A study was conducted to determine the optimum dosage of the exogenous cholesterol‐loaded cyclodextrins (CLC) to get maximum cryoprotection for bubaline spermatozoa. In the present study, 120 × 10⁶ spermatozoa were incubated in 2, 3 and 4 mg of CLC as grouped as Gr II, III and IV, respectively, and sperm progressive motility, intracellular Ca²⁺, capacitation status by protein tyrosine phosphorylation (PTP) assay and zona binding per cent (ZBP) and cleavage rate (CR) of the cryopreserved buffalo spermatozoa by in vitro fertility assay were assessed in comparison with an untreated control group (Gr I). Results revealed that there was a significant (p  < .05) linear decrease in percentage of sperm population with higher intracellular Ca²⁺ and percentage of sperm population with medium or high capacitated by PTP in CLC treated from 2 to 3 mg and then increased to 4 mg/120 × 10⁶ spermatozoa whereas sperm progressive motility, percentage of sperm population with low capacitated, ZBP and CR were increased significantly (p  < .05) in sperm population treated from 2 to 3 mg CLC and then decreased to 4 mg/120 × 10⁶ spermatozoa. The study has clearly indicated that CLC at 3 mg/120 × 10⁶ spermatozoa has maximum beneficial effects in protection of sperm progressive motility, membrane fluidity (low intracellular Ca²⁺); prevention of cryocapacitation (low capacitation pattern in immunolocalization) and enhancement of in vitro ZBP and CR. Post‐thaw motility of the CLC‐treated sperm has shown positively significant (p  < .05) correlation with sperm population with low intracellular Ca²⁺, low capacitated sperm population, ZBP and CR, whereas it was negatively (p  < .05) correlated with sperm population with high intracellular Ca²⁺, medium or high capacitated sperm. The present study has revealed for the first time that incubation of spermatozoa with CLC of higher dose (>3 mg/120 × 10⁶ spermatozoa) had adverse effects on sperm cryopreservation, although incubation of sperm with 3 mg/120 million prior to processing had minimised the freezing–thawing‐associated damages in bubaline species.

中文翻译：

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外源胆固醇可防止冷冻样变化，膜流动性，并增强大卵泡生子精子的体外受精能力。

进行了一项研究，以确定外源胆固醇负载的环糊精（CLC）的最佳剂量，以最大程度地获得对bubaline精子的冷冻保护。在本研究中，将120×10 ⁶精子分别在2、3和4 mg CLC中按Gr II，III和IV组进行孵育，并通过蛋白质酪氨酸磷酸化作用使精子进行性运动，细胞内Ca ^2+的获能状态（与未经处理的对照组（Gr I）相比，通过体外受精试验评估了冷冻保存的水牛精子的PTP）分析，透明带结合百分比（ZBP）和裂解率（CR）。结果显示， 细胞内Ca ^2+含量较高的精子群体百分比显着线性下降（p <.05）CLC中PTP中度或高容量精子群的百分比从2到3 mg，然后增加到4 mg / 120×10 ⁶精子，而精子进行性运动，低容量，ZBP和CR的精子群百分比增加 从2到3 mg CLC处理的精子数量显着（p <.05），然后降至4 mg / 120×10 ⁶精子。研究清楚表明，CLC在3 mg / 120×10 ⁶精子中对保护精子进行性运动，膜流动性（细胞内Ca ²⁺低）具有最大的有益作用。）; 预防冻融（免疫定位中的低获能模式）并增强体外ZBP和CR。在CLC-处理的精子在融化后运动已经显示出积极显著（p 具有低细胞内Ca精子群体<0.05）相关²⁺，低能精子群体，ZBP和CR，而这是负（p  <0.05）与具有高细胞内Ca ²⁺，中等或高获能精子的精子数量相关。本研究首次揭示了高剂量（> 3 mg / 120×10 ⁶ 精子）对精子的冷冻保存有不利影响，尽管在加工前将精子与3 mg / 1.2亿一起孵育可最大程度地减少了古巴物种的冻融相关损害。