RESUMO
Voltage-gated sodium channel alpha-subunits play a key role in pain pathophysiology, and are modulated by beta-subunits. We previously reported that beta1- and beta2-subunits were decreased in human sensory neurons after spinal root avulsion injury. We have now detected, by immunohistochemistry, beta3-subunits in 82% of small/medium and 67% of large diameter sensory neurons in intact human dorsal root ganglia: 54% of beta3 small/medium neurons were NGF receptor trkA negative. Unlike beta1- and beta2, beta3-immunoreactivity did not decrease after avulsion injury, and the beta3:neurofilament ratio was significantly increased in proximal injured human nerves. beta3-subunit expression may thus be regulated differently from beta1, beta2 and Nav1.8. Targeting beta3 interactions with key alpha-subunits, particularly Nav1.3 and Nav1.8, may provide novel selective analgesics.
Assuntos
Gânglios Espinais/citologia , Regulação da Expressão Gênica , Neurônios Aferentes/metabolismo , Subunidades Proteicas/metabolismo , Canais de Sódio/metabolismo , Adulto , Idoso , Western Blotting/métodos , Linhagem Celular , Embrião de Mamíferos , Gânglios Espinais/lesões , Humanos , Imuno-Histoquímica/métodos , Proteínas de Neurofilamentos/metabolismo , Subunidades Proteicas/genética , Canais de Sódio/genética , Fatores de Tempo , Transfecção/métodosRESUMO
The nociceptive C-fibers of the dorsal root ganglion express several sodium channel isoforms that associate with one or more regulatory beta-subunits (beta1-beta4). To determine the effects of individual and combinations of the beta-subunit isoforms, we co-expressed Nav1.8 in combination with these beta-subunits in Xenopus oocytes. Whole-cell inward sodium currents were recorded using the two-microelectrode voltage clamp method. Our studies revealed that the co-expression beta1 alone or in combination with other beta-subunits enhanced current amplitudes, accelerated current decay kinetics, and negatively shifted the steady-state curves. In contrast, beta2 alone and in combination with beta1 altered steady-state inactivation of Nav1.8 to more depolarized potentials. Co-expression of beta3 shifted steady-state inactivation to more depolarized potentials; however, combined beta1beta3 expression caused no shift in channel availability. The results in this study suggest that the functional behavior of Nav1.8 will vary depending on the type of beta-subunit that expressed under normal and disease states.
Assuntos
Proteínas do Tecido Nervoso/metabolismo , Canais de Sódio/metabolismo , Animais , Western Blotting , Canal de Sódio Disparado por Voltagem NAV1.8 , Proteínas do Tecido Nervoso/química , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Canais de Sódio/química , Xenopus laevisRESUMO
Resistin is a cysteine-rich protein postulated to be a molecular link between obesity and type 2 diabetes. The aim of this study was to investigate the role of PPAR gamma in the regulation of resistin expression in human primary macrophages. Fluorescent real-time PCR (Taqman) analysis of resistin expression across a range of human tissues showed that resistin is highly expressed in bone marrow compared to other tissues. Taqman analysis and Western blotting showed that rosiglitazone decreased resistin expression at both the mRNA and protein levels in human primary monocyte-derived macrophages in vitro. Resistin expression was reduced by up to 80% after exposure to 100 nM rosiglitazone for 96 h. Bioinformatics analysis of the genomic sequence upstream of the resistin coding sequence identified several putative PPAR response elements of which one was shown to bind PPAR gamma using electrophoretic mobility shift assays. Our data support a direct role for PPAR gamma in the regulation of resistin expression.