过表达IDO骨髓间充质干细胞分泌外泌体改善移植心脏存活

Exosomes excreted by bone mesenchymal stem cells with overexpressed IDO improves survival of transplanted heart

目的探讨过表达吲哚胺2,3双加氧酶(IDO)大鼠骨髓间充质干细胞(BMSCs)通过分泌外泌体(exosome)改善大鼠腹腔异位移植心脏存活机制。方法构建过表达IDO大鼠骨髓间充质干细胞,而后提取分泌的exosome。同时建立大鼠腹腔异位移植心脏模型,经尾静脉给予相应细胞分泌的exosome后检测如下指标:移植心脏心功能变化;移植心脏局部荧光强度;受体大鼠脾脏细胞表面CD40、CD86、CD80、主要组织相容性抗原-Ⅱ(MHCⅡ)、CD274(PDL-1)、CD45RA、CD45RA+CD45RB表达及Treg细胞数量。并采用苏木精-伊红(HE)染色评估移植心脏局部炎性细胞浸润情况。结果 (1)大鼠异位心脏移植模型建立后,给予相应细胞分泌exosome,过表达IDO-BMSCs-exosome组移植心脏的射血分数(EF)、短轴缩短率(FS)较其余各组有提高;(2)过表达IDO-BMSCs-exosome组移植心脏局部Dir荧光最强;(3)采用流式细胞技术证实过表达IDO-BMSCs-exosome组受体脾细胞CD86、CD40、CD80、主要组织相容性抗原-Ⅱ(MHCⅡ)、CD45RA+CD45RB、CD45RA、OX62表达量下降,而CD274(PDL-1)表达增高同时Treg细胞数量较其他组增多;(4)HE染色证实过表达IDO-BMSCs-exosome组移植心脏局部有较少炎性细胞浸润。结论大鼠BMSCs分泌的exosome可以有效调节DC细胞表面抗原分子表达、促进Treg细胞增生,从而在多个免疫层面改善心脏移植排斥。

Objective To investigate the mechanism of cardiac myocyte survival in rats by overexpression of indoleamine 2,3-dioxygenase（ IDO） in rat bone marrow mesenchymal stem cells（ BMSCs）. Methods Rat BMSCs overexpression IDO were constructed. The vector was transfected into rat BMSCs,and then the secreted exosomes were extracted. A rat heart model with abdominal heterotopic transplantation was also established. Exosomes secreted by corresponding cells were administered through the caudal vein. Cardiac function changes were also monitored after transplantation. Local fluorescence intensities of transplanted hearts was further evaluated. Flow cytometry was used to detect the expressions of differentiation CD40,CD86,CD80,major histocompatibility complex（ MHCII）,CD274,CD45 RA,CD45 RA ＋ CD45 RB,and Treg cell. Transplanted hearts in three exosome groups were removed to evaluate corresponding extents of inflammatory cell infiltration by hematoxylin-eosin（ HE） staining. Results Corresponding cell treatments were administered after heterotopic heart transplantation models were established in rats. Ejection fraction（ EF） and fractional shortening（ FS） of transplanted heart in rats with IDO-overexpressing BMSCs exosomes were higher than those of other groups. Assessment with Caliper IVIS Lumina II also showed the highest local fluorescence intensity in transplanted heart of the same group. An interference measure was provided after 2 days,and flow cytometry results revealed that spleen cells of rats with IDO-BMSCs exosomes presented downregulated expression levels of CD40,CD86,CD80,MHCII,CD45 RA,and CD45 RA ＋ CD45 RB and upregulated expression levels of CD274 and Treg cell. HE staining proved lower inflammatory cell infiltration in the same group than in other groups. Conclusion Exosome secreted by IDO-BMSCs can effectively regulate production of immune DC,T cells,and cytokines and consequently improve survival of transplanted hearts from multiple immunization levels.