过表达IDO大鼠骨髓间充质干细胞改善异位移植心脏存活

Improved survival of heterotopic heart transplant overexpressing IDO rat bone mesenchymal stem cells

目的:探讨过表达IDO大鼠骨髓间充质干细胞(BMSC)改善大鼠腹腔异位移植心脏的存活机制。方法:通过慢病毒载体GV308携带IDO基因转染大鼠骨髓间充质干细胞构建过表达IDO大鼠骨髓间充质干细胞。建立大鼠腹腔异位移植心脏模型,经尾静脉给予相应细胞处理:(1)利用超声心动图检测移植心脏心功能变化。(2)采用小动物活体成像系统评估移植心脏局部荧光强度。(3)再取各组受体大鼠的脾脏,采用流式细胞技术检测CD40、CD86、CD80、MHCⅡ、CD274、CD45RA、CD45RA+CD45RB、Treg细胞的表达情况。(4)取各组移植心脏,采用HE染色评估炎性细胞浸润情况。(5)利用液相芯片检测各组血清IL-1ɑ、IL-4、IL-1β、IL-2、IL-10、IFN-γ、IL-18、TGFβ1、TGFβ2、TGFβ3因子的变化情况。结果:(1)给予相应细胞处理,可见过表达IDO-BMSCs处理后2 d移植心脏的EF、FS较其余各组有提高。(2)采用小动物活体评估可见过表达IDO-BMSCs组移植心脏局部荧光强度最强。(3)给予干预措施后2 d可见过表达IDO-BMSCs组脾脏细胞CD40、CD86、CD80、MHCⅡ、CD45RA、CD45RA+CD45RB表达降低,而CD274、Treg细胞的表达增高。(4)采用液相芯片检测各组提取血清可见在2 d时,过表达IDOBMSCs组血清中IL-1α、IL-4、IL-1β、IL-2、IFN-γ、IL-18是降低的;IL-10、TGFβ1、TGFβ2、TGFβ3表达量是升高的。HE染色证实过表达IDO-BMSCs组炎性细胞浸润少于其他组。结论:过表达IDO的BMSCs可以通过有效调节免疫DC、T细胞及细胞因子,从多个免疫层面改善移植心脏存活。

Objective： To explores the improvement in survival mechanism of a rat model of enterocoelia heterotopic heart transplant with rat bone marrow mesenchymal stem cells（ BMSCs） IDO-overexpressing. Methods： IDO-overexpressing rat BMSCs were produced through transfection of rat BMSCs with IDO gene carried by the lentiviral vector GV308. A rat model of enterocoelia heterotopic heart transplantation was established. This rat model received a cell treatment via its tail veins,as follows：（1）Echocardiography was employed to detect the functional changes in the transplanted heart.（2）The fluorescence intensity of the different parts of the transplanted heart was evaluated using a body imaging system for small living animals.（3）Receptors rat spleens cells were obtained and used for a flow cytometric detection of the expression levels of CD40,CD86,CD80,MHCⅡ,CD274,CD45 RA,CD45RA＋CD45RB,and Treg cells.（4） A transplanted heart was obtained after injection to evaluate inflammatory cell infiltration through HE staining.（5）Liquid phase chips were used to detect changes in the serum factors IL-1ɑ,IL-4,IL-1β,IL-2,IL-10,IFN-γ,IL-18,TGFβ1,TGFβ2 and TGFβ3 in after injection cells. Results：（1）After the rat heterotopic heart transplantation model and the corresponding cell treatment were established,after over-expressed IDO-BMSCs treatment 2 days the EF and FS were higher in the transplanted heart than other groups.（2）The fluorescence intensity of the parts of the transplanted heart was highest in the IDO-BMSC overexpression group as revealed by small animal living body evaluation.（3） Two days after the interventions,spleen cells in the over-expressed IDO-BMSCs group showed reduced expression levels of CD40,CD86,CD80,MHCⅡ,CD45 RA,CD45RA＋CD45RB and increased expression levels of CD274 and Treg cells as revealed by flow cytometry.（4）Liquid phase chips were used to examine the serum obtained from each group2 days after the intervention,and the results showed that the expression levels of IL-1α,IL-4,IL-1β,IL-2,IFN-γ and IL-18 in the IDOBMSC overexpression group decrease. By contrast,the expression levels of IL-10,TGFβ1,TGFβ2 and TGFβ3 increase. HE staining results demonstrate that inflammatory cell infiltration was lower in IDO-BMSC overexpression group than in other groups. Conclusion：IDO-overexpressing BMSCs improve the survival of a transplanted heart through effective adjustment of immune DC and T cells,as well as cell factors.