74 research outputs found
Non-muscle invasive bladder cancer tissues have increased base excision repair capacity
No full textThe molecular mechanisms underlying the development and progression of bladder cancer (BC) are complex and have not been fully elucidated. Alterations in base excision repair (BER) capacity, one of several DNA repair mechanisms assigned to preserving genome integrity, have been reported to influence cancer susceptibility, recurrence, and progression, as well as responses to chemotherapy and radiotherapy. We report herein that non-muscle invasive BC (NMIBC) tissues exhibit increased uracil incision, abasic endonuclease and gap-filling activities, as well as total BER capacity in comparison to normal bladder tissue from the same patient (p<0.05). No significant difference was detected in 8-oxoG incision activity between cancer and normal tissues. NMIBC tissues have elevated protein levels of uracil DNA glycosylase, 8-oxoguanine DNA glycosylase, AP endonuclease 1 and DNA polymerase beta protein. Moreover, the fold increase in total BER and the individual BER enzyme activities were greater in high-grade tissues than in low-grade NMIBC tissues. These findings suggest that enhanced BER activity may play a role in the etiology of NMIBC and that BER proteins could serve as biomarkers in disease prognosis, progression or response to genotoxic therapeutics, such as Bacillus Calmette-Guerin.Muftuoglu, M (corresponding author), Acibadem Mehmet Ali Aydinlar Univ, Dept Med Biotechnol, TR-34752 Istanbul, Turkey ; Acibadem Mehmet Ali Aydinlar Univ, Dept Mol Biol & Genet, TR-34752 Istanbul, Turkey.
[email protected]
Evaluation of bony changes observed in mid-palatal suture after rapid maxillary expansion using fractal analysis
No full text
Werner syndrome protein: Functions in the response to DNA damage and replication stress in S-phase
No full textBacillus Calmette-Guerin Increases Base Excision Repair in Bladder Cancer Cells
No full textObjective:Most patients with non-muscle-invasive bladder cancer (NMIBC) do not respond to intravesical Bacillus Calmette-Guerin (BCG) immunotherapy and have high risk of NMIBC recurrence and progression. In addition to its therapeutic effect which increases the local immune response, BCG also exerts an anti-tumour effect by increasing oxidative stress, and producing reactive oxygen species and oxidative DNA damage in bladder cancer (BC) cells. The oxidative DNA damage is repaired by base excision repair (BER) mechanism. Thus, BER capacity of BC cells could be an important factor in response to BCG therapy. Effects of BCG on the activity of BER in BC transitional carcinoma cell line, T24 have been investigated.Materials and Methods:The uracil-initiated total BER and BER enzyme activities were measured in whole cell extracts with or without BCG treatment using a [γ-32P] adenosine triphosphate-labelled 51-mer DNA substrates.Results:BCG treatment increased the activities of uracil-initiated total BER and BER enzymes, uracil DNA glycosylase and DNA polymerase β in 6 h and 24 h repair periods and increased the activity of 8-oxoguanine DNA glycosylase in 6 h repair in T24 BC cell line.Conclusion:The enhanced BER activity in BC cells in response to BCG treatment could be an important factor in BCG resistance
Evaluation of anterior and posterior corneal aberrations in patients with keratoconus
No full textEvaluation of anterior and posterior corneal aberrations in patients with keratoconusPoster DetailsFirst Author: E.AltinkurtTURKEYCo Author(s): O. Muftuoglu S. Karaman Erdur Abstract DetailsPurpose:To evaluate the anterior and posterior corneal aberrations in patients with keratoconus.Setting:The study is made in a tertiary referral center.Methods:100 eyes of 57 patients with clinical keratoconus and 100 eyes of 50 control patients who were refractive surgery candidates were included in this study. All eyes were evaluated before and 6-months after surgery using Placido-Scheimpflug combined topo/tomographer. Visual acuity, keratometry, anterior, posterior, and total corneal aberrations were analyzed.Results:The mean Ksteep was 48.6 ± 4.8 D in keratoconus group, and 43.2 ±2.1 D in control group. The mean anterior corneal total higher order aberration, coma, and spherical aberration were 1.83±1.39 µm, 1.44 ± 0.92 µm, 0.31 ± 0.29 µm, respectively. The mean posterior corneal total higher order aberration, coma, and spherical aberration were 1.04 ± 1.63 µm, 0.36 ± 0.32 µm, 0.16 ± 0.21 µm, respectively. The mean anterior and posterior corneal total higher order aberration, coma, and spherical aberration were significantly higher than those of control groups (p˂0.05). The differences were more prominent in posterior corneal aberrations.Conclusions:Corneal higher order aberrations, particularly posterior corneal aberrations are significantly higher in eyes with keratoconus compared to normal controls
Acetylation regulates WRN catalytic activities and affects base excision DNA repair.
No full textThe Werner protein (WRN), defective in the premature aging disorder Werner syndrome, participates in a number of DNA metabolic processes, and we have been interested in the possible regulation of its function in DNA repair by post-translational modifications. Acetylation mediated by histone acetyltransferases is of key interest because of its potential importance in aging, DNA repair and transcription.Here, we have investigated the p300 acetylation mediated changes on the function of WRN in base excision DNA repair (BER). We show that acetylation of WRN increases in cells treated with methyl methanesulfonate (MMS), suggesting that acetylation of WRN may play a role in response to DNA damage. This hypothesis is consistent with our findings that acetylation of WRN stimulates its catalytic activities in vitro and in vivo, and that acetylated WRN enhances pol beta-mediated strand displacement DNA synthesis more than unacetylated WRN. Furthermore, we show that cellular exposure to the histone deacetylase inhibitor sodium butyrate stimulates long patch BER in wild type cells but not in WRN depleted cells, suggesting that acetylated WRN participates significantly in this process.Collectively, these results provide the first evidence for a specific role of p300 mediated WRN acetylation in regulating its function during BER
Parents of ataxia-telangiectasia patients display a distinct cellular immune phenotype mimickingATM-mutated patients
No full textBackground Heterozygous relatives of ataxia-telangiectasia (AT) patients are at an increased risk for certain AT-related manifestations. We also show that there is an increase of infection frequency in parents of AT patients. Thus, we hypothesized that the parents might exhibit immune alterations similar to their affected children. Methods Lymphocyte phenotyping to enumerate T- and B-cell subsets was performed. Functional analyses included in vitro quantified gamma-H2AX, poly (ADP-ribose) polymerase (PARP) and caspase-9 proteins. Chromosomal instability was determined by comet assay. Results We analyzed 20 AT patients (14F/6M), 31 parents (16F/15M), and 35 age-matched healthy controls. The AT patients' parents exhibited low frequency of naive CD4(+)T- (n = 14, 45%) and recent thymic emigrants (n = 11, 35%) in comparison with the age-matched healthy donors. Interestingly, parents with low naive T cells also demonstrated high rate of recurrent infections (9/14, 64%). In comparison with age-matched controls, parents who had recurrent infections and low naive T cells showed significantly higher baseline gamma-H2AX levels and H2O2-induced DNA damage as well as increased cleaved caspase-9 and PARP proteins. Conclusion Parents of AT patients could present with recurrent infections and display cellular defects that mimic AT patients. The observed immunological changes could be associated with increased DNA double-strand breaks
Simple, high-yield purification of xanthine oxidase from bovine milk
No full textXanthine oxidase, a commercially important enzyme with a wide area of application, was extracted from fresh milk, without added preservatives, using toluene and heat. The short purification procedure, with high yield, consisted of extraction, ammonium sulfate fractionation, and DEAE-Sepharose (fast how) column chromatography. Xanthine oxidase was eluted as a single activity peak from the column using a buffer gradient. The purification fold, specific activity and yield for the purified xanthine oxidase were 328, 10.161 U/mg and 69%, respectively. The enzyme was concentrated by ultrafiltration, although 31% of the activity was lost during concentration, no change in specific activity was observed. Activity and protein gave coincident staining bands on native polyacrylamide gels. The intensity and the number of bands were dependent on the oxidative state(s) of the enzyme; reduction by 2-mercaptoethanol decreased the intensity of the slow-moving bands and increased the intensity of the fastest-moving band. Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), two major bands (molecular masses of 152 and 131 kDa) were observed, accounting for greater than or equal to 95% of xanthine oxidase. Native- and SDS-PAGE showed that the purified xanthine oxidase becomes a heterodimer due to endogenous proteases. (C) 1999 Elsevier Science B.V. All rights reserved
Cosmic Whirl: Navigating the Comet Trail in DNA: H2AX Phosphorylation and the Enigma of Uncertain Significance Variants
No full textPathogenic variations in the BRCA2 gene have been detected with the development of next-generation sequencing (NGS)-based hereditary cancer panel testing technology. It also reveals an increasing number of variants of uncertain significance (VUSs). Well-established functional tests are crucial to accurately reclassifying VUSs for effective diagnosis and treatment. We retrospectively analyzed the multi-gene cancer panel results of 922 individuals and performed in silico analysis following ClinVar classification. Then, we selected five breast cancer-diagnosed patients’ missense BRCA2 VUSs (T1011R, T1104P/M1168K, R2027K, G2044A, and D2819) for reclassification. The effects of VUSs on BRCA2 function were analyzed using comet and H2AX phosphorylation (γH2AX) assays before and after the treatment of peripheral blood mononuclear cells (PBMCs) of subjects with the double-strand break (DSB) agent doxorubicin (Dox). Before and after Dox-induction, the amount of DNA in the comet tails was similar in VUS carriers; however, notable variations in γH2AX were observed, and according to combined computational and functional analyses, we reclassified T1001R as VUS-intermediate, T1104P/M1168K and D2819V as VUS (+), and R2027K and G2044A as likely benign. These findings highlight the importance of the variability of VUSs in response to DNA damage before and after Dox-induction and suggest that further investigation is needed to understand the underlying mechanisms
- …
