Study On Cationic Bovine Serum Albumin Induced Animal Model Of Immune Complex Renal Injury And Exploration Of Potential Kidney Injury Markers

Objective:Immune complex induced by biotechnological drugs often leads to renal toxicity in nonclinical animal experiments and affects renal function of animals.Therefore,to detect IC-induced renal injury in drug safety evaluation is very important.Our study aimed to establish the SD rats and BALB/c mice model of immune complex-induced renal injury and its biomarkers were studied to provide a reference for rapid,sensitive and efficient detection of IC-induced renal injury in nonclinical safety evaluation of biotechnological drugs.Methods:1、Male SD rats were randomly divided into vehicle control group,low dose model group(10mg/kg),medium dose model group(12.5 mg/kg)and high dose model group(15 mg/kg).A rat model of IC-induced renal injury was established by induction immunization with C-BSA for2 weeks followed by booster immunization for 6 weeks.The immune complex renal injury was evaluated by detecting general toxicity indexes,24 h urinary protein,complement indexes,renal gross pathology,histopathological examination,special staining examination and transmission electron microscope examination.2、Male BALB/c mice were randomly divided into vehicle control group,low dose model group(2.5 mg/kg),medium dose model group(5 mg/kg),high dose model group(10mg/kg).A mouse model of IC-induced renal injury was established by induction immunization with C-BSA for2 weeks followed by booster immunization for 2 weeks.The immune complex renal injury was evaluated by general toxicity index,24 h urine protein,renal gross pathology,histopathological examination,special staining examination and transmission electron microscope examination.3、The biomarkers of C3,Ig G,Desmin and Podocin in two different species of renal injury models with IC were examined by immunohistochemistry and immunofluorescence to evaluate their potential as biomarkers.Results:1、After booster immunization by C-BSA administrated via tail vein in SD rats,the rats in model groups showed decreased body weight and food consumption;significantly increased 24 h urine protein;significantly increased relative kidney weight;yellow discoloration of kidneys;increased glomerular mesangial matrixes,thicken glomerular basement membrane and infiltration of inflammatory cell;activation of the complement system;thickened basement membranes,flattened,disappeared or partially fused foot processes as shown by TEM;additionally,extensively fused of foot processes and hump-like deposition of electron dense substance in the model high dose group;granular deposition of complement C3 and Ig G along the basement membrane;significantly increased expression of Desmin and altered expression of Podocin.2、After booster immunization by C-BSA administrated via tail vein in BALB/c mice,the mice in model groups showed decreased body weight and food consumption;increased 24 h urine protein;significantly increased relative kidney weight;thickened basement membranes,flattened,disappeared or partially fused foot processes,hump-like deposition of electron dense substance as shown by TEM;additionally,extensively fused foot processes in the high dose model group;increased glomerular mesangial matrixes,thicken glomerular basement membrane and infiltration of inflammatory cell;granular deposition of complement C3 and Ig G along the basement membrane;increased expression of desmin and and expression of Podocin was not changed.Conclusion:1、IC-induced renal injury models in SD rats and BALB/c mice were successfully established by induction immunization followed by booster immunization of C-BSA in the study.2、C-BSA could induce hypersensitivity type Ⅲ in SD rats and BALB/c mice as a foreign protein,with IC developed and deposited at glomerular basement membrane,which could activate the complement system to initiate an inflammatory response leading to the damage of renal glomerulus.3、In this study,immunohistochemistry and immunofluorescence have the advantages of strong specificity and high sensitivity,which can be used as the first choice for the examination of kidney with immune complex renal injury.Special staining and transmission electron microscopy can be used as the auxiliary examination methods for immune complex renal injury.4、Complement C3,and Podocin might be biomarkers of IC-induced renal injury.The expression level of Desmin was positively correlated with the severity of the damage of renal glomerulus.