High Uric Acid Modulates The Functions Of Macrophages And Its Mechanisms

Hyperuricemia is a metabolic disease caused by disorders of purine metabolism.With the improvement of people’s living standards and changes in lifestyle,hyperuricemia has been one of the most common metabolic diseases.Currently,the prevalence of hyperuricemia in China is as high as 13.3%with the number of patients exceeding 120 million.Clinical studies have shown that hyperuricemia not only affects joints and causes gout,but also triggers and aggravates hypertension,diabetes,acute coronary syndrome,acute myocardial infarction,renal failure,stroke,and non-alcoholic fatty liver disease,etc.,which has become a major national health problem that needs to be addressed urgently.Hyperuricemia is implicated in mild tissue inflammation.Macrophages are ubiquitously present in all organs and their dysfunction is involved in multiple diseases.Dysregulated macrophages play a central role in the pathophysiology of inflammation,immunity and insulin resistance.Previous studies have focused on the effects and significance of monosodium urate crystals on macrophage activation,while the impacts of soluble urate on macrophages,including polarization,phagocytosis,migration,cytokine production,glucose metabolism,and insulin resistance,remain to be elucidated.Uricase(UOX)degrades uric acid to allantoin in rodents,which hinders the establishment of suitable rodent models of hyperuricemia.Here,we knocked out the UOX gene,established a mouse model of spontaneous hyperuricemia,and extracted peritoneal or bone marrow-derived macrophages to deeply investigate the intrinsic relationship between hyperuricemia and macrophages and its mechanism,which is valuable for the understanding of the pathophysiological significance of hyperuricemia.The underlying relationship between hyperuricemia and macrophages has been investigated in 3 aspects as follows:(1)High uric acid(HUA)regulates HMGB1-RAGE-ROS signaling to affect macrophage functionsBACKGROUND AND PURPOSE:Receptor for advanced glycosylation end products(RAGE)is a transmembrane protein belonging to the immunoglobulin receptor superfamily,widely expressed in a variety of cells.Multiple studies have suggested that the activation of RAGE signaling is profoundly engaged in the pathological processes of inflammation.The present study will examine the relationship between hyperuricemia,RAGE signaling and macrophage functions.METHODS:Peritoneal macrophages from WT or UOX-/-mice were harvested to evaluate inflammatory factor production,migration,and phagocytosis by flow cytometry(FC),RT-PCR,and western blot(WB).Bone marrow derived macrophage(BMDM)from WT or RAGE-/-mice were obtained and treated with HUA(10 and 15 mg/dL)for indicated time,and RNA-seq,WB,RT-PCR,or immunofluorescence(IF)were utilized to decipher the relationship between HUA,RAGE signaling and macrophage functions.RESULTS AND CONCLUSIONS:HUA induced the expression of RAGE ligands,activated the RAGE signaling,increased intracellular reactive oxygen species(ROS),triggered phosphorylation of ERK1/2,p38 and JNK,initiated NF-κB,increased secretion of pro-inflammatory factors and HMGB1,promoted migration and inhibited phagocytosis of macrophages.RAGE knockout alleviated HUA-induced secretion of pro-inflammatory factors,promotion of migration,and inhibition of phagocytosis in macrophages.These results suggest that HUA regulates HMGB1-RAGE-ROS signaling to affect macrophage functions.(1)High uric acid supports inflammatory responses by enhancing the pentose phosphate pathway in macrophages BACKGROUND AND AIMS:Macrophage polarization is a process under sophisticated control of cellular programming,regulated by metabolic adaptations besides dependence on cytokines,intrinsic signals and transcription factors.Glucose metabolism(glycolysis,tricarboxylic acid cycle,pentose phosphate pathway,etc.)plays a pivotal role in the pathophysiological processes of macrophages.We have illustrated the intrinsic signaling through which HUA affects macrophage functions in Part Ⅰ,but the underlying relationship between HUA,macrophage M1 polarization and glucose metabolism remains unclear,which is what this section will investigate.METHODS:BMDM of WT mice were isolated and subjected to HUA(15 mg/dL)for 24 h.The relationship between HUA,macrophage M1 polarization and glucose metabolism was examined using Seahorse,transmission electron microscopy,stable isotope tracing,metabolomics and FC.RESULTS AND CONCLUSIONS:HUA facilitated M1 polarization,decreased glycolysis,oxidative phosphorylation,enhanced the pentose phosphate pathway,disrupted the TCA cycle,and upregulated the NADPH/NADP+ratio of macrophages.These results indicate that HUA may bolster inflammatory response by potentiating the pentose phosphate pathway.Meanwhile,HUA downregulates glycolysis by upregulating the HIF1α/PFKFB3 signaling.(3)HUA induces insulin resistance in macrophages by promoting IRS2 degradation through the proteasome pathwayBACKGROUND AND AIMS:Studies have demonstrated a critical role of macrophage-induced inflammation in insulin resistance.Our previous study showed that HUA induces insulin resistance in liver,fat and skeletal muscle.However,the effect of HUA on the glucose uptake and insulin sensitivity of macrophages and its mechanisms are yet unknown.This study will evaluate the relationship between HUA,macrophages and insulin resistance.METHODS:Glucose/insulin tolerance test,metabolic cage test,tissue glucose uptake capacity test(18FDG SPECT/CT)were performed using WT or UOX-/-mice to assess systemic insulin resistance;F4/80 and glucose transporter 4(GLUT-4)were co-stained to detect the effect of hepatic circulating macrophage infiltration on glucose uptake;Percoll solution was used to isolate mouse hepatic macrophages and FC was used to detect the ratio of CD45+CD11 b+MHCII+inflammatory macrophages.BMDM was extracted from WT mice,treated with HUA,and subjected to the detection of insulin sensitivity and insulin signaling by 2-NBDG kit,WB and IF.RESULTS AND CONCLUSIONS:UOX-/-mice exhibited metabolic disorders,systemic insulin resistance,hepatic inflammatory macrophage infiltration,and increased circulating pro-inflammatory factors.Hepatic infiltrating macrophages showed an elevated CD45+CD11b+MHCII+ ratio.In vitro,HUA promotes IRS2 degradation via the proteasome pathway,thereby disrupting insulin signaling and inducing impaired glucose uptake and reduced insulin sensitivity in macrophages.We further showed that HUA promoted IRS2 degradation via the ubiquitination pathway in macrophages by IP and LC/MS enrichment analysis.These results suggest that UOX/-mice exhibit inflammatory macrophage accumulation in the liver and these macrophages show an inflammatory phenotype and impaired glucose uptake;mechanistically,HUA induces insulin resistance in macrophages by promoting IRS2 degradation through the proteasome pathway.