Sensing soluble uric acid by Naip1-Nlrp3 platform.

Uric acid (UA), a product of purine nucleotide degradation able to initiate an immune response, represents a breakpoint in the evolutionary history of humans, when uricase, the enzyme required for UA cleavage, was lost. Despite being inert in human cells, UA in its soluble form (sUA) can increase the level of interleukin-1ß (IL-1ß) in murine macrophages. We, therefore, hypothesized that the recognition of sUA is achieved by the Naip1-Nlrp3 inflammasome platform. Through structural modelling predictions and transcriptome and functional analyses, we found that murine Naip1 expression in human macrophages induces IL-1ß expression, fatty acid production and an inflammation-related response upon sUA stimulation, a process reversed by the pharmacological and genetic inhibition of Nlrp3. Moreover, molecular interaction experiments showed that Naip1 directly recognizes sUA. Accordingly, Naip may be the sUA receptor lost through the human evolutionary process, and a better understanding of its recognition may lead to novel anti-hyperuricaemia therapies.

Macrophage inflammatory state in Type 1 diabetes: triggered by NLRP3/iNOS pathway and attenuated by docosahexaenoic acid.

Type 1 diabetes mellitus (T1D) is a chronic autoimmune disease characterized by insulin-producing pancreatic ß-cell destruction and hyperglycemia. While monocytes and NOD-like receptor family-pyrin domain containing 3 (NLRP3) are associated with T1D onset and development, the specific receptors and factors involved in NLRP3 inflammasome activation remain unknown. Herein, we evaluated the inflammatory state of resident peritoneal macrophages (PMs) from genetically modified non-obese diabetic (NOD), NLRP3-KO, wild-type (WT) mice and in peripheral blood mononuclear cells (PBMCs) from human T1D patients. We also assessed the effect of docosahexaenoic acid (DHA) on the inflammatory status. Macrophages from STZ-induced T1D mice exhibited increased inflammatory cytokine/chemokine levels, nitric oxide (NO) secretion, NLRP3 and iNOS protein levels, and augmented glycolytic activity compared to control animals. In PMs from NOD and STZ-induced T1D mice, DHA reduced NO production and attenuated the inflammatory state. Furthermore, iNOS and IL-1ß protein expression levels and NO production were lower in the PMs from diabetic NLRP3-KO mice than from WT mice. We also observed increased IL-1ß secretion in PBMCs from T1D patients and immortalized murine macrophages treated with advanced glycation end products and palmitic acid. The present study demonstrated that the resident PMs are in a proinflammatory state characterized by increased NLRP3/iNOS pathway-mediated NO production, up-regulated proinflammatory cytokine/chemokine receptor expression and altered glycolytic activity. Notably, ex vivo treatment with DHA reverted the diabetes-induced changes and attenuated the macrophage inflammatory state. It is plausible that DHA supplementation could be employed as adjuvant therapy for treating individuals with T1D.

Macrophage inflammatory state in Type 1 diabetes: triggered by NLRP3/iNOS pathway and attenuated by docosahexaenoic acid

Type 1 diabetes mellitus (T1D) is a chronic autoimmune disease characterized by insulin-producing pancreatic β-cell destruction and hyperglycemia. While monocytes and NOD-like receptor family-pyrin domain containing 3 (NLRP3) are associated with T1D onset and development, the specific receptors and factors involved in NLRP3 inflammasome activation remain unknown. Herein, we evaluated the inflammatory state of resident peritoneal macrophages (PMs) from genetically modified non-obese diabetic (NOD), NLRP3-KO, wild-type (WT) mice and in peripheral blood mononuclear cells (PBMCs) from human T1D patients. We also assessed the effect of docosahexaenoic acid (DHA) on the inflammatory status. Macrophages from STZ-induced T1D mice exhibited increased inflammatory cytokine/chemokine levels, nitric oxide (NO) secretion, NLRP3 and iNOS protein levels, and augmented glycolytic activity compared to control animals. In PMs from NOD and STZ-induced T1D mice, DHA reduced NO production and attenuated the inflammatory state. Furthermore, iNOS and IL-1β protein expression levels and NO production were lower in the PMs from diabetic NLRP3-KO mice than from WT mice. We also observed increased IL-1β secretion in PBMCs from T1D patients and immortalized murine macrophages treated with advanced glycation end products and palmitic acid. The present study demonstrated that the resident PMs are in a proinflammatory state characterized by increased NLRP3/iNOS pathway-mediated NO production, up-regulated proinflammatory cytokine/chemokine receptor expression and altered glycolytic activity. Notably, ex vivo treatment with DHA reverted the diabetes-induced changes and attenuated the macrophage inflammatory state. It is plausible that DHA supplementation could be employed as adjuvant therapy for treating individuals with T1D.

DPP-4 Inhibition Leads to Decreased Pancreatic Inflammatory Profile and Increased Frequency of Regulatory T Cells in Experimental Type 1 Diabetes.

Sitagliptin is a dipeptidyl peptidase-4 inhibitor (iDPP-4), which has been used for type 2 diabetes treatment. Recently, iDPP-4 has been described as a promising treatment of type 1 diabetes (T1D) but is still necessary to evaluate immune effects of sitagliptin. C57BL/6 mice were induced by multiple low doses of streptozotocin. Diabetes incidence, insulin, glucagon, glucagon-like peptide-1 (GLP-1) serum levels, and inflammatory cytokine levels were quantified in pancreas homogenate after 30 and 90 days of treatment. In addition, frequencies of inflammatory and regulatory T cell subsets were determined in the spleen and in the pancreatic lymph nodes. iDPP-4 decreased blood glucose level while increased GLP-1 and insulin levels. After long-term treatment, treated diabetic mice presented decreased frequency of CD4+CD26+ T cells and increased percentage of CD4+CD25hiFoxp3+ T cells in the spleen. Besides, pancreatic lymph nodes from diabetic mice treated with iDPP-4 presented lower percentage of CD11b+ cells and decreased levels of inflammatory cytokines in the pancreas. Treatment of type 1 diabetic mice with iDPP-4 improved metabolic control, decreased inflammatory profile in the pancreatic microenvironment, and increased systemic regulatory T cell frequency. Therefore, we suggest the long-term use of sitagliptin as a feasible and effective therapy for T1D.

Combination of a high-fat diet with sweetened condensed milk exacerbates inflammation and insulin resistance induced by each separately in mice.

Obesogenic diets increase body weight and cause insulin resistance (IR), however, the association of these changes with the main macronutrient in the diet remains to be elucidated. Male C57BL/6 mice were fed with: control (CD), CD and sweetened condensed milk (HS), high-fat (HF), and HF and condensed milk (HSHF). After 2 months, increased body weight, glucose intolerance, adipocyte size and cholesterol levels were observed. As compared with CD, HS ingested the same amount of calories whereas HF and HSHF ingested less. HS had increased plasma AST activity and liver type I collagen. HF caused mild liver steatosis and hepatocellular damage. HF and HSHF increased LDL-cholesterol, hepatocyte and adipocyte hypertrophy, TNF-α by macrophages and decreased lipogenesis and adiponectin in adipose tissue (AT). HSHF exacerbated these effects, increasing IR, lipolysis, mRNA expression of F4/80 and leptin in AT, Tlr-4 in soleus muscle and IL-6, IL-1ß, VCAM-1, and ICAM-1 protein in AT. The three obesogenic diets induced obesity and metabolic dysfunction. HS was more proinflammatory than the HF and induced hepatic fibrosis. The HF was more detrimental in terms of insulin sensitivity, and it caused liver steatosis. The combination HSHF exacerbated the effects of each separately on insulin resistance and AT inflammatory state.

Soluble Uric Acid Activates the NLRP3 Inflammasome.

Uric acid is a damage-associated molecular pattern (DAMP), released from ischemic tissues and dying cells which, when crystalized, is able to activate the NLRP3 inflammasome. Soluble uric acid (sUA) is found in high concentrations in the serum of great apes, and even higher in some diseases, before the appearance of crystals. In the present study, we sought to investigate whether uric acid, in the soluble form, could also activate the NLRP3 inflammasome and induce the production of IL-1ß. We monitored ROS, mitochondrial area and respiratory parameters from macrophages following sUA stimulus. We observed that sUA is released in a hypoxic environment and is able to induce IL-1ß release. This process is followed by production of mitochondrial ROS, ASC speck formation and caspase-1 activation. Nlrp3-/- macrophages presented a protected redox state, increased maximum and reserve oxygen consumption ratio (OCR) and higher VDAC protein levels when compared to WT and Myd88-/- cells. Using a disease model characterized by increased sUA levels, we observed a correlation between sUA, inflammasome activation and fibrosis. These findings suggest sUA activates the NLRP3 inflammasome. We propose that future therapeutic strategies for renal fibrosis should include strategies that block sUA or inhibit its recognition by phagocytes.

Dynamic changes of the Th17/Tc17 and regulatory T cell populations interfere in the experimental autoimmune diabetes pathogenesis.

A balance between proinflammatory (Th17 and Tc17) and anti-inflammatory (regulatory T cells) subsets of T cells is essential to maintain immunological tolerance and prevent the onset of several autoimmune diseases, including type 1 diabetes. However, the kinetics of these subsets and disease severity during the streptozotocin (STZ)-induced diabetes course has not been determined. Thus, susceptible C57BL/6 mice were administrated with multiple low doses of STZ and we evaluated the frequency/absolute number of these T cell subsets in the pancreatic lymph nodes (PLNs) and spleen and Th1, Th17, Treg cytokine production in the pancreatic tissue. At different time points of the disease progression (6, 11, 18 and 25 days after the last STZ administration), the histopathological alterations were also evaluated by H&E and immunohistochemistry staining. During the initial phase of diabetes development (day 6), we noted increased numbers of CD4(+) and CD8(+) T cells in spleen and PLNs. At the same time, the frequencies of Th17 and Tc17 cells in PLNs were also enhanced. In addition, the early augment of interferon gamma (IFN-γ), tumoral necrosis factor (TNF-α), IL-6 and IL-17 levels in pancreatic tissue correlated with pancreatic islet inflammation and mild ß-cell damage. Notably, the absolute number of Treg cells increased in PLNs during over time when compared to control group. Interestingly, increased IL-10 levels were associated with control of the inflammatory process during the late phase of the type 1 diabetes (day 25). In agreement, mice lacking the expression of IL-17 receptor (Il17r) showed impairment in STZ-induced diabetes progression, reduced peri-insulitis and beta cells preservation when compared with wild-type mice. Our findings suggest that dynamic changes of pathogenic Th17/Tc17 and regulatory T cell subsets numbers is associated with early strong inflammation in the pancreatic islets followed by late regulatory profile during the experimental STZ-induced diabetes course.