| BackgroundHistone post-translation modifications (PTMs) are the points of convergence of signaling pathways and that this nodal event is crucial for gene expression. Signal transduction kinases can induce direct phosphorylation of specific residues of histone H3 at inducible promoters during interphase, cause rapid transcriptional induction of those genes. p300 is a transcription cofactor with histone acetyltransferas (HAT) activity, which is capable of acetylating all the 4 core histones as.well as at least 70 other proteins. Simultaneous induction of histone phosphorylation and acetylation are crucial for the activation of specific mammalian genes. Our previous studies showed that, in gastric cancer cells, octreotide (OCT), a somatostatin analogy, could down regulate the level of phospholated Akt, increase p300-HAT activity, and inhibit proliferation of gastric cancer cells.ObjectiveTo observe the effects of OCT on phosphorylation and acetylation of histone H3, and explore whether the phosphorylation and acetylation of histone H3 are involved in the inhibiting effects of OCT on the proliferation of gastric cancer cell.Materials and methods1.Experimental group Gastric cancer cell SGC7901 was randomly divided into four groups:the experimental groups 1-3 were cultured with medium containing OCT at a final concentration of 10nmol/L for 12h,24h and 48h, respectively; the control group was cultured with medium containing no OCT. The cells of the four groups were collected, and the nuclear proteins were extracted, respectively, and used for Western blot. The cells were seeded on the cover slides within the wells of 24-well-plate, grouped as mentioned above, and used for immunocytochemistry.2. Western blot The level of histone H3 with phosphorylated serine 10 (pS10-H3) and that with acetylated lysine 14 (acK14-H3) in the four groups was detected by using Western blot to observe the effects of OCT on the level of pS10-H3 and acK14-H3 protein in gastric cancer cell SGC7901.3. Immunocytochemistry The immunocytochemistry was used to observe the immuno-reactivity (IR) of pS10-H3 and acK14-H3 in the four groups, and detect the effects of OCT on pS10-H3 and acK14-H3 IR in gastric cancer cell SGC7901 by semi-quantitive analysis.4. Statistical analysis All data were showed as mean ± standard deviation (x±s) and analyzed with SPSS20.0 statistical software. P<0.05 was considered as statistically significant.Results1. Effects of OCT on the level of pS10-H3 protein in gastric cancer cell The relative level of pS10-H3 in control group, groups of OCT treated for 12h,24h and 48h was 1.005 ± 0.021,0.633 ± 0.022,0.401 ± 0.028 and 0.089 ± 0.010, respectively. A significant difference occurred between the relative level of pS10-H3 in control group and those in experimental groups (P< 0.01). OCT could significantly decrease the level of pS10-H3 protein in gastric cancer cell SGC7901.2. Effects of OCT on pS10-H3 IR in gastric cancer cell The pS10-H3 IR appeared as brownish granules, and mainly distributed in periphery of nucleus. The average level of pS10-H3 IR for weakly positive and positive cells in control group, groups of OCT treated for 12h,24h and 48h was 0.25±0.07 and 1.29±0.16, 0.23±0.04 and 1.08±0.12,0.23±0.07 and 0.54±0.16,0.26±0.05 and 0.12±0.02, respectively. The total average level of IR in the cells of 4 groups was 1.55±0.09, 1.32±0.09,0.77±0.09 and 0.38±0.07, respectively. The average level of pS10-H3 IR for positive cells in experimental groups was significantly lower than that in control group (P<0.01); however, there was no significant difference between the average level of pS10-H3 IR for weakly positive in experimental groups and that in control group (P>0.05). A significant difference occurred between the total average level of pS10-H3 IR in control group and those in experimental groups (P<0.01). OCT could significantly decrease pS10-H3 IR in gastric cancer cell SGC7901.3. Effects of OCT on the level of acK14-H3 protein in gastric cancer cell The relative level of acK14-H3 in control group, groups of OCT treated for 12h,24h and 48h was 0.109 ± 0.012,0.237 ± 0.013,0.492 ± 0.014 and 0.689 ± 0.009, respectively. There was a significant difference between the relative level of acK14-H3 in control group and those in experimental groups (P<0.01). OCT could significantly increase the the level of acK14-H3 protein in gastric cancer cell SGC7901.4. Effects of OCT on acK14-H3 IR in gastric cancer cell The acK14-H3 IR appeared as brownish granules, and mainly distributed in periphery of nucleus. The average level of acK14-H3 IR for weakly positive and positive cells in control group, groups of OCT treated for 12h,24h and 48h was 0.06±0.01 and 0.05±0.02, 0.18±0.04 and 0.35±0.08,0.30±0.04 and 0.48±0.03,0.32±0.06 and 1.06±0.07, respectively. The total average level of acK14-H3 IR in the cells of 4 groups was 0.15±0.03,0.53±0.11,0.78±0.06 and 1.39±0.03, respectively. The average level of acK14-H3 IR for weakly positive and positive cells in control group was significantly lower than that in control group (P<0.01). There was a significant difference between total average level of acK14-H3 IR in control group and those in experimental’ groups (P<0.01). OCT could significantly increase acK14-H3 IR in gastric cancer cell SGC7901.ConclusionsOCT can significantly decrease the phosphorylation of serine 10 and significantly increase the acetylation of lysine 14 in histone H3 in gastric cancer cell SGC7901. The results suggest that the phosphorylation of serine 10 and the acetylation of lysine 14 in histone H3 may be involved in the inhibitory effects of OCT on the proliferation of gastric cancer cell. |
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