World journal of gastroenterology · 2005
Induction of pancreatic duct cells of neonatal rats into insulin-producing cells with fetal bovine serum: a natural protocol and its use for patch clamp experiments.
СтатьяПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод пока не подготовлен
Обработка статьи · этапы, ошибки и управление
Загрузка состояния…
Ошибки и ограничения
История · было → стало
Состояние обновляется каждые 5 секунд, пока блок открыт. Процент показывается только там, где известен объём работы. Пауза останавливает продолжение этапа; текущий запрос к источнику может завершиться.
Leng San-Hua, Lu Fu-Er
Аннотация
AIM: To induce the pancreatic duct cells into endocrine cells with a new natural protocol for electrophysiological study.
METHODS: The pancreatic duct cells of neonatal rats were isolated, cultured and induced into endocrine cells with 15% fetal bovine serum for a period of 20 d. During this period, insulin secretion, MTT value, and morphological change of neonatal and adult pancreatic islet cells were comparatively investigated. Pancreatic beta-cells were identified by morphological and electrophysiological characteristics, while ATP sensitive potassium channels (K(ATP)), voltage-dependent potassium channels (K(V)), and voltage-dependent calcium channels (K(CA)) in beta-cells were identified by patch clamp technique.
RESULTS: After incubation with fetal bovine serum, the neonatal duct cells budded out, changed from duct-like cells into islet clusters. In the first 4 d, MTT value and insulin secretion increased slowly (MTT value from 0.024+/-0.003 to 0.028+/-0.003, insulin secretion from 2.6+/-0.6 to 3.1+/-0.8 mIU/L). Then MTT value and insulin secretion increased quickly from d 5 to d 10 (MTT value from 0.028+/-0.003 to 0.052+/-0.008, insulin secretion from 3.1+/-0.8 to 18.3+/-2.6 mIU/L), then reached high plateau (MTT value >0.052+/-0.008, insulin secretion >18.3+/-2.6 mIU/L). In contrast, for the isolated adult pancreatic islet cells, both insulin release and MTT value were stable in the first 4 d (MTT value from 0.029+/-0.01 to 0.031+/-0.011, insulin secretion from 13.9+/-3.1 to 14.3+/-3.3 mIU/L), but afterwards they reduced gradually (MTT value <0.031+/-0.011, insulin secretion <8.2+/-1.5 mIU/L), and the pancreatic islet cells became dispersed, broken or atrophied correspondingly. The differentiated neonatal cells were identified as pancreatic islet cells by dithizone staining method, and pancreatic beta-cells were further identified by both morphological features and electrophysiological characteristics, i.e. the existence of recording currents from K(ATP), K(V), and K(CA).
CONCLUSION: Islet cells differentiated from neonatal pancreatic duct cells with the new natural protocol are more advantageous in performing patch clamp study over the isolated adult pancreatic islet cells.
METHODS: The pancreatic duct cells of neonatal rats were isolated, cultured and induced into endocrine cells with 15% fetal bovine serum for a period of 20 d. During this period, insulin secretion, MTT value, and morphological change of neonatal and adult pancreatic islet cells were comparatively investigated. Pancreatic beta-cells were identified by morphological and electrophysiological characteristics, while ATP sensitive potassium channels (K(ATP)), voltage-dependent potassium channels (K(V)), and voltage-dependent calcium channels (K(CA)) in beta-cells were identified by patch clamp technique.
RESULTS: After incubation with fetal bovine serum, the neonatal duct cells budded out, changed from duct-like cells into islet clusters. In the first 4 d, MTT value and insulin secretion increased slowly (MTT value from 0.024+/-0.003 to 0.028+/-0.003, insulin secretion from 2.6+/-0.6 to 3.1+/-0.8 mIU/L). Then MTT value and insulin secretion increased quickly from d 5 to d 10 (MTT value from 0.028+/-0.003 to 0.052+/-0.008, insulin secretion from 3.1+/-0.8 to 18.3+/-2.6 mIU/L), then reached high plateau (MTT value >0.052+/-0.008, insulin secretion >18.3+/-2.6 mIU/L). In contrast, for the isolated adult pancreatic islet cells, both insulin release and MTT value were stable in the first 4 d (MTT value from 0.029+/-0.01 to 0.031+/-0.011, insulin secretion from 13.9+/-3.1 to 14.3+/-3.3 mIU/L), but afterwards they reduced gradually (MTT value <0.031+/-0.011, insulin secretion <8.2+/-1.5 mIU/L), and the pancreatic islet cells became dispersed, broken or atrophied correspondingly. The differentiated neonatal cells were identified as pancreatic islet cells by dithizone staining method, and pancreatic beta-cells were further identified by both morphological features and electrophysiological characteristics, i.e. the existence of recording currents from K(ATP), K(V), and K(CA).
CONCLUSION: Islet cells differentiated from neonatal pancreatic duct cells with the new natural protocol are more advantageous in performing patch clamp study over the isolated adult pancreatic islet cells.