Investigation Of Long Noncoding RNA In Urine Exosome Contributing To Early Diagnose Of Renal Cell Carcinoma

BACKGROUND & OBJECTIVERenal carcinoma is one of the few tumors with increasing incidence in the whole world,which has no significant clinical manifestation in early stage.It can not be directly detected by body fluid tumor markers without the help of ultrasound and CT imaging.About 30-40% of patients with kidney cancer have already been early distant metastasized at the time of diagnosis,whose 5-year survival rate is quite different from patients with early renal cell carcinoma.Therefore,early diagnosis and screening of renal cell carcinoma is of extremely clinical importance.The aim of this study was to investigate the effect of Lnc-RNA in urinary exosomes as a new diagnostic marker for early renal cancer.The exosome is a membranous vesicle with a diameter of about 30-120 nm,which is released into the extracellular microenvironment by fusion of the multelficular body（MVB）with the cell membrane.Exosomes contains protein and a variety of RNA and other ingredients.Including tumor cells,a variety of cells will secrete exosomes,which can be detected in the blood,urine,saliva,milk and other body fluids.Long non-coding RNA（LncRNA）is a non-coding RNA with a length greater than 200 nucleotides.Existing studies suggest that Lnc-RNA has a good potential as biomarker development.Methods1)Select 10 patients with renal clear cell carcinoma and 10 age,gender matched normal controls,collect their morning urine 50 ml.2)The exosomes was isolated by ultracentrifugation,and the obtained exosomes were examined by transmission electron microscopy,particle size distribution,protein markers（CD63、ALIX、TSG101）and so on.3)LncRNA,which was differentially expressed in urine of patients with renal clear cell carcinoma and normal controls,was analyzed by Next Generation of Sequence.Differently expressed LncRNAs were choosed according to the difference of multiple and P value.4)The most significant LncRNA was further validated in kidney cancer cell lines.5)Analyzed target LncRNA expression levels with more urinary exosomes samples.7)Make the ROC curve of the target LncRNAs,to evaluate its sensitivity and specificity and evaluated their clinical diagnostic values.Results1)Urinary exosomes can be successfully separated and enriched with the ultracentrifugation method.2)We have successfully screened LncRNA NR₀40448 and NR₀33390 in urinary exosomes,could probably indicate the early diagnosis of renal cell carcinoma,which are highly expressed in mature renal carcinoma cell lines and urinary exosomes of patients with RCC.ConclusionLncRNA NR₀40448 and NR₀33390 in urinary exosomes are promising markers for the early diagnosis of renal cell carcinoma.Their sensitivity and specificity also require larger samples of clinical data to verify.