| ObjectivesTo investigate the effect of two novel histone deacetylase inhibitors(HDACI) CC1030 and CC1054, on proliferation and apoptosis of human cervical carcinoma cell lines HeLa and Caski in vitro.Methods1. Human cervical carcinoma cell lines, HeLa and Caski were cultured and treated with different concentration of CC1030 and CC1054 in logarithmic growth phase, observation cell form under the optical microscope, record the celluar morphology change.2. HeLa and Caski were treated with different concentration of CC1030 and CC1054 for 24 hours,48 hours,72 hours,then cell proliferation inhibition were measured by MTT colorimetric assay, make the 50% inhibiting concentration (IC50) of cells and the concentration-inhibition rate curve and time-inhibition rate curve. Experimental repeat three times.3. HeLa and Caski were treated with CC1030 and CC1054 respectively for 48 hours, then observed the apoptosis of cells under the fluorescence microscope.4. HeLa and Caski treated with different concentration of CC1030 and CC1054 for 48 hours, doubly stained by Annexin V-FITC/propidiumiodide(PI), the apoptosis index was assessed by flow cytometry(FCM).Results1.The optical microscope characteristic were:both of them became shrinks, elongated,and cell division is reduced after treated with CC1030 and CC1054 for 24 hours; the characteristic elongated cells having filamentous protrusions induced by CC1030 and CC1054 for 48 hours, and majority of them were desquamated from the wall; after treated with CC1030 and CC1054 for 72 hours, cells almost desquamated from the wall in high concentration group, left some cell fragments. While, the control cell of HeLa was polygon or irregular shape and Caski was spindle.2. Cell proliferation measured by MTT colorimetric assay. HeLa and Caski treated with 1μM,10μM,20μM,30μM,40μM of CC1030 and CC1054 for 48 hours, respectively, and the proliferation inhibition rate of HeLa were:11.33%±1.24%,53.31%±2.90%,62.09%±3.21%, 70.84%±3.46%,80.87%±2.81%,9.99%±1.37%,44.08%±2.80%, 64.86%±1.49%,73.66%±2.75%,85.11%±1.19%, respectively; the proliferation inhibition rate of Caski were:6.83%±0.08%,33.84%±1.12%, 55.44%±1.09%,69.65%±1.43%,80.63%±0.31%,16.56%±0.69%, 36.35%±0.26%,68.51%±0.79%,70.52%±1.86%,89.61%±0.40%. the inhibition rate was in a time and dose dependent manner. Aftet treated with CC1030 and CC1054 for 48 hours, the IC50 of HeLa were 9.550μM and 10.066μM, the IC50 of Caski were 14.161μM and 9.072μM; While, the IC50 of HeLa were 4.266μM and 3.746μM, the IC50 of Caski were 5.042μM and 5.173μM for 72 hours, respectively.3. Aftet treated with CC1030 and CC1054 for 48 hours, under fluorescence, early apoptosis cells were seen to emit green fluorescent; late apoptosis cells were seen to emit green fluorescent and red fluorescent; necrotic cellular cells were were seen to emit red fluorescent.4. Doubly stained by Annexin V-FITC/PI, the apoptosis index was assessed by flow cytometry. HeLa and Caski treated with 10μM,20μM,30μM,40μM of CC1030 and CC1054 for 48 hours, respectively.After treated with CC1030, the total apoptosis index of HeLa were: 20.07%±3.59%,31.7%±3.85%.44.77%±7.92%,61.0%±11.67%, corresponding to CC1030 10μM,20μM,30μM,40μM, respectively. Compared with the control group, had statistic significance, P<0.05, The comparison of every treated groups, P<0.05; After treated with CC1054, the total apoptosis index were:18.1%±5.85%,43.73%±2.16%,45.5%±7.69%,62.27%±27.47%, respectively, compared with the control group, P<0.05, among the comparison of every treated groups, aslo had statistic significance, P<0.05,.except for 20μM group and 30μM group. The blank and after treated with DMSO the apoptosis index were: 6.9%±1.06%,6.33%±1.16%, compared the two P> 0.05, had no statistical significance.After treated with CC1030, the total apoptosis index of Caski were: 10.17%±4.09%,23.33%±2.28%,61.13%±4.39%,66.13%±3.89%, corresponding to 10μM,20μM,30μM,40μM, respectively. compared with the control group, P<0.05, had statistic significance, among the comparison of every treated groups, aslo had statistic significance, P< 0.05, except for 30μM group and 40μM group; After treated with CC1054, the total apoptosis index were:6.93%±2.61%,40.1%±6.72%,56.5%±6.19%,59.13%±4.46%, respectively, compared with the control group, P<0.05, had statistic significance, except for 10μM group.among the comparison of every treated groups, aslo had statistic significance, P <0.05, except for 30μM group and 40μM group. The blank and after treated with DMSO the apoptosis index were:5.73%±2.27%, 6.77%±2.89%, compared the two P>0.05, had no statistical significance.Conclusion1. CC1030 and CC1054 could inhibit the growth of human cervical carcinoma cell lines obviously, in a time and dose dependent manner.2. CC1030 and CC1054 could induce apoptosis of human cervical carcinoma cell lines effectively, in a dose dependent manner. |
