DIFFERENTIAL FREEZING TOLERANCE OF RAT PANCREATIC-ISLETS DEPENDING ONTHEIR SIZE VARIATION

We have investigated the freezing tolerance of rat pancreatic islets. Freshly isolated rat pancreatic islets were divided into three groups based on their longest diameter (small; 100-200 mu m, medium; 201-300 mu m, large; >300 mu m. They were then cryopreserved at a slow cooling rate (-0.3 degrees C/min) in the presence of dimethyl sulfoxide (Me(2 )SO) or ethylene glycol (EG). After storage at -196 degrees C for 1-4 weeks, they were thawed and their ability to secrete insulin in respon se to fluctuations in glucose concentration was examined during three consecutive static incubations in vitro (1st; 2.8 mM, 2nd; 16.7 mM, 3r d; 2.8 mM). Morphological examination of the beta-granule population w as determined by image analysis, and correlation with islets size was analyzed. The amount of insulin released from large-sized islets was s ignificantly suppressed in EG (p<0.05) and Me(2)SO (p<0.01) groups com pared to unfrozen islets. However, the mean volume of the large-sized islets isolated from one rat accounted for 43.0% of the total volume. On the other hand, the amount of insulin released from small- and medi um-sized islets did not differ from those of unfrozen islets, and thei r mean volumes were 13.2 and 43.8%, respectively. The percentage of ce lls with beta-granules was significantly correlated with size in both EG (r= -0.52) and Me(2)SO (r= -0.35) groups, but no significant correl ation was observed in the unfrozen islets groups. These findings sugge st that large-sized islets are more susceptible to freezing injury tha n small- or medium-sized islets. Moreover, the volume distribution of isolated islets indicated that it may be important to retain the abili ty of insulin secretion from the large-sized islets.