P-253

Function and viability of explanted islet-containing alginate sheets

M. Alexander¹, S. Li¹, M. Lamb¹, R. Storrs², R. Dorian², S. King², H. Ichii¹, C. Foster¹, J. Lakey^1
1 University of California Irvine Medical Center, Surgery, Orange, USA; ² Islet Sheet Medical LLC, San Francisco, USA

Objectives: Transplantation ofpancreatic islets without the use of pharmaceutical immunosuppression wouldimprove overall patient outcomes in the treatment of type 1 diabetes. We aim toencapsulate and implant islets in a semipermeable flat sheet of alginate anddemonstrate function in the islet implant after it was removed from therecipient.

Methods: Islets were isolated from maleFisher rats using standard techniques and encapsulated in thin alginatesheets. Recipientmale Fisher rats were rendereddiabetic by intravenous injection of streptozotocin (STZ, 25mg/kg) and sheetswere implanted subcutaneously after conformation of diabetes (blood sugar>300mg/dl on three consecutive measurements). Islet function was determinedby glucose stimulated insulin release (GSIR) assay. Stimulation index (SI) was calculated as insulin secretedfollowing a 1 hour culture in Krebs buffer supplemented with high glucose(28mM) + IBMX (50uM) over insulin secreted in low glucose (2.8mM). Cellviability was measured using FDA/PI fluorescence microscopy. Experimentalgroups were unencapsulated rat islets (100 islets, n=7), islet-containingalginate sheets (100 islets, n=3) and islet sheets that were explanted fromdiabetic rats after 4 weeks.

Results: Unencapsulated islets werefunctional with an SI of 3.5±0.8 (mean±SEM) and cell viability of 73.8±0.1%.Islets encapsulated in alginate sheets exhibited SI of 1.7±0.3 and viability of71.6±0.1%. Islet sheets explantedafter 4 weeks in vivo, retained in vivo function (SI=4.2±2) and viability(61.0±0.1%). Furthermore, the blood glucose and insulin requirement of thediabetic rats was reduced from 404±24mg/dL with 3U insulin prior to transplantto 110±18mg/dL with 1U insulin for 18 days after transplant.

Conclusions: This study demonstrated that both islet viability, measured usingmembrane integrity, and islet function measured by GSIR, can be maintained for4 weeks SC in diabetic rats when encapsulated in alginate sheets.

P-253

Function and viability of explanted islet-containing alginate sheets

M. Alexander¹, S. Li¹, M. Lamb¹, R. Storrs², R. Dorian², S. King², H. Ichii¹, C. Foster¹, J. Lakey^1
1 University of California Irvine Medical Center, Surgery, Orange, USA; ² Islet Sheet Medical LLC, San Francisco, USA

Objectives: Transplantation ofpancreatic islets without the use of pharmaceutical immunosuppression wouldimprove overall patient outcomes in the treatment of type 1 diabetes. We aim toencapsulate and implant islets in a semipermeable flat sheet of alginate anddemonstrate function in the islet implant after it was removed from therecipient.

Methods: Islets were isolated from maleFisher rats using standard techniques and encapsulated in thin alginatesheets. Recipientmale Fisher rats were rendereddiabetic by intravenous injection of streptozotocin (STZ, 25mg/kg) and sheetswere implanted subcutaneously after conformation of diabetes (blood sugar>300mg/dl on three consecutive measurements). Islet function was determinedby glucose stimulated insulin release (GSIR) assay. Stimulation index (SI) was calculated as insulin secretedfollowing a 1 hour culture in Krebs buffer supplemented with high glucose(28mM) + IBMX (50uM) over insulin secreted in low glucose (2.8mM). Cellviability was measured using FDA/PI fluorescence microscopy. Experimentalgroups were unencapsulated rat islets (100 islets, n=7), islet-containingalginate sheets (100 islets, n=3) and islet sheets that were explanted fromdiabetic rats after 4 weeks.

Results: Unencapsulated islets werefunctional with an SI of 3.5±0.8 (mean±SEM) and cell viability of 73.8±0.1%.Islets encapsulated in alginate sheets exhibited SI of 1.7±0.3 and viability of71.6±0.1%. Islet sheets explantedafter 4 weeks in vivo, retained in vivo function (SI=4.2±2) and viability(61.0±0.1%). Furthermore, the blood glucose and insulin requirement of thediabetic rats was reduced from 404±24mg/dL with 3U insulin prior to transplantto 110±18mg/dL with 1U insulin for 18 days after transplant.

Conclusions: This study demonstrated that both islet viability, measured usingmembrane integrity, and islet function measured by GSIR, can be maintained for4 weeks SC in diabetic rats when encapsulated in alginate sheets.