Preliminary Research on Restructuring ATP7B to the Hepatocyte of Wilson Patients
|School||Anhui Medical University,|
|Keywords||Hepatocyte Primary culture PAS ATP7B protein Ad - ATP7B|
Background The gene mutation of human ATP7B protein is the root causes of Wilson disease (liver degeneration), the metabolism of ATP7B protein is in practice by a series of copper-protein, ATP7B gene mutations caused ATP7B protein change ,the copper ions deposit in the body ,and lead to brain and liver pathological damage. consequently as a kind of single gene hereditary disease, liver degeneration of radical method when the most ideal to liver degeneration is gene therapy.Objective To study the expression of ATP7BcDNA in the the hepatocyte of Wilson patients.Obersve the expression of ATP7B protein in various periods.Methods Hepatocyte from Wilson patients were isolated and cultured in vitro, purpose gene ATP7BcDNA was transfection the hepatocyte through viral vectors , by immunocytochemistry and enzyme linked immunosorbent assay, to detect the expression of ATP7B protein in various periods.Results 1. Hepatocyte from Wilson patients were isolated and cultured in vitro successfully,and was proved by PAS.2. The purpose gene ATP7BcDNA was proved successfully transfection into the hepatocyte.3. The synthesis of ATP7B protein was confirmed by immunocytochemistryc(ICC).4.By enzyme linked immunosorbent assay,the expression of ATP7B protein began to increase at 3h,peaked at 3-5d,then decreased gradually after 15days.Conclusin 1. Hepatocyte from Wilson patients were isolated and cultured in vitro successfully,.2. pAdxsi-GFP-ATP7B was constructed successfully.3. The purpose gene ATP7BcDNA was proved successfully transfection into the hepatocyte .4. The synthesis of ATP7B protein was confirmed successfully .