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P.127 The influence of temperature conditions and reconstruction of the extracellular matrix environment on the viability of pancreatic islets

Michal Wszola, Poland

Foundation of Research and Science Development

Abstract

The influence of temperature conditions and reconstruction of the extracellular matrix environment on the viability of pancreatic islets

Michał Wszoła1, Marta Klak1, Grzegorz Tymicki1, Magdalena Gomółka1, Patrycja Kowalska1, Tomasz Bryniarski1, Andrzej Berman1.

1Foundation of Research and Science Development, Warsaw, Poland

Introduction: Pancreatic islet transplantation is one of the methods of treating patients with T1D. However, for the procedure to be successful, the high quality of the implanted islets is of key importance for a successful islet transplant. Following islet isolation, it has been shown that mainly β-cells undergo apoptosis due to vascular loss and hypoxia. The cellular stresses to which islets are subjected during isolation (for example disturbance of the cell-matrix relationship). Appropriate storage conditions for insulated islands are of key importance for their functionality. The most common method of islet storage is the introduction into a suspension culture. However, because these are micro organs, the storage of pancreatic islets is a more accurate term. Therefore, this study aimed to demonstrate whether for pancreatic islets it is better to cultivate them in standard conditions (37oC; 5% CO2), or maybe stored them at 4oC.

Material: The research material was porcine pancreatic islets. Islets were isolated from the native organ using collagenase and a Ricordi chamber. After isolation, the material was divided into 4 research groups: 1. control (islands suspended in CMRL 1066 medium) - traditional breeding conditions (temperature 37oC; 5% CO2); 2. CMRL 1066 suspended islets stored at 4oC; 3. Islets in CMRL 1066 medium with the addition of dECM (decellularized extracellular matrix) - in traditional culture conditions; 4. Islets in CMRL 1066 supplemented with dECM stored at 4oC. Each variant was tested in 3 replicates and each trial was 3000iEq. Additionally, each tested variant was tested on pancreatic islets that were cleaned or not immediately after isolation.

Results: The research showed that the storage temperature, reduced to 4oC, and the addition of ECM significantly influenced the functionality of the islets. This variant tested (uncleaned islands) showed functionality even 5 days after isolation. On the other hand, islets incubated at 37°C without the addition of dECM showed no functionality after 2 days of the experiment. The remaining variants showed a gradual loss of functionality. However, the 4oC variant was better in each case.

Conclusion: Pancreatic islets, as micro organs with their vascularization, need appropriate storage conditions after isolation to ensure their proper functioning. Based on these studies, it was shown that the islands should be stored and not cultivated as single-cell lines. In addition, the addition of dECM allows for the recreation of the matrix-cell environment, which is appropriate storage conditions - corresponding to the native environment.

This study was financially supported by The National Centre for Research and Development STRATEGMED3/305813/2/NCBR/2017.