GR-independent down-modulation on GM-CSF bone marrow-derived dendritic cells by the selective glucocorticoid receptor modulator Compound A – Scientific Reports – 2016 (http://dx.doi.org/10.1038/srep36646)
Abstract: Dendritic cells (DC) initiate the adaptive immune response. Glucocorticoids (GCs) down-modulate the function of DC. Compound A (CpdA, (2-(4-acetoxyphenyl)-2-chloro-N-methyl-ethylammonium chloride) is a plant-derived GR-ligand with marked dissociative properties. We investigated the effects of CpdA on in vitro generated GM-CSF-conditioned bone marrow-derived DC (BMDC). CpdA-exposed BMDC exhibited low expression of cell-surface molecules and diminution of the release of proinflammatory cytokines upon LPS stimulation; processes associated with BMDC maturation and activation. CpdA-treated BMDC were inefficient at Ag capture via mannose receptor-mediated endocytosis and displayed reduced T-cell priming. CpdA prevented the LPS-induced rise in pErk1/2 and pP38, kinases involved in TLR4 signaling. CpdA fully inhibited LPS-induced pAkt Ser473, a marker associated with the generation of tolerogenic DC. We used pharmacological blockade and selective genetic loss-of-function tools and demonstrated GR-independent inhibitory effects of CpdA in BMDC. Mechanistically, CpdA-mediated inactivation of the NF-κB intracellular signaling pathway was associated with a short-circuiting of pErk1/2 and pP38 upstream signaling. Assessment of the in vivo function of CpdA-treated BMDC pulsed with the hapten trinitrobenzenesulfonic acid showed impaired cell-mediated contact hypersensitivity. Collectively, we provide evidence that CpdA is an effective BMDC modulator that might have a benefit for immune disorders, even when GR is not directly targeted.
VACUOLE MEMBRANE PROTEIN 1 EXPRESSION IN BETA CELLS DURING NATURALLY OCCURRING DIABETES IN NOD MICE – Pancreatology – 2017
Abstract: Type 1 diabetes is an autoimmune disease in which insulin production is compromised due to dysfunction of B-cells by means of immune cells infiltrating islets (insulitis). We found that in response to experimentally-induced diabetes the autophagy pathway was activated to regulate cellular homeostasis. VMP1 is an autophagy-related protein that was found elevated during the course of several pathologies including diabetes. Based on this observation the Aim: of this work was to determine whether VMP1 is involved in the process of B-cell demise in the naturally progress of disease employing the non-obese diabetic (NOD) mice model of diabetes. Methods: NOD mice (4-28wk); immune-fluorescence microscopy. Results: Insulitis severity increased with age in female NOD mice while the intensity of immune-reactive insulin, assessed by IF, decreased. VMP1 co-expressed with insulin, and its expression was mainly detected in pancreatic B-cells. Local production of inflammatory cytokines during insulitis triggers B-cell death. Mimicking this process in vitro, we challenged the rat insulinoma INS-1E and isolated mouse islets with IL-1B+IFN-g (CYT). We detected an increase in the expression of VMP1 in both INS-1E cells and B-cells within islets after CYT exposure resembling the findings in NOD islets. Discussion: VMP1 expression is induced by proinflammatory cytokines during the process of B-cell apoptosis during autoimmune diabetes. Experiments are being carried out to unravel the mechanistic action of VMP1 in B-cells under autoimmunity in diabetes.
A selective glucocorticoid receptor modulator (CpdA) attenuates cytokine-induced endoplasmic reticulum stress in ß-cells – Pancreatology – 2017
Abstract: Introduction: Type 1 diabetes (T1D) is a T cell-mediated autoimmune disease that selectively destroys insulin producing ß-cells. ER stress and subsequent insulin secretory deficiency in ß-cells precede the onset of autoimmune diabetes. Pro-inflammatory cytokines (IL-1ß+IFN-g; CYT) signaling leads to activation of ER stress in ß-cells. We reported that Compound A (2-(4-acetoxyphenyl)-2-chloro-N-methyl-ethylammonium chloride; CpdA), a dissociative glucocorticoid receptor (GR)-ligand, is an effective modulator of T and dendritic cells. Aim: The aim of this study was to explore the beneficial effects of CpdA on cytokine-induced ß-cell ER stress. Results: CpdA significantly reduced CYT-induced NO secretion by INS-1E cells (p<0.05). CpdA treatment impaired eIF2a; phosphorylation enhancement and the increase of ER stress protein markers expression, such as ATF4 and CHOP, in CYT-challenged INS-1E cells (p<0.05). The expression of chaperones involved in protein folding and processing (PDI, ORP150) was enhanced in the presence of CpdA (p<0.05). CpdA administration (i.p.) to NODscid mice adoptively transferred with diabetogenic splenocytes (from diabetic NOD mice) led to a delay of disease onset (p<0.001 vs control). CpdA-treated mice showed a reduction in islet leukocytes infiltration and preserved insulin expression in comparison with veh-treated group, assessed by immunohistochemistry. Discussion: In summary, we demonstrate that CpdA directly improves the UPR attenuating ER stress in ß-cells challenged by CYT. The latter together with our previous report on immune cells modulation, might warrant the administration of CpdA as a novel therapeutic with dual activity on autoimmune diabetes.
