Incidence of sperm-tail tyrosine phosphorylation and hyperactivated motility in normozoospermic and asthenozoospermic human sperm samples

Our objective was to study the incidence of sperm-tail phosphotyrosine immunoreactivity in normozoospermic and asthenozoospermic human sperm samples, its association with sperm motion parameters, particularly hyperactivated motility, and its potential involvement in the pathogenesis of asthenozoospermia. The work was conducted as a prospective experimental study in the Sperm Biology and Andrology laboratories of the Jones Institute, a medical school-based fertility center. The study subjects were healthy fertile male donors (normozoospermic samples) and infertile patients (asthenozoospermic samples) attending the center. Recently ejaculated semen samples were washed twice to eliminate seminal plasma and a swim-up was performed to select the motile population which, in turn, was incubated up to 18 h at 37 degrees C in 3.5% human serum albumin-supplemented Ham's F10 to allow for capacitation. For evaluation, sperm aliquots were taken pre-swim-up (T0), immediately post swim-up (T1), at 6 h (T6), and 18 h (T18) of incubation. The main outcome measures were computer-analyzed sperm motion parameters and hyperactivated motility, and immunodetection of phosphotyrosine (PY)-containing proteins. During the capacitating incubation, normozoospermic samples displayed maximum motility, velocity, and hyperactivation at T6, significantly decreasing their values at T18. PY-proteins were located both at the tail and head of spermatozoa. Their expression increased progressively during the incubation, being present in about 70% of the sperm tails at T18. Asthenozoospermic samples showed an inability to respond to capacitation with an increase in motion parameters and PY-phosphorylation. At T6, both hyperactivation and PY-phosphorylation were significantly lower than in normal samples. Our results suggest that PY-phosphorylation of tail proteins is highly conspicuous in human spermatozoa, and increases its incidence in a time-dependent manner, as more sperm become capacitated. Asthenozoospermic samples displaying low percentages of motile sperm and altered motion characteristics showed a decreased incidence of PY-phosphorelated sperm. Tail protein PY-phosphorylation may be related to sperm movement, especially to hyperactivated motility and its deficiency may be associated to asthenozoospermia.

Golden hamster perivitelline spermatozoa do not show proacrosin/acrosin at the inner acrosomal membrane

It has been suggested that acrosin may function in penetration of the zona pellucida and of the highly structured extracellular matrix of the perivitelline space. In this study we investigated whether golden hamster perivitelline spermatozoa contain proacrosin/acrosin, as evidenced by the silver enhanced immunogold technique using the monoclonal antibody antiacrosin C2E5. None of the 197 spermatozoa recovered from the perivitelline space showed proacrosin/acrosin associated with the acrosomal region, suggesting that acrosin would not play a role in the penetration of the perivitelline extracellular matrix

Demonstration of anti-bradykinin compounds in callus cultures of Mandevilla velutina

Calluses from stems and leaves of Mandevilla velutina were grown in culture for 30,45 and 60 days. Thin-layer chromatography of ethyl acetate extracts of calluses from stems of M. velutina revealed the presence of 6 compounds. Two of them had RF values identical to those of the anti-bradykinin (BK) compounds MV8609 and MV8610 previously isolated from the natural plant. The ethyl acetate extract (20 to 80 microng/ml) from stem callus culture antagonized in a concentration-dependent and reversible manner contractions caused by BK (0.1-100 nM) in the isolated rat uterus. The extracts obtained from calluses cultured for 30,45 and 60 days were about 79-, 63-and 31-fold more potent, respectively, in antagonizing BK than the crude extract obtained from the rhizome of the plant

Blockade of the bradykinin-evoked diphasic response of isolated rat duodenum by the crude extract of and compounds obtained from Mandevilla velutina

The influence of an aqueous/ethanolic extract of M. velutina (CE) and of two compound (MV 8608 and MV 8612) purified from the plant on BK-, Ad- and K+ -induced responses of the rat duodenum was analyzed in vitro. In preparations precontracted with Ca2+, the CE (1 and 2 mg/ml) markedly inhibited BK -induced relaxation in a dose-dependent manner but was less effective against Ad-induced relaxation. The CE (0.5 mg/ml) also antagonized BK -induced relaxation and contraction of strips bathed in normal medium. The two compounds from M. velutina (10 and 20 microng/ml) also promoted a dose-dependent blockade of both responses to BK, only slightly depressing the response to K+