The importance of tryptic-like activity in purified enzyme blends for efficient islet isolation.
BACKGROUND
The isolation of islets from the human pancreas critically depends on an efficient enzyme blend. Previous studies have solely focused on the presence of collagenase and neutral protease/thermolysin. Despite improved characterization of these components, the lot-related variability in efficacy still persists suggesting that additional so far disregarded enzymes are required for efficient islet cleavage.


METHODS
Varying activities of a tryptic-like enzyme were identified within collagenase NB1 lots, which were selected according to a matched ratio between tryptic-like and collagenase activity (TLA-ratio). Rat and human pancreata were processed with current standard procedures.


RESULTS
Increasing the TLA-ratio from 1.3% to 10% reduced pancreas dissociation time in rats by 50% without affecting islet yield, viability, or posttransplant function in diabetic nude mice. Enhancing the TLA-ratio from 1.3% to 12.6% for human pancreas processing resulted in a significant reduction of recirculation time and increased incrementally human islet yield without affecting purity, in vitro function or recovery after culture. Optimized pancreas digestion correlated with a higher percentage of islet preparations fulfilling quality criteria for clinical transplantation.


CONCLUSIONS
We conclude that TLA is an effective component that should be included in moderate amounts in enzyme blends for human islet isolation to optimize the efficiency and minimize the lot-related variability.
Brandhorst
Heide
H
Department of Oncology, Radiology and Clinical Immunology, Uppsala University Hospital, Uppsala, Sweden. Daniel.Brandhorst@klinimm.uu.se
Friberg
Andrew
A
Andersson
Helena H
HH
Felldin
Maria
M
Foss
A
A
Salmela
Kaija
K
Lundgren
Torbjörn
T
Tibell
Annika
A
Tufveson
Gunnar
G
Korsgren
Olle
O
Brandhorst
Daniel
D
D000818:Animals; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival / Q000502:physiology*; D017364:Collagenases / Q000378:metabolism / Q000494:pharmacology; D003921:Diabetes Mellitus, Experimental / Q000601:surgery*; D005260:Female; D006085:Graft Survival / Q000502:physiology*; D006801:Humans; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000166:cytology*; D007515:Islets of Langerhans / Q000166:cytology*; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D008297:Male; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D051381:Rats; D011917:Rats, Inbred Lew; D013820:Thermolysin / Q000494:pharmacology; D014019:Tissue Donors; D014183:Transplantation, Heterologous; D014357:Trypsin / Q000378:metabolism / Q000494:pharmacology*
D007328:Insulin; D014357:Trypsin; D017364:Collagenases; D013820:Thermolysin