Paudel K P, Kumar S, Meur S K, Kumaresan A
Animal Health Research Division, Nepal Agricultural Research Division, Khumaltar, Lalitpur, Kathmandu, Nepal.
Reprod Domest Anim. 2010 Apr;45(2):256-62. doi: 10.1111/j.1439-0531.2008.01278.x. Epub 2008 Oct 30.
The present study evaluated the effectiveness of ascorbic acid, catalase, chlorpromazine and their combinations in reducing the cryodamages to crossbred bull (Bos taurus x Bos indicus) spermatozoa. A total of 32 ejaculates (eight each from four bulls) were diluted in Tris-citric acid-fructose-egg yolk-glycerol extender. Each ejaculate was split into six parts (five treatment and one control). Treatment groups included 10 mm ascorbic acid, 0.1 mm chlorpromazine, 200 IU/ml catalase, 10 mm ascorbic acid + 0.1 mm chlorpromazine or 200 IU/ml catalase + 0.1 mm chlorpromazine in the extender. Fluorescent probes (Fluorescein isothiocyanate--Pisum sativum agglutinin + Propidium iodide) were used for the assessment of spermatozoa viability and acrosomal status. The proportion of acrosome intact live (AIL), acrosome intact dead, acrosome reacted live and acrosome reacted dead sperm was assessed in fresh, equilibrated and frozen-thawed semen. The functional status of the sperm was assessed using hypo-osmotic sperm swelling test (HOSST). Activities of acrosin and hyaluronidase enzyme were also determined. Lipid peroxidation level was assayed based on the melonaldehyde (MDA) production. In cryopreserved semen, the values of AIL spermatozoa, HOSST response, hyaluronidase and acrosin activity were reduced by 53%, 47%, 34% and 54%, respectively from their initial values in fresh semen. However, MDA level was threefold higher in the frozen-thawed sperm compared with fresh sperm. Significant (p < 0.05) improvement in motility, viability, HOSST response, retention of hyaluonidase and acrosin and reduction in MDA was recorded in ascorbic acid, catalase, ascorbic acid + chlorpromazine and catalase + chlorpromazine incorporated groups. The percentage of AIL sperm was significantly (p < 0.05) higher in ascorbic acid, catalase and ascorbic acid + chlorpromazine incorporated groups compared with the control. Chlorpromazine alone did not improve the post-thaw semen quality but when combined with either ascorbic acid or catalase, improvement in semen quality was noticed. It was inferred that incorporation of ascorbic acid, catalase and ascorbic acid + chlorpromazine in semen extender improved the post-thaw semen quality in crossbred bulls.
本研究评估了抗坏血酸、过氧化氢酶、氯丙嗪及其组合在减轻对杂交公牛（瘤牛×印度野牛）精子的冷冻损伤方面的效果。总共32份射精样本（来自四头公牛，每头公牛各8份）用Tris - 柠檬酸 - 果糖 - 蛋黄 - 甘油稀释液进行稀释。每份射精样本被分成六份（五份处理样本和一份对照样本）。处理组在稀释液中分别添加10 mM抗坏血酸、0.1 mM氯丙嗪、200 IU/ml过氧化氢酶、10 mM抗坏血酸 + 0.1 mM氯丙嗪或200 IU/ml过氧化氢酶 + 0.1 mM氯丙嗪。使用荧光探针（异硫氰酸荧光素 - 豌豆凝集素 + 碘化丙啶）评估精子活力和顶体状态。在新鲜、平衡和冻融后的精液中评估顶体完整活精子（AIL）、顶体完整死精子、顶体反应活精子和顶体反应死精子的比例。使用低渗精子肿胀试验（HOSST）评估精子的功能状态。还测定了顶体蛋白酶和透明质酸酶的活性。基于丙二醛（MDA）的产生来测定脂质过氧化水平。在冷冻保存的精液中，AIL精子的值、HOSST反应、透明质酸酶和顶体蛋白酶活性分别比新鲜精液中的初始值降低了53%、47%、34%和54%。然而，冻融精子中的MDA水平比新鲜精子高三倍。在添加抗坏血酸、过氧化氢酶、抗坏血酸 + 氯丙嗪和过氧化氢酶 + 氯丙嗪的组中，精子活力、存活率、HOSST反应、透明质酸酶和顶体蛋白酶的保留率均有显著（p < 0.05）改善，MDA水平降低。与对照组相比，添加抗坏血酸、过氧化氢酶和抗坏血酸 + 氯丙嗪的组中AIL精子的百分比显著（p < 0.05）更高。单独使用氯丙嗪并不能改善解冻后精液质量，但与抗坏血酸或过氧化氢酶联合使用时，精液质量有改善。据推断，在精液稀释液中添加抗坏血酸、过氧化氢酶和抗坏血酸 + 氯丙嗪可改善杂交公牛解冻后精液质量。
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