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Optimizing Temperature and Oxygen Supports Long-term Culture of Human Islets

Komatsu, Hirotake, MD, PhD1; Rawson, Jeffrey1; Medrano, Leonard1; Cook, Colin A.2; Barriga, Alyssa1; Gonzalez, Nelson1; Salgado, Mayra1; Omori, Keiko, MD, PhD1; Kandeel, Fouad, MD, PhD1; Tai, Yu-Chong, PhD2; Mullen, Yoko, MD, PhD1

doi: 10.1097/TP.0000000000002280
Original Basic Science—General

Islet transplantation is a promising treatment for type-1 diabetes; however, donor shortage is a concern. Even when a pancreas is available, low islet yield limits the success of transplantation. Islet culture enables pooling of multiple low-yield isolations into an effective islet mass, but isolated islets rapidly deteriorate under conventional culture conditions. Oxygen (O2) depletion in the islet core, which leads to central necrosis and volume loss, is one of the major reasons for this deterioration.

Methods To promote long-term culture of human islets in PIM-R medium (used for islet research), we adjusted temperature (12°C, 22°C, and 37°C) and O2 concentration (21% and 50%). We simulated the O2 distribution in islets based on islet O2 consumption rate and dissolved O2 in the medium. We determined the optimal conditions for O2 distribution and volume maintenance in a 2-week culture and assessed viability and insulin secretion compared to noncultured islets. In vivo islet engraftment was assessed by transplantation into diabetic nonobese diabetic-severe combined immunodeficiency mouse kidneys. We validated our results using CMRL 1066 medium (used for clinical islet transplantation).

Results Simulation revealed that 12°C of 50% O2 PIM-R culture supplied O2 effectively into the islet core. This condition maintained islet volume at greater than 90% for 2 weeks. There were no significant differences in viability and function in vitro or diabetic reversal rate in vivo between 2-week cultured and noncultured islets. Similar results were obtained using CMRL 1066.

Conclusions By optimizing temperature and O2 concentration, we cultured human islets for 2 weeks with minimal loss of volume and function.

Culture of pancreatic islets pooled from multiple low-yield isolations could allow an effective islet mass, but islets rapidly deteriorate due to oxygen depletion in the islet core. Culture of islets in 12°C–50% oxygen maintain islet volume at >90% for 2 weeks with no significant differences in diabetic reversal rate in vivo compared with noncultured islets.

1 Department of Translational Research & Cellular Therapeutics, Beckman Research Institute of City of Hope, Duarte, CA.

2 Department of Electrical Engineering, California Institute of Technology, Pasadena, CA.

Received 5 February 2018. Revision received 12 April 2018.

Accepted 26 April 2018.

The authors declare no conflicts of interest.

This study was supported by a grant from the Nora Eccles Treadwell Foundation (Title of Grant: CURE OF DIABETES, Grant Period: July 1, 2012 to June 30, 2020, P.I.: Yoko Mullen, MD, PhD).

H.K. designed the study, collected and analyzed data, and wrote the article. J.R., L.M., C.A.C., A.B., N.G., and M.S. collected data. K.O., F.K., YC.T., and Y.M. reviewed and edited the article.

Correspondence: Hirotake Komatsu, MD, PhD, Department of Translational Research & Cellular Therapeutics, Beckman Research Institute of City of Hope, 1500 E. Duarte Rd., Duarte, CA 91010. (hkomatsu@coh.org).

Supplemental digital content (SDC) is available for this article. Direct URL citations appear in the printed text, and links to the digital files are provided in the HTML text of this article on the journal’s Web site (www.transplantjournal.com).

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