Effect of Uropathogenic <i>Escherichia coli</i> on Human Sperm Function and Male Fertility

Infections of the reproductive tract represent nearly 15% of male infertility cases. The most frequently isolated bacterium in the ejaculate of infertile men is Escherichia coli (E. coli), which causes between 60 and 85% of cases of chronic bacterial prostatitis leading to sperm damage. The aim of this chapter is to discuss the negative effects of E. coli on sperm quality and male fertility. The E. coli isolated from semen is uropathogenic (UPEC) and can damage sperm in different ways. UPEC induces activation of polymorphonuclear leukocytes with the release of cytokines and reactive oxygen species, the latter being harmful due to their ability to induce lipid peroxidation and early sperm capacitation. Also, UPEC decreases sperm motility, vitality and mitochondrial membrane potential through direct contact or mediated by its soluble metabolites. The negative effects are higher with strains with specific characteristics such as hemolytic capacity. In vivo studies with mice models have shown that UPEC inoculated into the epididymis induces inflammatory damage with testicular mass decrease and low sperm concentration. Future studies are needed to clarify the molecular mechanisms by which E. coli damages sperm. This knowledge will make it possible to take measures to avoid deleterious consequences on the fertilizing potential of men

Pharmacogenomics of vitamin K antagonists

There is an important interindividual variability in dose requirement for coumarinic anticoagulants, which could be explained by genetic and non-genetic factors. Among hereditary factors, there are gene polymorphisms that code the therapeutic target and the main enzyme responsible for their metabolism. However, there are other candidate genes that could modulate dose requirements. The is a paucity of pharmacogenomic platforms to determine dose requirements of coumarinics in the Chilean population. Therefore, algorithms considering different variables to adjust individual dosages are required. Herein, we analyze the available evidence about factors that can modify the effects of vitamin K antagonists and that should be incorporated to dosing algorithms

Effect of incubation temperature after devitrification on quality parameters in human sperm cells

Sperm cryopreservation is common in assisted reproduction laboratories, providing a therapeutic option for several clinical conditions. This process has been optimized; however, the effect of post-thaw incubation temperature has been poorly studied. This work analyzed the effect of incubation temperature after devitrification on human sperm quality. Spermatozoa from normozoospermic donors were cryopreserved by vitrification. After devitrification, the spermatozoa were separated into two aliquots: (i) incubated at room temperature (RT, 22-25 degrees C) and (ii) incubated at 37 degrees C. Reactive oxygen species (ROS), viability, mitochondrial membrane potential (AWM), phosphatidylserine externalization and motility were analyzed immediately after devitrification (control) and after 2, 4 and 6 h. Spermatozoa incubated at RT showed a conserved viability and Delta Psi M compared to the control, while the incubation at 37 degrees C promoted a decrease in these parameters. The ROS levels were increased at both incubation conditions. The progressive motility was decreased in all experimental groups and the decrease was more pronounced under incubation at RT. No increase in phosphatidylserine externalization was observed. In conclusion, prior to use in assisted reproduction procedures, devitrified spermatozoa at RT conserve a better viability and Delta Psi M than at 37 degrees C

Nitrosative stress in human spermatozoa causes cell death characterized by induction of mitochondrial permeability transition-driven necrosis

Peroxynitrite is a highly reactive nitrogen species and a potent inducer of apoptosis and necrosis in somatic cells. Peroxynitrite-induced nitrosative stress has emerged as a major cause of impaired sperm function; however, its ability to trigger cell death has not been described in human spermatozoa. The objective here was to characterize biochemical and morphological features of cell death induced by peroxynitrite-mediated nitrosative stress in human spermatozoa. For this, spermatozoa were incubated with and without (untreated control) 3-morpholinosydnonimine (SIN-1), in order to generate peroxynitrite. Sperm viability, mitochondrial permeability transition (MPT), externalization of phosphatidylserine, DNA oxidation and fragmentation, caspase activation, tyrosine nitration, and sperm ultrastructure were analyzed. The results showed that at 24 h of incubation with SIN-1, the sperm viability was significantly reduced compared to untreated control (P<0.001). Furthermore, the MPT was induced (P<0.01) and increment in DNA oxidation (P<0.01), DNA fragmentation (P<0.01), tyrosine nitration (P<0.0001) and ultrastructural damage were observed when compared to untreated control. Caspase activation was not evidenced, and although phosphatidylserine externalization increased compared to untreated control (P<0.001), this process was observed in <10% of the cells and the gradual loss of viability was not characterized by an important increase in this parameter. In conclusion, peroxynitrite-mediated nitrosative stress induces the regulated variant of cell death known as MPT-driven necrosis in human spermatozoa. This study provides a new insight into the pathophysiology of nitrosative stress in human spermatozoa and opens up a new focus for developing specific therapeutic strategies to better preserve sperm viability or to avoid cell death

Multiparameter Flow Cytometry Assay for Analysis of Nitrosative Stress Status in Human Spermatozoa

Male infertility is an increasing health problem, and oxidative/nitrosative stress plays an important role in the etiology of this condition. Nitrosative stress due to excessive levels of reactive nitrogen species (RNS) is associated with impaired male fertility. Flow cytometry may be a useful tool for semen evaluation, but the availability of multiparameter assays for analysis of sperm quality is limited. The present study standardized a multiparameter flow cytometry analysis for nitrosative stress status in human spermatozoa in a single assay. A suitable multicolor fluorochrome panel was designed and consisted of fluorescein-boronate to detect peroxynitrite, a highly RNS, propidium iodide to analyze viability, tetramethylrhodamine methyl ester perchlorate to detect mitochondrial membrane potential (MMP) and monobromobimane to analyze thiol oxidation. Proper positive and negative controls for each fluorochrome were used to establish the technique, and sperm cells of different qualities and spermatozoa subjected to cryopreservation were analyzed. The results showed that the controls clearly discriminated between the high and low fluorescence intensities for each fluorochrome. The analysis of sperm cells of different quality demonstrated that the assay properly detected differences in all parameters analyzed according to sperm quality. The results may be reported as the mean fluorescence intensity of each fluorochrome and the percentage of cells exhibiting different characteristics. In conclusion, a protocol was standardized to analyze nitrosative stress status, including peroxynitrite production, viability, MMP, and thiol oxidation, in a single analysis using flow cytometry. This protocol may be applied to research approaches and clinical andrology to improve the evaluation of sperm quality and provide a promising tool to increase the use of clinical flow cytometry. (c) 2020 International Society for Advancement of Cytometr

Obesity Is Associated with Changes in Laboratory Biomarkers in Chilean Patients Hospitalized with COVID-19

Background and aims: It is reported that patients with obesity are more frequently hospitalized for COVID-19, and evidence exists that obesity is a risk factor, regardless of other comorbidities. The objective of this study was to evaluate the association of obesity with changes in laboratory biomarkers in hospitalized Chilean patients. Materials and methods: A total of 202 hospitalized patients (71 with obesity and 131 without obesity) with a diagnosis of COVID-19 were included in the study. Demographic, clinical, and laboratory (days 1, 3, 7, 15) data were obtained. We performed a statistical analysis, assuming significance with a value of p &lt; 0.05. Results: Significant differences in chronic respiratory pathology are observed between patients with and without obesity. The inflammatory markers CPR, ferritin, NLR, and PLR are elevated during the evaluated period, while changes in leukocyte populations are present on day 1 (eosinophils) and day 3 (lymphocytes). Finally, a persistent elevation of D-dimer level is observed, presenting significant differences on day 7 between patients with and without obesity. Obesity had a positive correlation with admission to the critical patient unit, invasive mechanical ventilation, and length of hospital stay. Conclusion: Patients with obesity hospitalized for COVID-19 present marked elevations of inflammatory and hemostasis parameters, with a correlation between obesity, changes in laboratory biomarkers, and the risk of adverse clinical outcomes also observed

Dynamic changes of hematological and hemostatic parameters in COVID-19 hospitalized patients: Potential role as severity biomarkers for the Chilean population

Background: COVID-19 is still a global health issue, there is limited evidence in South America regarding laboratory biomarkers associated with severe disease. The objective of our study was to identify hematological and hemostatic changes associated with severe COVID-19. Methods: A total of 170 hospitalized patients with COVID19 were included in the study, defining their severity according to established criteria. Demographic, clinical, and laboratory (days 1, 3, 7, 15) data were obtained. We performed a statistical analysis, assuming significance with a value of p < 0.05. We analyzed the correlation between severity and biomarkers and established cut-off values for severe patients through ROC curves, estimating Odds Ratio associated with severe disease. Results: Day 1 was observed significant differences between moderate vs severe patients for leukocytes (WBC), Neutrophil-lymphocyte ratio (NLR), platelet-lymphocyte ratio (PLR) and D-dimer, establishing cut-off points for each of them. The markers we found associated to risk of severe disease were WBC (OR=3.2396; p = 0.0003), NLR (OR=5.7084; p < 0.0001), PLR (OR=4.4094; p < 0.0001), Neutrophil (OR=4.1193; p < 0.0001), D-dimer (OR=2.7827; p = 0.0124). Conclusions: The results allow to establish basic laboratory biomarkers associated to severe disease, which could be used as prognostic markers

Long-term survive of Aliarcobacterbutzleri in two models symbiotic interaction with Acanthamoebacastellanii

Aliarcobacter butzleri (formerly known as Arcobacter butzleri) is an emerging food-borne zoonotic pathogen that establishes in vitro endosymbiotic relationships with Acanthamoeba castellanii, a free-living amoeba. Previously, we described that this bacterium acts as an endocytobiont of A. castellanii, surviving for at least 10 days in absence of bacterial replication. Thus, the aim of this study was to evaluate the ability of A. butzleri to survive as a long-term endosymbiont of A. castellanii for 30 days in two models of symbiotic interaction with A. castellanii: (i) endosymbiotic culture followed by gentamicin protection assay and (ii) transwell co-culture assay. The results allow us to conclude that A. butzleri is capable of surviving as an endosymbiont of A. castellanii for at least 30 days, without multiplying, under controlled laboratory conditions. In addition, in the absence of nutrients and as both microorganisms remain in the same culture, separated by semi-permeable membranes, A. castellanii does not promote the survival of A. butzleri, nor does it multiply. Our findings suggest that the greater survival capacity of A. butzleri is associated with their endosymbiont status inside A. castellanii, pointing out the complexity of this type of symbiotic relationship