新鲜和冷冻人脐带来源间充质干细胞对大鼠骨关节炎的治疗作用比较

Comparison of therapeutic effects of fresh and frozen human umbilical cord-derived mesenchymal stem cells on osteoarthritis in rats

目的:比较新鲜和冷冻人脐带来源间充质干细胞对Ⅱ型胶原酶诱导的大鼠骨关节炎的治疗作用。方法:选用Sprague-Dawley (SD)大鼠,采用Ⅱ型胶原酶关节腔注射法建立大鼠骨关节炎模型,将大鼠随机分为正常组(Normal)、模型组(Model)、新鲜人脐带来源间充质干细胞组(Fresh hUC-MSC)、冷冻人脐带来源间充质干细胞组(Frozen hUC-MSC)和透明质酸组(HA),每组10只,于第一次注射Ⅱ型胶原酶后第7天给药。Fresh hUC-MSC和Frozen hUC-MSC组大鼠分别膝关节腔注射Fresh hUC-MSC和Frozen hUC-MSC 1×10^6/50 μL;HA组大鼠给予HA 50 μL;正常组和模型组大鼠给予等体积无菌生理盐水。给药后6周处死大鼠,观察各项指标。检测各组大鼠膝关节周长差值、关节软骨大体观察评分、膝关节X射线观察及评分、膝关节病理变化以及膝关节软骨基质金属蛋白酶-13(matrix metalloproteinase-13,MMP-13)和基质金属蛋白酶抑制因子-1(tissue inhibitor of metalloproteinase-1,TIMP-1)的表达情况。结果:与模型组相比,Fresh hUC-MSC和Frozen hUC-MSC组关节周长差值和关节软骨大体评分均显著减小,差异均有统计学意义(P<0.01);Fresh hUC-MSC和Frozen hUC-MSC组X射线的凯尔格伦-劳伦斯分级评分以及番红固绿染色病理变化的Mankin's评分差异均有统计学意义(P<0.05)。同时,Fresh hUC-MSC和Frozen hUC-MSC组的MMP-13的表达显著降低,TIMP-1的表达显著升高,MMP-13和TIMP-1的比值显著降低,并且差异均具有统计学意义(P<0.01)。结论:Fresh hUC-MSC和Frozen hUC-MSC均能改善大鼠骨关节的损伤,两组治疗骨关节炎差异无统计学意义,为临床应用hUC-MSC治疗提供实验依据。

AIM:To observe the therapeutic effect of fresh and frozen human umbilical cord-derived mesenchymal stem cells on type II collagenase-induced osteoarthritis in rats. METHODS:Sprague-Dawley (SD) rats with the model of osteoarthritis established by intra-articular injection of type II collagenase after one week of modeling, which were randomly divided into normal group (Normal), model group(Model), fresh human umbilical cord-derived mesenchymal stem cell group (Fresh hUC-MSC), and frozen human umbilical cord-derived mesenchymal stem cell group (Frozen hUC-MSC) and hyaluronic acid group (HA), with the number of 10 in each group. Rats in the Fresh hUC-MSC or Frozen hUC-MSC were injected into the knee joint cavity with 1×10^6/50 μL of fresh or frozen human umbilical cord-derived mesenchymal stem cells, respectively, in HA were given 50 μL of hyaluronic acid, and in Normal and Model group were given an equal volume of sterile physiological saline. The rats were sacrificed 6 weeks after the administration to observe various indexes.The knee circumference difference, articular cartilage gross observation score, knee X-ray, knee joint pathological changes and the expression of knee cartilage matrix metalloproteinase-13 (MMP-13) and matrix metalloproteinase inhibitor-1(TIMP-1) were detected in each group. RESULTS:Compared with the model group, the difference of joint circumference and the total score of articular cartilage in Fresh hUC-MSC and Frozen hUC-MSC group were significantly decreased, and the difference was statistically significant (P<0.01). The Kelgren-Lawrence grading scores of X-rays in Fresh hUC-MSC and Frozen hUC-MSC group and Mankin's scores in pathological changes of Safranin green staining were significantly different (P<0.05). At the same time, the expression of MMP-13 was significantly decreased in Fresh hUC-MSC and Frozen hUC-MSC group, the expression of TIMP-1 was significantly increased, the ratio of MMP-13 and TIMP-1 was significantly decreased, and the difference was statistically significant(P<0.01). CONCLUSION:Fresh hUC-MSC and Frozen hUC-MSC can improve the injury of bone and joint in rats, and the difference between the two groups was not significant. At the same time, it provides experimental basis for clinical application of hUC-MSC treatment.