual-specific CAR-T cell, where the same cell was triggered via TCR (tcrCTL) or CAR (carCTL). The carCTL immune synapse lacked distinct LFA-1 adhesion rings and was less reliant on LFA to form stable conjugates with target cells. carCTL receptors associated with the synapse were found to be disrupted and formed a convoluted multifocal pattern of Lck microclusters. Both proximal and distal receptor signaling pathways were induced more rapidly and subsequently decreased more rapidly in carCTL than in tcrCTL. The functional consequence of this rapid signaling in carCTL cells included faster lytic granule recruitment to the immune synapse, correlating with faster detachment of the CTL from the target cell. This study provides a mechanism for how CAR-T cells can debulk large tumor burden quickly and may contribute to further refinement of CAR design for enhancing the quality of signaling and programming of the T cell. Copyright © 2018 the Author(s). Published by PNAS.

Ex vivo expansion of bone marrow stromal cells by platelet-rich plasma: a promising strategy in maxillo-facial surgery.

Science.gov (United States)

Oliva, A; Passaro, I; Di Pasquale, R; Di Feo, A; Criscuolo, M; Zappia, V; Della Ragione, F; D'Amato, S; Annunziata, M; Guida, L

2005-01-01

The aim of our study is to evaluate in vitro the response of bone marrow stromal cells (BMSCs) to platelet-rich plasma (PRP), in order to clarify the potential role of their combined use in a preclinical phase preceding BMSCs transplantation for bone repair and regeneration procedures. The incubation of BMSCs with PRP promoted a remarkable, dose- and time- dependent, growth stimulation, that was paralleled to a strong increase in the quantity of type I collagen and to a significant decrease in the activity of the early osteoblastic differentiation marker, alkaline phosphatase (AP). Once PRP was removed and osteogenic inducers were added, AP returned to levels comparable to the control, while the late phenotypic markers, osteocalcin and matrix calcification, were enhanced to higher levels than in controls. Our data demonstrate that PRP induces a remarkable ex vivo enrichment of BMSCs maintaining their differentiative potential. Thus PRP represents a valid preclinical tool for obtaining an effective, rapid and safe ex vivo expansion of BMSCs prior to their clinical utilization in bone engineering.

Hematopoietic Stem and Progenitor Cell Expansion in Contact with Mesenchymal Stromal Cells in a Hanging Drop Model Uncovers Disadvantages of 3D Culture.

Science.gov (United States)

Schmal, Olga; Seifert, Jan; Schäffer, Tilman E; Walter, Christina B; Aicher, Wilhelm K; Klein, Gerd

2016-01-01

Efficient ex vivo expansion of hematopoietic stem cells with a concomitant preservation of stemness and self-renewal potential is still an unresolved ambition. Increased numbers of methods approaching this issue using three-dimensional (3D) cultures were reported. Here, we describe a simplified 3D hanging drop model for the coculture of cord blood-derived CD34(+) hematopoietic stem and progenitor cells (HSPCs) with bone marrow-derived mesenchymal stromal cells (MSCs). When seeded as a mixed cell suspension, MSCs segregated into tight spheroids. Despite the high expression of niche-specific extracellular matrix components by spheroid-forming MSCs, HSPCs did not migrate into the spheroids in the initial phase of coculture, indicating strong homotypic interactions of MSCs. After one week, however, HSPC attachment increased considerably, leading to spheroid collapse as demonstrated by electron microscopy and immunofluorescence staining. In terms of HSPC proliferation, the conventional 2D coculture system was superior to the hanging drop model. Furthermore, expansion of primitive hematopoietic progenitors was more favored in 2D than in 3D, as analyzed in colony-forming assays. Conclusively, our data demonstrate that MSCs, when arranged with a spread (monolayer) shape, exhibit better HSPC supportive qualities than spheroid-forming MSCs. Therefore, 3D systems are not necessarily superior to traditional 2D culture in this regard.

Studies on Rapidly Frozen Suspensions of Yeast Cells by Differential Thermal Analysis and Conductometry

Science.gov (United States)

Mazur, Peter

1963-01-01

Few, if any, yeast cells survived rapid cooling to -196°C and subsequent slow warming. After rapid freezing, the suspensions absorbed latent heat of fusion between -15° and 0°C during warming, and the relation between the amount of heat absorbed and the concentration of cells was the same as that in equivalent KCl solutions, indicating that frozen suspensions behave thermally like frozen solutions. The amount of heat absorbed was such that more than 80 per cent of the intracellular solution had to be frozen. The conductometric behavior of frozen suspensions showed that cell solutes were still inside the cells and surrounded by an intact cell membrane at the time heat was being absorbed. Two models are consistent with these findings. The first assumes that intracellular freezing has taken place; the second that all freezable water has left the cells and frozen externally. The latter model is ruled out because rapidly cooled cells do not shrink by an amount equal to the volume of water that would have to be withdrawn to prevent internal freezing. PMID:13934216

Observation of reversible, rapid changes in drug susceptibility of hypoxic tumor cells in a microfluidic device

Energy Technology Data Exchange (ETDEWEB)

Germain, Todd; Ansari, Megan; Pappas, Dimitri, E-mail: d.pappas@ttu.edu

2016-09-14

Hypoxia is a major stimulus for increased drug resistance and for survival of tumor cells. Work from our group and others has shown that hypoxia increases resistance to anti-cancer compounds, radiation, and other damage-pathway cytotoxic agents. In this work we utilize a microfluidic culture system capable of rapid switching of local oxygen concentrations to determine changes in drug resistance in prostate cancer cells. We observed rapid adaptation to hypoxia, with drug resistance to 2 μM staurosporine established within 30 min of hypoxia. Annexin-V/Sytox Green apoptosis assays over 9 h showed 78.0% viability, compared to 84.5% viability in control cells (normoxic cells with no staurosporine). Normoxic cells exposed to the same staurosporine concentration had a viability of 48.6% after 9 h. Hypoxia adaptation was rapid and reversible, with Hypoxic cells treated with 20% oxygen for 30 min responding to staurosporine with 51.6% viability after drug treatment for 9 h. Induction of apoptosis through the receptor-mediated pathway, which bypasses anti-apoptosis mechanisms induced by hypoxia, resulted in 39.4 ± 7% cell viability. The rapid reversibility indicates co-treatment of oxygen with anti-cancer compounds may be a potential therapeutic target. - Highlights: • Microfluidic system switches rapidly between normoxia and hypoxia (5 min). • Observation of rapid adaptation of PC3 cells to hypoxia and normoxia (30 min). • Drug susceptibility in tumor cells restored after chip switched to normoxia for 30 min.

Control of Cell Wall Extensibility during Pollen Tube Growth

OpenAIRE

Hepler, Peter K.; Rounds, Caleb M.; Winship, Lawrence J.

2013-01-01

Tip-growing pollen tubes achieve rapid elongation while maintaining cell wall integrity by balancing local expansion, controlled by local changes in wall viscosity, against exocytosis, influenced by the activity of the actin cytoskeleton, cellular energetics, and calcium and proton physiology.

Radial expansion and multifragmentation

International Nuclear Information System (INIS)

Angelique, J.C.; Bizard, G.; Bougault, R.; Brou, R.; Buta, A.; Colin, J.; Cussol, D.; Durand, D.; Kerambrun, A.; Le Brun, C.; Lecolley, J.F.; Lopez, O.; Louvel, M.; Meslin, C.; Nakagawa, T.; Patry, J.P.; Peter, J.; Popescu, R.; Regimbart, R.; Steckmeyer, J.C.; Tamain, B.; Vient, E.; Yuasa-Nakagawa, K.; Wieloch, A.

1998-01-01

The light systems 36 Ar + 27 Al and 64 Zn + nat Ti were measured at several bombarding energies between ∼ 35 and 95 MeV/nucleon. It was found that the predominant part of the cross section is due to binary collisions. In this paper the focus is placed on the properties of the quasi-projectile nuclei. In the central collisions the excitation energies of the quasi-projectile reach values exceeding largely 10 MeV/nucleon. The slope of the high energy part of the distribution can give only an upper limit of the apparent temperature (the average temperature along the decay chain). The highly excited quasi-projectile may get rapidly fragmented rather than sequentially. The heavy fragments are excited and can emit light particles (n, p, d, t, 3 He, α,...) what perturbs additionally the spectrum of these particles. Concerning the expansion energy, one can determine the average kinetic energies of the product (in the quasi-projectile-framework) and compare with simulation values. To fit the experimental data an additional radial expansion energy is to be considered. The average expansion energy depends slightly on the impact parameter but it increases with E * / A, ranging from 0.4 to 1,2 MeV/nucleon for an excitation energy increasing from 7 to 10.5 MeV/nucleon. This collective radial energy seems to be independent of the fragment mass, what is possibly valid for the case of larger quasi-projectile masses. The origin of the expansion is to be determined. It may be due to a compression in the interaction zone at the initial stage of the collision, which propagates in the quasi-projectile and quasi-target, or else, may be due, simply, to the increase of thermal energy leading to a rapid fragment emission. The sequential de-excitation calculation overestimates light particle emission and consequently heavy residues, particularly, at higher excitation energies. This disagreement indicates that a sequential process can not account for the di-excitation of very hot nuclei

Range expansion of heterogeneous populations.

Science.gov (United States)

Reiter, Matthias; Rulands, Steffen; Frey, Erwin

2014-04-11

Risk spreading in bacterial populations is generally regarded as a strategy to maximize survival. Here, we study its role during range expansion of a genetically diverse population where growth and motility are two alternative traits. We find that during the initial expansion phase fast-growing cells do have a selective advantage. By contrast, asymptotically, generalists balancing motility and reproduction are evolutionarily most successful. These findings are rationalized by a set of coupled Fisher equations complemented by stochastic simulations.

Continuous 4-1BB co-stimulatory signals for the optimal expansion of tumor-infiltrating lymphocytes for adoptive T-cell therapy.

Science.gov (United States)

Chacon, Jessica Ann; Pilon-Thomas, Shari; Sarnaik, Amod A; Radvanyi, Laszlo G

2013-09-01

Co-stimulation through members of the tumor necrosis factor receptor (TNFR) family appears to be critical for the generation of T cells with optimal effector-memory properties for adoptive cell therapy. Our work suggests that continuous 4-1BB/CD137 co-stimulation is required for the expansion of T cells with an optimal therapeutic profile and that the administration of 4-1BB agonists upon adoptive cell transfer further improves antitumor T-cell functions.

Wigner-Kirkwood expansion of the phase-space density for half infinite nuclear matter

International Nuclear Information System (INIS)

Durand, M.; Schuck, P.

1987-01-01

The phase space distribution of half infinite nuclear matter is expanded in a ℎ-series analogous to the low temperature expansion of the Fermi function. Besides the usual Wigner-Kirkwood expansion, oscillatory terms are derived. In the case of a Woods-Saxon potential, a smallness parameter is defined, which determines the convergence of the series and explains the very rapid convergence of the Wigner-Kirkwood expansion for average (nuclear) binding energies

Activation of intracellular angiotensin AT2 receptors induces rapid cell death in human uterine leiomyosarcoma cells

DEFF Research Database (Denmark)

Zhao, Yi; Lützen, Ulf; Fritsch, Jürgen

2015-01-01

The presence of AT2 receptors in mitochondria and their role in NO generation and cell aging were recently demonstrated in various human and mouse non-tumour cells. We investigated the intracellular distribution of AT2 receptors including their presence in mitochondria and the role in the induction...... agonist, Compound 21 (C21) penetrates the cell membrane of quiescent SK-UT-1 cells, activates intracellular AT2 receptors and induces rapid cell death; approximately 70% of cells died within 24 h. The cells, which escaped from the cell death, displayed activation of the mitochondrial apoptotic pathway, i...

Rapid replacement of bridge deck expansion joints study - phase I : [tech transfer summary].

Science.gov (United States)

2014-12-01

This initial research phase focused on documenting the current : means and methods of bridge expansion joint deterioration, : maintenance, and replacement and on identifying improvements : through all of the input gathered.

Investigation of droplet nucleation in CCS relevant systems - design and testing of the expansion chamber

Science.gov (United States)

Čenský, Miroslav; Hrubý, Jan; Vinš, Václav; Hykl, Jiří; Šmíd, Bohuslav

2018-06-01

A unique in-house designed experimental apparatus for investigation of nucleation of droplets in CCS relevant systems is being developed by the present team. The apparatus allows simulating various processes relevant to CCS technologies. Gaseous mixtures with CO2 are prepared in a Mixture Preparation Device (MPD) based on accurate adjustment of flow rates of individual components [EPJ Web of Conferences 143, 02140 (2017)]. The mixture then flows into an expansion chamber, where it undergoes a rapid adiabatic expansion. As a consequence of adiabatic cooling, the mixture becomes supersaturated and nucleation and simultaneous growth of droplets occurs. In this study, we describe the design and testing of the expansion part of the experimental setup. The rapid expansion was realized using two valve systems, one for low pressures (up to 0.7 MPa) and the other for high pressures (up to 10 MPa). A challenge for a proper design of the expansion system is avoiding acoustic oscillations. These can occur either in the mode of Helmholtz resonator, where the compressible gas in the chamber acts as a spring and the rapidly moving gas in the valve system as a mass, or in the "flute" mode, where acoustic waves are generated in a long outlet tubing.

Model test on partial expansion in stratified subsidence during foundation pit dewatering

Science.gov (United States)

Wang, Jianxiu; Deng, Yansheng; Ma, Ruiqiang; Liu, Xiaotian; Guo, Qingfeng; Liu, Shaoli; Shao, Yule; Wu, Linbo; Zhou, Jie; Yang, Tianliang; Wang, Hanmei; Huang, Xinlei

2018-02-01

Partial expansion was observed in stratified subsidence during foundation pit dewatering. However, the phenomenon was suspected to be an error because the compression of layers is known to occur when subsidence occurs. A slice of the subsidence cone induced by drawdown was selected as the prototype. Model tests were performed to investigate the phenomenon. The underlying confined aquifer was generated as a movable rigid plate with a hinge at one end. The overlying layers were simulated with remolded materials collected from a construction site. Model tests performed under the conceptual model indicated that partial expansion occurred in stratified settlements under coordination deformation and consolidation conditions. During foundation pit dewatering, rapid drawdown resulted in rapid subsidence in the dewatered confined aquifer. The rapidly subsiding confined aquifer top was the bottom deformation boundary of the overlying layers. Non-coordination deformation was observed at the top and bottom of the subsiding overlying layers. The subsidence of overlying layers was larger at the bottom than at the top. The layers expanded and became thicker. The phenomenon was verified using numerical simulation method based on finite difference method. Compared with numerical simulation results, the boundary effect of the physical tests was obvious in the observation point close to the movable endpoint. The tensile stress of the overlying soil layers induced by the underlying settlement of dewatered confined aquifer contributed to the expansion phenomenon. The partial expansion of overlying soil layers was defined as inversed rebound. The inversed rebound was induced by inversed coordination deformation. Compression was induced by the consolidation in the overlying soil layers because of drainage. Partial expansion occurred when the expansion exceeded the compression. Considering the inversed rebound, traditional layer-wise summation method for calculating subsidence should be

A two-parameter family of double-power-law biorthonormal potential-density expansions

Science.gov (United States)

Lilley, Edward J.; Sanders, Jason L.; Evans, N. Wyn

2018-05-01

We present a two-parameter family of biorthonormal double-power-law potential-density expansions. Both the potential and density are given in closed analytic form and may be rapidly computed via recurrence relations. We show that this family encompasses all the known analytic biorthonormal expansions: the Zhao expansions (themselves generalizations of ones found earlier by Hernquist & Ostriker and by Clutton-Brock) and the recently discovered Lilley et al. (2017a) expansion. Our new two-parameter family includes expansions based around many familiar spherical density profiles as zeroth-order models, including the γ models and the Jaffe model. It also contains a basis expansion that reproduces the famous Navarro-Frenk-White (NFW) profile at zeroth order. The new basis expansions have been found via a systematic methodology which has wide applications in finding other new expansions. In the process, we also uncovered a novel integral transform solution to Poisson's equation.

Exploring the Early Structure of a Rapidly Decompressed Particle Bed

Science.gov (United States)

Zunino, Heather; Adrian, R. J.; Clarke, Amanda; Johnson, Blair; Arizona State University Collaboration

2017-11-01

Rapid expansion of dense, pressurized beds of fine particles subjected to rapid reduction of the external pressure is studied in a vertical shock tube. A near-sonic expansion wave impinges on the particle bed-gas interface and rapidly unloads the particle bed. A high-speed video camera captures events occurring during bed expansion. The particle bed does not expand homogeneously, but breaks down into horizontal slabs and then transforms into a cellular-type structure. There are several key parameters that affect the particle bed evolution, including particle size and initial bed height. Analyses of this bed structure evolution from experiments with varying particle sizes and initial bed heights is presented. This work is supported by the U.S. Department of Energy, National Nuclear Security Administration, Advanced Simulation and Computing Program, as a Cooperative Agreement under the Predictive Science and Academic Alliance Program, under Contract No. DE-NA0002378.

Expansion of the uranium mines in the Elliot Lake area

International Nuclear Information System (INIS)

1979-11-01

This report forms the response of the government of Ontario to an earlier report issued by an Envrionmental Assessment Board. Specifically, the report deals with the rapid growth of the town of Elliot Lake due to expansion of several uranium mine-mill operations. Rapid growth of small communities presents considerable problems in providing housing, essential services, and educational facilities. Several specific actions taken by the government to help the town cope with rapid growth are presented. (O.T.)

IL-7 treatment augments and prolongs sepsis-induced expansion of IL-10-producing B lymphocytes and myeloid-derived suppressor cells.

Science.gov (United States)

Kulkarni, Upasana; Herrmenau, Christoph; Win, Stephanie J; Bauer, Michael; Kamradt, Thomas

2018-01-01

Immunological dysregulation in sepsis is associated with often lethal secondary infections. Loss of effector cells and an expansion of immunoregulatory cell populations both contribute to sepsis-induced immunosuppression. The extent and duration of this immunosuppression are unknown. Interleukin 7 (IL-7) is important for the maintenance of lymphocytes and can accelerate the reconstitution of effector lymphocytes in sepsis. How IL-7 influences immunosuppressive cell populations is unknown. We have used the mouse model of peritoneal contamination and infection (PCI) to investigate the expansion of immunoregulatory cells as long-term sequelae of sepsis with or without IL-7 treatment. We analysed the frequencies and numbers of regulatory T cells (Tregs), double negative T cells, IL-10 producing B cells and myeloid-derived suppressor cells (MDSCs) for 3.5 months after sepsis induction. Sepsis induced an increase in IL-10+ B cells, which was enhanced and prolonged by IL-7 treatment. An increased frequency of MDSCs in the spleen was still detectable 3.5 months after sepsis induction and this was more pronounced in IL-7-treated mice. MDSCs from septic mice were more potent at suppressing T cell proliferation than MDSCs from control mice. Our data reveal that sepsis induces a long lasting increase in IL-10+ B cells and MDSCs. Late-onset IL-7 treatment augments this increase, which should be relevant for clinical interventions.

IL-10 Production Is Critical for Sustaining the Expansion of CD5+ B and NKT Cells and Restraining Autoantibody Production in Congenic Lupus-Prone Mice.

Directory of Open Access Journals (Sweden)

Yuriy Baglaenko

Full Text Available The development and progression of systemic lupus erythematosus is mediated by the complex interaction of genetic and environmental factors. To decipher the genetics that contribute to pathogenesis and the production of pathogenic autoantibodies, our lab has focused on the generation of congenic lupus-prone mice derived from the New Zealand Black (NZB strain. Previous work has shown that an NZB-derived chromosome 4 interval spanning 32 to 151 Mb led to expansion of CD5+ B and Natural Killer T (NKT cells, and could suppress autoimmunity when crossed with a lupus-prone mouse strain. Subsequently, it was shown that CD5+ B cells but not NKT cells derived from these mice could suppress the development of pro-inflammatory T cells. In this paper, we aimed to further resolve the genetics that leads to expansion of these two innate-like populations through the creation of additional sub-congenic mice and to characterize the role of IL-10 in the suppression of autoimmunity through the generation of IL-10 knockout mice. We show that expansion of CD5+ B cells and NKT cells localizes to a chromosome 4 interval spanning 91 to 123 Mb, which is distinct from the region that mediates the majority of the suppressive phenotype. We also demonstrate that IL-10 is critical to restraining autoantibody production and surprisingly plays a vital role in supporting the expansion of innate-like populations.

IL-10 Production Is Critical for Sustaining the Expansion of CD5+ B and NKT Cells and Restraining Autoantibody Production in Congenic Lupus-Prone Mice.

Science.gov (United States)

Baglaenko, Yuriy; Manion, Kieran P; Chang, Nan-Hua; Gracey, Eric; Loh, Christina; Wither, Joan E

2016-01-01

The development and progression of systemic lupus erythematosus is mediated by the complex interaction of genetic and environmental factors. To decipher the genetics that contribute to pathogenesis and the production of pathogenic autoantibodies, our lab has focused on the generation of congenic lupus-prone mice derived from the New Zealand Black (NZB) strain. Previous work has shown that an NZB-derived chromosome 4 interval spanning 32 to 151 Mb led to expansion of CD5+ B and Natural Killer T (NKT) cells, and could suppress autoimmunity when crossed with a lupus-prone mouse strain. Subsequently, it was shown that CD5+ B cells but not NKT cells derived from these mice could suppress the development of pro-inflammatory T cells. In this paper, we aimed to further resolve the genetics that leads to expansion of these two innate-like populations through the creation of additional sub-congenic mice and to characterize the role of IL-10 in the suppression of autoimmunity through the generation of IL-10 knockout mice. We show that expansion of CD5+ B cells and NKT cells localizes to a chromosome 4 interval spanning 91 to 123 Mb, which is distinct from the region that mediates the majority of the suppressive phenotype. We also demonstrate that IL-10 is critical to restraining autoantibody production and surprisingly plays a vital role in supporting the expansion of innate-like populations.

Differences in Expansion Potential of Naive Chimeric Antigen Receptor T Cells from Healthy Donors and Untreated Chronic Lymphocytic Leukemia Patients

Directory of Open Access Journals (Sweden)

Jean-Marc Hoffmann

2018-01-01

Full Text Available IntroductionTherapy with chimeric antigen receptor T (CART cells for hematological malignancies has shown promising results. Effectiveness of CART cells may depend on the ratio of naive (TN vs. effector (TE T cells, TN cells being responsible for an enduring antitumor activity through maturation. Therefore, we investigated factors influencing the TN/TE ratio of CART cells.Materials and methodsCART cells were generated upon transduction of peripheral blood mononuclear cells with a CD19.CAR-CD28-CD137zeta third generation retroviral vector under two different stimulating culture conditions: anti-CD3/anti-CD28 antibodies adding either interleukin (IL-7/IL-15 or IL-2. CART cells were maintained in culture for 20 days. We evaluated 24 healthy donors (HDs and 11 patients with chronic lymphocytic leukemia (CLL for the composition of cell subsets and produced CART cells. Phenotype and functionality were tested using flow cytometry and chromium release assays.ResultsIL-7/IL-15 preferentially induced differentiation into TN, stem cell memory (TSCM: naive CD27+ CD95+, CD4+ and CXCR3+ CART cells, while IL-2 increased effector memory (TEM, CD56+ and CD4+ T regulatory (TReg CART cells. The net amplification of different CART subpopulations derived from HDs and untreated CLL patients was compared. Particularly the expansion of CD4+ CARTN cells differed significantly between the two groups. For HDs, this subtype expanded >60-fold, whereas CD4+ CARTN cells of untreated CLL patients expanded less than 10-fold. Expression of exhaustion marker programmed cell death 1 on CARTN cells on day 10 of culture was significantly higher in patient samples compared to HD samples. As the percentage of malignant B cells was expectedly higher within patient samples, an excessive amount of B cells during culture could account for the reduced expansion potential of CARTN cells in untreated CLL patients. Final TN/TE ratio stayed <0.3 despite stimulation condition for patients

Platelet released growth factors boost expansion of bone marrow derived CD34(+) and CD133(+) endothelial progenitor cells for autologous grafting.

Science.gov (United States)

Lippross, Sebastian; Loibl, Markus; Hoppe, Sven; Meury, Thomas; Benneker, Lorin; Alini, Mauro; Verrier, Sophie

2011-01-01

Stem cell based autologous grafting has recently gained mayor interest in various surgical fields for the treatment of extensive tissue defects. CD34(+) and CD133(+) cells that can be isolated from the pool of bone marrow mononuclear cells (BMC) are capable of differentiating into mature endothelial cells in vivo. These endothelial progenitor cells (EPC) are believed to represent a major portion of the angiogenic regenerative cells that are released from bone marrow when tissue injury has occurred. In recent years tissue engineers increasingly looked at the process of vessel neoformation because of its major importance for successful cell grafting to replace damaged tissue. Up to now one of the greatest problems preventing a clinical application is the large scale of expansion that is required for such purpose. We established a method to effectively enhance the expansion of CD34(+) and CD133(+) cells by the use of platelet-released growth factors (PRGF) as a media supplement. PRGF were prepared from thrombocyte concentrates and used as a media supplement to iscove's modified dulbecco's media (IMDM). EPC were immunomagnetically separated from human bone morrow monocyte cells and cultured in IMDM + 10% fetal calf serum (FCS), IMDM + 5%, FCS + 5% PRGF and IMDM + 10% PRGF. We clearly demonstrate a statistically significant higher and faster cell proliferation rate at 7, 14, 21, and 28 days of culture when both PRGF and FCS were added to the medium as opposed to 10% FCS or 10% PRGF alone. The addition of 10% PRGF to IMDM in the absence of FCS leads to a growth arrest from day 14 on. In histochemical, immunocytochemical, and gene-expression analysis we showed that angiogenic and precursor markers of CD34(+) and CD133(+) cells are maintained during long-term culture. In summary, we established a protocol to boost the expansion of CD34(+) and CD133(+) cells. Thereby we provide a technical step towards the clinical application of autologous stem cell

Scaling behavior of ground-state energy cluster expansion for linear polyenes

Science.gov (United States)

Griffin, L. L.; Wu, Jian; Klein, D. J.; Schmalz, T. G.; Bytautas, L.

Ground-state energies for linear-chain polyenes are additively expanded in a sequence of terms for chemically relevant conjugated substructures of increasing size. The asymptotic behavior of the large-substructure limit (i.e., high-polymer limit) is investigated as a means of characterizing the rapidity of convergence and consequent utility of this energy cluster expansion. Consideration is directed to computations via: simple Hückel theory, a refined Hückel scheme with geometry optimization, restricted Hartree-Fock self-consistent field (RHF-SCF) solutions of fixed bond-length Parisier-Parr-Pople (PPP)/Hubbard models, and ab initio SCF approaches with and without geometry optimization. The cluster expansion in what might be described as the more "refined" approaches appears to lead to qualitatively more rapid convergence: exponentially fast as opposed to an inverse power at the simple Hückel or SCF-Hubbard levels. The substructural energy cluster expansion then seems to merit special attention. Its possible utility in making accurate extrapolations from finite systems to extended polymers is noted.

Peruvian Higher Education: Expansions Amid Economic Crisis.

Science.gov (United States)

Post, David

1991-01-01

Development of the Peruvian university system is described, focusing on periods of rapid expansion. Enrollment declines in 1974-78 are analyzed in the context of the educational reform program of the military government. The 1983 new university law, following return to civilian government, and future prospects for higher education are discussed.…

Hematopoietic Stem and Progenitor Cell Expansion in Contact with Mesenchymal Stromal Cells in a Hanging Drop Model Uncovers Disadvantages of 3D Culture

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Olga Schmal

2016-01-01

Full Text Available Efficient ex vivo expansion of hematopoietic stem cells with a concomitant preservation of stemness and self-renewal potential is still an unresolved ambition. Increased numbers of methods approaching this issue using three-dimensional (3D cultures were reported. Here, we describe a simplified 3D hanging drop model for the coculture of cord blood-derived CD34+ hematopoietic stem and progenitor cells (HSPCs with bone marrow-derived mesenchymal stromal cells (MSCs. When seeded as a mixed cell suspension, MSCs segregated into tight spheroids. Despite the high expression of niche-specific extracellular matrix components by spheroid-forming MSCs, HSPCs did not migrate into the spheroids in the initial phase of coculture, indicating strong homotypic interactions of MSCs. After one week, however, HSPC attachment increased considerably, leading to spheroid collapse as demonstrated by electron microscopy and immunofluorescence staining. In terms of HSPC proliferation, the conventional 2D coculture system was superior to the hanging drop model. Furthermore, expansion of primitive hematopoietic progenitors was more favored in 2D than in 3D, as analyzed in colony-forming assays. Conclusively, our data demonstrate that MSCs, when arranged with a spread (monolayer shape, exhibit better HSPC supportive qualities than spheroid-forming MSCs. Therefore, 3D systems are not necessarily superior to traditional 2D culture in this regard.

Matrix expansion and syncytial aggregation of syndecan-1+ cells underpin villous atrophy in coeliac disease.

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Camilla Salvestrini

Full Text Available We studied the expression of sulphated glycosaminoglycans (GAGs in coeliac disease (CD mucosa, as they are critical determinants of tissue volume, which increases in active disease. We also examined mucosal expression of IL-6, which stimulates excess GAG synthesis in disorders such as Grave's ophthalmopathy.We stained archival jejunal biopsies from 5 children with CD at diagnosis, on gluten-free diet and challenge for sulphated GAGs. We then examined duodenal biopsies from 9 children with CD compared to 9 histological normal controls, staining for sulphated GAGs, heparan sulphate proteoglycans (HSPG, short-chain HSPG (Δ-HSPG and the proteoglycan syndecan-1 (CD138, which is expressed on epithelium and plasma cells. We confirmed findings with a second monoclonal in another 12 coeliac children. We determined mucosal IL-6 expression by immunohistochemistry and PCR in 9 further cases and controls, and used quantitative real time PCR for other Th17 pathway cytokines in an additional 10 cases and controls.In CD, HSPG expression was lost in the epithelial compartment but contrastingly maintained within an expanded lamina propria. Within the upper lamina propria, clusters of syndecan-1(+ plasma cells formed extensive syncytial sheets, comprising adherent plasma cells, lysed cells with punctate cytoplasmic staining and shed syndecan ectodomains. A dense infiltrate of IL-6(+ mononuclear cells was detected in active coeliac disease, also localised to the upper lamina propria, with significantly increased mRNA expression of IL-6 and IL-17A but not IL-23 p19.Matrix expansion, through syndecan-1(+ cell recruitment and lamina propria GAG increase, underpins villous atrophy in coeliac disease. The syndecan-1(+ cell syncytia and excess GAG production recapitulate elements of the invertebrate encapsulation reaction, itself dependent on insect transglutaminase and glutaminated early response proteins. As in other matrix expansion disorders, IL-6 is upregulated and

Matrix Expansion and Syncytial Aggregation of Syndecan-1+ Cells Underpin Villous Atrophy in Coeliac Disease

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Salvestrini, Camilla; Lucas, Mark; Lionetti, Paolo; Torrente, Franco; James, Sean; Phillips, Alan D.; Murch, Simon H.

2014-01-01

Background We studied the expression of sulphated glycosaminoglycans (GAGs) in coeliac disease (CD) mucosa, as they are critical determinants of tissue volume, which increases in active disease. We also examined mucosal expression of IL-6, which stimulates excess GAG synthesis in disorders such as Grave's ophthalmopathy. Methods We stained archival jejunal biopsies from 5 children with CD at diagnosis, on gluten-free diet and challenge for sulphated GAGs. We then examined duodenal biopsies from 9 children with CD compared to 9 histological normal controls, staining for sulphated GAGs, heparan sulphate proteoglycans (HSPG), short-chain HSPG (Δ-HSPG) and the proteoglycan syndecan-1 (CD138), which is expressed on epithelium and plasma cells. We confirmed findings with a second monoclonal in another 12 coeliac children. We determined mucosal IL-6 expression by immunohistochemistry and PCR in 9 further cases and controls, and used quantitative real time PCR for other Th17 pathway cytokines in an additional 10 cases and controls. Results In CD, HSPG expression was lost in the epithelial compartment but contrastingly maintained within an expanded lamina propria. Within the upper lamina propria, clusters of syndecan-1+ plasma cells formed extensive syncytial sheets, comprising adherent plasma cells, lysed cells with punctate cytoplasmic staining and shed syndecan ectodomains. A dense infiltrate of IL-6+ mononuclear cells was detected in active coeliac disease, also localised to the upper lamina propria, with significantly increased mRNA expression of IL-6 and IL-17A but not IL-23 p19. Conclusions Matrix expansion, through syndecan-1+ cell recruitment and lamina propria GAG increase, underpins villous atrophy in coeliac disease. The syndecan-1+ cell syncytia and excess GAG production recapitulate elements of the invertebrate encapsulation reaction, itself dependent on insect transglutaminase and glutaminated early response proteins. As in other matrix expansion disorders

Selective Destruction of Interleukin 23–Induced Expansion of a Major Antigen–Specific γδ T-Cell Subset in Patients With Tuberculosis

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Gu, Jin; Xiao, Heping; Liang, Shanshan; Yang, Enzhuo; Yang, Rui; Huang, Dan; Chen, Crystal; Wang, Feifei; Shen, Ling; Chen, Zheng W.

2017-01-01

Abstract A loss of antigen-specific T-cell responses due to defective cytokine signaling during infections has not been reported. We hypothesize that tuberculosis can destroy signaling effects of selective cytokine(s) and induce exhaustion of antigen-specific T cells. To test this hypothesis, mechanistic studies were performed to examine whether and how tuberculosis blocked interleukin 23 (IL-23) and interleukin 2 (IL-2) signaling effects on a major human γδ T-cell subpopulation, phosphoantigen HMBPP–specific Vγ2Vδ2 T cells. IL-23 and IL-2 significantly expanded HMBPP-stimulated Vγ2Vδ2 T cells from subjects with latent tuberculosis infection, and IL-2 synergized the effect of IL-23. IL-23–induced expansion of Vγ2Vδ2 T cells involved STAT3. Surprisingly, patients with tuberculosis exhibited a selective destruction of IL-23–induced expansion of these cells. The tuberculosis-driven destruction of IL-23 signaling coincided with decreases of expression and phosphorylation of STAT3. Interestingly, impairing of STAT3 was linked to marked increases in the microRNAs (miRNAs) hsa-miR-337-3p and hsa-miR-125b-5p in Vγ2Vδ2 T cells from patients with tuberculosis. Downregulation of hsa-miR-337-3p and hsa-miR-125b-5p by miRNA sponges improved IL-23–mediated expansion of Vγ2Vδ2 T cells and restored the ability of these cells to produce anti–tuberculosis cytokines. These results support our hypothesis that tuberculosis can selectively impair a cytokine effect while sparing another and can induce exhaustion of T cells in response to the respective cytokine. PMID:27789724

Contrasting patterns of urban expansion in Colombia, Ecuador, Peru, and Bolivia between 1992 and 2009.

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Alvarez-Berríos, Nora L; Parés-Ramos, Isabel K; Aide, T Mitchell

2013-02-01

The global urban population is increasing rapidly, but patterns of urban expansion differ greatly among countries. Urban transition theory predicts that the shift from low to high urbanization depends on a country's history and level of economic development. This study describes urban expansion in Colombia, Ecuador, Peru, and Bolivia between 1992 and 2009. Urban dynamics were analyzed by combining nighttime lights and census data from 4032 municipalities. High-lit areas (>52-63 pixel values) were correlated with urban populations across municipalities and years (R (2) > 0.90). Analyses showed that between 1992 and 2009 Bolivia and Ecuador had rapid population growth and rapidly increasing high-lit areas, while Peru and Colombia had lower rates of population growth and urbanization (i.e., expansion of high-lit areas). We demonstrate how nighttime lights can be a useful tool, providing a homogeneous platform for multi-scale analyses of urban growth.

MicroRNA-155 Modulates Acute Graft-versus-Host Disease by Impacting T Cell Expansion, Migration, and Effector Function.

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Zitzer, Nina C; Snyder, Katiri; Meng, Xiamoei; Taylor, Patricia A; Efebera, Yvonne A; Devine, Steven M; Blazar, Bruce R; Garzon, Ramiro; Ranganathan, Parvathi

2018-06-15

MicroRNA-155 (miR-155) is a small noncoding RNA critical for the regulation of inflammation as well as innate and adaptive immune responses. MiR-155 has been shown to be dysregulated in both donor and recipient immune cells during acute graft-versus-host disease (aGVHD). We previously reported that miR-155 is upregulated in donor T cells of mice and humans with aGVHD and that mice receiving miR-155-deficient (miR155 -/- ) splenocytes had markedly reduced aGVHD. However, molecular mechanisms by which miR-155 modulates T cell function in aGVHD have not been fully investigated. We identify that miR-155 expression in both donor CD8 + T cells and conventional CD4 + CD25 - T cells is pivotal for aGVHD pathogenesis. Using murine aGVHD transplant experiments, we show that miR-155 strongly impacts alloreactive T cell expansion through multiple distinct mechanisms, modulating proliferation in CD8 + donor T cells and promoting exhaustion in donor CD4 + T cells in both the spleen and colon. Additionally, miR-155 drives a proinflammatory Th1 phenotype in donor T cells in these two sites, and miR-155 -/- donor T cells are polarized toward an IL-4-producing Th2 phenotype. We further demonstrate that miR-155 expression in donor T cells regulates CCR5 and CXCR4 chemokine-dependent migration. Notably, we show that miR-155 expression is crucial for donor T cell infiltration into multiple target organs. These findings provide further understanding of the role of miR-155 in modulating aGVHD through T cell expansion, effector cytokine production, and migration. Copyright © 2018 by The American Association of Immunologists, Inc.

The double bromodomain protein Brd2 promotes B cell expansion and mitogenesis.

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Belkina, Anna C; Blanton, Wanda P; Nikolajczyk, Barbara S; Denis, Gerald V

2014-03-01

Bromodomain-containing transcriptional regulators represent new epigenetic targets in different hematologic malignancies. However, bromodomain-mediated mechanisms that couple histone acetylation to transcription in lymphopoiesis and govern mature lymphocyte mitogenesis are poorly understood. Brd2, a transcriptional coregulator that contains dual bromodomains and an extraterminal domain (the BET family), couples chromatin to cell-cycle progression. We reported previously the first functional characterization of a BET protein as an effector of mammalian mitogenic signal transduction: Eμ-Brd2 Tg mice develop "activated B cell" diffuse large B cell lymphoma. No other animal models exist for genetic or lentiviral expression of BET proteins, hampering testing of novel anti-BET anticancer drugs, such as JQ1. We transduced HSCs with Brd2 lentivirus and reconstituted recipient mice to test the hypothesis that Brd2 regulates hematopoiesis in BM and mitogenesis in the periphery. Forced expression of Brd2 provides an expansion advantage to the donor-derived B cell compartment in BM and increases mature B cell mitogenic responsiveness in vitro. Brd2 binds the cyclin A promoter in B cells, shown by ChIP, and increases cyclin A mRNA and protein levels, and S-phase progression in vitro in mitogen-stimulated primary B cells, but not T cells, reinforcing results from Eμ-Brd2 mice. The small molecule BET inhibitor JQ1 reduces B cell mitogenesis, consistent with the interpretation that BET inhibitors are antiproliferative. Brd2-specific knockdown experiments show that Brd2 is also required for hematopoiesis. We conclude that Brd2 plays a critical, independent role in regulation of mitogenic response genes, particularly cyclin A, in B cells.

Plume expansion of a laser-induced plasma studied with the particle-in-cell method

DEFF Research Database (Denmark)

Ellegaard, O.; Nedelea, T.; Schou, Jørgen

2002-01-01

energy as well as electron energy. We have estimated the time constant for energy transfer between the electrons and the ions. The scaling of these processes is given by a single parameter determined by the Debye length obtained from the electron density in the plasma outside the surface. (C) 2002......The initial stage of laser-induced plasma plume expansion from a solid in vacuum and the effect of the Coulomb field have been studied. We have performed a one-dimensional numerical calculation by mapping the charge on a computational grid according to the particle-in-cell (PIC) method of Birdsall...... et al. It is assumed that the particle ablation from a surface with a fixed temperature takes place as a pulse, i.e. within a finite period of time. A number of characteristic quantities for the plasma plume are compared with similar data for expansion of neutrals as well as fluid models: Density...

Sorafenib paradoxically activates the RAS/RAF/ERK pathway in polyclonal human NK cells during expansion and thereby enhances effector functions in a dose- and time-dependent manner.

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Lohmeyer, J; Nerreter, T; Dotterweich, J; Einsele, H; Seggewiss-Bernhardt, R

2018-03-24

Natural killer (NK) cells play a major role in host immunity against leukaemia and lymphoma. However, clinical trials applying NK cells have not been as efficient as hoped for. Patients treated with rapidly accelerated fibrosarcoma (RAF) inhibitors exhibit increased tumour infiltration by immune cells, suggesting that a combination of RAF inhibitors with immunotherapy might be beneficial. As mitogen-activated protein kinases (MAPKs) such as raf-1 proto-oncogene, serine/threonine kinase (CRAF) regulate NK cell functions, we performed an in-vitro investigation on the potential of clinically relevant short-acting tyrosine kinase inhibitors (TKIs) as potential adjuvants for NK cell therapy: NK cells from healthy human blood donors were thus treated with sorafenib, sunitinib or the pan-RAF inhibitor ZM336372 during ex-vivo expansion. Functional outcomes assessed after washout of the drugs included cytokine production, degranulation, cytotoxicity, apoptosis induction and signal transduction with/without target cell contact. Paradoxically, sorafenib enhanced NK cell effector functions in a time- and dose-dependent manner by raising the steady-state activation level. Of note, this did not lead to NK cell exhaustion, but enhanced activity against target cells such as K562 or Daudis mediated via the RAS/RAF/extracellular-regulated kinase (ERK) pathway, but not via protein kinase B (AKT). Our data will pave the path to develop a rationale for the considered use of RAF inhibitors such as sorafenib for pre-activation in NK cell-based adoptive immune therapy. © 2018 British Society for Immunology.

CAG Expansions Are Genetically Stable and Form Nontoxic Aggregates in Cells Lacking Endogenous Polyglutamine Proteins

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Ashley A. Zurawel

2016-09-01

Full Text Available Proteins containing polyglutamine (polyQ regions are found in almost all eukaryotes, albeit with various frequencies. In humans, proteins such as huntingtin (Htt with abnormally expanded polyQ regions cause neurodegenerative diseases such as Huntington’s disease (HD. To study how the presence of endogenous polyQ aggregation modulates polyQ aggregation and toxicity, we expressed polyQ expanded Htt fragments (polyQ Htt in Schizosaccharomyces pombe. In stark contrast to other unicellular fungi, such as Saccharomyces cerevisiae, S. pombe is uniquely devoid of proteins with more than 10 Q repeats. We found that polyQ Htt forms aggregates within S. pombe cells only with exceedingly long polyQ expansions. Surprisingly, despite the presence of polyQ Htt aggregates in both the cytoplasm and nucleus, no significant growth defect was observed in S. pombe cells. Further, PCR analysis showed that the repetitive polyQ-encoding DNA region remained constant following transformation and after multiple divisions in S. pombe, in contrast to the genetic instability of polyQ DNA sequences in other organisms. These results demonstrate that cells with a low content of polyQ or other aggregation-prone proteins can show a striking resilience with respect to polyQ toxicity and that genetic instability of repetitive DNA sequences may have played an important role in the evolutionary emergence and exclusion of polyQ expansion proteins in different organisms.

Seasonal hydroclimatic impacts of Sun Corridor expansion

International Nuclear Information System (INIS)

Georgescu, M; Mahalov, A; Moustaoui, M

2012-01-01

Conversion of natural to urban land forms imparts influence on local and regional hydroclimate via modification of the surface energy and water balance, and consideration of such effects due to rapidly expanding megapolitan areas is necessary in light of the growing global share of urban inhabitants. Based on a suite of ensemble-based, multi-year simulations using the Weather Research and Forecasting (WRF) model, we quantify seasonally varying hydroclimatic impacts of the most rapidly expanding megapolitan area in the US: Arizona’s Sun Corridor, centered upon the Greater Phoenix metropolitan area. Using a scenario-based urban expansion approach that accounts for the full range of Sun Corridor growth uncertainty through 2050, we show that built environment induced warming for the maximum development scenario is greatest during the summer season (regionally averaged warming over AZ exceeds 1 °C). Warming remains significant during the spring and fall seasons (regionally averaged warming over AZ approaches 0.9 °C during both seasons), and is least during the winter season (regionally averaged warming over AZ of 0.5 °C). Impacts from a minimum expansion scenario are reduced, with regionally averaged warming ranging between 0.1 and 0.3 °C for all seasons except winter, when no warming impacts are diagnosed. Integration of highly reflective cool roofs within the built environment, increasingly recognized as a cost-effective option intended to offset the warming influence of urban complexes, reduces urban-induced warming considerably. However, impacts on the hydrologic cycle are aggravated via enhanced evapotranspiration reduction, leading to a 4% total accumulated precipitation decrease relative to the non-adaptive maximum expansion scenario. Our results highlight potentially unintended consequences of this adaptation approach within rapidly expanding megapolitan areas, and emphasize the need for undeniably sustainable development paths that account for

Soluble CD54 induces human endothelial cells ex vivo expansion useful for cardiovascular regeneration and tissue engineering application

KAUST Repository

Malara, N.M.

2015-03-01

Aim: Consistent expansion of primary human endothelial cells in vitro is critical in the development of engineered tissue. A variety of complex culture media and techniques developed from different basal media have been reported with alternate success. Incongruous results are further confounded by donor-to-donor variability and cellular source of derivation. Our results demonstrate how to overcome these limitations using soluble CD54 (sCD54) as additive to conventional culture medium. Methods and results: Isolated primary fragment of different vessel types was expanded in Ham\\'s F12 DMEM, enriched with growth factors, Fetal Calf Serum and conditioned medium of Human Umbilical Vein Endothelial Cells (HUVEC) collected at different passages. Cytokine content of culture media was analyzed in order to identify the soluble factors correlating with better proliferation profile. sCD54 was found to induce the in vitro expansion of human endothelial cells (HECs) independently from the vessels source and even in the absence of HUVEC-conditioned medium. The HECs cultivated in the presence of sCD54 (50 ng/ml), resulted positive for the expression of CD146 and negative for CD45, and lower fibroblast contamination. Cells were capable to proliferate with an S phase of 25%, to produce vascular endothelial growth factor, VEGF, (10 ng/ml) and to give origin to vessel-like tubule in vitro. Conclusion: Our results demonstrate that sCD54 is an essential factor for the in-vitro expansion of HECs without donor and vessel-source variability. Resulting primary cultures can be useful, for tissue engineering in regenerative medicine (e.g. artificial micro tissue generation, coating artificial heart valve etc.) and bio-nanotechnology applications. © 2015 The Authors. Published by Elsevier Ireland Ltd.

Chimeric Antigen Receptor Therapy for B-cell Malignancies

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David L Porter, Michael Kalos, Zhaohui Zheng, Bruce Levine, Carl June

2011-01-01

Full Text Available We presented data showing that the CART-19 cells expressing the 4-1BB signaling domain can have unprecedented and massive in-vivo expansion, traffic to tumor sites, persist long term in vivo, and induce rapid and potent anti-tumor activity in chemotherapy refractory CLL patients.

Rapid Column-Free Enrichment of Mononuclear Cells from Solid Tissues

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Scoville, Steven D.; Keller, Karen A.; Cheng, Stephanie; Zhang, Michael; Zhang, Xiaoli; Caligiuri, Michael A.; Freud, Aharon G.

2015-01-01

We have developed a rapid negative selection method to enrich rare mononuclear cells from human tissues. Unwanted and antibody-tethered cells are selectively depleted during a Ficoll separation step, and there is no need for magnetic-based reagents and equipment. The new method is fast, customizable, inexpensive, remarkably efficient, and easy to perform, and per sample the overall cost is less than one-tenth the cost associated with a magnetic column-based method. PMID:26223896

Infliximab induces clonal expansion of γδ-T cells in Crohn's disease: a predictor of lymphoma risk?

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Jens Kelsen

Full Text Available BACKGROUND: Concominant with the widespread use of combined immunotherapy in the management of Crohn's disease (CD, the incidence of hepato-splenic gamma-delta (γδ-T cell lymphoma has increased sharply in CD patients. Malignant transformation of lymphocytes is believed to be a multistep process resulting in the selection of malignant γδ-T cell clones. We hypothesised that repeated infusion of anti-TNF-α agents may induce clonal selection and that concurrent treatment with immunomodulators further predisposes patients to γδ-T cell expansion. METHODOLOGY/PRINCIPAL FINDINGS: We investigated dynamic changes in the γδ-T cells of patient with CD following treatment with infliximab (Remicade®; n=20 or adalimumab (Humira®; n=26 using flow cytometry. In patients with a high γδ-T cell level, the γδ-T cells were assessed for clonality. Of these 46 CD patients, 35 had a γδ-T cells level (mean 1.6% comparable to healthy individuals (mean 2.2%, and 11 CD patients (24% exhibited an increased level of γδ-T cells (5-15%. In the 18 patients also receiving thiopurines or methotrexate, the average baseline γδ-T cell level was 4.4%. In three male CD patients with a high baseline value, the γδ-T cell population increased dramatically following infliximab therapy. A fourth male patient also on infliximab monotherapy presented with 20% γδ-T cells, which increased to 25% shortly after treatment and was 36% between infusions. Clonality studies revealed an oligoclonal γδ-T cell pattern with dominant γδ-T cell clones. In support of our clinical findings, in vitro experiments showed a dose-dependent proliferative effect of anti-TNF-α agents on γδ-T cells. CONCLUSION/SIGNIFICANCE: CD patients treated with immunomodulators had constitutively high levels of γδ-T cells. Infliximab exacerbated clonal γδ-T cell expansion in vivo and induced γδ-T cell proliferation in vitro. Overall, young, male CD patients with high baseline γδ-T cell

Infliximab induces clonal expansion of γδ-T cells in Crohn's disease: a predictor of lymphoma risk?

Science.gov (United States)

Kelsen, Jens; Dige, Anders; Schwindt, Heinrich; D'Amore, Francesco; Pedersen, Finn S; Agnholt, Jørgen; Christensen, Lisbet A; Dahlerup, Jens F; Hvas, Christian L

2011-03-31

Concominant with the widespread use of combined immunotherapy in the management of Crohn's disease (CD), the incidence of hepato-splenic gamma-delta (γδ)-T cell lymphoma has increased sharply in CD patients. Malignant transformation of lymphocytes is believed to be a multistep process resulting in the selection of malignant γδ-T cell clones. We hypothesised that repeated infusion of anti-TNF-α agents may induce clonal selection and that concurrent treatment with immunomodulators further predisposes patients to γδ-T cell expansion. We investigated dynamic changes in the γδ-T cells of patient with CD following treatment with infliximab (Remicade®; n=20) or adalimumab (Humira®; n=26) using flow cytometry. In patients with a high γδ-T cell level, the γδ-T cells were assessed for clonality. Of these 46 CD patients, 35 had a γδ-T cells level (mean 1.6%) comparable to healthy individuals (mean 2.2%), and 11 CD patients (24%) exhibited an increased level of γδ-T cells (5-15%). In the 18 patients also receiving thiopurines or methotrexate, the average baseline γδ-T cell level was 4.4%. In three male CD patients with a high baseline value, the γδ-T cell population increased dramatically following infliximab therapy. A fourth male patient also on infliximab monotherapy presented with 20% γδ-T cells, which increased to 25% shortly after treatment and was 36% between infusions. Clonality studies revealed an oligoclonal γδ-T cell pattern with dominant γδ-T cell clones. In support of our clinical findings, in vitro experiments showed a dose-dependent proliferative effect of anti-TNF-α agents on γδ-T cells. CD patients treated with immunomodulators had constitutively high levels of γδ-T cells. Infliximab exacerbated clonal γδ-T cell expansion in vivo and induced γδ-T cell proliferation in vitro. Overall, young, male CD patients with high baseline γδ-T cell levels may be at an increased risk of developing malignant γδ-T cell lymphomas

Investigation of droplet nucleation in CCS relevant systems – design and testing of the expansion chamber

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Čenský Miroslav

2018-01-01

Full Text Available A unique in-house designed experimental apparatus for investigation of nucleation of droplets in CCS relevant systems is being developed by the present team. The apparatus allows simulating various processes relevant to CCS technologies. Gaseous mixtures with CO2 are prepared in a Mixture Preparation Device (MPD based on accurate adjustment of flow rates of individual components [EPJ Web of Conferences 143, 02140 (2017]. The mixture then flows into an expansion chamber, where it undergoes a rapid adiabatic expansion. As a consequence of adiabatic cooling, the mixture becomes supersaturated and nucleation and simultaneous growth of droplets occurs. In this study, we describe the design and testing of the expansion part of the experimental setup. The rapid expansion was realized using two valve systems, one for low pressures (up to 0.7 MPa and the other for high pressures (up to 10 MPa. A challenge for a proper design of the expansion system is avoiding acoustic oscillations. These can occur either in the mode of Helmholtz resonator, where the compressible gas in the chamber acts as a spring and the rapidly moving gas in the valve system as a mass, or in the “flute” mode, where acoustic waves are generated in a long outlet tubing.

Correlations between Socioeconomic Drivers and Indicators of Urban Expansion: Evidence from the Heavily Urbanised Shanghai Metropolitan Area, China

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Jinghui Li

2017-07-01

Full Text Available Rapid urban expansion resulting in increased impervious surfaces causes a series of urban environmental problems, e.g., the urban heat island and urban forest fragmentation. Urban expansion is a serious threat to human quality of life and living environments. It has been studied from a variety of aspects, but its driving factors and time series expansion characteristics (i.e., expansion intensity, pattern and direction need to be better explained in order to devise more effective management strategies. This study examined how social and economic factors are linked in driving urban expansion. Based on multi-temporal aerial images, a rapid urban expansion period, 2000–2010, in Shanghai was analysed. The urban area expanded from 1770.36 to 2855.44 km2 in the period, with a mean annual expansion rate of 108.51 km2. Urban expansion in 2000–2005 (40.42% was much faster than in 2005–2010 (14.86%, and its direction was southeast, southwest and south. The main pattern was edge expansion in both sub-periods. Social factors, especially population density, significantly affected urban expansion. These findings can help understand the urban expansion process and its driving factors, which has important implications for urban planning and management in Shanghai and similar cities.

Evolution of density-dependent movement during experimental range expansions.

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Fronhofer, E A; Gut, S; Altermatt, F

2017-12-01

Range expansions and biological invasions are prime examples of transient processes that are likely impacted by rapid evolutionary changes. As a spatial process, range expansions are driven by dispersal and movement behaviour. Although it is widely accepted that dispersal and movement may be context-dependent, for instance density-dependent, and best represented by reaction norms, the evolution of density-dependent movement during range expansions has received little experimental attention. We therefore tested current theory predicting the evolution of increased movement at low densities at range margins using highly replicated and controlled range expansion experiments across multiple genotypes of the protist model system Tetrahymena thermophila. Although rare, we found evolutionary changes during range expansions even in the absence of initial standing genetic variation. Range expansions led to the evolution of negatively density-dependent movement at range margins. In addition, we report the evolution of increased intrastrain competitive ability and concurrently decreased population growth rates in range cores. Our findings highlight the importance of understanding movement and dispersal as evolving reaction norms and plastic life-history traits of central relevance for range expansions, biological invasions and the dynamics of spatially structured systems in general. © 2017 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2017 European Society For Evolutionary Biology.

Sub-cell balanced nodal expansion methods using S4 eigenfunctions for multi-group SN transport problems in slab geometry

International Nuclear Information System (INIS)

Hong, Ser Gi; Lee, Deokjung

2015-01-01

A highly accurate S 4 eigenfunction-based nodal method has been developed to solve multi-group discrete ordinate neutral particle transport problems with a linearly anisotropic scattering in slab geometry. The new method solves the even-parity form of discrete ordinates transport equation with an arbitrary S N order angular quadrature using two sub-cell balance equations and the S 4 eigenfunctions of within-group transport equation. The four eigenfunctions from S 4 approximation have been chosen as basis functions for the spatial expansion of the angular flux in each mesh. The constant and cubic polynomial approximations are adopted for the scattering source terms from other energy groups and fission source. A nodal method using the conventional polynomial expansion and the sub-cell balances was also developed to be used for demonstrating the high accuracy of the new methods. Using the new methods, a multi-group eigenvalue problem has been solved as well as fixed source problems. The numerical test results of one-group problem show that the new method has third-order accuracy as mesh size is finely refined and it has much higher accuracies for large meshes than the diamond differencing method and the nodal method using sub-cell balances and polynomial expansion of angular flux. For multi-group problems including eigenvalue problem, it was demonstrated that the new method using the cubic polynomial approximation of the sources could produce very accurate solutions even with large mesh sizes. (author)

In vitro expansion of Lin{sup +} and Lin{sup −} mononuclear cells from human peripheral blood

Energy Technology Data Exchange (ETDEWEB)

Norhaiza, H. Siti; Zarina, Z. A. Intan; Hisham, Z. A. Shahrul [School of Bioscience and Biotechnology, Faculty of Science and Technology, Universiti Kebangsaan Malaysia, 43600, Selangor (Malaysia); Rohaya, M. A. W. [Department of Orthodontics, Faculty of Dentistry, Universiti Kebangsaan Malaysia, 50300, Kuala Lumpur (Malaysia)

2013-11-27

Haematopoietic stem cells (HSCs) are used in the therapy of blood disorders due to the ability of these cells to reconstitute haematopoietic lineage cells when transplanted into myeloablative recipients. However, substantial number of cells is required in order for the reconstitution to take place. Since HSCs present in low frequency, larger number of donor is required to accommodate the demand of transplantable HSCs. Therefore, in vitro expansion of HSCs will have profound impact on clinical purposes. The aim of this study was to expand lineage negative (Lin{sup −}) stem cells from human peripheral blood. Total peripheral blood mononuclear cells (PBMNCs) were fractionated from human blood by density gradient centrifugation. Subsequently, PBMNCs were subjected to magnetic assisted cell sorter (MACS) which depletes lineage positive (Lin{sup +}) mononuclear cells expressing lineage positive markers such as CD2, CD3, CD11b, CD14, CD15, CD16, CD19, CD56, CD123, and CD235a to obtained Lin{sup −} cell population. The ability of Lin{sup +} and Lin{sup −} to survive in vitro was explored by culturing both cell populations in complete medium consisting of Alpha-Minimal Essential Medium (AMEM) +10% (v/v) Newborn Calf Serum (NBCS)+ 2% (v/v) pen/strep. In another experiment, Lin{sup +} and Lin{sup −} were cultured with complete medium supplemented with 10ng/mL of the following growth factors: stem cell factor (SCF), interleukin (IL)-3, granulocyte-macrophage colony stimulating factor (GM-CSF), 2IU/mL of Erythropoietin (Epo) and 20ng/mL of IL-6. Three samples were monitored in static culture for 22 days. The expansion potential was assessed by the number of total viable cells, counted by trypan blue exclusion assay. It was found that Lin{sup +} mononuclear cells were not able to survive either in normal proliferation medium or proliferation medium supplemented with cytokines. Similarly, Lin{sup −} stem cells were not able to survive in proliferation medium however

ERK/CANP rapid signaling mediates 17β-estradiol-induced proliferation of human breast cancer cell line MCF-7 cells.

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Wang, Guo-Sheng; Huang, Yan-Gang; Li, Huan; Bi, Shi-Jie; Zhao, Jin-Long

2014-01-01

17β-estradiol (E2) exerts its functions through both genomic and non-genomic signaling pathways. Because E2 is important in breast cancer development, we investigated whether its actions in promoting breast cancer cell proliferation occur through the non-genomic signaling pathway via extracellular signal-regulated kinase 1/2 (ERK1/2)/calcium-activated neutral protease (CANP). MCF-7 breast cancer cells were treated with ERKl/2 inhibitor (PD98059) or CANP inhibitor (calpeptin) before exposure to 1×10(-8) M E2. MTT colorimetry and flow cytometry were used to analyze effects on cell proliferation and cell cycle progression, respectively. Expression of phosphorylated-ERK (p-ERK), total ERK, and Capn4 proteins were assessed by Western blotting. Cell proliferation increased in cells treated with E2 for 24 h (P<0.05), and the proportion of cells in G0/G1 was decreased, accompanied by accelerated G1/S. Calpeptin pre-treatment significantly inhibited the E2-induced proliferation of MCF-7 cells (P<0.05), while also ameliorating the effects of E2 on cell cycle progression. Further, expression of p-ERK was rapidly up-regulated (after 10 min) by E2 (P<0.05), an effect that persisted 16 h after E2 exposure but which was significantly inhibited by PD98059 (P<0.05). Finally, expression of Capn4 protein was rapidly up-regulated in E2-exposed cells (P<0.05), but this change was significantly inhibited by PD98059 or calpeptin (P<0.05) pre-treatment. Thus, the rapid, non-genomic ERK/CANP signaling pathway mediates E2-induced proliferation of human breast cancer cells.

Cost-effective and rapid blood analysis on a cell-phone.

Science.gov (United States)

Zhu, Hongying; Sencan, Ikbal; Wong, Justin; Dimitrov, Stoyan; Tseng, Derek; Nagashima, Keita; Ozcan, Aydogan

2013-04-07

We demonstrate a compact and cost-effective imaging cytometry platform installed on a cell-phone for the measurement of the density of red and white blood cells as well as hemoglobin concentration in human blood samples. Fluorescent and bright-field images of blood samples are captured using separate optical attachments to the cell-phone and are rapidly processed through a custom-developed smart application running on the phone for counting of blood cells and determining hemoglobin density. We evaluated the performance of this cell-phone based blood analysis platform using anonymous human blood samples and achieved comparable results to a standard bench-top hematology analyser. Test results can either be stored on the cell-phone memory or be transmitted to a central server, providing remote diagnosis opportunities even in field settings.

Light-induced lattice expansion leads to high-efficiency perovskite solar cells

Energy Technology Data Exchange (ETDEWEB)

Tsai, Hsinhan; Asadpour, Reza; Blancon, Jean-Christophe; Stoumpos, Constantinos C.; Durand, Olivier; Strzalka, Joseph W.; Chen, Bo; Verduzco, Rafael; Ajayan, Pulickel M.; Tretiak, Sergei; Even, Jacky; Alam, Muhammad Ashraf; Kanatzidis, Mercouri G.; Nie, Wanyi; Mohite, Aditya D.

2018-04-05

Hybrid-perovskite based high-performance optoelectronic devices and clues from their operation has led to the realization that light-induced structural dynamics play a vital role on their physical properties, device performance and stability. Here, we report that continuous light illumination leads to a uniform lattice expansion in hybrid perovskite thin-films, which is critical for obtaining high-efficiency photovoltaic devices. Correlated, in-situ structural and device characterizations reveal that light-induced lattice expansion significantly benefits the performances of a mixed-cation pure-halide planar device, boosting the power conversion efficiency from 18.5% to 20.5%. This is a direct consequence of the relaxation of local lattice strains during lattice expansion, which results in the reduction of the energetic barriers at the perovskite/contact interfaces in devices, thus improving the open circuit voltage and fill factor. The light-induced lattice expansion stabilizes these high-efficiency photovoltaic devices under continuous operation of full-spectrum 1-Sun illumination for over 1500 hours. One Sentence Summary: Light-induced lattice expansion improves crystallinity, relaxes lattice strain, which enhances photovoltaic performance in hybrid perovskite device.

Comparison of the effects of sedation and general anesthesia in surgically assisted rapid palatal expansion.

Science.gov (United States)

Satilmis, Tulin; Ugurlu, Faysal; Garip, Hasan; Sener, Bedrettin C; Goker, Kamil

2011-06-01

To compare the effects of sedation and general anesthesia for surgically assisted rapid palatal expansion (SARPE). This randomized prospective study included 30 patients who were scheduled for SARPE, and was performed between January 2008 to February 2010 in the Department of Oral and Maxillofacial Surgery, Faculty of Dentistry, Marmara University, Istanbul, Turkey. Patients were allocated into Group S - midazolam + fentanyl sedation (n=15), and Group G - general anesthesia (n=15). Hemodynamic parameters, duration of anesthesia, surgery, recovery time, time to discharge, visual analogue scale (VAS) pain scores at 30 minutes (min), one hour (hr), 4 hours, 12 hours, and 24 hours, first consumption of analgesic time, total amount of consumption of analgesics, patient and surgeon satisfaction, nausea, and vomiting were recorded. Analgesic time was significantly longer in Group S (p=0.008), and total analgesic consumption was significantly lower in Group S than in Group G (p=0.031). Patient satisfaction was statistically higher in Group S (p=0.035). At 30 min, one hr, and 12 hrs, VAS satisfaction scores in Group S were statistically lower than those in Group G, and at 4 hrs and 24 hrs there was no statistical difference in VAS scores for both groups. The use of sedation for outpatient SARPE resulted in lower pain scores at discharge, lower analgesic consumption, and greater patient satisfaction.

The location of splenic NKT cells favours their rapid activation by blood-borne antigen

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Barral, Patricia; Sánchez-Niño, María Dolores; van Rooijen, Nico; Cerundolo, Vincenzo; Batista, Facundo D

2012-01-01

Natural killer T (NKT) cells play an important role in mounting protective responses to blood-borne infections. However, though the spleen is the largest blood filter in the body, the distribution and dynamics of NKT cells within this organ are not well characterized. Here we show that the majority of NKT cells patrol around the marginal zone (MZ) and red pulp (RP) of the spleen. In response to lipid antigen, these NKT cells become arrested and rapidly produce cytokines, while the small proportion of NKT cells located in the white pulp (WP) exhibit limited activation. Importantly, disruption of the splenic MZ by chemical or genetic approaches results in a severe reduction in NKT cell activation indicating the need of cooperation between both MZ macrophages and dendritic cells for efficient NKT cell responses. Thus, the location of splenic NKT cells in the MZ and RP facilitates their access to blood-borne antigen and enables the rapid initiation of protective immune responses. PMID:22505026

The location of splenic NKT cells favours their rapid activation by blood-borne antigen.

Science.gov (United States)

Barral, Patricia; Sánchez-Niño, María Dolores; van Rooijen, Nico; Cerundolo, Vincenzo; Batista, Facundo D

2012-05-16

Natural killer T (NKT) cells play an important role in mounting protective responses to blood-borne infections. However, though the spleen is the largest blood filter in the body, the distribution and dynamics of NKT cells within this organ are not well characterized. Here we show that the majority of NKT cells patrol around the marginal zone (MZ) and red pulp (RP) of the spleen. In response to lipid antigen, these NKT cells become arrested and rapidly produce cytokines, while the small proportion of NKT cells located in the white pulp (WP) exhibit limited activation. Importantly, disruption of the splenic MZ by chemical or genetic approaches results in a severe reduction in NKT cell activation indicating the need of cooperation between both MZ macrophages and dendritic cells for efficient NKT cell responses. Thus, the location of splenic NKT cells in the MZ and RP facilitates their access to blood-borne antigen and enables the rapid initiation of protective immune responses.

Separable pole expansions in four-nucleon bound state calculations

International Nuclear Information System (INIS)

Sofianos, S.A.; Fiedeldey, H.; Haberzettl, H.; Sandhas, W.

1982-04-01

We compare the utility of the Generalized Unitary Pole Expansion (GUPE) and the Energy-Dependent Pole Expansion (EDPE) for the three-body subsystem amplitudes in four-body state calculations for a variety of separable and local nucleon-nucleon interactions. It is found that, with the EDPE, the four-body binding energy is well reproduced with only two terms each for the (2+2)- and the (3+1)-subsystem, respectively, while the GUPE requires three terms for the (3+1)-channel and four terms for the (2+2)-channel. We thus conclude that pole dominance is of greater importance for the GUPE than for EDPE, which works equally well for both types of subsystems. It is found that both methods, in particular the EDPE, converge more rapidly with increasing repulsion in the two-body interaction, i.e. the more realistic the interaction becomes. Both expansions require similar computing times for a converged calculation and are about 15-20 times faster than the widely used Hilbert-Schmidt Expansion (HSE). The respective merits of the two pole expansions are discussed and compared with the HSE. (orig.)

Quantitative analysis of topoisomerase IIα to rapidly evaluate cell proliferation in brain tumors

International Nuclear Information System (INIS)

Oda, Masashi; Arakawa, Yoshiki; Kano, Hideyuki; Kawabata, Yasuhiro; Katsuki, Takahisa; Shirahata, Mitsuaki; Ono, Makoto; Yamana, Norikazu; Hashimoto, Nobuo; Takahashi, Jun A.

2005-01-01

Immunohistochemical cell proliferation analyses have come into wide use for evaluation of tumor malignancy. Topoisomerase IIα (topo IIα), an essential nuclear enzyme, has been known to have cell cycle coupled expression. We here show the usefulness of quantitative analysis of topo IIα mRNA to rapidly evaluate cell proliferation in brain tumors. A protocol to quantify topo IIα mRNA was developed with a real-time RT-PCR. It took only 3 h to quantify from a specimen. A total of 28 brain tumors were analyzed, and the level of topo IIα mRNA was significantly correlated with its immuno-staining index (p < 0.0001, r = 0.9077). Furthermore, it sharply detected that topo IIα mRNA decreased in growth-inhibited glioma cell. These results support that topo IIα mRNA may be a good and rapid indicator to evaluate cell proliferate potential in brain tumors

In vitro expansion of the mammary stem/progenitor cell population by xanthosine treatment

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Choudhary Ratan K

2012-06-01

Full Text Available Abstract Background Mammary stem cells are critical for growth and maintenance of the mammary gland and therefore are of considerable interest for improving productivity and efficiency of dairy animals. Xanthosine treatment has been demonstrated to promote expansion of putative mammary stem cells in vivo, and hepatic and hair follicle stem cells in vitro. In the latter, xanthosine promoted the symmetrical division of hepatic and hair follicle stem cells. The objective of this study was to determine if treating primary cultures of bovine mammary epithelial cells (MEC with xanthosine increases the stem/progenitor cell population by promoting symmetrical division of mammary stem cells. Results In vitro treatment with xanthosine increased the population of MEC during the exponential phase of cell growth, reducing the doubling time from 86 h in control cultures to 60 h in xanthosine-treated cultures. The bromodeoxyuridine (BrdU labeling index and the proportion of MEC in S-phase both were increased by xanthosine treatment, indicating that increased cell accretion was due to increased cell proliferation. Analysis of daughter-pairs indicated that xanthosine promoted a shift from asymmetric to symmetric cell division. Moreover, the 30 % increase in symmetric cell division was concomitant with an increase in the proportion of MEC that were positive for a putative stem cell marker (FNDC3B and a trend toward increased telomerase activity. These results suggest that xanthosine treatment in vitro can increase cell proliferation, promote symmetric cell division and enhance stem/progenitor cell activity. Conclusions Xanthosine treatment increased the proliferation rate of bovine MEC in vitro. This was likely to be mediated by an increase in the proportion of stem/progenitor cells in the MEC population due to promotion of symmetrical stem cell division by xanthosine.

Rapid host immune response and viral dynamics in herpes simplex virus-2 infection

Science.gov (United States)

Schiffer, Joshua T; Corey, Lawrence

2014-01-01

Herpes Simplex Virus-2 (HSV-2) is episodically shed throughout the human genital tract. While high viral load correlates with development of genital ulcers, shedding also commonly occurs even when ulcers are not present, allowing for silent transmission during coitus and contributing to high seroprevalence of HSV-2 worldwide. Frequent viral reactivation occurs despite diverse and complementary host and viral mechanisms within ganglionic tissue that predispose towards latency, suggesting that viral replication may be constantly occurring in a small minority of neurons within the ganglia. Within genital mucosa, the in vivo expansion and clearance rates of HSV-2 are extremely rapid. Resident dendritic cells and memory HSV-specific T cells persist at prior sites of genital tract reactivation, and in conjunction with prompt innate recognition of infected cells, lead to rapid containment of infected cells. Shedding episodes vary greatly in duration and severity within a single person over time: this heterogeneity appears best explained by variation in the densities of host immunity across the genital tract. The fact that immune responses usually control viral replication in genital skin prior to development of lesions provides optimism that enhancing such responses could lead to effective vaccines and immunotherapies. PMID:23467247

Relapse and stability of surgically assisted rapid maxillary expansion, an anatomical biomechanical study

NARCIS (Netherlands)

Koudstaal, M.J.; Smeets, J.B.J.; Kleinrensink, G.J.; Schulten, A.J.M.; van der Wal, K.G.H.

2009-01-01

Purpose: This anatomic biomechanical study was undertaken to gain insight into the underlining mechanism of tipping of the maxillary segments during transverse expansion using tooth-borne and bone-borne distraction devices. Materials and Methods: An anatomic biomechanical study was performed on 10

Correction of complete maxillary crossbite with severe crowding using Hyrax expansion and fixed appliance.

Science.gov (United States)

Wangsrimongkol, Tasanee; Manosudprasit, Montian; Pisek, Poonsak; Leelasinjaroen, Pornnapha

2013-09-01

An 18-year-old Thai man who presented with a secondary cleft palate, maxillary hypoplasia and severe crowding was treated by rapid maxillary expansion and fixed orthodontic appliances. Initial assessment found skeletal Class III malrelationship and dental Class II malocclusion with anterior and bilateral posterior crossbites. Camouflage orthodontic treatment was planned using a rapid maxillary expansion appliance and correcting crowding with extraction all four premolar teeth. A Hyrax appliance and vertical loop arch wire were placed for maxillary arch expansion of 9.5 mms at first molars and canines, and 5.5 mms at the premolars and obtained positive overjet. Both acceptable skeletal and soft tissue relationships and satisfactory occlusion were produced. After 14 months of postoperative follow-up, the occlusal result was stable and no skeletal reversals could be detected.

HEXIM1 controls satellite cell expansion after injury to regulate skeletal muscle regeneration

Science.gov (United States)

Hong, Peng; Chen, Kang; Huang, Bihui; Liu, Min; Cui, Miao; Rozenberg, Inna; Chaqour, Brahim; Pan, Xiaoyue; Barton, Elisabeth R.; Jiang, Xian-Cheng; Siddiqui, M.A.Q.

2012-01-01

The native capacity of adult skeletal muscles to regenerate is vital to the recovery from physical injuries and dystrophic diseases. Currently, the development of therapeutic interventions has been hindered by the complex regulatory network underlying the process of muscle regeneration. Using a mouse model of skeletal muscle regeneration after injury, we identified hexamethylene bisacetamide inducible 1 (HEXIM1, also referred to as CLP-1), the inhibitory component of the positive transcription elongation factor b (P-TEFb) complex, as a pivotal regulator of skeletal muscle regeneration. Hexim1-haplodeficient muscles exhibited greater mass and preserved function compared with those of WT muscles after injury, as a result of enhanced expansion of satellite cells. Transplanted Hexim1-haplodeficient satellite cells expanded and improved muscle regeneration more effectively than WT satellite cells. Conversely, HEXIM1 overexpression restrained satellite cell proliferation and impeded muscle regeneration. Mechanistically, dissociation of HEXIM1 from P-TEFb and subsequent activation of P-TEFb are required for satellite cell proliferation and the prevention of early myogenic differentiation. These findings suggest a crucial role for the HEXIM1/P-TEFb pathway in the regulation of satellite cell–mediated muscle regeneration and identify HEXIM1 as a potential therapeutic target for degenerative muscular diseases. PMID:23023707

Expansion of PD-1-positive effector CD4 T cells in an experimental model of SLE: contribution to the self-organized criticality theory.

Science.gov (United States)

Miyazaki, Yumi; Tsumiyama, Ken; Yamane, Takashi; Ito, Mitsuhiro; Shiozawa, Shunichi

2013-04-18

We have developed a systems biology concept to explain the origin of systemic autoimmunity. From our studies of systemic lupus erythematosus (SLE) we have concluded that this disease is the inevitable consequence of over-stimulating the host's immune system by repeated exposure to antigen to levels that surpass a critical threshold, which we term the system's "self-organized criticality". We observed that overstimulation of CD4 T cells in mice led to the development of autoantibody-inducing CD4 T cells (aiCD4 T) capable of generating various autoantibodies and pathological lesions identical to those observed in SLE. We show here that this is accompanied by the significant expansion of a novel population of effector T cells characterized by expression of programmed death-1 (PD-1)-positive, CD27(low), CD127(low), CCR7(low) and CD44(high)CD62L(low) markers, as well as increased production of IL-2 and IL-6. In addition, repeated immunization caused the expansion of CD8 T cells into fully-matured cytotoxic T lymphocytes (CTL) that express Ly6C(high)CD122(high) effector and memory markers. Thus, overstimulation with antigen leads to the expansion of a novel effector CD4 T cell population that expresses an unusual memory marker, PD-1, and that may contribute to the pathogenesis of SLE.

Evaluation of Peripheral Blood and Cord Blood Platelet Lysates in Isolation and Expansion of Multipotent Mesenchymal Stromal Cells

Directory of Open Access Journals (Sweden)

Ioanna Christou

2018-02-01

Full Text Available Background: Multipotent Mesenchymal Stromal Cells (MSCs are used in tissue engineering and regenerative medicine. The in vitro isolation and expansion of MSCs involve the use of foetal bovine serum (FBS. However, many concerns have been raised regarding the safety of this product. In this study, alternative additives derived either from peripheral or cord blood were tested as an FBS replacement. Methods: Platelet lysates (PL from peripheral and cord blood were used for the expansion of MSCs. The levels of growth factors in peripheral blood (PB and cord blood (CB PLs were determined using the Multiple Reaction Monitoring (MRM. Finally, the cell doubling time (CDT, tri-lineage differentiation and phenotypic characterization of the MSCs expanded with FBS and PLs were determined. Results: MSCs treated with culture media containing FBS and PB-PL, were successfully isolated and expanded, whereas MSCs treated with CB-PL could not be maintained in culture. Furthermore, the MRM analysis yielded differences in growth factor levels between PB-PL and CB-PL. In addition, the MSCs were successfully expanded with FBS and PB-PL and exhibited tri-lineage differentiation and stable phenotypic characteristics. Conclusion: PB-PL could be used as an alternative additive for the production of MSCs culture medium applied to xenogeneic-free expansion and maintenance of MSCs in large scale clinical studies.

Comparison of two protocols for maxillary protraction: bone anchors versus face mask with rapid maxillary expansion