小梁网干细胞的归巢及其在小梁创伤愈合中的作用

目的研究胶原蛋白在小梁网干细胞(trabecular meshwork stem cells,TMSC)归巢中的作用以及小梁网(trabecular meshwork,TM)细胞和TMSC在创伤愈合中的相互作用。设计实验研究。研究对象人原代TM细胞和TMSC。方法分离、培养和传代人TM细胞和TMSC,通过定量RT-PCR和地塞米松诱导方法明确细胞类型;通过细胞贴附实验评价TM细胞及胶原蛋白对TMSC的亲和力;利用划痕损伤实验和时差显微镜观察TMSC分布及其与TM的相互作用。主要指标 TMSC和TM细胞的鉴定、TMSC穿膜数量和划痕损伤实验中TM细胞的迁移状态。结果TMSC高表达OCT4而TM细胞高表达CHI3L1和MYOC,地塞米松刺激后TM细胞MYOC的表达显著增加。TMSC在含有TM细胞、胶原蛋白、TM细胞+胶原蛋白的培养皿中的穿膜数分别为(13.1±5.3)个/高倍视野、(22.6±10.1)个/高倍视野、(4.8±2.2)个/高倍视野,均明显高于在单纯细胞培养皿中的穿膜数(3.7±0.5)个/高倍视野(P=0.0009,<0.0001,<0.001)。用AMD3100抑制TMSC中CXCR4的表达后,TMSC在各种状态的迁移数量差异消失。在TM细胞划痕损伤模型中加入TMSC可促进TM细胞的分化和迁移。结论 TMSC可诱导小梁网细胞分裂向受损区域迁移,从而促进组织损伤修复。胶原蛋白在TMSC归巢过程中发挥重要作用。

Trabecular meshwork stem cells homing and effect on the wound-recovering of trabecular meshwork

Objective To investigate the role of collagen on the homing of trabecular meshwork stem cells (TMSCs) and the interaction of the TMSCs and trabecular meshwork (TM) cells in damage repairing. Design Experimental study. Participants Human TM cells and TMSCs. Methods Human primary TM cells and TMSCs were cultured and passaged. Then they were characterized by quantitative RT-PCR for gene expression and dexamethasone-induced experiment. TMSCs affinity was evaluated by comparing attached cell numbers in a co-culture system. The interaction of the TM cells and TMSCs was recorded by the wound-healing assay and time-lapse microscope. Main Outcome Measures The characterization of the TMSCs and TM cells. Affinity of TMSCs and TM status in the wound-healing assay. Results TMSCs expressed higher level of OCT4 while TM cells expressed higher levels of Chitinase 3-like1 (CHI3L1) and myocilin (MYOC). The expression of MYOC increased in response to dexamethasone.  The numbers of TMSCs attached to the wells with TM feeder (13.1±5.3, P=0.0009), with collagen (22.6±10.1, P<0.0001), with TM feeder and collagen (4.8±2.2, P<0.001) were all significant higher than the attached cells in the plain cell culture wells (3.7±0.5). With ADM3100, inhibition of CXCR4, the numbers of TMSCs attached to the wells were similar in various condition. In wound-healing model, TMSCs induced the division and migration of wounded TM cells. Conclusion TMSCs induced the migration of TM cells to the wounded region hence promoting wound healing. Collage plays a role in the homing of TMSCs.