The Study Of Tongmai Dasheng Pill Regulation On The TGF-β1/Smads Path Of The Ovary Granulose Cells From Female Rat In Vitro

Objective:Based on previous studies, in vitro cultured rat ovarian granulosa cells as a carrier, to study the influence of Tongmai Dasheng Tablets on TGF-β/Smads Signal transduction pathways of ovarian granular cells in rats,in order to confirm it act on the pathway, promoting follicular development, treating anovulatory infertility and laying the basis of experimental studies for drug research of TCM.Methods:Rat ovarian granulosa cells were cultured in DMEM/F12medium and put in37℃,5%CO2incubator.The cells were randomly divided into five groups: control group, high-dose group, medium-dose group, low-dose group and FSH group. Medicine was given in logarithmic growth phase and liquid supernatant was taken respectively1hour,2hours,6hours,12hours,24hours and48hours afterwards. The expression of TGF-β and TGF-β1R in the supernatant was measured by ELISA. Rat ovarian granulosa cells seeded were taken respectively6hours,12hours and24hours afterwards in each dose group of Tongmai Dasheng Tablets.The cells seeded were taken respectively1hours.2hours and6hours afterwards in FSH group.The cells seeded were fixed with formaldehyde,then the expression of smad3/4/7was measured by immunohistochemistry.The data was analyzed by SPSS13.0software.Results:1. There was no significant change on expression of TGF-β1and TGF-β1R in control group among different timing of medication. The expression of TGF-β1and TGF-β1R reached the peak in12hours after medication and declined afterwards to the lowest level in48hours in high-dose group, medium-dose group and low-dose group. The expression of TGF-β1reached the peak in2hours after medication and declined afterwards to the lowest level in48hours in the FSH group.There was no significant difference on peak expression of TGF-β1and TGF-β1R in medium-dose group and FSH group. There was no statistically significant(p>0.05).2. There was no significant change on the nuclear positive rate of Smad3in control group among different timing of medication. The nuclear positive rate of Smad3reached the peak in12hours after medication in high-dose group, medium-dose group and low-dose group,and it was highest in medium-dose group(p<0.05). The nuclear positive rate of Smad3reached the peak in2hours after medication in FSH group.There was no significant difference on peak nuclear positive rate of Smad3in medium-dose group and FSH group. There was no statistically significant(p>0.05).3. There was no significant change on the nuclear positive rate of Smad4in control group among different timing of medication.The nuclear positive rate of Smad4reached the peak in12hours after medication in high-dose group, medium-dose group and low-dose group.and it was highest in medium-dose group(p<0.05). The nuclear positive rate of Smad4reached the peak in2hours after medication in FSH group.There was no significant difference on peak nuclear positive rate of Smad4in medium-dose group and FSH group. There was no statistically significant(p>0.05).4. There was no significant change on the nuclear positive rate of Smad7in control group among different timing of medication.The nuclear positive rate of Smad7reached the least in12hours after medication in high-dose group, medium-dose group and low-dose group,and it was least in high-dose group(p<0.05). The nuclear positive rate of Smad7reached the least in2hours after medication in FSH group.There was significant difference on least nuclear positive rate of Smad7in high-dose group and FSH group. There was statistically significant(p>0.05).Conclusion:l.Tongmai Dasheng Tablets can increase the expression of TGF-β1and TGF-(31R in rat ovarian granulosa cells.2.Tongmai Dasheng Tablets can increase the expression of Smad3/4in rat ovarian granulosa cells and reduce the expression of Smad7.3.The expression of smad3/4/7dependented the concentration of TGF-β1...