Study Of The Characteristics Of The Esophageal Cancer Cells Expressing The P75^NTR In The Nucleus In Human Esophageal Squamous Cell Carcinoma

Objective To investigate the stem cell characteristics of p75^NTRTR nuclear positive cells in human esophageal carcinoma tissue.Methods 1.Nuclear positive expression of p75^NTRTR in esophageal squamous cell carcinoma samples was detected by immunohistochemistry and the correlation between the number of p75^NTRTR nuclear positive cell with various prognostic factors of esophageal cancer was analyzed by T-test.2.Eca109 cells were cultured by serum-free pellet-forming method to enrich the first generation tumor stem cell,which were then subcultured repeatly to obtain the third,fifth,seventh and ninth generation of tumor stem cells.3.The expression of p75^NTRin pellet-forming cells of different generations was detected by immunofluorescence.4.The cell cycle of pellet-forming cells of different generations was detected by flow cytometry.5.The expression of ABCG2 and ERCC1 in the pellet-forming cells was detected by Real time-PCR.6.CCK8 assay was used to detect the pellet-forming cells resistance to the chemotherapeutic drug.7.The tumorigenesis of pellet-forming cells of different generations was detected by animal tumorigenesis assay in vivo.Results 1.Among the 71 cases of esophageal cancer tissue specimens,42 cases of esophageal cancer cells expressed p75^NTR,and the expression of p75^NTRTR in a few of the tissues with positive expression of p75^NTRTR was in nucleus and the cells expressing p75NTR in nucleus is clustered.Statistical analysis showed that the number of cases expressing p75^NTRwas related to age,tumor diameter and pathological grade,but was not correlated with gender?31/42 in male,11/42 in female?,Ki67 expression?25/42?and distant metastasis?12/42?The cancer tissue with cancer cells positive expression of p75^NTRTR in nucleus were associated with the patient's age,tumor diameter and pathological grade,while there was no significant correlation with other relevant factors,such as gender?male 25/31,female 10/11?and lymph node metastasis?1/7?.The proportion of the cells withp75^NTRTR positive nuclear expression before the first generation of pellet-forming cells increased with the increase of pellet-forming cell passage,but was all positive nuclear after the fifth generation.2.The number of G0/G1cells increased with the increase of pellet-forming cell passage before 5th passage,but did not change significantly after the 5th passage.3.The expression levels of ABCG2 and ERCC1,and the tolerance to methotrexate,cisplatin and paclitaxel,and the tumorigenic ability of the pellet-forming cell increased gradually before the 5th passage,but did not change significantly after the 5th passage.Conclusion p75^NTRTR nuclear positive cells of esophageal cancer have the characteristics of stem cells,and we can get the pure esophageal cancer stem cells expressingp75^NTRTR in the nucleus with serum-free culture pellet-forming cells.