The role of autophagy in endoplasmic reticulum stress-induced pancreatic ß cell death.

In pancreatic ß-cells, the endoplasmic reticulum (ER) is the crucial site for insulin biosynthesis, as this is where the protein-folding machinery for secretory proteins is localized. Perturbations to ER function of the ß-cell, such as those caused by high levels of free fatty acid and insulin resistance, can lead to an imbalance in protein homeostasis and ER stress, which has been recognized as an important mechanism for type 2 diabetes. Macroautophagy (hereafter referred to as autophagy) is activated as a novel signaling pathway in response to ER stress. In this review, we outline the mechanism of ER stress-mediated ß-cell death and focus on the role of autophagy in ameliorating ER stress. The development of drugs to take advantage of the potential protective effect of autophagy in ER stress, such as glucagon like peptide-1, will be a promising avenue of investigation.

Induction of Galphas contributes to the paradoxical stimulation of cytosolic phospholipase A2alpha expression by cortisol in human amnion fibroblasts.

Cytosolic phospholipase A (cPLA(2alpha)) catalyzes the formation of arachidonic acid in prostaglandin synthesis. In contrast to the well-described down-regulation of cPLA(2alpha), up-regulation of cPLA(2alpha) by glucocorticoids has been reported in human amnion fibroblasts, which may play a key role in parturition. The mechanisms underlying this paradoxical induction of cPLA(2alpha) by glucocorticoids remain largely unknown. Using cultured human amnion fibroblasts, we found that the induction of cPLA(2alpha) by cortisol required ongoing transcription and synthesis of at least one other protein. The induction of cPLA(2alpha) by cortisol was abolished by mutagenesis of a glucocorticoid response element (GRE) in the promoter. The same GRE was found mediating the classical inhibition of cPLA(2alpha) expression by cortisol in human fetal lung fibroblasts (HFL-1). Cortisol increased Galpha(s) expression in amnion fibroblasts but not in HFL-1 cells. Inhibition of Galpha(s) with NF449 attenuated the phosphorylation of cAMP response element-binding protein-1 (CREB-1) and the induction of cPLA(2alpha) by cortisol in amnion fibroblasts. Both glucocorticoid receptor (GR) and CREB-1 were found bound to the GRE upon cortisol stimulation of amnion fibroblasts. The induction of cPLA(2alpha) by cortisol was blocked by GR antagonist RU486 or protein kinase A inhibitor H89 or dominant-negative CREB-1. In conclusion, cortisol activates the cAMP/protein kinase A/CREB-1 pathway via Galpha(s) induction, and the phosphorylated CREB-1 interacts with GR at the GRE to promote cPLA(2alpha) expression in amnion fibroblasts.

Induction of progesterone receptor A form attenuates the induction of cytosolic phospholipase A2alpha expression by cortisol in human amnion fibroblasts.

Cytosolic phospholipase A2alpha (cPLA(2alpha), now known as PLA2G4A) is the enzyme catalyzing the formation of the rate-limiting substrate, arachidonic acid, for prostaglandin (PG) synthesis. The increasing expression of PLA2G4A toward term gestation in human amnion fibroblasts is believed to be the crucial event in parturition. Human amnion fibroblasts produce cortisol, progesterone and express glucocorticoid receptor (GR), progesterone receptor A (PGRA) form at term. The roles of progesterone and PGRA in the induction of PLA2G4A by cortisol via GR in the amnion fibroblasts remain largely unknown. Using cultured human term amnion fibroblasts, we found that cortisol induced the expression of PGRA, which was attenuated by inhibiting PG synthesis with indomethacin. Knockdown of PGRA expression or inhibition of endogenous progesterone production with trilostane significantly enhanced the induction of PLA2G4A by cortisol, whereas overexpression of PGRA attenuated the induction of PLA2G4A by cortisol. Although exogenous progesterone did not alter PLA2G4A expression under basal conditions, it attenuated cortisol-induced PLA2G4A expression at concentrations about tenfold higher, which might be achieved by competition with cortisol for GR. In conclusion, PGRA in the presence of endogenous progesterone is a transdominant repressor of the induction of PLA2G4A by cortisol. High level of progesterone may compete with cortisol for GR, thus further inhibiting the induction of PLA2G4A by cortisol. Moreover, increased PG synthesis by cortisol may feed back on the expression of PGRA leading to attenuation of cortisol-induced PLA2G4A expression. The above findings may be pertinent to the inconsistent effects of glucocorticoids on parturition in humans.

Stimulation of 11ß-HSD1 expression by IL-1ß via a C/EBP binding site in human fetal lung fibroblasts.

Proinflammatory cytokines, just like glucocorticoids (GCs), have been reported to upregulate 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) expression in many cell types. This concerted regulation of 11ß-HSD1 by interleukin-1ß (IL-1ß) and GCs is in marked contrast to their antagonistic effects on inflammation. Further, the molecular mechanisms underlying the induction of 11ß-HSD1 by IL-1ß are not very well understood. In this study, we demonstrated that IL-1ß dramatically stimulated 11ß-HSD1 expression and enzyme activity as well as promoter activity including the -64 bp fragment upstream to the transcription start site in human fetal lung fibroblasts (HFL-1). Nucleotide mutations of the proximal CCAAT box within this region abolished the induction of 11ß-HSD1 promoter activity by IL-1ß. Western blotting analysis demonstrated that IL-1ß induced the expression of C/EBPß dramatically while C/EBPα was barely detectable in HFL-1 cells. Global inhibition of CCAAT/enhancer-binding proteins (C/EBPs) with transfection of C/EBP-specific dominant-negative expression plasmid (CMV500-A-C/EBP) significantly attenuated the induction of 11ß-HSD1 by IL-1ß, whereas over-expression of C/EBPß enhanced the expression of 11ß-HSD1. Chromatin immunoprecipitation assay revealed the recruitment of C/EBPß to the promoter region containing the C/EBP binding site. In conclusion, IL-1ß induces the expression of 11ß-HSD1 mRNA in the fetal lung tissue through mechanisms that involve C/EBPß binding to the promoter. This impact of IL-1ß on the expression of 11ß-HSD1 in human fetal lung cells may explain the alternate mechanism for the lung maturation that appears to occur when there is a risk of premature delivery of the fetus due to the presence of infection.

Expression of 11beta-hydroxysteroid dehydrogenase type 1 in human fetal lung and regulation of its expression by interleukin-1beta and cortisol.

CONTEXT: Glucocorticoids are crucial in fetal lung function. The amount of cortisol available to its receptors is increased by 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1). Glucocorticoids and IL-1beta are known to induce 11beta-HSD1 expression in a number of tissues, but controversial results were obtained with regard to 11beta-HSD1 expression in human fetal lung. OBJECTIVE: We examined the expression of 11beta-HSD1 and its regulation by cortisol and IL-1beta in human fetal lung. RESULTS: Immunohistochemistry revealed 11beta-HSD1 expression in the epithelium and mesenchymal layer of the small bronchus and bronchiole of human fetal lung at 8 months but not at 4 months gestation, which was confirmed by PCR revealing 11beta-HSD1 mRNA expression in the fetal lung tissue. By using a cell line derived from human fetal lung fibroblasts, we demonstrated that cortisol (10(-5) to 10(-3) mmol/liter) or IL-1beta (0.1 to 10 ng/ml) induced 11beta-HSD1 mRNA expression in a concentration-dependent manner. The induction of 11beta-HSD1 by IL-1beta was further increased by cortisol, whereas the induction of cyclooxygenase 2 by IL-1beta was inhibited by cortisol. Nuclear factor kappaB activation inhibitor could only block the induction of cyclooxygenase 2 but not 11beta-HSD1 by IL-1beta, suggesting that different mechanisms were utilized by IL-1beta in the regulation of 11beta-HSD1 versus proinflammatory mediators. Global inhibition of CCAAT-enhancer-binding proteins (C/EBPs) with transfection of C/EBP-specific dominant-negative expression plasmid could attenuate the induction of 11beta-HSD1 by IL-1beta, suggesting that C/EBPs may mediate the induction of 11beta-HSD1 by IL-1beta. CONCLUSIONS: 11beta-HSD1 is expressed in human fetal lung; cortisol and IL-1beta could synergistically induce its expression.