Andrology: Changes in chromatin condensation of human spermatozoa during epididymal transit as determined by flow cytometry

Abstract

Inasmuch as caput epididymal and even testicular spermatozoa are now being used to generate pregnancies by direct injection into the oocyte, differences in the chromatin of spermatozoa from proximal and distal locations in the epididymis were studied. Acridine Orange staining was used to investigate chromatin structure in human spermatozoa which had left the testis and were undergoing maturation in the epididymis. Measurement of green and red fluorescence intensities of human spermatozoa by flow cytometry demonstrated a decrease in binding of Acridine Orange to DNA as the spermatozoa traversed the epididymis. Using spermatozoa from the cauda epididymis as the standard, the percentages of spermatozoa from the efferent duct, proximal corpus epididymis, midcorpus epididymis, distal corpus epididymis, proximal cauda epididymis and distal cauda epididymis that had matured with regard to chromatin condensation were 28 +/- 5, 39 +/- 3, 49 +/- 5, 64 +/- 5, 69 +/- 6 and 74 +/- 4% respectively. It may be concluded that eggs fertilized by ejaculated spermatozoa receive a more highly condensed form of chromatin than that received by eggs inseminated with proximal epididymal or testicular spermatozoa.

Assessment of sperm chromatin condensation and ploidy status using flow cytometry correlates to fertilization, embryo quality and pregnancy following in vitro fertilization

“…AO staining followed by SFC [25] was performed to estimate the percentage of sperm cells with complete epididymal maturation and totally condensed nuclear chromatin. The progressive chromatin packaging produces a reduction in DNA stainability relative to round spermatids and a marked decrease in AO binding to DNA as human spermatozoa traverse the epididymis [8]. Lack of appropriate sperm maturation results in an increased DNA stainability [28], frequently observed in many ejaculated spermatozoa from infertile men [6].…”

“…Chromatin condensation and stability have been proposed as reliable markers of sperm maturity [7]. Indeed, the chromatin of spermatozoa from the distal cauda epididymis is more condensed than that from spermatozoa of the proximal sites of epididymis [8].…”

Protection post-testiculaire des gamètes mâles contre les dommages radicalaires

“…En effet, comme nous l'avons vu ci-dessus, les spermatozoïdes sont très sensibles aux attaques oxydantes et utilisent H 2 O 2 et/ou des dérivés lipidiques hydroperoxydés (LOOH) pour parfaire leur maturation. Plusieurs protéines spermatiques membranaires mais aussi internes au gamète sont soumises à une intense activité de pontage disulfure pendant la descente épididymaire [12][13][14][15][16][17][18][19]. L'évolution du rapport thiols libres/disulfides des protéines spermatiques en tête (10/1) et en queue d'épididyme (1/10) illustre ce phénomène.…”

Post-Meiotic DNA Damage and Response in Male Germ Cells

“…These modifications are known to modulate the affinity of histones for DNA, but also the affinity of other proteins for histones, such as chromatin remodelers, DNA repair proteins or the transcription machinery. After spermiation, this unique protamine-based chromatin is further stabilized by the creation of intraprotamine disulfide bonds during the transit through the epididymis (Golan et al, 1996). Therefore, protamination provides both chemical and mechanical protection to the male haploid genome.…”