Effects Of Ropivacaine On The Proliferation Inhibition And Apoptosis In Human Cervical Cancer HeLa Cell Line

Objective:To investigate the effects of ropivacaine on proliferation inhibition and apoptosis induction of human cervical cancer HeLa cell line in vitro through Caspase-3 signaling pathway,and to provide experimental basis for further study of the feasibility of ropivacaine used in clinical perioperative period of cervical cancer patients.Methods:The human cervical cancer HeLa cell line was inoculated into the culture plate for 24h and divided into control group(C group)and experimental group(ropivacaine group,R group).The R group was divided into the low concentration group(500μM,LD),the medium concentration group(1 000μM,MD),the high concentration group(1500μM,HD)after drug concentration screening experiment by MTT method.To evaluate the effect of ropivacaine with different concentrations of 24h on the proliferation and cell activity of HeLa cells,MTT assay was used.After DAPI staining,it was observed that the morphological and structural changes of cervical cancer HeLa cell line under fluorescence microscope,and the effects of different concentrations of ropivacaine on early and late apoptosis of HeLa cell line of cervical cancer were detected by flow cytometry.The expression levels of Bcl-2,Bax,Cytochrome C,Caspase-3,Caspase-8 and Cleaved-caspase 8 were detected by Western blot.Results:1.MTT assay showed that there was no significant difference in growth inhibition of cervical cancer HeLa cell lines treated with ropivacaine for 24 hours at 10μM,25μM,50μM,100μM and 200μM(P>0.05),but the activity of HeLa cell line in LD,MD and HD groups was significantly lower than that in C group(P<0.05).The cell inhibition rates in C group,LD,MD and HD groups were 2.0%,26.32%,38.99%and 51.84%,respectively.Compared with C group,the inhibition rate increased significantly with the increase of drug concentration(P<0.05).2.After the HeLa cell lines treated with ropivacaine were stained with DAPI,the morphological characteristics of apoptotic cells such as complete nucleus and plump cytoplasm were observed under inverted microscope in group C.Cell cytoplasm condensation,smaller size,broken nucleus and chromatin condensation were observed in group R.3.Flow cytometry showed that apoptotic cells accounted for 1.9%of total cells in group C,7.8%in LD,9.7%in MD and 12.3%in HD.Ropivacaine increased the apoptotic rate of HeLa cells in a concentration-dependent manner(P<0.05).After treatment with ropivacaine,the distribution of cervical cancer cells changed from living cells to early and late apoptotic cells.In group C,late apoptotic cells accounted for 1.6%,late apoptotic cells in LD group accounted for 6.5%,late apoptotic cells in MD group accounted for 8.8%,and late apoptotic cells in HD group accounted for 11.4%,which was concentration-dependent(P<0.05).4.Western blotting showed that ropivacaine promoted the expression of Caspase-3 in a concentration-dependent manner,increased the expression of Bax,reduced the expression of Bcl-2,induced the release of Cytochrome C,and promoted the expression of Caspase-8 and Cleaved-caspase 8(P<0.05).Conclusion:1.Ropivacaine significantly inhibits the proliferation and cell viability of human cervical cancer HeLa cell line.2.Ropivacaine induces apoptosis of human cervical cancer HeLa cell line in vitro.Its mechanism may be related to the regulation of Caspase-3 signaling pathway.