部分肝切除后的肝损伤血清和肝细胞生长因子促进大鼠骨髓细胞表达甲胎蛋白和白蛋白

Serum from partial hepatectomy rat and hepatocyte growth factor stimulate bone marrow cell expressing albumin and alpha fetoprotein

目的 检测大鼠骨髓中是否存在肝脏干细胞,并用肝损伤血清和肝细胞生长因子(HGF)刺激骨髓细胞向肝细胞转化。方法 SD大鼠骨髓单个核细胞分3组进行培养:(1)单纯培养基对照组;(2)肝损伤血清组(15％,血清来自于2-AAF+75％肝切除大鼠);(3)HGF(20ng/ml)组。用甲胎蛋白(AFP)和白蛋白作为细胞标志,用免疫组织化学、逆转录-聚合酶链反应(PCR)、巢式PCR和western blot方法,观察大鼠肝损伤血清和HGF对骨髓细胞转化的促进作用。结果 肝损伤血清组和HGF组培养后10d和20d AFP免疫组织化学和western blot染色阳性;RT-PCR AFP mRNA阳性,新鲜骨髓细胞和单纯IMDM/F12培养基组AFP蛋白和mRNA均阴性。新鲜骨髓细胞存在白蛋白mRNA表达,在肝损伤血清组和HGF组培养后10d和20d,白蛋白mRNA表达增强。结论 大鼠肝损伤血清和HGF可促使体外培养骨髓细胞表达AFP mRNA和AFP。骨髓中可能存在骨髓源性肝干细胞,并微弱表达白蛋白mRNA;肝损伤血清和HGF可以促进骨髓细胞表达白蛋白mRNA。

Objective To explore the effect of serum from partial hepatectomy (PH) rat and hepatocyte growth factor (HGF) on expression of albumin and AFP of bone marrow cells. Methods The bone marrow mono-nucleated cells were separated from SD rats and cultured in three groups: 1. The medium only group as control was added normal fetal bovine serum; 2. Rat hepatic injury serum group (was added 15% rat serum from 2-AAF+PH model); 3. HGF group (HGF 20 ng/ml). The role of these factors was determined by RT-PCR, immunohistochemistry (IHC) and Western blot, using AFP and albumin as special hepatocytic markers. Results By immunohistochemical staining and Western blot, the fresh bone marrow cells were AFP-negative, same as the cells cultured with medium only group. While bone marrow cells, cocultured with rat hepatic injury serum or HGF at day 10 and 20, expressed AFP protein. AFP mRNA expression could be found in bone marrow cells after 10 and 20 days cultured with rat hepatic injury serum or HGF, but not in fresh bone marrow cells and bone marrow cells cultured with medium only. Albumin mRNA expression was weak in fresh bone marrow cell and increased in groups 2 and 3. Conclusion The rat hepatic injury serum or HGF could stimulate the expression of AFP protein and it's mRNA of bone marrow cells. Also they can stimulate albumin mRNA expression. It seems that, in bone marrow, there is a kind of cells so called bone marrow derived liver stem cell which can express albumin mRNA in a weak style.