骨髓间充质干细胞胰系方向诱导分化的成瘤性及致瘤性  

The tumorigenicity and oncogenicity of bone mesenchymal stem cells during differentiation into pancreatic lineage cells in vitro

作　　者：姜达琳 刘国旺[1] 孙琢玉[2] 王心[3] Jiang Dalin;Liu Guowang;Sun Zhuoyu;Wang Xin(Department of Gastroenterology,the Second Renmin Hospital,Tianjin 300192,China;Department of Public Health,Tianjin Medical University,Tianjin 300251,China;Department of Emergency Surgery,the 4th Central Hospital,Tianjin 300140,China)

机构地区：[1]天津市第二人民医院消化内科,天津300192 [2]天津医科大学公共卫生学院,天津300251 [3]天津市第四中心医院急诊外科,天津300140

出　　处：《中华实验外科杂志》2023年第1期40-42,共3页Chinese Journal of Experimental Surgery

基　　金：天津市卫生与健康委员会科技人才培育项目(RC20093)。

摘　　要：目的探讨骨髓间充质干细胞(BMSCs)在体外诱导分化为胰系细胞过程中的潜在成瘤性以及体内移植后的潜在致瘤性。方法选取雄性SD大鼠(购自中科院上海动物实验中心),分离大鼠BMSCs进行体外培养及传代,选取生长良好的第2代细胞进行诱导,检测胰腺特征性蛋白的表达,同时检测端粒酶活性,c-myc和p16基因的表达;随后将经或未经诱导的第2代BMSCs制成单细胞悬液,调整浓度至3.0×10^(10)/L,采用随机数字法将SD大鼠分为3组,每组30只,分别为:BMSCs细胞组(注射未经诱导的BMSC细胞悬液0.2 ml),诱导BMSC细胞组(注射诱导后的BMSC细胞悬液0.2 ml),空白对照组(注射等量含胎牛血清的DMEM/F12培养液0.2 ml),6个月后留取活检组织,检测端粒酶活性,c-myc和p16基因的表达。多组间比较采用单因素方差分析,两组数据间比较采用t检验。结果诱导后可见胰腺特征性蛋白α-淀粉酶、细胞因子-7、C-肽、胎肝激酶-1的表达。随着BMSCs在体外传代、诱导分化及体内移植,端粒酶以及c-myc基因活性逐渐降低,体内移植后,BMSCs细胞组端粒酶活性低于空白对照组(0.45±0.12比1.32±0.41,P<0.05),诱导BMSCs细胞组端粒酶活性低于空白对照组(0.13±0.04比1.32±0.41,P<0.05),诱导BMSCs细胞组端粒酶活性低于BMSCs细胞组(0.13±0.04比1.32±0.41,P<0.05);同样,体内移植后,BMSCs细胞组c-myc活性低于空白对照组(3.26±1.33比5.85±1.42,P<0.05),诱导BMSCs细胞组c-myc活性低于空白对照组(1.42±1.15比5.85±1.42,P<0.05),而诱导BMSCs细胞组c-myc活性低于BMSCs细胞组(1.42±1.15比3.26±1.33,P<0.05);p16在各阶段基因功能正常,无突变及缺失。结论BMSCs在体外培养生长状态良好,细胞纯度高,可定向诱导能分化为胰系细胞,体外无成瘤性,植入动物体内后无致瘤性。Objective To study the potential tumorigenicity and oncogenicity of bone mesenchymal stem cells(BMSCs)during differentiation into pancreatic lineage cells in vitro and after implantation in vivo.Methods After adult Sprague-Dawley rats were killed,the BMSCs were isolated,cultured and passaged in vitro.At the 2nd passage,BMSCs were harvested for induction after which the pancreatic functional proteins,telomerase activity,and the expression of c-myc and p16 gene were measured.Induced or non-induced BMSCs were made into single cell suspension,and cell concentration was adjusted to 3.0×10^(10)/L.The Sprague-Dawley rats were divided into 3 groups:BMSCs group(0.2 ml suspension with non-induced BMSCs was subcutaneously incubated in mice),induced BMSCs group(0.2 ml suspension with induced BMSCs was subcutaneously incubated in mice),blank control group(0.2 ml suspension with DMEM/F12 containing fetal bovine serum was subcutaneously incubated in mice).After 6 months,biopsy tissues were retained for the detection of telomerase activity,and the expression of c-myc and p16 gene.Results After induction,the pancreatic functional proteins including α-Amy,CK7,C-peptide,and Flk-1 were increased.The telomerase activity decreased in the 2nd,3rd and 6th BMSCs passages,as well as in the induced BMSCs and still declined after implantation in vivo.After implantation,the telomerase activity in BMSCs group was lower than in the blank control group(0.45±0.12 vs.1.32±0.41,P<0.05).The telomerase activity in induced BMSCs group was lower than in the blank control group(0.13±0.04 vs.1.32±0.41,P<0.05).And the telomerase activity in the induced BMSCs group was lower than the BMSC group(0.13±0.04 vs.1.32±0.41,P<0.05).Also,after implantation,the c-myc gene expression in BMSCs group was lower than in the blank control group(3.26±1.33 vs.5.85±1.42,P<0.05).The c-myc gene expression in induced BMSCs group was lower than in the blank control group(1.42±1.15 vs.5.85±1.42,P<0.05).And the c-myc gene expression in the induced BMSCs group was low

关 键 词：骨髓间充质干细胞 胰系细胞 成瘤性 致瘤性 

分 类 号：R735.9[医药卫生—肿瘤]