The role of non-protein sulphydryls in determining the chemical repair rates of free radical precursors of DNA damage and cell killing in Chinese hamster V79 cells. I

Chinese hamster V79 fibroblasts were irradiated in the gas explosion apparatus and the chemical repair rates of the oxygen-dependent free radical precursors of DNA double-strand breaks (dsb) and lethal lesions measured using filter elution (pH 9.6) and a clonogenic assay. Depletion of cellular GSH levels, from 4.16 fmol/cell to 0.05 fmol/cell, by treatment with buthionine sulphoximine (50 mumol dm-3; 18 h), led to sensitization as regards DNA dsb induction and cell killing. This was evident at all time settings but was particularly pronounced when the oxygen shot was given 1 ms after the irradiation pulse. A detailed analysis of the chemical repair kinetics showed that depletion of GSH led to a reduction in the first-order rate constant for dsb precursors from 385 s-1 to 144 s-1, and for lethal lesion precursors from 533 s-1 to 165 s-1. This is generally consistent with the role of GSH in the repair-fixation model of radiation damage at the critical DNA lesions. However, the reduction in chemical repair rate was not proportional to the severe thiol depletion (down to almost-equal-to 1% for GSH) and a residual repair capacity remained (almost-equal-to 30%). This was found not to be due to compartmentalization of residual GSH in the nucleus, as the repair rate for dsb precursors in isolated nuclei, washed virtually free of GSH, was identical to that found in GSH-depleted cells (144 s-1), also the OER remained substantially above unity. This suggests that other reducing agents may have a role to play in the chemical repair of oxygen-dependent damage. One possible candidate is the significant level of protein sulphydryls present in isolated nuclei.

Kruppel-associated Box (KRAB)-associated co-repressor (KAP-1) Ser-473 phosphorylation regulates heterochromatin protein 1β (HP1-β) mobilization and DNA repair in heterochromatin

The DNA damage response encompasses a complex series of signaling pathways that function to regulate and facilitate the repair of damaged DNA. Recent studies have shown that the repair of transcriptionally inactive chromatin, named heterochromatin, is dependent upon the phosphorylation of the co-repressor, Krüppel-associated box (KRAB) domain-associated protein (KAP-1), by the ataxia telangiectasia-mutated (ATM) kinase. Co-repressors, such as KAP-1, function to regulate the rigid structure of heterochromatin by recruiting histone-modifying enzymes, such HDAC1/2, SETDB1, and nucleosome-remodeling complexes such as CHD3. Here, we have characterized a phosphorylation site in the HP1-binding domain of KAP-1, Ser-473, which is phosphorylated by the cell cycle checkpoint kinase Chk2. Expression of a nonphosphorylatable S473A mutant conferred cellular sensitivity to DNA-damaging agents and led to defective repair of DNA double-strand breaks in heterochromatin. In addition, cells expressing S473A also displayed defective mobilization of the HP1-ß chromodomain protein. The DNA repair defect observed in cells expressing S473A was alleviated by depletion of HP1-ß, suggesting that phosphorylation of KAP-1 on Ser-473 promotes the mobilization of HP1-ß from heterochromatin and subsequent DNA repair. These results suggest a novel mechanism of KAP-1-mediated chromatin restructuring via Chk2-regulated HP1-ß exchange from heterochromatin, promoting DNA repair.

Impact damage and repair of composite structures

This paper gives an overview of the work carried out in a GARTEUR (Group for Aeronautical Research and Technology in Europe) program, under the chairmanship of the author, to develop and validate analytical and numerical methods to characterise real impact damage in composite structures, particularly those designed to sustain load in a postbuckled state, and to study the durability of bonded repairs. GARTEUR is an inter-governmental agreement between the seven European countries with the largest direct employment in the Aerospace industry, to mobilise scientific and technical knowledge between the member countries. A number of Action Groups have been launched, since GARTEUR’s inception in the early 1970s, to address specific technical issues of interest to the participating members. The research presented in this paper was performed under Action Group 28 with partners from ONERA, EADS-CCR (France), DLR, AIRBUS-Deutschland, EADS-M (Germany), CIRA (Italy), INTA (Spain), SICOMP, Saab, (Sweden), NLR (The Netherlands), QinetiQ, BAE Systems, Imperial College London and the University of Sheffield (United Kingdom). The Action Group tasks were divided into four Work Elements (WEs): WE1-Prediction and characterisation of impact damage, WE2- Postbuckling with delamination, WE3-Repair and WE4-Fatigue. This paper outlines the main developments and achievements within each Work Element.

An Experimental and Numerical Study of the Static and Fatigue Performance of a Composite Adhesive Repair

Experimental static and fatigue tension-tension tests were carried out on 5HS/RTM6 composite intact coupons and coupons incorporating adhesively-bonded (FM300-2) stepped flush joints. The results show that the adhesive joint, which is widely used in repairs, significantly reduces the static strength as well as the fatigue life of the composite. Both, the static and the fatigue failure of the ‘repaired’ coupons occur at the adhesive joint and involve crack initiation and propagation. The latter is modelled using interface finite elements based on the decohezive zone approach. The material degradation in the interface constitutive law is described by a damage variable, which can evolve due to the applied loads as well as the number of fatigue cycles. The fatigue formulation, based on a published model, is adapted to fit the framework of the pseudotransient formulation that is used as a numerical tool to overcome convergence difficulties. The fatigue model requires three material parameters. Numerical tests show that a single set of these parameters can be used to recover, very accurately, the experimental S-N relationship. Sensitivity studies show that the results are not mesh dependent.

Hepatocellular carcinoma and polymorphisms in carcinogen-metabolizing and DNA repair enzymes in a population with aflatoxin exposure and hepatitis B virus endemicity

High rates of hepatocellular carcinoma (HCC) in The Gambia, West Africa, are primarily due to a high prevalence of chronic hepatitis B virus infection and heavy aflatoxin exposure via groundnut consumption. We investigated genetic polymorphisms in carcinogen-metabolizing (GSTM1, GSTT1, HYL1*2) and DNA repair (XRCC1) enzymes in a hospital-based case-control study. Incident HCC cases (n = 216) were compared with frequency-matched controls (n = 408) with no clinically apparent liver disease. Although the prevalence of variant genotypes was generally low, in multivariable analysis (adjusting for demographic factors, hepatitis B virus, hepatitis C virus, and TP53 status), the GSTM1-null genotype [odds ratio (OR), 2.45; 95% confidence interval (95% CI), 1.21-4.95] and the heterozygote XRCC1-399 AG genotype (OR, 3.18; 95% CI, 1.35-7.51) were significantly associated with HCC. A weak association of the HYL1*2 polymorphism with HCC was observed but did not reach statistical significance. GSTT1 was not associated with HCC. The risk for HCC with null GSTM1 was most prominent among those with the highest groundnut consumption (OR, 4.67; 95% CI, 1.45-15.1) and was not evident among those with less than the mean groundnut intake (OR, 0.64; 95% Cl, 0.20-2.02). Among participants who had all three suspected aflatoxin-related high-risk genotypes [GSTM1 null, HLY1*2 (HY/HH), and XRCC1 (AG/GG)], a significant 15-fold increased risk of HCC was observed albeit with imprecise estimates (OR, 14.7; 95% CI, 1.27-169). Our findings suggest that genetic modulation of carcinogen metabolism and DNA repair can alter susceptibility to HCC and that these effects may be modified by environmental factors.

Sca-1+ progenitors derived from embryonic stem cells differentiate into endothelial cells capable of vascular repair after arterial injury

Embryonic stem cells possess the ability to differentiate into endothelium. The ability to produce large volumes of endothelium from embryonic stem cells could provide a potential therapeutic modality for vascular injury. We describe an approach that selects endothelial cells using magnetic beads that may be used therapeutically to treat arterial injury.

The use of topical anaesthesia during repair of minor lacerations in Departments of Emergency Medicine:a literature review

There are currently a number of different methods available to obtain anaesthesia in minor dermatological procedures. Although intradermal infiltration of 1% lidocaine is the favoured method for anaesthesia induction in laceration repair, it can cause significant pain in itself. Topical anaesthesia has been investigated as an alternative to infiltration anaesthesia, with the majority of studies looking at preparations of either TAC (tetracaine, adrenaline and cocaine) or LAT (lidocaine, adrenaline and tetracaine).

DNA double strand break repair: a radiation perspective

SIGNIFICANCE:
Ionizing radiation (IR) can induce a wide range of unique deoxyribonucleic acid (DNA) lesions due to the spatiotemporal correlation of the ionization produced. Of these, DNA double strand breaks (DSBs) play a key role. Complex mechanisms and sophisticated pathways are available within cells to restore the integrity and sequence of the damaged DNA molecules.
RECENT ADVANCES:
Here we review the main aspects of the DNA DSB repair mechanisms with emphasis on the molecular pathways, radiation-induced lesions, and their significance for cellular processes.
CRITICAL ISSUES:
Although the main characteristics and proteins involved in the two DNA DSB repair processes present in eukaryotic cells (homologous recombination and nonhomologous end-joining) are reasonably well established, there are still uncertainties regarding the primary sensing event and their dependency on the complexity, location, and time of the damage. Interactions and overlaps between the different pathways play a critical role in defining the repair efficiency and determining the cellular functional behavior due to unrepaired/miss-repaired DNA lesions. The repair pathways involved in repairing lesions induced by soluble factors released from directly irradiated cells may also differ from the established response mechanisms.
FUTURE DIRECTIONS:
An improved understanding of the molecular pathways involved in sensing and repairing damaged DNA molecules and the role of DSBs is crucial for the development of novel classes of drugs to treat human diseases and to exploit characteristics of IR and alterations in tumor cells for successful radiotherapy applications.

Insulin-like growth factor binding protein-3 mediates vascular repair by enhancing nitric oxide generation

Insulin-like growth factor binding protein (IGFBP)-3 modulates vascular development by regulating endothelial progenitor cell (EPC) behavior, specifically stimulating EPC cell migration. This study was undertaken to investigate the mechanism of IGFBP-3 effects on EPC function and how IGFBP-3 mediates cytoprotection following vascular injury.