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Histone acetyltransferase inhibitor II induces apoptosis in glioma cell lines via the p53 signaling pathway.

Xu LX, Li ZH, Tao YF, Li RH, Fang F, Zhao H, Li G, Li YH, Wang J, Feng X, Pan J - J. Exp. Clin. Cancer Res. (2014)

Bottom Line: Here, we examined the effects and mechanism of action of HATi II in glioma cell lines.Upregulation of p53 signaling pathway-related genes in HATi II-treated cells was confirmed by quantitative RT-PCR and Western blotting.HATi II deserves further investigation as a novel treatment for glioma.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pediatric Research, Children's Hospital affiliated to Soochow University, Suzhou, 215006, China. xulixiao2013@hotmail.com.

ABSTRACT

Background: Histone acetyltransferase (HAT) inhibitors can inhibit proliferation and induce apoptosis in cancer cell lines. The novel cell-permeable p300/CREB-binding protein (CBP)-selective HAT inhibitor HATi II can reduce histone H3 acetylation and induce chromatin condensation in HeLa cells. Here, we examined the effects and mechanism of action of HATi II in glioma cell lines.

Methods: Cell viability was assessed using the CCK-8 assay. Cell cycle analysis was performed using flow cytometry. Apoptosis was evaluated using Annexin V staining and flow cytometry, Hoechst 33342 staining and the TUNEL assay. Expression and cleavage of caspase-3, caspase-9 and poly ADP-ribose polymerase (PARP) were assessed by Western blotting. Statistical analysis was performed using two-tailed Student's t-tests. The gene expression profiles of U251 glioma cells treated with HATi II or DMSO were analyzed using the Arraystar Human 8 x 60 K LncRNA/mRNA expression array; data was analyzed using MEV (Multi Experiment View) cluster software. Datasets representing genes with altered expression profiles (≥2-fold) derived from the cluster analyses were subjected to gene ontology and pathway analysis.

Results: HATi II inhibited the proliferation of U251, U87, HS683 and SHG44 cells in a dose-dependent manner. HATi II induced cell cycle arrest at the G2/M phase, and induced significant levels of apoptosis, apoptotic body formation and DNA fragmentation in HATi II-treated U251 and SHG44 cells. HATi II induced cleavage of caspase-3, caspase-9 and PARP in U251 and SHG44 cells. In HATi II-treated U251 cells, 965 genes were upregulated, 984 genes were downregulated and 3492/33327 lncRNAs were differentially expressed. GO analysis showed the differentially expressed genes with known functions are involved in a variety of processes; alcoholism, p53 signaling pathway, cytokine-cytokine receptor interaction and transcriptional mis-regulation in cancer were the four most significant pathways. Upregulation of p53 signaling pathway-related genes in HATi II-treated cells was confirmed by quantitative RT-PCR and Western blotting.

Conclusions: HATi II inhibits proliferation and induces apoptosis via the caspase-dependent pathway in human glioma cell lines, possibly by activating the p53 signaling pathway. HATi II deserves further investigation as a novel treatment for glioma.

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Growth inhibitory effects of HATi II in glioma cell lines. (A-D) Cell viability assay. Glioma cells cultured in 96-well plates were treated with different concentrations of HATi II or DMSO for 48 h, and cell viability was determined using the CCK-8 assay. Cell proliferation was calculated as a percentage of the DMSO-treated control cells; IC50 values were derived by plotting the proliferation values on a logarithmic curve. The experiments were repeated three times. (E) Phase-contrast microscopy of U251, HS683, U87 and SHG44 cells treated with HATi II for 24 or 48 h (×100).
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Fig1: Growth inhibitory effects of HATi II in glioma cell lines. (A-D) Cell viability assay. Glioma cells cultured in 96-well plates were treated with different concentrations of HATi II or DMSO for 48 h, and cell viability was determined using the CCK-8 assay. Cell proliferation was calculated as a percentage of the DMSO-treated control cells; IC50 values were derived by plotting the proliferation values on a logarithmic curve. The experiments were repeated three times. (E) Phase-contrast microscopy of U251, HS683, U87 and SHG44 cells treated with HATi II for 24 or 48 h (×100).

Mentions: U251, U87, SHG44 and HS683 cells were treated with different concentrations of HATi II (0, 2.5, 5, 7.5, 10, 12.5, 15, 20 or 25 μmol/L) for 48 h, then cell proliferation was assessed using the CCK-8 kit. HATi II inhibited proliferation in a dose-dependent manner in all four glioma cell lines; DMSO (≤0.1%) had negligible influence on cell proliferation. The IC50 values for HATi II at 48 h in U251, U87 and HS683 cells were approximately 12.17 ± 0.7489 μmol/L, 9.513 ± 0.8632 μmol/L and 9.558 ± 1.081 μmol/L; SHG44 cells were more sensitive to HATi II than the other three cell lines with an IC50 of 5.9688 ± 0.5351 μmol/L (Figure 1A-1D).Figure 1


Histone acetyltransferase inhibitor II induces apoptosis in glioma cell lines via the p53 signaling pathway.

Xu LX, Li ZH, Tao YF, Li RH, Fang F, Zhao H, Li G, Li YH, Wang J, Feng X, Pan J - J. Exp. Clin. Cancer Res. (2014)

Growth inhibitory effects of HATi II in glioma cell lines. (A-D) Cell viability assay. Glioma cells cultured in 96-well plates were treated with different concentrations of HATi II or DMSO for 48 h, and cell viability was determined using the CCK-8 assay. Cell proliferation was calculated as a percentage of the DMSO-treated control cells; IC50 values were derived by plotting the proliferation values on a logarithmic curve. The experiments were repeated three times. (E) Phase-contrast microscopy of U251, HS683, U87 and SHG44 cells treated with HATi II for 24 or 48 h (×100).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4321714&req=5

Fig1: Growth inhibitory effects of HATi II in glioma cell lines. (A-D) Cell viability assay. Glioma cells cultured in 96-well plates were treated with different concentrations of HATi II or DMSO for 48 h, and cell viability was determined using the CCK-8 assay. Cell proliferation was calculated as a percentage of the DMSO-treated control cells; IC50 values were derived by plotting the proliferation values on a logarithmic curve. The experiments were repeated three times. (E) Phase-contrast microscopy of U251, HS683, U87 and SHG44 cells treated with HATi II for 24 or 48 h (×100).
Mentions: U251, U87, SHG44 and HS683 cells were treated with different concentrations of HATi II (0, 2.5, 5, 7.5, 10, 12.5, 15, 20 or 25 μmol/L) for 48 h, then cell proliferation was assessed using the CCK-8 kit. HATi II inhibited proliferation in a dose-dependent manner in all four glioma cell lines; DMSO (≤0.1%) had negligible influence on cell proliferation. The IC50 values for HATi II at 48 h in U251, U87 and HS683 cells were approximately 12.17 ± 0.7489 μmol/L, 9.513 ± 0.8632 μmol/L and 9.558 ± 1.081 μmol/L; SHG44 cells were more sensitive to HATi II than the other three cell lines with an IC50 of 5.9688 ± 0.5351 μmol/L (Figure 1A-1D).Figure 1

Bottom Line: Here, we examined the effects and mechanism of action of HATi II in glioma cell lines.Upregulation of p53 signaling pathway-related genes in HATi II-treated cells was confirmed by quantitative RT-PCR and Western blotting.HATi II deserves further investigation as a novel treatment for glioma.

View Article: PubMed Central - PubMed

Affiliation: Institute of Pediatric Research, Children's Hospital affiliated to Soochow University, Suzhou, 215006, China. xulixiao2013@hotmail.com.

ABSTRACT

Background: Histone acetyltransferase (HAT) inhibitors can inhibit proliferation and induce apoptosis in cancer cell lines. The novel cell-permeable p300/CREB-binding protein (CBP)-selective HAT inhibitor HATi II can reduce histone H3 acetylation and induce chromatin condensation in HeLa cells. Here, we examined the effects and mechanism of action of HATi II in glioma cell lines.

Methods: Cell viability was assessed using the CCK-8 assay. Cell cycle analysis was performed using flow cytometry. Apoptosis was evaluated using Annexin V staining and flow cytometry, Hoechst 33342 staining and the TUNEL assay. Expression and cleavage of caspase-3, caspase-9 and poly ADP-ribose polymerase (PARP) were assessed by Western blotting. Statistical analysis was performed using two-tailed Student's t-tests. The gene expression profiles of U251 glioma cells treated with HATi II or DMSO were analyzed using the Arraystar Human 8 x 60 K LncRNA/mRNA expression array; data was analyzed using MEV (Multi Experiment View) cluster software. Datasets representing genes with altered expression profiles (≥2-fold) derived from the cluster analyses were subjected to gene ontology and pathway analysis.

Results: HATi II inhibited the proliferation of U251, U87, HS683 and SHG44 cells in a dose-dependent manner. HATi II induced cell cycle arrest at the G2/M phase, and induced significant levels of apoptosis, apoptotic body formation and DNA fragmentation in HATi II-treated U251 and SHG44 cells. HATi II induced cleavage of caspase-3, caspase-9 and PARP in U251 and SHG44 cells. In HATi II-treated U251 cells, 965 genes were upregulated, 984 genes were downregulated and 3492/33327 lncRNAs were differentially expressed. GO analysis showed the differentially expressed genes with known functions are involved in a variety of processes; alcoholism, p53 signaling pathway, cytokine-cytokine receptor interaction and transcriptional mis-regulation in cancer were the four most significant pathways. Upregulation of p53 signaling pathway-related genes in HATi II-treated cells was confirmed by quantitative RT-PCR and Western blotting.

Conclusions: HATi II inhibits proliferation and induces apoptosis via the caspase-dependent pathway in human glioma cell lines, possibly by activating the p53 signaling pathway. HATi II deserves further investigation as a novel treatment for glioma.

Show MeSH
Related in: MedlinePlus