Immuno-spin trapping of protein and DNA radicals: “Tagging” free radicals to locate and understand the redox process

Biomolecule-centered radicals are intermediate species produced during both reversible (redox modulation) and irreversible (oxidative stress) oxidative modification of biomolecules. These oxidative processes must be studied in situ and in real time to understand the molecular mechanism of cell adaptation or death in response to changes in the extracellular environment. In this regard, we have developed and validated immuno-spin trapping to tag the redox process, tracing the oxidatively generated modification of biomolecules, in situ and in real time, by detecting protein- and DNA-centered radicals. The purpose of this methods article is to introduce and update the basic methods and applications of immuno-spin trapping for the study of redox biochemistry in oxidative stress and redox regulation. We describe in detail the production, detection, and location of protein and DNA radicals in biochemical systems, cells, and tissues, and in the whole animal as well, by using immuno-spin trapping with the nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide.     

Crystal Structure of an RluF-RNA Complex: A Base-Pair Rearrangement Is the Key to Selectivity of RluF for U2604 of the Ribosome

Escherichia coli pseudouridine synthase RluF is dedicated to modifying U2604 in a stem-loop of 23S RNA, while a homologue, RluB, modifies the adjacent base, U2605. Both uridines are in the same RNA stem, separated by ∼ 4 Å. The 3.0 Å X-ray crystal structure of RluF bound to the isolated stem-loop, in which U2604 is substituted by 5-fluorouridine to prevent catalytic turnover, shows RluF distinguishes closely spaced bases in similar environments by a selectivity mechanism based on a frameshift in base pairing. The RNA stem-loop is bound to a conserved binding groove in the catalytic domain. A base from a bulge in the stem, A2602, has folded into the stem, forcing one strand of the RNA stem to translate by one position and thus positioning U2604 to flip into the active site. RluF does not modify U2604 in mutant stem-loops that lack the A2602 bulge and shows dramatically higher activity for a stem-loop with a mutation designed to facilitate A2602 refolding into the stem with concomitant RNA strand translation. Residues whose side chains contact rearranged bases in the bound stem-loop, while conserved among RluFs, are not conserved between RluFs and RluBs, suggesting that RluB does not bind to the rearranged stem loop.     

The Effect of Different Levels of Organic and Inorganic Chromium Supplementation on Production Performance, Carcass Traits and Some Blood Parameters of Broiler Chicken Under Heat Stress Condition

A total of 250 broilers in a completely randomized design selected to evaluate the effect of different levels of chromium (Cr) supplementation on performance, carcass traits and some blood parameters of heat-stressed broiler chicks. All birds were kept under heat stress temperature (33?±?3°C) and divided into five treatments groups. Each treatment consisted of five pens with 10 birds in each pen. The basal diets were supplemented with 0?ppb (control), 800?ppb Cr-L-Met (T(1)), 1,200?ppb Cr-L-Met (T(2)) or 800?ppb CrCl(3) (T(3)), and 1,200?ppb CrCl(3) (T(4)). The feed intake and body mass were measured at 10, 21, and 42?days of age. Blood samples were collected from two birds in each replicates to determine biological and hematological values at 28 and 42?days of age. There were no significant difference in mass gain and feed conversion of broilers that received Cr supplementations compared with controls. The serum glucose concentration decreased in broilers received organic chromium methionine supplements compared with other treatments groups. Slight but not significant increases were observed in serum high-density lipoprotein (HDL) concentration of treated groups than controls while the mean serum HDL concentration was significantly higher in T(2) group compared with control group. Serum low-density lipoprotein level decreased in broiler received organic Cr supplements (p?相似文献   

The effects of pulsed magnetic field exposure on the permeability of leukemia cancer cells

Purpose: The newer methods of cancer treatment require new idea of drug delivery in cancer cells. Due to numerous researches electromagnetic field affect on cell function and cell membrane for possible therapeutic and drug delivery. In this article, we determined in vitro uptake of fluorescent dyes into the attached K562 cells due to time-varying magnetic field exposure. Method and material: The K562 cells were exposed to magnetic pulses via Magstim stimulator and double 70?mm coil. The strength and duration of pulses in all experiments were the same and three different frequencies of 0.25, 1 and 10?Hz pulses for 56, 112 and 28 numbers of pulses were applied (nine experimental groups) and uptake of Ly and PI was measured in each group. Result: Our results show that magnetic field can efficiently increase permeability. Among the treatment groups, the system gives the optimal permeabilization when cells are exposed to a train of 28 pulses with 1?Hz frequency.     

Mesothelin Expression in Triple Negative Breast Carcinomas Correlates Significantly with Basal-Like Phenotype,Distant Metastases and Decreased Survival

Mesothelin is a cell surface associated antigen expressed on mesothelial cells and in some malignant neoplasms. Mesothelin-targeted therapies are in phase I/II clinical trials. The clinicopathologic and prognostic significance of mesothelin expression in triple negative breast carcinomas (TNBC) has not been fully assessed. We evaluated the expression of mesothelin and of basal markers in tissue microarrays of 226 TNBC and 88 non-TNBC and assessed the clinicopathologic features of mesothelin-expressing breast carcinomas. Furthermore, we investigated the impact of mesothelin expression on the disease-free and overall survival of patients with TNBC. We found that mesothelin expression is significantly more frequent in TNBC than in non-TNBC (36% vs 16%, respectively; p = 0.0006), and is significantly correlated with immunoreactivity for basal keratins, but not for EGFR. Mesothelin-positive and mesothelin-negative TNBC were not significantly different by patients’ race, tumor size, histologic grade, tumor subtype, lymphovascular invasion and lymph node metastases. Patients with mesothelin-positive TNBC were older than patients with mesothelin-negative TNBC, developed more distant metastases with a shorter interval, and had significantly lower overall and disease-free survival. Based on our results, patients with mesothelin-positive TNBC could benefit from mesothelin-targeted therapies.     

Citric Acid Cycle and Role of its Intermediates in Metabolism

The citric acid cycle is the final common oxidative pathway for carbohydrates, fats and amino acids. It is the most important metabolic pathway for the energy supply to the body. TCA is the most important central pathway connecting almost all the individual metabolic pathways. In this review article, introduction, regulation and energetics of TCA cycle have been discussed. The present study was carried out to review literature on TCA cycle.     

Molecular mechanism and therapy application of necrosis during myocardial injury

Necrosis is an ancient topic which gains new attraction in the research area these years. There is no doubt that some necrosis can be regulated by genetic manipulation other than an accidental cell death resulting from physical or chemical stimuli. Recent advances in the molecular mechanism underlying the programmed necrosis show a fine regulation network which indicates new therapy targets in human diseases. Heart diseases seriously endanger our health and have high fatality rates in the patients. Cell death of cardiac myocytes is believed to be critical in the pathogenesis of heart diseases. Although necrosis is likely to play a more important role in cardiac cell death than apoptosis, apoptosis has been paid much attention in the past 30 years because it used to be considered as the only form of programmed cell death. However, recent findings of programmed necrosis and the related signalling pathways have broadened our horizon in the field of programmed cell death and promote new pharmacological application in the treatment of heart diseases. In this review, we summarize the advanced progress in these signalling pathways and discuss the pathos‐physiological relevance and therapeutic implication of targeting necrosis in heart diseases treatment.     

The response of transgenic <Emphasis Type="Italic">Brassica</Emphasis> species to salt stress: a review

Salt stress is considered one of the main abiotic factors to limit crop growth and productivity by affecting morpho-physiological and biochemical processes. Genetically, a number of salt tolerant Brassica varieties have been developed and introduced, but breeding of such varieties is time consuming. Therefore, current focus is on transgenic technology, which plays an important role in the development of salt tolerant varieties. Various salt tolerant genes have been characterized and incorporated into Brassica. Therefore, such genetic transformation of Brassica species is a significant step for improvement of crops, as well as conferring salt stress resistance qualities to Brassica species. Complete genome sequencing has made the task of genetically transforming Brassica species easier, by identifying desired candidate genes. The present review discusses relevant information about the principles which should be employed to develop transgenic Brassica species, and also will recommend tools for improved tolerance to salinity.     

Kinetin and Indole Acetic Acid Promote Antioxidant Defense System and Reduce Oxidative Stress in Maize (Zea mays L.) Plants Grown at Boron Toxicity

Journal of Plant Growth Regulation - A study was conducted to assess the influence of boron (B) toxicity on functioning of antioxidant machinery to counteract oxidative stress in maize (Zea mays...