To explore the mechanism by which Huashen Tongfengtai ameliorates renal injury in a mouse model of urate nephropathy.

Sixty SPF-grade male C57BL/6J mice were used. Thirty mice were randomly divided into the normal (n=8), modeling (n=8), Hushen Tongfengtai low-dose (n=7, 33.47 g/kg), or Hushen Tongfengtai high-dose (n=7, 66.93 g/kg) groups using the random number table method. Mice in the normal group were administered 0.5% CMC-Na by gavage and intraperitoneal injection, urate nephropathy was induced by intraperitoneal injection of potassium oxonate (300 mg/kg) combined with oral hypoxanthine (250 mg/kg). During the model establishment, the normal and modeling groups received equal volumes of distilled water by gavage, whereas the treatment groups were administered the corresponding drugs once daily for 14 consecutive days. Serum uric acid (UA), creatinine (Cr), and blood urea nitrogen (BUN) contents were measured using an enzymatic colorimetric assay. Renal histopathological alterations were assessed through Masson staining. Gut metabonomics was conducted using UPLC-MS/MS with a self-constructed database to screen and identify key metabolites, followed by Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. The remaining 30 mice were subjected to identical modeling procedures and assigned to the model (n=8), Hushen Tongfengtai (n=8, 33.47 g/kg), 18β-glycyrrhetinic acid low-dose (n=7, 50 mg/kg), or 18β-glycyrrhetinic acid high-dose (n=7, 100 mg/kg) groups using the random number table method. During the model establishment, the mice in the model group received distilled water, whereas the other groups received the corresponding drugs via gavage once daily for 14 days. Serum UA was measured using an enzymatic colorimetric assay. The expression levels of tumor necrosis factor-α(TNF-α), interleukin (IL)-6, IL-1β, and IL-10 mRNA in the colon tissues were quantified by real-time PCR. Colonic morphology and mucus secretion were evaluated using Alcian blue-periodic acid-Schiff staining. Immunofluorescence was employed to examine the expression of zonula occludens-1(ZO-1), Occludin, and E-cadherin. The protein expressions of ZO-1, Occludin, E-cadherin, phosphorylated Toll-like receptor 4(p-TLR4), and phosphorylated myeloid differentiation primary response 88(p-MyD88) were determined via western blotting.

Compared to the normal group, the modeling group demonstrated increased UA, Cr, and BUN levels (P < 0.05), as well as renal interstitial fibrosis. Compared with the modeling group, UA, Cr, and BUN contents were reduced in the Hushen Tongfengtai low-and high-dose groups (P < 0.05) and renal histological injury was mitigated. Hushen Tongfengtai substantially modified the gut metabolic profile; 18β-glycyrrhetinic acid was the predominant upregulated metabolite, which is primarily associated with the TLR4/MyD88 signaling pathway. Compared to the model group, the serum UA levels were decreased in both 18β-glycyrrhetinic acid low-and high-dose groups, and IL-6 and IL-1β mRNA expressions were decreased in colon tissues. In contrast, IL-10 mRNA expression was increased, the colonic mucus secretion was enhanced, and ZO-1, Occludin, and E-cadherin protein expressions were increased, whereas that of p-TLR4 and p-MyD88 were reduced (P < 0.05).

Hushen Tongfengtai may ameliorate renal dysfunction and tissue injury in mice with urate nephropathy by upregulating 18β-glycyrrhetinic acid. This mechanism facilitates the restoration of intestinal barrier integrity and inhibits TLR4/MyD88-mediated inflammatory signaling.