| 【Background】Systemic lupus erythematosus(SLE)is a common autoimmune disease in childhood that involves multiple system organs.The kidney is one of the most frequently affected organs,and lupus nephritis(LN)is the most leading cause of death in children with SLE.Studies have confirmed tubulointerstitial fibrosis as a key link between impaired renal function and progression to end-stage renal disease in children with lupus nephritis.Epithelial mesenchymal transition is considered to be an important mechanism in the development of tubulointerstitial fibrosis.Angiopoietin-like 4(ANGPTL4),as a key molecule promoting the occurrence of hyperlipidemia,has recently been confirmed to also induce tumor epithelial mesenchymal transition.But what role does ANGPTL4 play in the development of tubulointerstitial fibrosis;Whether ANGPTL4 participates in the development of tubulointerstitial fibrosis by promoting tubuloepithelial mesenchymal transition has not been reported.【Objective】To investigate whether ANGPTL4 promotes tubulointerstitial fibrogenesis through epithelial to mesenchymal transition,which in turn affects renal function in children with SLE and provides a novel therapeutic target to clinically improve lupus nephritis.【Methods】1.Case inclusion and exclusion criteria: Fulfilled the SLE classification criteria established by the American College of Rheumatology in 1997;Pediatric patients(< 18years)with lupus nephritis diagnosed by renal biopsy;Complete clinical and pathological data.Patients with other autoimmune diseases,hepatitis B virus associated nephritis,or other severe systemic diseases were excluded.2.Analysis of case data: In this project,we retrospectively analyzed the clinical and pathological data of 101 children with lupus nephritis in a single center,including lipid parameters(triglycerides,total cholesterol),renal injury indicators(serum creatinine,urea nitrogen,Cystatin C),urinary protein levels,and glomerular filtration rate(e GFR).3.Detection of serum ANGPTL4: Using the biological sample bank where the researcher is located,serum samples were collected from 47 children with lupus nephritis and 27 healthy children(age and gender matched outpatient physical examination),and the level of ANGPTL4 was detected by ELISA.4.Correlation analysis: 1)analyze the association of tubulointerstitial lesions with clinical and pathological features in children with lupus nephritis;2)To analyze the correlation between ANGPTL4 and serum lipid and renal function in children with lupus nephritis.5.Immunofluorescence identifies the location of ANGPTL4 in kidney tissue in different pathologic types of lupus nephritis,and ANGPTL4 and α-smooth muscle actin(α-SMA)colocalization.6.After human recombinant ANGPTL4 protein stimulation of human renal epithelial cells,α-SMA expression changes were examined by RT-q PCR.7.Knockout mouse Construction: Angptl4 knockout mice were generated by CRISPR/cas9 on a C57BL/6JGpt background.8.Pristane was used to generate lupus nephritis models in Angptl4 knockout and wild-type C57BL/6J mice.Analysis of the effects on serum lipids and renal function and histopathology in lupus nephritis model after Angptl4 knockdown.【Results】1.Basic clinical features.101 children with lupus nephritis were included in the study with a mean age of 11.35 ± 2.56 years,including 73 females and 28 males.The median triglycerides were 2.61(2.09,3.49)mmol/L,the mean total cholesterol was6.39 ± 2.14 mmol/L,and the overall incidence of hyperlipidemia was 82.2%.Median serum creatinine 61(42.5,92μmol/L),median urea nitrogen 7.75(5.25,12.66)mmol/L,median e GFR 117.3(74.6,163.3)ml/min/1.73m2。2.Correlation between serum lipid and renal injury.triglycerides were positively correlated with urea nitrogen,Cystatin C,SLEDAI scores(r = 0.26,0.324,0.209,P values = 0.009,0.001,0.036,respectively)and negatively correlated with e GFR in children with lupus nephritis(r=-0.23,P=0.021).3.Renal pathological characteristics.The distribution of lupus nephritis pathology was type III in 8 cases(7.9%),type III + V in 9 cases(8.9%),type IV in 42 cases(41.6%),type IV + V in 36 cases(35.6%),and type V in 6 cases(5.9%).The frequency of tubulointerstitial lesions was 94.1%,including 73(72.3%)mild,21(20.8%)moderate,and 1(1%)severe tubulointerstitial lesions.4.Relationship between tubulointerstitial injury and clinical features and renal histopathology.1)Extent of tubulointerstitial lesions was positively correlated with the course,mean arterial pressure,triglycerides,24 h urine protein,serum creatinine,urea nitrogen,and cystatin C(r = 0.27,0.339,0.296,0.246,0.432,0.444,0.464,all P <0.05)and negatively correlated with serum albumin and e GFR(r =-0.208,-0.450,all P<0.05).2)The extent of tubulointerstitial lesions varied significantly among different pathological types of lupus nephritis.3)The extent of tubulointerstitial lesions correlated positively with both active and chronic indices of the kidney tissue and glomerulus(r = 0.646,0.598,all P <0.001).5.Association between serum ANGPTL4 and renal function in children with lupus nephritis.1)Expression levels of ANGPTL4: serum ANGPTL4 was significantly higher in the lupus nephritis group compared with the control group.2)Serum ANGPTL4 was positively correlated with serum creatinine,urea nitrogen,and cystatin C in the lupus nephritis group.6.Expression features of ANGPTL4 in kidney.ANGPTL4 expression was in the tubulointerstitial and there is colocalization with α-SMA.7.In vitro,recombinant ANGPTL4 upregulates human renal tubular epithelial cells α-SMA and fibronectin expression.8.Results of a mouse model of lupus nephritis with Angptl4 gene knockout.Knocking out Angptl4 can partially inhibit the elevation of serum creatinine and triglycerides in a mouse model of lupus nephritis,alleviate renal tissue pathological damage,and reduce immune complex deposition.【Conclusions】Hyperlipidemia was prevalent in children with lupus nephritis,and hypertriglyceridemia was positively correlated with the degree of tubulointerstitial lesions in children.ANGPTL4,a key molecule in lipid metabolism,was not only highly expressed in the serum of children with lupus nephritis,but also positively correlated with renal function indicators.ANGPTL4,which is localized in tubular epithelial cells,may contribute to the development of tubulointerstitial fibrosis through epithelial to mesenchymal transition.Knockout of Angptl4 gene can improve renal tissue pathological damage and renal function in models of lupus nephritis. |
PDF Full Text Request