蛋白激酶B调节骨桥蛋白在肝癌HepG2细胞中的表达

目的 转移相关基因骨桥蛋白(OPN)在肝癌中的表达方式、途径尚不清楚，检测OPN在HepG2细胞中转染蛋白激酶B(Akt)前后的表达，旨在探讨磷脂酰肌醇3激酶信号途径中的关键基因Akt与OPN表达的关系。方法 用脂质体介导的基因转染法将含有Akt基因的质粒转染HepG2细胞，并用Western blot鉴定；OPN的表达用Northern blot和Western blot方法检测。结果 Akt基因成功转染HepG2细胞，Western blot能检测到HepG2细胞中外源表达的Akt基因；Northern blot和Western blot检测发现，Akt在核酸和蛋白水平调节OPN的表达；在无血清培养条件下，OPN在HepG2中结构性表达量很少或无表达,转染活性型Akt后OPN表达升高；在有血清培养条件下,HepG2细胞转染缺陷型Akt基因后OPN表达下降。结论 Akt调节转移相关基因OPN在肝癌细胞中的表达，提示可通过使Akt基因失活来阻断OPN产生，从而抑制肝癌转移。

Osteopontin regulation by protein kinase B in HepG2 cells

Objective It has been unclear how osteopontin, one of metastasis-associated genes, could be regulated in HepG2 cells. The aim of this study was to investigate the effect of protein kinase B(Akt) on the expression of osteopontin in HepG2 cells, and to explore the relationship between Akt and osteopontin expression. Methods HepG2 cells were transfected with constitutively active Akt and dominant negative Akt by lipofectmine, and transfectants were confirmed using Western blot for Akt. Osteopontin expression was detected by Northern blot and Western blot. Results HepG2 cells were successfully transfected with Akt genes including constitutively active Akt and dominant negative Akt, and overexpression of exozegenes Akt could be detected in HepG2 cells by Western blot. Using Northern blot and Western blot, we found that Akt gene regulated osteopontin expression in RNA level and protein level. In serum-free condition, HepG2 cells constitutively expressed low levels of osteopontin. Transfection of constitutively active Akt increased osteopontin mRNA and protein expression. Transfection of dominant negative Akt decreased osteopontin expression.Conclusions The fact that osteopontin gene expression can be regulated by Akt indicates that osteopontin synthesis can be blocked by inactivation of Akt gene and that metastasis of liver cancer might be inhibited by this intervention.