骨髓间充质干细胞源性微囊泡对SD大鼠膝关节软骨损伤修复的影响

发布时间：2018-04-30 18:20

  本文选题：骨髓间充质干细胞 + 微囊泡　； 参考：《山西医科大学》2017年硕士论文

【摘要】：研究目的:构建大鼠膝关节软骨缺损模型,关节腔内分别注射生理盐水、BMSCs-MVs和BMSCs,通过检测修复软骨组织中的相关因子含量变化,比较各组软骨损伤的修复效果,探讨BMSCs-MVs对软骨组织损伤修复的影响。研究方法:1、全骨髓贴壁法分离、培养SD大鼠的BMSCs并鉴定;2、用超速离心法从上述BMSCs培养液中提取微囊泡(microvesicles,MVs)并用电镜观察其形态;3、构建SD大鼠膝关节软骨缺损模型,随机分为六个大组(每组6只):假手术组、生理盐水组、BMSCs组、BMSCs-MVs组(高、中、低3个剂量组)。每组双膝关节腔内注射分别注射不同实验试剂,每周注射1次,连续4、8次,每组分别于注射4周、8周后各处死3只,收集标本备用;4、修复组织的大体观察;组织切片染色(HE、糖原染色、甲苯胺蓝染色);免疫组化观察修复区Ⅱ型胶原、Ⅹ型胶原、MMP-13含量;修复组织Wakitani组织学评分;RT-PCR修复软骨组织Ⅱ型胶原、Ⅹ型胶原、MMP-13含量测定。5、统计学分析:本研究全部数据用SPSS18.0统计软件处理,计量资料用均数±标准差表示,用单因素方差分析行多组间比较,卡方检验行两组间比较,P0.05为差异有显著性意义。结果:1.大体观,BMSCs组、低剂量组缺损填充较完整、有光泽、边缘整齐,与周围组织界限难以区分;2.低剂量组Ⅱ型胶原表达为阳性,糖胺多糖含量较假手术组减少,较生理盐水组增多,Ⅹ型胶原、MMP-13含量较间充质干细胞组、生理盐水组减少;3.低剂量组改良Wakitani评分术后6周、10周低于生理盐水组,且差异有统计学意义(P0.05),高于间充质干细胞组,差异无统计学意义(P0.05);术后6周与术后10周相比,生理盐水组、低剂量组、间充质干细胞组Wakitani评分均降低,差异有统计学意义(P0.05);4.低剂量组术后6周、10周COL-II mRNA相对表达量与间充质干细胞组相比差异无统计学差异(P0.05)。5.低剂量组术后6周、10周改良Wakitani评分、免疫组化、RT-PCR均较高剂量组、中剂量组表现更优,差异有统计学意义(P0.05);结论:骨髓间充质干细胞源性微囊泡可以促进软骨缺损的修复。骨髓间充质干细胞可以通过旁分泌途径促进软骨修复。
[Abstract]:Objective: to establish the cartilage defect model of knee joint in rats, and to compare the repair effect of cartilage injury in each group by detecting the changes of relative factors in cartilage tissue by injecting normal saline into the articular cavity of BMSCs-MVs and BMSCs respectively. To investigate the effect of BMSCs-MVs on cartilage injury and repair. Methods BMSCs of SD rats was isolated and identified by the whole bone marrow adherent method. Microvesiclesus microvesicus MVss were extracted from the BMSCs medium by ultracentrifugation and the morphology of MVs3 was observed by electron microscope to establish the cartilage defect model of the knee joint of SD rats. They were randomly divided into six groups (6 rats in each group: sham operation group, normal saline group, BMSCs group, BMSCs-MVs group) (high, middle and low dose groups). Different experimental reagents were injected into the knee joint of each group once a week for 4 consecutive times. 3 rats in each group were killed after 4 weeks and 8 weeks of injection respectively. Tissue sections were stained with HEH, glycogen, toluidine blue staining; immunohistochemistry was used to observe the content of collagen 鈪，

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