E results showed that squamocin-treated cancer cells exhibited significant loss of
E results showed that squamocin-treated cancer cells exhibited significant loss of viability in dose-dependent manners (AZD4547 solubility Figure 4). The 50 inhibitory concentrations (IC50) of GBM8401, Huh-7, and SW620 cells were 46.1, 39.4, and 40.4 M, respectively.Squamocin arrested the cell cycle at the G1 phase and induced apoptosisTo further evaluate the potential relevance of histone H3 phosphorylation in cancer therapy, we examined the effects of squamocin on cell growth and viability. CellsLee et al. BMC Cancer 2011, 11:58 http://www.biomedcentral.com/1471-2407/11/Page 4 ofFigure 2 Squamocin decreased expression levels of RNA of aurora B and MSK1. GBM841, Huh-7 and SW620 cells were incubated with 30 and 60 M squamocin for 24 h. mRNA was extracted and detected by qRT-PCR. Data represent fold change versus controls, and values were normalized to GAPDH. Data are the mean of three independent experiments. *p < 0.05, compared to PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/28667899 the control.were treated with squamocin for 24 h, and the cell cycle distribution and apoptosis were measured by a flow cytometric analysis. Squamocin treatment significantly increased the population of G 1 phase cells (Figure 5). Also, high levels of apoptosis were detected in squamocin-treated cells (Figure 6). As shown in Figure 5, treatment of cells with 0, 15, 30, and 60 M of squamocin resulted in G1 phase accumulation of cells corresponding to 37.8 , 46.7 , 60.6 and 56.3 , respectively in GBM841 cells (Figure 5A), 41 , 59.7 , 53.6 , and 54.5 , respectively in Huh-7 cells (Figure 5B), and 53.2 , 64.9 , 59.1 , 54.5 , respectively in SW620 cells (Figure 5C). Moreover, squamocin-treated cells were stained with propidium iodide (PI) and annexin V to determine the apoptotic cells. Cells were differentiated among viable (annexin V, PI double negative), early-apoptotic (annexin V positive, PI negative) and late-apoptotic (annexin V, PI double positive) cells. Treatment of cells with 0, 15, 30, and 60 M of squamocin increased the percentage of early apoptosis from 0.7 to 14.1 , 4.3 , and 5.3 ,respectively and late apoptosis from 4.1 to 5.5 , 21.9 , and 49 , respectively in GBM8401 cells (Figure 6A), early apoptosis from 1.8 to 15.1 , 21 , and 7.6 , respectively and late apoptosis from 3.0 to 8.6 , 12.1 , and 62.9 , respectively in Huh-7 cells (Figure 6B), and early apoptosis from 3.0 to 21.2 , 20.1 , and 22.8 , respectively and late apoptosis from 1.2 to 2.4 , 20.2 , and 36.9 , respectively in SW620 cells (Figure 6C). Further, we extended our study to apoptosis-associated molecules and found that increasing levels of caspase-3, -8, and -9 activities and cleavage of poly ADP-ribose polymerase (PARP) were observed in squamocininduced apoptosis (Figure 7). From the results, it is evident that squamocin affected cell cycle progression and apoptosis.Effects of squamocin on mitogen-activated protein kinase (MAPK)The MAPK signaling pathway is implicated in a wide range of cellular functions, including cell proliferation,Figure 3 Downregulation of aurora B, pMSK1, H3S10p, and H3S28p protein expression levels was observed with squamocin treatment. Cells were incubated with 15, 30, and 60 M squamocin for 24 h. Proteins were extracted and analyzed by Western blotting. GAPDH was used as a loading control. (A) GBM841 cells. (B) Huh-7 cells. (C) SW620 cells. Data are representative of three independent experiments.Lee et al. BMC Cancer 2011, 11:58 http://www.biomedcentral.com/1471-2407/11/Page 5 ofFigure 4 Inhibition of cancer cell grow.
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