J Korean Diabetes Assoc.  2000 Apr;24(2):180-190.

Distension and Collagenase Digestion Time of The Pancreas are Critical Factors in Islet Isolation of Canine Pancreas

• Yang TY
• Jeong IK
• Oh SH
• Lee SH
• Kim DJ
• Hahm JR
• Park JH
• Kim JS
• Han JS
• Kim SJ
• Chung JH
• Min YK
• Lee MS
• Lee MK
• Kim KW

Abstract

BACKGROUNDS: One of the main problems conditioning the outcome of islet transplantation is the ability to separate a sufficient number of viable islets with preserved function. Islet purification is critically affected by all of the isolation stages, Thus, it is necessary to set up the standard isolation method that islets are separate well from acinar without compromising islet yield and viability.
METHODS
Twenty three adult mongrel dogs were used for the experiment of total pancreatectomy with islet isolation. The islets were properly isolated by a modified Recordi method. The obtained islets were further purified by centrifugation on discontinuous gradients using cell separation system (Model 2991, Cobe, Lakewood Colo). We evaluatad islet number (islet equivalent number, 150 gm equivalents/kg of recipient body weight, lEq/kg), purity. cell volume, viabilty, recovery rate, and comparison of outcome according to the isolation conditions.
RESULTS
1) The mean of islet numbers before purification were 13543+/-943 lEq/kg, digestion times were 13.8+/-2.6 min. digestion tamperature was b was 59,7+/-7.0%, viability was 90.0+/-2.1%, cell volume was 4.7+/-1.1 mL, islet number after purification were 4064+/-361 lEq/kg, and recovery rate was 29+2.9. 2) Isolated islet numbers were different according to the degree of pancreas distension with collagenase, digestion temperature, and digestion time. 3) The best conditions for islet isolation were above 37.5 degree C in temperature at recirculation of collagenase, within 12min in digestion time and well distended pancreas with collagenase. 4) According to multiple regression adjusted by variable factors, the degree of pancreas distension with collagenase and digestion time were independently associated factors for successful islet isolation.
CONCLUSIONS
In this study, we concluded that the degree of pancreas distension with collagenase and digestion time were independent factors for successful islet isolation and the best conditions for islet isolation were above 37.5 degree C in temperature at recirculation of collagenase, within 12 min in digestion time and well distended pancreas with collagenase.

Keyword

Pancreatic islet cell auto-transplantation; lslet cell survival; Diabetes; Portal pressure

MeSH Terms

Adult
Animals
Body Weight
Cell Separation
Cell Size
Centrifugation
Collagenases*
Digestion*
Dogs
Humans
Islets of Langerhans Transplantation
Pancreas*
Pancreatectomy
Portal Pressure
Collagenases