Chemical N‐desulfation of heparin negates its ability to capacitate bovine spermatozoa

Abstract

Heparin specifically and saturably binds to bovine spermatozoa and stimulates capacitation as assessed by the ability of spermatozoa to undergo a zona pellucida-induced acrosome reaction (AR) in vitro. However, the structural features of heparin important for capacitation are poorly understood. The purpose of this study was to determine the importance of the sulfate content of heparin for its potency to bind to bull spermatozoa and promote agglutination and capacitation. The pyridine salt of heparin was N-desulfated, which reduced its mean sulfate content from 19.7% to 11.6%. The N-desulfated heparin was then resulfated by incubation with trimethylamine sulfur trioxide for 6, 12, or 24 hr, raising sulfate to original concentrations. Heparin but not N-desulfated heparin competed with [3H]-heparin to bind to spermatozoa. Heparin at 11.6 micrograms/ml reduced [3H]-heparin binding by half when competing with a saturating concentration of the radiolabeled compound (12 micrograms/ml). N-desulfated heparin did not displace [3H]-heparin. Heparin, resulfated 6 hr or 12 hr, was equal to native heparin in binding potency. Heparin at 50, 100, or 250 micrograms/ml caused more than 40% of the cells to head-to-head agglutinate in aggregates of 8 or more. N-desulfated heparin did not cause agglutination. After spermatozoa were incubated with 0, 5, 10, 50, 100, or 250 micrograms/ml of heparin for 4 hr, 100 micrograms/ml of lysophosphatidylcholine (LPC)-induced AR within 20 min in 21.3, 37.7, 27.8, 45.3, 54.2, or 42.5% of the cells, respectively. Sperm exposed to the same concentrations of N-desulfated heparin exhibited AR of 17.7, 27.3, 24.3, 22.5, 27.7, or 33.8%, respectively, following exposure to LPC. Resulfated heparin did not agglutinate or capacitate spermatozoa. In conclusion, N-desulfation of heparin abolished heparin's ability to bind to, agglutinate, and capacitate bovine spermatozoa. Resulfation of N-desulfated heparin restored binding activity but not agglutination or capacitation activity.

Relationship between fertilizing ability of frozen human spermatozoa and capacity for heparin binding and nuclear decondensation

“…We conclude that certain motile spermatozoa that are resistant to nuclear decondensation induced by heparin or heparin-GSH lose their capacity to bind the oocyte. Although motility of spermato¬ zoa was inversely correlated with ability to bind heparin, heparin could modify the number of spermatozoa involved in the fertilization process by causing head-to-head sperm agglutination (Miller & Ax, 1989) or by decreasing capacity of spermatozoa to recognize the oocyte membrane after alterations in surface antigens as observed in bulls by Miller & Hunter (1986). We did not observe head-to-head sperm agglutination after treatments with heparin-GSH or heparin alone.…”

Influence of Semen‐collection Frequency on Sperm Quality in Boars, with Special Reference to Biochemical Markers

“…Heparin belongs to the glycosaminoglycane group (GAGS), present both in the seminal plasma and in the follicular fluid. The physiological function of heparin is connected with the stimulation of the capacitation process and acrosome reaction of sperm, with participation of specific plasmatic proteins, the so-called heparin-binding proteins (Miller and Ax 1989). These proteins, belonging to the 'sperrnadhesin' family, possess specific receptors to the plasmolemma of bull and boar spermatozoa (Ness et al 1990;Sanz et al 1993).…”

Phosphatidylcholine-binding proteins of bovine seminal plasma modulate capacitation of spermatozoa by heparin

“…Interestingly, the BSP proteins also bind to heparin, a glycosaminoglycan [20]. In the bovine species, heparin and other glycosaminoglycans (GAGs) present in the female reproductive tract appear to participate in the process of capacitation [21][22][23][24][25][26][27]. The BSP proteins also bind to the surface of spermatozoa upon ejaculation [4,28].…”