[Current knowledge on cryopreservation of spermatozoa, ovum cells and zygotes].

CELL INJURIES DURING FREEZING AND THAWING

The aim of various cryopreservation procedures is to minimize cell injuries during the freeze-thaw cycle (cryoinjuries). Generally, the cell damage during freezing and thawing procedures may be the results of: (a) extensive cellular dehydration (solution effect) and/or (b) intracellular ice crystallization/recrystallization (mechanical cell damage). Two independent mechanisms are involved. They can act simultaneously, leading to cytolysis. The first one is expressed primarily during low rate freezing, and the second one during rapid freezing. Thus, determination and use of the optimal cooling velocity, specific for each type of isolated cells, should be considered. Finally, a higher degree of cell destruction has been documented when the transition period from liquid to solid phase (release of the fusion heat) is prolonged. CRYOPROTECTIVE AGENTS: For successful cell cryopreservation, cryoprotectants are needed. They decrease the osmotic gradient and the vapor pressure difference between the intra- and extracellular area. Adequate choice of the most suitable type and concentration of cryoprotective agent is important for the required cell recovery after thawing. There are several well known protocols for obtaining cryopreservation of isolated cells using different cryoprotectants. Glycerol, dimethyl sulfoxide (DMSO) and propanediol sucrose are commonly used as cryoprotectants, though in different concentrations. Glycerol, a trihydric alcohol, is a clear, colorless fluid. Pharmacologically, it is relatively inert. DMSO is a colorless liquid with a sulphur-like smell and has several medical uses. It is highly polar and dissolves many water- and lipid-soluble substances. DMSO given intravenously may cause nausea, vomiting, local vasospasm and an objectionable garlic-like odor and taste. HUMAN SPERM, OVA AND EMBRYOS CRYOPRESERVATION: Despite the fact that cryopreservation procedures of spermatozoa, ova and embryos are already in routine clinical use, some questions related to the optimal cooling velocity during controlled-rate freezing and the choice of the most effective, either penetrating (glycerol, dimethyl sulfoxide) and/or non-penetrating (hydroxyethyl starch) cryoprotective agent at the appropriate concentration are not resolved.