PREVENTION OF OSMOTIC INJURY TO HUMAN SPERMATOZOA DURING ADDITION AND REMOVAL OF GLYCEROL

被引:255
作者
GAO, DY
LIU, J
LIU, C
MCGANN, LE
WATSON, PF
KLEINHANS, FW
MAZUR, P
CRITSER, ES
CRITSER, JK
机构
[1] METHODIST HOSP INDIANA INC,CRYOBIOL RES INST,INDIANAPOLIS,IN 46206
[2] INDIANA UNIV PURDUE UNIV,DEPT PHYS,INDIANAPOLIS,IN 46202
[3] UNIV ALBERTA,DEPT PATHOL,EDMONTON,AB T6G 2R8,CANADA
[4] UNIV LONDON ROYAL COLL VET SURG,DEPT VET BASIC SCI,LONDON NW1 0TU,ENGLAND
[5] OAK RIDGE NATL LAB,DIV BIOL,OAK RIDGE,TN 37831
[6] INDIANA UNIV,SCH MED,DEPT PHYSIOL & BIOPHYS,INDIANAPOLIS,IN 46202
[7] INDIANA UNIV,SCH MED,DEPT OBSTET & GYNECOL,INDIANAPOLIS,IN 46202
关键词
GLYCEROL; HUMAN SPERMATOZOA; OSMOTIC INJURY;
D O I
10.1093/oxfordjournals.humrep.a136103
中图分类号
R71 [妇产科学];
学科分类号
100211 ;
摘要
Use of a cryoprotective agent is indispensable to prevent injury to human spermatozoa during the cryopreservation process, However, addition of cryoprotective agents to spermatozoa before cooling and their removal after warming may create severe osmotic stress for the cells, resulting in injury. The objective of this study was to test the hypothesis that the degree (or magnitude) of human sperm volume excursion can be used as an independent indicator to evaluate and predict possible osmotic injury to spermatozoa during the addition and removal of cryo-protective agents. Glycerol was used as a model cryoprotective agent in the present study. To test this hypothesis, first the tolerance limits of spermatozoa to swelling in hypoosmotic solutions (iso-osmotic medium diluted with water) and to shrinkage in hyperosmotic solutions (iso-osmotic medium with sucrose) were determined. Sperm plasma membrane integrity was measured by fluorescent staining, and sperm motility was assessed by computer-assisted semen analysis before, during and after the anisosmotic exposure. The results indicate firstly that motility was much more sensitive to anisosmotic conditions than membrane integrity, and secondly that motility was substantially more sensitive to hypotonic than to hypertonic conditions. Based on the experimental data, osmotic injury as a function of sperm volume excursion (swelling or shrinking) was determined. The second step, using these sperm volume excursion limits and previously measured glycerol and water permeability coefficients of human spermatozoa, was to predict, by computer simulation, the cell osmotic injury caused by different procedures for the addition and removal of glycerol, The predicted sperm injury was confirmed by experiment, Based on this study, an analytical methodology has been developed for predicting optimal protocols to reduce osmotic injury associated with the addition and removal of hypertonic concentrations of glycerol in human spermatozoa.
引用
收藏
页码:1109 / 1122
页数:14
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