Composition of Stallion Seminal Plasma and Its Impact on Oxidative Stress Markers and Spermatozoa Quality

The composition of seminal plasma of individual sires varies and so does the fertilizing ability. Micro and macro elements along with seminal enzymes, hormones, proteins, and lipids contained in seminal plasma are essential for the proper physiological function of spermatozoa. However, elevated levels against the normal physiological values, especially in the case of trace metals, result in the production of reactive oxygen species. The deficiency of antioxidants in the seminal plasma that could scavenge free radicals causes an impairment of spermatozoa quality. Ejaculates were obtained from 19 stallions. The fresh semen was analyzed to evaluate qualitative parameters of spermatozoa in terms of the motility, viability, and integrity of DNA. Separated seminal plasma underwent the assessment of the chemical and biochemical composition and RedOx markers. Based on the obtained concentrations of individual chemical elements, the correlation analysis suggested a negative impact of Cu in seminal plasma on the SOD, GPx, and LPO. Contrary, positive correlation was detected between FRAP and motility features. While Cu negatively correlated with sperm motion parameters, the adverse effect on viability was suggested for Cd. Our data suggest that seminal plasma has a potential due to its availability to become the potential biomarker of the reproductive health of farm animals.

Evaluation of DNA integrity and mitochondrial membrane potential in catfish Pseudoplatystoma magdaleniatum, induced by exposure to different concentrations of ibuprofen

There are few studies to date that determine the effects of ibuprofen on mitochondrial membrane potential (ΔΨM) and DNA integrity in neotropical fish. The objective of this study is to determine if four months’ exposure to ibuprofen in different concentrations (25 and 50 µg/L) produces effects on ΔΨM and alters the integrity of DNA in striped catfish Pseudoplatystoma magdaleniatum. For this study, the fish were placed in tanks with water at constant concentrations of 0 (control), 25, and 50 µg/L of ibuprofen for four months. Subsequently, blood samples were taken for analysis of ΔΨM and DNA integrity, using a flow cytometer LSRFortessa BD Biosciences. After four months of exposure to ibuprofen at different concentrations, the results showed no increase in Low ΔΨM, indicating that there are no alterations in the mitochondrial membrane potential. On the other hand, the percentages of DNA damage were below 0.39, which indicates that there were no alterations in DNA integrity. It is possible that under the conditions in which this study was conducted (ibuprofen levels, exposure time), they are not sufficient to demonstrate the effects caused by this drug. Higher ibuprofen levels and/or longer exposures may be required to determine alteration in ΔΨM and DNA integrity. Flow cytometric analysis for these types of samples is a fast, specific, and reliable technique, compared to traditional methods.

Evaluation of mitochondrial membrane potential and DNA integrity in Catfish Pseudoplatystoma magdaleniatum, when exposed for prolonged times to different concentrations of ibuprofen.

There are few studies to date that determine the effects of ibuprofen on mitochondrial membrane potential (ΔΨM) and DNA integrity in neotropical fish. The objective of this study is to determine if four months’ exposure to ibuprofen in different concentrations (25 and 50 µg/L) produces effects on ΔΨM and alters the integrity of DNA in striped catfish Pseudoplatystoma magdaleniatum. For this study, the fish were placed in tanks with water at constant concentrations of 0 (control), 25, and 50 µg/L of ibuprofen for four months. Subsequently, blood samples were taken for analysis of ΔΨM and DNA integrity, using a flow cytometer LSRFortessa BD Biosciences. After four months of exposure to ibuprofen at different concentrations, the results showed no increase in Low ΔΨM, indicating that there are no alterations in the mitochondrial membrane potential. On the other hand, the percentages of DNA damage were below 0.39, which indicates that there were no alterations in DNA integrity. It is possible that under the conditions in which this study was conducted (ibuprofen levels, exposure time), they are not sufficient to demonstrate the effects caused by this drug. Higher ibuprofen levels and/or longer exposures may be required to determine alteration in ΔΨM and DNA integrity. Flow cytometric analysis for these types of samples is a fast, specific, and reliable technique, compared to traditional methods.

Base excision repair and its implications to cancer therapy

Base excision repair (BER) has evolved to preserve the integrity of DNA following cellular oxidative stress and in response to exogenous insults. The pathway is a coordinated, sequential process involving 30 proteins or more in which single strand breaks are generated as intermediates during the repair process. While deficiencies in BER activity can lead to high mutation rates and tumorigenesis, cancer cells often rely on increased BER activity to tolerate oxidative stress. Targeting BER has been an attractive strategy to overwhelm cancer cells with DNA damage, improve the efficacy of radiotherapy and/or chemotherapy, or form part of a lethal combination with a cancer specific mutation/loss of function. We provide an update on the progress of inhibitors to enzymes involved in BER, and some of the challenges faced with targeting the BER pathway.

The Integrity of DNA Extracted from Oryctolagus Cuniculus Carcasses Treated With Different Corrosive Chemical Substances

There is a gap in the literature concerning the use of corrosive chemicals to erode human body. The aim of this study is to determine the integrity of DNA samples extracted from rabbits’ carcasses immersed in different chemical. Twenty-one healthy rabbits were grouped into seven. Animals in group 1 were enclosed in transparent containers while groups 2 to 7 were submerged in 1% hydrochloric acid (HCl), sodium hypochlorite (NaClO), calcium hypochlorite (CaOCl2), 7.3% sodium hydroxide (NaOH), water and 35% HCl respectively. Tissue sample were collected on day(s) 1, 3, 7 and 12 for DNA extraction and analysis. The most destructive agent in this study was 35% HCl where the bones and almost all tissues degraded within 24hrs, followed by Sodium hydroxide while, there was no marked degradation in NaClO, CaOCl2, water and dilute HCl groups. Distinct, observable effects of each chemical on the extracted DNA samples revealed decreased DNA integrity and degradation at various time frames. Further studies need to be carried out to ascertain the concentration of these chemicals especially NaOH that will completely destroy the body of the rabbits within a short period of time. Chromosomal aberration analysis should also be included in future research.

Recent Synthetic Approaches towards Small Molecule Reactivators of p53

The tumor suppressor protein p53 is often called “the genome guardian” and controls the cell cycle and the integrity of DNA, as well as other important cellular functions. Its main function is to trigger the process of apoptosis in tumor cells, and approximately 50% of all cancers are related to the inactivation of the p53 protein through mutations in the TP53 gene. Due to the association of mutant p53 with cancer therapy resistance, different forms of restoration of p53 have been subject of intense research in recent years. In this sense, this review focus on the main currently adopted approaches for activation and reactivation of p53 tumor suppressor function, focusing on the synthetic approaches that are involved in the development and preparation of such small molecules.

Propylene glycol-based antifreeze as an effective preservative for DNA metabarcoding of benthic arthropods

Maintaining the integrity of DNA in bulk environmental samples from source to laboratory is crucial for capturing the true range of taxa present within an ecosystem. Preservation consideration of DNA is particularly important if samples are being collected in remote areas and by non-specialist ‘citizen scientists’ in nationwide programs. Traditionally, absolute ethanol is used as the preferred preservative for environmental samples collected for downstream DNA metabarcoding analyses. However, transport, shipping and DNA extraction of samples preserved in ethanol is a lengthy procedure due to safety restrictions and the requirement of full ethanol evaporation prior to extraction. We examined the efficacy of an easily accessible, non-toxic propylene glycol-based antifreeze as an alternative to absolute ethanol for preserving macroinvertebrate DNA from bulk-benthos DNA samples. We tested the differences in both cytochrome oxidase I (COI) exact sequence variants (ESVs) and COI taxonomic orders detected in both ethanol and antifreeze samples using two processing methods (no evaporation of preservative versus full evaporation). In addition, we assessed the detection of families and genera within the Arthropoda phylum for preservative type, site and processing method. Our results suggest that antifreeze is a suitable alternative to ethanol, a greater global ESV richness reported for antifreeze samples. Additionally, a higher proportion of arthropod reads in ESVs were detected in antifreeze (average 69%) compared with ethanol (average 53%). Finally, antifreeze samples produced similar results for the different processing methods, whereas ethanol samples failed to produce similar results without prior evaporation. Although ethanol is currently widely used for DNA preservation, our results demonstrate that by using antifreeze, it is possible to achieve similar taxonomic coverage and community assemblages of bulk-benthos DNA samples for macroinvertebrates, with the added simplicity and shorter laboratory processing time achieved using an easily available, unregulated preservative.

Formalin Fixation of Human Healthy Autopsied Tissues: The Influence of Type of Tissue, Temperature and Incubation Time on the Quality of Isolated DNA

Formalin fixation is a widely used method in histopathology that has certain limits. Formalin often leads to the degradation of DNA molecules in cancer tissues, which makes tissues unusable for molecular analysis. The other factors may also affect the quality of DNA isolated from fixed tissues. The aim of this study is to determine the impact of the incubation time and temperature on the quality of DNA molecules isolated from various healthy human tissues. The brain, lung and kidney tissues, excluded during the forensic autopsies of people who died of violent death, were fixed in phosphate-buffered formalin from 24h to two months. After the completion of the incubation period, the DNA was isolated using phenol-chloroform-isoamyl alcohol extraction method and the concentration and purity of the samples were determined spectrophotometrically. The degree of degradation of DNA was assessed by PCR reaction, by amplification of gene fragments which lengths were 150bp (GPD1) and 262bp (β-actin). The highest concentration, purity and preserved integrity of DNA were obtained from the brain samples. With prolonged tissue incubation times in formalin, the concentration and integrity of DNA decreased in all tissue samples, especially in the brain tissue, while the purity of DNA remained unchanged. Also, tissue fixation at +4°C contributed to a better quality of isolated DNA compared to DNA isolated from tissue fixed at room temperature. We can conclude that the type of human healthy tissue, temperature and the incubation time of formalin fixation have important influence on the concentration, purity and integrity of DNA during fixation of tissues excluded in the course of forensic autopsy.