Background: Chronic kidney disease (CKD) is a severe public health problem associated with a disordered gut microbiome. However, the functional alterations of microbiota and their cross talk with metabolism pathways based on disease severity remain unclear. Results: We performed metagenomics and untargeted metabolomics in a cohort of 68 patients with CKD of differing severities and 20 healthy controls to characterize the complex interplay between the gut microbiome and fecal and serum metabolites during CKD progression. We identified 26 microbial species that significantly changed in patients with CKD; 18 species changed as the disease progressed, and eight species changed only in a specific CKD group. These distinct changes in gut microbiota were accompanied by functional alterations in arginine and proline, arachidonic acid, and glutathione metabolism and ubiquinone and other terpenoid-quinone biosynthesis pathways during CKD progression. Further metabolomic analyses revealed that the distributions of toxic and pro-oxidant metabolites from these four essential metabolic pathways varied in the feces and serum as CKD progressed. Furthermore, we observed a complex co-occurrence between CKD severity-related bacteria and the characterized metabolites from the four essential metabolic pathways. Notably, Ruminococcus bromii, fecal hydroquinone, and serum creatinine were identified as the main contributors to the integrated network, indicating their key roles in CKD progression. Moreover, a noninvasive model including R. bromii and fecal hydroquinone, L-cystine, and 12-keto-tetrahydro-LTB4 levels classified the CKD severity (area under the curve [AUC]: > 0.9) and had better performance than the serum creatinine level for mild CKD (AUC: 0.972 vs. 0.896). Conclusions: Perturbed CKD severity-related gut microbiota may contribute to unbalanced toxic and pro-oxidant metabolism in the gut and host, accelerating CKD progression, which may be an early diagnostic and therapeutic target for CKD.

Objective Impaired adaptive response to oxidative injuries is a fundamental mechanism central to the pathogenesis of chronic hepatitis C (CHC). Glycogen synthase kinase (GSK) 3β is an indispensable regulator of the oxidative stress response. However, the exact role of GSK3β in CHC is uncertain and was examined. Design GSK3β and Nrf2 signalling pathways were examined in JFH1 HCV infected Huh7.5.1 hepatocytes, and also in liver biopsy specimens from CHC patients. Results HCV infection elicited prominent Nrf2 antioxidant response in hepatocytes, marked by elevated expression of the Nrf2-dependent molecule haem oxygenase-1 and subsequent protection from apoptotic cell death. Inhibitory phosphorylation of GSK3β seems to be essential and sufficient for HCV-induced Nrf2 response. Mechanistically, GSK3β associated and physically interacted with Nrf2 in hepatocytes. In silico analysis revealed that Nrf2 encompasses multiple GSK3β phosphorylation consensus motifs, denoting Nrf2 as a cognate substrate of GSK3β. In the presence of TGFβ1, the HCV-induced GSK3β phosphorylation was blunted via a protein phosphatase 1-dependent mechanism and the cytoprotective Nrf2 response drastically impaired. This effect was counteracted by lithium, a selective inhibitor of GSK3β. In liver biopsy specimens from CHC patients, the expression of phosphorylated GSK3β positively correlated with Nrf2 expression and was inversely associated with the degree of liver injury. Moreover, CHC patients who received long-term lithium carbonate therapy primarily for concomitant psychiatric disorders exhibited much less liver injury, associated with enhanced hepatic expression of Nrf2. Conclusions Inhibition of GSK3β exerts hepatoprotection in CHC possibly through its direct regulation of Nrf2 antioxidant response.

Evidence suggests that glycogen synthase kinase 3β (GSK3β) contributes to AKI; however, its role in post-AKI kidney repair remains uncertain. Here, delayed treatment with a single dose of lithium, a selective inhibitor of GSK3β and a US Food and Drug Administration–approved mood stabilizer, accelerated recovery of renal function, promoted repopulation of renal tubular epithelia, and improved kidney repair in murine models of cisplatin- and ischemia/reperfusion-induced AKI. These effects associated with reduced GSK3β activity and elevated expression of proproliferative molecules, including cyclin D1, c-Myc, and hypoxia-inducible factor 1α (HIF-1α), in renal tubular epithelia. In cultured renal tubular cells, cisplatin exposure led to transient repression of GSK3β activity followed by a prolonged upregulation of activity. Rescue treatment with lithium inhibited GSK3β activity, enhanced nuclear expression of cyclin D1, c-Myc, and HIF-1α, and boosted cellular proliferation. Similarly, ectopic expression of a kinase-dead mutant of GSK3β enhanced the expression of cyclin D1, c-Myc, and HIF-1α and amplified cellular proliferation after cisplatin injury, whereas forced expression of a constitutively active mutant of GSK3β abrogated the effects of lithium. Mechanistically, GSK3β colocalized and physically interacted with cyclin D1, c-Myc, and HIF-1α in tubular cells. In silico analysis revealed that cyclin D1, c-Myc, and HIF-1α harbor putative GSK3β consensus phosphorylation motifs, implying GSK3β-directed phosphorylation and subsequent degradation of these molecules. Notably, cotreatment with lithium enhanced the proapoptotic effects of cisplatin in cultured colon cancer cells. Collectively, our findings suggest that pharmacologic targeting of GSK3β by lithium may be a novel therapeutic strategy to improve renal salvage after AKI.

Nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB) is regulated by a myriad of signaling cascades including glycogen synthase kinase (GSK) 3β and plays a Janus role in podocyte injury. In vitro, lipopolysaccharide (LPS) or adriamycin (ADR) elicited podocyte injury and cytoskeletal disruption, associated with NFκB activation and induced expression of NFκB target molecules, including pro-survival Bcl-xL and podocytopathic mediators like MCP-1, cathepsin L, and B7-1. Broad-range inhibition of NFκB diminished the expression of all NFκB target genes, restored cytoskeleton integrity, but potentiated apoptosis. In contrast, blockade of GSK3β by lithium or 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8) mitigated the expression of podocytopathic mediators, ameliorated podocyte injury, but barely affected Bcl-xL expression or sensitized apoptosis. Mechanistically, GSK3β was sufficient and essential for RelA/p65 phosphorylation, specifically at serine 467, which specifies the expression of selective NFκB target molecules, including podocytopathic mediators, but not Bcl-xL. In vivo, lithium or TDZD-8 therapy improved podocyte injury and proteinuria in mice treated with LPS or ADR, concomitant with the suppression of podocytopathic mediators, but retained Bcl-xL in glomerulus. Broad-range inhibition of NFκB conferred similar but much weakened antiproteinuric and podoprotective effects accompanied with a blunted glomerular expression of Bcl-xL and marked podocyte apoptosis. Thus, the GSK3β-dictated fine-tuning of NFκB may serve as a novel therapeutic target for podocytopathy.

Gout commonly manifests as a painful, self-limiting inflammatory arthritis. Nevertheless, the understanding of the inflammatory and immune responses underlying gout flares and remission remains ambiguous. Here, based on single-cell RNA-Seq and an independent validation cohort, we identified the potential mechanism of gout flare, which likely involves the upregulation of HLA-DQA1 ⁺ nonclassical monocytes and is related to antigen processing and presentation. Furthermore, Tregs also play an essential role in the suppressive capacity during gout remission. Cell communication analysis suggested the existence of altered crosstalk between monocytes and other T cell types, such as Tregs. Moreover, we observed the systemic upregulation of inflammatory and cytokine genes, primarily in classical monocytes, during gout flares. All monocyte subtypes showed increased arachidonic acid metabolic activity along with upregulation of prostaglandin-endoperoxide synthase 2 (PTGS2). We also detected a decrease in blood arachidonic acid and an increase in leukotriene B4 levels during gout flares. In summary, our study illustrates the distinctive immune cell responses and systemic inflammation patterns that characterize the transition from gout flares to remission, and it suggests that blood monocyte subtypes and Tregs are potential intervention targets for preventing recurrent gout attacks and progression.