Fernández Silvina, Córdoba Mariana
Universidad de Buenos Aires, Facultad de Ciencias Veterinarias, Instituto de Investigación y Tecnología en Reproducción Animal (INITRA), Unidad Ejecutora de Investigaciones en Producción Animal UBA-CONICET (INPA), Cátedra de Química Biológica, Av. Chorroarín 280(1427), Ciudad Autónoma de Buenos Aires, Argentina.
Universidad de Buenos Aires, Facultad de Ciencias Veterinarias, Instituto de Investigación y Tecnología en Reproducción Animal (INITRA), Unidad Ejecutora de Investigaciones en Producción Animal UBA-CONICET (INPA), Cátedra de Química Biológica, Av. Chorroarín 280(1427), Ciudad Autónoma de Buenos Aires, Argentina.
Anim Reprod Sci. 2017 Apr;179:80-87. doi: 10.1016/j.anireprosci.2017.02.004. Epub 2017 Feb 20.
Hyaluronic acid, as well as heparin, is a glycosaminoglycan present in the female genital tract of cattle. The aim of this study was to evaluate oxidative metabolism and intracellular signals mediated by a membrane-associated adenylate cyclase (mAC), in sperm capacitation with hyaluronic acid and heparin, in cryopreserved bull sperm. The mAC inhibitor, 2',5'-dideoxyadenosine, was used in the present study. Lactate dehydrogenase (LDH) and creatine kinase (CK) activities and lactate concentration were determined spectrophotometrically in the incubation medium. Capacitation and acrosome reaction were evaluated by chlortetracycline technique, while plasma membrane and acrosome integrity were determined by trypan blue stain/differential interference contrast microscopy. Heparin capacitated samples had a significant decrease in LDH and CK activities, while in hyaluronic acid capacitated samples LDH and CK activities both increased compared to control samples, in heparin and hyaluronic acid capacitation conditions, respectively. A significant increase in lactate concentration in the incubation medium occurred in hyaluronic acid-treated sperm samples compared to heparin treatment, indicating this energetic metabolite is produced during capacitation. The LDH and CK enzyme activities and lactate concentrations in the incubation medium were decreased with 2',5'-dideoxyadenosine treatment in hyaluronic acid samples. The mAC inhibitor significantly inhibited heparin-induced capacitation of sperm cells, but did not completely inhibit hyaluronic acid capacitation. Therefore, hyaluronic acid and heparin are physiological glycosaminoglycans capable of inducing in vitro capacitation in cryopreserved bull sperm, stimulating different enzymatic pathways and intracellular signals modulated by a mAC. Hyaluronic acid induces sperm capacitation involving LDH and CK activities, thereby reducing oxidative metabolism, and this process is mediated by mAC.
透明质酸以及肝素是存在于母牛生殖道中的一种糖胺聚糖。本研究的目的是评估在冷冻保存的公牛精子中，透明质酸和肝素诱导精子获能过程中，由膜相关腺苷酸环化酶（mAC）介导的氧化代谢和细胞内信号。本研究使用了mAC抑制剂2',5'-二脱氧腺苷。通过分光光度法测定孵育培养基中的乳酸脱氢酶（LDH）和肌酸激酶（CK）活性以及乳酸浓度。采用金霉素技术评估精子获能和顶体反应，同时通过台盼蓝染色/微分干涉对比显微镜测定质膜和顶体的完整性。肝素诱导获能的样本中LDH和CK活性显著降低，而在透明质酸诱导获能的样本中，与对照样本相比，LDH和CK活性均增加，分别处于肝素和透明质酸诱导获能的条件下。与肝素处理相比，透明质酸处理的精子样本孵育培养基中的乳酸浓度显著增加，表明这种能量代谢产物是在获能过程中产生的。在透明质酸样本中，2',5'-二脱氧腺苷处理降低了孵育培养基中的LDH和CK酶活性以及乳酸浓度。mAC抑制剂显著抑制了肝素诱导的精子细胞获能，但并未完全抑制透明质酸诱导的获能。因此，透明质酸和肝素是能够诱导冷冻保存的公牛精子体外获能的生理性糖胺聚糖，刺激由mAC调节的不同酶途径和细胞内信号。透明质酸通过涉及LDH和CK活性诱导精子获能，从而降低氧化代谢，并且这个过程由mAC介导。
