The Effect And Mechanism Of LIGHT In Cisplatin Induced Acute Kidney Injury

【Background】Cisplatin is a chemotherapy drug used in the treatment of various malignant tumors,and its renal toxicity is an inevitable and serious adverse reaction in the course of tumor chemotherapy.About 30%of patients will suffer Cisplatin induced acute kidney injury during the chemotherapy.The renal injury induced with Cisplatin mainly caused by inflammatory response,mitochondrial damage,and oxidative stress.LIGHT is an important inflammatory cytokine,which has been shown to play an important role in inflammation-related diseases such as allergic asthma and psoriasis,as well as mitochondrial damage diseases such as the injury of pancreatic islet cells,tumor cells,and motor neuron cells.However,it is unclear the effects of LIGHT in Cisplatin induced acute kidney injury.【Objectives】1.To clarify the role of LIGHT in cisplatin induced acute kidney injury2.To explore the underlying mechanism of LIGHT in cisplatin induced acute kidney injury.【Methods】1.In vivo1.1 Male mice aged 8-12 weeks wild-type(LIGHT^+/+)mice and LIGHT gene knockout(LIGHT^-/-)C57BL/6 mice were selected and divided into four groups,cisplatin group was given a single intraperitoneal injection of cisplatin（20 mg/kg,200 u L）,while saline group was injected with equal volume of normal 9 g/L saline.After 72 h,the mice were sacrificed and blood was collected from the eyeball while renal tissues were collected.1.2 Blood urea nitrogen（BUN）and Serum creatinine（Scr）levels were detected by automatic biochemical analyzer.Renal histopathological change was detected by HE staining,The localization of LIGHT in renal tissues was measured with IHC and IHC.The mRNA levels of LIGHT and Kidney Injury Molecule 1（KIM-1）,Interleukin 6（IL-6）and MonocyteChemotactic Protein 1（MCP-1）,Interleukin 1β（IL-1β）,and tumor necrosis factorα（TNF-α）were detected by quantitative PCR,Transmission electron microscopy（TEM）wasused to observe mitochondrial damage in renal tubular epithelial cells.Apoptosis in renaltissues was detected by Tunel staining.Protein levels of LIGHT,LTβR,HVEM,Bcl-2,Bax,Cytochrome C,Cleaved caspase3,Cleaved caspase9,NF-KBp65,and NF-KBpp65 were detected by Western Blot or IHC.2.In vitro2.1 Human tubular epithelial cells（HK-2）were cultured in vitro,incubated in a constant temperature incubator at 37℃,5%CO2 and 95%air,and inoculated with a 24-well plate at5×10⁵/well.When the density of cell reached 80%-90%,the cells were treated with cisplatin 1mg/m L（20μL）or recombinant human LIGHT100ng/m L（10μL）,after 24 hours,the cells were collected for next testing.2.2 Cell apoptosis,Reactive Oxygen Species and Mitochondrial Potential were detected by flow cytometry.Quantitative PCR was used to detect the inflammatory factors and Western Blot was used to detect the apoptosis-related protein.【Results】1.In vivo1.1 After Cisplatin treatment,The expressions of LIGHT and its receptors LTβR andHVEM in renal tissues were increased,LIGHT major expressed in renal tubular cells,theseresults suggest LIGHT-LTβR/HVEM may be involved in the injury of renal tubular cellsinduced by Cisplatin.1.2 After treatment with Cisplatin,Compared with LIGHT^+/+mice,The renal injury was increased with the serum urea nitrogen,creatinine levels,the double-blind score of HE staining,KIM-1,and the inflammatory factors in LIGHT^-/-mice.1.3 LIGHT deficiency accelerated the renal injury induced by Cisplatin through decreasing the expression of anti-apoptosis molecule Bcl-2,increasing the expression of pro-apoptosis molecules Bax,CytC,Cleaved caspase-3,and Cleaved caspase-9,and activating the NF-kB pathway.2 in vitro2.1 The flow cytometry results indicated that the recombinant human LIGHT could reduce the apoptosis and oxidative stress in HK-2 cells after the cells were treated with Cisplatin.2.2 Western blot results indicated that the recombinant human LIGHT improve the cells apoptosis induced by Cisplatin via down-regulated mitochondria apoptosis【Conclusions】1.LIGHT plays an important role in Cisplatin induced acute kidney injury;2.In Cisplatin induced acute kidney injury,apoptosis caused by LIGHT gene defect is associated with oxidative stress and Bcl-2 family expression disorder;3.LIGHT alleviates the damage of renal tubular cells caused by Cisplatin through down-regulation the mitochondria apoptosis.4.In vitro the recombinant human LIGHT can significantly reduce Cisplatin induced inflammation response and apoptosis.