Fluorometric Viability Assessment of Capacitated and Acrosome-Reacted Boar Spermatozoa by Flow Cytometry
荧光容量受限和顶体反应的精子流式细胞仪的可行性评估作者机构：Departamento de Ciencias de la SaludUniversidad Autonoma Metropolitana-IztapalapaMexico CityMexico Laboratoire d’HistologieEmbryologie et Microscopie ElectroniqueUniversite LorraineNancyFrance
出 版 物：《Optics and Photonics Journal》 (光学与光子学期刊（英文）)
年 卷 期：2013年第3卷第1期
页 面：40-44页
学科分类：1002[医学-临床医学] 100214[医学-肿瘤学] 10[医学]
基 金：partially supported by the CONACYT(Mexico)grant 0105961/10110/194/09
主 题：Boar Sperm Flow Cytometry Propidium Iodide Rhodamine 123 Viability
摘 要：Sperm capacitation involves functional changes, such as the removal or appearance of specific molecules and changes in the plasma membrane;the acrosome reaction (AR) is an exocytotic event induced by calcium influx, enabling the spermatozoa to penetrate the zona pellucida. These processes can be achieved only if the spermatozoa have good viability;indeed, determination of sperm viability is used for the assessment of semen quality. Membrane integrity and mitochondrial activity are important viability parameters of spermatozoa and fluorescent techniques based on membrane permeability to dyes have been developed to determine these parameters. The aim of this work was to determine the viability of boar sperm (fresh, one hour of capacitation induction and 20 min of AR induction) by flow cytometry using propidium iodide (PI) (1.25 μg/mL) and rhodamine 123 (R123) (0.20 μg/mL). Aliquots of 5 × 105 sperm were incubated with each fluorochrome separately and simultaneously for 10 or 20 min, respectively, at 38℃. The proportion of labeled spermatozoa and their fluorescence intensities were measured using a flow cytometer. The fluorescence index (FI) with PI gradually increased during the incubation and we found significant differences between all the groups. With R123, the FI increased in the capacitated sperm but decreased in the acrosome-reacted sperm, with significant differences between the fresh and capacitated spermatozoa. Our results suggest that the increase in the R123 fluorescence intensity in capacitated spermatozoa is due to changes in the mitochondrial membrane activity because the spermatozoa experienced changes in membrane fluidity and flagellar activation during capacitation. The use of fluorochromes and flow cytometry is a good tool for monitoring many markers of sperm function. Although capacitation and AR processes have been well studied, there is still much information to be elucidated with regard to these complex processes.
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