RESUMO
In the vasculature, nitric oxide (NO) is produced in the endothelium by endothelial nitric oxide synthase (eNOS) and is critical for the regulation of blood flow and blood pressure. Blood flow may also be regulated by the formation of nitrite-derived NO catalyzed by hemoproteins under hypoxic conditions. We sought to investigate whether nitrite administration may affect tissue perfusion and systemic hemodynamics in WT and eNOS knockout mice. We found that global eNOS KO mice show decreased tissue perfusion compared to WT mice by using laser speckle contrast imaging. To study both the acute and long-term effects of sodium nitrite (0, 0.1, 1, and 10 mg/kg) on peripheral blood flow and systemic blood pressure, a bolus of nitrite was delivered intraperitoneally every 24 h over 4 consecutive days. We found that nitrite administration resulted in a dose-dependent and acute increase in peripheral blood flow in eNOS KO mice but had no effects in WT mice. The nitrite induced changes in tissue perfusion were transient, as determined by intraindividual comparisons of tissue perfusion 24-h after injection. Accordingly, 10 mg/kg sodium nitrite acutely decreased blood pressure in eNOS KO mice but not in WT mice as determined by invasive Millar catheterization. Interestingly, we found the vasodilatory effects of nitrite to be inversely correlated to baseline tissue perfusion. These results demonstrate the nitrite acutely recovers hypoperfusion and hypertension in global eNOS KO mice and suggest the vasodilatory actions of nitrite are dependent upon tissue hypoperfusion.
Assuntos
Camundongos Knockout , Óxido Nítrico Sintase Tipo III , Animais , Óxido Nítrico Sintase Tipo III/metabolismo , Camundongos , Hemodinâmica/efeitos dos fármacos , Nitrito de Sódio/farmacologia , Masculino , Pressão Sanguínea/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Nitritos/farmacologia , Fluxo Sanguíneo Regional/efeitos dos fármacosRESUMO
Heme is an iron-containing prosthetic group necessary for the function of several proteins termed "hemoproteins." Erythrocytes contain most of the body's heme in the form of hemoglobin and contain high concentrations of free heme. In nonerythroid cells, where cytosolic heme concentrations are 2 to 3 orders of magnitude lower, heme plays an essential and often overlooked role in a variety of cellular processes. Indeed, hemoproteins are found in almost every subcellular compartment and are integral in cellular operations such as oxidative phosphorylation, amino acid metabolism, xenobiotic metabolism, and transcriptional regulation. Growing evidence reveals the participation of heme in dynamic processes such as circadian rhythms, NO signaling, and the modulation of enzyme activity. This dynamic view of heme biology uncovers exciting possibilities as to how hemoproteins may participate in a range of physiologic systems. Here, we discuss how heme is regulated at the level of its synthesis, availability, redox state, transport, and degradation and highlight the implications for cellular function and whole organism physiology.
Assuntos
Fenômenos Fisiológicos Celulares , Heme , Animais , Humanos , Ritmo Circadiano/fisiologia , Heme/metabolismo , Hemeproteínas/metabolismo , Oxirredução , Transdução de Sinais , Espaço Intracelular/metabolismo , Fenômenos Fisiológicos Celulares/fisiologiaRESUMO
Targeted degradation regulates the activity of the transcriptional repressor Bcl6 and its ability to suppress oxidative stress and inflammation. Here, we report that abundance of endothelial Bcl6 is determined by its interaction with Golgi-localized pannexin 3 (Panx3) and that Bcl6 transcriptional activity protects against vascular oxidative stress. Consistent with data from obese, hypertensive humans, mice with an endothelial cell-specific deficiency in Panx3 had spontaneous systemic hypertension without obvious changes in channel function, as assessed by Ca2+ handling, ATP amounts, or Golgi luminal pH. Panx3 bound to Bcl6, and its absence reduced Bcl6 protein abundance, suggesting that the interaction with Panx3 stabilized Bcl6 by preventing its degradation. Panx3 deficiency was associated with increased expression of the gene encoding the H2O2-producing enzyme Nox4, which is normally repressed by Bcl6, resulting in H2O2-induced oxidative damage in the vasculature. Catalase rescued impaired vasodilation in mice lacking endothelial Panx3. Administration of a newly developed peptide to inhibit the Panx3-Bcl6 interaction recapitulated the increase in Nox4 expression and in blood pressure seen in mice with endothelial Panx3 deficiency. Panx3-Bcl6-Nox4 dysregulation occurred in obesity-related hypertension, but not when hypertension was induced in the absence of obesity. Our findings provide insight into a channel-independent role of Panx3 wherein its interaction with Bcl6 determines vascular oxidative state, particularly under the adverse conditions of obesity.
Assuntos
Hipertensão , Fatores de Transcrição , Animais , Humanos , Camundongos , Diferenciação Celular , Proliferação de Células/fisiologia , Conexinas/metabolismo , Peróxido de Hidrogênio/farmacologia , Obesidade , Estresse Oxidativo , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection is mediated by the entry receptor angiotensin-converting enzyme 2 (ACE2). Although attachment factors and coreceptors facilitating entry are extensively studied, cellular entry factors inhibiting viral entry are largely unknown. Using a surfaceome CRISPR activation screen, we identified human LRRC15 as an inhibitory attachment factor for SARS-CoV-2 entry. LRRC15 directly binds to the receptor-binding domain (RBD) of spike protein with a moderate affinity and inhibits spike-mediated entry. Analysis of human lung single-cell RNA sequencing dataset reveals that expression of LRRC15 is primarily detected in fibroblasts and particularly enriched in pathological fibroblasts in COVID-19 patients. ACE2 and LRRC15 are not coexpressed in the same cell types in the lung. Strikingly, expression of LRRC15 in ACE2-negative cells blocks spike-mediated viral entry in ACE2+ cell in trans, suggesting a protective role of LRRC15 in a physiological context. Therefore, LRRC15 represents an inhibitory attachment factor for SARS-CoV-2 that regulates viral entry in trans.
Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Humanos , Enzima de Conversão de Angiotensina 2/genética , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus/metabolismo , COVID-19/genética , Ligação Proteica , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismoRESUMO
SARS-CoV-2 infection is mediated by the entry receptor ACE2. Although attachment factors and co-receptors facilitating entry are extensively studied, cellular entry factors inhibiting viral entry are largely unknown. Using a surfaceome CRISPR activation screen, we identified human LRRC15 as an inhibitory receptor for SARS-CoV-2 entry. LRRC15 directly binds to the receptor-binding domain (RBD) of spike protein with a moderate affinity and inhibits spike-mediated entry. Analysis of human lung single cell RNA sequencing dataset reveals that expression of LRRC15 is primarily detected in fibroblasts and particularly enriched in pathological fibroblasts in COVID-19 patients. ACE2 and LRRC15 are not co-expressed in the same cell types in the lung. Strikingly, expression of LRRC15 in ACE2-negative cells blocks spike-mediated viral entry in ACE2+ cell in trans, suggesting a protective role of LRRC15 in a physiological context. Therefore, LRRC15 represents an inhibitory receptor for SARS-CoV-2 regulating viral entry in trans.