| Cervical cancer,an epithelial cell carcinoma is the world's most common female malignancy.Cervical cancer is the second incidence of malignant tumors for woman.Furthermore,the incidence of cervical cancer of Chinese woman was the first and the mortality rate in second.Studies have shown that the incidence of cervical cancer is growing at rate of 2%-3% every year.Western countries prevention and control experience shows that the incidence of cervical cancer can be reduced by 70%-90% through easy detection and early screening.Therefore,the study of the molecular mechanism of cervical cancer,screening and prevention of cervical cancer is critical and urgent.We investigated the effect of histone deacetylase inhibitor SAHA combined with paclitaxel on cervical cancer He La cells in vitro.The inhibitory rate of tumor cells was determined by MTT assay,the results showed that the survival rates from exposure to the combination of SAHA and paclitaxel(45.73±4.02%?22.98±3.86%)were significantly lower than the single treatment groups of SAHA(53.62±3.66%?34.74±3.03%)and paclitaxel(74.07±5.32%?70.88±3.09%)(P<0.01,P<0.01)after 24 h or 48 h.SPSS16.0 was used to calculate the effect of paclitaxel on IC50 in He La cells before and after SAHA treatment,IC50 dropped significantly,the half lethal dose of paclitaxel to He La cells decreased from(18.023±1.256)n M to(8.887±0.949)n M for 24 h and decreased from(12.119±1.083)n M to(3.994±0.601)n M for 48 h.The apoptotic rates of He La cells was detected by flow cytometry,and results showed that the apoptotic rates of SAHA group,paclitaxel group and SAHA combined with paclitaxel group was(6.44±0.86)%,(7.23±1.49)% and(16.22±3.38)% respectively.SAHA combination with paclitaxel can significantly increase the apoptotic rate of He La cells than SAHA or paclitaxel(P <0.05,P <0.05).Flow cytometry was also used to detected He La cells cycles,data showed He La cells were randomly divided into G0 / G1 phase(90.07 ± 1.39)% and S phase(3.88 ± 1.47)% by treated with SAHA for 24 h,randomly divided into G0 / G1 phase(33.48 ± 6.64)% and S phase(40.77 ± 4.43)% by treated with paclitaxel for 24 h,and randomly divided into G0 / G1 phase(84.22 ± 2.07)% and S phase(11.67 ± 1.28)% by treated with SAHA and paclitaxel for 24 h.The results suggested that SAHA combined with paclitaxel can significantly inhibit DNA synthesis and replication during He La cell mitosis.Flow cytometry assay was also performed to evaluate the ROS level in Hela cells.After 24-hour treating with SAHA,paclitaxel,SAHA and paclitaxel,the ROS level is(35.19±4.23)%?(27.26±3.74)% and(44.04±10.77)% respectively.While the control group is(1.65±0.84)%.The ROS level in cells treated with SAHA and paclitaxel increased significantly compared with the cells treated with paclitaxel separately(P<0.05).Immunofluorescence labeled microtubules and DNA of He La cell pointed out that SAHA combined with paclitaxel canshorten and thin microtubules within 24 h treatment.Additionally,the m RNA expression of P27 tumor suppressor gene in He La cells was detected by RT-PCR.The relative expression of p27 gene m RNA of paclitaxel group,SAHA group and SAHA combine with paclitaxel group was 0.978±0.117,5.845±0.548 and 10.601±0.673 respectively.The expression level of SAHA combine with paclitaxel group was significantly higher than paclitaxel and SAHA group(P<0.001,P <0.001).Western blot was used to detect the expression of caspase-3 and caspase-9 and acetylated histone H4(Ac-H4).And results indicated that SAHA combine with paclitaxel can activate caspase-3 and caspase-9 protein and enhance the acetylation of histone H4 level.In summary,in vitro,SAHA combined with paclitaxel can significantly inducing cervical cancer He La cells apoptosis,inhibiting cell proliferation,blocking cell cycle by activating the caspase protein pathway,enhance p27 tumor suppressor gene expression,increase histone H4 acetylation level and lead microtubule aberration in vitro,thereby enhancing their anti-tumor ability. |
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