右美托咪定对结肠癌模型大鼠的治疗作用及对RhoA/ROCK1信号通路的影响

目的:探讨右美托咪定对结肠癌模型大鼠的治疗作用及对Ras同源基因家族成员A(RhoA)/Rho相关卷曲螺旋蛋白激酶1(ROCK1)信号通路的影响。方法:通过1,2-二甲肼诱导建立结肠癌大鼠模型,将成模大鼠随机分为模型组,右美托咪定低(25μg/kg)、中(50μg/kg)和高(100μg/kg)剂量组,每组6只。另取6只未造模大鼠作为正常组。各组大鼠给予相应药物处理4周,观察大鼠的一般状况;检测大鼠血清肿瘤坏死因子α(TNF-α)、白细胞介素1β(IL-1β)水平;解剖观察肿瘤的生长和分布情况;采用苏木精-伊红染色观察病理学变化;检测正常结肠组织和结肠癌组织中RhoA和ROCK1蛋白表达水平。结果:正常组大鼠体重随时间增长,结肠组织结构正常,无炎症细胞浸润;模型组大鼠体重增长缓慢,肛门红肿溃破、大便稀溏,结肠上可见突出肿瘤组织,结肠组织结构异常,炎症细胞浸润程度严重;右美托咪定低、中和高剂量组大鼠体重均高于模型组,肠壁糜烂出血情况有不同程度的减轻,结肠组织结构相比于模型组较为完整,炎症细胞浸润程度有所降低。相较于正常组,模型组大鼠血清TNF-α、IL-1β水平以及结肠癌组织RhoA和ROCK1蛋白表达水平显著升高,差异均有统计学意义(P<0.05);相较于模型组,右美托咪定低、中和高剂量组大鼠血清TNF-α、IL-1β水平以及结肠癌组织RhoA和ROCK1蛋白表达水平显著降低,差异均有统计学意义(P<0.05)。结论:右美托咪定对结肠癌大鼠具有一定的治疗作用,可能与其抑制RhoA/ROCK1通路有关。

Effects of hexahydrocurcumin in combination with 5-fluorouracil on dimethylhydrazine-induced colon cancer in rats

AIM:To investigate the effects of hexahydrocurcumin(HHC),and its combination with 5-fluorouracil(5-FU) on dimethylhydrazine(DMH)-induced colon cancer in rats.METHODS:Male Wistar rats weighing 100-120 g were used as subject models.Aberrant crypt foci(ACF),early preneoplastic lesions of colon cancer,were induced by subcutaneous injection of DHM(40 mg/kg) twice a week for two weeks.After the first DMH injection,rats were treated daily with vehicle(n = 12),curcumin(CUR)(50 mg/kg)(n = 12),HHC(50 mg/kg) orally(n = 12),and treated weekly with an intraperitoneal injection of 5-FU(50 mg/kg)(n = 12),or a combination of 5-FU plus CUR(n = 12) and HHC(n = 12) at the same dosage(s) for 16 wk.The total number of ACF and large ACF were assessed.Cyclooxygenase(COX)-1 and COX-2 expression were detected by immunohistochemistry in colon tissues.The quantitative data of both COX-1 and COX-2 expression were presented as the percentage of number of positive-stained cells to the total number of cells counted.Apoptotic cells in colon tissues were also visualized using the dUTP-biotin nick end labeling method.Apoptotic index(AI) was determined as the percentage of labeled nuclei with respect to the total number of nuclei counted.RESULTS:The total number of ACF was highest in the DMH-vehicle group(1558.20 ± 17.37),however,the number of ACF was significantly reduced by all treatments,5-FU(1231.20 ± 25.62 vs 1558.20 ± 17.37,P < 0.001),CUR(1284.20 ± 25.47 vs 1558.20 ± 17.37,P < 0.001),HHC(1086.80 ± 53.47 vs 1558.20 ± 17.37,P < 0.001),DMH-5-FU + CUR(880.20 ± 13.67 vs 1558.20 ± 17.37,P < 0.001) and DMH-5-FU + HHC(665.80 ± 16.64 vs 1558.20 ± 17.37,P < 0.001).Interestingly,the total number of ACF in the combined treatment groups,the DMH-5-FU + CUR group(880.20 ± 13.67 vs 1231.20 ± 25.62,P < 0.001;880.20 ± 13.67 vs 1284.20 ± 25.47,P < 0.001) and the DMH-5-FU + HHC group(665.80 ± 16.64 vs 1231.20 ± 25.62,P < 0.001;665.80 ± 16.64 vs 1086.80 ± 53.47,P < 0.001) were significantly reduced as compared to 5-FU or each treatment alone.Large ACF were also significantly reduced in all treatment groups,5-FU(111.00 ± 7.88 vs 262.20 ± 10.18,P < 0.001),CUR(178.00 ± 7.33 vs 262.20 ± 10.18,P < 0.001),HHC(186.60 ± 21.51 vs 262.20 ± 10.18,P < 0.001),DMH-5-FU + CUR(122.00 ± 5.94 vs 262.20 ± 10.18,P < 0.001) and DMH-5-FU + HHC(119.00 ± 17.92 vs 262.20 ± 10.18,P < 0.001) when compared to the vehicle group.Furthermore,in the DMH-5-FU + CUR and DMH-5-FU + HHC groups the formation of large ACF was significantly reduced when compared to CUR(122.00 ± 5.94 vs 178.00 ± 7.33,P < 0.005) or HHC treatment alone(119.00 ± 17.92 vs 186.60 ± 21.51,P < 0.001),however,this reduction was not statistically different to 5-FU monotherapy(122.00 ± 5.94 vs 111.00 ± 7.88,P = 0.217;119.00 ± 17.92 vs 111.00 ± 7.88,P = 0.619,respectively).The levels of COX-1 protein after all treatments were not different from normal rats.A marked increase in the expression of COX-2 protein was observed in the DMH-vehicle group.Over-expression of COX-2 was not significantly decreased by 5-FU treatment alone(95.79 ± 1.60 vs 100 ± 0.00,P = 0.198).However,over-expression of COX-2 was significantly suppressed by CUR(77.52 ± 1.68 vs 100 ± 0.00,P < 0.001),HHC(71.33 ± 3.01 vs 100 ± 0.00,P < 0.001),5-FU + CUR(76.25 ± 3.32 vs 100 ± 0.00,P < 0.001) and 5-FU + HHC(68.48 ± 2.24 vs 100 ± 0.00,P < 0.001) in the treated groups compared to the vehicle group.Moreover,CUR(77.52 ± 1.68 vs 95.79 ± 1.60,P < 0.001),HHC(71.33 ± 3.01 vs 95.79 ± 1.60,P < 0.001),5-FU + CUR treatments(76.25 ± 3.32 vs 95.79 ± 1.60,P < 0.001) and 5-FU + HHC(68.48 ± 2.24 vs 95.79 ± 1.60,P < 0.001) markedly decreased COX-2 protein expression more than 5-FU alone.Furthermore,the AI in all treated groups,5-FU(38.86 ± 4.73 vs 23.56 ± 2.12,P = 0.038),CUR(41.78 ± 6.92 vs 23.56 ± 2.12,P < 0.001),HHC(41.06 ± 4.81 vs 23.56 ± 2.12,P < 0.001),5-FU + CUR(49.05 ± 6.75 vs 23.56 ± 2.12,P < 0.001) and 5-FU + HHC(53.69 ± 8.59 vs 23.56 ± 2.12,P < 0.001) significantly increased when compared to the DMH-vehicle group.However,the AI in the combination treatments,5-FU + CUR(49.05 ± 6.75 vs 41.78 ± 6.92,P = 0.192;49.05 ± 6.75 vs 38.86 ± 4.73,P = 0.771) and 5-FU + HHC(53.69 ± 8.59 vs 41.06 ± 4.81,P = 0.379;53.69 ± 8.59 vs 38.86 ± 4.73,P = 0.245) did not reach significant levels as compared with each treatment alone and 5-FU monotherapy,respectively.CONCLUSION:The combined effects of HHC with 5-FU exhibit a synergistic inhibition by decreasing ACF formation mediated by down-regulation of COX-2 expression.