Study On Potential Markers Of Nephrotic Syndrome Based On Metabolomics And Microbiology

Background: Nephrotic syndrome is a common glomerular disease with multiple types such as membranous nephropathy and Ig A nephropathy.If it cannot be treated timely and effectively,the disease will continue to deteriorate into renal failure and even uremia.At present,indicators such as urinary nitrogen and creatinine are commonly used in the clinical diagnosis of kidney diseases,but their low sensitivity is not conducive to the early diagnosis of renal diseases.At the same time,in clinical practice,the subtyp diagnosis of nephrotic syndrome mainly relies on renal biopsy,but it has disadvantages such as persistent hematuria,subcapsular hematoma,and infection at the puncture site.Patients with obvious coagulation dysfunction,isolated kidneys,and hypertension are prohibited from using renal biopsy for detection.In addition,glomerular filtration rate and proteinuria can reflect the progression of the disease to a certain extent,but they are easily affected by factors such as diet and body status,and cannot reflect the change of organism metabolism.It can be seen that there is no better early and pathological diagnostic method for nephrotic syndrome,and it is imperative to screen early diagnostic markers with high sensitivity for nephrotic syndrome,and it is also of great significance to search for pathological diagnostic biomarkers with small side effects and progressive biomarkers that can directly reflect the pathological state of nephrotic syndrome.Objective:(1)Screening for potential pathological biomarkers and potential progression biomarkers in the urine of patients with nephrotic syndrome.(2)Screening for potential pathological markers in serum of rats with nephrotic syndromeMethods:(1)Differential metabolites in the urine of healthy controls and patients with nephrotic syndrome(including membranous nephropathy and Ig A nephropathy)were screened and identified based on non-targeted metabolomics techniques.The metabolites that are not common between membranous nephropathy and Ig A nephropathy and meet AUC>0.5 were selected as their potential pathological biomarkers.At the same time,based on the non-targeted metabolomics technology,the differential metabolites in the urine of the healthy controls and uremic patients were screened and identified,and the metabolites that were significantly linear with the glomerular filtration rate were selected as potential progress biomarkers of membranous nephropathy or Ig A nephropathy into uremia.(2)The structural,composition,abundance and function of intestinal microflora in the colon contents of nephrotic syndrome model rats were analyzed by 16 s r DNA high-throughput sequencing technology,so as to analyze the flora changes of nephrotic syndrome model rats and screen potential flora markers.(3)Based on the non-targeted metabolomics,the changes of polar metabolites and non-polar metabolites in the serum of rats with nephrotic syndrome were analyzed.The differential variables between the control group and the nephrotic syndrome model group were screened based on the VIP and P values and identified through the HMDB online database combined with the mass spectrometry fragmentation rule of metabolites.Finally,the differential metabolites were focused according to the potential microbiota markers,and the potential pathological markers of nephrotic syndrome rats were obtained.Results:(1)Indolelactic acid,isoleucylproline,Dl-Indole-3-lactic acid,D-phenylalanine and L-tryptophan were identified as potential pathological biomarkers for membranous nephropathy and Ig A nephropathy.Alanylleucine,9-Decenoylcarnitine,gluconic acid,capryloylglycine,and sebacic acid were identified as potential progress biomarkers from membranous nephropathy to uraemia,and alanylleucine,9-Decenoylcarnitine,capryloylglycine,and sebacic acid were identified as potential progress biomarkers of Ig A nephropathy to uraemia.(2)In nephrotic syndrome rats,the Chao1 index of intestinal flora decreased significantly,indicating that the richness of intestinal flora was significantly reduced.The results of principal co-ordinates analysis and non-metric multilateral scaling showed that the control group and the model group could be completely separated,indicating that the structure and composition of the intestinal flora between the two groups had changed.Compared with the control group,the relative abundance of 15 microbiota in nephrotic syndrome rats changed significantly,among which 10 intestinal bacteria such as Firmicutes,Bacilli and Lactobacillales were significantly associated with the occurrence and development of the disease.In addition,36 metabolic pathways including L-tryptophan biosynthesis,superpathway of L-tyrosine biosynthesis,superpathway of L-phenylalanine biosynthesis and L-ornithine biosynthesis had undergone extremely significant changes(P<0.001)due to intestinal flora disorder.(3)This study screened 126 differential metabolites in the serum of control group and nephrotic syndrome model rats,and 46 metabolites meet AUC>0.85 and FC>1.5 or FC<0.8.There are 13 metabolites with an absolute value of correlation coefficient with potential microbial markers greater than 0.7,including LPC(17:0/0:0),LPC(22:5/0:0)and PC(O-16:0/20:5)which were preliminarily identified as serum potential pathological markers for nephrotic syndrome.Conclusion: In this study,the potential pathological type biomarkers of nephrotic syndrome and progression biomarkers of nephrotic syndrome to uraemia were screened based on kidney disease patient urine untargeted metabolomic techniques.Among them,potential pathological type biomarkers included indole derivatives and amino acid metabolites.The potential progression biomarkers of mainly involved amino acids and organic acids.At the same time,the potential bacterial markers were screened based on nephrotic syndrome model rats,the abundance of 10 intestinal bacteria,including Firmicutes,Bacilli and Lactobacillales,showed significant changes.Finally,the potential pathological markers in the serum of nephrotic syndrome model rats were focused combined with microbial markers,including 13 glycerophospholipid metabolites such as LPC(17:0/0:0),LPC(22:5/0:0),and PC(O-16:0/20:5).