摘要
目的观察胰腺应激蛋白PSP/reg对胰腺星状细胞(PSC)合成和分泌基质金属蛋白酶(MMPs)及其组织抑制剂(TIMPs)以及RECK表达的影响。方法分离纯化慢性胰腺炎患者纤维化区的PSC,基因重组胰腺应激蛋白PSP/reg,以终浓度为10和100 ng/mL对PSC进行干预,实时荧光定量PCR检测MMP1/2、TIMP1/2及RECK基因表达,Western blot测定MMP1/2、TIMP1/2及RECK蛋白,细胞免疫荧光观察细胞膜表面RECK分布。结果 PSP/reg对MMP1/2、TIMP1/2及RECK表达无明显影响;PSP/reg轻度抑制PSC培养上清中MMP2水平(P<0.05),而显著抑制TIMP1/2水平(P<0.01);PSC细胞膜表面发现有RECK蛋白,PSP/reg减少PSC的RECK含量(P<0.01)。结论胰腺应激蛋白PSP/reg能够降低TIMPs:MMPs比率、减少RECK蛋白水平表达,从而解除对MMPs的部分抑制,使MMPs活性相对增高,有利于纤维化的分解消散,促进胰腺损伤后的再生修复。
Objective To observe the effects of pancreatic stress protein (PSP/reg) on TIMPs : MMPs secretion and RECK expression of pancreatic stellate cell (PSC). Method PSC were obtained by outgrowth from fibrotic human pancreas tissue. PSP/reg was expressed in the yeast Pichia pastoris. PSP/reg was added at concentration of 10 and 100 ng/mL to cultured PSC. MMP1/2, TIMP1/2 and RECK mRNA level were determined by real-time quantitative RT-PCR. Their protein level were determined by Western blot. RECK was detected on the cell surface of PSC by immunofluorescence microscopy. Result No changes of MMP1/2, TIMP1/2 and RECK mRNA expressions were observed after PSP/reg application. PSP/reg slightly decreased the synthesis of MMP2 ( P 〈 0.05 ), while strongly decreased TIMP1/2 concentrations (P 〈 0.01 ) in PSC supernatants. Immunofluorescence staining showed RECK on the surface of human PSC. PSP/reg decreased RECK expression ( P 〈 0.01 ). Conclusion PSP/reg is upregulated significantly during acute and chronic pancreatitis. PSC plays a key role in fibrogenesis associated with pancreatitis. By decreasing TIMPs:MMPs ratio and RECK expression of PSC, PSP/reg may enhance MMPs activity and promote fibrolysis in the repair phase of acute and chronic pancreatitis.
出处
《中国微生态学杂志》
CAS
CSCD
2011年第12期1077-1081,共5页
Chinese Journal of Microecology
基金
国家自然科学基金项目(30900696
30971399
81170716)
江苏省自然科学基金项目(BK2009278)