Optimization of in vitro culture and transfection condition of bovine primary spermatogonial stem cells

The present study aimed to optimize the in vitro culture and transfection efficiency of bovine primary spermatogonial stem cells (SSCs). To this end, SSCs were obtained from newborn Holstein bull calves by two-step enzymatic digestion. After enrichment and culture, SSCs were characterized by using alkaline phosphatase (AP) staining and expression of vasa and thy1 genes as specific bovine SSC markers. To evaluate the effect of antioxidants on vitality, colony formation, and the expression of pro- and anti-apoptotic genes of bovine SSCs, various concentrations of vitamin C (5, 10, 25 and 50 μg/mL) and Trolox (a water soluble α-tocopherol analogue) (12.5, 25, 50 and 100 μg/mL) were added to the SSC culture medium. The results showed that SSCs treated with 50 μg/mL of vitamin C or 25 μg/mL of Trolox individually could increase cell viability and colony formation significantly in comparison with other concentrations and the control group. Additionally, the expressions of bax (as a pro-apoptotic gene) and bcl2 (as an anti-apoptotic gene) were significantly lower and higher than the control group, respectively. To optimize the transfection condition, the effective dosages of vitamin C or Trolox, with various concentrations of two transfection reagents (X-tremeGENE HP and Turbofect) and DNA, at day 8 of culture, were studied. Results showed that 1 μl X-tremeGENE HP or 0.5 μl Turbofect and 2 μg of DNA are the best concentrations for transfecting SSCs. However, X-tremeGENE HP expressed more potential for transfecting SSCs in comparison with Turbofect. Besides, no difference was observed between the use of defined doses of vitamin C or Trolox.Keywords: Apoptosis, gene transfer, primary cells, viability, vitamin C, Trolo

Acute Ultraviolet Light Exposure and Post-Resistance Exercise Serum Testosterone: A Pilot Study in Older Men

International Journal of Exercise Science 13(2): 607-614, 2020. Testosterone deficiency is linked to a multitude of conditions including depression, sexual dysfunction, and cognitive impairment. Although synthetic testosterone-boosting pharmaceuticals have gained wide use, recent data suggests that vitamin D, ingested orally, may positively impact serum testosterone. Furthermore, evidence tying ultraviolet (UV) light exposure to endogenous vitamin D synthesis presents an intriguing possibility for naturally enhancing serum testosterone. This investigation sought to determine the effect of acute UV light exposure on resistance exercise-induced acute testosteronemia and vitamin D in older men. Six older adult men of varying activity levels (age 62 ± 1.79 yrs., height 179.92 ± 1.12 cm., body mass 83.79 ± 3.12 kg., BMI 25.95 ± 1.15 kg/m2) participated in two testing sessions separated by one week: 1) Resistance exercise followed by standard passive recovery (RERC) and 2) RE plus UV light exposure during the first 10-minutes of RE passive recovery (RERC-UV). The RE protocol was identical in both sessions and consisted of four sets of 10RM on leg press, chest press, and back row with 1-minute of rest between sets followed by 30-minutes of post-RE passive recovery. Serum testosterone and vitamin D were measured pre- and post-RE in 5-minute increments during the 30-minute recovery. Analysis of variance revealed neither RE or RERC-UV significantly affected serum testosterone or vitamin D. These findings suggest that acute UV light exposure may be insufficient to positively impact serum testosterone and vitamin D following a single bout of RE in older adult men

Livestock slaughterhouses waste management in urban environment

The current study presents an investigation on the waste management in the cattle slaughterhouse by the following objectives: a) to identify the existing waste management practices in relation to sources, quantity and characteristic of wastes; b) to identify the situation of production, collection, storage, transportation, processing and recycling, and final disposal of wastes and the problems of existing waste management practices. In order to obtain reliable information and filling the check list, site surveys were conducted when the management of the slaughter-house was interviewed in waste management practices. The total produced industrial waste in studying units' was found to be 10252 tons/year. The per capita waste generations were reported to be 54.6 kg/cattle/day and 11.1 kg/sheep/day. Also, more than 98% of the hazardous waste produced can be infectious. However, it is important to keep in mind that setting an operational program and careful monitoring of its optimal execution by the slaughterhouse manager is necessary. Consequently, findings provide useful inputs for decision making processes around construction slaughterhouses waste management.  </strong

Wastewater Based Epidemiology Perspective as a Faster Protocol for Detecting Coronavirus RNA in Human Populations: A Review with Specific Reference to SARS-CoV-2 Virus

Abstract: Wastewater-based epidemiology (WBE) has a long history of identifying a variety of viruses from poliovirus to coronaviruses, including novel Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The presence and detection of SARS-CoV-2 in human feces and its passage into the water bodies are significant public health challenges. Hence, the hot issue of WBE of SARS-CoV-2 in the coronavirus respiratory disease (COVID-19) pandemic is a matter of utmost importance (e.g., SARS-CoV-1). The present review discusses the background, state of the art, actual status, and prospects of WBE, as well as the detection and quantification protocols of SARS-CoV-2 in wastewater. The SARS-CoV-2 detection studies have been performed in different water matrixes such as influent and effluent of wastewater treatment plants, suburban pumping stations, hospital wastewater, and sewer networks around the globe except for Antarctica. The findings revealed that all WBE studies were in accordance with clinical and epidemiological data, which correlates the presence of SARS-CoV-2 ribonucleic acid (RNA) with the number of new daily positive cases officially reported. This last was confirmed via Reverse Transcriptase-quantitative Polymerase Chain Reaction (RT-qPCR) testing which unfortunately is not suitable for real-time surveillance. In addition, WBE concept may act as a faster protocol to alert the public health authorities to take administrative orders (possible re-emerging infections) due to the impracticality of testing all citizens in a short time with limited diagnostic facilities. A comprehensive and integrated review covering all steps starting from sampling to molecular detection of SARS-CoV-2 in wastewater has been made to guide for the development well-defined and reliable protocols

Evaluating bovine sperm transfection using a high-performance polymer reagent and assessing the fertilizing capacity of transfected spermatozoa using an in vitro fertilization technique

Sperm-mediated gene transfer (SMGT) has been considered as an innovative device for transgenesis on a mass scale by taking advantage of live spermatozoa to transfer exogenous DNA. However, the fertilizing ability of transfected sperm cells and the poor reproducibility of this method are still matters of controversy. Hence, the current study was conducted to evaluate transfecting the enhanced green fluorescent protein (EGFP) as the source of exogenous DNA into bovine spermatozoa using a high-performance polymer reagent as well as assessing the fertilizing capacity of transfected sperm cells by in vitro fertilization (IVF). In the first experiment, three different concentrations of rhodamine-labeled DNA and high-performance polymer transfection reagent, X-tremeGENE HP, were used to transfect bovine spermatozoa. In the second experiment, IVF and fluorescence microscopy methods were utilized to assess the fertilizing capacity of sperm cells carrying exogenous DNA when X-tremeGENE HP was used either alone or with dimethyl sulfoxide (DMSO) treatment. Findings revealed that at 1&thinsp;µL X-tremeGENE HP and 1&thinsp;µg of DNA concentration, approximately one-third of total spermatozoa were transfected. However, following IVF and fluorescence microscopy, no EGFP expression was detected in zygotes and morula-stage embryos. Results of this study showed that, although X-tremeGENE HP could transfer EGFP to bovine spermatozoa, transfected sperm cells were unable to transfer foreign DNA to matured bovine oocytes. Under our experimental conditions, we hypothesized that the absence of the EGFP fluorescence signal in embryos could be due to the detrimental effects of transfection treatments on sperm cells' fertility performance as well as incompetency of IVF to produce transgenic embryos using transfected sperm cells.</p

The influence of the absolute surface area on the NO2 and H2 gas responses of ZnO nanorods prepared by hydrothermal growth

In this paper we report the influence of the absolute surface area (ASA) and aspect ratio (L/D) of hydrothermally grown ZnO nanorods (NRs) on the NO2 and H2 gases relative responses (RRs) in the 20 ppb − 1 ppm and 50 ppm–1000 ppm concentrations range. We used films of NRs characterized by different geometrical features, such as diameter, length, L/D and ASA, obtained using different growth times, but all showing a well-defined hexagonal shape and an alignment perpendicular to the substrate. Values of L/D up to 129 and NRs lengths up to 14 μm have been obtained; one of the highest reported for quasi-one-dimensional ZnO nanostructures obtained through solution-based one-pot syntheses. Electrical tests at 150 °C (NO2) and 200 °C (H2) carried out using films of NRs presenting different L/D evidenced a direct relationship between the increase of the ASA and L/D and gas RRs. The influence of geometrical and chemical characteristics, such as diameter, size and surface-defects concentration is also discussed in light of the existing literature. Cross sensitivity, investigated measuring the H2 (target) response in the presence of NO2 (interfering) gases as well as dynamic and cumulative NO2 gas reproducibility test are also presented. Beside the considerable interference played by NO2 to the H2 response, our long NRs demonstrate an excellent reproducibility of the electrical signal and fast recovery of the base line, evidencing the absence of irreversible chemisorption or sample-surface “history-related” phenomena