Copper Transport Function of Mutant-type and Wild-type ATP7B in Skin Fibroblasts

Objective: Mutant-type and Wild-type ATP7B were transfected and expressed in skin fibroblasts Me32aT22/2L cell lines. Copper transport function of mutant-type and wild-type ATP7B were studied and made the basement for further gene therapy of hepatolenticular degeneration. Methods: pRc/CMV-WD containing cDNA of mutant-type or wild-type ATP7B was transfected into Me32aT22/2L cell using liposome transfection methods,respectively. Intracellular distribution of mutant-type and wild-type ATP7B were observed by immunofluorescence histochemistry methods and copper transport function was studied in copper incubation experiments. Results: Expression of ATB7B gene was detected and located around the nuclei in Me32aT22/2L cell. At 24 and 48 hours,in wild-type ATP7B group,copper/protein value is 335.33±49.86 and 477.38±30.95,respectively,however,606.14± 45.72 vs 901.84±53.18 in mutant-type ATP7B group. Significant statistical difference could be obtained (P 0.05). Conclusion: Wild-type ATP7B could transport intracellular redundant copper and mutant-type ATP7B lost this function. Mutant R778L gene is a pathogenic gene of hepatolenticular degeneration.