Modulation of the circadian clock by glucocorticoid receptor isoforms in the H295R cell line.

Peripheral clocks are set by different nervous, hormonal and metabolic stimuli, and regulate the circadian expression of several genes. We investigated whether a peripheral clock could be induced in the human adrenocortical cell line H295R and whether glucocorticoid receptor isoforms (GRα and GRß) are involved in this clock system. After synchronization of cells with serum shock, the rhythmic oscillation of clock genes PER1, PER2, REV-ERBα, and ARNTL was confirmed. In addition, H295R cells even without serum shock showed rhythmic expression of PER1, PER2, CRY1 and ARNTL. Glucocorticoid treatment induced a rapid response of PER1, PER2 and CRY1 in a GRα-dependent manner. Continuous glucocorticoid stimulation after 6h caused suppression of REV-ERBα. Administration of a GR antagonist, RU486, disrupted the circadian oscillation of clock genes and prevented the acute changes in PER1, PER2 and CRY1 levels. Overexpression of the GRß isoform alone did not alter the expression of the examined clock genes, but did prevent the GRα-related suppression of REV-ERBα. These alterations occurred independently from ACTH and CRH. Our data demonstrate that a peripheral clock system is present in a human adrenocortical cell line and that periodic oscillations of clock genes are influenced by glucocorticoids, mainly through GRα.

Overexpression of GRß in colonic mucosal cell line partly reflects altered gene expression in colonic mucosa of patients with inflammatory bowel disease.

The glucocorticoid receptor (GR) plays a crucial role in inflammatory responses. GR has several isoforms, of which the most deeply studied are the GRα and GRß. Recently it has been suggested that in addition to its negative dominant effect on GRα, the GRß may have a GRα-independent transcriptional activity. The GRß isoform was found to be frequently overexpressed in various autoimmune diseases, including inflammatory bowel disease (IBD). In this study, we wished to test whether the gene expression profile found in a GRß overexpressing intestinal cell line (Caco-2GRß) might mimic the gene expression alterations found in patients with IBD. Whole genome microarray analysis was performed in both normal and GRß overexpressing Caco-2 cell lines with and without dexamethasone treatment. IBD-related genes were identified from a meta-analysis of 245 microarrays available in online microarray deposits performed on intestinal mucosa samples from patients with IBD and healthy individuals. The differentially expressed genes were further studied using in silico pathway analysis. Overexpression of GRß altered a large proportion of genes that were not regulated by dexamethasone suggesting that GRß may have a GRα-independent role in the regulation of gene expression. About 10% of genes differentially expressed in colonic mucosa samples from IBD patients compared to normal subjects were also detected in Caco-2 GRß intestinal cell line. Common genes are involved in cell adhesion and cell proliferation. Overexpression of GRß in intestinal cells may affect appropriate mucosal repair and intact barrier function. The proposed novel role of GRß in intestinal epithelium warrants further studies.

A novel SPECT-based approach reveals early mechanisms of central and peripheral inflammation after cerebral ischemia.

Inflammation that develops in the brain and peripheral organs after stroke contributes profoundly to poor outcome of patients. However, mechanisms through which inflammation impacts on brain injury and overall outcome are improperly understood, in part because the earliest inflammatory events after brain injury are not revealed by current imaging tools. Here, we show that single-photon emission computed tomography (NanoSPECT/CT Plus) allows visualization of blood brain barrier (BBB) injury after experimental stroke well before changes can be detected with magnetic resonance imaging (MRI). Early 99mTc-DTPA (diethylene triamine pentaacetic acid) signal changes predict infarct development and systemic inflammation preceding experimental stroke leads to very early perfusion deficits and increased BBB injury within 2 hours after the onset of ischemia. Acute brain injury also leads to peripheral inflammation and immunosuppression, which contribute to poor outcome of stroke patients. The SPECT imaging revealed early (within 2 hours) changes in perfusion, barrier function and inflammation in the lungs and the gut after experimental stroke, with good predictive value for the development of histopathologic changes at later time points. Collectively, visualization of early inflammatory changes after stroke could open new translational research avenues to elucidate the interactions between central and peripheral inflammation and to evaluate in vivo 'multi-system' effects of putative anti-inflammatory treatments.