Browsing by Author "Flint, Margot Gwyneth"

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    Gamete proteomic profile of male patients suffering from sexually transmitted infections
    (Stellenbosch : Stellenbosch University, 2016-03) Flint, Margot Gwyneth; Du Plessis, Stefan; Van der Horst, Gerhard; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences. Medical Physiology.
    ENGLISH ABSTRACT: The aim of this study was to provide better insight into the effects of Neisseria gonorrhoea, Chlamydia trachomatis, and Trichomonas vaginalis on semen characteristics. In addition, a crucial focus was also to determine the protein profile of spermatozoa isolated from these infected semen samples. Identification of semen samples positive for bacterial colonisation of N. gonorrhoea and T. vaginalis were done through the use of differential isolation mediums. For the detection of samples positive for C. trachomatis, the ImmunoComb® was employed, which is a quantitative indirect enzyme immunoassay. Macro- and microscopic semen and spermatozoa parameters were assessed, including: volume, pH, viscosity, concentration, motility, morphology, viability, acrosome reaction, leukocyte count, and PMN-elastase concentration. To assess prostate and seminal vesicle functioning, the seminal concentrations of citric acid and fructose were photometrically quantified. The quantification of the level of reactive oxygen species (ROS) production was determined by means of flow cytometry and the DNA fragmentation was detected using a commercially available assay. For proteomic analysis, samples were iTRAQ (isobaric tags for relative and absolute quantification) labelled and underwent liquid chromatography-mass spectrometry, followed by data analysis, protein identification, and quantification. Results of the study showed that amongst the three microorganisms, the most prevalent occurrence rate in the population tested was that of T. vaginalis. The effects of the studied sexually transmitted infections (STIs) on the spermatozoa parameters demonstrate the negative impact of the microorganisms on the fertility outcome of the male partner. The study has shown the relationship between leukocyte-derived ROS and DNA fragmentation, which can significantly impair the fertility outcome. Additionally, the findings of decreased concentrations of fructose in the ejaculates positive for N. gonorrhoea, demonstrates the effect of an STI on the glandular functioning of the secretory activity of the seminal vesicles. The observed negative correlation between ROS and DNA fragmentation in samples positive for N. gonorrhoea demonstrates the impact of a pro-oxidant overload on spermatozoa DNA integrity. A considerable number of 178 differentially expressed proteins (DEPs) were identified in the STI positive sample groups. The study subsequently focused on specific proteins according to their role in male fertility. Insight into the role DEPs may play in spermatozoa metabolism and the impact on the motility was provided. Amongst structural related proteins, the down-regulation of outer dense fibre 2, in samples positive for C. trachomatis, was evaluated. Oxidative stress related proteins included the up-regulated superoxide dismutase 1 in the ejaculates positive for N. gonorrhoea and down-regulated peroxiredoxin 5 in the C. trachomatis positive group. From the proteins involved in the response to physiological stress, heat shock proteins was discussed, with the down-regulation of the 70-kDa heat shock protein and the up-regulated HSP 90-kDa-beta member 1, both identified in samples positive for N. gonorrhoea. Among immune response proteins, prolactin-inducible protein (C. trachomatis) was found to be down-regulated, while azurocidin (N. gonorrhoea) and filamin-B (T. vaginalis) were up-regulated. Proteins involved in DNA condensation included down-regulated protamine-2 (C. trachomatis). The identification of proteins, which are differentially expressed between spermatozoa from samples positive for STIs, can provide crucial insight into their possible influence on male fertility and role as potential biomarkers for further research. Research on a proteomic and molecular level could allow for the prevention of the long-term obstacles facing partners experiencing compromised fertility. Despite the ongoing research focusing on the sperm proteome, it can be stated with reasonable certainty, that this is the first study into the proteomic profile and corresponding parameters of spermatozoa isolated from semen samples positive for STIs.