Research On Regulation Of Lipid Metabolism Of Hepatocellular Carcinoma Cells By CC3/TIP30

Aims : Hepatocellular carcinoma (HCC) is one of the most common malignancies, its occurrence and development is a multi-factor involved in the complex regulatory process. Tumor lipid metabolism provide the necessary energy to the rapid growth of tumor cells for proliferation and we also noticed that lipid reprogramming has been considered as an important characteristic in the development of HCC. However, the detailed molecular mechanisms have yet to be clearly defined.We found that up-regulation of CC3 / TIP30 expression could inhibit the expression of lipogenic enzymes FASN, SCD and SREBP1 who is one of the key transcription factors regulating liver lipid metabolism ,that is in turn reducing lipid metabolism levels and inhibiting tumor growth. Thus revealing the molecular mechanism of lipid metabolism regulation in HCC, bringing anew ideas for targeting lipid metabolism of tumor diagnosis and treatment.Methods: In this study, the expression of CC3 / TIP30 was successfully upregulated and overexpressed by lentivirus packaging plasmid.The result of CC3 /TIP30 was involved in the metabolism especially in the fatty acids metabolism of hepatocarcinoma cells by using RNA-Seq chip analysis,that is verifird by using quantitative real time polymerase chain reaction(Q - PCR) technology. The key enzymes that regulate the metabolism of fatty acids were detected by Q-PCR and western blotting (WB).The technology of immunofluorescence and WB demonstrated that CC3 / TIP30 increased phosphorylated AKT and mTOR expression, enabling AKT pathway activation to increase SREBP1 expression and nuclear aggregation?Using MTS and plate colony experiments confirmed the way through CC3 / TIP30 and SREBP1 regulation of lipid metabolism can affect the proliferation and growth of HCC cells in vitro. Nude mice subcutaneously transplanted tumor model established revealed that lipid metabolism disorder can affect the growth of the nude mice tumor .IHC was used to explored the relationship between lipogenic enzymes and CC3 / TIP30. 80 HCC patients' tissue sample were collected for finding the relative expression of TIP30 / CC3 and SREBP1 on the prognosis of HCC patients using the method of IHC.Results : In this study, the expression of CC3 / TIP30 was successfully upregulated and overexpressed by lentivirus packaging plasmid. The result of CC3 /TIP30 was involved in the metabolism especially in the fatty acids metabolism of hepatocarcinoma cells by using RNA-Seq chip analysis.We verified the conclusion by using Q-PCR and we also found that CC3 / TIP30 regulates the lipid metabolism in HCC mainly by regulating the synthesis of fatty acids. Down-regulation of CC3 /TIP30 resulted in increased of lipid droplet uptake and increased triglycerides in HCC cells. Using Q-PCR and immunoblotting, It was confirmed that SREBP1 could be induced by the expression of CC3 / TIP30 and guide the expression of FASN and SCD in the lipid metabolic regulation.Down-regulating CC3/TIP30 activated the Akt/mTOR signaling pathway to up-regulate the expression of SREBP1, which in turn directly activated the transcription of the major lipogenic genes FASN and SCD to promote de novo lipogenesis.We also noticed that the lipid droplets and triglycerides stored in HCC cells increased obviously. This conclusion can be reversed by the blocking effect of AKT inhibitor.Down-regulation of CC3 / TIP30 on SREBP1-mediated Lipid Metabolism Pathway Promotes HCC Cell Growth. The method of MTS to detect HCC cell proliferation and plate colony detection of cell growth have shown that downregulation of SREBP1 can inhibit the reduction of CC3 / TIP30 cells caused by the proliferation and growth of hepatocellular carcinoma in vitro. The expression of CC3 / TIP30 and SREBP1 was observed by lentivirus in nude mice subcutaneously transplanted model and the different down-regulation combinations were grouped according to the experiment. It was confirmed that the expression of SREBP1 in vivo blocked the tumor growth caused by CC3 / TIP30 deficiency .That also can be causing the same trend of triglyceride levels in tumor tissue. The expression of SASBP1 and FASN and SCD in nude mice were confirmed to be inversely proportional to the expression of CC3 / TIP30 by IHC analysis.Analysis of the expression of SREBP1 and CC3 / TIP30 proteins can be used to assess the survival and prognosis of patients with HCC more accurately. Our work show that the combined expression of SREBP1 and CC3 / TIP30 proteins can be used to predict the survival and prognosis of HCC patients by collecting and combining the clinical data of patients.Conclusion : TIP30 negatively regulates lipid metabolism in hepatocarcinoma cells. SREBP1 is essential for CC3 / TIP30 deficiency to promote lipogenesis and is regulated by Akt / mTOR pathway. Down-regulation of CC3 / TIP30 action on SREBP1-mediated lipid metabolism pathway promotes HCC cell growth. TIP30 expression is negatively associated with SREBP1 expression in tumor tissues of HCC patients and the combinational biomarkers provide powerful prognostic value for HCC. This new mechanism of CC3 / TIP30 regulates SREBP1-directed lipid metabolism may becomea new target for the treatment of HCC.