Nonsimultaneous administration of pancreas dissociation enzymes during islet isolation.
BACKGROUND
Successful islet isolation relies heavily on enzyme products. Among them, Liberase was used in islet transplantation programs until the islet community was notified of the use of a bovine brain component during the manufacturing process. To minimize potential risk of prion disease transmission, many islet isolation facilities switched to Serva enzyme, which is considered to pose less risk. However, this conversion significantly affected the field in transplant activity. Here, we report our successful conversion from Liberase to Serva collagenase with the use of a modified digestion protocol.


METHODS
We compared the quality of Serva versus Liberase enzyme using chromatography and collagenase activity assay. On the basis of the findings, we developed a pancreas digestion protocol optimized for Serva enzyme, where only collagenase was injected into the pancreas through the duct, and then neutral protease was added to the circulating system during the digestion phase.


RESULTS
Class I collagenase activity of Serva was remarkably reduced compared with Liberase. Chromatography of Serva demonstrated suspected degradation of class I collagenase. When the modified protocol was applied to donor pancreata more than 35 years of age, we recovered 3119+/-147 islet equivalent/g pancreas with a success rate of 51% (35/68), whereas the standard method yielded only 1809+/-266 islet equivalent/g pancreas (P=0.02) with a success rate of 11% (1/9). This beneficial effect of the modified method was, however, diminished when applied to younger donor pancreata.


CONCLUSIONS
Our study brings new insight into the role of collagenase and noncollagenolytic protease on pancreas dissociation.
Kin
Tatsuya
T
Clinical Islet Transplant Program, University of Alberta and Alberta Health Services, Edmonton, Alberta, Canada. tkin@ualberta.ca
O'Gorman
Doug
D
Zhai
Xiaojun
X
Pawlick
Rena
R
Imes
Sharleen
S
Senior
Peter
P
Shapiro
A M James
AM
D000328:Adult; D015992:Body Mass Index; D002102:Cadaver; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange / Q000379:methods; D017364:Collagenases / Q000378:metabolism; D006801:Humans; D007515:Islets of Langerhans / Q000166:cytology*; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D008875:Middle Aged; D009929:Organ Size; D010179:Pancreas / Q000033:anatomy & histology / Q000166:cytology* / Q000201:enzymology; D013820:Thermolysin; D014019:Tissue Donors; D020858:Tissue and Organ Harvesting / Q000379:methods*
D017364:Collagenases; C106635:Liberase; D013820:Thermolysin