The Expression And Effects Of Homeobox Gene HOXA10 On The Proliferation And Invasion Of Ovarian Clear Cell Adenocarcinoma

Homeobox gene HOXA10 belongs to HOXA cluster of homebox family. HOXA10 was reported to participat in the development of female gential tract and was found to express in normal endometrium and vagina muliebris. Recently,it has been found that HOXA10 may be involved in the carcinogenesis of ovarian carcinoma generating from endometriosis(EMs) besides its effect on endometrium epithelial differentiation and stroma decidualization.The cellular structure and immunophenotype of ovarian clear cell adenocarcinoma(OCCA) is very similar with those of endometrium and vagina clear cell adenocarcinoma.Most scholars considered OCCA came from the Muller's duct.According to the high percentage(25%—55%) of EMs found in the OCCA patients,a part of OCCA was considered to come from the malignant EMs and atypical endometriosis(AEMs) which could be a kind of precancerous lesion.But little is known about the relationship between HOXA10 in malignant EMs and ovarian carcinogenesis.Our experiments are divided into two parts as follows:(1) observeing the expression of homebox gene HOXA10 in ovarian clear cell adenocarcinoma and endometriotic tissues and its correlative analysis of clinical prognosis.(2) Establishing HOXA10 expression ES-2 cell line,an ovarian clear cell adenocarcinoma cell line,and investigating the effects of HOXA10 on proliferation,motility and invasion.SectionⅠExpression of HOXA10 in ovarian clear cell adenocarcinomaObjective To examine the expression of HOXA10 in ovarian clear cell adenocarcinoma,normal ovarian,normal endometrial,ovarian endometriotic and corresponding endometrial tissues in mRNA and protein level.The relationships between expression of HOXA10 and its correlative clinical prognosis were analyzed.Methods RT-PCR was used to detect mRNA expression level of HOXA10 in ovarian clear cell adenocarcinoma,normal endometrial,ovarian endometriotic and corresponding endometrial tissues. Immunohistochemistry staining was used to investigate the expression of HOXA10 protein in ovarian clear cell adenocarcinoma,normal ovarian, normal endometrial,ovarian endometriotic and corresponding endometrial tissues.Results(1) The expression of HOXA10 gene was found in ovarian clear cell adenocarcinoma tissues but not in ovarian endometriosis.Different expression of HOXA10 was found in corresponding normal endometrium in different menstrual cycle.The menstrual cycle of positive expression patients were most in ovulation period and were confirmed to be in secretory phase by pathematology detection.HOXA10 expression of endometrium in patients without endometriosis was the same as that in patients with endometriosis.(2) HOXA10 gene expression level was positively correlated with the stage and grade of ovarian cancers(P＜0.05),but was negatively correlatated with five year survival rate (R=-0.442,P=0.043).Conclusions Overexpression of HOXA10 may be related to the tumorigenesis of epithelial ovarian cancer and canceration of EMs.SectionⅡThe effects of HOXA10 on proliferation,motility and invasion of ovarian clear cell adenocarcinoma ES-2Objective To evaluate the effects of HOXA10 on proliferation,motility and invasion of human ovarian clear-cell carcinoma ES-2.Methods A stable HOXA10-expressed ES-2 cell line was established and the high positive expression clone was screening out.Enhancement of HOXA10 expression was confirmed by Western blot.ES-2 cells were divided into two groups:HOXA10-ES-2 and pcDNA-ES-2.Cell proliferation of each group were evaluated by Sulforhodamine B assay.The motility and invasion of two groups were evaluated by Wound assay and transwell.Results(1) The stable HOXA10 expression clones of ES-2 cells were established by Lipofectamine transfection.The transfection efficiency was satisfactory when ratio between plasmid and Lipofectaminewas 1:2.5.(2) The difference of cell proliferation rates between HOXA10 transfected groups and negative control groups cell was significant(0.446±0.0024 vs 0.281±0.002,P＜0.01) by SRB assay.(3) Using wound healing assays,cell motility for HOXA10 transfection groups were faster than that of empty groups at the fifth hour after scratch.(0.47±0.07mm vs 0.25±0.03 mm,P＜0.05).(4) The invasion activity of HOXA10 transfection cells were significantly higher compared with those of control cells(446±14 vs 227±12,P＜0.01) by transwell.Conclusions HOXA10 could stimulate the carcinogenesis of ovarian clear cell adenocarcinoma by up-regulation proliferation,motility and invasion of cells.The present study has shown that(1) Homebox gene HOXA10 is over-expressed in ovarian clear cell adenocarcinoma.A reverse correlation between prognosis and HOXA10 expression exists in ovarian clear cell carcinoma.(2) HOXA10 could stimulate the carcinogenesis of ovarian clear cell adenocarcinoma by induce the proliferation,motility and invasion of cells.