间充质干细胞对辐射诱发胸腺瘤β-catenin和c-myc基因影响

目的 探讨骨髓间充质干细胞(MSCs)对辐射诱发胸腺瘤过程中β-catenin和c-myc mRNA表达影响,为阐明MSCs在肿瘤发生发展中作用提供理论依据。方法全骨髓贴壁培养C57BL/6小鼠MSCs,X射线照射C57BL/6小鼠建立胸腺淋巴瘤模型,将小鼠随机分成对照组、辐射组及MSCs组,每组6只;逆转录-聚合酶链反应技术检测各组小鼠胸腺β-catenin和c-myc mRNA表达。结果 辐射组小鼠胸腺瘤β-catenin mRNA表达量(0.92±0.23)明显高于对照组(0.64±0.21)(P<0.05),MSCs组β-catenin mRNA表达量(0.77±0.24)下降;与对照组(1.48±0.34)比较,辐射组c-myc mRNA表达量(1.61±0.37)上调,MSCs组c-myc mRNA表达量(1.18±0.34)明显低于辐射组(P<0.05)。结论 MSCs可通过抑制β-catenin和c-myc基因异常表达,抑制肿瘤的增殖。     

BMP2诱导骨骼肌成骨的研究进展

20世纪90年代以来,组织工程技术不断发展,在组织工程化骨领域取得了较大进展,然而,迄今为止,组织工程化骨修复缺损,离临床的要求还有较大距离。目前认为,主要问题是种子细胞在一定传代后发生衰老而不能继续增殖。骨组织工程种子细胞多集中在骨髓基质干细胞的研究上,实际上机体骨髓中所含的基质干细胞数量非常有限^[1],     

骨髓间质干细胞对血管性痴呆大鼠BDNF和NGF的影响

目的:观察静脉注射骨髓间质干细胞(marrowstromalcell,MSC)对血管性痴呆(vasculardementia,VD)大鼠脑组织脑源性神经营养因子(BDNF)和神经生长因子(NGF)表达的影响。方法:体外分离培养、扩增成年大鼠MSC。利用结扎双侧颈总动脉方法制备慢性前脑缺血动物模型,利用穿梭箱、水迷宫试验、ELISA方法检测注射MSC后30,60dVD大鼠认知功能以及脑组织BDNF和NGF含量变化。结果:与正常对照组相比,VD大鼠主动回避反应、游泳时间、错误次数和盲端时间,在造模后30和60d差异有显著性意义(P<0.01)。静脉注射MSC后30和60d,VD大鼠认知功能显著改善。造模后30,60d大鼠脑BDNF含量犤(96±27),(108±26)ng/g犦,NGF含量犤(73±22),(89±24)ng/g犦较对照组显著增高犤(51±13)ng/g,(36±11)ng/g犦,P<0.01;注射MSC后30,60d大鼠脑BDNF含量犤(180±46),(185±5)ng/g犦,NGF含量犤(115±29),(108±34)ng/g犦较模型组显著升高。结论:静脉注射MSC可显著改善VD大鼠认知功能,增加VD大鼠脑组织BDNF和NGF含量。     

骨髓间充质干细胞体外诱导为神经细胞的研究

本实验观察了大鼠MSCs向神经细胞方向的诱导分化情况,以期为MSCs在神经移植领域的临床应用提供理论基础。用含10 ng／ml bFGF+20％FBS的DMEM对MSCs进行预诱导24 h后,以含200μmol／L的BHA+2％DMSO的无血清DMEM对MSCs进行诱导,观察诱导后细胞的光、电镜形态学变化,通过免疫组织化学法对诱导后细胞进行神经细胞表型及神经递质合成酶鉴定。结果显示:MSCs经BHA和DMSO诱导后,80％以上的细胞表现出神经元样形态,胞浆内可见较多Nissl体,并表达nestin、NSE、NF、MAP、SYN,部分诱导后的细胞表达ChAT、TH、GAD;电镜下观察,诱导后细胞核大而圆,核仁明显,胞浆内细胞器发达,可见大量粗面内质网和游离核糖体。提示,MSCs体外可被诱导分化为神经元样细胞,诱导后的细胞有合成某些神经递质的能力并具有发育早期神经元的超微结构特点。     

兔MSCs-PGA复合物在RCCS中的成骨潜能

目的：探讨联合应用兔骨髓基质干细胞（MSCs）、聚乙醇酸（PGA）支架材料和成骨诱导因子在旋转式细胞培养系统（RCCS）中构建组织工程化骨的可行性。方法：利用密度梯度离心法分离培养兔骨髓基质干细胞，采用动力接种方法形成骨髓基质干细胞-PGA支架复合物，以成骨条件培养液为生长介质，在RCCS中进行培养。用趋骨性荧光素四环素标记、茜素红染色以及扫描电镜、透射电镜观察，研究细胞-PGA支架在RCCS中的成骨潜能。结果：组织化学研究显示，兔骨髓基质干细胞-PGA支架结构中RCCS中培养第7d时，茜素红钙质染色呈弱阳性反应，至第21d及28d时，茜素红钙质染色呈强阳性反应。荧光显微锏上，培养组织趋骨性荧光素四环素标记呈现金黄色荧光，荧光亮度与分泌物堆积程度成正相关，与钙质染色阳性反应结果一致。超微结构观察，见骨髓基质干细胞合成分泌胶原纤维，释放基质小泡，并出现钙盐沉积，形成丛毛球状钙球，最终形成骨组织。结论：在旋转式细胞培养系统中，骨髓基质干细胞-PGA支架结构具有形成类似活体骨组织特征的组织工程化骨的潜能。     

对比研究经颈动脉和立体定位注射骨髓间充质干细胞治疗缺血性脑卒中

目的探讨安全高效移植骨髓间充质干细胞(mensenchymal stem cells,MSCs)治疗脑缺血的方法.方法采用连续传代培养的方法纯化MSCs,Brdu标记后分别经颈动脉(1×104/μl,200 μl)和立体定位(1×105/μl,10 μl)注射至大鼠脑缺血2 h再灌注1 d模型,各自设立对照组,观察术后神经功能评分以及脑部Brdu阳性细胞的分布.结果经颈动脉注射组较立体定位注射组爬杆实验评分低(P＜0.05),Brdu阳性细胞存活多,迁移范围更广.结论 MSCs具有卒中后脑保护作用,而经颈动脉的给药方式优于立体定位注射.     

Isolation of the multipotent MSC subpopulation from human gingival fibroblasts by culturing on chitosan membranes

Literature has different opinions regarding the percentage of mesenchymal stem cell (MSC)-like population in human gingival tissue. Isolation of these cells is thus important for clinical applications. In this study, two typical but distinct types of gingival fibroblasts (GF), GF-A and GF-B, were grown from human gingival biopsies. They were characterized for surface markers by flow cytometry as well as the expressions of stemness and neural crest marker genes by RT-PCR. The two types of GF were slightly different in their surface markers; however, they had dramatic difference in the expression levels of stemness marker genes and neural crest marker genes. They also demonstrated distinct differentiation capacity. Upon the appropriate induction, GF-A were capable of osteogenic, adipogenic, chondrogenic, and neurogenic differentiation while GF-B only underwent osteogenic differentiation. By culturing either type of GF on chitosan membranes for 24 h, we were able to isolate two distinct subpopulations in each type of GF, i.e. cells with spheroid-forming ability (GF-AS and GF-BS) or those remained flat and attached (GF-AN and GF-BN). We further characterized these cells, and determined the common properties shared by the spheroid-forming subpopulation “S”, as well as those shared by the non-spheroid-forming subpopulation “N”. The subpopulation “S” was capable of the multilineage differentiation, while the subpopulation “N” was only efficient in osteogenic differentiation. GF-A and GF-B had different proportions of subpopulations. Chitosan as the cell culture substratum up-regulated the N-cadherin expression of the “S” but not “N” subpopulation, which may account for the cell sorting effect. This study showed that chitosan membranes could be used for isolation of the spheroid forming subpopulation in human GF that contained multipotent adult stem cells of which the number varied among donors and sites.     

加载机械振动对骨髓间质干细胞粘附和碱性磷酸酶活性的影响

目的是探索通过加载机械振动对骨髓间质干细胞（Mesenchymal stemcells,MSCs）的粘附、增殖和碱性磷酸酶活性的影响。材料和方法将频率（F）为300 Hz,加速度（α）为1.4 gm/s2的垂直的机械振动加载至成骨诱导培养中的MSCs。细胞传代培养后立即加载机械振动,并于1、2、3、6、12、24、48 h后观察细胞贴壁粘附形态变化,并检测培养14 d后细胞碱性磷酸酶（alkaline phosphatase,ALP）活性;对照组不加载机械振动。结果在48小时内实验组可见细胞集落,而在对照组未发现细胞集落的形成;相同时间点实验组的细胞贴壁数量明显多于对照组（P〈0.05）。但是两组在第14 d时的碱性磷酸酶水平无明显差异。结论通过加载机械振动促进了细胞粘附,这种方法有可能有用于促进种子细胞与基质材料之间的粘附,从而获得分布均一的组织工程细胞-材料复合物。     

MSCs局部注射对豚鼠光老化皮肤组织SOD、MDA的表达影响

目的 观察SOD、MDA在骨髓间充质干细胞(marrow mesenchymal stem cells,MSCs)修复皮肤光老化表达.方法 采用密度梯度离心法分离培养MSCs,利用UVB灯管照射豚鼠制备皮肤光老化模型,随机分为正常组、模型组及治疗组.检测皮肤组织中SOD、MDA表达.结果 MSCs细胞在豚鼠皮肤照射区局部多点皮内注射后,MDA表达较模型组明显下降(P＜0.01),但高于正常组.治疗组中SOD表达明显高于模型组(P＜0.01),低于正常组.结论 MSCs注射可以清除体内的氧自由基等有害物质,从而使抗氧化平衡系统保持稳定,MSCs对皮肤光老化有治疗作用.

Abstract：

Objective To observe the expressions of SOD, MDA on photoaging skin when it was repaired by bone marrow mesenchymal stem cells (MSCs) . Methods MSCs were isolated and cultured according to density gradient centrifugation. The model of photoaging skin was prepared in guinea pigs by exposure to UVB lamp. The animals were divided into the normal group, model group and treatment group. The expressions of SOD and MDA were detected in skin tissue. Results The expression of MDA in treatment group was significantly decreased compared with that of the model ( P ＜ 0. 01 ), but higher than that in the normal group. The expression of SOD in treatment group was significantly higher than that of model group ( P ＜ 0. 01 ), but lower than that of normal group. Conclusions MSCs can remove the harmful substances such as oxygen free radicals and keep the antioxidant balance of the system. MSCs play the role on the treatment of skin photoaging.     

骨髓间充质干细胞定向诱导分化成骨细胞移植修复腭裂骨缺损

目的：探讨骨髓间充质干细胞（MSCs）定向诱导分化成骨细胞修复先天性腭裂骨缺损的可行性。方法：将16只2月龄的日本大耳白兔,在其腭部制作骨组织缺损和采集胫骨MSCs培养,并随机分为2组。实验组：将骨髓间充质干细胞扩增后复合明胶海绵支架并行成骨细胞定向诱导分化后,植入腭骨缺损区;对照组：进行与实验组相同的手术操作,腭骨缺损区仅植入明胶海绵支架。于4、6、8和12周采集标本行大体观察、X线片观察等。结果：实验组4周时骨缺损部分有骨痂形成,第6周时实验组骨缺损表面有较多骨痴形成,与骨端连接紧密,有骨桥形成;第8、12周时实验组骨缺损完全修复。对照组骨缺损未见骨组织修复。结论：通过MSCs扩增并向成骨细胞定向诱导分化后,以明胶海绵支架为载体移植腭骨缺损中,能较好的修复骨组织缺损。