High dosage of Exendin-4 increased early insulin secretion in differentiated beta cells from mouse embryonic stem cells

Abstract

Aim: To investigate early insulin release (EIR) and late insulin release (LIR) upon glucose challenge as well as important insulin signaling factors in differentiated insulin-producing cells from embryonic stem cells (ESCs). Methods: A recently published protocol was modified by increasing the concentration of Exendin-4 (from 0.1 nmol/L to 10 nmol/L) together with an additional 5-day culture in low glucose (5.5 mmol/L) medium after differentiation. Gene expression profile, insulin content, C-peptide, EIR and LIR were determined. Results: Compared to a lower concentration of Exendin-4 (0.1 nmol/L), a higher concentration of Exendin-4 (10 nmol/L) increased glucose-responsive insulin secretion, especially EIR. Moreover, 10 nmol/L Exendin-4 increased the expression of the following genes: insulin 1, Pdx-1 (an important transcription factor, newly recognized insulin signaling factors), Epac1 and Epac2 (exchange proteins directly activated by cAMP 1 and 2), and sulfonylurea receptor 1 (SUR1, the subunit of the K ATP channel). Conclusion: According to current knowledge, our modified protocol with a higher concentration of Exendin-4 (10 nmol/L) together with an additional 5-day 5.5 mmol/L glucose culture after differentiation improved the efficiency of differentiation toward the beta cell phenotype, which was possibly the result of stimulated expression of Pdx-1, Epac 1, and Epac 2, which in turn inhibited the K(ATP) channel through combination with SUR1, leading to increased EIR upon glucose challenge.

Exchange protein activated by cAMP 1(Epac1)‐deficient mice develop β‐cell dysfunction and metabolic syndrome

“…Previously, treatment with exendin‐4 (Ex‐4), a dipeptidyl peptidase IV (DPP‐IV)‐resistant glucagon‐like peptide‐1 (GLP‐1) analog, has been shown to promote differentiation of fetal pancreatic tissue, pancreatic progenitors, and intestinal stem cells into insulin‐producing cells and ameliorate hyperglycemia (11, 12). Exogenous treatment of Ex‐4 also leads to increased insulin secretion in β cells differentiated from mouse embryonic stem (ES) cells, with increased expression of insulin‐1, pancreatic and duodenal homeobox 1 (PDX‐1), sulfonylurea receptor 1 [SUR1; a subunit of the ATP‐sensitive K + (K ATP ) channel], Epac1, and Epac2 (13). However, the significance of the increased level of Epac1 and Epac2 expression is not clear.…”

Chorionic and amniotic placental membrane‐derived stem cells, from gestational diabetic women, have distinct insulin secreting cell differentiation capacities

“…To explore induction further, we supplemented the culture medium with additional molecules to improve their maturation: GLP‐1, exendin‐4, and betacellulin. GLP‐1 was incorporated at Stage 3 of our original protocol to promote IPC maturation, whereas exendin‐4 and betacellulin have been widely used to promote the maturation and differentiation of MSCs into functional pancreatic cells (Kumar et al, ; Li, Lam, Xu, Lam, & Chung, ; Wong, ).…”

“…Gene expression was examined at the end of the three-stage differentiation process and increased fold change of each gene calculated by direct comparison to undifferentiated H-/G-CMSCs/AMSCs. With endocrine progenitor markers, NEUROG3 and ILS1, expression was (Kumar et al, 2014;Li, Lam, Xu, Lam, & Chung, 2010;Wong, 2011). (Figure 4f).…”

Stem Cell Therapies for Type I Diabetes

“…Hepatocyte Growth Factor (HGF) is an inducer of cell proliferation that is secreted by mesenchymal cells, and has been used for human ESC-IPC differentiation (Otonkoski, Cirulli et al 1996;D'Amour, Bang et al 2006;Phillips, Hentze et al 2007) as well as murine fetal liver differentiation into IPCs (Feng, Du et al 2005). Exendin-4 stimulates cell proliferation (Xu, Stoffers et al 1999) and it has been used for murine and human ESC-IPC differentiation (Tang, Cao et al 2004;Wu, Liu et al 2007;Gabr, Sobh et al 2008;Gao, Wu et al 2008;Li, Lam et al). A combination of insulin, transferrin, selenium and fibronectin (ITSFn) has been a common cocktail in murine ESC-IPC differentiation, although the presence of insulin in the medium brought some controversy about the endogenous insulin production of IPCs (Lumelsky, Blondel et al 2001;Hori, Rulifson et al 2002;Blyszczuk, Asbrand et al 2004;Hansson, Tonning et al 2004).…”