移植间充质干细胞在胶原诱导性关节炎大鼠关节的分布和作用

目的 探讨移植人的异基因骨髓间充质干细胞(BM-MSCs)在类风湿关节炎(RA)动物模型胶原诱导性关节炎(CIA)大鼠炎症关节局部的转归以及对关节局部组织损伤的修复作用.方法 分别选取5只Wistar幼年大鼠和30只Wistar成年雌性大鼠,5只Wistar幼年大鼠用以提取BM-MSCs,30只Wistar成年雌性大鼠随机分为CIA造模A组、B组和正常C组各10只.BM-MSCs分离、传代培养后用5-溴脲嘧啶脱氧核苷(5-BrdU)进行标记.CIA大鼠成功造模后,采取尾静脉注射移植于A组和C组大鼠,移植细胞数为1.0×107/kg.移植后1-4周观察各组动物一般状况和关节局部肿胀度的变化;于移植4周后用关节组织病理切片局部免疫组织化学抗5-BrdU和骨保护素的方法观察BM-MSCs向受体关节局部的趋化和聚集以及对关节组织中骨保护素表达的影响.2组间比较采用t检验.结果 BM-MSCs移植后,受试动物关节肿胀开始消退[(200±350)μl],移植后2、3、4周时与未移植BM-MSCs的B组动物[(320±110)μl]之间比较差异有统计学意义(P＜0.05);在受者关节滑膜、血管内膜及软骨组织中均可见5-BrdU阳性细胞,滑膜组织处的平均灰度值CIA造模A组(85±9)低于正常C组(110±6),差异有统计学意义(P＜0.05);BM-MSCs移植后CIA造模A组关节滑膜中骨保护素阳性率增加,平均灰度值为54±4,明显低于未移植的CIA造模B组(77±6),差异有统计学意义(P＜0.05).结论 通过外周静脉移植的BM-MSCs可以特异性趋化和聚集到炎症关节局部,并可以减轻关节损害,而这种局部组织损伤修复作用可能是通过增加局部炎症关节组织中骨保护素的含量而发挥的.

Abstract：

Objective To study the distribution of allogenic bone marrow-derived mesenchymal stem cells (BM-MSCs) on joints of collagen-induced arthritis (CIA) rats and to investigate their repair effects on joint damages. Methods Five Wistar rats were used for extraction of mesenchymal stem cells and 30 adult female Wistar rats were divided into 3 groups: the CIA rats group A (n=10), CIA rats group B (n=10) and normal rats control group C (n=10). BM-MSCs of Wistar rats were isolated, cultured in vitro routinely and the fourth passages was taken for identification of specific surface antigens by flow cytometry, then the cells were labeled with 5-bromodeoxyuridine (5-BrdU) in vitro. The models of CIA rats were established. 5-BrdU labeled BM-MSCs (1.0×107 cells/kg) were imfused from through tail vein to CIA rats group A and control group C. During the first 4 weeks after BM-MSCs transplantation, changes of general condition and left hind paw swelling were examined. At the fourth week, immunohistochemical examination of 5 -BrdU and osteoprotegerin (OPG) were performed to investigate BM-MSCs aggregation around the knee joints. The contribution of BM -MSCs to repairing of joint damages was identified. Comparisons between groups were performed by t-test. Results After BM-MSCs transplantation, left hindpaw swelling of group A were relieved compared with group B (P＜0.05) and the mobility of the joints was significantly improved. At the fourth week, much more implanted cells (5-BrdU positive cells.) were detected in the damaged knee joints than those in normal knee joints. The average grey scale values on synovium of knee joints in the CIA group A (85±9) was significantly lower than that of the normal group C (110±6, P＜0.05). At the same time, OPG expression was increased in damaged knee joints. The average grey scale values on synovium of knee joints in CIA group A (54±4) was significantly lower than that of the CIA group B (77±6, P＜0.05). Conclusion The transplanted allogeneic bone marrow mesenchy-mal stem cells can migrate to sites of damaged tissue in arthritis. They can prevent tissue damage and repair the damaged joints tissue by increasing OPG expression. This study has provided some evidence for developing effective therapy for rheumatoid arthritis.

The distribution and effect of allogenic mesenchymal stem cells on joints of collagen induced arthritis rats

Objective To study the distribution of allogenic bone marrow-derived mesenchymal stem cells (BM-MSCs) on joints of collagen-induced arthritis (CIA) rats and to investigate their repair effects on joint damages. Methods Five Wistar rats were used for extraction of mesenchymal stem cells and 30 adult female Wistar rats were divided into 3 groups: the CIA rats group A (n=10), CIA rats group B (n=10) and normal rats control group C (n=10). BM-MSCs of Wistar rats were isolated, cultured in vitro routinely and the fourth passages was taken for identification of specific surface antigens by flow cytometry, then the cells were labeled with 5-bromodeoxyuridine (5-BrdU) in vitro. The models of CIA rats were established. 5-BrdU labeled BM-MSCs (1.0×107 cells/kg) were imfused from through tail vein to CIA rats group A and control group C. During the first 4 weeks after BM-MSCs transplantation, changes of general condition and left hind paw swelling were examined. At the fourth week, immunohistochemical examination of 5 -BrdU and osteoprotegerin (OPG) were performed to investigate BM-MSCs aggregation around the knee joints. The contribution of BM -MSCs to repairing of joint damages was identified. Comparisons between groups were performed by t-test. Results After BM-MSCs transplantation, left hindpaw swelling of group A were relieved compared with group B (P＜0.05) and the mobility of the joints was significantly improved. At the fourth week, much more implanted cells (5-BrdU positive cells.) were detected in the damaged knee joints than those in normal knee joints. The average grey scale values on synovium of knee joints in the CIA group A (85±9) was significantly lower than that of the normal group C (110±6, P＜0.05). At the same time, OPG expression was increased in damaged knee joints. The average grey scale values on synovium of knee joints in CIA group A (54±4) was significantly lower than that of the CIA group B (77±6, P＜0.05). Conclusion The transplanted allogeneic bone marrow mesenchy-mal stem cells can migrate to sites of damaged tissue in arthritis. They can prevent tissue damage and repair the damaged joints tissue by increasing OPG expression. This study has provided some evidence for developing effective therapy for rheumatoid arthritis.