The Role And Mechanism Of PGC1A/UCP1 Mediated Lipid Browning In Renal Clear Cell Carcinoma

Renal Cell Carcinoma(RCC)is one of the most common malignant tumor in urinary system,Renal Clear Cell Carcinoma(cc RCC)is the most common and lethal Cell type,which has a characteristic of strong death,invasion,metastasis and drug resistance.Clinical studies have shown that some of RCC were already had distant metastases when it was diagnosed.Surgery was the main way to treat early RCC,and advanced renal cancer was mainly treated with drugs which were targeted at VEGF and m TOR.To sum up,due to the high degree of malignancy,easy to miss the diagnose and metastasis of RCC,new prognostic molecules of renal cancer,new treatment directions and strategies need to be further studied.Lipid droplets accumulation and abnormal lipid metabolism are the most common features in cc RCC cells,which can be considered as a metabolic disease.At present,the mechanism of abnormal cc RCC lipid metabolism and its causality and biological significance in cc RCC development are not studied systematically.In order to further understand the pathogenesis and develop new treatment of RCC,we explored biological function when overexpressing or downregulating of related metabolic gene in cc RCC,and explore the mechanism of abnormal lipid accumulation in this study.This reserch is divided into three parts:Part Ⅱ: Screening and expression of lipid metabolism related genes and clinical value in renal clear cell carcinomaObjective: to explore the expression of lipid browning gene in cc RCC and to determine whether the target gene has the role of tumor markers and its clinical value.Methods: Screening genes according to the classification of gene: lipid,mitochondria,thermogenesis and beige related genes,and then use Oncomine database(https://www.oncomine.org)and The Cancer Genome Atlas(TCGA)database(https://cancergenome.nih.gov)to demonstrates PGC1 A expression in renaltissue and cancer tissue,and the relation between clinical stage,pathological and prognosis.Western and real-time quantitative PCR(RT-PCR)was used to detected the expression of PGC1A(peroxisome proliferator activated receptor γ coactivator-la,PGC1A)in renal cell lines,and western,real-time quantitative PCR(RT-PCR)and immunohistochemistry(IHC)was used to detected the expression of PGC1 A in clinical samples of cc RCC.Results: PGC1 A is one gene in classification of lipid,mitochondria,thermogenesis and beige related genes,and the expression in the TCGA and Oncomine database have obvious difference in carcinoma and beside(p < 0.0001),m RNA and protein levels were low expression in cc RCC clinical samples and cell line,lipid of brown related gene expression in TCGA database were lower and there have a positive correlations,the expression of PGC1 A had a statistically significant with cancer clinical stage,pathological and prognosis.Conclusion: The expression of lipid brown gene PGC1 A in cancer tissue is low,and it will be a new and excellent diagnostic and prognostic marker of cc RCC.Part Ⅱ: The inhibition role and mechanism of PGC1 A in the renal cell carcinoma.Objective: to explore the specific role and the mechanism of lipid browning gene PGC1 A in cc RCC and the biological effect after PGC1 A in vitro.Methods: Lenti virus infection build overexpressed PGC1 A in cc RCC cell lines A498(A498-PGC1A/A498 – NC)and Caki-1(Caki-1-PGC1A/ Caki-1-NC),q RT-PCR to detect PGC1 A expression in PGC1 A stable expression cell lines,western and q RT-PCR detected the expression of PGC1 A and UCP1.Whether high expression of PGC1 A can affect UCP1 and lipid metabolism or not.Transfection of small RNA(si RNA)siUCP1 after express PGC1 A cc RCC cell lines and detect the influence of si-UCP1 in over express PGC1 A in cc RCC cell lines.Transwell Assay and scratch test after highexpression of PGC1 A and si-UCP1 in cell migration and invasion ability and of A498 and Caki-1.The change of lipid content was detected by oil red staining.Results: Overexpression of PGC1 A can promote the expression of UCP1,reduce lipid accumulation within the tumor cells,thus inhibiting renal cancer cell migration ability,silence UCP1 after overexpression PGC1 A and can increase lipid accumulation,and reverse invasion and migration ability of cc RCC cells.Conclusion: The lipid brown gene PGC1 A can promote the browning of renal cell lipids and promote lipid consumption through UCP1,thereby inhibiting the growth,migration and invasion of cc RCC cells.Part Ⅲ: The effect of PGC1 A on the growth and metastasis of cc RCC in nude mice.Objective: To explore the effect of PGC1 A on renal clear cell carcinoma in vivo.Methods: Mouse tail intravenous Caki-1-PGC1A/ Caki-1-NC cells were used to observe tumor invasion and metastasis of cc RCC cells,and then detected with HE staining.Subcutaneous tumor experiment was used for the detection of growth observation of cc RCC cells,immunohistochemical detected the expression of PGC1 A and UCP1.Results: There was a significant decrease in the formation of metastatic tumor in PGC1 A group compared with NC group,the formation of metastatic tumor was detected by HE staining.The formation of subcutaneous tumor was significantly inhibited by PGC1 A group compared with the NC group,and the higher expression of PGC1 A and UCP1 in PGC1 A group which was detected with immunohistochemical.Conclusion: Over express PGC1 A in renal cell carcinoma cells can reduce the hepatic metastasis and inhibit the growth of subcutaneous tumor.