Heparan sulfate, present on the equatorial segment of human spermatozoa, does not participate in in vitro nuclear sperm decondensation

GALOTTO C; CAMBIASSO MY; PIÑEIRO L; CALVO JC; ROMANATO M

Congreso; LXIII Reunión Anual de la Sociedad Argentina de Investigación Clìnica; 2018

Sociedad Argentina de Investigación Clínica (SAIC)

We have previously reported that Heparan Sulfate (HS) is present in the ooplasm and perivitelline space of human oocytes and participates in sperm decondensation in vivo. Flow cytometry experiments revealed that HS is also present on the surface of human spermatozoa. The aim of this study was to 1)determine the localization of HS on the surface of human spermatozoa and 2)evaluate whether this HS could be involved in sperm nuclear decondensation. 1)Spermatozoa were obtained from normozoospermic donors (WHO), swam up in HTF medium (W), incubated in capacitating conditions (overnight, 37°C, 5% CO2, C)and exposed to A23187 calcium ionophore (IAR). Immunocytochemistry was performed using a monoclonal anti HS antibody and Rhodamine labeled second antibody. Acrosomal status was determined with Pisum sativum lectin. 2) Spermnuclear decondensation was evaluated in vitro in the presence of murine oocytes with or without the addition of 20 mUI/mL heparinase III (Hase). 1) HS was present on 77±6% W, 62±10% C and 46±6 % IAR spermatozoa (n=3), localizing mainly (67±4% in W; 48±8% in C, and 38±11% in IAR) over the equatorial segment. Acrosomal labeling was observed in 10±4% sperm throughout. Double stainingwith Pisum sativum and anti-HS revealed that HS localization was not related to acrosomal status. 2) Sperm in vitro decondensation in murine oocytes was not affected by prior incubation of spermatozoa with Hase (62±3 % control vs 55±8 %, NS) treated but decreased significantly when Hase was added to oocytes (25±2 %, p=0.024, ANOVA+Tukey, n=3). Results demonstrate that HS localizes primarilyover the equatorial segment of human spermatozoa, regardless of physiologicalstatus and does not participate in nuclear sperm decondensation in vitro. Its possible involvement in sperm interaction with the epithelium of the femaletract is currently under investigation.