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Calcium : A Crucial Potentiator for Efficient Enzyme Digestion of the Human Pancreas

Eich, Torsten; Ståhle, Magnus; Gustafsson, Bengt; Horneland, Rune; Lempinen, Marko; Lundgren, Torbjörn; Rafael, Ehab LU ; Tufveson, Gunnar; Zur-Mühlen, Bengt von and Olerud, Johan, et al. (2018) In Cell Transplantation 27(7). p.1031-1038
Abstract

Background: Effective digestive enzymes are crucial for successful islet isolation. Supplemental proteases are essential because they synergize with collagenase for effective pancreatic digestion. The activity of these enzymes is critically dependent on the presence of Ca2+ ions at a concentration of 5–10 mM. The present study aimed to determine the Ca2+ concentration during human islet isolation and to ascertain whether the addition of supplementary Ca2+ is required to maintain an optimal Ca2+ concentration during the various phases of the islet isolation process. Methods: Human islets were isolated according to standard methods and isolation parameters. Islet quality control and the number... (More)

Background: Effective digestive enzymes are crucial for successful islet isolation. Supplemental proteases are essential because they synergize with collagenase for effective pancreatic digestion. The activity of these enzymes is critically dependent on the presence of Ca2+ ions at a concentration of 5–10 mM. The present study aimed to determine the Ca2+ concentration during human islet isolation and to ascertain whether the addition of supplementary Ca2+ is required to maintain an optimal Ca2+ concentration during the various phases of the islet isolation process. Methods: Human islets were isolated according to standard methods and isolation parameters. Islet quality control and the number of isolations fulfilling standard transplantation criteria were evaluated. Ca2+ was determined by using standard clinical chemistry routines. Islet isolation was performed with or without addition of supplementary Ca2+ to reach a Ca2+ of 5 mM. Results: Ca2+ concentration was markedly reduced in bicarbonate-based buffers, especially if additional bicarbonate was used to adjust the pH as recommended by the Clinical Islet Transplantation Consortium. A major reduction in Ca2+ concentration was also observed during pancreatic enzyme perfusion, digestion, and harvest. Additional Ca2+ supplementation of media used for dissolving the enzymes and during digestion, perfusion, and harvest was necessary in order to obtain the concentration recommended for optimal enzyme activity and efficient liberation of a large number of islets from the human pancreas. Conclusions: Ca2+ is to a large extent consumed during clinical islet isolation, and in the absence of supplementation, the concentration fell below that recommended for optimal enzyme activity. Ca2+ supplementation of the media used during human pancreas digestion is necessary to maintain the concentration recommended for optimal enzyme activity. Addition of Ca2+ to the enzyme blend has been implemented in the standard isolation protocols in the Nordic Network for Clinical Islet Transplantation.

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publishing date
type
Contribution to journal
publication status
published
subject
keywords
calcium, clinical islet transplantation, diabetes, islet isolation
in
Cell Transplantation
volume
27
issue
7
pages
8 pages
publisher
Cognizant Communication Corporation
external identifiers
  • scopus:85050929587
ISSN
0963-6897
DOI
10.1177/0963689718779350
language
English
LU publication?
no
id
309b1956-c3b6-4fe7-a544-58a15dfbb037
date added to LUP
2018-09-12 14:49:19
date last changed
2019-02-20 11:26:41
@article{309b1956-c3b6-4fe7-a544-58a15dfbb037,
  abstract     = {<p>Background: Effective digestive enzymes are crucial for successful islet isolation. Supplemental proteases are essential because they synergize with collagenase for effective pancreatic digestion. The activity of these enzymes is critically dependent on the presence of Ca<sup>2+</sup> ions at a concentration of 5–10 mM. The present study aimed to determine the Ca<sup>2+</sup> concentration during human islet isolation and to ascertain whether the addition of supplementary Ca<sup>2+</sup> is required to maintain an optimal Ca<sup>2+</sup> concentration during the various phases of the islet isolation process. Methods: Human islets were isolated according to standard methods and isolation parameters. Islet quality control and the number of isolations fulfilling standard transplantation criteria were evaluated. Ca<sup>2+</sup> was determined by using standard clinical chemistry routines. Islet isolation was performed with or without addition of supplementary Ca<sup>2+</sup> to reach a Ca<sup>2+</sup> of 5 mM. Results: Ca<sup>2+</sup> concentration was markedly reduced in bicarbonate-based buffers, especially if additional bicarbonate was used to adjust the pH as recommended by the Clinical Islet Transplantation Consortium. A major reduction in Ca<sup>2+</sup> concentration was also observed during pancreatic enzyme perfusion, digestion, and harvest. Additional Ca<sup>2+</sup> supplementation of media used for dissolving the enzymes and during digestion, perfusion, and harvest was necessary in order to obtain the concentration recommended for optimal enzyme activity and efficient liberation of a large number of islets from the human pancreas. Conclusions: Ca<sup>2+</sup> is to a large extent consumed during clinical islet isolation, and in the absence of supplementation, the concentration fell below that recommended for optimal enzyme activity. Ca<sup>2+</sup> supplementation of the media used during human pancreas digestion is necessary to maintain the concentration recommended for optimal enzyme activity. Addition of Ca<sup>2+</sup> to the enzyme blend has been implemented in the standard isolation protocols in the Nordic Network for Clinical Islet Transplantation.</p>},
  author       = {Eich, Torsten and Ståhle, Magnus and Gustafsson, Bengt and Horneland, Rune and Lempinen, Marko and Lundgren, Torbjörn and Rafael, Ehab and Tufveson, Gunnar and Zur-Mühlen, Bengt von and Olerud, Johan and Scholz, Hanne and Korsgren, Olle},
  issn         = {0963-6897},
  keyword      = {calcium,clinical islet transplantation,diabetes,islet isolation},
  language     = {eng},
  month        = {07},
  number       = {7},
  pages        = {1031--1038},
  publisher    = {Cognizant Communication Corporation},
  series       = {Cell Transplantation},
  title        = {Calcium : A Crucial Potentiator for Efficient Enzyme Digestion of the Human Pancreas},
  url          = {http://dx.doi.org/10.1177/0963689718779350},
  volume       = {27},
  year         = {2018},
}