糖皮质激素促间充质干细胞的成骨效应

背景：糖皮质激素是有效的免疫抑制和抗炎症药物而广泛应用于临床,然而长期服用后也会引起一些不良反应如骨质疏松症。糖皮质激素通过促进成骨细胞与骨细胞的凋亡,同时抑制间充质干细胞向成骨细胞分化,导致成骨细胞数量减少;然而在一定的条件下,糖皮质激素又能够促进间充质干细胞表达成骨细胞标志基因诱导其向成骨方向分化。 目的：对以地塞米松为代表的糖皮质激素在间充质干细胞成骨分化中发挥的作用以及可能的分子机制进行综述。 方法：由第一作者检索PubMed数据库1978至2012年有关地塞米松和间充质干细胞成骨分化的文献,英文关键词“Mesenchymal stem cell,dexamethasone,osteogenesis,osteoporosis,Runx2,BMP,Noggin,GILZ,glucocorticoid receptor”以不同的组合方式查找,排除重复性的研究,最终保留55篇进行归纳综述。 结果与结论：地塞米松能够调节Runx2,Noggin以及亮氨酸拉链蛋白等基因调控间充质干细胞成骨分化。当糖皮质激素作用于间充质干细胞时,可能由糖皮质激素受体介导其生理作用,也可能在糖皮质激素与其受体结合之前就受到11β-羟甾类脱氢酶的调节。另外,生理剂量(10^-8 mol/L)的地塞米松能促进间充质干细胞成骨分化,药理剂量(≥10^-7 mol/L)有抑制作用。     

Cotton wool-like nanocomposite biomaterials prepared by electrospinning: in vitro bioactivity and osteogenic differentiation of human mesenchymal stem cells

The present study evaluates the in vitro biomedical performance of an electrospun, flexible, and cotton wool-like poly(lactide-co-glycolide) (PLGA)/amorphous tricalcium phosphate (ATCP) nanocomposite. Experiments on in vitro biomineralization, applicability in model defects and a cell culture study with human mesenchymal stem cells (hMSC) allowed assessing the application of the material for potential use as a bone graft. Scaffolds with different flame made ATCP nanoparticle loadings were prepared by electrospinning of a PLGA-based composite. Immersion in simulated body fluid showed significant deposition of a hydroxyapatite layer only on the surface of ATCP doped PLGA (up to 175% mass gain within 15 days for PLGA/ATCP 60:40). Proliferation and osteogenic differentiation of hMSC on different nanocomposites were assessed by incubating cells in osteogenic medium for 4 weeks. Proper adhesion and an unaffected morphology of the cells were observed by confocal laser scanning microscopy for all samples. Fluorometric quantification of dsDNA and analysis of ALP activity revealed no significant difference between the tested scaffolds and excluded any acute cytotoxic effects of the nanoparticles. The osteocalcin content for all scaffolds was 0.12-0.19 ng/ng DNA confirming osteogenic differentiation of human mesenchymal stem cells on these flexible bone implants.     

低氧影响干细胞成骨分化的研究进展

氧稳态是机体或细胞正常功能所必需的, 低氧是人正常发育的一个重要的生理因素,并参与许多疾病的发生、发展.在骨发育过程中, 组织和细胞缺氧是经常存在的.体内生长板软骨增殖区氧浓度仅为 2%-5%, 肥大区氧浓度为 0.15%-1%[1].机体损伤后,由于损伤部位血流中断或血肿形成,造成局部氧分压降低.在骨折血流中断处,中央部位氧分压甚至降低到0%-2%[2,3].骨折修复需要机体在骨折部位重建血运,以便吸收损伤组织和转运营养物质.     

体外冲击波通过三磷酸腺苷诱导人骨髓间充质干细胞向成骨细胞分化

目的 研究体外冲击波是否通过三磷酸腺苷（ATP）激活P2X7受体,诱导人骨髓间充质干细胞(human mesenchymal stem cells,hMSCs)向成骨细胞分化。方法 培养hMSCs细胞,检测冲击波是否引起其向外释放ATP;通过检测碱性磷酸酶(ALP)活性、骨钙素表达和钙结节形成,判断骨化形成和钙质沉积;用实时定量PCR检测P2X7受体的mRNA表达;用ATP水解酶、P2X7受体的siRNA以及 P2受体的抑制剂评估ATP释放和P2X7受体在冲击波诱导hMSCs成骨分化中的作用。结果 冲击波可引起细胞内ATP向外释放,冲击波和细胞外ATP能够诱导hMSCs向成骨分化,采用ATP水解酶、P2X7受体的siRNA和抑制剂能够抑制冲击波引起的hMSCs成骨化作用。结论 冲击波通过引起细胞内ATP向外释放,激活P2X7受体传导信号通路,促进hMSCs向成骨细胞分化。本研究结果为冲击波促进骨折愈合和治疗骨不连疗法提供了理论依据。     

miR-195对人骨髓间充质干细胞成骨分化的影响

BACKGROUND:Previous studies have found that the expression level of miR-195 in differentiated human bone marrow mesenchymal stem cells (hBMMSCs) is significantly higher than that in undifferentiated hBMMSCs. However, miR-195 effect during this differentiation process and possible mechanism remain unclear. OBJECTIVE:To explore the effect of miR-195 on osteogenic differentiation of hBMMSCs and possible mechanism. METHODS:hBMMSCs were isolated and cultured in vitro. Alkaline phosphatase activity, Runx2, osteopontin and SMAD7 protein expression and miR-195 expression level during osteogenic differentiation of hBMMSCs were determined by alkaline phosphatase kit, western blot and real-time PCR, respectively. miR-195-downexpressed hBMMSCs were constructed by lipofection transfection, and were used to investigate the effect of miR-195 was on osteogenic differentiation of hBMMSCs. Dual luciferase reporter assay was used to identify whether the 3’UTR of SMAD7 mRNA was a binding target of miR-195. In addition, we transfected hBMMSCs with SMAD7 cDNA (pcDNA-SMAD7), and investigated the effect of SMAD7 on osteogenic differentiation of hBMMSCs. RESULTS AND CONCLUSION:The isolated and cultured hBMMSCs had good osteogenic differentiation ability in vitro. Expression level of miR-195 was increased with the increasing of induction time, and the expression level of SMAD7 was reversed. miR-195 promoted osteogenic differentiation of hBMMSCs. Luciferase assay confirmed that miR-195 targeted SMAD7 directly, and overexpression of SMAD7 inhibited the osteogenic differentiation of hBMMSCs. Taken together, miR-195 promotes osteogenic differentiation of hBMMSCs by targeting SMAD7.     

辛伐他汀可刺激骨髓间充质干细胞的成骨分化

背景:辛伐他汀可显著刺激骨髓间充质干细胞成骨分化,但机制不明.近年研究证实p38 MAPK信号通路参与调控骨髓间充质干细胞向成骨细胞分化的过程.目的:观察p38丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路在辛伐他汀体外干预刺激骨髓间充质干细胞成骨分化中的作用....     

纤维蛋白凝胶促进大鼠间充质干细胞的成骨分化

背景：纤维蛋白是一种天然的可生物降解、组织相容性好的高分子材料,是一种能促进细胞和外源性生长因子释放的载体,其中血纤维蛋白稳定因子ⅩⅢ已证明有利于未分化的间充质干细胞在高度交联的凝胶支架内迁移,并且促进这些细胞的增殖与分化能力。 目的：观察大鼠间充质干细胞在纤维蛋白凝胶内的行为。 方法：无菌条件下分离大鼠胎肢细胞获得间充质干细胞,取第3代细胞分别接种于0,5,10,20 g/L纤维蛋白凝胶内,用倒置相差显微镜和激光扫描共聚焦显微镜分析细胞在凝胶内的形态学变化; 酶标仪和Von Kossa染色分析碱性磷酸酶活性和钙盐沉积。 结果与结论：5 g/L低浓度纤维蛋白凝胶有利于细胞形态的发生,20 g/L高浓度凝胶有利于细胞的成骨分化。20 g/L纤维蛋白凝胶碱性磷酸酶活性高于对照组,10和20 g/L浓度纤维蛋白凝胶矿化结节出现在21至28 d,而对照组无矿化结节出现。提示大鼠间充质干细胞的形态与成骨分化依赖于纤维蛋白凝胶浓度,提示纤维蛋白凝胶有助于间充质干细胞的成骨分化。中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程全文链接：     

Nitric oxide production during the osteogenic differentiation of human periodontal ligament mesenchymal stem cells

The critical tissues that require regeneration in the periodontium are of mesenchymal origin; therefore, the ability to identify, characterize and manipulate mesenchymal stem cells within the periodontium is of considerable clinical significance. In particular, recent findings suggest that periodontal ligament cells may possess many osteoblast-like properties. In the present study, periodontal ligament mesenchymal stem cells obtained from healthy volunteers were maintained in culture until confluence and then induced to osteogenic differentiation. Intracellular calcium ([Ca²⁺]_i) concentration and nitric oxide, important signalling molecules in the bone, were measured along with cell differentiation. Alkaline phosphatase activity was assayed and bone nodule-like structures were evaluated by means of morphological and histochemical analysis. Our results showed that the periodontal ligament mesenchymal stem cells underwent an in vitro osteogenic differentiation, resulting in the appearance of active osteoblast-like cells together with the formation of calcified deposits. Differentiating cells were also characterized by an increase of [Ca²⁺]_i and nitric oxide production. In conclusion, our data show a link between nitric oxide and the osteogenic differentiation of human periodontal ligament mesenchymal stem cells, thus suggesting that local reimplantation of expanded cells in conjugation with a nitric oxide donor could represent a promising method for treatment of periodontal defects.     

Continuous stimulation with differentiation factors is necessary to enhance osteogenic differentiation of human mesenchymal stem cells in-vitro

Bone defect treatment belongs to the most challenging fields in orthopedic surgery and requires the well-coordinated application of mesenchymal stem cells (MSC) and differentiation factors. MSC isolated from reaming material (RMSC) and iliac crest (BMSC) in combination with bone morphogenetic protein-7 (BMP-7) and insulin-like growth factor-1 (IGF-1) have been used. The short half-life of both factors limit their applications: a burst release of the factor can probably not induce sustainable differentiation. We stimulated MSC in osteogenic differentiation medium with three different concentrations of BMP-7 or IGF-1: Group A was stimulated continuously, group B for 24?h and group C remained without any stimulation. Osteogenic differentiation was measured after seven and 14?days by alizarin red staining and alkaline phosphatase (ALP) activity. Continuous stimulation led to higher levels of osteogenic differentiation than short-term stimulation. This could lead to a reconsideration of established application forms for differentiation factors, aiming to provide a more sustained release.     

力学因素对间充质干细胞成骨分化的影响

背景：随着间充质干细胞的广泛研究,间充质干细胞能够向成骨方向分化已得到充分证实。近年来,许多学者致力于研究力学因素对体外培养间充质干细胞增殖和定向诱导分化的影响。 目的：综述力学因素对间充质干细胞成骨分化产生的影响。 方法：应用计算机检索PubMed,ScienceDirect,ISI-SCIE,中国期刊全文数据库,中国期刊全文数据库_世纪期刊,中国博士学位论文全文数据库,中国优秀硕士学位论文全文数据库数据库中1999-01/2012-01关于力学因素对间充质干细胞成骨分化影响的文章,在标题和摘要中以“力学因素, 间充质干细胞,成骨分化”或“mechanical factors,Mesenchymal Stem Cell,Osteogenic Differentiation”为检索词进行检索。选择文章所述内容涉及力学因素对间充质干细胞中成骨分化的作用,并且选择近期发表或发表在权威杂志文章。初检索到文献104篇,最终入选30篇文献进行综述。 结果与结论：近年的各项实验都证明,力学因素对间充质干细胞的增殖和分化、形态、发育和功能起着重要的调节作用。而力学刺激的大小、方式以及作用时间对间充质干细胞定向分化的调节、功效作用不尽相同。应选择最佳的力学刺激促使间充质干细胞的成骨分化,以达到最佳的临床治疗效果。     

骨髓间充质干细胞成骨分化调控的研究进展

骨髓间充质干细胞具有较强的自我更新能力和多向分化潜能 ,其应用是目前国际上组织工程领域中重要的研究内容之一。近年来 ,许多实验室从分子、生化、物理等水平对其成骨分化调控进行了深入研究 ,取得了较大的进展     

胰岛素促进脐带间充质干细胞成骨分化的作用

BACKGROUND: How to effectively and rapidly induce the osteogenic differentiation of human umbilical cord mesenchymal stem cells is the focus of the current stem cell research. Increasing evidence has demonstrated some growth factors, such as bone morphogenetic protein-2, have important effects on the transdifferentiation of umbilical cord mesenchymal stem cells into osteoblasts in vitro. However, widespread use of growth factors is limited because of high cost. Insulin is widely used in the cell culture and induction, but there is no report about the effect of insulin on the osteogenic differentiation of human umbilical cord mesenchymal stem cells. OBJECTIVE: To observe the effect of insulin on osteogenic differentiation of human umbilical cord mesenchymal stem cells and to explore the feasibility of human umbilical cord mesenchymal stem cell transplantation in the treatment of diabetic delayed fracture healing. METHODS: The passage 3 human umbilical cord mesenchymal stem cells were inoculated in two flasks, denoted as experimental group and control group. The insulin (10-7 mmol/L) was added to the experimental group but not to the control group. The proliferative capacity of human umbilical cord mesenchymal stem cells was evaluated by cell count kit-8 and alkaline phosphatase activity. The osteogenic differentiation capacity of human umbilical cord mesenchymal stem cells was evaluated by measuring the protein and mRNA expressions of type I collagen as well as osteocalcin mRNA level. RESULTS AND CONCLUSION: After 1-2 weeks of induction, compared with the control group, insulin could significantly increase the number of human umbilical cord mesenchymal stem cells in the experimental group, the activity of alkaline phosphatase and expressions of type I collagen osteocalcin mRNA (P < 0.05). These data indicate that insulin can promote the proliferation and osteogenic differentiation of human umbilical cord mesenchymal stem cells.       

人骨髓间充质干细胞的体外培养、鉴定及成骨分化

背景：国内外对骨髓间充质干细胞的体外成骨诱导分化研究手段、测定指标均不够全面。 目的：建立并完善一整套人骨髓间充质干细胞的分离培养及鉴定方法,探讨其体外成骨分化能力。 方法：采用密度梯度离心法分离培养人骨髓间充质干细胞,流式细胞仪鉴定细胞表面表型。传至第3代时更换成骨诱导培养基进行成骨分化诱导。 结果与结论：人骨髓间充质干细胞生长旺盛,传代后增殖旺盛,第3代骨髓间充质干细胞表面表型CD44、CD73、CD90表达阳性,CD34表达阴性。诱导后的成骨细胞碱性磷酸酶活性增加,Gomori、Von kossa、茜素红染色均阳性。RT-PCR检测诱导后细胞有Ⅰ型胶原、碱性磷酸酶、骨钙素、骨唾液酸蛋白、骨桥蛋白及骨连接蛋白基因的表达,证明了人骨髓间充质干细胞成功向成骨方向分化。表明实验建立了一整套稳定、成熟的骨髓间充质干细胞分离、培养、扩增方案。     

Study of hydroxyapatite osteoinductivity with an osteogenic differentiation of mesenchymal stem cells

Osteoinductivity of hydroxyapatite (HA) was investigated using uncommitted pluripotent mouse stem cells, C3H10T1/2 in an in vitro differentiation assay. For comparative analysis, the cells were cultured on substrates made of osteoinductive HA, with biocompatible titanium and plastics as the negative control. HA exhibited the ability to induce expression of osteo-specific genes in C3H10T1/2, including alkaline phosphatase (ALP), type I collagen, and osteocalcin; compared with its insignificant up-regulation of the same genes in osteoblast-like cells, Saos-2. HA osteoinductivity exhibited in C3H10T1/2 was comparable to that of a bone morphogenetic protein (BMP) with reference to the up-regulation of osteo-specific genes except the core binding factor 1 (Cbfa1, Runx). This result implies a difference in osteogenic induction pathway initiated by HA and BMP. Using this mesenchymal stem cells (MSC) culture assay, osteoinductivity was also demonstrated to be present in the conditioned medium derived from MSC cultured on HA substrates. This conditioned medium exhibited excellent ability to up-regulate ALP in the absence of HA and BMP. The results suggest that the HA can interact with the cells and generate potent inductive substance released into the medium. Such substance in turn is able to induce uncommitted cells to differentiate into the osteolineage.     

中药诱导骨髓间充质干细胞的成骨分化

背景：某些中药可以诱导骨髓间充质干细胞的成骨分化。骨髓间质干细胞向成骨分化的潜能,与中药治疗骨质疏松症、骨折、骨坏死、骨缺损等骨相关疾病有着理论上的相通性。 目的：了解中药诱导骨髓间充质干细胞成骨分化的发展现状,为进一步的研究奠定基础。 方法：由第一作者检索2000-01/2010-06中国期刊全文数据库(CNKI)(http://www.cnki.net/)及Pubmed数据库(http://www.ncbi.nlm.nih.gov/PubMed)。中文检索词为“中药,骨髓间充质干细胞,成骨分化”。英文检索词为“chinese herb, mesenchymal stem cells, osteogenic differentiation”。文献检索语种限定为中文和英文,纳入中药单体、单味中药、中药复方等及其含药血清在体内或体外对人或动物骨髓间充质干细胞成骨分化作用的研究文献,排除重复研究。 结果与结论：共纳入32篇文献,有关骨髓间充质干细胞研究背景的文献2篇;有关单味中药的文献5篇,其中补肾药4篇,补气药1篇;有关中药复方的文献10篇,其中补肾方6篇,补肾活血方4篇;有关中药有效组分的文献15篇。补肾、补气及活血类中药可以诱导骨髓间充质干细胞向成骨细胞分化,但以补肾类中药为主,在一定程度上阐释了“肾主骨”理论的科学内涵,并为体外大量扩增骨髓间充质干细胞、促成骨分化及组织工程骨提供了更多的种子细胞来源。     

大鼠骨髓间充质干细胞成骨诱导分化的超微结构变化

背景：目前对骨髓间充质干细胞成骨诱导分化的超微结构观察的报道甚少。 目的：采用全骨髓贴壁法分离培养大鼠骨髓间充质干细胞,成骨诱导并染色鉴定,利用电镜观察诱导前后细胞超微结构变化特点。 方法：全骨髓贴壁法体外分离、培养、纯化大鼠骨髓间充质干细胞,流式细胞仪检测细胞表面标记物,成骨染色鉴定向成骨方向诱导分化,扫描电镜及透射电镜观察成骨诱导前后细胞超微结构变化。 结果与结论：培养的第3代骨髓间充质干细胞纯度高、活力强,成骨诱导后的碱性磷酸酶活性染色、钙化结节染色均呈阳性。扫描电镜及透射电镜观察显示,经向成骨细胞诱导分化后,细胞形态铺展,不规则,其线粒体、粗面内质网、空泡明显增多,表明细胞功能活跃。