Acceleration of bone regeneration by overexpression of vascular endothelial growth factor gene in bone marrow mesenchymal stem cells

(1)将小鼠sox9 cDNA转染骨髓间充质干细胞（MSCs），观察其成软骨分化情况。这些sox9转染的MSCs的体外高密度微质量培养表明，富含基质的聚集体被阿利新蓝和II型胶原阳性染色。接下来，将sox9转染的MSCs装入扩散室，移植到胸腺小鼠体内。移植4周后腔内可见大量组织形成。组织学检查显示，肿块的细胞外基质中Alcian blue和II型胶原蛋白均呈阳性染色，而X型胶原蛋白未染色。(2)评价串联传代大鼠间充质干细胞成骨潜能。通过碱性磷酸酶和骨钙素的浓度和mRNA表达来评价体外成骨分化。为了在体内成骨，将不同分化程度的MSCs植入胸腺小鼠体内。虽然成骨标志物水平的升高在连续培养的较少传代的MSCs中是显著的，但它们随着传代而降低。通过P2细胞观察到P0膜内有丰富的骨和软骨形成。然而，在P3细胞中，腔室充满纤维组织。(3)回顾了3例软骨发育不全患者和2例腿长不齐患者的5个股骨和3个胫骨的MSCs +富血小板血浆（PRP）牵张成骨的临床结果。用成骨剂培养髂骨骨髓间充质干细胞并分化为成骨细胞样细胞。移植前准备PRP。将培养的扩增成骨样细胞和自体PRP与凝血钙混合物一起注射到分散的愈伤组织中，使PRP凝胶在注射部位发育。每条腿均获得靶长度，无重大并发症，平均愈合指数为24.0天/厘米。少

(1)Mouse sox9 cDNA was transfected into bone marrow derived mesenchymal stem cells(MSCs) and chondrogenic differentiation was evaluated. In vitro high density micromass culture of these sox9 transfected MSCs demonstrated that a matrix-rich aggregate was positively stained by Alcian blue and type II collagen. Next, sox9 transfected MSCs were loaded into the diffusion chamber and transplanted into athymic mice. A massive tissue formation was visible in the chamber after 4 weeks transplantation. Histological examinations demonstrated that both Alcian blue and type II collagen were positively stained in the extracellular matrix of the mass while type X collagen was not stained.(2)Osteogenic potential of serially passaged rat MSCs was evaluated. Osteogenic differentiation in vitro was evaluated by the concentration and mRNA expression of alkaline phosphatase and osteocalcin. For in vivo osteogenesis, MSCs in various degrees of differentiation were implanted into the athymic mice. Although e … More levated levels of osteogenic markers were prominent in the less passaged MSCs continuously cultured with osteogenic supplements, they decreased with passaging. Abundant bone and cartilage formations inside the membrane were observed in the P0 through P2 cells. In the P3 cells, however, the chambers were filled with fibrous tissues.(3)Clinical results of distraction osteogenesis with transplantation of MSCs and platelet rich plasma(PRP) were reviewed in five femora and three tibiae of the three patients with achondroplasia and two patients with leg length discrepancy. MSCs derived from the iliac crest were cultured with osteogenic supplements and differentiated into osteoblast-like cells. PRP was prepared just before transplantation. Culture expanded osteoblast-like cells and autologous PRP were injected into the distracted callus with the thrombin-calcium mixture so that the PRP gel might develop within the injected site. The target lengths were obtained in every leg without major complications and the average healing index was 24.0 day/cm. Less