Overexpression of Bcl-x_Lin β-cells prevents cell death but impairs mitochondrial signal for insulin secretion

Abstract

To study effects of Bcl-x(L) in the pancreatic beta-cell, two transgenic lines were produced using different forms of the rat insulin promoter. Bcl-x(L) expression in beta-cells was increased 2- to 3-fold in founder (Fd) 1 and over 10-fold in Fd 2 compared with littermate controls. After exposure to thapsigargin (10 microM for 48 h), losses of cell viability in islets of Fd 1 and Fd 2 Bcl-x(L) transgenic mice were significantly lower than in islets of wild-type mice. Unexpectedly, severe glucose intolerance was observed in Fd 2 but not Fd 1 Bcl-x(L) mice. Pancreatic insulin content and islet morphology were not different from control in either transgenic line. However, Fd 2 Bcl-x(L) islets had impaired insulin secretory and intracellular free Ca(2+) ([Ca(2+)](i)) responses to glucose and KCl. Furthermore, insulin and [Ca(2+)](i) responses to pyruvate methyl ester (PME) were similarly reduced as glucose in Fd 2 Bcl-x(L) islets. Consistent with a mitochondrial defect, glucose oxidation, but not glycolysis, was significantly lower in Fd 2 Bcl-x(L) islets than in wild-type islets. Glucose-, PME-, and alpha-ketoisocaproate-induced hyperpolarization of mitochondrial membrane potential, NAD(P)H, and ATP production were also significantly reduced in Fd 2 Bcl-x(L) islets. Thus, although Bcl-x(L) promotes beta-cell survival, high levels of expression of Bcl-x(L) result in reduced glucose-induced insulin secretion and hyperglycemia due to a defect in mitochondrial nutrient metabolism and signaling for insulin secretion.

Mcl-1 downregulation by pro-inflammatory cytokines and palmitate is an early event contributing to β-cell apoptosis

“…3,[5][6][7]13,18,32 Here, we show that cytokines, palmitate and thapsigargin, induce Bax translocation, cytochrome c release and caspase-3 cleavage, independently of changes in Bcl-2, Bcl-XL, Bax and Bak expression levels, but partly as a consequence of Mcl-1 downregulation (Figure 8). Our data are in agreement with previous studies showing that Mcl-1, in spite of its preferential mitochondrial localization, 33 is able to prevent Bax activation and translocation.…”

“…Bcl-XL overexpression improves b-cell survival, but impairs glucose oxidation and GSIS, 7 and the BH3-only protein BAD is also an important modulator of GSIS, 41 suggesting that some members of the Bcl-2 family have 'day' and 'night' jobs in b-cells, affecting both cell survival and function. 42 This is apparently not the case for Mcl-1 as neither Mcl-1 knockdown nor overexpression altered GSIS.…”

“…4 The Bax/Bak activation is regulated by a delicate equilibrium between antiapoptotic Bcl-2 proteins and pro-apoptotic BH3-only proteins. Among the antiapoptotic proteins, overexpression of Bcl-2 and Bcl-XL have been shown to prevent b-cell apoptosis in a variety of models, [5][6][7] but they may hamper b-cell function. 7 The molecular role of Mcl-1 in b-cells remains to be characterized.…”

“…Among the antiapoptotic proteins, overexpression of Bcl-2 and Bcl-XL have been shown to prevent b-cell apoptosis in a variety of models, [5][6][7] but they may hamper b-cell function. 7 The molecular role of Mcl-1 in b-cells remains to be characterized. Mcl-1 is structurally unique among the Bcl-2 family; it contains two PEST sequences (polypeptide sequences enriched in proline (P), glutamic acid (E), serine (S) and threonine (T)), which favor Mcl-1 degradation, thereby reducing its half-life.…”

Dual role of proapoptotic BAD in insulin secretion and beta cell survival

“…These findings are consistent with the observation that loss of BAD in beta cells does not entirely phenocopy BCL-X L overexpression. Transgenic islets expressing BCL-X L had a general secretory impairment, including a lack of response to KCl, tolbutamide and KIC 36 . This is consistent with the function of BCL-X L at the endoplasmic reticulum and probably reflects perturbations in Ca 2+ homeostasis associated with BCL-X L overexpression 4 .…”