An evaluation of the activation of endogenous pancreatic enzymes during human islet isolations.
Despite advances in human islet isolations, there remain inconsistencies in human islet yield and viability after collagenase digestion. It has been suggested that trypsin may contribute to the proteolysis of collagenase and the destruction of islet cells, or possibly exert indirect effects on the pancreas by activating other endogenous serine proenzymes. This study evaluated the effects of serine proteases on collagenase activity and profiled the kinetics of serine protease activity throughout human islet isolations with and without addition of Pefabloc, a serine protease inhibitor. Cadaveric pancreases were perfused in the presence (n = 12) and absence of Pefabloc (0.4 mmole; n = 8). Samples were collected before and throughout the digestion process and were assayed for trypsin, chymotrypsin, and elastase activity. A study of the enzyme kinetics of serine proteases throughout human islet isolations showed an increase in activity levels throughout the digestion period. There was a significant difference in the chymotrypsin (1342 +/- 503 and 384 +/- 71 units) and elastase (7.94 +/- 1.1 and 2.761 +/- 0.69 units) levels between the control and Pefabloc-supplemented isolations, respectively. There was no significance difference noted among the trypsin (88 +/- 27 and 54 +/- 18 units) levels between the control and Pefabloc-supplemented isolations, respectively. This demonstrates that serine proteases are effectively inhibited by Pefabloc during the islet isolation process. These data show that the presence of serine proteases may likely damage the islets upon prolonged digestion of the pancreatic tissue.
Rose
N L
NL
Department of Surgery, University of Alberta, Edmonton, Alberta, Canada.
Palcic
M M
MM
Shapiro
A M J
AM
Lakey
J R T
JR
D002102:Cadaver; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D002918:Chymotrypsin / Q000378:metabolism; D017364:Collagenases; D010450:Endopeptidases / Q000378:metabolism*; D006801:Humans; D007515:Islets of Langerhans / Q000166:cytology*; D007700:Kinetics; D010179:Pancreas / Q000201:enzymology* / Q000378:metabolism; D010196:Pancreatic Elastase / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D014019:Tissue Donors; D014357:Trypsin
D010450:Endopeptidases; D012697:Serine Endopeptidases; D002918:Chymotrypsin; D010196:Pancreatic Elastase; D014357:Trypsin; D017364:Collagenases