肠三叶因子对氨甲喋呤所致肠黏膜细胞损伤的保护作用  

Protective effect of intestinal trefoil factor on methotrexate-induced injury in intestinal mucosa

作　　者：苏华芳[1] 邹阮敏[2] 俞康[3] 江松福[3] 

机构地区：[1]温州医学院附属第一医院放射化学治疗科,325003 [2]温州医学院附属第一医院妇产科,325003 [3]温州医学院附属第一医院血液科,325003

出　　处：《中华消化杂志》2011年第3期164-168,共5页Chinese Journal of Digestion

基　　金：浙江省自然科学基金资助项目（Y207422）;浙江省医药卫生科学研究基金资助项目（2007A137）;温州市科技局科技合作资助项目（H20090010）

摘　　要：目的 探讨肠三叶因子（ITF）保护甲氨喋呤（MTX）所致肠黏膜损伤的可能机制.方法 IEC-6细胞分为仅予培养液的空白对照组、经MTX处理的MTX对照组、仅予ITF预处理的ITF对照组和经不同浓度ITF预处理后再予MTX的ITF实验组.RT-PCR法检测E-cadherinmRNA表达变化.明胶酶谱法检测基质金属蛋白酶（MMP）-2和MMP-9活性.分光光度计法检测Caspase-3活性.细胞计数试剂盒-8法检测细胞增殖情况.改良的Boyden趋化小室法观测细胞迁移情况.结果 0.1 mg/mlITF实验组、1 mg/ml ITF实验组E-cadherin mRNA的相对表达量（分别为0.538±0.109和0.528±0.132）与MTX对照组（0.763±0.139）比较差异有统计学意义（P值分别=0.021和0.025）.各组间MMP-2和MMP-9活性差异均无统计学意义（P值均＞0.05）.0.1 mg/ml ITF实验组和1.0 mg/ml ITF实验组Caspase-3活性分别为0.077±0.009和0.044±0.009,较MTX对照组（0.090±0.011）下降（P值分别=0.032和0.005）.各ITF实验组的细胞增殖情况与MTX对照组比较差异均无统计学意义（P值分别=0.132、0.150、0.114、0.367）.1.0 mg/ml ITF实验组、0.1 mg/mlITF实验组Boyden小室穿膜细胞数[分别为（224.33±34.67）和（143.56±25.23）个]与MTX对照组[（50.11±7.77）个]比较差异均有统计学意义（P值均＜0.05）,两实验组间亦差异有统计学意义（P＜0.05）.结论 ITF的保护作用与其促细胞迁移和抗细胞凋亡功能有关,且不会导致明显的细胞增殖.其促细胞迁移功能可能与下调E-cadherin基因转录水平有关.Objective To investigate the potential mechanism of intestinal trefoil factor（ITF）against methotrexate （MTX）- induced injury in intestinal mucosa. Methods Cultured IEC-6 cells were divided into groups as follows： blank group, MTX treated group, ITF treated group and experimental group treated with gradient concentrations of ITF plus MTX. Expression of E-cadherin mRNA was determined by Real-Time polymerase chain reaciton （RT- PCR）. The activity of matrix metalloproteinase（MMP）-2 and MMP-9 was measured by gelatin zymogramphy. Caspases-3 activity was measured by colorimetric assay. Cell proliferation was assessed by cell counting kit-8 （CCK-8）assay. Migration of IEC-6 in vitro was observed using modified Boyden chamber assay. Results The expression of E-cadherin mRNA in experimental group （treated with 0.1 mg/ml or 1 mg/ml of ITF） was significantly down-regulated （0. 538±0. 109 or 0. 528±0. 132, respectively） in comparison with MTX treated group （0. 763±0. 139） with significant difference （P=0. 021 or P=0. 025, respectively）. There was no significant difference in activity of MMP-2 and MMP-9 among groups （P＞0. 05）. When compared with MTX treated group （0. 090 ±0. 011 ）, the activity of Caspase3 in experimental group （treated with 0. 1 mg/ml or 1 mg/ml of ITF） was significantly decreased （0. 077±0. 009, P=0. 032 or 0. 044±0. 009,P=0. 005, respectively）. There was no statistical difference in cell proliferation between experimental group （treated with 1 μg/ml, 0.01 mg/ml, 0. 1 mg/ml or 1.0 mg/ml of ITF） and MTX treated group （P=0. 132,0. 150,0. 114 or 0. 367, respectivley）. More migratory cells attached to the bottom surface of the membrane in experiment group （treated with 0. 1 mg/ml or 1 mg/ml of ITF） in comparison with MTX treated group （P ＜0. 001 ）. Moreover, more migratory cells were found in experimental group treated with 1.0 mg/ml of ITF than those in group treated with 0. 1 mg/ml of ITF （P＜0. 001）. Conclusions Without cell

关 键 词：氨甲喋呤 肠三叶因子 细胞迁移 细胞凋亡 E-钙粘蛋白 

分 类 号：R965[医药卫生—药理学]