Celecoxib Inhibits Proliferation Of Leiomyoma Cells With Its Molecular Mechanism

Objective:Cyclooxygenase -2(COX-2) inhibitors have a certain degree of anti-tumor effects,one of the selective COX-2 inhibitor celecoxib has been the United States FDA approved the treatment of familial adenomatous polyposis(FAP).In uterine leiomyoma research is not yet perfect,has reported that COX-2 inhibitors can inhibit the uterine leiomyoma cell proliferation.In order to further explore the COX-2 inhibitor on the role of uterine leiomyoma, we cultured human uterine leiomyoma cells,and use selective COX-2 inhibitor Celecoxib in vitro toxicity testing,and to uterine leiomyoma cell signaling pathway related molecules study to examine Cyclooxygenase -2(COX-2) selective inhibitor celecoxib on the uterine leiomyoma cell proliferation and the molecular mechanism of inhibition.Materials and Methods:We collected 15 cases which were been normal human uterus smooth muscles and uterine leiomyoma by the pathologic confirmation to culture in vitro.The cases were hysteromyoma patients whose uterus were excises completely from No.1 Hospital of Shanxi Medical University gynecology and obstetrics department in from March,2007 to June, They are 35-45 years old.Then identified the human uterus smooth muscle cell and uterine leiomyoma cell by means of immunohistochemical method with mouse anti-person desmin and mouse anti-person a-actin.All cells were divided into 3 groups:normal uterine smooth muscle cells groups;uterine leiomyoma cell groups;added drugs uterine leiomyoma cell groups (simply add Celecoxib,final concentration of 12.5,25,50μmol /L),each group has 6 bottles.We observed cytomorphology change with inverted microscope.We carried on the cell counting by means of MTT method,then drawed the growth cures.After cultured 48 hours,we collected all cells and analyzed the expression of Wnt Signaling Pathway by RT-PCR method.Using the ratio of sample with GAPDH,we calculated the content of relative mRNA expression.Duplicate the experiment 3 times and averaged.SPSS13.0 statistical software are used analysis of data,the results of that test are showed by(?)±s and the significance using one-way ANOVA.To P＜0. 05 for the difference was statistically significant.Results:The uterus smooth muscle cells which were digested well are long spindle-shaped or the ellipse.The cells cultured within the first 24h were well spread.They were found to grow into the cell clone about 48 hours.The cell clones fused about a week.The cell shape is the typical long spindle-shaped.The fasciculated smooth muscle cells were parallel permutation. The partial cells were overlap to multilayer.The partial areas were typical "peak -valley".The results by means of immunohistochemical method are anticipated.The dyeing result with a-action is masculine,and the endochylema is brownish yellow,and the cell nucleus is blue.The dyeing result with desmin is negative.The cell nucleus in negative control is blue,and the cytoblastema does not to put color on.From the growth curve,we could see different concentrations of Celecoxib(6.25,12.5,25 and 50μmol / L) role in uterine leiomyoma cells 24,48,72 h after celecoxib on cultured human uterine leiomyoma cells shows proliferation inhibitory effect,and its role in a dose-and time-dependent(24,48,72 h various concentrations of the experimental group compared with the corresponding control group,P＜0.05).Cell culture 3d add drug and added drugs uterine leiomyoma cells were collected after 48h and carry out RT-PCR analysis.Semi-quantitative RT-PCR results showed that uterine leiomyoma cells exist Wnt5b,β-catenin,c-myc gene and the normal myometrium group without these expression. Celecoxib(12.5,25,50μmol / L) affect uterine leiomyoma cells 48 h,will reduce the expression of these genes,and its role is a dose and time-dependent manner,showing apparent activation inhibition.Conclusion:1.Wnt signaling pathway exists in human uterine leiomyoma cells.2. Celecoxib significantly inhibited the in vitro cultured human uterine leiomyoma cell proliferation, with the increase in drug dose and the extension of the time,uterine leiomyoma cell proliferation and activation are inhibited,showing a dose-and time-dependent.3,celecoxib inhibits human uterine leiomyoma cells by the cox-2 pathway and the non cox-2 pathway,the later is achieved by the Wnt signal pathway.Uterine leiomyoma cells Wnt5b,β-catenin,c-myc gene expression was negatively correlated Celecoxib concentrations...