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New Method Simplifies Sample Preparation for Diabetes-Related Measurements

By iednewsdesk On May 27, 2024

USP researchers have developed a method that expands the possibilities of evaluating the functioning of pancreatic islets, cells that control sugar levels in the body and whose poor performance is linked to diabetes. The technique simplifies the preparation of islet samples, allowing oxygen consumption to be measured and an idea of their functionality using various equipment, without the need for exclusive materials, which could improve diagnostics and drug testing.

The results of the study are in an article in the scientific journal Molecular Metabolism in March this year. “Pancreatic islets are clusters of cells present in our pancreas, an organ that participates in the digestive system and also in the production of hormones”, says professor Eloisa Aparecida Vilas-Boas, from the Faculty of Pharmaceutical Sciences (FCF) at USP, one of the authors of the work. “Islet cells produce several hormones responsible for controlling glucose (sugar) levels in our body and regulating the formation and use of our energy stores.”

“One of the main hormones is insulin, produced by the beta cells of the islets. Insulin directs glucose to be stored and inhibits the use of our energy stores when we have a large energy supply”, reports the professor. “In this way, stocks can be used in situations where we need more energy, such as during physical exercise, and in fasting situations.”

“Disorders in the function of pancreatic islets can lead to the development of a disease called diabetes mellitus , in which our body’s glucose levels are not well controlled”, highlights Eloísa Aparecida Vilas-Boas. “The person will have excess glucose in the bloodstream, but will not be able to use it to generate energy due to the absence of insulin or resistance to its action.”

Energy

The professor notes that the function of islets can be assessed in numerous ways. “Within our cells, we have a very important compartment (organelle) that is responsible for generating energy, called mitochondria, whose activity is closely linked to the function of the cells that produce insulin”, she describes. “Mitochondria use oxygen to generate energy and their oxygen consumption can be used to evaluate their activity.”

“As islets are very large spherical structures, formed by clusters of several cells, this makes it difficult to use conventional methods to measure oxygen consumption”, says Eloisa Aparecida Vilas-Boas. “Most studies use beta cell tumor lines, which are interesting but less robust models.”

To measure oxygen consumption, the Oroboros can be used, equipment in which isolated cells or mitochondria are kept in suspension. “The disadvantages are the use of a very large quantity of samples, extracted from many animals, which needs to be shaken all the time, damaging the islets”, emphasizes the professor. “To avoid this, an isolation chamber was created that keeps the islets safe, but this device is not yet commercialized and does not eliminate the problem of using large numbers of animals.”

Another measuring device is the Seahorse, in which cells are adhered to the bottom of special culture plates. “They are of two types, one developed exclusively for islets and the other only for spherical structures, which was adapted, both much more expensive than the traditional ones, and each one compatible with only one model of the device”, reports Eloisa Aparecida Vilas-Boas. “There is a third piece of equipment available to measure oxygen consumption, Resipher, which had never been adapted for islets.”

Measure

“Our proposal was to develop a new, practical and robust method that would make it possible to measure oxygen consumption in islets using the standard plate, not the special one, and that would be compatible with any model of equipment”, highlights the professor. “The new protocol proposes the prior and controlled dispersion of islet cells, which are then adhered to the standard plate.”

The method developed in the research allowed the assessment of oxygen consumption using both Seahorse models, and also the Resipher, for the first time. “The islets showed a robust response for up to a week, with very high viability”, emphasizes Eloisa Aparecida Vilas-Boas. “Our method was also validated by other functional assays and the cells responded adequately to the different stimuli tested.”

“Based on the assessment of islet function, it is possible to evaluate islets from animal models and patients with insulin production disorders and better understand how these cells fail”, plans the professor. “In addition, it will be useful for translational investigations, such as discovery of pharmacological candidates and islet transplantation protocols.”

The study was designed by professor Eloisa Aparecida Vilas-Boas, from the Department of Clinical and Toxicological Analysis at FCF, corresponding author, and by Débora Santos Rocha, post-doctoral student in the Department of Biochemistry at the Institute of Chemistry (IQ) at USP, first author of the article. The study was carried out with the support of professors Alicia Kowaltowski, Alexandre Bruni-Cardoso and doctoral student Antonio Carlos Manucci, from the Department of Biochemistry at IQ.

University of São Paulo