A stirred microchamber for oxygen consumption rate measurements with pancreatic islets

Abstract

Improvements in pancreatic islet transplantation for treatment of diabetes are hindered by the absence of meaningful islet quality assessment methods. Oxygen consumption rate (OCR) has previously been used to assess the quality of organs and primary tissue for transplantation. In this study, we describe and characterize a stirred microchamber for measuring OCR with small quantities of islets. The device has a titanium body with a chamber volume of about 200 µL and is magnetically stirred and water jacketed for temperature control. Oxygen partial pressure (pO 2 ) is measured by fluorescence quenching with a fiber optic probe, and OCR is determined from the linear decrease of pO 2 with time. We demonstrate that measurements can be made rapidly and with high precision. Measurements with βTC3 cells and islets show that OCR is directly proportional to the number of viable cells in mixtures of live and dead cells and correlate linearly with membrane integrity measurements made with cells that have been cultured for 24 h under various stressful conditions.

Improvement of islet function in a bioartificial pancreas by enhanced oxygen supply and growth hormone releasing hormone agonist

“…Data were analyzed using Labview 7.2 software (National Instruments). OCR was calculated as described (60).…”

Quantitative Assessment of Islets of Langerhans Encapsulated in Alginate

“…37 The time-dependent pO 2 within the chamber was recorded, and data in the region <160 mm Hg and >60 mm Hg, which yielded the highest slope of pO 2 versus time, were fit to a straight line. OCR was evaluated from…”

“…[34][35][36] We developed a stirred microchamber that permits rapid, accurate OCR measurements with a small volume of islet tissue. 37 This method was used in studies with human 38 and rat 39 islets transplanted into immunocompromised mice. Transplantation outcome correlated with two parameters: OCR, a measure of the amount of viable tissue, and OCR/cell (which can be determined with DNA or nuclei measurements), a measure of the relative viability of the tissue.…”

Mitochondrial Dysfunction Contributes to Impaired Insulin Secretion in INS-1 Cells with Dominant-negative Mutations of HNF-1α and in HNF-1α-deficient Islets

“…Oxygen Consumption Rates-Oxygen consumption rates of freshly trypsinized cells or encapsulated cells were measured using a fiber optic oxygen monitor (model 210, Instech Laboratories, Plymouth Meeting, PA) (23). A 250-l chamber was loaded with ϳ2 ϫ 10 6 cells or ϳ5 beads, and changes in the oxygen concentration were monitored under buffer conditions and substrate additions identical to 31 P NMR experiments for measurement of ATP synthesis rates.…”