Excellent Islet Yields after 18-h Porcine Pancreas Preservation by Ductal Injection, Pancreas Preservation with MK Solution, Bottle Purification, and Islet Purification Using Iodixanol with UW Solution and Iodixanol with MK Solution

Abstract

Successful islet isolation is the key to successful islet transplantation. Our group recently modified the islet isolation protocol to include pancreatic ductal injection of the preservation solution, pancreas storage in modified extracellular-type trehalose-containing Kyoto (MK) solution, and use of an iodixanol-based purification solution and bottle purification. In this study, we applied these methods to porcine islet isolation after 18-h pancreas preservation and compared two solutions with different compositions in bottle purification. Islet yield before purification was 651,661 ± 157,719 islet equivalents (IE) and 5576 ± 1538 IE/g pancreas weight. An IU solution was made by adding iodixanol to University of Wisconsin solution and an IK solution was made by adding iodixanol to MK solution. The efficacy of the two solutions for islet isolation was compared. There were no significant differences between the two purification methods with regard to islet yield, survival rate, purity, score, or stimulation index. These results indicate that our isolation protocol produces efficient islet yields from prolonged cold-stored pancreas and that IU and IK solutions are equally useful for islet purification.

Pancreas preservation in extracellular‐type p38 inhibitor‐containing solution improves islet yield for porcine islet isolation

“…A gradient was generated using a gradient marker (Biorep Technologies) and candy cane‐shaped Stainless Steel Pipes (length 30 cm; UMIHIRA) to enable loading from the low‐density solution to high‐density solution, leaving the stainless steel pipe in place. After generating a continuous gradient, the digested tissue was loaded as the top layer 25 . The bottles were centrifuged at 1000 rpm (235 × g ) for five min at 4°C.…”

“…Therefore, 1500 IE and 2000 IE of porcine islets in each group were processed for transplantation. Diabetes induction, transplantation into 6‐week‐old male severe combined immunodeficiency (SCID) mice (Charles River Laboratories Japan, Inc, Kanagawa, Japan), and intraperitoneal glucose tolerance testing (IPGTT) were performed as described previously 22,25,28,32 . All animal studies were approved by the Institutional Animal Care and Use Committee of the University of the Ryukyus.…”

“…We compared the performance of EP solution with that of UW solution in porcine islet isolation. We used porcine islets in this study because the use of human islets for research is illegal in Japan, and porcine islets are expected to be used in islet xenotransplantation 6,9,22‐26 …”

Pancreatic Islet Purification from Large Mammals and Humans Using a COBE 2991 Cell Processor versus Large Plastic Bottles

“…Therefore, some groups including our group have used controlled osmolality solutions for islet purification [11,17,25,[56][57][58][59]. Islet purification using large plastic bottles has some advantages over COBE purification [18,19,[22][23][24]. The bottle purification method is simple and easy to perform, while the COBE 2991 cell processor is high in cost and is not easy to The method was to mix the digested tissue with a low-density solution and then to make a continuous gradient.…”

“…A semiautomated computerized COBE 2991 cell processor is commonly used as is considered the gold standard method [1,2]. We and other groups have reported islet purification from large mammals and humans using large cylindrical plastic bottles [18][19][20][21][22][23][24], which substantially improved the efficacy in comparison to standard purification using a COBE 2991 cell processor [18]. This review describes current advances in islet purification from the pancreas of large mammals and humans using a COBE 2991 cell processor versus large plastic bottles.…”