结直肠癌中缺氧诱导因子1-α与FasL 表达相关性的研允

目的 探讨结直肠癌侵袭转移过程中缺氧诱导因子1-α(alpha,HIF-1α)与FasL表达的相关性.方法 采用分子克隆方法,将我室已构建的FasL-pcD-NA3.1(+)质粒与pcDNA3.1(一)质粒进行重组,得到新的FasL-pcDNA3.1(一)质粒并加以鉴定;通过脂质体转染法将空质粒、FasL-pcDNA3.1(+)与FasL-pcDNA3.1(一)质粒分别转染人直肠癌HR-8348细胞,构建侵袭力不同的结直肠癌细胞HR-8348L、HR一8348F和HR一8348As,未转染细胞HR一8348B为空白对照,应用Tran-swell,小室检测各组细胞的侵袭能力;采用化学缺氧法构建四组细胞的缺氧模型,Western blot方法定量检测缺氧0h、6h、12h及24h各组细胞内HIF-1α的表达.结果 FasL-pcDNA3.1(一)质粒符合要求,FasL片段大小约900bp,测序结果正确率99.2%;单层细胞体外侵袭实验见HR一8348F细胞穿透Transwell滤膜的细胞数目为(12.930±2.434),显著多于HR-8348B(8.133±1.959)、HR-8348L(7.670±2.093)和HR-8348As(7.870±1.685)细胞(P<0.05);Western blot检测示HIF-1α蛋白于120kD处显色,缺氧0h与6h,各组样品中HIF-α仅表达微量,HIF-1α水平无显著性差异(P>0.05);缺氧12h与24h,HR一8348F细胞内HIF-1α水平较0h和6h时明显增高(P<0.05),而HR-8348B、HR-8348L及HR-8348As细胞内HIF-1α表达与6h时无明显变化(P>0.05),HR-8348F细胞HIF-1α水平显著高于HR-8348B、HR-8348L及HR-8348As细胞(P<0.01).结论 缺氧环境中结直肠癌细胞FasL表达增强是除低氧分压外另一个诱导HIF-1α表达增高的因素,FasL与 HIF-1α水平呈正相关,高侵袭能力的结直肠癌细胞对缺氧的适应能力加强,促进肿瘤的远处转移.

Correlation of hypoxia inducible factor-1 alpha and FasL in colorectal carcinoma

0bjective To explore the correlation of hypoxia inducible factor-1 alpha(HIF-1α) and FasL in the invasion and metastasis of colorectal carcinoma.was reconstructed and verificated.The empty plasmid,FasL-pcDNA3.1(+)plasmid with lipid infection.Then HR-8348 cells with different invasive abilities,HR-8348L,HR-8348F and HR-8348As were constructed and verificated,and the non-transfected cell HR-8348B was blank,in all of which the invasion abilities were detected.Hypoxia models for HR-8348 B,HR-8348L, HR-8348F and HR-8348As were constructed with chemical modeling.Then the expression of HIF-1αin all groups at Oh,6h,12h and 24h after hypoxia was quantitated with Western blot.Results The new constructed plasmid actitude ratio of sequencing was 99.2%.The number of HR-8348F cells(12.930±2.434) which penetrated Transwell was significantly more than that of HR-8348B (8.133±1.95),HR-8348L(7.670±2.093)and HR-8348As cells(7.870±1.685),respectively(P<0.05)in cell monolayer intrafiltration experiment in vitro.HIF-1αprotein was coloration at 120kD by Western blot,and the expression level of HIF-1α was not markedly different in all groups at 0h and 6h after hypoxia(P>0.05),but was significantly higher in HR-8348F cell than in HR-8348B,HR-8348L and HR-8348As cell(P<0.01)at 12h and 24h after hypoxia.Conclusions The expression enhance-ment of intracellular FasL in colorectal carcincoma in hypoxia could induce the expression of HIF-1α,which promote tumor cells to adapt hypoxia,and the level of FasL is posi-tively carrelated with HIF-1α.