Hyperuricemia contributes to renal inflammation. We aimed to investigate the role of Na+–K+–ATPase (NKA) in hyperuricemia-induced renal tubular injury. Human primary proximal tubular epithelial cells (PTECs) were incubated with uric acid (UA) at increasing doses or for increasing lengths of time. PTECs were then stimulated by pre-incubation with an NKA α1 expression vector or small interfering RNA before UA (100 μg ml−1, 48 h) stimulation. Hyperuricemic rats were induced by gastric oxonic acid and treated with febuxostat (Feb). ATP levels, the activity of NKA and expression of its α1 subunit, Src, NOD-like receptor pyrin domain-containing protein 3 (NLRP3) and interleukin 1β (IL-1β) were measured both in vitro and in vivo. Beginning at concentrations of 100 μg ml−1, UA started to dose-dependently reduce NKA activity. UA at a concentration of 100 μg ml−1 time-dependently affected the NKA activity, with the maximal increased NKA activity at 24 h, but the activity started to decrease after 48 h. This inhibitory effect of UA on NKA activity at 48 h was in addition to a decrease in NKA α1 expression in the cell membrane, but an increase in lysosomes. This process also involved the subsequent activation of Src kinase and NLRP3, promoting IL-1β processing. In hyperuricemic rats, renal cortex NKA activity and its α1 expression were upregulated at the 7th week and both decreased at the 10th week, accompanied with increased renal cortex expression of Src, NLRP3 and IL-1β. The UA levels were reduced and renal tubular injuries in hyperuricemic rats were alleviated in the Feb group. Our data suggested that the impairment of NKA and its consequent regulation of Src, NLRP3 and IL-1β in the renal proximal tubule contributed to hyperuricemia-induced renal tubular injury.

Adiponectin (APN) is a protein hormone that is primarily derived from adipocytes. It can also be secreted by renal cells. Hypoadiponectinemia has been documented in patients with hyperuricemia, however, whether soluble uric acid (SUA) regulates the expression of APN and APN receptor 1 (AdipoR1) in renal proximal tubule epithelial cells (PTECs) remains to be elucidated. The present study investigated the expression of APN and AdipoR1 in cultured PTECs that were exposed to SUA through immunofluorescence and western blot analysis. In addition, Sprague-Dawley rats with oxonic acid-induced hyperuricemia (HUA) with or without febuxostat treatment were employed as an animal model to measure 24 h urine protein, serum creatinine, urea nitrogen, uric acid and homeostasis model assessment of insulin resistance. Renal pathology was evaluated using hematoxylin and eosin and immunohistochemical staining. APN and AdipoR1 expression in the renal cortex were evaluated by western blotting. The results demonstrated that, in PTECs, the expression of APN and AdipoR1 was constant and increased upon SUA exposure. Similar observations were made within the proximal renal tubules of rats, and the oxonic acid-induced increases in APN and AdipoR1 were offset by febuxostat treatment. Furthermore, SUA-treated PTECs exhibited an increase in the expression of NLR family pyrin domain-containing (NLRP) 3, which was dose-dependent. NLRP3 expression was also significantly increased in the renal cortex of HUA rats compared with control and febuxostat-treated rats. In conclusion, SUA enhanced the expression of APN and AdipoR1 in PTECs, which was associated with an increase in NLRP3 expression. The APN-AdipoR1 pathway was demonstrated to have an important role in in vitro and in vivo models of renal proximal tubule inflammatory injury. Therefore, this pathway may be a potential therapy target in urate nephropathy.

Adiponectin (APN) exerts anti-inflammatory effects in various cells. Uric acid (UA) induces inflammation in proximal renal tubular epithelial cells (PTEcs). It remains unknown whether APN protects against UA-induced inflammation. In the present study, human PTEcs were incubated with 100 µg/ml soluble (S) UA in the presence or absence of globular (g) APN, APN receptor 1 (AdipoR1)-short hairpin RNA lentivirus or compound c. Reverse transcription-quantitative polymerase chain reaction (RT-qPcR) assays were performed to assess APN mRNA expression. Immunoblotting was used to assess the protein expression of APN, AdipoR1, NAcHT, leucine rich repeat and pyrin domain-containing protein 3 (NLRP3) and the activation of tumor necrosis factor (TNF) α and adenosine monophosphate-activated protein kinase (AMPK). ELISA analyses were performed to assess supernatant levels of interleukin (IL)-1β and TNFα. It was observed that SUA significantly enhanced APN mRNA and protein expression (both P<0.05) and increased NLRP3 (P<0.001) and TNFα (P<0.05) protein levels, as well as supernatant levels of IL-1β (P<0.01) and TNFα (P<0.001) compared with untreated cells. gAPN administration significantly limited TNFα synthesis and secretion (both P<0.001), significantly decreased IL-1β release (P<0.01), impacted NLRP3 protein expression and augmented AdipoR1 protein (P<0.01) and AMPK phosphorylation (P<0.05) levels compared with SUA-treated cells. AdipoR1 knockdown significantly promoted the synthesis (P<0.05) and release of TNFα (P<0.001), significantly increased IL-1β supernatant levels (P<0.01) and exhibited little influence on NLRP3 production (P>0.05) compared with the SUA-treated cells. Secreted TNFα levels were significantly increased upon the inhibition of AMPK (P<0.05) and protein levels of IL-1β, NLRP3 and TNFα in cell lysates were not significantly affected (P>0.05). In summary, the data demonstrated that SUA promoted APN expression in PTECs and that gAPN attenuated SUA-induced inflammation through the AdipoR1/AMPK signaling pathway. AdipoR1 knockdown and AMPK inactivation increased SUA-induced inflammatory damage in PTECs. These findings may help to further understand and regulate UA-associated inflammation in proximal renal tubules.