Heterogenecity of stromal cell precursers isolated from rat bone marrow

Bone marrow stroma contains mesenchymal stem cells (MSC) which are progenitor cells, at least for tissues arising from mesechyma. The study of MSC biology yields controversial data. Therefore further experiments are needed to characterize these cells. The aim of our research was to compare primary cultures and subcultures of stromal precursor cells isolated from rat bone marrow. Long-term cultures of these cells isolated from 5 animals have been obtained. Morphological, immunophenotypic, and functional (capacity to osteogenic differentiation) characteristics of the cells have been investigated. We show that the cell morphology in the cultures is highly heterogenic. Morphological cell types are described. Heterogeneity of stromal cells declines on late passages. Cell cultures isolated from different animals have the same immunophenotypic markers (CD90, CD44, CD54, CD106, CD45, CD11b) but different morphological characteristics and a different capacity to osteogenic differentiation during long-term cultivation. The data show that more specific markers and functional tests should be applied to identify MSC.     

Mesenchymal stem cells induce dermal fibroblast responses to injury

Although bone marrow-derived mesenchymal stem cells have been shown to promote repair when applied to cutaneous wounds, the mechanism for this response remains to be determined. The aim of this study was to determine the effects of paracrine signaling from mesenchymal stem cells on dermal fibroblast responses to injury including proliferation, migration and expression of genes important in wound repair. Dermal fibroblasts were co-cultured with bone marrow-derived mesenchymal stem cells grown in inserts, which allowed for paracrine interactions without direct cell contact. In this co-culture model, bone marrow-derived mesenchymal stem cells regulate dermal fibroblast proliferation, migration and gene expression. When co-cultured with mesenchymal stem cells, dermal fibroblasts show increased proliferation and accelerated migration in a scratch assay. A chemotaxis assay also demonstrated that dermal fibroblasts migrate towards bone marrow-derived mesenchymal stem cells. A PCR array was used to analyze the effect of mesenchymal stem cells on dermal fibroblast gene expression. In response to mesenchymal stem cells, dermal fibroblasts up-regulate integrin alpha 7 expression and down-regulate expression of ICAM1, VCAM1 and MMP11. These observations suggest that mesenchymal stem cells may provide an important early signal for dermal fibroblast responses to cutaneous injury.     

Prion diseases and adult neurogenesis: How do prions counteract the brain's endogenous repair machinery?

Scientific advances in stem cell biology and adult neurogenesis have raised the hope that neurodegenerative disorders could benefit from stem cell-based therapy.Adult neurogenesis might be part of the physiological regenerative process; however, it might become impaired by the disease''s mechanism and therefore contribute to neurodegeneration. In prion disorders this endogenous repair system has rarely been studied. Whether adult neurogenesis plays a role or not in brain repair or in the propagation of prion pathology remains unclear. We have recently investigated the status of adult neural stem cells isolated from prion-infected mice. We were able to show that neural stem cells accumulate and replicate prions thus resulting in an alteration of their neuronal destiny. We also reproduced these results in adult neural stem cells, which were infected in vitro. The fact that endogenous adult neurogenesis could be altered by the accumulation of misfolded prion protein represents another great challenge. Inhibiting prion propagation in these cells would thus help the endogenous neurogenesis to compensate for the injured neuronal system. Moreover, understanding the endogenous modulation of the neurogenesis system would help develop effective neural stem cell-based therapies.     

骨髓间充质干细胞移植对心衰大鼠心肌结构和功能的影响

研究骨髓间充质干细胞(MSC)移植对心力衰竭(简称心衰)大鼠心肌结构和功能的影响以及在病损心肌体内分化为心肌细胞的情况。将96只Wistar大鼠,用阿霉素成功诱导了54只心衰模型,随机分成3组,移植组为左室前壁注射MSC,对照组注射培养基,心衰组不给予任何干预措施。由彩色超声心动图(TTE)监测左室心功能参数。8周检测完成后取出心脏标本,做冰冻切片脏染色观察病损心肌结构的变化及免疫荧光检查植入MSC心肌肌球蛋白重链(MHC)及心肌特有的连接蛋白(Cx43)表达情况。结果表明植入的MSC存活并表达了MHC及Cx43,其周围宿主心肌细胞肿胀明显减轻。在移植MSC2周后,心功能开始改善,至8周时,心功改善能更明显。由此得出结论：MSC在病损心肌体内不仅能存活、分化为心肌细胞,使病损心肌组织病变减轻。而且可显著改善心衰大鼠的心功能。     

Development of novel monoclonal antibodies that define differentiation stages of human stromal (mesenchymal) stem cells

Human mesenchymal stem cells (hMSC) are currently being introduced for cell therapy, yet, antibodies specific for native and differentiated MSCs are required for their identification prior to clinical use. Herein, high quality antibodies against MSC surface proteins were developed by immunizing mice with hMSC, and by using a panel of subsequent screening methods. Flow cytometry analysis revealed that 83.5, 1.1, and 8.5% of primary cultures of hMSC were double positive for STRO-1 and either of DJ 3, 9, and 18, respectively. However, none of the three DJ antibodies allowed enrichment of clonogenic hMSC from BMMNCs as single reagents. Using mass-spectrometric analysis, we identified the antigen recognised by DJ3 as CD44, whereas DJ9 and DJ18 recognized HLA-DRB1 and Collagen VI, respectively. The identified proteins were highly expressed throughout in vitro osteogenic- and adipogenic differentiation. Interestingly, undifferentiated cells revealed a sole cytoplasmic distribution pattern of Collagen VI, which however changed to an extracellular matrix appearance upon osteogenic- and adipogenic differentiation. In relation to this, we found that STRO-1^+/−/Collagen VI⁻ sorted hMSC contained fewer differentiated alkaline phosphatase⁺ cells compared to STRO-1^+/−/Collagen VI⁺ hMSC, suggesting that Collagen VI on the cell membrane exclusively defines differentiated MSCs. In conclusion, we have generated a panel of high quality antibodies to be used for characterization of MSCs, and in addition our results may suggest that the DJ18 generated antibody against Collagen VI can be used for negative selection of cultured undifferentiated MSCs.     

Mechanisms of bone anabolism regulated by statins

Osteoporosis is a common disease in the elderly population. The progress of this disease results in the reduction of bone mass and can increase the incidence of fractures. Drugs presently used clinically can block the aggravation of this disease. However, these drugs cannot increase the bone mass and may result in certain side effects. Statins, also known as HMG-CoA (3-hydroxy-3-methylglutaryl-CoA) reductase inhibitors, have been widely prescribed for CVD (cardiovascular disease) for decades. Nonetheless, several studies have demonstrated that statins exert bone anabolic effect and may be helpful for the treatment of osteoporosis. Several experiments have analysed the mechanisms of bone anabolism regulated by statins. In the present paper, we review the mechanisms of promoting osteogenesis, suppressing osteoblast apoptosis and inhibiting osteoclastogenesis.     

调控骨骼形成的信号通路

骨骼形成后会处于不断的分解与重建中.通过骨骼形成与骨骼吸收之间的动态平衡来维持骨量.如果二者间的平衡被打破,骨吸收大于骨形成时,骨量会减少,骨骼微环境随之发生改变,脆性增加,进而引发骨质疏松、骨折等疾病.其中,骨骼形成是成骨细胞的重要功能.成骨细胞由间充质干细胞(mesenchymal stem cells,MSCs)...     

Do microRNAs regulate bone marrow stem cell niche physiology?

The adult bone marrow, situated within the bone cavity, comprises three distinct stem cell populations: hematopoietic stem cells (HSCs), mesenchymal stromal/stem cells (MSCs) and endothelial progenitor/stem cells (EPCs). HSCs are a well-characterized population of self-renewing cells that give rise to all blood cells. The definition of MSCs is more complex due to the limited understanding of MSC properties. In general, MSCs are considered multipotent stromal cells that are able to differentiate into various cell types, including osteoblasts, chondrocytes and adipocytes. Compared to HSCs and MSCs, EPCs are a newly discovered population of stem/progenitor cells with the capacity to differentiate into endothelial cells, the cells forming the inner lining of a blood vessel.     

MiR-31促进人牙髓间充质干细胞增殖和迁移

牙髓炎和根尖周炎是口腔科目前较为常见的2种疾病,现有的治疗方案主要包括根管治疗和牙髓血运重建,能有效控制住炎症进而保存患牙,然而同时也会导致牙髓组织的永久失活,发生结构故障和继发感染。近年来,结合干细胞和生物材料等的组织工程技术使牙髓再生的研究逐渐进入人们的视野,其中从恒牙或乳牙中分离的牙髓间充质干细胞(dental pulp stem cells, DPSCs),因其多向分化和高增殖等特性已成为牙本质或者髓样组织再生的重要的干细胞来源。但是由于干细胞不能高效募集到损伤区域,影响受损区的活细胞数量和存活时间,显著降低了其应用疗效,因此,需要提高牙髓干细胞的迁移和增殖能力,本研究旨在探讨微核糖核酸31(microRNA-31, miR-31)能否有效提高DPSCs的增殖迁移能力。通过组织块酶消化法从牙髓组织中成功分离培养了DPSCs,比较了分别取自健康牙与炎症牙的牙髓组织和DPSCs中 miR-31水平的差异,利用实时荧光定量PCR(RT-qPCR)技术检测的结果显示,与健康牙比较,炎症牙来源的牙髓组织和DPSCs中 miR-31 表达水平明显降低(P<0.05)。干扰和过表达DPSCs中的miR-31表达,具体分为NC组、miR-31 agomir(过表达)组和miR-31 antagomir(抑制剂)3个组,RTq-PCR 结果显示,其转染成功(P<0.001)。CCK-8、细胞划痕以及Transwell迁移结果表明,与对照组相比,过表达miR-31成功提高了牙髓干细胞的增殖和迁移能力(P<0.05),且进一步通过 Western印迹结果发现,过表达miR-31后增殖关键蛋白质增殖抗原(Ki67)、增殖细胞核抗原(proliferating cell nuclear antigen, PCNA)以及迁移关键蛋白质CXC趋化因子受体4型(CXC chemokine receptor type 4, CXCR4)、基质金属蛋白酶2(matrix metalloproteinase2, MMP2)的蛋白质水平均显著升高(P<0.05)。本研究表明,miR-31能有效提高DPSCs的增殖迁移能力,以期为DPSCs更好应用于再生医学提供了有力的理论支持。