Prolonged Cryopreservation of Purified Human Pancreatic Islets

Abstract

Long-term islet storage would facilitate many aspects of islet research and clinical islet transplantation. Collagenase-isolated, Ficoll-purified islets from eight cadaveric pancreases were stored in liquid nitrogen for 44 +/- 9 days after dimethyl sulfoxide equilibration and slow cooling. Rapid rewarming and 48 h of culture preceded repeat evaluation of recovery by islet counts, insulin extraction, and glucose-stimulated perifusion. Islet recovery was 94 +/- 4% by count and 90 +/- 22% by insulin extraction immediately after thawing. After an additional 48 h in culture, recovery was 74 +/- 12% by insulin extraction and 79% by quantitative perifusion culture. Perifusion demonstrated normal baseline and first-phase insulin secretion with decreased second-phase insulin secretion after cryopreservation. Insulin-stained sections and electron microscopy revealed preserved islet morphology and ultrastructure. Granulated islets with preserved morphology were recovered 14 days after renal subcapsular xenografting into nude mice. This study demonstrates high recovery and good functional activity of human islets after prolonged cryopreservation.

Cloning and functional expression of a human pancreatic islet glucose-transporter cDNA.

“…Pancreatic islets obtained from cadaver donors maintain viability even after prolonged cold time of ischemia and the isolation process (25); condition of the RNA that could be obtained from these islets was not known, however. Furthermore, the donors had suffered from hours to weeks of either shock or chronic disease or both before death.…”

Effect of thawing rate and post-thaw culture on the cryopreserved fetal rat islets: Functional and morphological correlation

“…Insulin and C-peptide release from slow-thawed islets was lower than that of the control islets, and were nearly similar to that of the control in fast-thawed islets. Whereas extensive work has documented the effect of fast thawing (Sander et al, 1986;Simonsen et al, 1987;Dawidson et al, 1988;Kneteman et al, 1989;Beattie et al, 1993;Liu et al, 1993;Andereggen et al, 1996;GonzalezClemente et al, 1997), few reports could be located in the literature comparing the effect of slow and fast thawing temperature on the cryopreserved islets (Taylor and Benton, 1987;Tze and Tai, 1990;Rich et al, 1993). Taylor and Benton (1987) found by physiological studies that survival of the isolated rat islets was greater than 50% with fast warming rate, compared to slow warming.…”

“…The value of tissue culture before cryopreservation or after thawing has been suggested by several authors (McKay and Karow, 1983;Sandler and Andersson, 1987;Foreman and Taylor, 1989;Vasir et al, 1989;Kneteman et al, 1989;Foreman et al, 1993;Lakey et al, 1997;Duvivier et al, 1999;Gatto et al, 2003). McKay and Karow (1983) have found that islet function can be significantly improved by increasing the duration of post-thaw culture.…”

“…Vasir et al (1989) showed that the best results could be obtained when cryopreservation of adult inbred Lewis rat islets was followed by rapid thawing to 37 -C and then a 24 h culture. Kneteman et al (1989) found that insulin release from the cryopreserved human islets could be improved from 79% to 87% with postthaw 48 h culture period. Foreman et al (1993) found that cryopreserved rat islets required 72 h of culture for a survival rate of more than 50%.…”

Human Islet Allotransplantation: World Experience and Current Status

“…Further, cryopreservation can facilitate clinical islet transplantation by providing a practical means for maintaining a tissue bank of islets of various ABO groups. Since 1972, when it appeared that rat islets could be successfully cryopreserved, there have been many reports on freezing and thawing of islets [47, 48, 49, 50]. It is difficult to establish an optimum procedure for islet cryopreservation because different protocols have been used.…”