CAJ | 학술논문

目的观察蓝莓对大鼠肝星状细胞(HSC)增殖、活化的影响及其机制.方法采用Ⅳ型胶原酶消化和Nycodenz密度梯度离心的方法,分离、培养大鼠HSC,加入10%蓝莓低剂量及10%蓝莓高剂量动物血清干预体外培养的HSC,并设10%丹芍化纤阳性对照血清组(阳性对照血清组)和10%生理盐水血清对照组.用MTT比色法测定各组HSC的增殖效应,ELISA方法测培养液上清Ⅰ型胶原(Col Ⅰ)的含量,免疫细胞染色法检测各组HSC内α-平滑肌肌动蛋白(α-SMA)的表达,采用Western印迹检测HSC、核转录相关因子(Nrf2)、Ⅰ型血红素氧化酶(HO-1)的表达.结果与生理盐水血清组比较,蓝莓血清组均可显著抑制大鼠HSC的增殖(P＜0.05),其抑制率与阳性对照血清组接近;蓝莓低剂量、高剂量血清组培养液上清Col Ⅰ的含量明显减低(P＜0.05);HSC内α-SMA的表达强度降低(P＜0.05),与阳性对照血清组相当;Western印迹检测显示,与生理盐水血清组比较,蓝莓低剂量、高剂量血清组与阳性对照血清组HSC Nrf2和HO-1蛋白表达增多,以监莓低剂量、高剂量血清组增高更为明显(P＜0.05).结论蓝莓可抑制体外大鼠HSC的增殖和活化,减少细胞外基质的合成,对大鼠肝纤维化有潜在的干预作用,其机制可能与激活HSC Nrf2增加HO-1的表达有关.
목적 관찰람매대대서간성상세포(HSC)증식、활화적영향급기궤제.방법 채용Ⅳ형효원매소화화Nycodenz밀도제도리심적방법,분리、배양대서HSC,가입10%람매저제량급10%람매고제량동물혈청간예체외배양적HSC,병설10%단작화섬양성대조혈청조(양성대조혈청조)화10%생리염수혈청대조조.용MTT비색법측정각조HSC적증식효응,ELISA방법측배양액상청Ⅰ형효원(Col Ⅰ)적함량,면역세포염색법검측각조HSC내α-평활기기동단백(α-SMA)적표체,채용Western인적검측HSC、핵전록상관인자(Nrf2)、Ⅰ형혈홍소양화매(HO-1)적표체.결과 여생리염수혈청조비교,람매혈청조균가현저억제대서HSC적증식(P＜0.05),기억제솔여양성대조혈청조접근;람매저제량、고제량혈청조배양액상청Col Ⅰ적함량명현감저(P＜0.05);HSC내α-SMA적표체강도강저(P＜0.05),여양성대조혈청조상당;Western인적검측현시,여생리염수혈청조비교,람매저제량、고제량혈청조여양성대조혈청조HSC Nrf2화HO-1단백표체증다,이감매저제량、고제량혈청조증고경위명현(P＜0.05).결론 람매가억제체외대서HSC적증식화활화,감소세포외기질적합성,대대서간섬유화유잠재적간예작용,기궤제가능여격활HSC Nrf2증가HO-1적표체유관.
Objective To investigate the effects of blueberry on the proliferation and activation of hepatic stellate cell (HSC) and its mechanism. Methods Rat HSC were isolated by type Ⅳ collagenase digestion and Nycodenz density gradient centrifugation. The cultured HSC was incubated at different concentrations of 10% blueberry serum. The 10% DSHX serum was used as positive control and 10%normal rat serum group as control. MTT colorimetric assay was used to detect the HSC proliferation. Col Ⅰ of culture supernatant was detected by ELISA. The expression of α-SMA in HSC was measured by immunocytochemistry staining while the expressions of Nrf2 and HO-1 were determined by Western blot. Results Compared with controls, the low and high-dose blueberry serum groups significantly inhibited the HSC proliferation (P ＜0. 05 ). It had the same inhibitory effects as the positive control serum group( P ＞0. 05). Col Ⅰ of culture supernatant obviously decreased ( P ＜ 0. 05 ). And the expression levels of α-SMAin low and high-dose blueberry serum groups decreased significantly ( P ＜ 0. 05 ). And there were similareffects in low & high-dose blueberry serum and positive control serum groups. Western blot showed that theexpressions of Nrf2 and HO-1 in blueberry and positive control serum groups were higher than that in control group. And the increment was more significant in the low and high-dose blueberry serum groups( P ＜0.05 ).Conclusion Blueberry can significantly inhibit the proliferation and activation of HSC and reduce the synthesis of extra-cellular matrix. It may have potential preventive and protective effects on hepatic fibrosis.The mechanism may be related to an elevated expression of HO-1 through the Nrf2 pathway.