Medical Physiology

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Browsing Medical Physiology by Author "Agarwal, Ashok"

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    Influence of ejaculation frequency on seminal parameters
    (BioMed Central, 2015-05-21) Mayorga-Torres, B. Jose Manuel; Camargo, Mauricio; Agarwal, Ashok; Du Plessis, Stefan S.; Cadavid, Angela P.; Maya, Walter D. Cardona
    Background: Several factors have been shown to influence semen parameters, one of which is sexual abstinence; a clinical criteria included in the semen evaluation to provide maximum sperm quality. The aim of the present study was to assess the effect of a daily ejaculation frequency on conventional and functional semen parameters. Methods: Semen samples were collected daily over a period of two weeks of which every second sample per person was processed and analyzed according to the World Health Organization guidelines. Furthermore, mitochondrial function, intracellular reactive oxygen species production and sperm DNA fragmentation were evaluated by flow cytometry. Results: Total sperm count and seminal volume per ejaculation declined and remained decreased for the duration of the daily ejaculation period. However, conventional parameters such as sperm concentration, motility, progressive motility, morphology, vitality and functional parameters such as sperm plasma membrane integrity, mitochondrial membrane potential and DNA fragmentation was not significantly affected and remained similar to the initial measurement throughout the daily ejaculation period. Despite intra- and inter individual variations, the average values of the basic semen parameters remained above the WHO (2010) reference values throughout the daily ejaculation period. Interestingly, a decreasing trend in intracellular ROS production was observed, although statistically not significant. Conclusions: The study shows that an extended 2 week period of daily ejaculation does not have major clinical effects on conventional and functional seminal parameters.
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    Proteomic analysis of human spermatozoa proteins with oxidative stress
    (BioMed Central, 2013-05) Sharma, Rakesh; Agarwal, Ashok; Mohanty, Gayatri; Hamada, Alaa J.; Gopalan, Banu; Willard, Belinda; Yadav, Satya; Du Plessis, Stefan
    Background: Oxidative stress plays a key role in the etiology of male infertility. Significant alterations in the sperm proteome are associated with poor semen quality. The aim of the present study was to examine if elevated levels of reactive oxygen species cause an alteration in the proteomic profile of spermatozoa. Methods This prospective study consisted of 52 subjects: 32 infertile men and 20 normal donors. Seminal ejaculates were classified as ROS+ or ROS- and evaluated for their proteomic profile. Samples were pooled and subjected to LC-MS/MS analysis through in-solution digestion of proteins for peptide characterization. The expression profile of proteins present in human spermatozoa was examined using proteomic and bioinformatic analysis to elucidate the regulatory pathways of oxidative stress. Results Of the 74 proteins identified, 10 proteins with a 2-fold difference were overexpressed and 5 were underexpressed in the ROS+ group; energy metabolism and regulation, carbohydrate metabolic processes such as gluconeogenesis and glycolysis, protein modifications and oxidative stress regulation were some of the metabolic processes affected in ROS+ group. Conclusions We have identified proteins involved in a variety of functions associated with response and management of oxidative stress. In the present study we focused on proteins that showed a high degree of differential expression and thus, have a greater impact on the fertilizing potential of the spermatozoa. While proteomic analyses identified the potential biomarkers, further studies through Western Blot are necessary to validate the biomarker status of the proteins in pathological conditions.
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    Proteomic analysis of seminal fluid from men exhibiting oxidative stress
    (BioMed Central, 2013-09) Sharma, Rakesh; Agarwal, Ashok; Mohanty, Gayatri; Du Plessis, Stefan S.; Gopalan, Banu; Willard, Belinda; Yadav, Satya P.; Sabanegh, Edmund
    ABSTRACT: Background: Seminal plasma serves as a natural reservoir of antioxidants. It helps to remove excessive formation of reactive oxygen species (ROS) and consequently, reduce oxidative stress. Proteomic profiling of seminal plasma proteins is important to understand the molecular mechanisms underlying oxidative stress and sperm dysfunction in infertile men. Methods This prospective study consisted of 52 subjects: 32 infertile men and 20 healthy donors. Once semen and oxidative stress parameters were assessed (ROS, antioxidant concentration and DNA damage), the subjects were categorized into ROS positive (ROS+) or ROS negative (ROS-). Seminal plasma from each group was pooled and subjected to proteomics analysis. In-solution digestion and protein identification with liquid chromatography tandem mass spectrometry (LC-MS/MS), followed by bioinformatics analyses was used to identify and characterize potential biomarker proteins. Results A total of 14 proteins were identified in this analysis with 7 of these common and unique proteins were identified in both the ROS+ and ROS- groups through MASCOT and SEQUEST analyses, respectively. Prolactin-induced protein was found to be more abundantly present in men with increased levels of ROS. Gene ontology annotations showed extracellular distribution of proteins with a major role in antioxidative activity and regulatory processes. Conclusions We have identified proteins that help protect against oxidative stress and are uniquely present in the seminal plasma of the ROS- men. Men exhibiting high levels of ROS in their seminal ejaculate are likely to exhibit proteins that are either downregulated or oxidatively modified, and these could potentially contribute to male infertility.
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    Proteomics : a subcellular look at spermatozoa
    (BioMed Central, 2011-03) Du Plessis, Stefan S.; Kashou, Anthony H.; Benjamin, David J.; Yadav, Satya P.; Agarwal, Ashok
    Abstract. Background. Male-factor infertility presents a vexing problem for many reproductively active couples. Many studies have focused on abnormal sperm parameters. Recent advances in proteomic techniques, especially in mass spectrometry, have aided in the study of sperm and more specifically, sperm proteins. The aim of this study was to review the current literature on the various proteomic techniques, and their usefulness in diagnosing sperm dysfunction and potential applications in the clinical setting. Methods Review of PubMed database. Key words: spermatozoa, proteomics, protein, proteome, 2D-PAGE, mass spectrometry. Results. Recently employed proteomic methods, such as two-dimensional polyacrylamide gel electrophoresis, mass spectrometry, and differential in gel electrophoresis, have identified numerous sperm-specific proteins. They also have provided a further understanding of protein function involved in sperm processes and for the differentiation between normal and abnormal states. In addition, studies on the sperm proteome have demonstrated the importance of post-translational modifications, and their ability to bring about physiological changes in sperm function. No longer do researchers believe that in order for them to elucidate the biochemical functions of genes, mere knowledge of the human genome sequence is sufficient. Moreover, a greater understanding of the physiological function of every protein in the tissue-specific proteome is essential in order to unravel the biological display of the human genome. Conclusion Recent advances in proteomic techniques have provided insight into sperm function and dysfunction. Several multidimensional separation techniques can be utilized to identify and characterize spermatozoa. Future developments in bioinformatics can further assist researchers in understanding the vast amount of data collected in proteomic studies. Moreover, such advances in proteomics may help to decipher metabolites which can act as biomarkers in the detection of sperm impairments and to potentially develop treatment for infertile couples. Further comprehensive studies on sperm-specific proteome, mechanisms of protein function and its proteolytic regulation, biomarkers and functional pathways, such as oxidative-stress induced mechanisms, will provide better insight into physiological functions of the spermatozoa. Large-scale proteomic studies using purified protein assays will eventually lead to the development of novel biomarkers that may allow for detection of disease states, genetic abnormalities, and risk factors for male infertility. Ultimately, these biomarkers will allow for a better diagnosis of sperm dysfunction and aid in drug development.
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    Utility of antioxidants during assisted reproductive techniques : an evidence based review
    (BioMed Central, 2014-11-24) Agarwal, Ashok; Durairajanayagam, Damayanthi; Du Plessis, Stefan S.
    Assisted reproductive technology (ART) is a common treatment of choice for many couples facing infertility issues, be it due to male or female factor, or idiopathic. Employment of ART techniques, however, come with its own challenges as the in vitro environment is not nearly as ideal as the in vivo environment, where reactive oxygen species (ROS) build-up leading to oxidative stress is kept in check by the endogenous antioxidants system. While physiological amounts of ROS are necessary for normal reproductive function in vivo, in vitro manipulation of gametes and embryos exposes these cells to excessive ROS production either by endogenous or exogenous environmental factors. In this review, we discuss the sources of ROS in an in vitro clinical setting and the influence of oxidative stress on gamete/embryo quality and the outcome of IVF/ICSI. Sources of ROS and different strategies of overcoming the excessive generation of ROS in vitro are also highlighted. Endogenously, the gametes and the developing embryo become sources of ROS. Multiple exogenous factors act as potential sources of ROS, including exposure to visible light, composition of culture media, pH and temperature, oxygen concentration, centrifugation during spermatozoa preparation, ART technique involving handling of gamete/embryo and cryopreservation technique (freeze/thawing process). Finally, the use of antioxidants as agents to minimize ROS generation in the in vitro environment and as oral therapy is highlighted. Both enzymatic and non-enzymatic antioxidants are discussed and the outcome of studies using these antioxidants as oral therapy in the male or female or its use in vitro in media is presented. While results of studies using certain antioxidant agents are promising, the current body of evidence as a whole suggests the need for further well-designed and larger scale randomized controlled studies, as well as research to minimize oxidative stress conditions in the clinical ART setting.