Department of Reproduction and Artificial Insemination, Faculty of Veterinary Medicine, University of Adnan Menderes, 09016 Aydin, Turkey.
Anim Reprod Sci. 2010 Jul;120(1-4):166-72. doi: 10.1016/j.anireprosci.2010.02.014. Epub 2010 Feb 23.
The effects of cholesterol-loaded cyclodextrin (CLC) treatment on the osmotic tolerance and ability to undergo the acrosome reaction of rabbit spermatozoa, with an unusually high cholesterol/phospholipid ratio in plasma membranes, were examined in two successive experiments. In the first experiment, CLC-pretreated and untreated sperm cells were exposed for 15min to one of five fructose solutions, adjusted to 20, 80, 290, 500 or 1500mOsm/L. After the anisoosmotic challenge, the integrity of sperm membranes in the CLC-supplemented (at a dose level of 3mg/120x10(6)spermatozoa) and control groups was estimated by a modified hypoosmotic swelling test (HOST) associated with a supravital eosin staining test (HE-test). In the second part of the study, the influence of cholesterol supplementation on the acrosome reaction of sperm cells stimulated by either calcium ionophore A23187 (CI) or lysophosphatidylcholine (LPC) was evaluated. CLC pretreatment increased viable and live-HOST-responsive sperm rates (P<0.01) after incubation in anisoosmotic solutions varying from 80 to 1500mOsm/L. However, CLC supplementation did not influence the percentage of HOST-responsive sperm cells (P>0.05). A significant interaction was determined between CLC pretreatment and the level of osmotic pressure in maintaining the functional and physical integrities of sperm membranes undergoing osmotic challenges. Both CI and LPC successfully induced the acrosome reaction in rabbit spermatozoa (P<0.001). Compared with CI, LPC was more effective (P<0.0001). CLC pretreatment resulted in a significant reduction (P<0.01) in the percentage of acrosome reacted sperm cells irrespective of the inducing agent, either CI or LPC. In conclusion, CLC treatment enhanced the anisoosmotic tolerance of rabbit spermatozoa and reduced their ability to undergo the acrosome reaction after stimulation by CI or LPC.
本研究分两部分进行，旨在探讨胆固醇负载环糊精（CLC）处理对兔精子渗透耐受性和顶体反应能力的影响。兔精子质膜胆固醇/磷脂比值异常高。第一部分实验中，将 CLC 预处理和未经处理的精子细胞分别暴露于五种果糖溶液（渗透压分别为 20、80、290、500 和 1500mOsm/L）15min。非等渗刺激后，通过改良的低渗肿胀试验（HOST）联合伊红活细胞染色试验（HE 试验）评估 CLC 补充（剂量水平为 3mg/120x10(6)精子）和对照组精子质膜的完整性。第二部分实验中，评估胆固醇补充对钙离子载体 A23187（CI）或溶血磷脂酰胆碱（LPC）刺激的精子顶体反应的影响。CLC 预处理可提高渗透压 80-1500mOsm/L 范围内的精子活率和活 HOST 反应率（P<0.01）。然而，CLC 补充并不影响 HOST 反应性精子的比例（P>0.05）。CLC 预处理与渗透压水平之间存在显著的交互作用，可维持渗透压刺激下精子质膜的功能和物理完整性。CI 和 LPC 均可成功诱导兔精子发生顶体反应（P<0.001）。与 CI 相比，LPC 更为有效（P<0.0001）。无论诱导剂是 CI 还是 LPC，CLC 预处理均导致顶体反应的精子比例显著降低（P<0.01）。结论：CLC 处理可提高兔精子的非等渗耐受性，并降低其在 CI 或 LPC 刺激后发生顶体反应的能力。
