Protective effect of intestinal trefoil factor on methotrexate-induced injury in intestinal mucosa / 中华消化杂志

ABSTRACT

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 proliferation, the protective effect of ITF is related to its functions of promoting cell migration and inhibiting cell apoptosis, which may down-regulate expression of E-cadherin mRNA.