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Objective To observe the effect of hypoxia on the expression of epithelial growth factor receptor (EGFR) and cell apoptosis of breast and cervical cancer xenografts in nude mouse models.Methods The nude mouse models with MCF-7 and HeLa xenografts were established.The degree of hypoxia and EGFR expression were observed by confocal microscopy.The influence of EGFR expression on cell apoptosis under hypoxia was observed by TUNEL assay.Results EGFR expression was either up-regulated or down-regulated in the MCF-7 and HeLa cells with high degree of hypoxia.Furthermore,the degree of apoptosis was reduced in tumor tissues with high EGFR expression compared with that in those with low expression of EGFR.Conclusion The hypoxia in MCF-7 and HeLa cells exerts heterogeneous effect on EGFR expression.Under hypoxic conditions,EGFR exoression is negatively correlated with cell apoptosis.
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Objective@#To evaluate the effect of long-chain non-coding RNA TUG1 on the radiosensitivity of cervical cancer cells and explore its underlying mechanism.@*Methods@#The expression of TUG1 and miR-145 in cervical cancer cells XB1702 and normal endometrial stromal cells (ESCs) was detected by qRT-PCR. The transfected si-NC, transfected si-TUG1, transfected si-NC combined with irradiation, transfected si-TUG1 combined with irradiation, si-TUG1 and anti-miR-NC co-transfected and, si-TUG1 and anti-miR-145 co-transfected groups were established, which were transfected into XB1702 cells by liposome method. The survival fraction of each group was detected by colony formation assay. The cell apoptosis of each group was detected by flow cytometry. The fluorescence activity of each group was assessed by dual luciferase reporter gene assay.@*Results@#Compared with the normal ESCs, the expression of TUG1 was significantly up-regulated, whereas that of miR-145 was significantly down-regulated in the cervical cancer cells XB1702. Silencing TUG1 significantly increased the survival fraction of XB1702 cells, promoted cell apoptosis and enhanced the radiosensitivity of irradiation to XB1702 cells. TUG1 could target and regulate the expression of miR-145. Suppressing miR-145 reversed the silencing effect of TUG1 on inhibiting proliferation, accelerating apoptosis promotion and enhancing sensitization of XB1702 cells.@*Conclusions@#Silencing long-chain non-coding RNA TUG1 can enhance the radiosensitivity of cervical cancer cells. The mechanism may be related to targeting miR-145, which will provide a target for radiotherapy of cervical cancer.
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An important molecular target for cancer therapy is the possible reactivation of tumor suppressor genes that have been silenced by promoter methylation. It was observed that the treatment of an adenocarcinoma cervical cancer cell line, HeLa with 20 μg/ml of the ethanolic extract of Withania somnifera for 6 days resulted in demethylation of promoter of RARβ2 gene. However, treatment with Ocimum sanctum and Azadirachta indica (20μg/ml) did not cause the reversal of hypermethylation after 6 days of treatment. This is the first report to show the reversal of hypermethylation of RARβ2 gene by Withania somnifera extract in a cervical cancer cell line.
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Background and purpose:Loss or altered expression of Ras association domain family 1A gene (RASSF1A) through DNA methylation has been associated with the pathogenesis of a variety of cancers, which suggests the tumor suppressor function of this gene. This study aimed to explore the effect of DNA methyltransferase inhibitor 5-Aza-2’deoxycytidine (5-Aza-dc) on demethylation and expression of RASSF1A in cervical cancer cell lines. Methods:HPV positive cervical cancer cell lines HeLa and Caski, HPV negative cell lines HT-3 and C-33A were treated with two different concentration of 5-Aza-dc (5 μmol/L, 10 μmol/L). MSP (methylation-specific PCR) and Bisulfite genomic sequencing PCR (BGS) combined with TA clone were used to investigate methylation status of RASSF1A gene promoter and exon 1 before and after treatment of 5-Aza-dc. RASSF1A gene mRNA expression was detected by RT-PCR. Results:Two HPV positive cell lines showed hypomethylated RASSF1A promoters and expressed RASSF1A mRNA, and after treatment with 5-Aza-dc, the mRNA expression of RASSF1A did not change significantly (FHeLa=3.003, P=0.125; FCaski=0.045, P=0.956). Two HPV negative cervical cancer cell lines showed hypermethylation status of RASSF1A promoter and silenced RASSF1A. After treatment with 5-Aza-dc, demethylation occurred in the promoter region of RASSF1A gene, which subsequently induced re-expression of this gene in HT-3 and C-33A. The F test (FHT-3=18.002, P=0.03;FC-33A=17.179, P=0.03) and LSD-t test (P<0.05) demonstrated that significant difference in the expression of RASSF1A was found upon two different concentrations drug treatment.Conclusion:The methylation status of promoter and exon 1 of RASSFIA gene in HPV positive and HPV negative cervical cancer cell lines are different. The promoter hypermethylation is correlated with RASSF1A gene expression in HPV negative cervical cancer cell line HT-3 and C-33A, and plays a key role in RASSF1A silencing. 5-Aza-dc may effectively reverse the methylation status of RASSFIA gene promoter in cervical cancer HT-3 and C-33A cells and reactivate gene expression silenced by aberrant hypermethylation in a dose-dependent manner within certain extent.
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Background: Vitamin E is a potent growth inhibitor of various cancer cell types in vitro and in vivo. The cell death mechanism is believed to be via cell cycle blockage, differentiation, and apoptosis. Objectives: To determine the possible involvement of protein expression of MEK-2 and ERK-2 in the cell death mechanism induced by palm oil ?-tocotrienol and ?-tocopherol in human cervical cancer cell line, CaSki cells. Methods: In this study, we tested the effect of ?-tocotrienol and ?-tocopherol on the proliferation and apoptosis in CaSki cells. Western blot analysis was used to determine the involvement of MEK-2 and ERK-2 in regulating the cell death mechanism. Results: Gamma-tocotrienol and α-tocopherol efficiently inhibited the proliferation of CaSki cells by 85.2% to 90.8% (p<0.01, n=4) and 10.2% to 39.1% (p<0.01, n=4) beginning at 100 μM and 50 μM, respectively. The possible cell death mechanism induced by both compounds may be due to apoptosis as confirmed by the presence of cellular DNA fragments separated by electrophoresis and enhancement of apoptotic activity. Treatment with γ-tocotrienol at 150 μM markedly decreased the protein expression of MEK-2 and ERK-2 at 12 hours and 18 hours. In contrast, treatment with α-tocopherol at 300μM has no effect on both protein expressions. Conclusion: The transient decreases in the protein expression of MEK-2 and ERK-2 suggested that the anti proliferative effect of γ-tocotrienol might involve alteration of the proliferative signaling cascade.
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OBJECTIVE: To estimate the difference in gene expression related to carcinogenesis between HPV 16 positive squamous cell carcinoma and HPV 16 positive adenocarcinoma of cervix. METHODS: We used cDNA microarray technology to identify alterations in gene expression of human cervical cancers. Gene expression of three cell lines, CaSki and SiHa (HPV 16 positive squamous cell carcinoma) and HeLa (HPV 16 positive adenocarcinoma) were compared with HT3 (HPV 16 negative squamous cell carcinoma). The microarray contains twin spots for 344 cancer-associated genes. RESULTS: The analysis showed several interesting findings: (1) In all three squamous cell lines, CD4, CSF1, MMP15 and TNFR6 were increased, whereas SLC3A2 were decreased, (2) Only in adenocarcinoma cell line HeLa, CDC25A, CDK2, CDK9, IL2, PF4, MAD, FCER2, MAP4K1 and MS4A1 were increased, and PLAU, IL8, IL9R and ATK were decreased. (3) In both squamous cell carcinoma cell lines CaSki and SiHa, 61 genes which belong to chemokine, cell cycle, growth factor, interleukin, adhesion molecule, protein kinase and TNF were increased, whereas 10 genes which are associated with apoptosis and cytokine were increased only in SiHa, and 97 genes which are associated with a variety of cell functions were increased only in CaSki. CONCLUSION: We suggest that there might be common, but also different carcinogenic mechanisms involved in HPV 16 related cervical cancers according to the histologic subtypes and different tumors.
Subject(s)
Female , Humans , Adenocarcinoma , Apoptosis , Carcinogenesis , Carcinoma, Squamous Cell , Cell Cycle , Cell Line , Cervix Uteri , DNA, Complementary , Gene Expression , Human papillomavirus 16 , Interleukin-2 , Interleukin-8 , Interleukins , Oligonucleotide Array Sequence Analysis , Protein Kinases , Uterine Cervical NeoplasmsABSTRACT
OBJECTIVE: To obtain information on the growth inhibition effect of arsenic compounds and gene expression profiles using cDNA microarray technique in SiHa cell lines. METHODS: We cultured 103 SiHa cell in 96 well plate and we investigated growth inhibition effects using MTT assay and also we performed gene expression profile experiment using 384 cDNA chip in SiHa cell after exposure of arsenics (As2O3, As4O6 - 1 (micro)M) for 48 hrs. RESULTS: Arsenics (As2O3, As4O6) inhibit the growth of SiHa cells (As2O3: 0.5, 1, 2, 3, 4, 5 (micro)M - 9.2, 56, 89, 93, 96, 96%, As4O6: 0.5, 1, 2, 3, 4, 5 (micro)M- 54, 84, 84, 85, 85, 87%) in 4 days culture. As2O3 and As4O6 induced apoptosis in SiHa cells. After exposure of As2O3, 47 genes were changed more than 2 times (eg, thymidylate synthetase, cyclin B1, CDC 20). In case of As4O6, 78 genes were changed more than 2 times (eg, CDC 20, cyclin B1, primase, proliferating cell nuclear antigen). CONCLUSION: we observed arsenic compound (As2O3, As4O6) inhibit the growth of SiHa cell. In gene expression profiling experiment, 78 genes was changed the expression level 2 times more than that of reference RNA after treatment of As4O6 and 47 genes after treatment of As2O3. Through these result, we thought more study need in functional genomics after arsenic treated cervical cancer cells.