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【目的】观察苦杏仁苷对血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)刺激下HSC-T6细胞Ca2+效应的抑制作用。【方法】将HSC-T6细胞系分为以下5组:正常组、AngⅡ组、苦杏仁苷高剂量(10-3 mol/L)组、苦杏仁苷中剂量(10-4 mol/L)组、苦杏仁苷低剂量(10-5 mol/L)组。正常组采用含体积分数2%胎牛血清的DMEM培养,其余各组在DMEM培养液的基础上,AngⅡ组加入10-7 mol/L AngⅡ溶液;高、中、低剂量苦杏仁苷组均加入10-7 mol/L AngⅡ溶液,在此基础上各组分别加入不同浓度的苦杏仁苷。钙荧光探针Fluo-3/AM负载后置于LSCM SP2激光共聚焦显微镜(LSCM)下检测Ca2+。【结果】LSCM下Ca2+荧光成像的HSC-T6细胞仍可部分表现倒置显微镜下的形态学特征,各组细胞均染色良好,且各细胞间染色强度基本一致。AngⅡ组Ca2+荧光强度非常明显,清晰可见;正常组强度也较明显,稍次于AngⅡ组;苦杏仁苷高剂量组强度也较强,稍次于AngⅡ组和正常组,但区别不甚明显;苦杏仁苷中、低剂量组强度明显减弱,并可见大量Ca2+噬斑出现,其中中剂量组图像噬斑更为清晰。AngⅡ组荧光强度较正常组稍强,但差异无统计学意义(P>0.05)。苦杏仁苷高剂量荧光强度稍低于AngⅡ组与正常组,但差异均无统计学意义(P>0.05);苦杏仁苷中、低剂量组荧光强度显著低于AngⅡ组和正常组,差异均有统计学意义(P<0.05)。【结论】苦杏仁苷可抑制HSC内Ca2+升高,从而抑制HSC的活化与增殖,产生抗肝纤维化作用。
【Objective】 To observe the inhibitory effect of amygdaloside on Ca2 + in HSC-T6 cells stimulated by angiotensinⅡ (AngⅡ). 【Methods】 HSC-T6 cells were divided into the following 5 groups: normal group, AngⅡgroup, amygdalin high dose (10-3 mol / L) group, amygdalin medium dose (10-4 mol / L) , Amygdalin low dose (10-5 mol / L) group. The normal group was cultured in DMEM containing 2% fetal bovine serum. The other groups were added with 10-7 mol / L AngⅡ solution on the basis of DMEM culture medium. The high, middle and low doses of amygdalin were added 10-7 mol / L AngⅡ solution, on the basis of each group were added different concentrations of amygdalin. Calcium fluorescence probe Fluo-3 / AM was placed in LSCM SP2 confocal laser scanning microscopy (LSCM) detection of Ca2 +. 【Results】 HSC-T6 cells stained by Ca2 + fluorescence in LSCM can still partially reflect the morphological features under inverted microscope. The cells in each group stained well, and the staining intensities of all cells were basically the same. Ang Ⅱ group Ca2 + fluorescence intensity is very clear, clearly visible; the intensity of the normal group is also more obvious, slightly inferior to the Ang Ⅱ group; amygdalin high dose group intensity is also strong, slightly inferior to Ang Ⅱ group and normal group, but the difference is not obvious; In amygdalin, the intensity of low dose group was significantly weakened, and a large number of Ca2 + plaques appeared, of which the middle dose group image plaque was more clear. The fluorescence intensity of Ang Ⅱ group was slightly stronger than that of normal group, but the difference was not statistically significant (P> 0.05). The high-dose fluorescence intensity of amygdalin was slightly lower than that of AngⅡgroup and normal group, but the difference was not statistically significant (P> 0.05). The fluorescence intensity of amygdalin in low-dose group was significantly lower than that in AngⅡgroup and normal group There was statistical significance (P <0.05). 【Conclusion】 Amygdalin can inhibit the increase of intracellular Ca2 + in HSC, thereby inhibiting the activation and proliferation of HSC and producing anti-hepatic fibrosis.