P53 Gene Exons Research Articles

SINGLE AND COMBINED EFFECTS OF 5GHZ FREQUENCY WAVES AND COMPUTED TOMOGRAPHY(CT) SCAN DOSES ON WHOLE BLOOD PARAMETRES, MEMORY AND P53 GENE EXPRESSION IN RATS

AbstractBackgroundExponential increase has emerged globally in the use of non‐ionizing and ionizing radio‐frequency electromagnetic fields(Rf‐emfs) for a wide range of applications, daily activities, communication, diagnostics , therapeutics to mention but a few. However, a significant public concern is how these technologies interacts and affects the biota as electromagnetic field (EMF) emissions is linked to dysregulation of genes involved in neurotransmission,oxidative stress and normal cellular function.MethodsStandard methods (y‐maze, novel object recognition tests, automated hematology analyzer, PCR etc), were used to study how the combination of EM waves from 5Ghz radio and CT irradiation affects whole blood parameters, neurobehavioural profiles (memory and cognition), DNA fragmentation and p53 gene expression in Wistar rats which were grouped into five namely: I‐Negative control, II‐sham, III‐5Ghz only , IV‐ 5Ghz+CT, V‐CT only for fourteen days. Genomic DNA was isolated from rats’ cerebral cortex while target gene and internal control primers (GAPDH) were synthesized for p53 gene exons expression and electrophoresed on a 2.5% agarose gel.ResultsWe found that CT irradiation had higher modulatory effects on platelets, white blood cell counts and memory impairments compared to 5GHz only group. However, there was gross DNA damage and complete loss of p53 (exons 5‐7) gene expression profiles in the 5GHz while the CT irradiated rats expressed mutant p53 gene. The effect of CT irradiation in pre‐exposed 5GHz rats revealed mutant p53(exons 5‐7), elevated memory and cognition impairments amongst others .ConclusionDeleterious single effects of 5GHz was observed in p53 gene expression profiles and cognition with an interferential interaction in its combination with CT irradiation leading to mutant p53 gene expression . The impaired neurobehavioural and p53 gene profiles as a result of the single and combined 5GHz and CT exposure predicts functional radiation waves emission effects, changes in resultant protein and dysregulated cellular functions.

Mutations in p53 Gene Exons in a Sample from the South of Spain in Oral Cancer.

BackgroundCancer is a genetic disease caused by mutations in DNA and epigenetic alterations that control gene expression. The majority of epidermoid carcinomas develop within the fields of epithelial genetic alterations. The mechanisms underlying tumorigenesis of epidermoid carcinoma are as yet unknown; therefore, precise identification of the risk factors is needed. Aim: The main aim of this study is to analyse and identify the emergence of the mutations described in the literature of the p53 gene with regard to the emergence of cancer in a sample of dysplastic and cancerous lesions in oral cavity mucosa in the population of the south of Spain, in order to determine the presence of said mutations and the percentage of them in our population. Material and MethodsA cross-sectional study was carried out, with a sample size of 22 patients with potentially malignant oral lesions ancillary to biopsy. All were patients, of both sexes, over 18 years of age from the Virgen del Rocío University Hospital with potentially malignant lesions in oral mucosa ancillary to biopsy (leukoplakias, erythroplasias or leukoerythopkias). An anatomopathological study was performed on all the samples and the lesions were divided into three types: low-grade dysplasia, high-grade dysplasia and squamous cell carcinoma. In respect of the genome study process, a complete search or scan for mutations in exons 5, 6, 8 and 9 of the p 53 gene was carried out, given that in the IARC database we observed that the 5 and 6 as well as the 8 and 9 exon sizes can be scanned completely in this way, since they have amplificon sizes of 476 and 445 base pairs respectively. ResultsIn the scan for the complete exons 5, 6, 8 and 9 only a single result of interest was found to be described. In patient NBI 57 a change was observed in the TAT triplet by ATT of EXON 6, the change being of the T nucleotide by the A and in both directions both in Forward and Reverse. The exact location in the NCBI is GR Ch 37 p13 on chromosome 17, EXON 6 of the P53 gene and the change is in the C.613 T>A nucleotide; NM_000546. ConclusionsOn reviewing this genetic variant in different scientific databases, such as ENSEMBL among others, in at least 6 different biocomputing tools it is described as a pathogen, therefore we can conclude that it is a pathogenic mutation for this case in particular. The rest of the mutations described in the literature on exons 5, 6, 8 and 9 of the p53 gene have not been found in our sample. Key words:Oral cancer, p53, Mutations, Exon.

The hypermethylation of p16 gene exon 1 and exon 2: potential biomarkers for colorectal cancer and are associated with cancer pathological staging

BackgroundTumor suppressor gene p16 promoter hypermethylation has been widely studied in colorectal cancer (CRC), yet its clinicopathological significance remains controversial. The methylation alterations of other regions within p16 gene are still rarely researched. The present study aimed to explore the methylation changes of p16 gene body in CRC and to find whether they were associated with clinicopathological staging of CRC.MethodsPaired colorectal cancer tissues and corresponding adjacent normal tissues from 30 CRC patients were collected. The methylation levels of two CpG islands within p16 gene body, exon 1 and exon 2, were accurately assessed simultaneously by a LC-MS/MS method. The p16 protein expressions were assessed by immunohistochemistry assay. Statistical analyses were carried out using SPSS 17.0 software. Heat-map analysis was carried out by HemI 1.0 software.ResultsIn the present study, CRC tissues showed more highly methylated than adjacent normal tissues at both CpG islands of p16 gene. And exon 2 hypermethylation was higher and more frequent than exon 1. The ROC curve analysis showed that the simultaneous use of both indicators had excellent sensitivity and specificity for distinguishing CRC tissues and adjacent normal tissues. Following, the methylation level of p16 exon 1/2 was negatively related to p16 protein expression. Further correlation analysis revealed that p16 exon 1 hypermethylation was associated with N/Dukes staging (p = 0.033), and p16 exon 2 hypermethylaiton was associated with T staging (p = 0.035).ConclusionsThe p16 gene body was remarkably hyper-methylated in CRC tissues and associated with p16 protein expression and cancer clinicopathological staging. The combination of p16 exon 1 and exon 2 could better reflect the overall methylation status of p16 gene body and provide potential biomarkers of CRC.

Design of Novel Putative Peptide Fragments Derived from Exons of P53 Isoform G and P53 Promoter Region ORF Sequence Targeting HDM2/MDM2-P53 Interaction

p53 is a potent endogenous tumour suppressor agent which guards against DNA damage and prevents transformation of normal cells into malignant cells. In most forms of cancers except malignant melanoma, mutation occurs in p53 gene rendering it dysfunctional or suppressed. This research attempts to design novel putative bioactive oligopeptides from tumour suppressor p53 gene sequences using its promoter sequence and deriving several primer-based oligomers from the exons of p53 gene. p53 induces cells to undergo repair or apoptosis in response to DNA-damage induced stress and thus acts as the Warden of the Cell. We devise some novel methods to derive our de novo sequences from the exons of this gene and report discovery of two new oligopep-tides such as HRGRES and PAAPA (P 7 TA 7) peptide being two putative oligopeptides.

Pathways of Metabolite-Related Damage to a Synthetic p53 Gene Exon 7 Oligonucleotide Using Magnetic Enzyme Bioreactor Beads and LC-MS/MS Sequencing.

Reactive metabolites of environmental chemicals and drugs can cause site specific damage to the p53 tumor suppressor gene in a major pathway for genotoxicity. We report here a high-throughput, cell-free, 96-well plate magnetic bead-enzyme system interfaced with LC-MS/MS sequencing for bioactivating test chemicals and identifying resulting adduction sites on genes. Bioactivated aflatoxin B1 was reacted with a 32 bp exon 7 fragment of the p53 gene using eight microsomal cytochrome (cyt) P450 enzymes from different organs coated on magnetic beads. All cyt P450s converted aflatoxin B1 to aflatoxin B1-8,9-epoxide that adducts guanine (G) in codon 249, with subsequent depurination to give abasic sites and then strand breaks. This is the first demonstration in a cell-free medium that the aflatoxin B1 metabolite selectively causes abasic site formation and strand breaks at codon 249 of the p53 probe, corresponding to the chemical pathway and mutations of p53 in human liver cells and tumors. Molecular modeling supports the view that binding of aflatoxin B1-8,9-epoxide to G in codon 249 precedes the SN2 adduction reaction. Among a range of metabolic enzymes characteristic of different organs, human liver microsomes and cyt P450 3A5 supersomes showed the highest bioactivation rate for p53 exon 7 damage. This method of identifying metabolite-related gene damage sites may facilitate predictions of organ specific cancers for test chemicals via correlations with mutation sites.

Correlation between mutation of p53 gene 2-4 exons from peripheral blood and HPV16 positive cervical cancer susceptibility and clinical significance

Objective: To detect mutations of p53 gene 2-4 exons from peripheral blood and to explore their relevance in HPV16-positive cervical cancer susceptibility and clinical significance. Methods: Collected firstly cases from the Third Affiliated Hospital of Kunming Medical University from October 2012 to April 2014, included 167 cases HPV16-postive cervical cancer and 160 cases HPV-negative healthy women. Genomic DNA from the host peripheral venous blood was taken, mutations of p53 gene 2-4 exons were analyzed with software DNAstar after PCR and bidirectional sequencing. Meanwhile, mutations of p53 gene 2-4 exons among different clinicopathological characteristics in HPV16-postive cervical cancer were distinguished. Results: (1) Three mutations and an 16-bp insertion/deletion sequences were found in p53 gene exons 2-4, included C/G mutation of single nucleotide polymorphism (SNP) 11827 in intron2, A/C mutation of SNP11992 in intron3, C/G mutation in codon 72 (rs1042522) of exon4 and 16-bp (acctggagggctgggg) repeat insertion or deletion in intron3 (rs17878362), while deletion recorded as A1, insertion recorded as A2. No significant differences were found in each point allele and genotype frequency (P>0.05) . (2) Stratified analysis for cervical cancer group resulted with some differences. Compared group of non-squamous carcinoma with squamous carcinoma group, there were obviously decreased in allete A2 [11.8% (4/34) vs 3.5% (10/284) ; χ(2)=4.90, P=0.027], genotype A1A2 [4/17 vs 7.0% (10/142) ; χ(2)=5.14, P=0.023], and haplotype C-A2 [11.8% (4/34) vs 3.5% (10/284) ; χ(2)=4.91, P=0.027]. Compared with poorly differentiated group, allele C of SNP11827 and rs1042522 were obviously decreased in medium high differentiation group [50.8% (61/120) vs 38.8% (62/160) ; χ(2)=4.07, P=0.044], while haplotype G-A1 were apparently higher [49.2% (59/120) vs 61.2% (98/160) ; χ(2)=4.07, P=0.044], genotype GG of SNP11827 and rs1042522 were obviously decreased in superficial myometrial invasion depth group than that in deep myometrial invasion depth group [46.3% (25/54) vs 21.1% (8/38) ; χ(2)=7.06, P=0.029]. No significant differences were found between stage Ⅰ and Ⅱ, pelvic lymph node metastasis or not (all P>0.05) . Conclusions: No obvious correlation is found between polymorphisms in exons 2-4 of p53 gene and susceptibility of HPV16-postive cervical cancer. But the patient with allete C and A2, genotype GG and A1A2, haplotype C-A2 and G-A1 may be increase risk of poorly differentiation, deep muscular invasion and bad pathological type. Analysis of p53 gene polymorphism may be provide a basis for the prognosis evaluation and individualized treatment of cervical cancer.

Mutation of p53 Gene and Its Correlation with the Clinical Outcome in Dogs with Lymphoma.

Background p53 plays a key role in the apoptotic event induced by chemotherapeutic agents. Mutation of p53 gene has been observed in various spontaneous tumors in humans and is associated with a poor prognosis. p53 abnormalities have been evaluated in several tumors in dogs; however, the association of p53 gene mutation with clinical outcome in dogs with lymphoma has not been documented.Hypothesis/ObjectivesThe aim of this study was to examine p53 mutation in canine lymphoma cells and its association with the clinical outcome.AnimalsForty‐three dogs with previously untreated high‐grade lymphoma referred to the University of Tokyo were included in this study.MethodsProspective cohort study. We examined p53 gene (exon 4–8) mutation in the tumor tissues from 43 dogs with lymphoma using PCR‐SSCP (polymerase chain reaction – single‐strand conformational polymorphism) analysis, followed by nucleotide sequencing of the abnormal bands.ResultsOf the 43 dogs, 7 dogs (16%) had p53 mutation, whereas 36 dogs (84%) were devoid of p53 mutation. Overall response rate after remission induction was significantly lower (33% versus 88%, P = .002) in dogs with lymphomas having p53 mutation than those with lymphomas devoid of p53 mutation. Overall survival time was significantly shorter (67 days versus 264 days, P = .004) in dogs with lymphoma with p53 mutation than those with lymphoma retaining wild‐type p53.Conclusion and Clinical ImportanceMutations of p53 gene were detected in a proportion of canine lymphoma cells from untreated dogs and can be associated with a poor prognosis.

Molecular mechanisms of lung cancer development at its different stages in nuclear industry workers

to assess mutational events in exons 5, 7, and 8 of the p53 gene and to reveal mutant p53 protein in verified cases of morphologically altered (proliferative and precancerous changes, lung cancer) and histologically unaltered, lung tissues in workers exposed to occupational radiation. The investigation used formalin-fixed paraffin-embedded unaltered and altered lung tissue blocks (FFPBs) obtained from the human radiobiological tissue repository. The shelf-life of FFPBs was 5-31 years. An immunohistochemical technique using mouse antibodies against p53 protein (<<DAKO>>, Denmark), stained with diaminobenzidine (DAB) chromogen, was employed to determine p53 protein. DNA was isolated from lung tissue FFPBs with QIAmp DNA FFPE Tissue Kit, (<<QIAGEN>>, USA). Polymerase chain reaction (PCR) was performed to amplify the p53 gene exons 5, 7, and 8 selected for examination, by applying the sequences of genes and primers, the specificity of which was checked using the online resource (http://www.ncbi.nlm.nih.gov/blast). PCR products were detected by temporal temperature gradient gel-electrophoresis and the Sanger sequencing method. The obtained DNA fragments were analyzed on a sequencer ABI Prism 3100 Genetic Analizer (<<Applied Biosystems>>, USA). Computer-aided DNA analysis was made using the BLAST program. A package of applied Statistica 6.0 programs was employed for statistical data processing. Results. Immunohistochemical analysis showed that mutant p53 protein was absent in the cells of unaltered lung tissue and the number of cells with mutant p53 protein increased in all the patients with proliferative and precancerous changes and lung cancer, suggesting p53 protein dysfunction. The total number of p53 gene mutations in exons 5, 7, and 8, if there were proliferative and precancerous lung tissue changes and lung cancer, were 25, 20, and 40%, respectively. All the found mutations were transversions (the substitution of purine for pyrimidine or, conversely), indicating the action of exogenous mutagens. The results of this investigation have confirmed other investigators' data showing that p53 gene mutations in lung cancer are observed in 40-70% of cases. The differences in the number of cases of altered lung tissue with mutations in the p53 gene (not more than 40%) and in those of p53 protein expression were found in 100%, suggesting the regulation of p53 gene function in the cell at multiple levels.

Differences in the mutation of the p53 gene in exons 6 and 7 in cervical samples from HIV- and HPV-infected women

BackgroundHuman Papillomavirus (HPV) infection is a serious problem for human immunodeficiency virus (HIV)-infected women, increases their risk of cervical lesions and cancer. In cervical carcinogenesis, mutations in the p53 gene occur most frequently within exons 5–8. To our knowledge, no previous studies have analyzed mutations in exons 5–8 of the p53 gene in HIV- and HPV-infected women. In our study, we verified these mutations in women with and without cervical abnormalities.FindingsThe study included 160 women, divided into three groups: (1) 83 HPV- and HIV-infected women (HIV group); (2) 37 HPV-infected/HIV-uninfected (control group); and (3) 40 normal cytology/DNA-HPV negative/HIV-uninfected women (negative control p53 reactions). HPV-DNA was detected using polymerase chain reaction (PCR) and genotyping by PCR-restriction fragment length polymorphism analysis. Using primers for exons 5–8, the mutation of the p53 gene was verified by PCR-single strand conformational polymorphism. The total mutation of the p53 gene in exons 5–8 was not significantly associated with the HIV and control groups. The mutations in exon 7 were the highest in the HIV group (43.8%) and in exon 6 in the control group (57.2%) (p = 0.0793) suggesting a tendency toward differential mutation in exon 7 in the HIV group.ConclusionsOur study provides preliminary evidence that the mutation in exon 7 might be an important differentiating factor for cervical carcinogenesis in HIV-infected women. This aspect deserves an additional cross-sectional and longitudinal study using a larger sample size with a higher number of High-grade squamous intraephitelial lesion (HSIL) to observe the evolution of cervical lesions.

Association of intron and exon polymorphisms of p53 gene in Iranian patients with gastritis.

The propose of this study was to evaluate the probable correlation between exon and intron polymorphisms of p53 gene and their association with clinicopathological aspects of gastritis. Regarding to the decisive role of p53 in the development of a variety of human cancers, a comprehensive study concerning probable correlation between polymorphisms in the p53 intron and exon in gastritis lesions, may open new insight toward gastric cancer development and prevention. PCR-Sequencing was done for exons and introns 2-7 on the 97 gastritis and normal samples, age range of 15-83 years. Also, microsatellite status was evaluated using five mono nucleotide repeat markers. Variation at codon 72 was associated with IVS2 + 38, p53INS3 and IVS3-29. In addition, IVS2 + 38 had association with polymorphism at codon 36 & 245. Gastritis samples had stable microsatellite except nine patients showing polymorphism for NR-21 and one for Bat-25. Most of patients with stable microsatellites (83.9%) had allele G at codon72 without p53INS3. In addition, all patients with GA and CG at codon 36 / IVS2 + 38 had stable microsatellites. Severity and activity of gastritis were in association with genotypes combined of codon 36/IVS2 + 38 and 245/IVS2 + 38 respectively. In addition, the profiles of combined variation at codon 72/IVS3-29 and codon 72/IVS6 + 31 were different between patients with ages less and greater than 45 years. As, some exon variations of p53 gene specially codon 72, were in association with alterations at introns and their combined genotypes were correlated with microsatellite status, pathological findings and age, therefore, it could be inferred that the these combinations of p53 gene polymorphisms work as a whole, not as single.

Role of p53 codon 72 polymorphism in chromosomal aberrations and mitotic index in patients with chronic hepatitis B

Polymorphisms of the p53 gene, which participates in DNA repair, can affect the functioning of the p53 protein. The Arg and Pro variants in p53 codon 72 were shown to have different regulation properties of p53-dependent DNA repair target genes that can affect various levels of cytogenetic aberrations in chronic hepatitis B patients. The present study aimed to examine the frequency of chromosomal aberrations and the mitotic index in patients with chronic hepatitis B and their possible association with p53 gene exon 4 codon 72 Arg72Pro (Ex4+119 G>C; rs1042522) polymorphism. Fifty-eight patients with chronic hepatitis B and 30 healthy individuals were genotyped in terms of the p53 gene codon 72 Arg72Pro polymorphism by PCR-RFLP. A 72-h cell culture was performed on the same individuals and evaluated in terms of chromosomal aberrations and mitotic index. A high frequency of chromosomal aberrations and low mitotic index were detected in the patient group compared to the control group. A higher frequency of chromosomal aberrations was detected in both the patient and the control groups with a homozygous proline genotype (13 patients, 3 control subjects) compared to patients and controls with other genotypes [Arg/Pro (38 patients, 20 control subjects) and Arg/Arg (7 patients, 7 control subjects)]. We observed an increased frequency of cytogenetic aberrations in patients with chronic hepatitis B. In addition, a higher frequency of cytogenetic aberrations was observed in p53 variants having the homozygous proline genotype compared to variants having other genotypes both in patients and healthy individuals.

Blinded study determination of high sensitivity and specificity microchip electrophoresis-SSCP/HA to detect mutations in the p53 gene

Knowledge of the genetic changes that lead to disease has grown and continues to grow at a rapid pace. However, there is a need for clinical devices that can be used routinely to translate this knowledge into the treatment of patients. Use in a clinical setting requires high sensitivity and specificity (>97%) in order to prevent misdiagnoses. Single-strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) are two DNA-based, complementary methods for mutation detection that are inexpensive and relatively easy to implement. However, both methods are most commonly detected by slab gel electrophoresis, which can be labor-intensive, time-consuming, and often the methods are unable to produce high sensitivity and specificity without the use of multiple analysis conditions. Here, we demonstrate the first blinded study using microchip electrophoresis (ME)-SSCP/HA. We demonstrate the ability of ME-SSCP/HA to detect with 98% sensitivity and specificity >100 samples from the p53 gene exons 5-9 in a blinded study in an analysis time of <10 min.

Dietary Selenomethionine Increases Exon-Specific DNA Methylation of the p53 Gene in Rat Liver and Colon Mucosa ,

The regulation of site-specific DNA methylation of tumor suppressor genes has been considered as a leading mechanism by which certain nutrients exert their anticancer property. This study was to investigate whether selenium (Se) affects the methylation of globe genomic DNA and the exon-specific p53 gene. Three groups of rats ( n = 6–7/group) were fed the AIN-93G basal diet supplemented with 0 [Se deficient (D)], 0.15 [Se adequate (A)], or 4 mg [Se supranutritional (S)] (Se as L-selenomethionine)/kg diet for 104 d, respectively. Rats fed the A or S diet had greater plasma and liver glutathione peroxidase activity, liver thioredoxin reductase activity, and plasma homocysteine concentration than those fed the D diet. However, compared with the A diet, rats fed the S diet did not further increase these Se-dependent enzyme activities or homocysteine concentration. In contrast, Se concentrations in kidney, liver, gastrocnemius muscle, and plasma were increased in a Se-dose–dependent manner. Interestingly, rats fed the S diet had significantly less global liver genomic DNA methylation than those fed the D diet. However, the S diet significantly increased the methylation of the p53 gene (exons 5–8) but not the β-actin gene (exons 2–3) DNA in liver and colon mucosa compared with those fed the D diet. Taken together, long-term Se consumption not only affects selenoprotein enzyme activities, homocysteine, tissue Se concentrations, and global genomic DNA methylation but also increases exon-specific DNA methylation of the p53 gene in a Se-dose–dependent manner in rat liver and colon mucosa.

Relationship between the methylation and mutation of p53 gene and endemic arsenism caused by coal-burning

To explore the influence of arsenic pollution caused by coal-burning on methylation (promoter and exon 5) and mutation (exon 5) of human p53 gene, and to analyze the relationship between methylation, mutation and arsenism. According to the diagnostic criteria of endemic arsenism, 112 patients with arsenism (including 38 mild cases, 43 moderate cases and 31 severe cases) were selected in the areas with endemic arsenism from Xingren, Guizhou province. Among the subjects, 43 cases were diagnosed by dermatopathological methods, and they were divided into non-cancerous group (24 cases) and cancerous group (19 cases). 90 controls were selected from the non-arsenic polluted areas. Under the principle of informed consent, blood samples were collected from individuals. The methylation of p53 gene in promoter region and exon 5 were detected by extinction enzyme-PCR, the mutation of p53 gene (exon 5) was detected by PCR-SSCP, PCR products cloning and sequencing technology. The positive rates of methylation of p53 gene in promoter region were 13.16% (5/38), 27.91% (12/43) and 45.16% (14/31) respectively among mild, moderate and severe arsenism group, which were obviously higher than the rates in the control group (1.11% (1/90), χ² values were 8.679, 23.690, 41.199, respectively, both P values < 0.017). The positive rates of methylation of p53 gene were 25.00% (6/24) and 63.16% (12/19) in non-cancerous and cancerous group respectively, which were obviously higher than those in the control group (1.11% (1/90), χ² values were 18.762, 57.497, respectively, both P values < 0.025). The positive rates of methylation of p53 gene (exon 5) were 55.26% (21/38), 51.16% (22/43) and 48.39% (15/31) respectively among mild, moderate and severe arsenism group, which were obviously lower than the rates in the control group (88.88% (80/90), χ² values were 18.151, 23.168, 22.420, respectively, both P values < 0.017). The positive rates of methylation of p53 gene (exon 5) were 54.17% (13/24) and 42.11% (8/19) in non-cancerous and cancerous group respectively, which were obviously lower than those in the control group (88.88% (80/90), χ² values were 15.201, 22.075, respectively, both P values < 0.025). The mutation rates of p53 gene (exon 5) were respectively 5.26% (2/38), 16.28% (7/43) and 25.81% (8/31) among mild, moderate and severe arsenism group; while the results in moderate and severe arsenism group were obviously higher than in the control group (0.00%, χ² values were 15.465, 24.870, respectively, both P values < 0.017). The positive rate of mutation of p53 gene (exon 5) were respectively 16.67% (4/24) and 31.58% (6/19) in non-cancerous and cancerous group, which were obviously higher than it in the control group (0.00%, χ² values were 15.545, 30.077, both P values < 0.025). The hypermethylation of p53 gene in promoter region was related with the mutation of p53 gene (exon 5) (coefficient of association was 0.294, P value < 0.05); and the hypomethylation of p53 gene (exon 5) was related with the its mutation (coefficient of association was 0.410, P value < 0.05). Arsenic pollution caused by coal-burning can cause the hypermethylation of p53 gene in promoter region, hypomethylation and mutation of p53 gene (exon 5), and the changes of methylation of p53 gene are related with its mutation and might be one of the important etiological factors of arsenic pathogenicity or carcinogenesis.

A combined histologic and molecular approach identifies three groups of gastric cancer with different prognosis

The limited prognostic value of currently used histologic classifications of gastric cancer and their failure to account for the complexity of the disease as revealed by more recent investigations prompted a combined reinvestigation of histologic, molecular, and clinicopathologic patterns in 294 extensively sampled, invasive gastric cancers representing all main histotypes and stages of the disease and followed for a median of 150 months. Among histologic parameters tested, only cellular atypia, angio-lympho- or neuroinvasion, Ki67 proliferation index, expansile/infiltrative type growth, and T8 cell-rich high lymphoid intra-/peritumor response (HLR) proved to be stage-independent predictors of patient survival. Among molecular tests, p53 gene exon 7 (loop 3) and 8 (loop-sheet-helix motif and S-10 band), but not p53 protein overexpression, TP53 LOH or 18qLOH, were found to worsen prognosis. Microsatellite DNA instability was a favorable prognostic factor when coupled with HLR. Patient survival analysis of the main histotypes and their subtypes confirmed the favorable prognosis of HLR, well-differentiated tubular, muconodular, and low grade diffuse desmoplastic cancers, and highlighted the worse prognosis of anaplastic and infiltrative-lymphoinvasive mucinous cancers compared to ordinary cohesive and diffuse cancers. Distinct roles of individual morphologic and molecular factors in tumor progression of the different histotypes have been recognized. The combination of survival-predictive histotypes and individual histologic or molecular parameters allowed us to develop a classification of all gastric cancers into three grades of increasing malignancy which proved to be of high prognostic value.

Molecular analysis of mutations in thyroid tumors with TGGE.

Immunohistochemical demonstration of overexpression of the p53 protein indicates a mutational alteration of the gene. Our own investigations of 59 differentiated thyroid carcinomas revealed an overexpression in 15% of the tumors. A correlation to unfavourable tumor prognosis was found (stage I and II: 0/11 (0%); stage III: 4/26 (14%); stage IV: 5/22 (23%)). For screening of one out of more than 300 possible mutations temperature gradient gel electrophoresis was employed. Analysis of the highly-conserved regions of the p53 gene (exon 5 to 8) could demonstrate a mutation in only 1 out of 31 differentiated thyroid carcinomas. The question arises whether accumulation of the protein is due to a mutational event or rather other molecular mechanisms.

Transformation analysis of oral epithelial dysplasia to carcinoma in-situ and squamous cell carcinoma by p53 expression and gene mutations

Background : It is known that oral squamous epithelial dysplasia (SED) and carcinoma in-situ (CIS) are precancerous lesion and it could transform to squamous cell carcinoma (SCC). We had reported p53-Protein Over-Expression and Gene Mutational of Oral CIS, such as basaloid, verrucous, and acanthothic/atrophic types, but demarcated between SED to CIS and CIS to SCC and how their transformation is still unclear. It is considered that their molecular behavior related one another. Purpose : To understand the molecular behavior of them we examined p53 exon 5-8 gene mutation and their protein expression in the sequential cases. Methods : Using 10 cases formalin–fixed paraffin sections that composed SED appearance, CIS and SCC in the same case were subjected to P53 immunohistochemistry. Then all cases were subjected to p53 gene mutations analysis. By laser capturing microdissection dysplasia part, CIS part and SCC part were cutted, and followed by direct sequencing of PCR product for exon 5-8. Result : SED p53-protein over-expression in some cells, and the expression was increased to CIS and SCC. Mutational analysis for p53 gene showed that 60% of p53 gene mutation in CIS also found in SCC, therefore SCC had additional mutation in other exon of p53 gene. While no particular mutations were found in SED part of all cases. Conclusion : Carcinoma in-situ is a squamous cell carcinoma eventhough not invasive yet, but squamous epithelial dysplasia is an early step to malignancy. It needs other genes examination to know any genes are involved in the precancerous to cancer transformation process.

FREQUENCY OF THE CORRELATION BETWEEN MUTATION IN 7TH EXON OF P53 GENE AND THE PROGNOSIS OF COLORECTAL CANCER

Introduction: Colorectal cancer (CRC) is one of the most prevalent cancers in the world. Genetic disorders and dietary habits play a part in CRC. The most important cancer suppressor gene is P53. Different mutations rates in 7th exon of p53 gene are reported in different areas which may have a relationship with prognosis of CRC. This study examines the relationship between mutation in 7th exon of P53 gene and the prognosis of CRC. Materials and Methods: This descriptive analytical study included 80 CRC cases hospitalized in Isfahan between 2003 and 2007. The DNA samples were extracted through phenol chloroform isoamil alcohol. In genetic department of Isfahan medical faculty. The 7th exon of P53 gene was amplified using specific primers in a PCR assay. After that mutation was detected by gel electrophoresis and SSCP analysis. P53 gene expression was detected by immunohistochemical methods. Data was analyzed using chi-square test. Results: From the total of 80 specimens, 4 (5%) had mutation in 7th exon. The positive specimens demonstrated over expression for p53 gene. There was a significant (p 0.05). Two samples with mutations as well as one sample without mutation showed metastases in the liver tissue. The prognosis of CRC was 3 years after diagnosis. And follow-up was less in patients with mutation compared to those without mutation. Conclusion: The result showed that all mutations in 7th exon of p53 are accompanied by p53 over expression. The mutation in 7th exon of p53 gene is associated with poor prognosis in CRC.

Expression of immunoreactivity and genetic mutation in eosinophilic and ciliated metaplastic changes of endometrial glandular and stromal breakdown: cytodiagnostic implications

Various metaplastic changes may be present in endometrium, in which also cellular atypias may often be observed. Particularly, eosinophilic and ciliated changes (ECCs) occur in both nonneoplastic and neoplastic endometrium. This may cause confusion in the cytodiagnosis. This study was enterprised to investigate the possible help of immunocytochemical and cytogenetic study in the diagnostic and biologic assessment of ECC cells. In immunocytochemistry for p53 protein, Ki-67, and cyclin A, the material consists of 40 cases of cytologic smears examined by direct sampling of the endometrial cavity comprising 30 cases of ECC in endometrial glandular and stromal breakdown (EGBD) and 10 cases of well-differentiated adenocarcinoma (G1). After cytodiagnosis, immunostaining for p53 protein, Ki-67, and cyclin A was performed on multiple wet-fixed slides from each single case to evaluate the immunoreactivity, intensity of nuclear staining, and nuclear labeling index (N-LI). The intensity of nuclear staining was scored as negative (0), weak (1), moderate (2), or strong (3), and the N-LI was scored as less than 10% (0), from 10% to 25% (1), from 26% to 50% (2), or more than 50% (3), and the final score was calculated by adding both partial scores. A statistical significance test was performed by using Mann-Whitney U test, and the result was judged as significant when the P value was less than .05. For genetic mutation analysis of p53, the material comprised 6 cases of EGBD in which a high score was measured with immunocytochemistry for p53 protein, and the presence of ECC was confirmed on the hematoxylin and eosin. The ECC cells on paraffin-embedded specimens were captured using laser capture microdissection technology. Mutations in p53 gene (exons 5-8) were examined using DNA sequencing analysis. In immunocytochemistry for p53 protein, Ki-67, and cyclin A, the proportions of immunoreactive cells for p53 were statistically higher in ECC compared with those of G1 (P < .05). The proportions of the immunoreactive cells for Ki-67 and cyclin A were statistically higher in G1 compared with those of ECC (P < .05). (2) In genetic mutation analysis of p53, DNA sequencing of p53 in 6 cases revealed no mutations. The percentage of immunoreactive cells for p53 protein were higher in ECC than in G1, but the mutation point was not confirmed in genetic mutation analysis. The differential expression of these biologic parameters in ECC cells could be considered of possible relevance to the cytopathologic diagnosis in the future.

P53 gene mutational rate, Gleason score, and BK virus infection in prostate adenocarcinoma: Is there a correlation?

Prostate cancer represents the second leading cause of cancer deaths in Western countries. Viral infections could play a role in prostate carcinogenesis. Human polyomavirus BK (BKV) is a possible candidate because of its transforming properties. In this study, BKV sequences in urine, blood, fresh, and paraffin-embedded prostate cancer samples from 26 patients were searched using Q-PCR analysis. T antigen (TAg) and p53 localization in neoplastic cells were evaluated by immunohistochemical analysis. Also, the presence of mutations in 5-9 exons of p53 gene was analyzed. Results showed that BKV-DNA was found in urine (54%), plasma (31%), and in fresh prostate cancer specimens (85%). The analysis of p53 gene evidenced several mutations in high Gleason patients, according to tumor advanced stage. Immunohistochemical analysis results evidenced the localization of p53 and TAg into cytoplasm, whereas in TAg-negative tumors, p53 was nuclear. This study suggests that BKV acts as cofactor in the pathogenesis of prostate cancer. These observations emphasize previous studies regarding the cellular pathways that may be deregulated by BKV.