RESUMEN
Regulator of G protein signaling (RGS) proteins, and notably members of the RGS-R4 subfamily, control vasocontractility by accelerating the inactivation of Gα-dependent signaling. RGS5 is the most highly and differently expressed RGS-R4 subfamily member in arterial smooth muscle. Expression of RGS5 first appears in pericytes during development of the afferent vascular tree, suggesting that RGS5 is a good candidate for a regulator of arterial contractility and, perhaps, for determining the mass of the smooth muscle coats required to regulate blood flow in the branches of the arterial tree. Consistent with this hypothesis, using cultured vascular smooth muscle cells (VSMCs), we demonstrate RGS5 overexpression inhibits G protein-coupled receptor (GPCR)-mediated hypertrophic responses. The next objective was to determine which physiological agonists directly control RGS5 expression in VSMCs. GPCR agonists failed to directly regulate RGS5 mRNA expression; however, platelet-derived growth factor (PDGF) acutely represses expression. Downregulation of RGS5 results in the induction of migration and the activation of the GPCR-mediated signaling pathways. This stimulation leads to the activation of mitogen-activated protein kinases directly downstream of receptor stimulation, and ultimately VSMC hypertrophy. These results demonstrate that RGS5 expression is a critical mediator of both VSMC contraction and potentially, arterial remodeling.
Asunto(s)
Músculo Liso Vascular/metabolismo , Miocitos del Músculo Liso/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Proteínas RGS/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores del Factor de Crecimiento Derivado de Plaquetas/metabolismo , Transducción de Señal , Angiotensina II/metabolismo , Animales , Becaplermina , Línea Celular , Movimiento Celular , Regulación de la Expresión Génica , Hipertrofia , Ligandos , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Músculo Liso Vascular/efectos de los fármacos , Miocitos del Músculo Liso/efectos de los fármacos , Proteínas Proto-Oncogénicas c-sis , Proteínas RGS/deficiencia , Proteínas RGS/genética , Interferencia de ARN , Ratas , Receptores Acoplados a Proteínas G/agonistas , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Transfección , VasoconstricciónRESUMEN
The engraftment of human stem cell-derived cardiomyocytes (hSC-CMs) is a promising treatment for remuscularizing the heart wall post-infarction, but it is plagued by low survival of transplanted cells. We hypothesize that this low survival rate is due to continued ischemia within the infarct, and that increasing the vascularization of the scar will ameliorate the ischemia and improve hSC-CM survival and engraftment. An adenovirus expressing the vascular growth factor Sonic Hedgehog (Shh) was injected into the infarcted myocardium of rats immediately after ischemia/reperfusion, four days prior to hSC-CM injection. By two weeks post-cell injection, Shh treatment had successfully increased capillary density outside the scar, but not within the scar. In addition, there was no change in vessel size or percent vascular volume when compared to cell injection alone. Micro-computed tomography revealed that Shh failed to increase the number and size of larger vessels. It also had no effect on graft size or heart function when compared to cell engraftment alone. Our data suggests that, when combined with the engraftment of hSC-CMs, expression of Shh within the infarct scar and surrounding myocardium is unable to increase vascularization of the infarct scar, and it does not improve survival or function of hSC-CM grafts.
Asunto(s)
Proteínas Hedgehog/metabolismo , Células Madre Embrionarias Humanas/metabolismo , Infarto del Miocardio/terapia , Miocitos Cardíacos/trasplante , Adenoviridae/genética , Animales , Diferenciación Celular , Vasos Coronarios/diagnóstico por imagen , Modelos Animales de Enfermedad , Vectores Genéticos/genética , Corazón/diagnóstico por imagen , Proteínas Hedgehog/genética , Humanos , Masculino , Infarto del Miocardio/diagnóstico por imagen , Infarto del Miocardio/etiología , Infarto del Miocardio/mortalidad , Miocardio/citología , Miocitos Cardíacos/metabolismo , Neovascularización Fisiológica , Ratas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Daño por Reperfusión/complicaciones , Tasa de Supervivencia , Transfección , Resultado del Tratamiento , Regulación hacia Arriba , Microtomografía por Rayos XRESUMEN
Liver fibrosis is mediated by hepatic stellate cells (HSCs), which respond to a variety of cytokine and growth factors to moderate the response to injury and create extracellular matrix at the site of injury. G-protein coupled receptor (GPCR)-mediated signaling, via endothelin-1 (ET-1) and angiotensin II (AngII), increases HSC contraction, migration and fibrogenesis. Regulator of G-protein signaling-5 (RGS5), an inhibitor of vasoactive GPCR agonists, functions to control GPCR-mediated contraction and hypertrophy in pericytes and smooth muscle cells (SMCs). Therefore we hypothesized that RGS5 controls GPCR signaling in activated HSCs in the context of liver injury. In this study, we localize RGS5 to the HSCs and demonstrate that Rgs5 expression is regulated during carbon tetrachloride (CCl4)-induced acute and chronic liver injury in Rgs5LacZ/LacZ reporter mice. Furthermore, CCl4 treated RGS5-null mice develop increased hepatocyte damage and fibrosis in response to CCl4 and have increased expression of markers of HSC activation. Knockdown of Rgs5 enhances ET-1-mediated signaling in HSCs in vitro. Taken together, we demonstrate that RGS5 is a critical regulator of GPCR signaling in HSCs and regulates HSC activation and fibrogenesis in liver injury.