Acidosis Activation of the Proton-Sensing GPR4 Receptor Stimulates Vascular Endothelial Cell Inflammatory Responses Revealed by Transcriptome Analysis

Acidic tissue microenvironment commonly exists in inflammatory diseases, tumors, ischemic organs, sickle cell disease, and many other pathological conditions due to hypoxia, glycolytic cell metabolism and deficient blood perfusion. However, the molecular mechanisms by which cells sense and respond to the acidic microenvironment are not well understood. GPR4 is a proton-sensing receptor expressed in endothelial cells and other cell types. The receptor is fully activated by acidic extracellular pH but exhibits lesser activity at the physiological pH 7.4 and minimal activity at more alkaline pH. To delineate the function and signaling pathways of GPR4 activation by acidosis in endothelial cells, we compared the global gene expression of the acidosis response in primary human umbilical vein endothelial cells (HUVEC) with varying level of GPR4. The results demonstrated that acidosis activation of GPR4 in HUVEC substantially increased the expression of a number of inflammatory genes such as chemokines, cytokines, adhesion molecules, NF-κB pathway genes, and prostaglandin-endoperoxidase synthase 2 (PTGS2 or COX-2) and stress response genes such as ATF3 and DDIT3 (CHOP). Similar GPR4-mediated acidosis induction of the inflammatory genes was also noted in other types of endothelial cells including human lung microvascular endothelial cells and pulmonary artery endothelial cells. Further analyses indicated that the NF-κB pathway was important for the acidosis/GPR4-induced inflammatory gene expression. Moreover, acidosis activation of GPR4 increased the adhesion of HUVEC to U937 monocytic cells under a flow condition. Importantly, treatment with a recently identified GPR4 antagonist significantly reduced the acidosis/GPR4-mediated endothelial cell inflammatory response. Taken together, these results show that activation of GPR4 by acidosis stimulates the expression of a wide range of inflammatory genes in endothelial cells. Such inflammatory response can be suppressed by GPR4 small molecule inhibitors and hold potential therapeutic value

Differential endothelial cell gene expression by African Americans versus Caucasian Americans: a possible contribution to health disparity in vascular disease and cancer

<p>Abstract</p> <p>Background</p> <p>Health disparities and the high prevalence of cardiovascular disease continue to be perplexing worldwide health challenges. This study addresses the possibility that genetic differences affecting the biology of the vascular endothelium could be a factor contributing to the increased burden of cardiovascular disease and cancer among African Americans (AA) compared to Caucasian Americans (CA).</p> <p>Methods</p> <p>From self-identified, healthy, 20 to 29-year-old AA (n = 21) and CA (n = 17), we established cultures of blood outgrowth endothelial cells (BOEC) and applied microarray profiling. BOEC have never been exposed to <it>in vivo </it>influences, and their gene expression reflects culture conditions (meticulously controlled) and donor genetics. Significance Analysis of Microarray identified differential expression of single genes. Gene Set Enrichment Analysis examined expression of pre-determined gene sets that survey nine biological systems relevant to endothelial biology.</p> <p>Results</p> <p>At the highly stringent threshold of False Discovery Rate (FDR) = 0, 31 single genes were differentially expressed in AA. <it>PSPH </it>exhibited the greatest fold-change (AA > CA), but this was entirely accounted for by a homolog (<it>PSPHL</it>) hidden within the <it>PSPH </it>probe set. Among other significantly different genes were: for AA > CA, <it>SOS1, AMFR, FGFR3; and for AA < CA, ARVCF, BIN3, EIF4B. </it>Many more (221 transcripts for 204 genes) were differentially expressed at the less stringent threshold of FDR <.05. Using the biological systems approach, we identified shear response biology as being significantly different for AA versus CA, showing an apparent tonic increase of expression (AA > CA) for 46/157 genes within that system.</p> <p>Conclusions</p> <p>Many of the genes implicated here have substantial roles in endothelial biology. Shear stress response, a critical regulator of endothelial function and vascular homeostasis, may be different between AA and CA. These results potentially have direct implications for the role of endothelial cells in vascular disease (hypertension, stroke) and cancer (via angiogenesis). Also, they are consistent with our over-arching hypothesis that genetic influences stemming from ancestral continent-of-origin could impact upon endothelial cell biology and thereby contribute to disparity of vascular-related disease burden among AA. The method used here could be productively employed to bridge the gap between information from structural genomics (for example, disease association) and cell function and pathophysiology.</p

Mechanisms of enhanced thrombus formation in cerebral microvessels of mice expressing hemoglobin-S

The microvasculature assumes an inflammatory and procoagulant state in a variety of different diseases, including sickle cell disease (SCD), which may contribute to the high incidence of ischemic stroke in these patients. This study provides evidence for accelerated thrombus formation in arterioles and venules in the cerebral vasculature of mice that express hemoglobin-S (βs mice). Enhanced microvascular thrombosis in βs mice was blunted by immunologic or genetic interventions that target tissue factor, endothelial protein C receptor, activated protein C, or thrombin. Platelets from βs mice also exhibited enhanced aggregation velocity after stimulation with thrombin but not ADP. Neutropenia also protected against the enhanced thrombosis response in βs mice. These results indicate that the cerebral microvasculature is rendered vulnerable to thrombus formation in βs mice via a neutrophil-dependent mechanism that is associated with an increased formation of and enhanced platelet sensitivity to thrombin