Summary Classical activation of macrophages (M(LPS+IFNγ)) elicits the expression of inducible nitric oxide synthase (iNOS), generating large amounts of NO and inhibiting mitochondrial respiration. Upregulation of glycolysis and a disrupted tricarboxylic acid (TCA) cycle underpin this switch to a pro-inflammatory phenotype. We show that the NOS cofactor tetrahydrobiopterin (BH 4 ) modulates IL-1β production and key aspects of metabolic remodeling in activated murine macrophages via NO production. Using two complementary genetic models, we reveal that NO modulates levels of the essential TCA cycle metabolites citrate and succinate, as well as the inflammatory mediator itaconate. Furthermore, NO regulates macrophage respiratory function via changes in the abundance of critical N-module subunits in Complex I. However, NO-deficient cells can still upregulate glycolysis despite changes in the abundance of glycolytic intermediates and proteins involved in glucose metabolism. Our findings reveal a fundamental role for iNOS-derived NO in regulating metabolic remodeling and cytokine production in the pro-inflammatory macrophage.

Objective. To investigate the endogenous involvement of transient receptor potential vanilloid 1 (TRPV1) in a model of knee joint inflammation in the mouse.Methods. Following characterization of wild-type (WT) and TRPV1-knockout mice, inflammation was induced via intraarticular (IA) injection of Freund's complete adjuvant (CFA). Knee swelling was assessed as diameter, and inflammatory heat hyperalgesia was determined using the Hargreaves technique, for up to 3 weeks. At 18 hours, acute hyperpermeability was measured with 125 I-albumin, and cytokines and myeloperoxidase activity, a marker of neutrophils, were assayed in synovial fluid extracts. The possibility that exogenous tumor necrosis factor ␣ (TNF␣) was involved in influencing TRPV1 activation was investigated in separate experiments.Results. Increased levels of knee swelling, hyperpermeability, leukocyte accumulation, and TNF␣ were found in WT mice 18 hours after IA CFA treatment compared with saline treatment. Significantly less knee swelling and hyperpermeability were found in TRPV1 ؊/؊ mice, but leukocyte accumulation and TNF␣ levels were similar in WT and TRPV1 ؊/؊ mice. Knee swelling in response to CFA remained significantly higher for a longer period in WT mice compared with TRPV1 ؊/؊ mice, with thermal hyperalgesic sensitivity observed at 24 hours and at 1 week in WT, but not TRPV1 ؊/؊ , mice. Knee swelling was attenuated (P < 0.05) in TRPV1 ؊/؊ compared with WT mice 4 hours after IA administration of TNF␣.Conclusion. Our findings indicate that TRPV1 has a role in acute and chronic inflammation in the mouse knee joint. Thus, selective antagonism of TRPV1 should be considered as a potential target for treatment of acute and chronic joint inflammation.

Transient potential vanilloid 1 (TRPV1) receptor is an ion channel receptor primarily localized on sensory nerves and activated by specific stimuli to initiate and amplify pain and inflammation, as typified by murine models of scald and arthritis. Little is known of the role of TRPV1 in sepsis, an infective disease associated with inflammation. Through use of a sublethal murine model of lipopolysaccharide-induced peritoneal sepsis, we provide novel evidence that genetic deletion of TRPV1 leads to an enhanced onset of various pathological components of systemic endotoxemia. Paired studies of TRPV1 knockout (KO) and wild-type mice demonstrate significantly enhanced hypotension (56+/-2% vs. 38+/-6% decrease in blood pressure, n=12), hypothermia (13+/-3% vs. 7+/-1% decrease in core temperature, n=6), and peritoneal exudate mediator levels (TNF-alpha, 0.78+/-0.2 vs. 0.38+/-0.1 ng/ml; nitrite, for NO, 35+/-10 vs. 15+/-3 microM; n=8) in TRPV1 KO mice, indicating loss of protective effect. Findings correlated with liver edema and raised plasma levels of aspartate aminotransferase in TRPV1 KO mice. These data suggest that TRPV1 may play an important regulatory role in sepsis independent of the major sensory neuropeptide substance P. The findings are relevant to developing strategies that increase the beneficial, and reduce the harmful, components of sepsis to prevent and treat this often fatal condition.

These studies provide in vivo evidence that ROS are involved in mediating TRPV1- and neuropeptide-dependent neurogenic vasodilatation. An essential role of NADPH oxidase-dependent ROS is revealed that may be of fundamental importance to the neurogenic vasodilator component involved in circulatory homeostasis and the pathophysiology of certain cardiovascular diseases.