Pyridoxamine

Diabetic kidney disease (DKD) is the leading cause of chronic kidney disease and end-stage renal disease (ESRD), necessitating novel therapies beyond conventional approaches. Emerging evidence indicates that gut microbiota dysbiosis promotes DKD progression through metabolite-mediated renal injury. Jinlida (JLD) is a clinically validated traditional Chinese medicine with antidiabetic activity, but its microbiota-mediated renoprotective mechanism remains unclear.

This study investigates whether JLD alleviates DKD by modulating gut microbiota and vitamin B6 metabolism, and elucidates the renoprotective mechanism of its key metabolite, pyridoxamine (PM).

To assess JLD's microbiota-dependent effects, we employed antibiotic-induced pseudo-germ-free mice and fecal microbiota transplantation (FMT). Metagenomics and untargeted metabolomics delineated gut microbiota and metabolite compositional changes. Renal PM levels were quantified by LC-MS/MS. The renoprotective effects and mechanisms of direct PM supplementation against DKD were further evaluated in vivo and in vitro.

JLD's therapeutic effects on proteinuria and glomerulosclerosis were shown to partially depend on microbiota homeostasis. Metabolomic analysis demonstrated that JLD significantly upregulated the vitamin B6 metabolic pathway and increased levels of related metabolites, including PM and pyridoxine (PN). Metagenomic analyses indicated that JLD remodeled the gut microbiota composition and enriched pathways related to cofactor biosynthesis, and markedly increased the relative abundance of key enzyme genes involved in the de novo (DXP-dependent) vitamin B6 biosynthesis pathway - namely pdxJ, pdxB, dxs and dxr. Genes related to vitamin B6 activation and conversion (pdxH, aldH) showed no significant changes, suggesting that JLD may promote PM accumulation by enhancing the microbiota's capacity for vitamin B6 biosynthesis rather than its subsequent activation/conversion. Source-tracking pinpointed Paramuribaculum intestinale as the core functional species. In vitro culture experiments showed that JLD markedly promoted the growth of this strain and elevated PM production, and that the strain's conditioned culture medium effectively inhibited formation of advanced glycation end-products (AGEs). Notably, direct supplementation with PM recapitulated the renoprotective effects of JLD in vivo. Mechanistically, PM inhibited the AGEs-RAGE-NF-κB-AP-1 axis and TGF-β receptor signaling, thereby suppressing NF-κB-driven inflammation and Smad2-mediated fibrosis.

JLD remodels the gut microbiota and enhances its de novo vitamin B6 biosynthetic capacity, leading to accumulation of PM. Gut-derived PM enters the circulation and functions as an effector molecule targeting the kidney; through PM's direct carbonyl-trapping activity it scavenges AGEs and suppresses the AGEs-RAGE axis as well as downstream inflammatory and profibrotic signaling, thereby exerting renoprotective effects. This study reveals PM as a microbially derived metabolite with therapeutic potential in DKD and offers a new metabolism-directed strategy for DKD treatment.

The typical western diet contributes to the accumulation of Advanced Glycation End-products (AGEs), reactive compounds involved in metabolic alterations and low-grade inflammation, referred as metaflammation. AGEs affect sphingolipid metabolism increasing sphingosine-1-phosphate (S1P). S1P is in turn induces pro-inflammatory M1 macrophage polarization through activation of RhoA/ROCK. The current study seeks to determine whether inhibition of AGEs by the antiglycating vitamin B6 analog pyridoxamine prevents metaflammation via modulation of S1P/RhoA/ROCK signalling. Gastrocnemius muscle was obtained from mice fed a standard diet (SD) or a western diet (WD), supplemented or not with pyridoxamine (WD+PYR, 150mg/kg bw/day). Metabolic and inflammatory profiles have been investigated in plasma, while accumulation of AGE-modified proteins, expression of RAGE, S1P-synthesizing enzymes and S1P receptors, as well as activation of RhoA/ROCK/IRS-1 pathway have been analysed in gastrocnemius muscle. Markers of local inflammatory cells infiltration and macrophage polarization were also explored. In comparison to SD mice, WD mice developed obesity, glucose intolerance, and muscle lipid accumulation with increased plasma proinflammatory cytokines and muscle M1-polarized macrophage infiltration. WD mice muscle showed higher content of AGE-modified proteins with increased S1P synthesis and S1PR1 expression, paralleled by activation of RhoA/ROCK and impairment of insulin signalling. By inhibiting AGEs production through pyridoxamine supplementation, all the WD-induced metabolic and inflammatory signalling alterations were significantly reversed towards the SD state. The present findings confirm a relevant role for AGE/S1P/RhoA/ROCK signalling in diet-induced metaflammation, suggesting the formulation of pharmacological and nutraceutical tools to prevent AGEs accumulation as a useful strategy against obesity-related metabolic and inflammatory diseases.