Exposure to toxicologically relevant atrazine concentrations impair the glycolytic function of mouse Sertoli cells through the downregulation of lactate dehydrogenase.

Atrazine (ATZ), a widely used herbicide with potent endocrine-disrupting properties, has been implicated in hormonal disturbances and fertility issues. Sertoli cells (SCs) play a crucial role in providing mechanical and nutritional support of spermatogenesis. Herein, we aimed to study the effects of environmentally relevant ATZ concentrations on the nutritional support of spermatogenesis provided by SCs. For that, mouse SCs (TM4) were exposed to increasing ATZ concentrations (in µg/L: 0.3, 3, 30, 300, or 3000). After 24 h, cellular proliferation and metabolic activity were assessed. Mitochondrial activity and endogenous reactive oxygen species (ROS) production were evaluated using JC-1 and CM-H2DCFDA probes, respectively. We also analyzed protein levels of lactate dehydrogenase (LDH) using Western Blot and live cells glycolytic function through Seahorse XF Glycolysis Stress Test Kit. ATZ exposure decreased the activity of oxidoreductases in SCs, suggesting a decreased metabolic activity. Although ATZ is reported to induce oxidative stress, we did not observe alterations in mitochondrial membrane potential and ROS production across all tested concentrations. When we evaluated the glycolytic function of SCs, we observed that ATZ significantly impaired glycolysis and the glycolytic capacity at all tested concentrations. These results were supported by the decreased expression of LDH in SCs. Overall, our findings suggest that ATZ impairs the glycolytic function of SCs through LDH downregulation. Since lactate is the preferential energetic substrate for germ cells, exposure to ATZ may detrimentally impact the nutritional support crucial for spermatogenesis, hinting for a relationship between ATZ exposure and male infertility.

Signatures of metabolic diseases on spermatogenesis and testicular metabolism.

Diets leading to caloric overload are linked to metabolic disorders and reproductive function impairment. Metabolic and hormonal abnormalities stand out as defining features of metabolic disorders, and substantially affect the functionality of the testis. Metabolic disorders induce testicular metabolic dysfunction, chronic inflammation and oxidative stress. The disruption of gastrointestinal, pancreatic, adipose tissue and testicular hormonal regulation induced by metabolic disorders can also contribute to a state of compromised fertility. In this Review, we will delve into the effects of high-fat diets and metabolic disorders on testicular metabolism and spermatogenesis, which are crucial elements for male reproductive function. Moreover, metabolic disorders have been shown to influence the epigenome of male gametes and might have a potential role in transmitting phenotype traits across generations. However, the existing evidence strongly underscores the unmet need to understand the mechanisms responsible for transgenerational paternal inheritance of male reproductive function impairment related to metabolic disorders. This knowledge could be useful for developing targeted interventions to prevent, counteract, and most of all break the perpetuation chain of male reproductive dysfunction associated with metabolic disorders across generations.

Mitochondrial uncoupling proteins regulate the metabolic function of human Sertoli cells.

In brief: Mitochondrial uncoupling proteins (UCPs) regulate mitochondrial activity and reactive oxygen species production through the transport of protons and metabolites. This study identified the expression of UCPs in human Sertoli cells, which proved to be modulators of their mitochondrial activity. Abstract: Mitochondrial uncoupling proteins (UCPs) are mitochondrial channels responsible for the transport of protons and small molecular substrates across the inner mitochondrial membrane. Altered UCP expression or function is commonly associated with mitochondrial dysfunction and increased oxidative stress, which are both known causes of male infertility. However, UCP expression and function in the human testis remain to be characterized. This study aimed to assess the UCP homologs (UCP1-6) expression and function in primary cultures of human Sertoli cells (hSCs). We identified the mRNA expression of all UCP homologs (UCP1-6) and protein expression of UCP1, UCP2, and UCP3 in hSCs. UCP inhibition by genipin for 24 h decreased hSCs proliferation without causing cytotoxicity (n = 6). Surprisingly, the prolonged UCP inhibition for 24 h decreased mitochondrial membrane potential, oxygen consumption rate (OCR), and endogenous reactive oxygen species (ROS) production. The metabolism of hSCs was also affected as UCP inhibition shifted their metabolism toward an increased pyruvate consumption. Taken together, these findings demonstrate that UCPs play a role as regulators of the mitochondrial function in hSCs, emphasizing their potential as targets in the study of male (in)fertility.

Impact of Different Treatment Regimens and Timeframes in the Plasmatic Metabolic Profiling of Patients with Lung Adenocarcinoma.

In recent years, the treatment of advanced non-small cell lung cancer (NSCLC) has suffered a variety of alterations. Chemotherapy (CTX), immunotherapy (IT) and tyrosine kinase inhibitors (TKI) have shown remarkable results. However, not all patients with NSCLC respond to these drug treatments or receive durable benefits. In this framework, metabolomics has been applied to improve the diagnosis, treatment, and prognosis of lung cancer and particularly lung adenocarcinoma (AdC). In our study, metabolomics was used to analyze plasma samples from 18 patients with AdC treated with CTX or IT via 1H-NMR spectroscopy. Relevant clinical information was gathered, and several biochemical parameters were also evaluated throughout the treatments. During the follow-up of patients undergoing CTX or IT, imaging control is recommended in order to assess the effectiveness of the therapy. This evaluation is usually performed every three treatments. Based on this procedure, all the samples were collected before the beginning of the treatment and after three and six treatments. The identified and quantified metabolites in the analyzed plasma samples were the following: isoleucine, valine, alanine, acetate, lactate, glucose, tyrosine, and formate. Multivariate/univariate statistical analyses were performed. Our data are in accordance with previous published results, suggesting that the plasma glucose levels of patients under CTX become higher throughout the course of treatment, which we hypothesize could be related to the tumor response to the therapy. It was also found that alanine levels become lower during treatment with CTX regimens, a fact that could be associated with frailty. NMR spectra of long responders' profiles also showed similar results. Based on the results of the study, metabolomics can represent a potential option for future studies, in order to facilitate patient selection and the monitoring of therapy efficacy in treated patients with AdC. Further studies are needed to improve the prospective identification of predictive markers, particularly glucose and alanine levels, as well as confer guidance to NSCLC treatment and patient stratification, thus avoiding ineffective therapeutic strategies.

The progression from mild to severe hyperglycemia coupled with insulin resistance causes mitochondrial dysfunction and alters the metabolic secretome of epithelial kidney cells.

Diabetic nephropathy (DN) and insulin resistance (IR) in kidney cells are considered main causes for end-stage renal failure. However, it is unclear how IR affects early stages of the disease. Here, we investigate the impact of mild (11 mM) and severe (22 mM) hyperglycemia, with and without induced IR, on cellular metabolism and mitochondrial bioenergetics in a human kidney cell line (HK-2). IR in HK-2 cells was induced with palmitic acid and cellular cytotoxicity was studied. We evaluated the impact of mild and severe hyperglycemia with and without IR on the metabolic secretome of the cells, their live-cell mitochondria function, mitochondrial membrane potential, and mitochondrial complex activities. Furthermore, we measured fatty acid oxidation and lipid accumulation. Cells cultured under mild hyperglycemic conditions exhibited increased mitochondrial bioenergetic parameters, such as basal respiration, ATP-linked production, maximal respiration capacity, and spare respiration capacity. However, these parameters decreased when cells were cultured under higher glucose concentrations when IR was induced. Our data suggests that progression from mild to severe hyperglycemia induces a metabolic shift, where gluconeogenic amino acids play a crucial role in supplying the energy requirements of HK-2. To our knowledge, this is the first study to evaluate the progression from mild to severe hyperglycemia allied to IR in human kidney cells. This work highlights that this progression leads to mitochondrial dysfunction and alters the metabolic profile of kidney cells. These results identify possible targets for early intervention in DN.

Dysglycemia Shapes Visceral Adipose Tissue's Response to GIP, GLP-1 and Glucagon in Individuals with Obesity.

Visceral adipose tissue (VAT) metabolic fingerprints differ according to body mass index (BMI) and glycemic status. Glucagon-like peptide 1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and glucagon are gut-associated hormones that play an important role in regulating energy and glucose homeostasis, although their metabolic actions in VAT are still poorly characterized. Our aim was to assess whether GLP-1, GIP and glucagon influence the VAT metabolite profile. To achieve this goal, VAT harvested during elective surgical procedures from individuals (N = 19) with different BMIs and glycemic statuses was stimulated with GLP-1, GIP or glucagon, and culture media was analyzed using proton nuclear magnetic resonance. In the VAT of individuals with obesity and prediabetes, GLP-1 shifted its metabolic profile by increasing alanine and lactate production while also decreasing isoleucine consumption, whereas GIP and glucagon decreased lactate and alanine production and increased pyruvate consumption. In summary, GLP-1, GIP and glucagon were shown to distinctively modulate the VAT metabolic profile depending on the subject's BMI and glycemic status. In VAT from patients with obesity and prediabetes, these hormones induced metabolic shifts toward gluconeogenesis suppression and oxidative phosphorylation enhancement, suggesting an overall improvement in AT mitochondrial function.

Inhibition of Mitochondrial Uncoupling Proteins Arrests Human Spermatozoa Motility without Compromising Viability.

Mitochondrial uncoupling proteins (UCPs) are central in the regulation of mitochondrial activity and reactive oxygen species (ROS) production. High oxidative stress is a major cause of male infertility; however, UCPs expression and function in human spermatozoa are still unknown. Herein, we aimed to assess the expression and function of the different homologs (UCP1-6) in human spermatozoa. For this purpose, we screened for the mRNA expression of all UCP homologs. Protein expression and immunolocalization of UCP1, UCP2, and UCP3 were also assessed. Highly motile spermatozoa were isolated from human normozoospermic seminal samples (n = 16) and incubated with genipin, an inhibitor of UCPs (0, 0.5, 5, and 50 µM) for 3 h at 37 °C. Viability and total motility were assessed. Mitochondrial membrane potential and ROS production were evaluated. Media were collected and the metabolic profile and antioxidant potential were analyzed by 1H-NMR and FRAP, respectively. The expression of all UCP homologs (UCP1-6) mRNA by human spermatozoa is herein reported for the first time. UCP1-3 are predominant at the head equatorial segment, whereas UCP1 and UCP2 are also expressed at the spermatozoa midpiece, where mitochondria are located. The inhibition of UCPs by 50 µM genipin, resulting in the UCP3 inhibition, did not compromise sperm cell viability but resulted in irreversible total motility loss that persisted despite washing or incubation with theophylline, a cAMP activator. These effects were associated with decreased mitochondrial membrane potential and lactate production. No differences concerning UCP3 expression, however, were observed in spermatozoa from normozoospermic versus asthenozoospermic men (n = 6). The inhibition of UCPs did not increase ROS production, possibly due to the decreased mitochondrial activity and genipin antioxidant properties. In sum, UCPs are major regulators of human spermatozoa motility and metabolism. The discovery and characterization of UCPs' role in human spermatozoa can shed new light on spermatozoa ROS-related pathways and bioenergetics physiology.

CFTR modulates aquaporin-mediated glycerol permeability in mouse Sertoli cells.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel that is crucial for fluid homeodynamics throughout the male reproductive tract. Previous evidence shed light on a potential molecular partnership between this channel and aquaporins (AQPs). Herein, we explore the role of CFTR on AQPs-mediated glycerol permeability in mouse Sertoli cells (mSCs). We were able to identify the expression of CFTR, AQP3, AQP7, and AQP9 in mSCs by RT-PCR, Western blot, and immunofluorescence techniques. Cells were then treated with CFTRinh-172, a specific CFTR inhibitor, and its glycerol permeability was evaluated by stopped-flow light scattering. We observed that CFTR inhibition decreased glycerol permeability in mSCs by 30.6% when compared to the control group. A DUOLINK proximity ligation assay was used to evaluate the endogenous protein-protein interactions between CFTR and the various aquaglyceroporins we identified. We positively detected that CFTR is in close proximity with AQP3, AQP7, and AQP9 and that, through a possible physical interaction, CFTR can modulate AQP-mediated glycerol permeability in mSCs. As glycerol is essential for the control of the blood-testis barrier and elevated concentration in testis results in the disruption of spermatogenesis, we suggest that the malfunction of CFTR and the consequent alteration in glycerol permeability is a potential link between male infertility and cystic fibrosis.

Evaluation of Human Spermatozoa Mitochondrial Membrane Potential Using the JC-1 Dye.

Mitochondria are fundamental for human spermatozoa motility and fertilizing ability. Mitochondria participate not only in ATP production, but also in reactive oxygen species production, redox equilibrium, and calcium regulation, all of which are central for human spermatozoa motility, capacitation, acrosome reaction, and ultimately, oocyte fertilization. Mitochondrial membrane potential is a key indicator of mitochondrial health and activity. Most commonly used methods for the study of mitochondrial membrane potential, however, cannot be applied to human spermatozoa due to their unique characteristics, including high motility and time-dependent decay of quality, limiting the study of this important parameter in these cells. Here, we describe an easy, fast, and cheap protocol for the quantitative evaluation of human spermatozoa mitochondrial membrane potential, using the fluorescent cationic dye 5,5,6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimi-dazoylcarbocyanine iodide (JC-1). JC-1 is a sensitive marker for mitochondrial membrane potential, exhibiting a potential-dependent accumulation in the mitochondria. At high mitochondrial membrane potential, JC-1 forms J-aggregates, which emit red fluorescence, whereas at low mitochondrial membrane potential, JC-1 remains at its monomer state, which emits green fluorescence. We first describe how to evaluate human spermatozoa mitochondrial membrane potential using JC-1 and a fluorescence plate reader, for high-throughput studies. The calculation of the JC-1 ratio (indicative of the J-aggregates/monomers ratio) is then used to quantitatively evaluate mitochondrial health and activity. In addition, we describe an imaging protocol for the qualitative evaluation of human spermatozoa mitochondrial membrane potential using a fluorescence microscope. This allows for a visual analysis of the results that can complement the quantitative data. These protocols can be used to study the effects of spermatozoa exposure to compounds of interest, and alterations due to diseases or different conditions. While these protocols are illustrated with human spermatozoa, they can be adapted and used on spermatozoa of different species. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Quantitative evaluation of human spermatozoa mitochondrial membrane potential using the JC-1 dye and a fluorescence plate reader Basic Protocol 2: Qualitative evaluation of human spermatozoa mitochondrial membrane potential using the JC-1 dye and fluorescence microscopy Support Protocol: Preparation of the JC-1 working solution.

Carbamate Pesticides: Shedding Light on Their Impact on the Male Reproductive System.

Carbamates are widely used and known around the world as pesticides in spite of also having medical applications. This class of chemicals is classified as acetylcholinesterase inhibitors, blocking acetylcholine hydrolyzation in a reversible manner. Their lack of species selectivity and their reported high toxicity can induce, upon exposure, adverse outcomes in male fertility that may lead to infertility. In addition, they are also considered endocrine-disrupting chemicals and can interfere with the hypothalamic-pituitary-testicular axis, essential for the normal function of the male reproductive system, thus being able to provoke male reproductive dysfunctions. Although the molecular mechanisms are not fully understood, various signaling pathways, such as those mediated by acetylcholine or kisspeptin, are affected by exposure to carbamates, thus compromising steroidogenesis and spermatogenesis. Over the last decades, several studies, both in vitro and in vivo, have reported a myriad of negative effects of carbamates on the male reproductive system. In this review, an up-to-date overview of the impact of carbamates on the male reproductive system is discussed, with an emphasis on the role of these compounds on acetylcholine regulation and the male endocrine system.

Antioxidants and Male Infertility.

Oxidative stress has been associated with decreased sperm quality and male infertility [...].

Animal models of male reproductive ageing to study testosterone production and spermatogenesis.

Ageing is the time-dependent gradual decline of the functional characteristics in an organism. It has been shown that it results in the loss of reproductive health and fertility. The age-dependent decline of fertility is a potential issue as the parenthood age is increasing in Western countries, mostly due to socioeconomic factors. In comparison to women, for whom the consequences of ageing are well documented and general awareness of the population is extensively raised, the effects of ageing for male fertility and the consequences of advanced paternal age for the offspring have not been widely studied. Studies with humans are welcome but it is hard to implement relevant experimental approaches to unveil the molecular mechanisms by which ageing affects male reproductive potential. Animal models have thus been extensively used. These models are advantageous due to their reduced costs, general easy maintenance in laboratory facilities, rigorous manipulation tools, short lifespan, known genetic backgrounds, and reduced ethical constraints. Herein, we discuss animal models for the study of male reproductive ageing. The most well-known and studied reproductive ageing models are rodents and non-human primates. The data collected from these models, particularly studies on testicular ageing, steroidogenesis, and genetic and epigenetic changes in spermatogenesis are detailed. Notably, some species challenge the currently accepted ageing theories and the concept of senescence itself, which renders them interesting animal models for the study of male reproductive ageing.

Metabolomics as a tool for the early diagnosis and prognosis of diabetic kidney disease.

Diabetic kidney disease (DKD) is one of the most prevalent comorbidities of diabetes mellitus and the leading cause of the end-stage renal disease (ESRD). DKD results from chronic exposure to hyperglycemia, leading to progressive alterations in kidney structure and function. The early development of DKD is clinically silent and when albuminuria is detected the lesions are often at advanced stages, leading to rapid kidney function decline towards ESRD. DKD progression can be arrested or substantially delayed if detected and addressed at early stages. A major limitation of current methods is the absence of albuminuria in non-albuminuric phenotypes of diabetic nephropathy, which becomes increasingly prevalent and lacks focused therapy. Metabolomics is an ever-evolving omics technology that enables the study of metabolites, downstream products of every biochemical event that occurs in an organism. Metabolomics disclosures complex metabolic networks and provide knowledge of the very foundation of several physiological or pathophysiological processes, ultimately leading to the identification of diseases' unique metabolic signatures. In this sense, metabolomics is a promising tool not only for the diagnosis but also for the identification of pre-disease states which would confer a rapid and personalized clinical practice. Herein, the use of metabolomics as a tool to identify the DKD metabolic signature of tubule interstitial lesions to diagnose or predict the time-course of DKD will be discussed. In addition, the proficiency and limitations of the currently available high-throughput metabolomic techniques will be discussed.

Aquaporins and Animal Gamete Cryopreservation: Advances and Future Challenges.

Cryopreservation is globally used as a method for long-term preservation, although freeze-thawing procedures may strongly impair the gamete function. The correct cryopreservation procedure is characterized by the balance between freezing rate and cryoprotective agents (CPAs), which minimizes cellular dehydration and intracellular ice formation. For this purpose, osmoregulation is a central process in cryopreservation. During cryopreservation, water and small solutes, including penetrating cryoprotective agents, cross the plasma membrane. Aquaporins (AQPs) constitute a family of channel proteins responsible for the transport of water, small solutes, and certain gases across biological membranes. Thirteen homologs of AQPs (AQP0-12) have been described. AQPs are widely distributed throughout the male and female reproductive systems, including the sperm and oocyte membrane. The composition of the male and female gamete membrane is of special interest for assisted reproductive techniques (ART), including cryopreservation. In this review, we detail the mechanisms involved in gamete cryopreservation, including the most used techniques and CPAs. In addition, the expression and function of AQPs in the male and female gametes are explored, highlighting the potential protective role of AQPs against damage induced during cryopreservation.

Hyperoside Supplementation in Preservation Media Surpasses Vitamin C Protection Against Oxidative Stress-Induced Damages in Human Spermatozoa.

BACKGROUND/AIMS: Oxidative Stress (OS) is reported as one of the main causes of male infertility. Infertile couples often resort to assisted reproductive technology (ART) to achieve parenthood. However, preparation for ART protocols increases the exposer of gametes to OS. Thus, it is crucial to find suitable preservation media that can counteract the OS-induced damages in spermatozoa. In this work, we tested and compared the efficiency of vitamin C (VC) and hyperoside (HYP) as potential antioxidant supplements for sperm preservation media. METHODS: We evaluated the cytotoxicity of HYP (0, 5, 50, 100, and 500 µM) in spermatozoa. After incubation of sperm cells with VC (600 µM) and HYP (100 and 500 µM), in the presence and absence of H2O2 (300 µM), the following parameters were assessed: total sperm motility and vitality, OS biomarkers expression, total antioxidant capacity (TAC) of the media, percentage of DNA fragmentation, mitochondrial membrane potential (MMP), and metabolite quantification of the media by proton nuclear magnetic resonance (1H-NMR). RESULTS: The supplementation with VC (600 µM) and HYP (100 and 500 µM) did not induce any deleterious effects to the physiology and metabolism of the spermatozoa, after 1-hour of treatment. In the presence of H2O2 (300 µM), both VC and HYP were able to prevent some of the deleterious effects of H2O2 in sperm, which were represented by an increase in sperm motility, a decrease in DNA fragmentation, and a decreasing trend in lipid peroxidation levels. However, these antioxidants were not able to prevent the decrease of MMP associated with H2O2 treatment, nor were able to prevent the conversion of pyruvate into acetate (a reaction promoted by H2O2). CONCLUSION: The supplementation of sperm preservation media with VC and HYP could be beneficial for the preservation of sperm physiology. From the antioxidant conditions tested, the supplementation of media with HYP (100 µM) demonstrated the best results regarding sperm preservation, evidencing the higher antioxidant capacity of HYP compared to VC. Nevertheless, none of the antioxidants used was able to prevent the metabolic alterations promoted by H2O2 in spermatozoa.

Molecular mechanisms regulating spermatogenesis in vertebrates: Environmental, metabolic, and epigenetic factor effects.

The renewal of the natural resources is one of the most concerning aspects of modern farming. In animal production, there are many barriers breeders and researchers have to overcome to develop new practices to improve reproductive potential and hasten sexual maturation of the commercially viable species, while maintaining meat quality and sustainability. With the utilization of molecular biology techniques, there have been relevant advances in the knowledge of spermatogenesis, especially in mammals, resulting in new possibilities to control male fertility and the selection of desirable characteristics. Most of these discoveries have not been implemented in animal production. In this review, recent studies are highlighted on the molecular pathways involved in spermatogenesis in the context of animal production. There is also exploration of the interaction between environmental factors and spermatogenesis and how this knowledge may revolutionize animal production techniques. Furthermore, new insights are described about the inheritance of desired characteristics in mammals and there is a review of nefarious actions of pollutants, nutrition, and metabolism on reproductive potential in subsequent generations. Even though there are these advances in knowledge base, results from recent studies indicate there are previously unrecognized environmental effects on spermatogenesis. The molecular mechanisms underlying this interaction are not well understood. Research in spermatogenesis, therefore, remains pivotal as a pillar of animal production sustainability.

Mitochondrial Uncoupling Proteins (UCPs) as Key Modulators of ROS Homeostasis: A Crosstalk between Diabesity and Male Infertility?

Uncoupling proteins (UCPs) are transmembrane proteins members of the mitochondrial anion transporter family present in the mitochondrial inner membrane. Currently, six homologs have been identified (UCP1-6) in mammals, with ubiquitous tissue distribution and multiple physiological functions. UCPs are regulators of key events for cellular bioenergetic metabolism, such as membrane potential, metabolic efficiency, and energy dissipation also functioning as pivotal modulators of ROS production and general cellular redox state. UCPs can act as proton channels, leading to proton re-entry the mitochondrial matrix from the intermembrane space and thus collapsing the proton gradient and decreasing the membrane potential. Each homolog exhibits its specific functions, from thermogenesis to regulation of ROS production. The expression and function of UCPs are intimately linked to diabesity, with their dysregulation/dysfunction not only associated to diabesity onset, but also by exacerbating oxidative stress-related damage. Male infertility is one of the most overlooked diabesity-related comorbidities, where high oxidative stress takes a major role. In this review, we discuss in detail the expression and function of the different UCP homologs. In addition, the role of UCPs as key regulators of ROS production and redox homeostasis, as well as their influence on the pathophysiology of diabesity and potential role on diabesity-induced male infertility is debated.

Sperm selection strategies and their impact on assisted reproductive technology outcomes.

The application of assisted reproductive technologies (ART) has revolutionised the treatment of human infertility, giving hope to the patients previously considered incapable of establishing pregnancy. While semen analysis is performed to access whether a sample has an adequate number of viable, motile and morphologically normal sperm cells able to achieve fertilisation, sperm selection techniques for ART aim to isolate the most competent spermatozoon which is characterised by the highest fertilising potential. Based on the semen analysis results, the correct sperm selection technique must be chosen and applied. In this review, different sperm selection strategies for retrieving spermatozoa with the highest fertilising potential and their impact on ART outcomes are discussed. In addition, advantages and disadvantages of each method and the best suited techniques for each clinical scenario are described.

Mitochondrial Activation and Reactive Oxygen-Species Overproduction during Sperm Capacitation are Independent of Glucose Stimuli.

Spermatozoa capacitation is a complex process that requires specific ionic and energetic conditions to support biochemical alterations leading to motility hyperactivation. However, human sperm capacitation is still poorly understood. Herein, we studied the effects of glucose on human sperm capacitation. Healthy men seminal samples (n = 55) were submitted to a density gradient centrifugation and incubated in capacitating conditions in the absence or presence of increasing glucose concentrations (0, 5.5, 11, and 22 mM). Viability and total motility were accessed. Phosphotyrosine levels were measured. Mitochondrial activity and endogenous ROS production were evaluated. Oxidative stress-induced damage was analyzed. Culture media was collected and analyzed by 1H-NMR. Our results show that glucose is essential for human sperm capacitation and motility. Notably, we observed that mitochondrial activity increased even in the absence of glucose. This increased mitochondrial activity was followed by a ROS overproduction, although no oxidative stress-induced damage was detected. Our results show that glucose is essential for capacitation but mitochondrial activation is independent from its stimuli. ROS overproduction may take part on a finely regulated signaling pathway that modulates or even activates capacitation. Taken together, our results constitute a paradigm shift on human sperm capacitation physiology.

CFTR regulation of aquaporin-mediated water transport.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel responsible for the direct transport of bicarbonate and chloride. CFTR-dependent ionic transport is crucial for pH regulation and fluid homeodynamics among epithelial surfaces. Particularly, CFTR performs an essential role in the male reproductive tract, which requires a tight regulation of water and electrolytes in order to produce healthy spermatozoa. The absence or malfunction of CFTR results in cystic fibrosis, the most common lethal disease among Caucasians, that is characterized by an impaired fluid and ionic homeostasis in the whole organism. Due to the wide expression and importance of CFTR, the male reproductive tract is highly affected by cystic fibrosis, resulting in male infertility. Although CFTR is not permeable to water, this protein acts as a regulator of other protein channels, such as aquaporins. In fact, CFTR acts as a molecular partner of aquaporins in epithelial cells, regulating fluid homeodynamics. Herein, up-to-date data concerning the regulation of aquaporin-mediated water transport by CFTR will be discussed, highlighting the role of both channels in the male reproductive tract.