A new enzyme mixture to increase the yield and transplant rate of autologous and allogeneic human islet products.
BACKGROUND
The optimal enzyme blend that maximizes human islet yield for transplantation remains to be determined. In this study, we evaluated eight different enzyme combinations (ECs) in an attempt to improve islet yield. The ECs consisted of purified, intact or truncated class 1 (C1) and class 2 (C2) collagenases from Clostridium histolyticum (Ch), and neutral protease (NP) from Bacillus thermoproteolyticus rokko (thermolysin) or Ch (ChNP).


METHODS
We report the results of 249 human islet isolations, including 99 deceased donors (research n=57, clinical n=42) and 150 chronic pancreatitis pancreases. We prepared a new enzyme mixture (NEM) composed of intact C1 and C2 collagenases and ChNP in place of thermolysin. The NEM was first tested in split pancreas (n=5) experiments and then used for islet autologous (n=21) and allogeneic transplantation (n=10). Islet isolation outcomes from eight different ECs were statistically compared using multivariate analysis.


RESULTS
The NEM consistently achieved higher islet yields from pancreatitis (P<0.003) and deceased donor pancreases (P<0.001) than other standard ECs. Using the NEM, islet products met release criteria for transplantation from 8 of 10 consecutive pancreases, averaging 6510 ± 2150 islet equivalent number/gram (IEQ/g) pancreas and 694,681 ± 147,356 total IEQ/transplantation. In autologous isolation, the NEM yielded more than 200,000 IEQ from 19 of 21 pancreases (averaging 422,893 ± 181,329 total IEQ and 5979 ± 1469 IEQ/kg recipient body weight) regardless of the severity of fibrosis.


CONCLUSIONS
A NEM composed of ChNP with CIzyme high intact C1 collagenase recovers higher islet yield from deceased and pancreatitis pancreases while retaining islet quality and function.
Schulze Diabetes Institute, University of Minnesota, Minneapolis, MN 55455, USA. bala@umn.edu
A N
AN
Balamurugan
Gopalakrishnan
G
Loganathan
Melena D
MD
Bellin
Joshua J
JJ
Wilhelm
James
J
Harmon
Takayuki
T
Anazawa
Sajjad M
SM
Soltani
David M
DM
Radosevich
Takeshi
T
Yuasa
Mukesh
M
Tiwari
Klearchos K
KK
Papas
Robert
R
McCarthy
David E R
DE
Sutherland
Bernhard J
BJ
Hering
D000328:Adult; D001406:Bacillaceae / Q000201:enzymology*; D016009:Chi-Square Distribution; D047009:Hathewaya histolytica / Q000201:enzymology*; D010450:Endopeptidases / Q000302:isolation & purification / Q000378:metabolism*; D005260:Female; D006801:Humans; D007515:Islets of Langerhans / Q000378:metabolism*; D016381:Islets of Langerhans Transplantation; D008297:Male; D003012:Microbial Collagenase / Q000302:isolation & purification / Q000378:metabolism*; D008875:Middle Aged; D008910:Minnesota; D015999:Multivariate Analysis; D012044:Regression Analysis; D013820:Thermolysin / Q000302:isolation & purification / Q000378:metabolism*; D014022:Tissue Survival; D020858:Tissue and Organ Harvesting / Q000379:methods*; D014182:Transplantation, Autologous; D014184:Transplantation, Homologous; D016896:Treatment Outcome; D055815:Young Adult
D010450:Endopeptidases; D013820:Thermolysin; D003012:Microbial Collagenase