Acute Microcystin Exposure Induced Kidney Damage And Ferroptosis In Rats

Objective:To investigate the toxic effects and the regulatory mechanism of ferroptosis on kidneys in rats after acute microcystin-LR（MC-LR）exposure,a dose-effect model and a time-effect model of renal injury induced by MC-LR were constructed.Changes of histopathology,blood biochemistry and transcription expression of genes related to ferroptosis were detected.This study will provide a theoretical basis for the prevention and treatment of renal toxicity induced by microcystin exposure in human.Methods:（1）Dose-effect model:healthy male Wistar rats were randomly divided into 4 groups（n=6）and given a single intravenous injection of MC-LR for 24 h:control group（0.9%Na Cl）,low dose group(50μg/kg MC-LR bw,0.5 LD₅₀),medium dose group(75μg/kg MC-LR bw,0.75 LD₅₀)and high dose group(100μg/kg MC-LR bw,1 LD₅₀).（2）Time-effect model:healthy male Wistar rats were randomly divided into two groups:control group（0.9%Na Cl）and MC-LR group(75μg/kg MC-LR bw,0.75 LD₅₀),with 6 rats in each group.The rats were administrated with MC-LR via intravenous injection for 2 h,6 h,12 h,and 24 h.Serum and kidney tissue were collected.The toxic effect of MC-LR on kidney was analyzed by HE staining,serum biochemical indexes,antioxidant system enzymes and real-time quantitative PCR.Results:（1）After 24 h of acute exposure to MC-LR,different degrees of pathological changes were observed in kidney tissues,mainly manifested as the appearance of protein casts,glomerular swelling,renal tubular degeneration and necrosis.Serum biochemical results showed that the concentrations of UREA,CREA,UA,CO₂,P,MG,Cys-c and e GFR were changed after MC-LR exposure,indicating that MC-LR induced kidney function damage in rats..MC-LR could induce changes of activities of GST and GPX and higher on the concentrations of GSH and MDA,suggesting that acute exposure to MC-LR induced oxidative stress.The m RNA levels of slc3a2,gpx4,fth1,alox15,acls4,tfr,fpn1 and ncoa4 were up-regulated or down-regulated,suggesting that ferroptosis might be involved in kidney injury induced by acute exposure to MC-LR.（2）Histopathological results showed that after exposure for 2 h,6 h,and12 h,there were no significant pathological changes in both the control group and the 75μg/kg group.After 24 h of exposure,the 75μg/kg group showed swelling and necrosis of glomeruli in the cortical area,exudation of red blood cells,and degeneration and necrosis of renal tubules.Serum biochemical results showed that the concentrations of UREA,CREA and P increased significantly and CO₂decreased at 2 h.The concentrations of UREA,Na,P and MG increased significantly and K decreased significantly at 6 h.The concentrations of Cys-c decreased and e GFR increased at 12 h.The concentrations of UREA and CREA increased significantly,while UA and P decreased at 24 h.These results suggested that the disturbance of electrolyte acid hydrolysis occurred at2 h after MC-LR exposure,and the glomerular filtration function may be affected at 12 h after MC-LR exposure.The transcription levels of ferroptosis-related genes did not change at 2 h after MC-LR exposure.The transcriptional levels of alox15 and ncoa4 were significantly down-regulated at6 h after MC-LR exposure.The transcriptional levels of slc3a2,gpx4,alox15,tfr,fth1 and ncoa4 were significantly down-regulated at 12 h after MC-LR exposure.The transcriptional levels of slc3a2,alox15,acsl4 and tfr were up-regulated while the transcriptional levels of gpx4,fth1,ncoa4 and fpn1 were down-regulated at 24 h after MC-LR exposure.Conclusion:（1）Acute exposure to 0.5,0.75 or 1 LD₅₀dose of MC-LR for24 h could induce histopathological changes in the kidney of rats,leading to kidney function damage.Oxidative stress and ferroptosis might be involved in the kidney toxicity induced by MC-LR in rats.（2）After acute exposure to 75μg/kg MC-LR(0.75 LD₅₀)for 6 h and 12 h,renal function and the transcription of genes related to ferroptosis were changed,especially at 24 h,suggesting that MC-LR exposure induced changes of renal function and ferroptosis.