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Adipose tissue-derived mesenchymal stem cells (ADMSCs) enhance tissue healing and approximation in stomach: 脂肪組織來源的間充質幹細胞促進胃損傷愈合的相關性研究 / Liu, Liu / 脂肪組織來源的間充質幹細胞促進胃損傷愈合的相關性研究 / CUHK electronic theses & dissertations collection / Adipose tissue-derived mesenchymal stem cells (ADMSCs) enhance tissue healing and approximation in stomach: Zhi fang zu zhi lai yuan de jian chong zhi gan xi bao cu jin wei sun shang yu he de xiang guan xing yan jiu / Zhi fang zu zhi lai yuan de jian chong zhi gan xi bao cu jin wei sun shang yu he de xiang guan xing yan jiuJanuary 2014 (has links)
Introduction. Safe closure of gastric luminal defects remains a big challenge for development of gastric endoscopic surgery. The aims of this thesis are to assess the effect and efficiency of Eagle Claw VIII (endoscopic suturing device)and adipose tissue-derived mesenchymal stem cells (ADMSCs) for closure and enhancing healing of gastric luminal defects. / Methods and Results. 1. Endoscopic suturing is superior to endoclips for closure of gastrotomy after NOTES. A 2cm linear incision on the body of porcine stomach was closed by hand suturing, Eagle Claw VIII or endoclips, respectively (n=17 for each group). The results indicated that all gastrotomies were successfully closed. Closure time was significantly longer in Eagle Claw VIII group. Bursting pressure of gastrotomies for Eagle Claw VIII was significantly higher than endoclips, but lower than hand suturing. Besides, both Eagle Claw VIII and endoclip closure encountered significantly technical challenges. This study suggested that Eagle Claw VIII had potential for endoscopic closure of gastrotomies, but need further refinement. / 2. ADMSCs for Acceleration of Healing of Sutured Gastric Perforation(SGP). ADMSCs were isolated and expanded in vitro, and characterized by stromal differentiations and cell surface markers. A 2cm SGP was produced on gastric body of rats. 5×10⁶ ADMSCs were transplanted into SGP by local injection (LI-ADMSCs) or topical spraying (TS-ADMSCs). Healing of SGP was assessed. LI-ADMCs significantly decreased peritoneal adhesion and wound dehiscence, and increased bursting pressure of SGP, when comparing to other experimental groups. Histologic analysis indicated that SGPs in LI-ADMSCs group had more re-epithelialization and collagen regeneration, and less inflammation. Expression of TGF-β1 was up-regulated, while IL-6 was down-regulated in LI-ADMSCs group, when comparing to fibrin and control groups. This study suggested that local injection of ADMSCs is an effective approach for accelerating the healing of SGP. / 3. Promoting Effect of ADMSCs on Healing of Gastric Ulcer is abrogated by NSAIDs. Gastric ulcer model in rats was successfully produced by using 70% acetic acid. A total of 1×10⁷ ADMSCs was locally injected into ulcer lesion. Ulcer area was measured at different time points. Therapeutic potentialof ADMSCs was assessed when NSAIDs was simultaneously administrated. The results demonstrated that ADMSCs significantly decreased ulcer area. Histologic assessment indicated that ADMSCs increased re-epithelialization, angiogenesis and collagen deposition, and suppressed inflammation. Transplanted ADMSCs homed into gastric ulcer lesion and differentiated into endothelial and smooth muscle cells. In addition, ADMSCs treatment increased the gene expressions for wound healing, and activated COX-2-PGE₂ and Erk1/2-MAPK signaling pathways. Repeated administration of Indomethacin reduced cell proliferation and angiogenesis, and eliminated ADMSCs-induced ulcer healing on day 10. The results suggested that ADMSCs promoted the healing of peptic ulcer, which is eliminated by NSAIDs. / Conclusions. Endoscopic suturing by Eagle Claw VIII is feasible for closure of gastrotomy, when comparing to endoclips. ADMSC promotes the healing of gastric luminal defects including SGP and ulcer. The promoting effect of ADMSC is PGE₂-dependent, and attenuated by NSAIDs. These evidences implied that combined use of endoscopic suturing and ADMSCs is a helpful approach for safe closure of gastrotomy and gastric perforation. / 引言:胃傷口癒合是胃消化內鏡手術發展的障礙之壹。本課題之目的是評價和探索Eagle Claw VIII和脂肪幹細胞(ADMSCs)縫合和促進胃內傷口癒合的效果和作用。 / 方法和結果:1. 內鏡縫合器Eagle Claw VIII閉合經胃自然腔道手術後傷口的效果評價體外豬胃體上造2cm的胃傷口模型,使用手工縫合、內鏡下Eagle Claw VIII縫合或內鏡夾閉合胃傷口;每組17個樣本。本研究提示所有胃傷口均成功閉合。Eagle Claw VIII縫合胃傷口時間顯著長於其他兩組的閉合時間。Eagle Claw VIII縫合的胃傷口破裂壓顯著高於內鏡夾閉組,但是明顯低於手工縫合組。此外,內鏡縫合和夾閉都面臨較大的技術難度。本研究提示Eagle Claw VIII有臨床運用的潛在價值,但需要進壹步改進。 / 2. 局部移植脂肪幹細胞促進胃穿孔癒合的實驗性研究:建立大鼠2cm胃體穿孔模型,局部註射或傷口表面塗抹法移植ADMSCs,觀察胃傷口癒合情況。局部註射移植ADMSCs顯著減輕胃傷口粘連和裂開發生率,增加胃傷口破裂壓。組織學分析提示ADMSCs治療促進傷口上皮和肉芽組織再生,抑制炎癥反應。此外,局部註射ADMSCs增加TGF-β1抑制IL-6表達。本研究提示局部註射移植ADMSCs是促進胃穿孔傷口癒合的有效方法。 / 3. 局部移植脂肪幹細胞促進胃饋瘍癒合的實驗性研究:使用70%醋酸建立大鼠胃體饋瘍模型;饋瘍病竈內局部註射移植1×107 ADMSCs。研究提示第10和15天ADMSCs顯著減小饋瘍面積。組織學研究提示ADMSCs增加饋瘍傷口上皮和血管再生,促進膠原蛋白分泌和抑制炎癥反應。移植的ADMSCs能夠在饋瘍病竈內成活,並分化成血管內皮細胞和平滑肌細胞。ADMSCs顯著提高促傷口癒合相關基因表達水準。此外,ADMSCs啟動COX-2-PGE2和Erk1/2-MAPK信號通路。第10天,和對照組相比,引哚美辛/ADMSCs組潰瘍病竈內細胞增殖和血管再生顯著降低、饋瘍癒合延遲。本研究提示脂ADMSCs促進胃饋瘍癒合;非甾體抗炎藥顯著減弱ADMSCs的促胃饋瘍癒合作用。 / 結論:與內鏡夾閉相比,Eagle Claw VIII內鏡縫合胃創口有可行性。ADMSCs促進胃穿孔和饋瘍癒合,且依賴於前列腺素E2;引哚美辛抑制前列腺素E2合成從而抑制ADMSCs促胃組織癒合之效能。本研究提示聯合使用內鏡縫合器和ADMSCs是促進胃傷口癒合的潛在有效方法。 / Thesis Ph.D. Chinese University of Hong Kong 2014. / Includes bibliographical references (leaves 152-162). / Abstracts also in Chinese. / Title from PDF title page (viewed on 11, October, 2016). / Liu, Liu. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.
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Exploring the Role of Hypoxia-related Parameters in the Vascularization of Modular TissuesLam, Gabrielle 29 November 2013 (has links)
Modular tissue engineering involves assembling tissue constructs with integral vasculature from units containing adipose-derived mesenchymal stromal cells (adMSCs) and endothelial cells. Here, the effects of implant volume and adMSC density on the vascularization of modular tissues were explored. Both parameters affected the contributions of host- and graft-derived vessels, without affecting total vessel density. Increasing implant volume from 0.01 to 0.10 mL increased HIF1α expression and graft-derived vessel density, suggesting a role of hypoxia in graft-derived vessel formation. However, increasing adMSC density within small-volume implants did not increase HIF1α expression. Vascularization of small-volume implants of high (4.3•10^6 cells/mL) and low (1.0•10^6 cells/mL) adMSC densities was dominated by host vessel ingrowth at day 7. By increasing adMSC density, a high proportion of host-derived vessels was maintained to day 14, presumably via paracrine effects. Further dissection of the role of hypoxia in modular tissue engineering remains a promising avenue to pursue.
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Exploring the Role of Hypoxia-related Parameters in the Vascularization of Modular TissuesLam, Gabrielle 29 November 2013 (has links)
Modular tissue engineering involves assembling tissue constructs with integral vasculature from units containing adipose-derived mesenchymal stromal cells (adMSCs) and endothelial cells. Here, the effects of implant volume and adMSC density on the vascularization of modular tissues were explored. Both parameters affected the contributions of host- and graft-derived vessels, without affecting total vessel density. Increasing implant volume from 0.01 to 0.10 mL increased HIF1α expression and graft-derived vessel density, suggesting a role of hypoxia in graft-derived vessel formation. However, increasing adMSC density within small-volume implants did not increase HIF1α expression. Vascularization of small-volume implants of high (4.3•10^6 cells/mL) and low (1.0•10^6 cells/mL) adMSC densities was dominated by host vessel ingrowth at day 7. By increasing adMSC density, a high proportion of host-derived vessels was maintained to day 14, presumably via paracrine effects. Further dissection of the role of hypoxia in modular tissue engineering remains a promising avenue to pursue.
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Bone Marrow Microenvironment in Acute Myleoid LeukemiaChandran, Priya 09 July 2013 (has links)
Acute myeloid leukemia (AML) often remains refractory to current chemotherapy and transplantation approaches despite many advances in our understanding of mechanisms in leukemogenesis. The bone marrow “niche” or microenvironment, however, may be permissive to leukemia development and studying interactions between the microenvironment and leukemia cells may provide new insight for therapeutic advances. Mesenchymal stem cells (MSCs) are central to the development and maintenance of the bone marrow niche and have been shown to have important functional alterations derived from patients with different hematological disorders. The extent to which MSCs derived from AML patients are altered remains unclear. The aim of this study was to detect changes occurring in MSCs obtained from human bone marrow in patients with AML by comparing their function and gene expression pattern with normal age-matched controls.
MSCs expanded from patients diagnosed with acute leukemia were observed to have heterogeneous morphological characteristics compared to the healthy controls. Immunohistochemistry and flow data confirmed the typical cell surface immunophenotype of CD90+ CD105+ CD73+ CD34- CD45-, although MSCs from two patients with AML revealed reduced surface expression of CD105 and CD90 antigens respectively. Differentiation assays demonstrated the potential of MSCs from AML patients and healthy donors to differentiate into bone, fat and cartilage. However, the ability of MSCs from AML samples to support hematopoietic function of CD34+ progenitors was found to be impaired while the key hematopoietic genes were found to be differentially expressed on AML-MSCs compared to nMSCs.
These studies indicate that there exist differences in the biologic profile of MSCs from AML patients compared to MSCs derived from healthy donors. The results described in the thesis provide a formulation for additional studies that may allow us to identify new targets for improved treatment of AML.
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Investigation of Inhaled Nitric Oxide and Mesenchymal Stromal Cells as Novel Therapeutic Strategies to Improve Clinical Outcome in Experimental Severe InfluenzaDarwish, Ilyse 21 November 2012 (has links)
Severe influenza, recognized as a clinical syndrome characterized by hyper-induction of pro-inflammatory cytokine production, results in approximately 250–500 thousand deaths annually worldwide. Current influenza research is focused on therapeutics to target the influenza virus or modulate influenza virus-induced inflammation as potential treatment options to improve clinical outcome in experimental influenza A (H1N1) virus infection. The goals of this work were: (1) to evaluate the utility of inhaled nitric oxide (iNO) for decreasing influenza virus production in the lungs, and (2) investigate the use of mesenchymal stromal (stem) cells (MSCs) for mitigating deleterious host responses to influenza infection. Here, we report that MSCs and iNO, administered alone either prophylactically or post-influenza virus infection, fail to modulate host inflammation, fail to improve acute lung injury, fail to dampen lung viral load, and fail to improve survival of infected mice.
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Investigation of Inhaled Nitric Oxide and Mesenchymal Stromal Cells as Novel Therapeutic Strategies to Improve Clinical Outcome in Experimental Severe InfluenzaDarwish, Ilyse 21 November 2012 (has links)
Severe influenza, recognized as a clinical syndrome characterized by hyper-induction of pro-inflammatory cytokine production, results in approximately 250–500 thousand deaths annually worldwide. Current influenza research is focused on therapeutics to target the influenza virus or modulate influenza virus-induced inflammation as potential treatment options to improve clinical outcome in experimental influenza A (H1N1) virus infection. The goals of this work were: (1) to evaluate the utility of inhaled nitric oxide (iNO) for decreasing influenza virus production in the lungs, and (2) investigate the use of mesenchymal stromal (stem) cells (MSCs) for mitigating deleterious host responses to influenza infection. Here, we report that MSCs and iNO, administered alone either prophylactically or post-influenza virus infection, fail to modulate host inflammation, fail to improve acute lung injury, fail to dampen lung viral load, and fail to improve survival of infected mice.
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Estudo dos mecanismos envolvidos no processo de diferenciação em linhagem osteogênica de células-tronco mesenquimais da medula óssea de ratos Wistar e ratos espontaneamente hipertensos (SHR)Barros, Thamine Landim de [UNESP] 07 March 2014 (has links) (PDF)
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000799295.pdf: 1434919 bytes, checksum: 36275b6266ae49abeed7a7bba084d604 (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Células-tronco mesenquimais (CTMs) obtidas a partir da medula óssea são capazes de se diferenciarem, sobretudo, em condrócitos, adipócitos e osteoblastos. Durante a osteogênese in vitro, alguns parâmetros são utilizados para caracterizar este processo, tais como atividade da fosfatase alcalina (FAL), mineralização e expressão de proteínas associadas à osteoblastos. Ratos espontaneamente hipertensos (SHR) são um modelo animal de hipertensão essencial humana e desenvolvem hipertensão após 4 semanas de idade. Esta linhagem apresenta alterações significativas no metabolismo ósseo. O objetivo do presente estudo foi investigar se, o genótipo hipertensivo poderia interferir na diferenciação osteoblástica das CTMs de ratos SHR e qual mecanismo está alterado quando comparadas com a linhagem progenitora, ratos Wistar. Para isso, nós obtivemos CTMs da medula óssea de ratos Wistar e SHR com 4 semanas de idade, sem a hipertensão estabelecida, afim de avaliar somente o possível efeito do genótipo hipertensivo na diferenciação osteogênica in vitro. Nós induzimos, ou não, a diferenciação osteogênica in vitro por meio da utilização dos indutores osteogênicos: ácido ascórbico, ?-glicerofosfato e dexametasona. Os resultados demonstraram que, CTMs indiferenciadas de SHR (SHRC) demonstraram taxa de proliferação aumentada em comparação a CTMs, na mesma condição, de Wistar (WC), e após a indução da osteogênica, a taxa de proliferação apresentou uma diminuição acentuada no grupo SHR (SHRMO) do que no grupo Wistar na mesma condição (WMO). Embora não fora observada diferença significativa na atividade da FAL entre SHRMO e WOM no 7° dia, a mineralização e a diferenciação osteoblástica foram menores no grupo SHRMO no mesmo período experimental. Os fatores de transcrição Osterix e ?-catenina parecem estar envolvidos na diferenciação reduzida no grupo SHRMO... / Mesenchymal stem cells (MSCs) from bone marrow are able to differentiate mainly into chondrocytes, adipocytes and osteoblasts. During in vitro osteogenesis, some parameters are used to characterize this process, such as the activity of alkaline phosphatase (ALP), mineralization and osteoblast-associated proteins expression. Spontaneously hypertensive rats (SHR) is an animal model of human essential hypertension. This animals developing hypertension after 4 weeks of age. This strain shows significant changes in bone metabolism. The aim of this study was to investigate whether the hypertensive genotype could influence the osteoblastic differentiation of MSCs from SHR and which mechanism are altered when compared to the parental strain, Wistar rats. For that, we have obtained bone marrow MSCs from Wistar and SHR rats at 4 weeks of age, without hypertension established in order to evaluate only the possible effect of hypertensive genotype on osteogenic differentiation in vitro. We induced or non-osteogenic differentiation in vitro using osteogenic inducers: ascorbic acid, dexamethasone and ?-glycerophosphate. The results demonstrate that undifferentiated MSCs SHR (SHRC) showed increased proliferation rate compared to MSCs, in the same condition Wistar (WC) and after osteogenic induction, proliferation rate showed a marked decrease in SHR (SHRMO) than in Wistar group in the same condition (WMO). Although it was not observed significant difference in ALP activity between WMO and SHRMO on day 7, mineralization and osteoblast differentiation were lower on group SHRMO in the same experimental period. The transcription factors Osterix and ?-catenin appear to be involved in reduced differentiation in SHRMO group because they showed lower expression in this experimental group. Furthermore, the decreased... / FAPESP: 12/01924-9 / FAPESP: 11/06070-5 / FAPESP: 11/19458-1
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Mesenchymal stromal cell migration is regulated by fibronectin through integrin-mediated activation of PDGFR-βVeevers, Jennifer January 2010 (has links)
Human adult mesenchymal stem cells (MSCs) derived from bone marrow have the capacity to self-renew and to differentiate into a variety of cells and tissues. They can leave their niche to migrate to remote tissues where they play a critical role in angiogenesis, wound repair and tissue regeneration. A major goal in adult stem cell research is to define how MSC fate is controlled by the pericellular extracellular matrix (ECM) and soluble factors that largely constitute their tissue-specific niches. Defining crucial regulatory signals that control the fate and function of MSCs in vitro will contribute to the development of therapeutic strategies to improve tissue regeneration. The objective of this study was to investigate the molecular relationships between cell-ECM integrin receptors and platelet-derived growth factor receptor (PDGFR) tyrosine kinases, which are crucial in modulating MSC expansion, recruitment, and differentiation towards a number of different cell lineages. This study reports that ECM-directed cross-talk between PDGFR-β and alpha5β1 integrin controls the migration of MSCs. Cell adhesion to fibronectin induced integrin alpha5β1-dependent phosphorylation of PDGFR-β in the absence of growth factor stimulation. Phosphorylated PDGFR-β co-immunoprecipitated with integrin alpha5 and co-localised with alpha5β1 in a transient tidemark of focal adhesions. Adhesion to fibronectin also strongly potentiated platelet-derived growth factor (PDGF)-BB-stimulated PDGFR-β phosphorylation, in an alpha5β1-dependent manner. PDGFR-β-activated phosphatidylinositol 3 ́-kinase (PI3-kinase) and Akt activity, actin reorganisation and cell migration were all regulated by fibronectin engagement of alpha5β1 integrin. This synergistic relationship between integrin alpha5β1 and PDGFR-β is a fundamental determinant of mesenchymal cell migration. Thus, fibronectin-rich matrices can prime PDGFR-β to recruit mesenchymal cells at sites of tissue remodelling.
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Culture of human umbilical cord mesenchymal stromal cells in a three-dimensional human platelet lysate gelJirakittisonthon, Thitikan January 1900 (has links)
Master of Science in Biomedical Sciences / Department of Anatomy and Physiology / Mark L. Weiss / The traditional cell culture method after isolation from the body involves growing cells in 2 dimensions on plastic culture plate. However, the natural structure and physiology is 3 dimensions. To mimic in vivo environment, there has an increasing interest to find the way to maintain physiological properties. Here, we describe culturing human umbilical cord mesenchymal stromal cells (HUC-MSCs_in 3D setting using human platelet lysate gel. This gel is a fibrin-based structure like a blood clot. The preparation step of human platelet lysate (HPL) is by freeze- thaw cycles in order to release factors important for cells to grow and expand. Using of HPL to substitute for fetal bovine serum reduces potential cross contamination between species and xenogenicity. To maintain HPL media as a liquid, we add the anticoagulant heparin. Without adding anticoagulant, the gel will form. The aim of this study is to retrieve HUC-MSCs from HPL gel using Nattokinase, to characterize HUC-MSCs following the International Society for Cell Therapy’s MSC criteria, and to test a 3D invasion model with HPL-gel based structure. The result shows that using 1.75% Nattokinase at 60 minutes can recover the cells without reducing cell number and viability. After Nattokinase treatment, cells are able to attach to plastic and to increase in number. Moreover, they are able to differentiate into fat, bone, and cartilage no different from cells grown in 2D culture. However, to test surface markers by flow cytometry, all MSC markers are positive except CD 105. They are also positive of cell surface markers that should be negative. When seeded back to 2D culture for an additional passage, the MSCs meet ISCT criteria the same as control.
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Estudo dos mecanismos envolvidos no processo de diferenciação em linhagem osteogênica de células-tronco mesenquimais da medula óssea de ratos Wistar e ratos espontaneamente hipertensos (SHR) /Barros, Thamine Landim de. January 2014 (has links)
Orientador: Sandra Helena Penha de Oliveira / Banca: Gustavo Pompermaier Garlet / Banca: Willian Fernando Zambuzzi / Resumo: Células-tronco mesenquimais (CTMs) obtidas a partir da medula óssea são capazes de se diferenciarem, sobretudo, em condrócitos, adipócitos e osteoblastos. Durante a osteogênese in vitro, alguns parâmetros são utilizados para caracterizar este processo, tais como atividade da fosfatase alcalina (FAL), mineralização e expressão de proteínas associadas à osteoblastos. Ratos espontaneamente hipertensos (SHR) são um modelo animal de hipertensão essencial humana e desenvolvem hipertensão após 4 semanas de idade. Esta linhagem apresenta alterações significativas no metabolismo ósseo. O objetivo do presente estudo foi investigar se, o genótipo hipertensivo poderia interferir na diferenciação osteoblástica das CTMs de ratos SHR e qual mecanismo está alterado quando comparadas com a linhagem progenitora, ratos Wistar. Para isso, nós obtivemos CTMs da medula óssea de ratos Wistar e SHR com 4 semanas de idade, sem a hipertensão estabelecida, afim de avaliar somente o possível efeito do genótipo hipertensivo na diferenciação osteogênica in vitro. Nós induzimos, ou não, a diferenciação osteogênica in vitro por meio da utilização dos indutores osteogênicos: ácido ascórbico, β-glicerofosfato e dexametasona. Os resultados demonstraram que, CTMs indiferenciadas de SHR (SHRC) demonstraram taxa de proliferação aumentada em comparação a CTMs, na mesma condição, de Wistar (WC), e após a indução da osteogênica, a taxa de proliferação apresentou uma diminuição acentuada no grupo SHR (SHRMO) do que no grupo Wistar na mesma condição (WMO). Embora não fora observada diferença significativa na atividade da FAL entre SHRMO e WOM no 7° dia, a mineralização e a diferenciação osteoblástica foram menores no grupo SHRMO no mesmo período experimental. Os fatores de transcrição Osterix e β-catenina parecem estar envolvidos na diferenciação reduzida no grupo SHRMO... / Abstract: Mesenchymal stem cells (MSCs) from bone marrow are able to differentiate mainly into chondrocytes, adipocytes and osteoblasts. During in vitro osteogenesis, some parameters are used to characterize this process, such as the activity of alkaline phosphatase (ALP), mineralization and osteoblast-associated proteins expression. Spontaneously hypertensive rats (SHR) is an animal model of human essential hypertension. This animals developing hypertension after 4 weeks of age. This strain shows significant changes in bone metabolism. The aim of this study was to investigate whether the hypertensive genotype could influence the osteoblastic differentiation of MSCs from SHR and which mechanism are altered when compared to the parental strain, Wistar rats. For that, we have obtained bone marrow MSCs from Wistar and SHR rats at 4 weeks of age, without hypertension established in order to evaluate only the possible effect of hypertensive genotype on osteogenic differentiation in vitro. We induced or non-osteogenic differentiation in vitro using osteogenic inducers: ascorbic acid, dexamethasone and β-glycerophosphate. The results demonstrate that undifferentiated MSCs SHR (SHRC) showed increased proliferation rate compared to MSCs, in the same condition Wistar (WC) and after osteogenic induction, proliferation rate showed a marked decrease in SHR (SHRMO) than in Wistar group in the same condition (WMO). Although it was not observed significant difference in ALP activity between WMO and SHRMO on day 7, mineralization and osteoblast differentiation were lower on group SHRMO in the same experimental period. The transcription factors Osterix and β-catenin appear to be involved in reduced differentiation in SHRMO group because they showed lower expression in this experimental group. Furthermore, the decreased... / Mestre
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