TLR7 mediates increased vulnerability to ischemic acute kidney injury in diabetes

SUMMARY OBJECTIVE Diabetes is a risk factor for acute kidney injury (AKI). However, its mechanism of pathogenesis has not been elucidated. The aim of the study was to investigate the role of inflammation and the toll-like receptor 7 (TLR7) in ischemic AKI for diabetes. METHODS A high glucose hypoxia-reoxygenation model of human renal tubular epithelial (HK-2) cells was used to generate AKI induced by ischemia-reperfusion in diabetes. The activity of cells was measured by CCK-8 assay and LDH activity. Inflammatory cytokines were assessed by ELISA. TLR7, MyD88, and NF-κB expressions were examined by western blotting. Apoptosis was evaluated by flow cytometry. RESULTS The high glucose group and low glucose group were subjected to hypoxia-reoxygenation. The low glucose group developed only mild cell damage, apoptosis, and inflammatory response. In contrast, an equivalent hypoxia-reoxygenation injury provoked severe cell damage, apoptosis, and inflammatory response in the high glucose group. Expression of TLR7 and its related proteins were measured in the high glucose group before and after hypoxia-reoxygenation. The high glucose group exhibited more significant increases in TLR7 expression following hypoxia-reoxygenation than the low glucose group. In addition, the expression of TLR7 and its related proteins after hypoxia-reoxygenation were higher in the high glucose group than in the low glucose group. Inhibition of TLR7 provides significant protection against ischemic injury in diabetes. CONCLUSION Our results suggest that diabetes increases the vulnerability to ischemia-induced renal injury. This increased vulnerability originates from a heightened inflammatory response involving the TLR7 signal transduction pathway.

RESUMO OBJETIVO O diabetes é um fator de risco para a lesão renal aguda (LRA). No entanto, seu mecanismo de patogênese não foi elucidado. O objetivo do estudo foi investigar o papel da inflamação e do receptor Toll-like 7 (TLR7) na LRA isquêmica no diabetes. MÉTODOS Um modelo de hipóxia-reoxigenação de células epiteliais tubulares renais humanas (HK-2) na presença de concentrações altas de glicose foi utilizado para gerar LRA induzida por isquemia-reperfusão em diabetes. A atividade das células foi medida pelo ensaio Cell Counting Kit-8 (CCK-8) e pela atividade da lactato desidrogenase (LDH). As citocinas inflamatórias foram avaliadas por ensaio imunoenzimático (Elisa). A expressão de TLR7, do fator de diferenciação mieloide 88 (MyD88) e do fator de transcrição nuclear-κB (NF-κB) foi examinada por Western blotting. A apoptose foi avaliada por citometria de fluxo. RESULTADOS Os grupos glicose alta e glicose baixa foram submetidos à hipóxia-reoxigenação. O grupo de baixa glicose desenvolveu apenas danos celulares ligeiros, apoptose e uma resposta inflamatória. Em contraste, no grupo de alta glicose, uma lesão equivalente de hipóxia-reoxigenação provocou danos celulares graves, apoptose e uma resposta inflamatória. A expressão de TLR7 e suas proteínas relacionadas foi medida no grupo de alta glicose antes e após a hipóxia-reoxigenação. O grupo de alta glicose exibiu maiores aumentos na expressão de TLR7 após hipóxia-reoxigenação do que o grupo de baixa glicose. Além disso, a expressão de TLR7 e suas proteínas relacionadas após a hipóxia-reoxigenação foi maior no grupo com alto nível de glicose do que no grupo com baixo nível de glicose. A inibição do TLR7 fornece proteção significativa contra a lesão isquêmica no diabetes. CONCLUSÃO Nossos resultados sugerem que o diabetes aumenta a vulnerabilidade à lesão renal induzida por isquemia. Essa vulnerabilidade acrescida tem por origem uma resposta inflamatória aumentada envolvendo a via de transdução de sinal do TLR7.

LBP reduces theinflammatory injuryof kidney in septic rat and regulates the Keap1-Nrf2∕ARE signaling pathway

Abstract Purpose: To investigate the influence of lycium barbarum polysaccharides (LBP), a functional derivative from lycium barbarum, on septic kidney injury. Methods: The SD male rats were randomly divided into 8 groups. The concentration of IL-1β, IL-6, IL-8, TNF-α, NF-κB and ROS, in kidney cortex homogenates after 12 h treatments were determined by enzyme-linked immunosorbent assay and ROS test kit, respectively. Morphology observation of kidney tissue was conducted with HE staining. The mRNA and protein expression levels of Nrf2, HO-1, NQO1, NF-κB, and Keap1 in kidney tissues were determined by qRT-PCR and Western blot, respectively. Results: LPS treatment significantly increased the oxidative stress. After LBP treatment, the ROS content reduced significantly in a dose-depend manner. However, the levels of HO-1, NQO1 and Nrf2 as molecular elements that respond to oxidative stress were further increased. Also, administration of LBP increased the levels of NF-κB and Keap1, and decreased the levels of Nrf2 in the Keap 1-Nrf2∕ARE signaling pathway. By administrating the brusatol, the inhibition of Nrf2 enhanced the expression of NF-κB, inhibits the antioxidant responses, and further reverse the protective effect of LBP on the LPS induced septic kidney injury. Conclusion: Lycium barbarum polysaccharides can reduce inflammation and activate the antioxidant responses via regulating the level of pro-inflammatory cytokines and the Keap1-Nrf2/ARE signaling pathway.