Improvement in islet yield from a cold-preserved pancreas by pancreatic ductal collagenase distention at the time of harvesting.
This study tried to improve the number of viable islets isolated from a pancreas because a sufficient number cannot be obtained when the organ is preserved in the manner used for pancreas transplantation. The mechanism involved in the decrease in islet yield during preservation was studied to try to develop a better method for islet preparation. First, the integrity of the ductal system was compared between fresh and 6-hr simply preserved (in Hanks' balanced salt solution) rat pancreases. The ductal pressure after ductal injection of HBSS reached a plateau earlier and was significantly lower for the preserved pancreases (0.073 +/- 0.026 min, 410 +/- 17 mmHg, n = 5) than for the fresh ones (0.176 +/- 0.086 min, 561 +/- 103 mmHg, n = 7, P less than 0.05). Second, the extent of pancreatic distention was examined following ductal injection of barium gelatin solution. Solution leakage occurred earlier and distention was less in the preserved pancreas. In addition, the gelatin was found in the capillaries within some islets of the preserved pancreas. These results indicated that the preservation led to a rapid loss of integrity of the ductal system before collagenase injection. We therefore tested the efficacy of ductal collagenase injection at the time of harvesting: 15 ml of 1.0 mg/ml collagenase HBSS was intraductally injected and the pancreas was preserved at 4 degrees C for 2, 4, 6, and 24 hr. The isolation procedure was similar to that used for the fresh pancreas. The yield was significantly better than that of the simply preserved pancreas at 4 hr (241 +/- 22, n = 3, vs. 140 +/- 58, n = 3, P less than 0.05) and at 6 hr (171 +/- 58, n = 14, vs. 32 +/- 33, n = 6, P less than 0.01). These isolated islets were spherical-oval and their viability was confirmed by the ability to reverse STZ-induced diabetes in mice. These results indicated that the integrity of the ductal system, which is necessary for distention of the whole pancreas, was lost during preservation. To solve this problem, ductal collagenase injection should be done at the time of pancreas harvesting and then followed by simple preservation. This method is recommended to obtain viable islets from a preserved pancreas.
Ohzato
H
H
Department of Surgery II, Osaka University Medical School, Japan.
Gotoh
M
M
Monden
M
M
Dono
K
K
Kanai
T
T
Mori
T
T
D000818:Animals; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D003080:Cold Temperature; D007511:Ischemia; D007515:Islets of Langerhans / Q000166:cytology*; D008297:Male; D003012:Microbial Collagenase; D009926:Organ Preservation / Q000379:methods; D010179:Pancreas / Q000098:blood supply / Q000166:cytology*; D010183:Pancreatic Ducts / Q000502:physiology; D011312:Pressure; D051381:Rats; D011919:Rats, Inbred Strains; D013997:Time Factors
D003012:Microbial Collagenase