Studies On The Characteristics Of HepG2 Stem Cells Induced By Hypoxia Inducible Factor 1?

Background Liver cancer is one of the most common malignant tumors in china.The treatment of liver cancer is still mainly surgical,but only some patients can receive surgical treatment.Although the treatment technology has been improving in recent years and the quality of life has improved after treatment,the survival rate has not improved significantly.Therefore,exploring the development of liver cancer,its exact mechanism and effective treatment is still the key point of basic and clinical research.Studies have shown that the onset and development of tumor are related to a small group of cancer stem cells with multidirectional differentiation and self-renewing abilities in tumor tissues,which are the sauce of tumorigenesis,metastasis and recurrence.Many scholars at home and abroad have proved that there are tumor stem cells in many solid tumors,including liver cancer,this study provides a new way to explore the pathogenesis and effective treatment of liver cancer from the perspective of cancer stem cells(CSCs).Hypoxia is a common phenomenon in solid tumors.Pathological study showed that as a lack of blood supply tumor tissue,there is a large area of anoxic zone in liver cancer tissue.Oxygen deficiency can cause a series of changes in molecular biology,cytology and physiology,and make tumor cells adapt to the hypoxia microenvironment while increasing their own invasiveness and resistance to treatment.In this process,the transcription factor hypoxia inducible factor-1?(HIF-1?)acts as a central link.HIF-1?,as a transcription factor,can regulate the expression of more than 100 genes.By regulating a series of downstream target genes and corresponding protein products to enhance the adaptability of cells to hypoxia,in order to facilitate tumor growth.Tumor stem cells are a cell population in tumor tissues and are also in an hypoxic microenvironment.So whether HIF-1? induces proliferation and differentiation of tumor stem cells and whether hypoxia-mediated therapeutic resistance is related to tumor stem cells become our research point.In this study,high expression plasmid HIF-1? was proposed to be introduced into HepG2 cell line to analyze whether they are related to tumor stem cells,in order to provide theoretical basis for HIF-1? targeted liver cancer therapy.Objective To investigate the effect of HIF-1? on the characteristics of hepatocellular carcinoma cells HepG2,and to analyze its resistance to chemotherapeutic drugs.Methods 1 The plasmids were introduced into HepG2 cells using the LIPO3000 kit and divided into three groups: HepG2 blank group,no-load control group(HepG2-pcDNA)and experimental group(HepG2-HIF-1?).2 Collect positive cells containing HIF-1? plasmids by G418 screening.3 The expression of HIF-l? mRNA in three groups of cells was detected by qRT-PCR kit.4 The content of HIF-1? protein was detected by Western blot(Western blot test).5 After adding 0?5?10?25?50?mol/L of antitumor drug epirubicin,the cell proliferation and cell activity were detected by MTT method to study the effect of different expression levels of HIF-1? on its efficacy.6 Detection of apoptosis after 50?mol/L of epirubicin which had a significant effect on the three groups by flow cytometry.7 Detection of expression of cell marker CD133 by flow cytometry.Results 1 qRT-PCR showed that the amount of HIF-1? mRNA in HepG2-HIF-1? group was significantly higher than that in HepG2 group and HepG2-pcDNA group.2 Western blotting showed that HIF-1? protein expression level in HepG2-HIF-1? group was significantly higher than that in HepG2 group and HepG2-pcDNA group.3 The results of MTT showed that the value-added activity of the cells in the three groups decreased significantly with the addition of different concentrations of epirubicin,and the proliferation activity of the cells in HepG2-HIF-1? group was significantly higher than that in HepG2 group and HepG2-pcDNA group at the concentrations of 25 and 50?mol/L.4 Under the action of 50?mol/L epirubicin,the apoptosis rate of HepG2-HIF-1? group was significantly lower than that of HepG2 group and HepG2-pcDNA group.5 Flow cytometry showed that the positive expression of CD133 in HepG2-HIF-1? group was 20.11% higher than 0.04% in HepG2 group and 0.03% in HepG2-pcDNA group.Conclusion High expression plasmid HIF-1? can induce HepG2 cells to produce more CD133 positive tumor stem cells,enhance the proliferation of tumor cells,and strengthen the resistance of tumor stem cells to the chemotherapeutic drug epirubicin.