The Role And Mechanism Of Remdesivir In Acute Kidney Injury By Inhibiting The Activation Of NLRP3 Inflammasome

Background:Acute kidney injury(AKI)is a common clinical complication in critically ill patients,which can progress to renal failure with high morbidity and mortality.However,other than dialysis,there is no effective therapeutic intervention to limit kidney damage and improve patient survival.Remdesivir(RDV,GS-5734),as a broad-spectrum antiviral nucleotide prodrug,has good in vitro antiviral effect and can inhibit the viral load in mice infected with severe acute respiratory syndrome-associated coronavirus.More and more researchers are interested in RDV due to the good effect of RDV in the treatment of COVID-19 patients.Studies have confirmed that RDV can protect rodents from acute lung injury by reducing neutrophil infiltration and interferon levels.However,the underlying mechanism remains unclear,and the role of RDV in other inflammatory diseases remains to be explored.As the core of inflammatory response,NLRP3 inflammasome is an important component of innate immunity,which can be activated by various types of pathogens or danger signals,and plays an important role in the body’s immune response and disease occurrence.Therefore,we propose that in AKI,RDV can regulate the activation of the NLRP3 inflammasome to improve AKI by effectively suppressing the inflammatory immune response.Methods:C57BL/6 male mice were selected;obtain mouse primary peritoneal macrophages:C57BL/6J mice(4-6 weeks old)were intraperitoneally injected with 3%thioglycolate broth.Three days later,peritoneal exudate cells were harvested and incubated for two hours,non-adherent cells were discarded,and adherent monolayers were used as peritoneal macrophages.Mouse primary peritoneal macrophages were treated with RDV/LPS/RDV+LPS,and the effect of RDV on NLRP3 inflammasome and pro-inflammatory factors was detected by RT-PCR.Western blotting was used to detect the effect of RDV on the activation of NLRP3 inflammasome,regulation of NF-κB and MAPK signaling pathways and pro-inflammatory factors;immunofluorescence was used to detect the effect of RDV on the expression of p-p65 protein;ELISA was used to detect the inhibition of RDV on pro-inflammatory factors effect.At the same time,the murine macrophage cell lines RAW264.7 and RAW264.7-RFP-ASC were selected as the research objects.After LPS and LPS+RDV treatment,the differences in the formation of ASC spots between the two groups were studied.Mice were randomly divided into four groups(n=6 per group):control,RDV,LPS,and RDV+LPS groups.There are two ways to administer RDV.The first way:mice in the RDV and RDV+ LPS groups were subcutaneously injected with a dose of 25 mg/kg of RDV at 12-hour intervals for 7 days,and the remaining two groups received an equal volume of normal saline.After 7 days,mice in the LPS and RDV+LPS groups were intraperitoneally injected with 10 mg/kg LPS to induce AKI,and mice in the control and RDV groups were intraperitoneally injected with an equal volume of normal saline.Mice were sacrificed 12 h after LPS injection,and kidney tissue,blood and urine were collected.Alternatively,mice in the RDV and RDV+LPS groups were first treated with 10 mg/kg LPS,followed by a subcutaneous injection of 25 mg/kg RDV 1 hour later.Twelve hours after LPS injection,the mice were sacrificed,and kidney tissue,blood and urine were collected.We used the first mode of administration(It was discussed in the discussion of the specific reasons).The effect of RDV on LPS-induced AKI was observed by HE and PAS staining of kidney tissue sections.The levels of Scr,BUN and urinary albumin in urine samples of mice in different treatment groups were detected by ELISA.Western blotting detected the effect of RDV on inflammasome activation and NF-κB and MAPK signaling pathways in renal tissue.The expression of related pro-inflammatory factors TNF-α,IL-6,IL-1β and IL-12 protein in the serum of mice in different treatment groups was detected by ELISA.Results:RDV could selectively inhibit LPS-induced NLRP3 inflammasome activation in macrophages,and at the same time inhibit NF-kB and MAPK signaling pathways in inflammatory macrophages.RDV can suppress the inflammatory response of macrophages by inhibiting the expression of pro-inflammatory cytokines.RDV can effectively inhibit the inflammatory pathological state and improve the renal function of AKI mice.RDV may alleviate LPS-induced AKI by inhibiting NF-kB and MAPK signaling pathways,thereby reducing the expression of NLRP3 and the release of inflammatory cytokines.Conclusion:Remdesivir can improve AKI by regulating the activation of macrophage inflammasome and inflammatory immune response through NF-κB and MAPK signaling pathways,and may have a potential therapeutic effect on patients with AKI in clinical practice.