Evaluation of human islet-specific functional quality cultured on different gas-permeable membranes.
The aim of this study was to investigate different gas-permeable membranes for culturing human islets. Dynamic insulin release was used to assess islet functional quality. Islets isolated from cadaveric pancreata (n = 8) using standard isolation methods were stained with dithizone, counted, and cultured on five different commercially available medical-grade membranes reported to have high permeability to O2, CO2, and other gases. Fraction 1 (20,000 islet equivalents [IEQ] purity >70%; viability >85%) was cultured using serum-free medium in nonadherence tissue culture flasks (group I) and custom-made chambers with membranes (group II). Each vessel contained 5000 IEQ at a density of 30 IEQ/cm2 and 69 IEQ/cm2 for groups I and II, respectively. Islets were cultured for 48 to 90 hours at 37 degrees C in 5% CO2. In vitro dynamic insulin response to low glucose (3 mmol/L), high glucose (16.7 mmol/L), and 25 mmol/L KCI was measured. Stimulation indices were calculated by dividing average of initial response over basal insulin release; basal insulin release defined as average of the first seven values. Islets cultured on MG7 (n = 3) showed a higher stimulation index (3.49 +/- 0.37) compared with flasks (2.44 +/- 0.22), indicating better specific functional quality. Islets cultured on other membranes proved to show similar or worse functional quality than those cultured in flasks. In fact, islets cultured on MG6 (n = 2) were not tested owing to complete disintegration. Islet functional quality was improved when cultured on selected biocompatible gas-permeable membranes; however, finding the best membrane requires further investigation before clinical application.
Bentsi-Barnes
K
K
City of Hope National Medical Center, and Beckman Research Institute, Duarte, California 91010, USA.
Kandeel
F
F
Al-Abdullah
I H
IH
D002102:Cadaver; D002245:Carbon Dioxide / Q000032:analysis; D002452:Cell Count; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D016895:Culture Media, Serum-Free; D005740:Gases / Q000032:analysis*; D006801:Humans; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism / Q000502:physiology*; D016381:Islets of Langerhans Transplantation; D008567:Membranes, Artificial; D010100:Oxygen / Q000032:analysis; D010539:Permeability; D014019:Tissue Donors
D016895:Culture Media, Serum-Free; D005740:Gases; D007328:Insulin; D008567:Membranes, Artificial; D002245:Carbon Dioxide; D010100:Oxygen