Devices and methods for maintenance of temperature and pressure during islet shipment.

UNLABELLED: Exposure to extreme temperatures and pressure fluctuations during shipment by air may have a detrimental impact on islet quality. In this study, we sought to assess the ability of methods and devices to provide better control of the internal environment of islet shipping containers in terms of temperature and pressure. METHODS: Experimental islet shipping containers were packed with 21 panels of commercially available TCP Phase 22 Phase Change Material (TCP). The containers were then exposed for at least 15 hours to three constant external temperature conditions, namely, -20 degrees C, 4 degrees C, and 40 degrees C, and then evaluated for their ability to maintain an internal temperature close to the desired value of 22 degrees C. Custom-designed pressure regulated gyroscopic shipping containers (PRGSC) placed in a vacuum chamber were exposed to an absolute pressure of 250 mm Hg (substantially lower than that experienced during shipment by air) for 25 minutes to assess their ability to control internal pressure under vacuum. Electronic data loggers were used to monitor internal and external temperatures and pressures under all conditions. RESULTS: Twenty-one TCP panels placed in a single islet shipping container were able to maintain the internal temperature between 17 degrees C and 24 degrees C for a minimum of 15 hours at all three external temperatures. The PRGSC tested were able to maintain a constant internal pressure of 760 mm Hg when exposed to vacuum. CONCLUSIONS: Our results demonstrated that the use of containers equipped with TCP and PRGSC exert excellent environmental control over islet shipments by minimizing temperature and eliminating pressure fluctuations.

High-density culture of human islets on top of silicone rubber membranes.

Islet culture has emerged as a standard practice prior to clinical transplantation. However, culturing large numbers of islets requires low islet density (number of islets per unit surface area) and, consequently, 20 to 30 flasks per pancreas in order to avoid hypoxia-induced death (HID). There is a need for a simple, practical, small-footprint culture vessel that will accommodate aseptic maintenance of entire human islet isolations while avoiding HID. In this communication, we examine the hypothesis that by improving oxygen transfer through culture of islets on silicone rubber membranes (SRM), we may increase islet surface coverage and reduce the number of flasks required while avoiding HID. Our results demonstrate that islets cultured for up to 48 hours in vessels with SRM bottoms at 2000 to 4000 islet equivalents (IE)/cm(2), a surface coverage 10- to 20-fold higher than the standard culture protocol, displayed no significant loss of viability. In contrast, islets cultured for 48 hours at 4000 IE/cm(2) in flasks with gas-impermeable bottoms suffered a 60% to 70% reduction in viability. The data suggest that it is possible to culture all islets isolated from a human pancreas on SRM in a single, standard-sized vessel while maintaining the same viability as with the current, standard culture protocols that require 20 to 30 flasks. This approach may lead to substantial improvements in islet culture for research and clinical transplantation.

Human islet viability and function is maintained during high-density shipment in silicone rubber membrane vessels.

BACKGROUND: The shipment of human islets (IE) from processing centers to distant laboratories is beneficial for both research and clinical applications. The maintenance of islet viability and function in transit is critically important. Gas-permeable silicone rubber membrane (SRM) vessels reduce the risk of hypoxia-induced death or dysfunction during high-density islet culture or shipment. SRM vessels may offer additional advantages: they are cost-effective (fewer flasks, less labor needed), safer (lower contamination risk), and simpler (culture vessel can also be used for shipment). METHOD: IE were isolated from two manufacturing centers and shipped in 10-cm(2) surface area SRM vessels in temperature- and pressure-controlled containers to a distant center after at least 2 days of culture (n = 6). Three conditions were examined: low density (LD), high density (HD), and a microcentrifuge tube negative control (NC). LD was designed to mimic the standard culture density for IE preparations (200 IE/cm(2)), while HD was designed to have a 20-fold higher tissue density, which would enable the culture of an entire human isolation in 1-3 vessels. Upon receipt, islets were assessed for viability (measured by oxygen consumption rate normalized to DNA content [OCR/DNA)]), quantity (measured by DNA), and, when possible, potency and function (measured by dynamic glucose-stimulated insulin secretion measurements and transplants in immunodeficient B6 Rag(+/-) mice). Postshipment OCR/DNA was not reduced in HD vs LD and was substantially reduced in the NC condition. HD islets exhibited normal function postshipment. Based on the data, we conclude that entire islet isolations (up to 400,000 IE) may be shipped using a single, larger SRM vessel with no negative effect on viability and ex vivo and in vivo function.

Bioengineering in development of the hybrid artificial pancreas.

The hybrid artificial pancreas for treatment of diabetes consists of insulin-secreting pancreatic tissue which is surrounded by a membrane that protects the tissue from rejection by the immune system following implantation. In this paper, we review the alternative therapeutic approaches for diabetes under study and then discuss the technical requirements that must be met by a hybrid device useful to humans. Previous work on intravascular and extravascular immunoisolation devices is reviewed from the standpoint of these requirements, and three critical unresolved issues are discussed: biocompatibility, oxygen supply limitations, and prevention of immune rejection.