Large variability of the intracellular ATP content of human islets isolated from different donors

Abstract

Observations in experimental heart, liver, kidney and pancreas transplantation indicated that graft function and survival correlates significantly with ATP content of transplanted tissue. The ATP content of cells can be reduced by several factors i.e. the nutritional donor status, storage technique, warm ischemia and cold ischemia time. This study investigates the intracellular ATP content of isolated human islets for the first time.

Quantified samples of freshly isolated (digestion-filtration, continuous ficoll gradient purification) and cultured (22°C, CMRL+10% FCS) islet equivalents (IEQ) of consecutively processed human pancreata from multiorgan donors (UW vascular flush) were shock frozen in liquid nitrogen and stored at –196°C until rapid thawing, sonification and subsequent luminometric determination of ATP (Luciferin-Luciferase-reaction) and assessment of islet protein (IP). The ATP content was analysed for freshly isolated and subsequently 5±1 days cultured islets (n=10).

The ATP content of freshly isolated human islets was 130.4±53.4 pg/µg IP (mean ± SEM) corresponding to 20.7±6.3 pg/IEQ. After culture ATP content increased to 265.5±113.3 pg/µg IP (204.2±41.5%) corresponding to 43.7±15.3 pg/IEQ (216.1±34.9%; p<0.05). The coefficient of variation was 129.5%, 96.5% (fresh) and 135.0%, 111.0% (cultured) for ATP/µg IP and ATP/IEQ, respectively.

The present data show that: (1) the ATP content of freshly isolated human islets varies enormously; (2) intraislet ATP levels increase significantly during 22°C culture suggesting that the capacity to produce ATP is maintained despite hypothermic environment. More data are necessary to clarify the relevance of intraislet ATP content for graft function and survival after islet transplantation.