Leboeuf B, Le Vern Y, Furstoss V, Kerboeuf D, Guillouet P, Magistrini M
Unité Expérimentale d'Insémination Artificielle Caprine et Porcine, INRA, Centre Poitou-Charentes, 86480 Rouillé, France.
Anim Reprod Sci. 2006 Feb;91(3-4):265-74. doi: 10.1016/j.anireprosci.2005.04.014. Epub 2005 Jun 27.
Hypoosmotic swelling test (HOS) has been proposed by many authors to evaluate the functional integrity of the sperm membrane. Our approach in this experiment has consisted in exposing spermatozoa to a wide range of osmotic pressures then evaluating the reacted sperm cells by flow cytometry and finally modelling the sperm cell responses. Semen samples were diluted in skim milk or NPPC (native phosphocaseinate) extenders, and stored at 4 degrees C for 3 days. At D0 and D3 aliquots from each ejaculate (n=12) were submitted to seven hypoosmotic solutions varying from 230 to 10mOsm/kg. Sperm samples were analyzed using flow cytometry to determine two populations of spermatozoa identified by propidium iodide (PI): PI+ (including PI, red fluorescence) and PI- (excluding PI, no fluorescence). Spermatozoa PI+ were considered as spermatozoa with membrane damages. PI+ exhibited a high variation from 230 to 10mOsm/kg which was considered as a dose-response curve. Data were modelled using Mixed procedure and probit analysis to a sigmoid curve. Each model curve characterized the profile of response of the variable PI+ to the range of osmotic pressure from 230 to 10mOsm/kg. The estimated parameters modelling the sigmoid curves are discussed in order to evaluate the effect of extender (skim milk versus NPPC) and duration of preservation (D0 versus D3). Such modelling could help to differentiate storage method ejaculates within males or between male, contributing therefore to improve semen technology.
许多作者提出了低渗肿胀试验（HOS）来评估精子膜的功能完整性。我们在本实验中的方法包括将精子暴露于广泛的渗透压范围内，然后通过流式细胞术评估反应后的精子细胞，最后对精子细胞反应进行建模。精液样本用脱脂牛奶或NPPC（天然磷酸酪蛋白）稀释剂稀释，并在4℃下储存3天。在第0天和第3天，从每个射精样本（n = 12）中取出的等分试样被置于七种渗透压从230到10mOsm/kg不等的低渗溶液中。使用流式细胞术分析精子样本，以确定由碘化丙啶（PI）识别的两个精子群体：PI +（包括PI，红色荧光）和PI -（不包括PI，无荧光）。PI +精子被认为是膜受损的精子。PI +在230到10mOsm/kg之间表现出很大的变化，这被视为剂量反应曲线。数据使用混合程序和概率分析建模为S形曲线。每条模型曲线表征了变量PI +对230到10mOsm/kg渗透压范围的反应概况。讨论了对S形曲线进行建模的估计参数，以评估稀释剂（脱脂牛奶与NPPC）和保存时间（第0天与第3天）的影响。这种建模有助于区分雄性个体内或雄性个体间的精液保存方法，从而有助于改进精液技术。
