Saturday September 25, 2021 - 15:50 to 17:05
Comparison of islet isolation result and clinical applicability according to GMP-grade collagenase enzyme blend in adult porcine islet isolation and culture
Kyungmin Kwak1, Jae-kyung Park1, Joohyun Shim1, Nayoung Ko1, Hyoung-Joo Kim1, Yongjin Lee1, Jun-Hyeong Kim1, Michael Alexander2, Jonathan Lakey2,3, Hyunil Kim1, Kimyung Choi1.
1Optipharm Inc, Osong, Korea; 2Department of Biomedical Engineering, University of California Irvine, Irvine, CA, United States; 3Department of Surgery, University of California Irvine, Orange, CA, United States
Introduction: Islet transplantation is promising treatment for type 1 diabetes as an alternative of long term insulin and drug therapy. However, due to the shortage of human organ donors, several research groups have studied pig islets. Islet quality is very important for successful islet xenotransplantation. Producing well-functioning and viable porcine islets requires the establishment of good and consistent isolation and digestion techniques, but still requires appropriate enzyme types and blends. We evaluated the effect of enzyme quality on islet isolation and subsequent islet function as part of a validation plan for the usefulness of adult porcine islet isolate quality control research techniques.
Methods: Adult pig islets were isolated from 16 to 17 month old Yucatan miniature pigs according to standard rapid procurement. Both GMP grade enzymes (Collagenase AF-1 GMP grade, Liberase MTF C/T GMP grade) and crude enzyme (Collagenase P) were evaluated after ductal cannulation. Islet quality control assessments including islet yield, islet size (IEQ), membrane integrity, and functional viability were assessed three times 1 day after islet isolation
Results: The islet yield was highest in the group of adult pigs in which collagenase AF-1 GMP grade was used: the mean islet yield was 16,586 ± 1,391 IEQ/g versus pancreas isolated using crude collagenase P, from 8,302 ± 986 IEQ/g. The islet sizes are as follows: Collagenase AF-1 GMP grade with neutral protease is greater compared to Collagenase P and Liberase MTF C/T GMP grade. Although no significant differences were observed between the experimental groups, in vitro islet function after overnight tissue culture was significantly higher in the collagenase AF-1 GMP grade with neutral protease and Liberase MTF C/T GMP grade compared to the crude control enzyme group. As expected, the GMP grade enzyme had significantly lower endotoxin levels compared to the crude control enzyme group.
Conclusions: Our group confirmed that the GMP grade enzyme had a higher islet isolation yield compared to the crude enzyme. Additionally, GMP grade enzymes are highly effective in increasing islet size, viability, function and lowering acceptable endotoxin levels.Although the enzyme grade may have a limited effect on the viability of islet cells and the function of islets after transplantation. Nevertheless, islets of good quality can be a good criteria in the initiation of xenogeneic islet transplantation and a good criteria to illustrate the complexity of xenogeneic islet transplantation.
This research was supported by a grant of the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number : HI20C0056). .