Enrichment Pattern Of Liver Cancer Stem Cells In Vitro Based On The Resistancecharacteristics And Holoclone

Background:cancer stem cells (CSCs) refers to the tumor tissue,very little has unlimited self-renewal potential and real tumorigenic ability is the most important feature of the tumor cell population. The most common malignant tumor in China liver cancer, malignant progression and high degree of disease quickly, disease after the survival of only 6 months is called "the king of cancer", its incidence has accounted for 40% of cases worldwide, it was included in the focus of prevention and treatment of illness. How can a liver cancer stem cell isolation and purification of active and effective, has been the focus of academia and research focus the research group, select the hepatoblastoma cell line HepG2 as the research object, which is derived from a 15 year old white liver cell adenoma tissue, as may be the earliest isolated and clear background and advantages of phenotypic stability, has become widely used in hepatocytes The cell lines used in liver cancer research field. According to a single cell clone enrichment method combined with the advantages of clinical broad-spectrum cytotoxic anticancer drugs, adriamycin intervention, the cloning of hepatocellular carcinoma cells will get the xenograft experiments to detect the tumor formation, in order to establish a modified single cell clone of hepatocellular carcinoma with adriamycin the enrichment of stem cells. The intervention mode of study found:single cell clone culture dilution method to get the total tumor formation rate of 100% cloned cells in nude mice, and the single cell clone in different concentrations of adriamycin intervention under the mode of training, the total ratio is not the same as the cloning and tumor formation in nude mice was also different. Research shows that single cell culture. The method of cloning and adriamycin intervention may become a great improvement of liver cancer new stem cells.Objective:To observe the formation of three clones and adriamycin intervention under all clones in nude mice in vivo tumor cultured human hepatic blastoma HepG2 individual cells and study based on cancer stem cells resistant properties and in vitro single cell culture of the clones, the establishment of a simple of liver cancer stem cells in vitro enrichment pattern.Methods:human hepatoblastoma cell line HepG2, with limited dilution method in vitro single cell clone culture system, the following two part:1 to observe the cloning of single cell culture in vitro growth curve and the formation of the whole part of cloning, cloning and cloning of three clones beside the proportion after the selected clone step by step amplified experiment mode finally take 106 HepG2 cells in nude mice subcutaneous xenograft experiment.2. adriamycin intervention HepG2 cancer cells clone:1. The single cell clone culture, and then only for the whole cell clone plus 0.5 g/mL,0.3 g/mL and 0.2 g/mL,,0.1 g/mL,48hrs 0.05 and g/mL adriamycin intervention 0.01 g/mL concentration gradient, the observation of clonal growth; all step by step amplification and cloning of nude mice for survival. After the intervention. The first plus 0.1 g/mL,0.05 g/mL and 0.01 g/mL concentration gradient 48hrs cells were induced by adriamycin in HepG2 cells, and the clones were formed by single cell clone culture. Some clones were amplified by step by step and nude mice were implanted with tumor.Results:1 HepG2 cells in single cell culture can form a whole part of cloning, cloning and cloning by three kinds of cancer cell expansion growth pattern, the total colony formation rate was (32.78+9.03)%, which is composed of all clones cloning (7.12+2.96)% and (62.67+13.39) partial cloning humancloning%, next the percentage of (30.21+14.87)%; all clones can be amplified in 106 cells and step by step, the level of the tumorigenicity rate was 100%(8/8).2. The first adriamycin intervention single cell clone culture, found only 0.05 g/mL and 0.01 g/mL group total clone survival and proliferation, and step by step 100%(3/3,8/8). The tumor was full of adriamycin intervention single cell culture after cloning, found only 0.1 g/mL,0.05 g/mL and 0.01 g/mL group colongenic, and stepwise Amplification Culture and 100%(5/5,5/5,5/5) tumor.Conclusion:HepG2 cells cultured in vitro with a single cell culture and the growth and tolerance of doxorubicin induced killer cells, most likely to be rich in liver cancer stem cells, which may be a simple and effective in vitro tumor stem cell enrichment model.