RESUMEN
Collecting duct-derived endothelin (ET)-1 is an autocrine inhibitor of Na(+) and water reabsorption; its deficiency causes hypertension and water retention. Extracellular fluid volume expansion increases collecting duct ET-1, thereby promoting natriuresis and diuresis; however, how this coupling between volume expansion and collecting duct ET-1 occurs is incompletely understood. One possibility is that volume expansion increases tubular fluid flow. To investigate this, cultured IMCD3 cells were subjected to static or flow conditions. Exposure to a shear stress of 2 dyn/cm(2) for 2 h increased ET-1 mRNA content by â¼2.3-fold. Absence of perfusate Ca(2+), chelation of intracellular Ca(2+), or inhibition of Ca(2+) signaling (calmodulin, Ca(2+)/calmodulin-dependent kinase, calcineurin, PKC, or phospholipase C) prevented the flow response. Evaluation of possible flow-activated Ca(2+) entry pathways revealed no role for transient receptor potential (TRP)C3, TRPC6, and TRPV4; however, cells with TRPP2 (polycystin-2) knockdown had no ET-1 flow response. Flow increased intracellular Ca(2+) was blunted in TRPP2 knockdown cells. Nonspecific blockade of P2 receptors, as well as specific inhibition of P2X7 and P2Y2 receptors, prevented the ET-1 flow response. The ET-1 flow response was not affected by inhibition of either epithelial Na(+) channels or the mitochondrial Na(+)/Ca(2+) exchanger. Taken together, these findings provide evidence that in IMCD3 cells, flow, via polycystin-2 and P2 receptors, engages Ca(2+)-dependent signaling pathways that stimulate ET-1 synthesis.
Asunto(s)
Endotelina-1/metabolismo , Médula Renal/metabolismo , Túbulos Renales Colectores/metabolismo , Receptores Purinérgicos/metabolismo , Canales Catiónicos TRPP/metabolismo , Animales , Calcio/metabolismo , Señalización del Calcio , Línea Celular , Diuresis , Canales Epiteliales de Sodio/metabolismo , Masculino , Ratones , Natriuresis , ARN Mensajero/metabolismo , Ratas Sprague-Dawley , Sodio/metabolismo , Canales de Potencial de Receptor Transitorio/metabolismo , Agua/metabolismoRESUMEN
The important role of CD8(+) T cells in controlling HIV-1 infection through the innate as well as the adaptive immune system is well established. In addition to the major histocompatibility complex (MHC)-dependent cytotoxic activity of CD8(+) T cells, they produce soluble factors that suppress HIV-1 replication in an MHC-independent manner. Several of those factors have been identified, including beta-chemokines, Rantes, MIP-1alpha, MIP-1beta, and MDC. We previously identified that prothymosin alpha (ProTalpha) in the conditioned medium of HVS transformed CD8(+) T cells was a potent inhibitor of HIV-1 replication following proviral integration. In this report we further characterize the anti-HIV-1 activity of ProTalpha by demonstrating its target-cell specificity, distinction from additional inhibitors of HIV-1 transcription in CD8(+) T cell supernatants, as well as the differential regulation of host cell antiviral genes that could impact HIV-1 replication. These genes include a number of transcription factors as well IFN-alpha-inducible genes including PKR, IRF1, and Rantes, in the absence of induction of IFN-alpha. These data suggest that the anti-HIV-1 activity of ProTalpha is mediated through the modulation of a number of genes that have been reported to suppress HIV-1 replication including the dysregulation of transcription factors and the induction of PKR and Rantes mRNA.