Assessment of six different collagenase-based methods to isolate feline pancreatic islets.
Isolation of pancreatic islets is necessary to study the molecular mechanisms underlying beta-cell demise in diabetic cats. Six collagenase-based methods of isolation were compared in 10 cat pancreata, including single and double course of collagenase, followed or not by Ficoll centrifugation or accutase, and collagenase plus accutase. Morphometric analysis was performed to measure the relative area of islet and exocrine tissue. Islet specific mRNA transcripts were quantified in isolates by real-time PCR. The single and double course of collagenase digestion was successful in each cat and provided similar islet-to-exocrine tissue ratio. Quantities of insulin mRNA did not differ between the two methods. However, on histological examination either method yielded only approximately 2% of pure islets. The other methods provided disrupted islets or insufficient samples in 1-7 cats. Although pancreas digestion with single and double course of collagenase was superior, further studies are needed to improve islet isolation in cats.
Zini
Eric
E
Clinic for Small Animal Internal Medicine, Vetsuisse Faculty, University of Zürich, Winterthurerstr. 260, 8057 Zürich, Switzerland. ezini@vetclinics.uzh.ch
Franchini
Marco
M
Guscetti
Franco
F
Osto
Melania
M
Kaufmann
Karin
K
Ackermann
Mathias
M
Lutz
Thomas A
TA
Reusch
Claudia E
CE
D000818:Animals; D002415:Cats; D018929:Cell Culture Techniques / Q000662:veterinary; D017364:Collagenases / Q000378:metabolism*; D007515:Islets of Langerhans / Q000166:cytology*; D010179:Pancreas / Q000201:enzymology / Q000378:metabolism
D017364:Collagenases