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Aims:Non-small cell lung cancer (NSCLC) accounts for high lung cancer death that is mostly associated with advanced disease stage at diagnosis and resistance to chemotherapy. In the present study, we investigated whether xanthohumol, a prenylated flavonoid of hop plant, induces metastatic lung cancer H1299 cell death, and whether in combination with cisplatin there are additive effects. Methodology:H1299 cells were grown and treated with xanthohumol (6.25, 12.5, or 25 μM), cisplatin (12.5, 25, or 50 μM) and the combination of cisplatin and xanthohumol for 24 h. Cell viability, cell morphology, chromatin condensation, ɣH2AX, cPARP-1, capsase-3, p21WAF1/CIP1and p14ARFgenes were analyzed Results:Xanthohumol, cisplatin, and the combination of cisplatin and xanthohumol inhibited H1299 cells viability. Cisplatin growth inhibitory effects were potentiated by xanthohumol. Xanthohumol induced chromatin condensation and apoptosis and potentiated cisplatin’s effect vs cisplatin alone. Further investigation of growth inhibitory effects, xanthohumol alone induced γH2AX foci formation and the combination potentiated γH2AX foci formation. Cisplatin, xanthohumol at 25 μM, and the combination of cisplatin and xanthohumol at 6.25 and 12.5 μM increased cPARP-1 level. Active caspase-3 was increased by cisplatin, 12.5 μM of xanthohumol, and the combination of xanthohumol and cisplatin. Xanthohumol at 6.25 or 12.5 μM potentiated cisplatin effect on active caspase-3 and cPARP-1, respectively. Xanthohumol at 25 μM significtly induced the expression cell cycle control genes p21WAF1/CIP1and p14ARF. These results indicate that xanthohumol inhibits proliferation of H1299 cells and induces cell death through cleavage of PARP-1 and activation of caspase-3. The combination of cisplatin and xanthohumol potentiated cytotoxic effects of each other compound.Conclusion:The present study suggests that xanthohumol poses apoptotic effects and potentiates cisplatin’s growth inhibitory effects on metastatic lung cancer cells
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Objective To investigate the clinical diagnostic and differential diagnostic values of antisense non-coding RNA in the INK4 locus (ANRIL) and tumor suppressors (p14ARF, p15INK4b and p16INK4a) mRNA expression levels in peripheral blood lymphocytes of patients with cirrhosis and hepatocellular carcinoma. Methods The patients with hepatocellular carcinoma and cirrhosis admitted in Shantou Central Hospital from October 2013 to April 2014 were selected. The real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect ANRIL, p14ARF, p15INK4b and p16INK4a mRNA expression levels of peripheral blood lymphocytes. The subjects having taken physical health examinations in outpatient clinics were assigned in the healthy control group. Results During the study period, 19 cases of hepatocellular carcinoma, 24 cases of cirrhosis, and 31 healthy controls were finally enrolled. In the hepatocellular carcinoma group, the mRNA expression level of ANRIL was significantly higher than that of the healthy control group (?Ct:13.07±0.62 vs. 12.45±0.84, P0.05). There were also no statistically significant differences in p14ARF and p16INK4a mRNA expressions among the three groups (all P>0.05). Conclusion The elevation of ANRIL and descent of p15INK4b mRNA expression levels in peripheral blood lymphocytes in patients with liver lesion can be used as the reference indicators for the early diagnosis of hepatocellular carcinoma and to predict their prognoses.
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Purpose To explore the expression of Bmi-1, p14ARF and Mdm2 in colorectal carcinoma and to find out their association with clinicopathological features. Methods The expression of Bmi-1, p14ARF and Mdm2 was detected by immunohistochemistry in 125 cases of colorectal carcinoma and 20 cases of normal colorectal tissues. Results ( 1 ) The positive rates of Bmi-1 and Mdm2 were 56. 8% and 62. 4% in colorectal carcinoma, respectively, which were much higher than that in normal colorectal tissues (20. 0% and 15. 0%). The positive rates of p14ARF was 47. 2% in colorectal carcinoma, which was much lower than that 75. 0% in normal colorec-tal tissues ( P<0. 05 ) . ( 2 ) The expression of Bmi-1 was significantly correlated with serosal invasion, lymph node metastasis and Dukes stage. The expression of p14ARF was significantly correlated with serosal invasion and Dukes stage. The expression of Mdm2 was significantly correlated with histologic grade, lymph node metastasis and Dukes stage (P<0. 05). (3) In colorectal carcinoma, the expression of Bmi-1 and Mdm2 was negatively correlated with p14ARF, respectively (P<0. 05). Conclusions Abnormal expression of Bmi-1, p14ARF and Mdm2 is involved in the occurrence, development and metastasis of colorectal carcinoma, which could be as the reference factors in judgement of biological behavior and provide a new target for clinical treatment.
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Cervical cancer is one of the most common gynecological malignancies and is the only one which has certain pathogenesis in all malignancies currently.p14ARF is one of the tumor suppressor gene discovered recently and highly expressed in almost all cervical cancer.p14ARF has high specificity and sensitivity and it is related to invasion and prognosis of cervical cancer.Therefore,p14ARF is an ideal maker that can be used to early diagnose,screening precancerous lesions and predict prognosis in cervical cancer.
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Objective To determine the protein expression of P14ARF,MDM2 and mutant type P53 (P53mt) in skin specimens of coal-burning-type of endemic arseniasis patients and to reveal the molecular mechanism of the disease.Methods Sixty skin specimens from 60 endemic arseniasis patients including 35 of skin lesions patients,19 of precancerous lesion and 6 of skin cancer and 9 normal skin specimens from non-cancer patients were studied.Expression of P14~,MDM2 and P53mt was evaluated by immunohistochemistry using corresponding monoclonal antibodies.Results There was significant difference in the positive rates of P14ARF,MDM2 and P53mt among the 4 groups(x2 =9.39,6.21,20.64,all P ＜ 0.05).The positive rates of P14ARF in precancerous lesion and skin cancer specimens were 46.1％ (6/19) and 33.3％ (2/6),respectively,which were significantly lower than that of the normal skin specimens [88.9％(8/9),all P ＜ 0.05].Decreased expression of P14ARF was correlated with the development of dermopathy (P ＜ 0.05).The positive rates of MDM2 and P53mt in skin lesions,precancerous lesion and skin cancer specimens were 54.2％ ( 19/35 ),63.2％ (10/19),66.7％ (4/6) and 25.7％(9/35),73.7％(14/19),83.3％(5/6),respectively,which were significantly higher than those of the control (0,0,all P＜ 0.05).The expression of MDM2 and P53mt increased with the development of dermopathy(all P ＜ 0.05).Conclusions P53mt protein in skin tissue of coal-burning-type of endemic arseniasis patients is over expressed.Abnormal expression of P14ARF and MDM2 may be one of the reasons lead to abnormal cell cycle control disorders and may play a role in the development of endemic arseniasis.
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O câncer de esôfago encontra-se entre os dez tipos de câncer mais incidentes no mundo, sendo o sexto tipo mais mortal. Fatores genéticos, como mutações no gene TP53, e epigenéticos como, por exemplo, a hipermetilação das ilhotas CpG na região promotora de determinados genes, são eventos importantes no desenvolvimento do câncer e podem causar a inativação de genes supressores de tumor. Neste trabalho, avaliamos o perfil de mutação no gene TP53 em 101 pacientes com carcinoma epidermóide de esôfago (CEE) residentes na região sudeste do Brasil. Destes pacientes, 33,7% apresentaram mutações nos éxons 5, 6, 7 e 8 com prevalência nos códons 248, 179 e 220. A metilação das citosinas das ilhotas CpG resulta da atividade de uma família de enzimas, as DNA metiltransferases (DNMTs). A hipermetilação destas ilhotas leva à formação de um complexo de proteínas incluindo proteínas que têm afinidade por CpG metilado (MBDs e MeCP) impedindo que ocorra a transcrição. Neste trabalho nós investigamos, por RT-PCR semi-quantitativo, a expressão das DNMT1, DNMT3A, DNMT3B, MBD1, MBD3, MDB4 e MeCP2 em mucosa esofágica normal. Em seguida, analisamos a expressão das DNMTs, MBDs esofagina, p14ARF, p16INK4a e E-caderina em 17 amostras pareadas, tecidos normal e tumoral, de pacientes com carcinoma epidermóide de esôfago (CEE). Todas as enzimas foram constitutivamente expressas na mucosa esofágica normal. Nos tumores, foi observado um aumento significativo na expressão da DNMT3B (p=0,0038) e da MBD4 (p=0,0197) em relação à mucosa normal adjacente. A expressão dos genes esofagina, p14ARF e p16INK4a, no tecido tumoral, foi ausente ou reduzida em 64,7%, 52,9% e 58,8% das amostras, respectivamente. Apenas 11,7% das amostras de CEE mostraram níveis reduzidos de E-caderina. Quando a correlação entre a expressão da DNMTs com esofagina, p14ARF, p16INK4a e E-caderina, foi analisada pelo teste de Spearman foi observada uma correlação inversamente proporcional entre a expressão de DNMT3B e esofagina...
Esophageal cancer is one of the ten most common malignancies and it is the sixth cause of cancer-related death in the world. Genetic alterations, such as TP53 mutations and epigenetic modifications, such as the hypermethylation of CpG islands, are important events in cancer development and are a common way of inactivating tumor suppressor genes. in this study, we analyzed the spectrum of TP53 mutations in 101 patients with esophageal squamous cell carcinoma (ESCC) living in Southeastern Brazil. Among those patients, 33.7% showed mutations in exons 5, 6, 7 e 8 and these alterations are prevalent in codons 248, 179 and 220. Cytosine methylation is established and maintained by a family of DNA methiltransferase enzymes (DNMTs). Hypermethylation of CpG dinucleotides initiates the formation of a protein complex, including proteins who bind these methylated residues (MBDs and MeCP2), leading to transcriptional repression. We investigated, by RT-PCR, the expression of human DNMT1, DNMT3A, DNMT3B, MBD1, MBD2, MBD3, MBD4 and MeCP2 in normal esophageal mucosa. Then, we analyzed the mRNA expression of these DNMTs, MBDs, esophagin, p14ARF, p16INK4a and E-cacherin in 17 esophageal squamous cell carcinoma (ESCC) samples as well as their adjacent normal epithelial tissues. The expression of esophagin, p14ARF and p16INK4a was absent of reduced in 64.7%, 52.9% and 58.8% of the ESCC samples, respectively. Only 11.7% of the ESCC samples showed reduced levels of E-cadherin mRNA. When the correlation between mRNA expression of the DNMTs and these genes was analyzed by the Spearman rank test we observed that it was inversely correlated for DNMT3B and esophagin (p=0.0112), p14ARF (p=0.0384) and p16INK4a (p=0.0378). The results suggest that DMNT3B overexpression may be involved in the suppression or in the lower expression of p14ARF and p16INK4a seen in esophageal ESCC. Consequently, we selected DNMY3B, MBD4, p14ARF e p16 INK4a to be analyzed by real time PCR...
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Humans , Carcinoma, Squamous Cell/genetics , /analysis , /genetics , Epigenesis, Genetic , Gene Silencing , /genetics , DNA Methylation/genetics , Esophageal Neoplasms/genetics , DNA, Neoplasm/genetics , Sequence Analysis, DNAABSTRACT
Objective: To investigate the correlation of the expression of pokemon with p 14ARF and bcl-2 in non small cell lung cancer (NSCLC) tissues, and their values for determining the clinicopathological characteristics and prognosis of NSCLC. Methods: Immunohistochemical SP staining was used to detect the expressions of pokemon, p 14 ARF, and bcl-2 proteins in 62 NSCLC tumor tissues and 20 adjacent normal tissues. Their relationship with the pathological characteristics of NSCLC was analyzed. The correlation of abnormal expressions of pokemon and p 14 ARF and bcl-2 was analyzed. The effects of the expressions of the three proteins on the long-term prognosis of NSCLC patients were observed based on the follow-up data. Results: In normal tissues there was no pokemon expression but the positive rates of p 14 ARF and bcl-2 expression were 95.0% and 15.0%, respectively. In NSCLC tissues, the positive rate of pokemon, p 14 ARF,and bcl-2 protein were 72.6%, 66.1%, and 53.2%, respectively. The expression of pokemon had negative correlation with expression of p 14 ARF (r = - 0.287, P < 0.05), but had positive correlation with expression of bcl-2 (r = 0.293, P < 0.05). The expression of pokemon and p 14 ARF was associated with TNM staging (P < 0.05). However, the expression of bcl-2 correlated with pathological classification (P < 0.05). In pokemon and bcl-2 positive patients, the five-year survival rates were 10.95% and 13.74%, respectively, which were significantly lower than that in pokemon and bcl-2 negative patients (P < 0.05). In contrast, the five-year survival rate was 21.68% in patients with positive expression of p 14 ARF, which was significantly higher than that in patients with negative expression of p 14 ARF (P < 0.05). Conclusion: Pokemon, p 14 ARF, and bcl-2 are all expressed in NSCLC tissues. There is a negative correlation between the expression of pokemon and p 14 ARF and a positive correlation between the expression of pokemon and bcl-2, which has a certain clinical significance for evaluating the prognosis of NSCLC patients.
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The cancer is one of the most common and severe problems in clinical medicine, and nervous system tumors represent about 2% of the types of cancer. The central role of the nervous system in the maintenance of vital activities and the functional consequences of the loss of neurons can explain how severe brain cancers are. The cell cycle is a highly complex process, with a wide number of regulatory proteins involved, and such proteins can suffer alterations that transform normal cells into malignant ones. The INK4 family members (CDK inhibitors) are the cell cycle regulators that block the progression of the cycle through the R point, causing an arrest in G1 stage. The p14ARF (alternative reading frame) gene is a tumor suppressor that inhibits p53 degradation during the progression of the cell cycle. The PTEN gene is related to the induction of growth suppression through cell cycle arrest, to apoptosis and to the inhibition of cell adhesion and migration. The purpose of the present study was to assess the mutational state of the genes p14ARF, p15INK4b, p16INK4a, and PTEN in 64 human nervous system tumor samples. Homozygous deletions were found in exon 2 of the p15INK4b gene and exon 3 of the p16INK4a gene in two schwannomas. Three samples showed a guanine deletion (63 codon) which led to a loss of heterozygosity in the p15 gene, and no alterations could be seen in the PTEN gene. Although the group of patients was heterogeneous, our results are in accordance with other different studies that indicate that homozygous deletion and loss of heterozygosity in the INK4 family members are frequently observed in nervous system tumors.
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Humans , /genetics , /genetics , Nervous System Neoplasms/genetics , /genetics , DNA Mutational Analysis/methods , Gene Deletion , Homozygote , Loss of Heterozygosity , Nervous System Neoplasms/pathology , Polymerase Chain Reaction , PTEN PhosphohydrolaseABSTRACT
0.05).Conclusion: P14ARF and E2F1 can suppress colorectal tumorigenesis.The protein dysfunction is merely an early event in the tumorigenic process,correlated neither with such invasive activities as lymphonode and distant organ metastases nor with clinicopathologic characteristics.P14ARF and E2F1 have a synergenic action in suppressing cell growth and inducing cell apoptosis,and therefore can be used as potentially active gene therapeutic agents for colorectal cancer.
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PURPOSE: In this study, the level of expression of p14(ARF), p16(INK4a), p53 and pRb was immunohistochemically examined according to the stage of gastric cancer, lymph node metastasis, cell differentiation and Lauren's classification. The effect on survival rate and the associations between the components were examined as well. METHODS: One hundred and fourteen patients who underwent surgery for gastric cancer were studied retrospectively using their paraffin embedded tissue and medical records. Using antibodies of p14(ARF), p16(INK4a), p53 and pRb, immunohistochemical stain was applied and their level of expression examined. RESULTS: The level of p53 expression was high when the stage of gastric cancer was more progressed, the invasiveness higher, lymph node metastasis present and cell differentiation poorer. In contrast, the level of p14ARF expression tended to be lower as the stage was more progressed, but this was not statistically significant. Expression of p16 and pRb did not show any association with stage or other pathologic findings. Expression of p53 and p14(ARF) also had a significant association with survival rate. Survival rate was lower in patients who expressed p53 than in those who did not, but it was higher in who expressed p14ARF than in those who did not. When these two were combined, patients with p14(ARF)(+)/p53(-) had the highest survival rate, whereas those with p14(ARF)(-)/p53(+)had the lowest. This demonstrated that the expressions of p14ART and p53 have value as prognostic indicators. CONCLUSION: From these results, p53 seems closely related to stage and other pathologic findings. Furthermore, p14(ARF) and p53 showed a statistically significant relationship with survival rate, making them valuable as prognostic indicators after surgery. In combination, it would be possible to predict a more accurate prognosis.
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Humans , Antibodies , Cell Differentiation , Classification , Cyclin-Dependent Kinase Inhibitor p16 , Lymph Nodes , Medical Records , Neoplasm Metastasis , Paraffin , Prognosis , Retrospective Studies , Stomach Neoplasms , Survival Rate , Tumor Suppressor Protein p14ARFABSTRACT
PURPOSE: The methylation status of the CpG promoter regions of the p16INK4A and p14ARF genes, mutations of 4 exons of the CDKN2A gene, and the expression of the corresponding proteins were examined. Prognostic implications were assessed in osteosarcoma. MATERIALS AND METHODS: Methylation-specific PCR, sequence analysis, and immunohistochemical staining were performed upon 32 frozen osteosarcoma tissues. RESULTS: Methylation of p16INK4A was found in 16%, and methylation of p14ARF in 47%. Metastasis and poor survival was statistically related to the methylation of p14ARF. The methylation of p14ARF correlated with the repression of the corresponding protein, and repression of p14ARF with the repression of p21 and of wild type of p53. No sequence alterations were found in the four exons of the CDKN2A gene. Methylation of p14 showed highest hazard ratio by multivariate survival analysis. CONCLUSION: Our data suggest that methylation of the CDKN2A gene seems to be the main mechanism of protein repression. For p14ARF, the methylation of its promoter region was related to the repression of p21 and wild type p53, distant metastasis and a poor prognosis. Further study regarding cell cycle regulatory factors should shed light on oncogenesis and the possibility of a new treatment strategy for osteosarcoma.
Subject(s)
Carcinogenesis , Cell Cycle , Exons , Genes, p16 , Methylation , Neoplasm Metastasis , Osteosarcoma , Phosphotransferases , Polymerase Chain Reaction , Prognosis , Promoter Regions, Genetic , Repression, Psychology , Sequence Analysis , Tumor Suppressor Protein p14ARFABSTRACT
Objective To investigate the influence of co-expression state of p14 ARF and p16 INK4a protein on radiochemotherapy and length of survival period in patients with non-small cell lung cancer(NSCLC)after resection. Methods Thirty-five patients with negative p14 ARF and p16 INK4a co-expression and 20 patients with positive p14 ARF and p16 INK4a co-expression were enrolled for the study. The co-expression of the said proteins were previously determined by immunohistochemistry (S-P). Clinical pathological characteristics were compared between two groups, and the survival time and the results of radiochemotherapy of patients were respectively recorded and analysed. Results No significant differences were found in age, TNM stages, degree of differentiation, recurrence/metastasis and radiochemotherapy between two groups. However, there was significant differences in sex, smoking index, and pathological classification. It was found that 2-year, 3-year and total survival rate were significantly lower in patients with p14 ARF and p16 INK4a co-expression than those with positive co-expression (P
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Purpose:To study the effect of P14ARF gene on human glioma cell line (U 251) Methods:We transfected P14ARF gene into human malignant glioma cell line with liposomal transfection reagent, we assessed cell growth properties,cell cycle.Results:After successful transfection, the human malignant glioma cell line U 251 proliferation rate was slower. The cell cycle was suppressed at G 1 and G 2.Conclusions:The findings suggest that p14ARF gene may be able to inhibit the growth of glioma cell in vitro.
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AIM: To investigate the genetic and epigenetic alterations of p14~ ARF gene and mutation status of p53 gene in human primary colorectal carcinomas and to analyze the relationship between the two gene changes and the role of abrogation of the p14~ ARF -p53 pathway in colorectal carcinogenesis. METHODS: The homozygous deletions, mutations, methylation of 5′ CpG islands, mRNA expression of p14~ ARF gene and mutations of p53 gene were assessed by PCR, direct sequencing, methylation-specific PCR, and RT-PCR in the tumorous and matched adjacent normal colorectal tissues from 56 patients with colorectal carcinoma. RESULTS: ① p14~ ARF alterations were detected in 27% (15/56) of colorectal carcinoma tissues studied, of which 1 case showed homozygous deletion, 14 cases showed 5′ CpG island methylation, and no mutation was found in any tumor. ②15 colorectal carcinomas with p14~ ARF alterations indicated lack of (13 cases) or at low level of expression (2 cases) of p14~ ARF mRNA, while expression of the p14~ ARF transcript was detected in the remaining 41 colorectal carcinomas and any matched adjacent normal colorectal tissues. ③ The mutations of p53 gene were detected in 48% (27/56) of colorectal carcinomas investigated. ④ Of these 56 cases, 12 had p14~ ARF alterations alone, 24 had p53 mutations alone, 3 had both p53 mutations and p14~ ARF methylation, and 17 had neither. 70% (39/56) of the samples had either or both abnormalities of the two genes, and p14~ ARF hypermethylation was related to wildtype p53 (P