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Objective:To investigate the methylation status of O6-methylguanine-DNA methyltransferase (MGMT) in meningioma tissue and its effect on the growth and metastasis of human meningioma cell line IOMM-Lee cells.Methods:The specimens of 34 patients with meningioma were collected. Methylation-specific polymerase chain reaction (MSP) method was used to detect MGMT methylation in meningioma tissue and normal brain tissue; immunohistochemical staining was used to detect the expression of MGMT in meningioma tissue and normal brain tissue. The cultured human glioma cell line IOMM-Lee cells were divided into blank control group (normal cultured IOMM-Lee cells), negative control group (empty vector virus transfected IOMM-Lee cells) and RNAi lentivirus transfection group (transfected with RNAi lentivirus vector to down-regulate the expression of MGMT). The expression of MGMT in IOMM-Lee cells was silenced by RNAi technology. The expression levels of MGMT mRNA and protein in cells were detected by real-time fluorescence quantitative PCR and Western blot. The proliferation activity of cells was detected by cell counting kit-8 (CCK-8) test, the colony forming ability was detected by cell clone formation test, and the invasion and migration ability of cells in each group were detected by Transwell test and cell scratch test.Results:The methylation of MGMT in meningioma tissue reached 88.23% (30/34). MGMT methylation was not detected in normal brain tissue; the staining intensity of MGMT in meningioma tissue was significantly higher than that in normal brain tissue. Compared with the blank control group and the negative control group, the relative expression of MGMT mRNA and protein in IOMM-Lee cells of the RNAi lentiviral transfection group were significantly decreased (all P<0.05). After 24, 48 and 72 hours of transfection, the proliferation activity of IOMM-Lee cells decreased significantly (all P<0.05), with reduced number of cell clone formation and cell invasion (all P<0.05). The rate of scar healing decreased significantly ( P<0.05). Conclusions:MGMT is mostly hypermethylated in human meningiomas. Silencing the expression of MGMT in meningiomas can inhibit the growth and metastasis of meningiomas.
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Objective@#To investigate the prognostic impact of alterations of epidermal growth factor receptor(EGFR) and MGMT in glioblastoma.@*Methods@#The retrospective study included 161 supratentorial glioblastomas diagnosed in the Department of Pathology, Xuanwu Hospital, Capital Medical University from 2009 to 2015. EGFR and EGFRvⅢ protein expression was detected by immunohistochemistry; EGFR amplification was detected by fluorescence in situ hybridization; MGMT promoter methylation was detected by pyrosequencing. The change of molecular genetics EGFR and MGMT and outcome were assessed statistically.@*Results@#There were 161 patients, including 85 (52.8%) males and 76 (47.2%) females. The mean age was 53 years, and the median overall survival was 13 months. The integrated classification of glioblastoma included 16 IDH-mutant, 134 wild type, and 11 NOS. The rate of overexpression of EGFR protein was 32.9%(53/161), and that of EGFR amplification was 37.5%(18/48). There was high concordance between immunohistochemistry and FISH(85.4%, Kappa=0.475, P<0.01) and between the level of EGFR protein and EGFR amplification (P<0.01). Twelve cases showed EGFRvⅢ expression, and all also showed EGFR protein overexpression; 149 cases were EGFRv Ⅲ wild type, and EGFR protein overexpression was seen in 27.5%(41/149) of cases. There was no correlation between EGFR and EGFRv Ⅲ expression. Of all cases, 70.2%(106/151) showed MGMT promoter methylation by pyrosequencing. The changes of molecular genetics of EGFR and MGMT were not related. EGFR amplification and protein overexpression had no significant relationship with prognosis. Patients with EGFRv Ⅲ-mutant had shorter survival time than the EGFRv Ⅲ-wild type(P=0.014); patients with MGMT promoter methylation had better prognosis than without (PFS:P=0.002,OS:P=0.006),and MGMT promoter methylation was an independent predictor for overall survival (HR=0.269, 95%CI 0.124-0.583, P=0.001).@*Conclusions@#EGFR protein expression by immunohistochemistry correlates with the status of EGFR amplification. Patients with EGFRv Ⅲ-mutant tumors have poorer prognosis than that with EGFRv Ⅲ-wild type tumors. MGMT promoter methylation is closely associated with prognosis and an independent predictor for overall survival.
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Objective@#To investigate the expressions of histone lysine-specific demethylase 1 (LSD1), O6-methylguanine DNA methyltransferase (MGMT) and cell proliferation-associated antigen Ki-67 in high-grade glioma and their influences on prognosis.@*Methods@#Sixty-five cases of grade Ⅲ and Ⅳ glioma confirmed by pathology from January 2011 to June 2017 in the First Affiliated Hospital of Xinjiang Medical University were selected. Immunohistochemistry (SP method) was used to detect the expressions of LSD1, MGMT and Ki-67 in pathological specimens. The therapeutic effect was evaluated by long-term follow-up. The relationships between the three markers and pathological grade, progression-free survival (PFS) and overall survival (OS) were analyzed.@*Results@#The overall positive rates of LSD1, MGMT and Ki-67 in the 65 high-grade glioma specimens were 70.8% (46/65), 60.0% (39/65) and 100.0% (65/65), respectively. There were no significant differences in the expressions of LSD1 and MGMT in grade Ⅲ and Ⅳ glioma (χ2=1.588, P=0.208, χ2=0.013, P=0.908). Ki-67 expression (+ ), (+ + ), (+ + + ) in grade Ⅳ glioma were observed in 18, 19 and 11 cases, respectively. Ki-67 expression (+ ), (+ + ) in grade Ⅲ glioma were observed in 11, 5 cases, and 1 case was (+ + + ), and the difference in expression intensity between the two groups was statistically significant (Z=-2.083, P=0.037). Log-rank test showed that the positive expressions of LSD1, MGMT and Ki-67 were negatively correlated with the PFS of patients with high-grade glioma (χ2=12.217, P=0.007; χ2=4.446, P=0.035; χ2=12.536, P=0.002), also were negatively correlated with OS (χ2=11.708, P=0.008; χ2=6.637, P=0.010; χ2=11.807, P=0.003). Grade Ⅳ patients were more likely to have relapse progression than grade Ⅲ patients (χ2=6.573, P=0.010), and OS was shorter (χ2=3.974, P=0.046). Cox proportional hazards model analysis showed that the expressions of LSD1 (HR=1.361, 95%CI: 1.094-1.694, P=0.006; HR=1.406, 95%CI: 1.117-1.771, P=0.004) and Ki-67 (HR=1.703, 95%CI: 1.175-2.468, P=0.005; HR=1.778, 95%CI: 1.209-2.616, P=0.003) were the independent prognostic risk factors for PFS and OS of patients with high-grade glioma. Correlation analysis results showed that the expression of MGMT was positively correlated with the expression of LSD1 (r=0.406, P=0.001).@*Conclusion@#LSD1, MGMT and Ki-67 have higher positive expression rates in high-grade glioma. MGMT is a prognostic factor for high-grade glioma, and LSD1 and Ki-67 can be used as independent predictors of prognosis for high-grade gliomas.
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Objective To investigate the expressions of histone lysine-specific demethylase 1 (LSD1),O6-methylguanine DNA methyltransferase (MGMT) and cell proliferation-associated antigen Ki-67 in high-grade glioma and their influences on prognosis.Methods Sixty-five cases of grade Ⅲ and Ⅳ glioma confirmed by pathology from January 2011 to June 2017 in the First Affiliated Hospital of Xinjiang Medical University were selected.Immunohistochemistry (SP method) was used to detect the expressions of LSD1,MGMT and Ki-67 in pathological specimens.The therapeutic effect was evaluated by long-term follow-up.The relationships between the three markers and pathological grade,progression-free survival (PFS) and overall survival (OS) were analyzed.Results The overall positive rates of LSD1,MGMT and Ki-67 in the 65 high-grade glioma specimens were 70.8％ (46/65),60.0％ (39/65) and 100.0％ (65/65),respectively.There were no significant differences in the expressions of LSD1 and MGMT in grade Ⅲ and Ⅳ glioma (x2 =1.588,P =0.208,x2 =0.013,P=0.908).Ki-67 expression (+),(++),(+++) in grade Ⅳ glioma were observed in 18,19 and 11 cases,respectively.Ki-67 expression (+),(++) in grade Ⅲ glioma were observed in 11,5 cases,and 1 case was (+++),and the difference in expression intensity between the two groups was statistically significant (Z =-2.083,P =0.037).Log-rank test showed that the positive expressions of LSD1,MGMT and Ki-67 were negatively correlated with the PFS of patients with high-grade glioma (x2 =12.217,P =0.007;x2=4.446,P =0.035;x2=12.536,P =0.002),also were negatively correlated with OS (x2 =11.708,P =O.008;x2 =6.637,P =0.010;x2 =11.807,P =0.003).Grade Ⅳ patients were more likely to have relapse progression than grade Ⅲ patients (x2 =6.573,P =0.010),and OS was shorter (x2 =3.974,P=0.046).Cox proportional hazards model analysis showed that the expressions of LSD1 (HR =1.361,95％CI:1.094-1.694,P=0.006;HR=1.406,95％CI:1.117-1.771,P =0.004) and Ki-67 (HR=1.703,95％ CI:1.175-2.468,P =0.005;HR =1.778,95％ CI:1.209-2.616,P =0.003) were the independent prognostic risk factors for PFS and OS of patients with high-grade glioma.Correlation analysis results showed that the expression of MGMT was positively correlated with the expression of LSD1 (r =0.406,P =0.001).Conclusion LSD1,MGMT and Ki-67 have higher positive expression rates in high-grade glioma.MGMT is a prognostic factor for high-grade glioma,and LSD1 and Ki-67 can be used as independent predictors of prognosis for high-grade gliomas.
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O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation is an important molecular biomarker which plays a key role in tumor development and determine the molecular subtypes and prognosis of gliomas. Radiomics and imaging examinations like MR and PET can obtain information of morphology, function, metabolism and molecular alterations of gliomas, which may help comprehensively and non-invasively predict MGMT promoter methylation status in gliomas. The relationships between imaging features and MGMT promoter methylation status of gliomas were reviewed in this article.
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Background: Patients with Glioblastoma multiforme (GBM) have a five years survival of less than 5%, but the response to chemotherapy with alkylating agents can vary depending on the methylation status of O6-methylguanine-DNA-methyltransferase (MGMT). Genetic testing has limitations for routine use, while immunohistochemistry (IHC) offers a fast and affordable technique but with heterogeneous results in the literature. Aim: To evaluate MGMT expression by IHC in tumor tissue of Chilean patients with GBM. Material and Methods: Tumor samples of 29 patients with a pathological diagnosis of GBM were studied. We performed IHC staining and manual analysis of positive and negative cells for MGMT expression. A cut-off of at least 10% of cells expressing MGMT was used. Demographic and clinical features of patients were obtained from clinical records. Results: The median number of cells counted per case was 692 (interquartile range [IQR] 492-928). Fifteen cases (52%) were positive for MGMT expression. Median overall survival was 5.3 months (IQR 3.4-12-8). The effect of MGMT expression on the therapeutic response was not studied since only 3 patients received chemotherapy. Conclusions: Our results are similar to international reports, but we were not able to determine the association between MGMT expression and therapeutic response.
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Humans , Male , Female , Adult , Middle Aged , Aged , Brain Neoplasms/enzymology , Biomarkers, Tumor/metabolism , Glioblastoma/enzymology , O(6)-Methylguanine-DNA Methyltransferase/metabolism , Prognosis , Brain Neoplasms/genetics , Immunohistochemistry , Biomarkers, Tumor , Gene Expression Regulation, Neoplastic , Chile , Survival Rate , Retrospective Studies , Glioblastoma/genetics , O(6)-Methylguanine-DNA Methyltransferase/geneticsABSTRACT
Objective To establish a mouse model for short-term exposure to ambient PM and to investigate the impact on the Cytochrome P450 1A1(CYP1A1)and O6-methylguanine-DNA methyltransferase(MGMT)mRNA expression. Methods Twenty 6-week-old BALB/c mice were randomly assigned to one of two groups, each consisting of 5 male and 5 female animals. These mice were then housed in situ concurrently for 2 weeks in our lab located in urban area of Hangzhou. The first group was kept inside an individual ventilated caging(IVC)system equipped with a high-efficiency particulate-air(HEPA)filter, whereas the second was housed inside a IVC with HEPA filter removed. Then it's allowed flow-through of ambient air freely via a pipeline outside. Mice inside the HEPA filtration chamber were therefore protected from exposure to all airborne particulate. The other was in fact exposed to ambient air directly. After the exposure, the bronchoalveolar lavage(BAL)fiuid was collected for each animal and the differentials and percentages of BAL cells were determined. Paraffin sections of lungs of the mice were made and were examined for any inflammation changes. CYP1A1 and MGMT mRNA levels in the lungs were then detected by RT-qPCR. Results The mean concentration of PM2.5was(99.7±51.6)μg/m3in the exposure group. Weight increases were similar between the two groups(P>0.05). The number of total cells and macrophages in BALF from exposure mice was significantly greater than control.A mild inflammation was observed from light photomicrographs of the lung after PM exposure. CYP1A1 and MGMT mRNA levels were significantly up-regulated in the lung from the exposure group. Conclusion A mouse model for short-term exposure to ambient PM was established. CYP1A1 and MGMT may mediate the toxic effect of PM exposure.
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High grade gliomas are always the research focus in the field of neurosurgery due to their poor prognosis despite the current standard therapeutic regimen of surgical resection followed by radiation therapy and chemotherapy.Alkylating agent temozolomide has been established as the standard chemotherapy while its resistance inevitable during treatment.This phenomenon seriously influences the prognosis of patients suffering from high grade gliomas.This review aims to elucidate temozolomide chemoresistance mechanisms through three chapters including O6-methylguanine-DNA methyltransferase (MGMT) methylation,mismatch repair mutation and epigenetic regulation consisting of p21,chromatin and histone,Y-box binding protein-1 and microRNAs.
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High grade gliomas are always the research focus in the field of neurosurgery due to their poor prognosis despite the current standard therapeutic regimen of surgical resection followed by radiation therapy and chemotherapy. Alkylating agent temozolomide has been established as the standard chemotherapy while its resistance inevitable during treatment. This phenomenon seriously influences the prognosis of patients suffering from high grade gliomas. This review aims to elucidate temozolomide chemoresistance mechanisms through three chapters including O6-methylguanine-DNA methyltransferase (MGMT) methylation, mismatch repair mutation and epigenetic regulation consisting of p21, chromatin and histone, Y-box binding protein-1 and microRNAs.
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Background:Studies have shown that promoter methylation of MGMT gene is closely related to many malignant tumor including gastric cancer.DNA methyltransferase 1 (DNMT1) is highly expressed in many malignant tumor tissues.However, studies on relationship between methylation of MGMT gene and DNMT1 expression in gastric cancer are rare.Aims:To investigate the relationship between methylation of MGMT gene, protein expression of DNMT1 and gastric cancer.Methods:Promoter methylation status of MGMT gene in 60 gastric adenocarcinoma tissues and corresponding paracancerous tissues were detected by methylation-specific PCR (MSP).RT-PCR and immunohistochemistry were used to measure mRNA and protein expressions of MGMT and DNMT1, respectively.Results:Methylation rate of MGMT gene promoter in gastric cancer was significantly higher than that in paracancerous tissue (45.0% vs.13.3%, P<0.001).The positivity rate of MGMT mRNA in gastric cancer was significantly lower than that in paracancerous tissue (41.7% vs.93.3%, P<0.001), while the positivity rate of DNMT1 mRNA expression in gastric cancer was significantly higher than that in paracancerous tissue (76.7% vs.18.3%, P<0.001).Methylation rate of MGMT promoter in MGMT mRNA-negative expressed gastric cancer tissue was significantly higher than that in MGMT mRNA-positive expressed gastric cancer tissue (57.1% vs.28.0%, P<0.05).It showed a negative correlation between MGMT protein expression and DNMT1 protein expression in gastric cancer tissue (r=-0.795, P<0.01).Conclusions:Promoter methylation of MGMT gene and high expression of DNMT1 may be associated with the development and progression of gastric cancer.
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Objective To observe the influences of NaAsO2 on H3K36me3 modifications,mRNA transcription of O6-methylguanine-DNA methyltransferase gene (MGMT) in HaCaT cells,and to explore the relationship between the transcription of MGMT gene regulated by H3K36me3 and DNA damage induced by arsenic,in order to provide new ideas and scientific basis for prevention and intervention of arsenism.Methods HaCaT cells were treated with 1.25,2.50,5.00 and 10.00 μmol/L NaAsO2 for 24 h,and were also treated with 10.00 μmol/L NaAsO2 for 6,12 and 24 h.HaCaT cells that treated with 0.00 pmol/L NaAsO2 and 0 h were used as blank control group.The degree of DNA damage in peripheral blood cells was detected by single cell gel electrophoresis.The level of H3K36me3 modifications was detected using Western blotting.Quantitative real-time polymerase chain reaction was used to detect the mRNA levels of MGMT gene.Quantitative chromatin immuno-precipitation was used to detect the level of H3K36me3 modifications in the coding regions (ChIP1 and ChIP2) of MGMT gene.Results ①Among the groups of HaCaT cells treated with 2.50,5.00 and 10.00 μmol/L NaAsO2,the levels of tail DNA％ (11.83 ± 1.15,16.85 ± 2.52,24.23 ± 2.75) and olive tail moment (OTM,10.90 ± 1.13,16.19 ± 2.26,23.83 ± 2.79)were significantly increased compared with those of the control group (0.00 μmol/L,2.40 ± 0.51,2.26 ± 0.40,all P ＜ 0.05).After treated with 10.00 μmoFL NaAsO2 for 12 and 24 h,compared with the control group (0 h,3.66 ± 1.02,3.38 ± 1.00),the degrees of tail DNA％ (15.51 ± 1.92,24.18 ± 2.42) and OTM (13.58 ± 2.04,23.14 ± 2.11)were significantly increased (all P ＜ 0.05).②Compared with the control group (0.00 μmol/L,100.00 ± 0.00),the levels of H3K36me3 modifications (60.59 ± 9.75,57.82 ± 11.28,39.45 ± 7.09) were lower at the dosages of 2.50,5.00 and 10.00 μmol/L NaAsO2 (all P ＜ 0.05).Compared with the control group (0 h,100.00 ± 0.00),the levels of H3K36me3 modifications (48.47 ± 9.67,47.75 ± 6.98) were lower after treated with 10.00 μ mol/L NaAsO2 for 12 and 24 h (all P ＜ 0.05).③The levels of H3K36me3 modifications in HaCaT cells exposed to different doses of NaAsO2 were negatively associated with the tail DNA％ and OTM (r =-0.897,-0.903,all P ＜ 0.05).④Compared with the control group (0.00 μmol/L,100.00 ± 0.00),the mRNA levels of MGMT gene were lower at the dosages of 2.50,5.00 and 10.00 pmol/L NaAsO2 (78.20 ± 3.50,61.40 ± 2.60,49.15 ± 4.70,all P ＜ 0.05).⑤There was no observed H3K36me3 enrichmem regularity in the gene encoding ChIP1 and ChIP2 regions of MGMT gene in all doses of NaAsO2 groups (all P ＞ 0.05).Conclusions H3K36me3 may be involved in the regulation of arsenicinduced DNA damage in HaCaT cell.Amenic could inhibit the mRNA transcription of MGMT gene in HaCaT cells,but the transcription of MGMT gene regulate by H3K36me3 is not closely related to DNA damage induced by arsenic.
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Objective To investigate the differences of clinical efficacy and untoward reaction of different chemotherapy regi‐mens for patients with malignant glioma on different expression levels of O6‐methylguanine‐DNA‐methyltransferase(MGMT) ,in order to provide references for clinical treatment .Methods Totally 90 cases of patients with malignant glioma in our hospital from January 2011 to January 2013 were selected ,among them ,64 cases of MGMT negative expressing patients were divided into group A and group B with 32 cases in each group ,and 26 cases of MGMT positive expressing patients were enrolled into the group C . Group A was treated with combination of radiotherapy ,teniposide and nimustine ,group B was treated with radiotherapy‐temozolo‐mide combination regimen ,group C was treated with combination of radiotherapy ,teniposide and nimustine .The untoward reactions of the three groups were compared ,and the survival rate was observed after one year follow‐up .Results The hemoglobin ,leuko‐cyte ,granulocyte ,platelet ,bleeding ,alanine aminotransferase ,creatinine ,urea nitrogen ,peripheral neuritis ,untoward reactions a‐mong the three groups had no statistically significant differences (P>0 .05);the incidence rates of nausea and vomiting ,diarrhea , constipation among the three groups had statistically significant differences(P<0 .05) ,in which group C was significantly higher than that of group A and group B(P<0 .05) .Only one case in the group C was lost in the one year follow‐up .The median survival time was 10 months in group A and group B ,and was 7 months in group C .The median survival time in group C was significantly lower when compared with that in group A and group B(χ2 =7 .673 ,P=0 .006 ;χ2 =6 .395 ,P=0 .011) ,while there was no signifi‐cant difference of median survival time between group A and group B(χ2 =0 .063 .P=0 .802) .Conclusion The long‐term prognosis of patients with negative MGMT expression might be significantly worse than that of patients with negative MGMT expression in glioma .
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Female adnexal tumor of probable Wolffian origin (FATWO) is a rare disease entity that arises from the mesonephric duct system. FATWO is different than other gynecological cancers in terms of embryology. Here, we describe the case of a 52-year-old woman with malignant FATWO. The patient underwent explorative laparotomy and surgical staging after a frozen section revealed malignancy. Detailed examination of the pathologic findings were consistent with FATWO. Counseling and further testing were provided to the patient to assess the risk of germline mutation and epigenetic change. An O-6-methylguanine-DNA methyltransferase gene methylation test was positive, and all other tests were normal. This is the first study to report a case of O-6-methylguanine-DNA methyltransferase methylation with FATWO in Korea.
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Female , Humans , Middle Aged , Counseling , Embryology , Epigenesis, Genetic , Epigenomics , Frozen Sections , Germ-Line Mutation , Korea , Laparotomy , Methylation , Rare Diseases , Wolffian DuctsABSTRACT
OBJECTIVE: Malignant gliomas with neuronal marker expression (MGwNM) are rare and poorly characterized. Increasingly diverse types of MGwNM have been described and these reported cases underscore the dilemmas in the classification and diagnosis of those tumors. The aim of this study is to provide additional insights into MGwNM and present the clinicopathological features of 18 patients. METHODS: We reviewed the medical records of 18 patients diagnosed as MGwNM at our institute between January 2006 and December 2012. Macroscopic total resection was performed in 11 patients (61%). We evaluated the methylation status of O6-methylguanine-DNA methyltransferase (MGMT) and expression of isocitrate dehydrogenase 1 (IDH-1) in all cases, and deletions of 1p and 19q in available cases. RESULTS: The estimated median overall survival was 21.2 months. The median progression-free survival was 6.3 months. Six patients (33%) had MGMT methylation but IDH1 mutation was found in only one patient (6%). Gene analysis for 1p19q performed in nine patients revealed no deletion in six, 19q deletion only in two, and 1p deletion only in one. The extent of resection was significantly correlated with progression free survival on both univariate analysis and multivariate analysis (p=0.002 and p=0.013, respectively). CONCLUSION: In this study, the overall survival of MGwNM was not superior to glioblastoma. The extent of resection has a significant prognostic impact on progression-free survival. Further studies of the prognostic factors related to chemo-radio therapy, similar to studies with glioblastoma, are mandatory to improve survival.
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Humans , Classification , Diagnosis , Disease-Free Survival , Glioblastoma , Glioma , Isocitrate Dehydrogenase , Medical Records , Methylation , Multivariate Analysis , NeuronsABSTRACT
Despite multimodality treatment protocol including surgical resection, radiotherapy, and chemotherapy in patients with glioblastoma multiforme (GBM), most suffer from treatment failure and tumor recurrence within a few months of initial surgery. The effectiveness of temozolomide (TMZ), the most commonly used chemotherapeutic agent, is largely dependent on the methylation status of the promoter of the gene O6‑methylguanine‑DNA methyltransferase (MGMT) and the integrity of the mismatch repair (MMR) system. Changes in these regulatory mechanisms at the time of recurrence may influence response to therapy. Deciphering the molecular mechanisms of resistance to these drugs may in future lead to improvised patient management. In this article, we provide an update of the spectrum of molecular changes that occur in recurrent GBMs, and thus may have an impact on patient survival and treatment response. For review, electronic search for the keywords “Recurrent GBM”, “Recurrent GBM AND MGMT” “Recurrent glioma AND MGMT”, “Recurrent GBM AND MMR” and “Recurrent glioma AND MMR”, “Recurrent GBM AND MMR” and “Recurrent glioma AND MMR” was done on PubMed and relevant citations were screened including cross‑references.
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Objective To detect the expression of DNA repair enzyme O6-methylguanine DNA methyl-transferase(MGMT),cell proliferation-related nuclear protein(Ki-67)and P53 protein in gliomas,and inves-tigate the relationship and clinical significance among them and gloma grade. Methods 61 cases of brain glio-ma specimes and 16 cases of internal decompression of brain trauma were used to detect the expression of MGMT,Ki-67 and P53 by using immunohistochemical SP method. Results MGMT protein expression (19. 67%∶0,χ2 =3. 729,P=0. 062),Ki-67 protein expression(39. 34% ∶0,χ2 =5. 722,P=0. 016)and P53 protein expression(27. 87% ∶0,χ2 =9. 146,P=0. 002)showed significant differences in gliomas com-pared to normal brain tissues,the expression of Ki-67 was significantly higher in high-grade gliomas(Ⅲ-Ⅳ) than that in low-grade gliomas(Ⅰ-Ⅱ)(14 ∶10,χ2 =11. 718,P =0. 001). No significant difference was found between the MGMT and P53. Conclusion MGMT protein can be used as a biomarker in gliomas detec-tion;Ki-67 has a positive correlation with tumor grade,and can be used as a reference indicator of pathological grade;P53 protein expression may be used as a potential target for the treatment of gliomas.
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Background: Oral squamous cell carcinoma (OSCC) is a common cancer world‑wide that is highly lethal due to its recurrence and metastasis. Methylation is a common epigenetic mechanism that leads to gene silencing in tumors and could be a useful biomarker in OSCC. The prevalence of P16, death‑associated protein kinase (DAPK) and O6‑methylguanine‑DNA‑methyltransferase (MGMT) promoter hypermethylation in OSCC has been evaluated for several years while the results remain controversial. Objective: The aim of this systematic review is to critically analyze and perform a meta‑analysis on the various studies in the literature that have reported the promoter hypermethylation of P16, DAPK and MGMT genes in OSCC. Search Strategy: Articles were searched and selected through PubMed. Hand search from the relevant journals was also performed. Articles were reviewed and analyzed. Results: The estimated prevalence of P16 methylation was 43%, DAPK methylation was 39.7% and MGMT methylation was 39.8%. Heterogeneity in methylation prevalences and correlations with the clinical outcomes of the disease prevailed in various studies. Conclusion: We can conclude from our systematic review that a higher prevalence of methylation of P16, DAPK and MGMT occur in OSCC. Further studies are required to substantiate the role of methylation of P16, DAPK and MGMT as a marker in OSCC.
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Carcinoma, Squamous Cell/analysis , Death-Associated Protein Kinases/metabolism , Genes, p16 , Genes, Reporter , Humans , MethylationABSTRACT
Objective To establish a drug-resistance cell line of human glioma with temozolomide ( TMZ) ,investigate its resistance mechanisms, and provide experimental evidence for optimal TMZ therapy. Methods A TMZ-resistant human glioma cell line,U251/TR,was established by stepwise exposure of human parental U251 cells to TMZ. Resistance index and cell viability were accessed by MTT assay. Western-Blot,RT-PCR,immunohistochemistry and immunofluorescence were used to detect MGMT expression for the analysis of resistance mechanism. Results A TMZ-resistant human glioma cell line,U251/TR,was developed after 8 months of stepwise induction with 0. 25-16. 00 μg·mL-1 TMZ. IC50 in U251/TR cells was approximately 7 times higher compared with that in U251 cells (P=0. 00 ). The MGMT expression was significantly increased in U251/TR cells compared with that in parental U251 cells (P=0. 00) . Conclusion A TMZ-resistant human glioma cell line,U251/TR,was established by stepwise exposure of human parental U251 cells to TMZ. The primary mechanism of TMZ resistance is associated with increased activity of MGMT.
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Objective: To observe the effect of Sanguisorba tannins on the expression of MGMT (O6-methylguanine-DNA methyltransferase) gene and protein in myelosuppressed mouse model induced by CTX (cyclophosphamide). Methods: Forty Chinese Kunming mice were randomly divided into 4 groups: normal group (NR group), model group (MD group), positive group (G-CSF group) and Sanguisorba tannins group (ST group), with 10 mice in each group. Mouse model of bone marrow suppression was made by intraperitoneal injection of CTX. The mice in G-CSF group were intraperitoneally injected with 30 μg/kg colony-stimulating factor after CTX induction for 3 days. The mice in ST group started to take 20 mg/kg Sanguisorba tannins orally 3 days before CTX induction. The mice in NR group and MD group were given purified water orally. At the end of the treatment, blood and bone marrow of all mice were taken out. Then the flow cytometry technology was used to examine the DNA content in bone marrow cells. The RT-PCR (reverse transcription-PCR) and Western blotting technologies were used to determine the expression changes of MGMT gene and protein in bone marrow cells. Results: Compared with the NR group, the DNA content (P < 0.01) and the expressions of MGMT gene and protein (P < 0.01) were significantly reduced in bone marrow cells of mice in MD group. Compared with the MD group, bone marrow DNA content and the expression levels of MGMT gene and protein were significantly increased in ST group (P < 0.01). Conclusion: Sanguisorba tannins can promote the expression of MGMT gene and protein in bone marrow cells of myelosuppressed mice. Copyright © 2013 by TUMOR.
ABSTRACT
Objective To investigate the effect of NaAsO2 on the binding levels of methyl CpG binding protein 2(MeCP2),DNA methyltransferase 1 (DNMT1) and histone deacetylase 1(HDAC1) to the hypermethylation promoter region of MGMT gene in HaCaT cells,in order to provide a basis to deepen the interpretation of the role of arsenic poisoning mechanism.Methods HaCaT cells were treated repeatedly and interval with different concentrations of NaAsO2(3.13,6.25,12.50,25.00 μnol/L,respectively) for 72 h.Untreated HaCaT was used as blank control group and human epidermal squamous carcinoma cell line(A431 cells) as positive control group.The binding levels to the two transcription regulatory regions(ChIP1,ChIP2) and to the coding region(ChIP3) of MGMT 8ene were detected by chromatin immuno-precipitation combined with quantitative PCR.Results The differences of binding levels of MeCP2,DNMT1 and HDAC1 to ChIP1 and ChIP2 in each group were significant (F=7.387,84.634,78.442 and 19.263,69.649,26.546,all P ＜ 0.05).The binding levels of MeCP2,DNMT1 and HDAC1 to ChIP1 and ChIP2 in each NaAsO2 exposed group[3.13 μmol/L NaAsO2 exposed group:(136.00 ±16.97)％,(145.00 ± 2.83)％,(88.50 ± 19.09)％ and (106.50 ± 37.48)％,(112.34 ± 8.73)％,(59.71 ± 8.49)％;6.25 μmol/L NaAsO2 exposed group:(130.00 ± 42.43)％,(154.50 ± 4.95)％,(101.00 ± 1.27)％ and (88.50 ±3.54)％,(134.32 ± 2.82)％,(102.75 ± 19.91)％ ; 12.50 μmol/L NaAsO2 exposed group:(141.50 ± 23.33)％,(161.50 ± 7.78)％,(125.00 ± 11.31)％ and (119.50 ± 24.75)％,(171.59 ± 3.54)％,(167.61 ± 10.61)％; 25.00μmol/L NaAsO2 exposed group:(134.50 ± 43.13)％,(472.50+ 50.20)％,(383.50 ± 30.41)％ and (180.09 ±12.73)％,(348.50 ± 27.58)％,(158.45 ± 12.02)％] were higher than that in the blank control group[(51.50 ±9.19)％,(82.00 ± 12.73)％,(25.03 ± 2.91)％ and (37.02 ± 4.24)％,(91.56 ± 26.16)％,(19.09 ± 2.90)％,all P ＜ 0.05].The differences of binding levels of MeCP2 to ChIP3 in each group were not significant(F =1.670,P ＞0.05),but the differences of binding levels of DNMT1 and HDAC1 to ChIP3 were significant (F =4.404,9.863,all P ＜ 0.05),and only the binding levels in the 25.00 μmol/L NaAsO2 exposed group [(615.85 ± 29.63)％,(306.09 ± 59.40)％] were higher than that in the blank control group[(99.70 ± 12.02)％,(92.45 ± 48.79)％,all P ＜ 0.05].Conclusions MeCP2 can bind to the methylated MGMT gene transcriptional regulatory regions which are induced by arsenic and leads to histone deacetylation by the recruitment of DNMT1 and HDAC1 and,meanwhile,DNMT1 can bind to the coding region of MGMT gene to recruit HDAC1 in a methyl DNA binding protein(MBD) independence manner and media MGMT gene silencing through the chromatin remodeling way,which might be the early molecular events of arsenic poisoning.