IMPROVED FUNCTIONAL SURVIVAL OF HUMAN ISLETS OF LANGERHANS IN 3-DIMENSIONAL MATRIX CULTURE

The current study evaluates functional survival of human islets mainta ined In tissue culture for up to 4 wk in suspension media (CMRL-1066 w ith supplements) and contrasts these results with immobilizing three-d imensional matrices (agarose or alginate). The absolute number and vol ume of islets retrieved from agarose is significantly higher after two and four wk of culture compared to conventional free-floating media. In vitro function of islets, assessed by insulin/DNA content, insulin secretion into the culture media over 24 h and glucose-theophylline st imulated insulin release in a dynamic perifusion system, was not signi ficantly different between free-floating and matrix preserved islets. In vivo islet function was evaluated by the effectiveness for reversal of insulin-dependent diabetes mellitus by transplantation of the isle ts under the kidney capsule of nude mice. Although adequate insulin re sponses to glucose were seen after culture in conventional or matrix m edia, only agarose embedded islets were consistently able to induce no rmoglycemia in diabetic recipients after 14 days of culture. Additiona l transplantation experiments defined the threshold level required to reverse diabetes to be between 1,000 and 1,500 agarose preserved islet s. Our data suggest improved engraftment of human islets after agarose culture. This culture method may be of benefit for the accumulation o f functionally competent human islets, thus facilitating the implement ation of clinical protocols that utilize freshly isolated islets from multiple donors without the need for cryopreservation.