| dc.description.abstract | The disequilibrium of reactive oxygen/nitrogen species (ROS/RNS) is one of the causes of male infertility. To examine the role of ROS/RNS in reproduction, we modulated the cell concentration of О2•– and NO using the superoxide dismutase (SOD) mimic, M40403, which selectively removes О2•– and consequently increases the NO bioavailability. The fine chromatin changes are difficult to observe via routine light/electron microscopy, therefore we used fractal analysis to analyze DNA compaction. We used normospermic semen samples from ten human subjects. After purification in Cook density gradient, the sperm-rich fraction was rinsed in modified Tyrod medium (TM). Purified samples were divided into three groups. One group was evaluated immediately after resuspension in TM and served as the control. For the other two groups, untreated and SOD mimic-treated group (50 μM), the sperm was resuspended in TM and evaluated after incubation at 37 ºC / 6% CO2 for 3 hours. Air-dried and methanol fixed sperm smears were stained with toluidine blue and analyzed on Leica DMLB microscope. Images from ten randomly selected fields were analyzed in ImageJ plugin FracLac to get fractal dimension defined as a measure of complexity. Average values of fractal dimensions (mean ± SEM) are: control group – 1.016 ± 0.553; untreated group – 0.955 ± 0.291; treated group – 1.146 ± 0.096. Also, average values of lacunarity are: control group – 0.734 ± 0.129; untreated group – 0.727 ± 0.129; treated group – 0.901 ± 0.297. Results show that SOD mimic remodels chromatin condensation by increasing the euchromatin area, potentially leading to a resume in transcriptional activity. This hypothesis would be consistent with our published findings of up-regulated mRNA expression of eNOS, MnSOD, and catalase in SOD mimic-treated spermatozoa ex vivo (Otasevic et al., 2013). Therefore, SOD mimic modulates the redox environment via direct chromatin remodeling in spermatozoa. Fractal analysis has proven to be a good method for the analysis of chromatin condensation in human sperm nuclei. Ref: Otasevic V, Korac A, Vucetic M, Macanovic B, Garalejic E, Ivanovic-Burmazovic I, Filipovic MR, Buzadzic B, Stancic A, Jankovic A, Velickovic K, Golic I, Markelic M, Korac B. Is manganese (II) pentaazamacrocyclic superoxide dismutase mimic beneficial for human sperm mitochondria function and motility? Antioxid Redox Signal. 2013 Jan 10;18(2):170-8. doi: 10.1089/ars.2012.4684. Epub 2012 Jun 12. PMID: 22563824; PMCID: PMC3513981 | sr |
