DNA fragmentation factors 40 and 45 (DFF40/DFF45) and B-cell lymphoma 2 (Bcl-2) protein are underexpressed in uterine leiomyosarcomas and may predict survival Tomasz Banas,1 Kazimierz Pitynski,1 Krzysztof Okon,2 Aleksandra Czerw3,4 1Department of Gynecology and Oncology, 2Department of Pathomorphology, Jagiellonian University Medical College, Krakow, 3Department of Public Health, Faculty of Health Science, Medical University of Warsaw, 4Department of Health Promotion and Postgraduate Education, National Institute of Public Health – National Institute of Hygiene, Warsaw, Poland Objectives: DNA fragmentation factors 40 and 45 (DFF40 and DFF45) are responsible for final DNA-laddering during apoptosis, whereas Bcl-2 (B-cell lymphoma 2) is an apoptosis inhibitor. Our aim was to investigate the expression of DFF40, DFF45, and Bcl-2 in uterine leiomyosarcomas (uLMS), leiomyomas (uLM), and the normal myometrium. Furthermore, the correlation between DFF40, DFF45, and Bcl-2 expression and
Sperm DNA fragmentation (sDF) has been proved to be an important parameter in order to predict in vitro the potential fertility of a semen sample. Colloid centrifugation could be a suitable technique to select those donkey sperm more resistant to DNA fragmentation after thawing. Previous studies have shown that to elucidate the latent damage of the DNA molecule, sDF should be assessed dynamically, where the rate of fragmentation between treatments indicates how resistant the DNA is to iatrogenic damage. The rate of fragmentation is calculated using the slope of a linear regression equation. However, it has not been studied if sDF dynamics fit this model. The objectives of this study were to evaluate the effect of different after-thawing centrifugation protocols on sperm DNA fragmentation and elucidate the most accurate mathematical model (linear regression, exponential or polynomial) for DNA fragmentation over time in frozen-thawed donkey semen. After submitting post-thaw semen samples to no
CIP-induced chromosomal DNA fragmentation was assayed in situ in E. coli using the Micro-Halomax® kit [15]. We grew the samples in LB agar because this is simpler and is used routinely in clinical microbiology laboratories. The sample is scratched, diluted in LB broth to an OD600 of 0.05, and incubated with CIP in 4 ml of liquid LB in a 15 ml Falcon tube at 37°C with aeration. Incubation in a 1.5 ml Eppendorf tube with 24 μl of LB broth at room temperature (22°C) and without aeration does not modify the kinetics of DNA fragmentation induced by 1 μg/ml of CIP. We observed similar results in the TG1 strain and in three other E. coli-sensitive samples. Further confirmation in other sensitive strains could simplify the protocol for assessing E. coli sensitivity or resistance to CIP in the clinic.. Incubating TG1 with CIP for 40 min before technical processing produced a clear dose-response effect in chromosomal DNA fragmentation, and the damage level was similar in the different nucleoids. The ...
Paddenberg, P., Wulf, S., Weber, A., Heimann, P., Beck, L. A., & Mannherz, H. G. (1996). Internucleosomal DNA fragmentation in cultured cells under conditions reported to induce apoptosis may be caused by mycoplasma endonucleases. Eur J Cell Biol, 71(1), 105-119 ...
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To evaluate the apoptotic propensity to high and low cell densities, C2C12 myoblasts were seeded in six-well plates at either 2.0×103/cm2 to obtain a low density (~20-30% confluent) or 2.1×104/cm2 to reach a high cell density (~100% confluent) within 48 hours. The phase-contrast images in the top panel of Fig. 1A show typical morphology of C2C12 myoblasts at low or high cell densities. The protein abundance of cleaved caspase-3 and Poly (ADP-ribose) polymerase (PARP) was markedly increased in fully confluent cells when compared with cells that were plated at a low density (Fig. 1, middle panel). The cleavage of PARP by caspases leads to nuclear DNA fragmentation. Furthermore, a cell-death ELISA assay showed an elevation of cytosolic nucleosomes at full cell confluence (Fig. 1A, bottom panel). This provided additional evidence for an increase in apoptotic DNA fragmentation in confluent cells compared with non-confluent cells. We next explored the expression pattern and level of M-cadherin at ...
Apoptosis Protocol - posted in Apoptosis, Necrosis and Autophagy: Introduction to apoptosishttp://www.abcam.com...g...e&rid=11367Annexin V detection protocolhttp://www.abcam.com...g...e&rid=11369Apoptosis induction protocolhttp://www.abcam.com...g...e&rid=11368Caspase detection protocolhttp://www.abcam.com...g...e&rid=11370DNA fragmentation analysis protocolhttp://www.abcam.com...g...e&rid=11371
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In the present study, the major findings were that 1) KR-31378 effectively protected HUVECs from LPS-induced cell death accompanied by oligonucleosomal DNA fragmentation; 2) KR-31378 and KR-31612 significantly reduced the increased production of intracellular ROS and TNF-α that are stimulated by LPS, and KR-31378 more effectively scavenged peroxyl radical than α-tocopherol; and 3) incubation with LPS markedly decreased the Bcl-2 protein and increased the Bax protein in association with increase in cytochrome c release, which was significantly reversed by KR-31378 and to a lesser degree by KR-31612 and by α-tocopherol.. LPS, a bacterial endotoxin, is a proinflammatory mediator that induces the production of significant amounts of endogenous TNF-α, and both may mediate endothelial cell damage (Egido et al., 1993). The importance of TNF-α and ROS endogenously released after exposure to LPS has been associated with the genesis of apoptosis and cell death (Böhler et al., 2000). TNF-α also may ...
Question - Is it necessary to undergo DNA fragmentation and histoscopy after a failed IUI?. Ask a Doctor about In vitro fertilization, Ask an Infertility Specialist
A comparison of distinct modes of tumor cell death in Hodgkins disease using morphology and in situ DNA fragmentation.: The study examined the morphology and f
Submission: There are two things of interest in this paper. First is confirmation that even in women with normal ovarian reserve, elevated sperm DNA fragmentation (SDF) is related to increased rates of miscarriage. It seems that clinics who do NOT carry out an SDF test and treatment results in miscarriage may be exposing themselves to complaint. Unfortunately, in this study, IVF and ICSI produced the same outcomes. The authors identified a SDF level of 27% as the critical cutoff point. However the main feature of the article was that the SDF level significantly influenced the rate that live birth rates were diminished in women with decreased ovarian reserve. That is, when addressing the problem of age related success rates, the degree of male related sperm damage needs to be also considered. The authors suggested this may be interpreted as an indication that poor oocyte quality may be unable to correct the sperm DNA damage. While the concept of oocyte restoration of sperm quality as been around ...
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The instrument is particularly useful for structural characterization of species due to its ability to perform multiple stage (MSn) ion fragmentation analyses (up to 10 stages), allowing ion-mapping experiments to be conducted. It is also useful for analysis of high mass species that form multiply charged ions with ESI (such as proteins, peptides) due to the post-acquisition de-convolution capability of the operating software.. Applications include:. Full-scan MS/MS analysis, analysis of low-abundance compounds in matrices, consecutive reaction monitoring.. ...
Hi ladies! I know that I dont post in here much (sorry!) but I have just gotten some more bad news and was looking for help. My DHs Sperm DNA Fragmentation has come back quite high at 30%, and I have been googling a lot and found - page 59
What is DNA Fragmentation? I recently went to a medical conference where one of the brightest urologists dedicated to Assisted Reproduction, Craig Niederberger
Programmed cell death or apoptosis is a widespread biological phenomenon. Apoptosis is characterized by typical cell features such as membrane blebbing, chromatin condensation, and DNA fragmentation....
Fragmentation and growth-fragmentation equations is a family of problems with varied and wide applications. This paper is devoted to description of the long time time asymptotics of two critical cases of these equations, when the division rate is constant and the growth rate is linear or zero. The study of these cases may be reduced to the study of the following fragmentation equation: $$\frac{\partial}{\partial t} u(t,x) + u(t,x)=\int\limits_x^\infty k_0(\frac{x}{y}) u(t,y) dy.$$ Using the Mellin transform of the equation, we determine the long time behavior of the solutions. Our results show in particular the strong dependence of this asymptotic behavior with respect to the initial data.
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A ladder. The red arrow is the weight of the person climbing the ladder. Obviously, that person can be anywhere between the bottom rung and the top of the ladder. The top of the ladder is treated as a roller. It can only push perpendicular to the ladder ...
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The SCSA®is one of the most widely utilized tests of sperm DNA damage. There are now a number of commercial kits available for testing of sperm DNA fragmentation in which great variations of...
METHODS: The cytotoxic effect of deoxyelephantopin on A549 cells and normal lymphocytes was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and 50% inhibitory concentration (IC50) value was determined. The self-renewal and proliferating potential of A549 cells after treatment with deoxyelephantopin were examined by colony formation assay. Cellular morphology of deoxyelephantopin-treated cells was observed using phase-contrast microscopy. The induction of apoptosis was evaluated using acridine orange and ethidium bromide staining, Hoechst 33342 staining, terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end-labeling (TUNEL) assay, DNA fragmentation analysis and Annexin V-fluorescein isothiocyanate staining by flow cytometry. Activation of caspases was detected using fluorogenic substrate specific to caspases 2, 3, 8 and 9 and flow cytometric analysis. The total cellular DNA content and expression of cleaved poly (ADP-ribose) polymerase was ...
Our team in Birmingham has shown that sperm DNA damage more than doubles the risk of miscarriage.. This is a crucial finding; until now, miscarriage has generally been considered an exclusively female problem, with investigations and management targeting only women.. Yet the role of sperm DNA damage in miscarriage is not surprising. This is because while most cell types are able to repair damaged DNA, sperm lose this ability during development and have to rely on repair mechanisms in the egg. As the level of damage in the sperm DNA increases it also becomes increasingly likely that any repairs by the egg may create genetic mutations that could increase the risk of miscarriage.. Most existing tests for sperm DNA damage are insufficiently sensitive to be clinically useful; we are developing a more accurate combined assay system and therapies to achieve repair. One potential cause of sperm DNA damage is exposure to Reactive Oxygen Species (ROS) during production and transit. We are investigating ...
Random, unbiased fragmentation of DNA is necessary for next-generation sequencing (NGS) and chromatin immunoprecipitation (ChIP). Since DNA fragmentation can be a very problematic step for both NGS and ChIP, any technology that increased the efficiency and consistency of this step will be highly desirable for both research laboratories and in clinical diagnostics. Also, a technology that could make this step easier with no new equipment and very little cost to the laboratory would be ideal. We propose to apply the use of lipid encapsulated microbubbles to the fragmentation of DNA from both purified genomic and formaldehyde crosslinked samples. We have recently explored the feasibility of this technology, and our results were impressive. Preliminary data indicate that microbubbles can greatly improve the consistency of acoustic DNA fragmentation. Additionally, these bubbles are added in microliter volumes to the DNA or cell suspension at a cost ranging from one to ten cents per well, and can be ...
Exposure of fetal mouse brain cocultures to cocaine results selectively in the loss of neurites followed by neuronal death. By using enriched neuronal cultures, we here demonstrate that disappearance of neurons, when cultured with cocaine, is caused by apoptosis, based on (1) characteristic morphology of apoptotic nuclei at the level of neurons but not of glial cells by optic microscopy, and on total cell pellets by electron microscopy; (2) fragmentation of total DNA with a typical "ladder" pattern on agarose gels; (3) extensive in situ DNA fragmentation labeling (TUNEL method); and (4) prevention of cell loss by cycloheximide. The major metabolites of cocaine have no detectable effects on neurons, indicating that apoptosis is due to cocaine itself. Inappropriate neuronal apoptosis in cocaine-exposed fetal brain could perturb the neurodevelopmental program and contribute to the quantitative neuronal defects that are too frequently reported in the offspring of cocaine-abusing pregnant women. ...
Metacaspases are novel cysteine proteases found in apicomplexan whose function is poorly understood. Our earlier studies on Plasmodium falciparum metacaspase-2 (PfMCA-2) revealed that the caspase inhibitor, Z-FA-FMK efficiently inhibited PfMCA-2 activity and, expression, and significantly blocked in vitro progression of the parasite developmental cycle via apoptosis-like parasite death. Building on these findings, we synthesized a set of novel inhibitors based on structural modification of Z-FA-FMK with the amides of piperic acid and investigated their effect on PfMCA-2. One of these analogues, SS-5, specifically inhibited the activity and expression of PfMCA-2. The activities of some other known malarial proteases (falcipains, plasmepsins, & vivapain), and human cathepsins-B, D and L, and caspase-3 and -7 were not inhibited by SS-5. SS-5 blocked the development of P. falciparumin vitro (IC50 1µM) and caused prominent morphological distortions. Incubation with SS-5 led to persistent parasite ...
Atshan M, et al. - Semen samples were collected from 31 infertile men with immotile short-tail sperm (ISTS) as case group and 31 normozoospermic men as a con...
The ApopLadder Ex Kit is a sensitive, specific and rapid method for isolation of fragmented DNA from apoptotic cells. The kit contains all reagents for selective extraction of fragmented DNA and the protocol minimizes contamination by intact chromatin.
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Hydrogen peroxide (H(2)O(2)) is the major reactive oxygen species (ROS) produced in sperm. High concentrations of H(2)O(2) in sperm induce nuclear DNA fragmentation and lipid peroxidation and result in cell death. The respiratory chain of the mitochondrion is one of the most productive ROS generating systems in sperm, and thus the destruction of ROS in mitochondria is critical for the cell. It was recently reported that H(2)O(2) generated by the respiratory chain of the mitochondrion can be efficiently destroyed by the cytochrome c-mediated electron-leak pathway where the electron of ferrocytochrome c migrates directly to H(2)O(2) instead of to cytochrome c oxidase. In our studies, we found that mouse testis-specific cytochrome c (T-Cc) can catalyze the reduction of H(2)O(2) three times faster than its counterpart in somatic cells (S-Cc) and that the T-Cc heme has the greater resistance to being degraded by H(2)O(2). Together, these findings strongly imply that T-Cc can protect sperm from the ...
Browse our 17 DNA fragmentation resource search results. Abcam provides a large range of protocols, pathways, webinars, guides, articles, videos and …
Apoptosis From Wikipedia, the free encyclopedia Apoptosis Apoptosis increasing from normal cells (top) to apoptotic ones (bottom). Apoptosis ( /ˌæpəˈtoʊsɪs/)[1][2] is the process of programmed cell death (PCD) that may occur in multicellular organisms.[3] Biochemical events lead to characteristic cell changes (morphology) and death. These changes includeblebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation. (See alsoapoptotic DNA fragmentation.) In contrast to necrosis, which is a form of…
In the present study we tested whether N-acetyl-L-cysteine (LNAC) affects apoptotic death of neuronal cells caused by trophic factor deprivation. LNAC, an antioxidant, elevates intracellular levels of glutathione. We used serum-deprived PC12 cells, neuronally differentiated PC12 cells deprived of serum and NGF, and NGF-deprived neonatal sympathetic neurons. In each case LNAC prevents apoptotic DNA fragmentation and maintains long-term survival in the absence of other trophic support. Unlike NGF, LNAC does not induce or maintain neurite outgrowth or somatic hypertrophy. To rule out actions of LNAC metabolic derivatives, we assessed N-acetyl-D-cysteine (DNAC). DNAC also prevents death of PC12 cells and sympathetic neurons. However, other antioxidants were ineffective in this regard. Since it has been hypothesized that trophic factors prevent neuronal death by either preventing or coordinating cell cycle progression, we tested whether LNAC or DNAC treatment can affect cell cycle. We found that both ...
The sperm DNA fragmentation involves damage in the genetic material of sperm. At Institut Marquès we analyze and treat all causes.
We studied MSI in 205 tumours from 152 patients with HNPCC. Of these, 37 patients fulfilled the Amsterdam criteria, 72 patients were familial and 43 were sporadic cases. We used the method of fragmentation analysis (ABI Prism 310 Genetic Analyzer) with fluorescent labelled primers; three mononucleotide (BAT-RII, BAT-25, BAT-26) and five dinucleotide (D2S123, D3S1029, D5S346, D17S250, D18S58) repeat loci were analysed. We detected 75 tumours with a high degree of MSI (MSI-H), 12 tumours with a low degree of MSI (MSI-L) and 118 tumours with stable microsatellites (MSS). We found a loss of heterozygozity (LOH) in 44 MSS tumours. In 30 patients with MSI-H tumours mutation in one of mismatch repair genes was detected ...
Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology. (BMS) ...
We supply a wide range of Invitrogen DNA ladders and markers for accurate size and mass estimation (quantitation) of DNA fragments. DNA ladders and markers are available for sizing double-stranded, single-stranded, or supercoiled DNA.
TUNEL is a common method for detecting DNA fragmentation that results from apoptotic signaling cascades. The assay relies on the presence of nicks in the DNA which can be identified by terminal deoxynucleotidyl transferase or TdT, an enzyme that will catalyze the addition of dUTPs that are secondarily labeled with a marker. It may also label cells that have suffered severe DNA damage. This examination is going to be performed the 3-5 days after the first clinical ...
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New, license-free DNA ladders will allow researchers to estimate the size of fragments of DNA for a fraction of the cost of currently available methods.
An analysis of radiotherapy (RT) services in 33 European countries showed wide disparities in access, multiple areas of fragmentation and unmet needs, according to a study published online Jan. 23 in The Lancet Oncology.
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Highlights: • Aging process increases ROS accumulation. • Aging process increases DNA damage levels. • Absence of SOD activity does not cause DNA damage in young cells. • Absence of SOD activity accelerate aging and increase oxidative DNA damages during the aging process. - Abstract: Superoxide dismutases (SOD) serve as an important antioxidant defense mechanism in aerobic organisms, and deletion of these genes shortens the replicative life span in the budding yeast Saccharomyces cerevisiae. Even though involvement of superoxide dismutase enzymes in ROS scavenging and the aging process has been studied extensively in different organisms, analyses of DNA damages has not been performed for replicatively old superoxide dismutase deficient cells. In this study, we investigated the roles of SOD1, SOD2 and CCS1 genes in preserving genomic integrity in replicatively old yeast cells using the single cell comet assay. We observed that extend of DNA damage was not significantly different among the ...
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Butyrate, an intestinal microbiota metabolite of dietary fiber, exhibits chemoprevention effects on colon cancer development. However, the mechanistic action of butyrate remains to be determined. We hypothesize that butyrate inhibits cancerous cell proliferation but to a lesser extent in noncancerous cells through regulating apoptosis and cellular-signaling pathways. We tested this hypothesis by exposing cancerous HCT116 or non-cancerous NCM460 colon cells to physiologically relevant doses of butyrate. Cellular responses to butyrate were characterized by Western analysis, fluorescent microscopy, acetylation, and DNA fragmentation analyses. Butyrate inhibited cell proliferation, and led to an induction of apoptosis, genomic DNA fragmentation in HCT116 cells, but to a lesser extent in NCM460 cells. Although butyrate increased H3 histone deacetylation and p21 tumor suppressor expression in both cell types, p21 protein level was greater with intense expression around the nuclei in HCT116 cells when compared
In this study, a compressional-puffing process (CPP) was used to pretreat Sargassum hemiphyllum (SH) and then fucoidan was extracted from SH by hot water. Three fucoidan extracts, namely SH1 (puffing at 0 kg/cm2); SH2 (puffing at 1.7 kg/cm2); and SH3 (puffing at 10.0 kg/cm2) were obtained, and their compositions and biological activities were evaluated. The results indicate that CPP increased the extraction yield, total sugar content, and molar ratios of sulfate/fucose of fucoidan and decreased molecular weight and impurities of fucoidan. The SH1-SH3 extracts exhibited characteristics of fucoidan as demonstrated by the analyses of composition, FTIR spectroscopy, NMR spectroscopy, and molecular weight. All SH1-SH3 extracts showed antioxidant activities. The SH1-SH3 extracts protected SH-SY5Y cells from 6-hydroxydopamine (6-OHDA)-induced apoptosis as illustrated by cell cycle distribution, cytochrome c release, activation of caspase-8, -9, and -3, and DNA fragmentation analyses. Additional experiments
Simultaneous deficiencies in two genes have been proposed to introduce lethality in biologic systems that would otherwise tolerate the loss of one of the genes (22). According to this theory, the inhibition of PARP is a potential therapeutic strategy for the treatment of types of cancer that have specific DNA repair defects, including those that arise in individuals who are carriers of mutations in BRCA1 or BRCA2 (23). Other important genes involved in HRR include MRE11, XRCC2 and PARP1. In the present study, it was hypothesized that other components of this pathway may predict an increase in PARP1 inhibitor sensitivity. XRCC2 has roles in the HRR pathway, which repairs chromosomal fragmentation, deletions and translocations. Numerous studies have reported an association between XRCC2 SNPs and the cancer incidence risk; the most common mutation studied being the XRCC2 rs3218536 SNP (10-17). The present study aimed to investigate the effect of this SNP on PARP1 inhibitor sensitivity in CRC cell ...