肉桂醛调节Shh/Gli1信号通路对幽门螺旋杆菌胃炎大鼠胃黏膜损伤的影响

Effect of cinnamaldehyde on gastric mucosal damage in rats with Helicobacter pylori gastritis by regulating the Shh/Gli1 signaling pathway

目的 探讨肉桂醛调节音猬因子(Shh)/Gli家族锌指蛋白1(Gli1)信号通路对幽门螺旋杆菌(Hp)胃炎大鼠胃黏膜损伤的影响。方法 将大鼠随机分为对照组、模型组、肉桂醛低剂量组、肉桂醛中剂量组、肉桂醛高剂量组、替普瑞酮组、肉桂醛高剂量+Shh激活剂Purmorphamine(PUR)组,每组18只。通过灌胃Hp的方法构建胃炎大鼠模型。建模成功后,肉桂醛低、中、高剂量组大鼠分别灌胃12.5 mg/kg、25 mg/kg、50 mg/kg肉桂醛,替普瑞酮组大鼠灌胃0.13 g/kg替普瑞酮,肉桂醛高剂量+PUR组大鼠给予50 mg/kg肉桂醛灌胃和6.67 mg/kg PUR腹腔注射,每天给药1次,持续2周。HE染色检测大鼠胃黏膜组织病理变化;透射电镜观察大鼠胃黏膜细胞形态。将GES-1细胞分为vitro-对照组、vitro-模型组、vitro-肉桂醛低剂量组、vitro-肉桂醛中剂量组、vitro-肉桂醛高剂量组、vitro-替普瑞酮组、vitro-肉桂醛高剂量+PUR组。除vitro-对照组外,其他组GES-1细胞均需被Hp感染,感染结束后,vitro-肉桂醛低剂量组、vitro-肉桂醛中剂量组、vitro-肉桂醛高剂量组分别用5μmol/L、10μmol/L、20μmol/L肉桂醛处理24 h;vitro-替普瑞酮组用1μmol/L替普瑞酮处理24 h;vitro-肉桂醛高剂量+PUR组用20μmol/L肉桂醛和1μmol/L PUR共同处理24 h,CCK-8法检测GES-1细胞增殖抑制率;ELISA法检测大鼠胃黏膜组织及GES-1细胞上清中白细胞介素1β (IL-1β)、肿瘤坏死因子-α(TNF-α)水平;Western blot检测大鼠胃黏膜组织及GES-1细胞中Shh、Gli1蛋白表达。结果 与对照组比较,模型组大鼠胃黏膜组织病理损伤严重,胃黏膜表面上皮细胞固缩,细胞膜破损,IL-1β、TNF-α水平及Shh、Gli1蛋白表达升高(P<0.05);与模型组比较,肉桂醛低剂量组、肉桂醛中剂量组、肉桂醛高剂量组、替普瑞酮组大鼠胃黏膜组织病理损伤、胃黏膜表面上皮细胞结构及形态有所改善,IL-1β、TNF-α水平及Shh、Gli1蛋白表达降低(P<0.05);与肉桂醛高剂量组比较,肉桂醛高剂量+PUR组大鼠胃黏膜组织病理损伤加剧,胃黏膜表面上皮细胞结构及形态破环严重,IL-1β、TNF-α水平及Shh、Gli1蛋白表达升高(P<0.05)。与vitro-对照组比较,vitro-模型组GES-1细胞增殖抑制率、细胞上清中IL-1β、TNF-α水平及细胞中Shh、Gli1蛋白表达升高(P<0.05);与vitro-模型组比较,vitro-肉桂醛低剂量组、vitro-肉桂醛中剂量组、vitro-肉桂醛高剂量组、vitro-替普瑞酮组GES-1细胞增殖抑制率、细胞上清中IL-1β、TNF-α水平及细胞中Shh、Gli1蛋白表达降低(P<0.05);与vitro-肉桂醛高剂量组比较,vitro-肉桂醛高剂量+PUR组GES-1细胞增殖抑制率、细胞上清中IL-1β、TNF-α水平及细胞中Shh、Gli1蛋白表达升高(P<0.05)。结论 肉桂醛可能通过抑制Shh/Gli1通路减轻Hp诱导的胃炎大鼠胃黏膜损伤。

Objective To investigate the effect of cinnamaldehyde on gastric mucosal damage in rats with Helicobacter pylori (Hp) gastritis by regulating the sonic hedgehog (Shh)/Gli family zinc finger protein 1 (Gli1) signaling pathway.Methods The rats were randomly grouped into the control group,model group,low-dose cinnamaldehyde group,medium-dose cinnamaldehyde group,high-dose cinnamaldehyde group,teprenone group,and high-dose cinnamaldehyde+Shh activator Purmorphamine (PUR) group,with 18 rats in each group.The rat model of gastritis was established by intragastric Hp.After successful modeling,the rats in the low,medium and high dose groups were given12.5 mg/kg,25 mg/kg and 50 mg/kg cinnamaldehyde,and the rats in the tepredone group were given 0.13 g/kg tepredone;rats in the high dose of cinnamaldehyde+PUR group were given 50 mg/kg cinnamaldehyde by gavage and 6.67 mg/kg PUR by intraperitoneal injection,once a day for two weeks.HE staining was applied to detect the pathological changes in gastric mucosa tissue;transmission electron microscopy was applied to observe the morphology of gastric mucosal cells in rats.GES-1 cells were separated into vitro-control group,vitro-model group,vitrolow-dose cinnamaldehyde group,vitro-medium-dose cinnamaldehyde group,vitro-high-dose cinnamaldehyde group,vitro-teprenone group,and vitro-high-dose cinnamaldehyde+PUR group.Except for the vitro-control group,GES-1 cells in other groups were infected with Hp,after the infection ended,vitro-low-dose group,vitro-medium-dose group and vitro-high-dose group were treated with 5μmol/L,10μmol/L and20μmol/L cinnamaldehyde for 24 h,respectively;vitro-tepredone group was treated with 1μmol/L tepredone for 24 h;vitro-high-dose cinnamaldehyde+PUR group was treated with 20μmol/L cinnamaldehyde and 1μmol/L PUR for 24 hours;CCK-8 method was applied to detect the inhibition rate of proliferation of GES-1 cells;ELISA method was applied to detect the levels of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in gastric mucosal tissue and GES-1 cell supernatant;Western blot was applied to detect the expression of Shh and Gli1 proteins in gastric mucosal tissue and GES-1 cells.Results Compared with the control group,the pathological damage of the gastric mucosal tissue in the model group was severe,with pyknosis of epithelial cells on the surface of the gastric mucosa and damage to the cell membrane,and the levels of IL-1β,TNF-α,and the expression of Shh and Gli1 proteins increased (P<0.05);compared with the model group,the pathological damage of gastric mucosal tissue,the structure and morphology of gastric mucosal surface epithelial cells were improved in the the low-dose cinnamaldehyde group,the medium-dose cinnamaldehyde group,the high-dose cinnamaldehyde group,and teprenone group,and the levels of IL-1β,TNF-α,and the expression of Shh and Gli1 proteins decreased (P<0.05);compared with the high-dose cinnamaldehyde group,the pathological damage to the gastric mucosal tissue in the high-dose cinnamaldehyde+PUR group was exacerbated,and the structure and morphology of the gastric mucosal surface epithelial cells were severely disrupted,and the levels of IL-1β,TNF-α,and the expression of Shh and Gli1 proteins increased (P<0.05).Compared with the vitro-control group,the inhibition rate of GES-1 cell proliferation,the levels of IL-1βand TNF-α in the cell supernatant,and the expression of Shh and Gli1 proteins in the cells in vitro-model group increased (P<0.05);compared with the vitro-model group,the inhibition rate of GES-1 cell proliferation,the levels of IL-1β and TNF-α in the cell supernatant,and the expression of Shh and Gli1 proteins in the cells in vitro-low-dose cinnamaldehyde group,the vitro-medium-dose cinnamaldehyde group,the vitro-high-dose cinnamaldehyde group,and vitro-teprenone group decreased (P<0.05);compared with the vitro-high-dose cinnamaldehyde group,the inhibition rate of GES-1 cell proliferation,the levels of IL-1β and TNF-α in the cell supernatant,and the expression of Shh and Gli1 proteins in the cells in vitro-high-dose cinnamaldehyde+PUR group increased (P<0.05).Conclusion Cinnamaldehyde may alleviate gastric mucosal damage induced by Hp in rats with gastritis by inhibiting the Shh/Gli1 pathway.