The Therapeutic Effect And Mechanism Study Of Astaxantin In Acute Kidney Injury And Renal Fibrosis

Research Purpose:Acute kidney injury(AKI)is a common clinical critical illness.Renal fibrosis is the end-result of injury and the primary cause of the end-stage chronic renal failure.However,there is limited effective treatment methods.Astaxanthin(ATX)is a powerful antioxidant in the carotenoid family,and the antioxidant activity of ATX has been confirmed in a wide variety of living organisms.This research mainly explores the therapeutic effect and the mechanism of natural compounds ATX in acute kidney injury and renal fibrosis.The patients might benefit from a new therapeutic option.Methods:Part One:The therapeutic effect and the mechanism of ATX in acute kidney injury.HE staining was examined to evaluate the degree of renal ischemia-reperfusion injury(RIRI)in mice.Blood serum creatinine(Cr)and urea nitrogen(BUN)were assessed to measure renal function in mice.Malondialdehyde(MDA)and superoxide dismutase(SOD)were used to evaluate oxidative stress in kidney.TUNEL staining of kidney sections was used to detect the apoptosis cells.Immunohistochemical was used to evaluate the level of inflammation and fibrosis degree.In vitro,cells vitality was detected by CCK-8;cells apoptosis was analyzed by AnnexinV-FITC/PI.Malondialdehyde(MDA)and reactive oxygen species(ROS)were used to detect cell oxidative stress levels.RT-qPCR and Immunoblotting were examined to detect CHOP and Nrf2 signaling pathways.Part Two:The therapeutic effect and the mechanism of ATX in renal fibrosis.HE staining was examined to evaluate renal tubular atrophy,Masson and Sirius red staining were used to detect the collagen deposition.RT-qPCR was used to detect mRNAs of chemokines,inflammatory factors and M2 macrophage related genes.ELISA was used to test cytokines in cell supernatants.Immunohistochemical was used to detect extracellular matrix and macrophage infiltration.Flow cytometry was used to detect lymphocytes in peripheral blood and kidney tissues.Immunoblotting was examined to detect the fiber formation and EMT related proteins.Results:Astaxanthin significantly preserves renal function and histology after RIRI.The SOD increased and MDA decreased significantly after ATX treatment in kidney tissues.Serum Cr and BUN were obviously lower after ATX treatment.Immunohistochemical display ATX suppress the expression of SMA,F4/80 and NF kappa b significantly.ATX can increase the cells vitality of oxidative stress stimulated HK2,TUNEL staining and flow cytometry both proved that ATX can effectively inhibit the cell apoptosis in vivo and in vitro.ATX can inhibite CHOP signaling,and enhance Nrf2 signaling.ATX can also reduce renal tubular atrophy in renal fibrosis model,decrease extracellular collagen deposition,significantly reduce the SMA,Fibronectin and collagen I in kidney tissues.Chemokines and inflammatory factors were reduced.The percentage of CD8+T cells and CD163+M2c macrophages were obviously increased in ATX pre-treated mice.In vitro experiments established that ATX can restrain the activation of the M1 macrophages,and the M2 related genes increased.ATX can inhibit TGF-β inducing the expression of alpha SMA,fibronectin,and EMT related Vimentin,SNAIL,β-catenin.Conclusion:ATX preserves renal function and histology after RIRI by inhibiting CHOP signaling and promoting Nrf2 signaling.ATX can also inhibit EMT of renal tubular epithelial and renal fibrosis by affecting macrophages transformation,the raise of the CD8+T cells in kidney.This may supply a new strategy for clinical treatment of acute kidney injury and renal fibrosis.