外周血干细胞输注对肺腺癌免疫行为的影响

Immunological impact of autologous peripheral blood stem cell infusion on lung adenocarcinoma

背景:目前临床上自体外周血干细胞分离鉴定方法相对较多,尚缺乏统一的标准。同时自体外周血干细胞在肺腺癌移植治疗过程中对免疫的影响也缺乏报道。目的:探讨自体外周血干细胞的分离、鉴定方法,观察自体外周血干细胞移植对肺腺癌大鼠免疫功能的影响。方法:采用密度梯度离心法从Wistar大鼠外周血分离单个核细胞并进行鉴定。将18只Wistar大鼠随机分为常规处理组和细胞移植组,每组9只,建立肺腺癌模型,常规处理组不给予任何方法治疗,细胞移植组经尾静脉注入0.2 mL含有2.5×106 L^(-1)的自体外周血干细胞悬液,大鼠治疗后检测其免疫指标。2组大鼠分别于治疗4,10,14 d后麻醉处死(n=3),观察自体外周血干细胞移植治疗对肺腺癌免疫行为的影响。结果与结论:(1)从大鼠外周血新分离的单个核细胞呈现圆形或者椭圆形,且体积相对较小;自体外周血干细胞培养5 d后细胞生长迅速,圆形细胞开始向两端伸展,并呈现梭形,从而能够形成较小的细胞集落;自体外周血干细胞体外诱导培养后先后经历了生长滞缓期、对数增殖期以及生长平台期;(2)流式细胞仪分选纯化的细胞,干细胞因子受体CD90、CD133及Flk-1均有表达,提示分离培养获得的细胞为自体外周血干细胞;(3)自体外周血干细胞培养4,10,14 d后检测其表面抗原表达结果显示,CD90,CD133呈现先升高后下降趋势;Flk-1呈现上升趋势;(4)在移植治疗后第4,10,14天,细胞移植组免疫指标肿瘤坏死因子诱导蛋白6显著高于常规处理组(P<0.05),而核因子κB显著低于常规处理组(P<0.05);(5)结果提示,自体外周血干细胞分离、鉴定方法比较简单,并且应用于治疗肺腺癌能够分泌大量营养因子,治疗过程中能够增强机体免疫。

BACKGROUND： Although clinical methods for autologous peripheral blood stem cell separation and identification are relatively more, there is no unified standard. Moreover, little is reported on immune effects on lung adenocarcinoma during autologous peripheral blood stem cell transplantation.OBJECTIVE：To study the effect of autologous peripheral blood stem cell transplantation on immunological function of lung adenocarcinoma rats by exploring the separation and identification methods of autologous peripheral blood stem cells. METHODS：The mononuclear cells were isolated from the peripheral blood of Wistar rats by density gradient centrifugation method and then identified.Eighteen Wistar rats were randomly divided into routine treatment group and cell transplantation group,with nine rats in each group.Animal models of lung adenocarcinoma were established in the rats,and underwent no treatment in the routine treatment group but injection of autologous peripheral blood stem cell suspension（0.2 m L,2.5×10~6/L）via the tail vein in the cell transplantation group.Immune indicators were detected in two groups after treatment.Three rats from each group were anesthetized and scarified at 4,10,14 days after treatment, respectively.Afterwards,we detected the effects of autologous peripheral blood stem cell transplantation on immunological function of lung adenocarcinoma rats.RESULTS AND CONCLUSION：Newly isolated mononuclear cells from rat peripheral blood showed a round or oval shape and were relatively small in size.After 5 days of culture,autologous peripheral blood stem cells grew rapidly,and the round cells began to spread to both ends and became spindle-shaped,thereby forming small colonies.Autologous peripheral blood stem cells induced in vitro experienced three stages successively：cell arrest,logarithmic and plateau phases.Flow cytometry showed that sorted cells were identified as autologous peripheral blood stem cells by expressing CD90,CD133 and Flk-1.After 4,10 and 14 days culture,the expression of CD133 increased first and then descended,and Flk-1 showed an increasing trend.Compared with the routine treatment group,the expression of tumor necrosis factor-αinduced protein 6 in the cell transplantation group was significantly higher（P〈0.05）,while the expression of nuclear factorκB was significantly lower（P〈0.05）.In conclusion,the isolation and identification methods of autologous peripheral blood stem cells are simple and can be used to secrete a large number of nutritional factors in the treatment of lung adenocarcinoma,which can enhance body immunity in the treatment.