Significant progress in porcine islet mass isolation utilizing liberase HIfor enzymatic low-temperature pancreas digestion

Background. Frequent success in human islet isolation is prevented by the l arge variability of scarce organ donors; this favors the future utilization of pigs as donors for clinical islet xenotransplantation. Porcine-specific difficulties of islet isolation are attributed to the intrinsic fragility of islets during pancreas digestion. Methods. To preserve islet integrity during efficient pancreas dissociation , porcine pancreata (n = 48) were distended after cold storage with cold Un iversity of Wisconsin solution containing Liberase HI and digested at 24-28 degrees C using digestion-filtration. Pancreata distended with University of Wisconsin solution containing well-proven crude collagenase and digested at 32-34 degrees C served as controls (n = 46). Monolayer Ficoll-diatrizoa te gradient purification was performed in a Cobe 2991. Results. Purified yield of islet equivalents per pancreas (mean +/- SEM) wa s almost doubled by Liberase HI compared with crude collagenase (526,480 +/ - 46,560 vs. 270,270 +/- 19,420; P < 0.0001) and also significantly increas ed comparing islet equivalents per gram of pancreas (4,210 +/- 320 vs. 2,64 0 +/- 245; P = 0.0004). Islet integrity was better preserved during Liberas e HI digestion compared with crude collagenase digestion as indicated by is olation index (2.1 +/- 0.1 vs. 1.4 +/- 0.1; P < 0.0001). Purity, viability, and in vitro function of islets did not differ between experimental groups . Preserved in vivo function of islets isolated by Liberase HI was demonstr ated after subcapsular transplantation into 16 diabetic nude rats. Conclusions. If the problems related to xenograft rejection and xenosis cou ld be solved, low-temperature digestion of porcine pancreata using Liberase HI could serve as an essential prerequisite for successful 1:1 xenotranspl antation of pig islets into type 1 diabetic human recipients.