供体骨髓间充质干细胞干预非协调性异种肝移植免疫排斥反应的实验研究  

The study on immunomodulation of donor mesenchymal stem cells on discordant liver xenotransplantation

作　　者：王建伟[1] 刘颖斌[2] 许斌[3] 李江涛[2] 钱浩然[3] 张茂[4] 彭淑牖[2] 

机构地区：[1]浙江大学医学院附属第二医院肿瘤外科,杭州310009 [2]浙江大学医学院附属第二医院外科,杭州310009 [3]浙江大学医学院附属邵逸夫医院普外科 [4]浙江大学医学院附属第二医院急诊中心,杭州310009

出　　处：《中华外科杂志》2005年第19期1254-1258,共5页Chinese Journal of Surgery

基　　金：浙江省自然科学基金(M303820)

摘　　要：目的研究骨髓间充质干细胞(MSCs)对非协调性异种肝移植排斥反应的免疫干预作用。方法密度梯度离心法分离培养豚鼠MSCs,流式细胞仪检测其膜表面CD34、CD45、CD44和CD29的表达,脂肪细胞诱导液培养诱导MSCs向脂肪细胞分化。豚鼠MSCs输注大鼠后检测不同时相免疫球蛋白IgG、IgM、IgA以及补体C3、C4的浓度变化。豚鼠和大鼠各30只随机分为3组,所有大鼠受体经环磷酰胺预处理,A组为MSCs输注组;B组为生理盐水输注组;C组为地塞米松输注组。各组行豚鼠大鼠原位肝脏移植手术,观察受体的存活情况及排斥反应情况。结果MSCs膜表面CD34阴性、CD45阴性、CD44阳性、CD29阳性。脂肪细胞诱导液培养后细胞内出现脂滴沉着。MSCs细胞悬液输注受体后各个时相的免疫球蛋白IgG、IgM和补体C3较输注前有显著降低(P<0.01),而IgA和补体C4与输注前相比无明显变化(P>0.05),输注后的免疫球蛋白IgG、IgM和补体C3各个时相点之间的浓度变化无显著差异(P>0.05)。肝脏移植模型A组受体存活时间为(431±27)min,较B组和C组的存活时间[(148±16)min、(141±22)min]显著延长(P<0.01)。A组受体超急性排斥反应发生较迟,程度轻。结论MSCs的鉴定可根据细胞形态,膜表面标志物和分化能力来确定,MSCs可干预异种肝脏移植的超急性排斥反应的发生。Objective To observe the influence of guinea pig mesenchymal stem cells （MSCs） culture in vitro on the immunological status in heterogenous recipient, and to evaluate the immunomodulation of MSCs to the immune rejection of liver xenotransplantation. Methods The mononuclear cell fraction obtained by centrifugation over Percoll was cultured in vitro. Cell surface epitopes were analyzed by flow cytometry technique, and MSCs were incubated at the addition of adipocyte induction media, and stained in Oil Red O. The immunoglobulin and complement in rat recipient serum were assayed after infusion of donor MSCs. Thirty guinea pigs and thirty Wistar rats were randomly divided into three groups. All rat recipients undertook cyclophosphamide （CTX） infusion. Group A was MSCs infusion group, group B was saline infusion group and group C was dexamethasone （ DXM ） infusion group. Orthotopic guinea pig to rat liver transplantation model was established. The survival time and the immunopathology of graft were observed. Results According to flow cytometry assay, MSCs were positive for CD44 and CD29, while negative for CD34 and CD45. Lipid droplets were found in the MSCs cytoplasm after being incubated in adipogenic inducation media. The concentrations of IgG, IgM, C3 at all time point after MSCs infusion were significantly decreased than before （ P 〈 0.05 ） , while the concentrations of IgA and CA did not changed. The survival time of group A（431 ±27 min） was obviously longer than group B（ 148 ± 16 min） and group C （ 141 ± 22 min） （ P 〈 0.01 ）. The intense hyperacute rejection rapidly occurred of group B and C after blood re-perfusion. The severe immunopathological injury could be observed at the grafts of group B and C. However, the hyperacute rejection of group A occurred slightly. Conclusions MSCs can be identified by virtue of cell morphology, membrane phenotype and differential potential. MSC may play a role in the immunosuppression in hyperacute rejection in the liver xenotransplantatio

关 键 词：问充质干细胞 移植 异种 移植物排斥 移植免疫排斥反应 骨髓间充质干细胞 肝移植排斥反应 非协调性 细胞干 异种 免疫球蛋白IGG 

分 类 号：R657.3[医药卫生—外科学]