Effect of varicocele on chromatin condensation and DNA integrity of ejaculated spermatozoa using cytochemical tests

Abstract

Varicocele occurs in approximately 15% to 20% of the general male population and it is the most common cause of poor semen production and decreased semen quality. It has been demonstrated that patients with varicocele have a significantly higher DNA fragmentation index (DFI) and spermatozoa with nuclear anomalies than healthy fertile men. Therefore, the aim of this study was to evaluate sperm chromatin integrity in these patients. Sixty men referring to the andrology laboratory were categorised into three different groups: 20 infertile men with varicocele, 20 infertile men with abnormal semen parameters and 20 fertile men who had normal spermatogram were considered as control group. Semen analysis was performed according to WHO criteria. To evaluate sperm chromatin quality and DNA integrity, after fixation of sperm smears, aniline blue, toluidine blue, chromomycin A(3) and acridine orange staining were applied in three groups. The slides were analysed by light and fluorescent microscopy and to determine the percentage of mature or immature spermatozoa, 200 spermatozoa were counted in each slide. The results showed that the rates of aniline blue-reacted spermatozoa were significantly higher in infertile and varicocele patients than in the normal group (P < 0.001). In addition, with regard to chromomycin A(3), acridine orange and toluidine blue staining, there was a significant difference between the three groups (P < 0.001). The results showed that the varicocele samples contain a higher proportion of spermatozoa with abnormal DNA and immature chromatin than those from fertile men as well as infertile men without varicocele. Therefore, varicocele results in the production of spermatozoa with less condensed chromatin and this is one of the possible causes of infertility due to varicocele.

Sperm chromatin condensation, DNA integrity, and apoptosis in men with spinal cord injury

“…The spermatozoa with unstained or pale-blue nuclei were recorded as normal and those with dark-blue nuclei were regarded as abnormal ones. 11,22 Furthermore, 87.5% of patients with SCI showed higher AB-reacted spermatozoa than "cut-off" value (30%) (Fig. 1).…”

“…The percentage of green fluorescence spermatozoa (normal double-stranded DNA) and orange/red fluorescence spermatozoa (abnormally denatured DNA) per sample were calculated. 11,22,26 Only 29.2%…”

“…The slides were rinsed thrice in distilled water for 2 minutes and finally stained with 0.05% TB in 50% McIlvaine buffer ( pH 3.5) for 10 minutes at room temperature. 22,23 The chromatin quality of spermatozoa was determined according to metachromatic staining of sperm heads with the aid of light microscopy at ×100 magnification.…”

Sperm chromatin condensation in infertile men with varicocele before and after surgical repair

“…Fuse et al (19) reported significant increase of altered sperm chromatin structure and DNA damage in infertile men with varicocele compared with infertile men without varicocele or even fertile subjects. Talebi et al (20) showed that stained sperm head percentage in men with varicocele was significantly higher than in controls. Blumer et al (21) added that men with varicocele had a higher percentage of cells with DNA fragmentation and sperm with inactive mitochondria compared with healthy controls.…”

SEPT12 deficiency causes sperm nucleus damage and developmental arrest of preimplantation embryos

“…The experiment was performed according to standard protocols (6). These three assays unanimously showed increased DNA damage in the sperm isolated from Septin 12 þ/-chimeric mice, compared with that from the wild type mice.…”