IL-10基因修饰骨髓源性肝干细胞移植对肝纤维化大鼠细胞外基质积聚的影响  

Transplantation of interleukin 10-modif ied bone marrow-derived liver stem cells reduces accumulation of extracellular matrix in fi brotic liver in rats

作　　者：兰玲[1] 陈源文[2] 孙超[2] 刘博伟[1] 孙巧玲[3] 

机构地区：[1]河南省人民医院二内消化科,河南省郑州市450003 [2]上海交通大学医学院附属新华医院消化内科,上海市200092 [3]上海交通大学医学院附属第六人民医院老年科,上海市200233

出　　处：《世界华人消化杂志》2009年第22期2231-2236,共6页World Chinese Journal of Digestology

基　　金：国家自然科学基金资助项目;No.30500236~~

摘　　要：目的:评价IL-10基因修饰骨髓源性肝干细胞(BDLSCs)移植对肝纤维化大鼠细胞外基质(ECM)积聚的影响.方法:利用免疫磁珠细胞分选(magnetic bead cell sorting,MACS)方法分选大鼠β2m-/Thy-1+BDLSCs.将腺病毒介导的IL-10基因转导至BDLSCs,采用ELISA法检测IL-10蛋白分泌水平.将大鼠随机分为4组:正常组、模型组、BDLSCs组及BDLSCs/IL-10组.采用PCR法检测Y染色体性别决定基因Sry;采用VG染色法观测肝组织胶原沉积面积;Western blot法检测肝组织α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)表达;采用碱水解法检测肝组织羟脯氨酸(hydroxyproline,Hyp)浓度;ELISA法检测血清透明质酸(HA)、层粘连蛋白(LN)和Ⅲ型前胶原(PCⅢ)等细胞外基质含量.结果:MASC法能成功分选纯化BDLSCs.IL-10修饰BDLSCs持续高浓度分泌IL-10蛋白至胞外,移植后能顺利种植于肝内,减轻胶原沉积,减少α-SMA表达从而抑制肝星状细胞活化,与BDLSCs组比较,BDLSCs/IL-10组大鼠肝组织Hyp水平显著降低(255.0±50.5μg/gvs373.0±26.7μg/g,P<0.01),血清HA,LN及PCⅢ水平也有所降低,差异具有统计学意义(40.5±7.7μg/Lvs79.4±10.3μg/L,61.5±16.4μg/Lvs77.7±12.6μg/L,14.3±0.8μg/Lvs14.9±1.5μg/L,P<0.01或0.05).结论:IL-10基因修饰BDLSCs移植能有效减少肝纤维化大鼠ECM的积聚,为治疗肝纤维化提供了新思路.AIM：To investigate the effects of transplanting interleukin 10（IL-10）-modified bone marrow- derived liver stem cells （BDLSCs） on accumulation of extracellular matrix in fibrotic liver in rats.METHODS： Rat beta-2 microglobulin（β2m）^-/ Thy-1^＋ BDLSCs were isolated by magnetic bead cell sorting （MACS）, and transduced with adenovirus-mediated IL-10 gene. The level of IL-10 protein secretion by BDLSCs was assessed by ELISA. The rats were divided randomly into four groups： normal group, model group, BDLSCs group and BDLSCs plus IL-10 group. IL-10 gene-modified BDLSCs from male rats were transplanted into female liver fibrosis rats via a branch of the portal vein. Sry gene was amplified by PCR to evaluate the implantation of BDLSCs in liver. Collagen area in liver tissues was detected by Van Gieson＇s （VG） staining. Expression of α-smooth muscle actin （α-SMA） protein in liver tissue was determined by Western blot. Hydroxyproline （Hyp） in liver tissues was quantified by the alkaline hydrolysis method. Extracellular matrix （ECM） proteins in serum were quantified by ELISA.RESULTS： BDLSCs were successfully isolated by MACS. IL-10 secreted by IL-10 gene-modified BDLSCs presented persistently at a high level. Transduced BDLSCs were implanted success- fully into impaired liver. Transplantation of IL-10 gene-modified BDLSCs lessened deposi- tion of collagen, decreased α-SMA expression and thereby suppressed activation of hepatic stellate cells. Compared with the BDLSCs group, the level of Hyp in liver tissue decreased markedly in the BDLSCs plus IL-10 group （255.0±50.5μg/g vs 373.0±26.7μg/g,P〈0.01）, and the levels of ECM proteins in serum also decreased （40.5±7.7μg/L vs 79.4±10.3μg/L,61.5±16.4μg/L vs 77.7±12.6μg/L,14.3±0.8μg/L vs 14.9±1.5μg/L,P〈0.01 or 0.05）.CONCLUSION： Transplantation of IL-10 gene-modified BDLSCs can effectively decrease accumulation of ECM in fibrotic liver in rats, suggesting the potential utility of this novel combined strategy of

关 键 词：骨髓源性肝干细胞 白介素10 肝纤维化 细胞外基质 

分 类 号：R575.2[医药卫生—消化系统]