DIFFERING EFFECTS OF VARIOUS CRYOPROTECTANTS ON INTERCELLULAR-JUNCTIONS OF EPITHELIAL (MDCK) CELLS

The effects of 30 min exposure to 10% v/v cryoprotectant (glycerol, di methyl sulfoxide, or propanel-1,2-diol) on intercellular junctions wer e investigated in monolayers of epithelial (MDCK) cells. The cells wer e grown on microporous membrane filters and transepithelial resistance was monitored as an index of junction integrity. At 22 degrees C, Me( 2)SO had the least effect of the three cryoprotectants, causing a grad ual fall in transepithelial resistance of only 17% in 30 min. This com pares with falls of 50% in 15 min and 37% over 30 min, respectively, w ith glycerol and propane-1,2-diol. At 0 degrees C, however, propane-1, 2-diol had no effect on transepithelial resistance, whereas the effect of Me(2)SO was now similar to that of glycerol, viz. a sustained redu ction in resistance of approximately 30% throughout the 30-min exposur e. The results could not be explained solely in terms of osmotic effec ts and suggested that the cryoprotectants were directly affecting comp onents of the junctional complex or associated elements of the cytoske leton. During removal of the cryoprotectants by two-step dilution, cha nges in transepithelial resistance showed little consistency between t he different cryoprotectants. This suggested that the pattern of chang e was dictated more by the state of the junctions immediately before r emoval of the cryoprotectants, with each cryoprotectant having affecte d the junctions to different extents and perhaps by different mechanis ms, than by the removal of cryoprotectant itself. Thus, intercellular junctions in epithelial sheets were affected by even modest concentrat ions of cryoprotectant, but the effect was influenced by the type of c ryoprotectant and by temperature of exposure. The changes in transepit helial resistance were not reversed immediately following removal of t he cryoprotectants but they were reversed when the monolayers were ret urned to 37 degrees C. (C) 1995 Academic Press, Inc.