Oxidative DNA damage, resulting from ROS, increases with age and can accumulate as a variety ofoxidative modifications in purines and pyrimidines [27–28]. Oxidized bases may lead to mutagenesis, block DNA replication, or alter the affinity of DNA binding proteins, which can, in turn, attenuate cell viability or promote tumorigenesis [29–31]. BER is the primary DNA repair pathway for the repairof non-bulky damaged bases, and the initial step in BER is base removal by a DNA glycosylase. Several DNA glycosylases with distinct, but overlapping substrate specificities have been characterized, and OGG1 primarily excises 8-oxoG and FapyG paired with cytosine in duplex DNA [30–31]. OGG1 is well conserved from bacterial to mammals, implying its significant functional importance in maintaining genome integrity [27–28]. If 8-oxoG is unrepaired, it becomes highly mutagenic, because it can pair with adenine and lead to GC to TA transversions after two rounds of replication [32–34]. The removal of adenine opposite 8-oxoG is via the adenine-specific mismatch DNA glycosylase, MYH . Mice lacking these repair genes exhibit an increased spontaneous mutation rate and a marked increase in tumor predisposition [35–38]. In addition, Ogg1 and Myh deficient murine cells are sensitive to oxidative stress [39–41]. These studies are consistent with the idea that oxidative base lesions contribute to genome instability, neoplastic transformation, and cell death.
Reactive oxygen species (ROS) are continuously produced in all living beings, especially in higher organisms, as a result of normal cellular metabolism, phagocytises, inflammation, and exogenous factors such as ionizing radiations and xenobiotics. 1 ROS can induce cell damage by reacting with biomolecules (proteins, lipids) and cause serious lesions in the DNA molecule, 2 such as strand breaks, DNA-protein cross-linking, and base-free sites. 3 The mammalian body has certain endogenous antioxidant defense mechanisms to combat and reduce oxidativedamage such as enzymatic systems, and exogenous antioxidant systems, such as as vitamins, minerals, and proteins. Antioxidants, which can inhibit or delay the oxidation of a substrate in a chain reaction, therefore, appear to be very important in the prevention of many diseases. 4 Foodstuffs constitute an excellent exogenous source of natural antioxidants. It is known that vegetables, fruits, whole-grain, and some beverages (tea, juice, wine) are rich in antioxidants and bioactive compounds. Examples of antioxidants present in food are vitamins (particularly C and E), phenolic compounds (flavonoids, catechins, flavones, flavonols, anthocyanins), and carotenoids including β-carotene. 5 A healthy diet should provide an adequate and continuous supply of these antioxidants. Other antioxidants, such as ubiquinol and thiol compounds, produced in small amounts by the organism, can be obtained in higher amounts by dietary supplements. 6 Consequently, interest is increasing in new effective natural antioxidants as well as in the chemical and biochemical characterizationof foodstuffs and beverages to evaluate them with regard to their antioxidant profiles.
TbPolβ characterization also showed that, in addition to a mitochondrial localization, it is active as a DNA polymerase and as a lyase . The cellular localization of these polymerases highlights an important feature of the Tritryps: the presence of kDNA. The kDNA structure is so complex that it requires an unusual replication mechanism, which differs from higher eukaryotes [ 55, 56]. This complexity is reflected in the DNA repairand replication machinery that can be localized to this organelle . Polβ is an example of a polymerase that shows a nuclear localization in higher eukaryotes  but is addressed to the kinetoplast in the Tritryps [21, 22]. The L. major Polβ has not yet been exper- imentally characterized; however, a Polβ from L. infantum was shown to have a nuclear localization , which could indicate that the L. major polymerase is also nuclear, as their primary protein sequences showed 100% identity. The possibility that L. major possesses a nuclear Polβ, combined with the fact that this parasite does not have the PARP enzyme [7, 15], suggests that short-patch BER could play an important role in nuclear DNA repair for this organism. As Leishmania proliferates inside macrophage phagolysosomes, a well-coordinated nuclear short-patch BER is essential to combat oxidative DNA damage during parasite nuclear DNA replication . The Tritryps genomes apparently do not encode for the other X-family polymerases, DNA polymerase lambda (Polλ), and mu (Polµ)[7; 15]; thus, L. major may be the only Tritryps parasite that has an X-family polymerase in the nucleus, reinforcing the importance of short-patch BER in this organelle.
The mechanism of metformin effects on DNA mole- cules is unknown. Notwithstanding, these might possibly be mediated through its activation of AMPK, thereby in- creasing nitric oxide synthase (Zhou et al., 2001; Davis et al., 2006; Za’tara et al., 2008). Zou et al., (2004) specu- lated that mitochondria-derived reactive-nitrogen-species mediate AMPK activation by way of metformin. De- pending on the dose, nitric oxide is capable of inducing beneficial effects by playing a role in the gene regulation and signal transduction pathways possibly involved in de- fensive mechanisms against oxidative stress (Miyamoto et al., 2003). Nevertheless, high levels (nanomoles) can damage macromolecules, such as lipids, proteins and DNA, thereby leading to mutagenesis and carcinogenesis (Bishop and Cashman, 2003). Moreover, it has been shown that nitric oxide can induce mutation by impairing the repair-enzyme system (Jaiswal et al., 2001). Metformin is known to accumulate in tissues. Studies have indicated that doses insufficient for lowering hyperglycemia in diabetic animals concentrate in tissues (Wilcock and Bailey, 1994). Hirsch et al., (2009) showed that low doses of metformin inhibits cellular transforma- tion and selectively kills cancer-stem cells in four geneti- cally different types of breast cancer. The authors propose combining metformin with chemotherapy as a novel treat- ment, not only for breast cancer, but possibly others. Fur- thermore, in vitro lymphocytes challenged with CumOOH showed that high concentrations of metformin potentially induce DNA damage (Onaran et al., 2006). Taken to- gether, the data suggest that chronic metformin exposure may be potentially genotoxic. Thus, pregnant woman un- der treatment with metformin should be properly evalu- ated as to vulnerability to DNA damage.
At the Foundrys of Drawski M łyn and “WSK–Rzeszów” Metallurgical Plant in Rzeszów, a special technique of the nodularising (or vermicularising ) treatment was implemented. It was based on the use of cored wires, one cored with magnesium, and another with inoculant.
We were particularly interested in G. biloba tree species because standard Ginkgo biloba L. leaf extract (EGb 761) has been one of the best-selling medicinal products worldwide due to its antioxidant activity. Memory improvement, decrease of cerebral insufficiency, increase of cerebral blood flow and circulation, and beneficial effects in patients of Alzheimer's disease are some of the effects of GBE, which can be explained by its antioxidant activity [39, 41, 42, 43, 44, 64]. In addition, in vivo andin vitro experiments have demonstrated the efficacy of EGb 761 in protecting against age- related processes such as increase ofoxidative stress, brain mitochondrial dysfunction  and chronic age-dependent neurological disorders [66, 67]. Moreover, its antioxidant and antigenotoxic effects in S. cerevisiae cells were previously investigated and confirmed in our laboratory . The analyses of DNA (microarrays) made possible the discrimination of genes regulated by G. biloba L. leaf extract [68, 69], however, the mechanisms of its action are still unclear. Recently, a set of genes, modulated by EGb761 extract have been identified , some of them are involved cell cycle regulation. Thus to investigate such activity, the budding index approach used to identify yeast cells with phases of the cell cycle was applied with an extract from the leaves of G. biloba tree species. Results suggest that GBE protects cells from H 2 O 2 ,
In the present study, patients treated with second-generation TKIs had signiicantly higher levels of nitrite and MDA when compared to patients treated with imatinib. Such results suggest that oxidative stress parameters can be used to predict lack of response to 1st generation TKIs. Patients who are refractory to imatinib had higher levels ofoxidativedamage markers, possibly due to a high rate of additional mutations, the leading causes of resistance to irst-line therapy. Studies show that the occurrence of mutations is closely associated with oxidative stress and disease progression (39) . Therefore, it is likely that reactive oxygen species
with the TIP regimen (paclitaxel 175 mg/m 2 on day 1 + ifosfamide 2500 mg/m 2 on days 1 and 2 + cisplatin 50 mg/m 2 on day 2 every 21 days for three or four cycles) followed by radical sur- gery. Patients were considered eligible if they completed the planned treatment, data on clinical features and treatment outcomes were available, and the amount of biological materials in their biopsies was sufficient for molecular analyses. pCR was defined as no residual disease in surgi- cal samples. The immunohistochemical assessment of pWee1, γ-H2AX, and pChk1 was per- formed in formalin-fixed paraffin-embedded (FFPE) tissues, obtained from the biological specimens collected through bioptic procedures in untreated patients, using the following anti- bodies: anti-phospho-H2AX (Ser139) (clone JBW301) mouse monoclonal antibody (MAb) (Upstate, NY, USA) at the dilution of 1:500, anti-phospho-Wee1 (Ser642) (clone D47G5) rab- bit MAb (Cell Signaling, Danvers, MA, USA) at the dilution of 1:100, and anti-phospho-Chk1 (Ser345) (clone 133D3) rabbit MAb (Cell Signaling, Danvers, MA, USA) at the dilution of 1:100. Immunohistochemical staining was performed in an automated autostainer (BOND-III, Leica, Milan, Italy) by a biotin-free polymeric horseradish peroxidase (HRP)-linker antibody conjugate system (Leica, Milan, Italy). For each tumor, three different, 3 μm paraffin sections were analyzed and examined by light microscopy. Immunoreaction of tumor cells was counted in four high-power fields (400x magnification) per section. pWee1 and pChk1 were considered positive when 10% of the neoplastic cells showed a distinct nuclear immunoreactivity. pWee1 and pChk1 were graded on a four-grade scale based on staining intensity (0: negative, 1+: weak, 2+: moderate, 3+: strong). Tumors were classified as negative (0 = pWee1 neg and pChk1 neg ) or positive (1–3 = pWee1 pos and pChk1 pos ).The Allred scores were obtained as pre- viously described , considering staining intensity and percentage
Saturation dives involve long-duration (several days to weeks) stays in a closed environment with an increased partial pressure of oxygen. Saturation diving and exposure to oxygen-rich environ- ments in general lead to oxidativedamage [1–3]. Increased oxygen exposure in 10- to 30-d saturation dives can also lead to increases in body iron stores [1,3–6], and evidence exists that the status of vitamins—in particular, B vitamins involved in 1-carbon metabolism (for example, folate and vitamins B6 and B12)—could be affected by the oxidative environment . In support of this concept, homocysteine was elevated during 10- to 12-d saturation dives . Circulating homocysteine, a neurotoxin , increases in situations where folate, vitamin B12, or vitamin B6 status is decreased. An abundance of literature supports the notion that excess iron can also be toxic, through the formation of oxygen free radicals or increased availability of iron to pathogens or cancer cells, both of which are usually limited by iron availability [8–10]. Over the past 10 years, NASA has conducted NASA Extreme Environment Mission Operations (NEEMO) [3,5] missions as an analog to simulate several aspects of spaceflight. In these 7-to 14-d missions, divers live in a habitat 19 m below the ocean surface in an atmosphere with an ambient pressure of 2.5 atm (253 kPa) and 21% oxygen. This hyperbaric, and thus oxygen-rich, environment associated with the NEEMO missions is ideal to identify and
Waterbodies are a major sink for industrial, domestic and other anthropogenic compounds (Canli, Ay, Kalay, 1998). The aquatic pollution has far reaching impacts on organisms in the recipient environment. Fish as inhabitant of aquatic system cannot avoid the inimical efects of the pollutants. A set of biomarkers is generally used to evaluate the biological effects of pollutants. Such biomarkers act as an early warning of a specific detrimental biological endpoint. Oxidative stress and histopathologic biomarkers are used in ecotoxicology (Pandey et al ., 2003). Histopathology of ish tissues is a reliable monitoring tool, which allows the assessment of the environmental stressor’s efects. It is one of the most reliable indicator of the health impairment induced by the anthropogenic stressors in aquatic organism (Fernandes et al., 2008; Leonardi, Tarifeno, Vera, 2009). Enzymatic
In brief, 100 patients from the register of the Sevillian Fibromyalgia Association (AFIBROSE) and 45 healthy matched controls were enrolled into our study. Informed consent and the approval of the local ethical committee were obtained. The inclusion criterion during this study was: Patients diagnosed of FM in the last 2–3 years, based on the current ACR diagnostic criteria . Exclusion criteria were acute infectious diseases in the previous 3 weeks; past or present neurological, psychiatric, metabolic, autoimmune, allergy-related, dermal or chronic inflammatory disease; undesired habits (e.g., smoking, alcohol, etc.); medical conditions that required glucocorticoid treatment, use of analgesics, antidepressants drugs; past or current substance abuse or dependence; and pregnancy or current breastfeeding. Sixty-five potential participants met the inclusion criteria and were enrolled into the study (males/5, females/60), and 35 patients were excluded: 15 were smoker, 13 were using antidepressant treatment, 5 had rheumatoid arthritis, and 2 had hepatitis c. Forty-five healthy volunteers (males/5, females/40) were included in the study matching the age range, gender, ethnicity, and demographics (completion of at least 9 years of education and part of the middle socioeconomic class) of the recruited female FM subjects. Healthy controls had no signs or symptoms of FM and were free of any medication for at least 3 weeks before the study began. All patients and controls had not taken any drug or vitamin/nutritional supplement during a 3 weeks period before the collection of the blood samples. All patients and controls reported followed a standard balanced diet (carbohydrate 50–60%, protein 10–20% and fat 20–30%) that was established by a diet program during 3 weeks before blood collection. The diagnosis of FM was established by an experienced rheumatologist according to ACR criteria . Clinical data were obtained from physical examina-
Another mechanism involved in the activation of the autophagy is increased endoplasmic reticulum stress, which has been associated with pathological conditions of excessive nutrients, such as obesity. In vivo andin vitro studies indicate that excess energy and nutrients induces endoplasmic reticulum stress in adipose tissue cells, which is sufficient stimulus to promote autophagy (10, 40). In the MC3T3-E1 pre-osteoblasts, the increase in autophagy, either by endoplasmic reticulum stress or by the action of drugs, have paradoxical effects that may negatively influence cell proliferation or even to induce a protective effect against apoptosis (12, 41). When cells were treated with excess of Leu, we were not able to induce autophagy, although it was possible to identify the antiproliferative effect, since a lower number of cells per field were visible. The images of the cells stained with acridine orange obtained by fluorescence microscopy showed that, under normal intracellular conditions, it emits green fluorescence, but starts emitting red fluorescence during the autophagic process where the dye binds to organelles acidic vacuolar as lysosomes and autolissomes. While it is possible to identify small red dots on the images of cells treated with Leu, we could not find significant differences in the quantification of the percentage of autophagic cells, evaluated by flow cytometry. This result indicates that the decrease in cell proliferation caused by supplementation with Leu in MC3T3-E1 cells is not caused by increased autophagy.
Finally, despite the acute and chronic benefits from phytotherapic medicines associated with physical exercises presented here, they should be prescribed by qua- lified professionals. Such benefits can be superseded by the adverse reactions of self- -ingestion without proper guidance, which may even cause intoxications. The idea is that physical exercise programs aimed at sportsmen could count on the participation of nutritionists with deep knowledge on the application of these phytotherapic me- dicines. Likewise, undergraduate and graduate courses in nutrition should include Exercise Physiology discipline for a deeper understanding of the effects of physical exercise on the physiology of different systems. This symbiosis, if well planned and executed, can benefit everyone involved with the practice of physical exercises.
seeds weight, samples were taken randomly from harvested seeds of each subplot, the samples were weighed and the mean weight was obtained. On the other hand, weed population and percent ground cover were measured four weeks after emergence using 0.1 m 2 quadrant at six random positions per plot, two days before the scheduled weeding. 2.5 Performance of Chisel Plow and Disc Harrow Technical performance was conducted to compare chisel plow, the newly introduced implement in the irrigated schemes, and the disc harrow the widely used in the Gezira scheme, the comparison included measurement of draft, travel reduction (slippage), drawbar power determination, measurements of actual field capacities and efficiencies, and measurement of fuel consumption rates. For draft measurement, two tractors with the same horse-power were used as a test and auxiliary to estimate draft requirement for the chisel and the tandem disc harrow using hydraulic dynamometer at the assigned depths (10 cm and 20 cm), the measurements were performed according to Bukhari et al.
Figure 2. The effect of transcription inhibition on UV sensitivity is decreased after serum starvation. Clonogenic survival assay in AA8 (A), UV5 (B) and irs1SF (C) cell lines after synchronization by serum starvation. The cells were serum starved for 48 h and then treated with increasing doses of UV, incubated in the presence or absence of DRB for 24 h and cultivated in fresh medium. In the case of the UV5 cell line (B), the doses of UV were considerably lower, since the cell line is highly sensitive to UV-irradiation. The graphs depict the mean and standard deviation of at least three independent experiments. Flow cytometry plots of unsynchronized (upper row, AA8 (D), UV5 (E) and irs1SF (F)) and synchronized (lower row, AA8 (G), UV5 (H) and irs1SF (I)) cells, showing reduction in S-phase population after serum starvation. BrdU incorporation was used as a marker of ongoing replication. S-phase cut-off is shown as a percentage of total number of cells.
In the present study, we analyzed the damage profile andrepairof DNA lesions induced by the photodynamic action of phy- cocyanin in vitro andin a cellular system. The damage profile of DNA exposed to PHY plus VL was determined using a set of DNA repair endonucleases purified from E. coli. The DNA damage profile of PHY-induced lesions was different from that of MB-in- duced lesions. While MB plus VL induces a large excess of Fpg-sensitive sites, PHY pho- tosensitization mostly induced single strand breaks (Figure 1). The major difference be- tween these two profiles is in the number of Fpg-sensitive sites generated by the treat- ments. The excess of Fpg-sensitive sites gen- erated by MB photosensitization can be ex- plained by the production of 8-oxodGuo, which was not detected after PHY plus VL treatment. In fact, the profile of DNA dam- age induced by phycocyanin was quite simi- lar to that induced by ionizing radiation or by hydrogen peroxide in the presence of transi- tion metals (3,35), but was different from that induced by molecules which involve solely singlet oxygen as the major species responsible for the DNA modifications, such as heat decomposition of 3,3'-(1,4 naphthyl- idene)-dipropionate endoperoxide, NDPO 2
device was studied. The study took into account the effects of machine scale, wear surface structure of the rolls, grinding pressures and rolls speed, gap settings, feed size distribution and moisture content for a range of ores. The authors proposed a prevailing wear mechanism and a methodology for minimising wear of the grinding rolls, specific to the high pressure grinding device only. An example of a direct method, Bond (1964) and Buchi (1995) developed testing apparatus that determine rock abrasiveness in a low abrasion/medium impact mode of wear where rock abrasiveness is measured as the amount of material lost by a standard steel paddle which rotates on a shaft in a sample of loose rock particles of a certain specified size range. As can be seen from the above examples, the indirect methods of rock abrasivity assessment have the advantage of using data which is either readily available or relatively straightforward to obtain. However, they do not take into account process variables for specific modes of wear. Hence, they are normally not used in isolation, but rather in combination with direct methods, or holistic approaches, to supplement or confirm other more relevant direct measures. However, there is no universally accepted one standard test to determine the rock abrasivity although a large number of different tests are in use. All the studies about rock abrasiveness are concentrated on the amount of quartz, grain size and cementation degree of quartz, the geometry of the abrasive mineral and mechanical strength of rock.
The centromere is the chromosomal locus that directs kinetochore formation in order to secure faithful segregation of sister chromatids during mitosis. Nucleosomes that contain CENP-A (centromere protein A), an H3-histone variant, are thought to be the epigenetic mark indicating active centromeres, and its assembly into chromatin requires the Mis18 complex and its dedicated chaperone HJURP. This process is restricted in the cell cycle to telophase/early G1 phase and is a consequence of low Cdk1 and Cdk2 activity. Outside G1 phase, these kinases inhibit CENP-A assembly by acting on assembly factors Mis18 complex and HJURP. This model is derived from the observation that inhibition of Cdk activity can induce assembly before mitosis in G2 phase. However, induction of assembly following Cdk inhibition is not observed in S phase. In this study, I sought to investigate what are the mechanisms of inhibition in this cell cycle stage. I pursued several hypotheses, namely whether protein levels of HJURP and Mis18BP1, a subunit of Mis18 complex, were here diminished, which was not observed. Furthermore, I investigated a possible role of DNA damagein the prevention of CENP-A assembly. DNA replication occurs in S phase, which constitutes an important source of DNA damage. In addition, some studies postulated the participation of CENP-A and HJURP in the DNA damage response, which supported the hypothesis that DNA damage could constitute an inhibitory mechanism. This was tested in two different ways. First, I confirmed a negative correlation between the presence of DNA damageand CENP-A assembly, but inhibition of upstream kinases involved in DNA damage signaling did not lead to CENP-A assembly in S phase. Second, induction of DNA damage by two different means was not inhibitory in G1 and G2 phase. Unexpectedly, instead of blocking assembly, DNA damage induction was sufficient to induce CENP-A assembly in G2 phase. A possible explanation may be the alleviation of Cdk inhibition by activation of G2/M checkpoint. Together, these results suggest that the refractory nature of S phase is not a consequence of lack of key components, nor is it due to the existence of DNA damage. I will discuss further hypothesis and future experiments that may shed light on this phenomenon.
A cross-sectional study was conducted. First, 74 patients were recruited from the Endocrine Clinic of the Hospital Clementino Fraga Filho (HUCFF, Rio de Janeiro, Brazil) that consisted of 6 euthyroid control individuals and 68 patients with exogenous SCH due to TSH suppressive therapy with LT4 after total thyroidectomy for DTC. All patients were 18 years of age or older and exhibited a stable health profile over the last year of follow-up exams. After a clinical examination, 22 subjects were excluded due to characteristics that could affect the oxidative stress measurements including: detectable thyroglobulin, positive whole body scan (PCI), C-reactive protein greater than 3.0 ng/L, possible current malignancy or serious illnesses, smokers, alcoholics and those on medications that could influence the oxidative stress.
Part of the liver was removed by modified freeze clamping and placed in liquid nitrogen for ATP and adenosine diphosphate (ADP) measurements and part was placed in isolation buffer. This was done before exsanguination to ensure mitochondrial viability and minimize ATP degradation. Mitochondria were isolated by differential centrifugation steps and respiration was measured polarographically at 37uC using a respiratory system (System S 200A, Strathkelvin Instruments, Glasgow, Scotland). ATP and ADP levels were analysed by HPLC method . The expression of VDAC and its regulators, including a–tubulin and phosphor PKC-e were measured at protein level in the mitochondrial fraction by western blot. Expression of VDAC and a-tubulin was normalized to expression of prohibitin. Expression of phosphor- ylated PKCe is shown as a ratio to total PKCe. Expression of PGC1-a, and mtDNA (COX1, NADH1 and NADH6) were measured in liver and lung tissue as well as in plasma and BALF. See methods S1 for more detailed description.