Serine-protease inhibition during islet isolation increases islet yield from human pancreases with prolonged ischemia

Background. Islet isolation from the pancreatic tissue matrix remains highl y variable. Recent evidence suggests that intrinsic human pancreatic protea ses, including trypsin, may inhibit effective collagenase enzymatic activit y during islet isolation, thereby impairing the isolation success. In this study we have hypothesized that serine protease inhibition applied during p ancreatic digestion, could improve yield and/or functional viability of isl ets isolated from human pancreases. Methods. Twelve organ donor pancreases with 12.9 +/-0.6 hr cold storage (me an SEM) were perfused via their ducts with Liberase-HI enzyme in the presen ce n=6 or absence (n=6) of 0.4 mM Pefabloc. All were then gently dissociate d and their purified islets separated with Ficoll density gradient centrifu gation. Results. Donor-related factors (age, gender, cold storage time, body mass i ndex, and pancreas weight) did not differ significantly between the two exp erimental groups. Pefabloc supplementation did not affect the digestion tim e, islets remaining trapped in exocrine tissue, or final islet purity. Isle t recovery was increased in the Pefabloc-treated group (mean SEM yield 323. 8 +/- 80.8x10(3) islet equivalents vs. 130.8 +/- 13.6x10(3) islet equivalen ts, P <0.05). Cellular composition, DNA and insulin content, and insulin se cretory activity of the isolated islets was similar. Conclusions. Inhibition of intrinsic protease activity within pancreases af ter prolonged cold storage improves isolation of viable islets.