M Shimoda1, S Chen, H Noguchi, S Matsumoto, P A Grayburn. 1. Division of Cardiology, Department of Internal Medicine, Baylor University Medical Center, Baylor Heart and Vascular Institute, Dallas, Texas, USA.
Abstract
BACKGROUND: It has been reported that the human pancreatic nonendocrine fraction, which remains after islet isolation, can be differentiated toward beta cells. However, the optimal method to accomplish this goal has not been established. In this study, we introduced the human neurogenic differentiation 1 (NeuroD1) gene into human nonendocrine pancreatic epithelial cells (NEPECs) and promoted insulin-producing cells in vitro. METHODS: The human pancreatic nonislet fractions were obtained from brain-dead donors and cultured in suspension for 2-3 days followed by culture with G418 for 4 days. These cells (NEPECs) were then plated on dishes. The NEPECs spread into a cell monolayer within 7 days and all of the cells were cytokeratin-19 (CK19) positive. Seven days after plating, plasmids encoding human NeuroD1 gene under human CK19 promoter were transfected 3 times every other day (termed NEPEC+ND). Seven days after starting induction, these cells were characterized. RESULTS: Seven days after starting the induction of human NeuroD1, NEPEC+ND strongly expressed NeuroD1 and insulin mRNA. The ratio of NeuroD1-positive cells in NEPEC+ND was significantly higher than in NEPEC. Human insulin-positive cells in NEPEC+ND were also significantly greater than in NEPEC. Human insulin and C-peptide levels in culture medium in NEPEC+ND were significantly higher than in NEPEC. CONCLUSIONS: These findings demonstrated that human NeuroD1 under control of the CK19 promoter can induce the differentiation of CK19-positive NEPECs into insulin-producing cells. Copyright 2010 Elsevier Inc. All rights reserved.
BACKGROUND: It has been reported that the humanpancreatic nonendocrine fraction, which remains after islet isolation, can be differentiated toward beta cells. However, the optimal method to accomplish this goal has not been established. In this study, we introduced the humanneurogenic differentiation 1 (NeuroD1) gene into human nonendocrine pancreatic epithelial cells (NEPECs) and promoted insulin-producing cells in vitro. METHODS: The humanpancreatic nonislet fractions were obtained from brain-dead donors and cultured in suspension for 2-3 days followed by culture with G418 for 4 days. These cells (NEPECs) were then plated on dishes. The NEPECs spread into a cell monolayer within 7 days and all of the cells were cytokeratin-19 (CK19) positive. Seven days after plating, plasmids encoding humanNeuroD1 gene under humanCK19 promoter were transfected 3 times every other day (termed NEPEC+ND). Seven days after starting induction, these cells were characterized. RESULTS: Seven days after starting the induction of humanNeuroD1, NEPEC+ND strongly expressed NeuroD1 and insulin mRNA. The ratio of NeuroD1-positive cells in NEPEC+ND was significantly higher than in NEPEC. Humaninsulin-positive cells in NEPEC+ND were also significantly greater than in NEPEC. Humaninsulin and C-peptide levels in culture medium in NEPEC+ND were significantly higher than in NEPEC. CONCLUSIONS: These findings demonstrated that humanNeuroD1 under control of the CK19 promoter can induce the differentiation of CK19-positive NEPECs into insulin-producing cells. Copyright 2010 Elsevier Inc. All rights reserved.
Authors: Shruti Mohan; Ryan Lafferty; Neil Tanday; Peter R Flatt; R Charlotte Moffett; Nigel Irwin Journal: PLoS One Date: 2021-12-20 Impact factor: 3.240