RESPONSE OF EPITHELIAL (MDCK) CELL-JUNCTIONS TO CALCIUM REMOVAL AND OSMOTIC-STRESS IS INFLUENCED BY TEMPERATURE

Intercellular junctions could be targets of injury during both hypothe rmic storage and cryopreservation. Monolayers of Madin-Darby canine ki dney (MDCK) cells, an epithelial cell line, were grown on microporous membrane filters, and junction integrity was monitored by transepithel ial electrical resistance (TER). At 37 degrees C, TER was 196 ohm.cm(2 ) (SD 58, n = 59): this rose with falling temperature to 278 (SD 54) a nd 409 (SD 110) ohm.cm(2), respectively, at 22 and 0 degrees C. This i nitial increase was accounted for solely by the change in conductivity of the bathing medium, which declined with falling temperature. Prolo nged exposure to reduced temperature, however, did cause a gradual dec line in TER. Removal of calcium at 22 degrees C caused a rapid fall in TER owing to dissociation of calcium-dependent components of the junc tional complex. This was followed by a gradual increase in TER over se veral hours when calcium was restored to the bathing medium. At 0 degr ees C, TER declined slowly regardless of the presence or absence of ca lcium, which suggested that reduced temperature stabilized junctions w ith respect to their calcium dependence, but that low temperature itse lf was causing dissociation of junctions. Hypertonic stress (597 mOsmo l/kg) caused a reduction in TER both at 22 and 0 degrees C, which, unl ike the reduction caused by calcium removal at 22 degrees C, was rever sed relatively rapidly on return to isotonic conditions. Hypotonic str ess (201 mOsmol/kg) increased TER at 0 degrees C, but had no effect at 22 degrees C. The lack of response at the higher temperature might ha ve been the result of a cell volume regulatory mechanism. This techniq ue provides a straightforward method for investigating the effects of some of the stresses encountered during hypothermic storage and cryopr eservation on intercellular junctions. (C) 1994 Academic Press, Inc.