Studies On Circulating MDSCs Of Primary Hepatic Cancer Patients And Gemcitabine Induced Autophagy Of Hepatocellular Carcinoma HepG2Cells

Primary hepatic cancer is the third fatal tumor disease and is one of China’s treatment-resistant refractory. Primary hepatocellular carcinoma accounts for more than90%of it.In primary hepatic cancer occurrence, development and treatment processes are related to immune responses, and thus the associated immune messengers may be useful tumor marker. Additionally, the efficiency of primary hepatic cancer chemotherapy is not satisfactory. Cell autophagy is closely related to the tumor occurrence, development, treatment, and has been the tumor research focus in recent years.For HCC the above two key problems will be addressed " To Detect the Circulating MDSCs in Peripheral Blood of Primary Hepatic Cancer Patients" and " Study on Gemcitabine Induced Autophagy of Hepatocellular Carcinoma HepG2Cells". Its characteristic is that peripheral blood detection is a noninvasive procedure and multiple material collections can be easily achieved with dynamic observation attainable, the MDSCs are related with tumor immune escape; Gemcitabine autophagy mechanism study of HCC has not been reported. Part OneTo Detect the Circulating MDSCs in the Peripheral Blood of Primary Hepatic Cancer PatientsObjective:Myeloid derived suppressor cells (MDSCs) can be induced or expanded in tumor-bearing mice and cancer patients. The frequency of MDSCs denoted here as Lin-/low CD33+HLA-DR-was investigated in hepatocellular carcinoma (HCC) patients. The clinical relevance of MDSCs and patients’characteristics were examined. And MDSC-related immune regulatory pathway in these patients were discussed.Methods:The population of MDSCs was tested in peripheral blood of patients with HCC (n=63) and healthy donors (n=56). The expressions of EFN-γ, VEGF, COX-2, MMP-13, NOS-2and ARG-1were analyzed. Coculturing with anti-CD3/CD28-stimulated T lymphocytes was used to determine the suppressive effect of MDSCs on the T lymphocytes.Results:Patients with treatment-naive HCC had an increased subpopulation of Lin-/low CD33+HLA-DR-cells in the PBMCs with characteristics of MDSCs and associated to the stage (P=0.0004). Patients with splenomegaly had a higher frequency of circulating MDSCs. Also COX-2, MMP-13and VEGF were expressed differently associated with the alteration of MDSCs.Conclusion:Our study provide evidence showing an increased population of Lin-/lowCD33+HLA-DR-MDSCs in the peripheral blood of HCC patients. Our data also suggest that MMP-13and COX-2in PBMC may play a new important role companied with MDSCs in HCC patients. Part TwoA Study on the Gemcitabine Induced Autophagy of Hepatocellular Carcinoma HepG2CellsObjective:Study the effects of gemcitabine on hepatocellular carcinoma HepG2cells induced autophagy and molecular mechanism.Methods: The cellular responses of HepG2cells after gemcitabine treatment are correlated with microtubule associated protein1light chain3(MAP1-LC3) changes and autophagic vacuoles, which are observed by western blots and transmission electron microscopy (TEM). The anti-apoptotic protein Bcl-XL levels are shown through western blots; and autophagy inhibition with chloroquine or si-BECN1interference is utilized to study the cytotoxic effects of gemcitabine.Results:Gemcitabine effectively inhibit the proliferation of hepatocellular carcinoma HepG2cells in a time and concentration-dependent manner. Gemcitabine also induced enhanced autophagy level of HepG2cells and the anti-apoptotic protein Bcl-XL level increased significantly.Conclusion:The inhibition of gemcitabine-induced autophagy can promote the toxicity of gemcitabine on hepatocellular carcinoma HepG2cells and the inhibition of HepG2cellular proliferation. Clinically, the efficacy of incorporating autophagy inhibition treatment remains to be further verified.