Ya-Wun Yang1, Chih-Kai Chao. 1. School of Pharmacy, College of Medicine, National Taiwan University, 1 Jen-Ai Road, Section 1, Taipei 100, Taiwan. ywyang@ha.mc.ntu.edu.tw
Abstract
BACKGROUND: Increased efficiency of transgene expression is desired for virus-mediated gene delivery. In the present study, we examined the effect of calcium phosphate (CaPi) on recombinant adeno-associated virus (rAAV)-mediated insulin therapy in diabetic animals. METHODS: The rAAV vector, rAAV.PEPCK.Ins.EGFP, containing the human insulin gene under control of the phosphoenolpyruvate carboxykinase (PEPCK) promoter and the enhanced green fluorescence protein (EGFP) gene driven by the cytomegalovirus (CMV) IE promoter, was employed in this study. C57BL/6J mice were made diabetic with streptozotocin (STZ), followed by injection into the livers with either rAAV alone, or noncovalent complexes with calcium phosphate. Body weight and blood glucose levels of the animals were routinely monitored after 6 h fasting. Secretion of human insulin in the rAAV-transduced animals was determined by radioimmunoassay (RIA). Expression of human insulin in the livers of the animals was detected by immunohistochemical staining. RESULTS: Compared with the STZ-treated control mice, administration of rAAV containing the human insulin gene significantly decreased blood glucose levels and maintained body weight of the diabetic animals. Complexation of rAAV with calcium phosphate enhanced the hypoglycemic effect of rAAV-mediated gene transfer. Results obtained from both RIA and immunohistochemical staining demonstrated that incorporation of calcium phosphate enhanced rAAV-mediated gene transfer in vivo, leading to higher expression and secretion of human insulin. CONCLUSIONS: Administration of rAAV harboring the human insulin gene into livers of the STZ-diabetic mice improved blood glucose levels, maintained body weight of the diabetic animals, and resulted in human insulin secretion. Complexation of rAAV with calcium phosphate significantly potentiated the efficiency of rAAV-mediated diabetic gene therapy. Copyright 2002 John Wiley & Sons, Ltd.
BACKGROUND: Increased efficiency of transgene expression is desired for virus-mediated gene delivery. In the present study, we examined the effect of calcium phosphate (CaPi) on recombinant adeno-associated virus (rAAV)-mediated insulin therapy in diabetic animals. METHODS: The rAAV vector, rAAV.PEPCK.Ins.EGFP, containing the humaninsulin gene under control of the phosphoenolpyruvate carboxykinase (PEPCK) promoter and the enhanced green fluorescence protein (EGFP) gene driven by the cytomegalovirus (CMV) IE promoter, was employed in this study. C57BL/6J mice were made diabetic with streptozotocin (STZ), followed by injection into the livers with either rAAV alone, or noncovalent complexes with calcium phosphate. Body weight and blood glucose levels of the animals were routinely monitored after 6 h fasting. Secretion of humaninsulin in the rAAV-transduced animals was determined by radioimmunoassay (RIA). Expression of humaninsulin in the livers of the animals was detected by immunohistochemical staining. RESULTS: Compared with the STZ-treated control mice, administration of rAAV containing the humaninsulin gene significantly decreased blood glucose levels and maintained body weight of the diabetic animals. Complexation of rAAV with calcium phosphate enhanced the hypoglycemic effect of rAAV-mediated gene transfer. Results obtained from both RIA and immunohistochemical staining demonstrated that incorporation of calcium phosphate enhanced rAAV-mediated gene transfer in vivo, leading to higher expression and secretion of humaninsulin. CONCLUSIONS: Administration of rAAV harboring the humaninsulin gene into livers of the STZ-diabeticmice improved blood glucose levels, maintained body weight of the diabetic animals, and resulted in humaninsulin secretion. Complexation of rAAV with calcium phosphate significantly potentiated the efficiency of rAAV-mediated diabetic gene therapy. Copyright 2002 John Wiley & Sons, Ltd.
Authors: Maria Luisa Jaén; Laia Vilà; Ivet Elias; Veronica Jimenez; Jordi Rodó; Luca Maggioni; Rafael Ruiz-de Gopegui; Miguel Garcia; Sergio Muñoz; David Callejas; Eduard Ayuso; Tura Ferré; Iris Grifoll; Anna Andaluz; Jesus Ruberte; Virginia Haurigot; Fatima Bosch Journal: Mol Ther Methods Clin Dev Date: 2017-04-05 Impact factor: 6.698