A new study by Alicia Kowaltowski’s group shows that renal mitochondria from rats submitted to caloric restriction generate more free radicals and are more prone to calcium-induced mitochondrial permeability transition, a process in which the mitochondrial membrane loses selectivity, compromising ATP synthesis and leading to cell death. The study also demonstrated, for the first time, that caloric restriction regulates calcium uptake into mitochondria by modulating the MICU2 protein. According to the researchers, the data suggest that the kidneys respond differently to restricted diets and, in the long term, are susceptible to damage. Kowaltowski is a professor at the Instituto de Química, Universidade de São Paulo and a member of the RIDC Redoxoma.

“Interestingly, we have shown the mechanism by which increased production of oxidants causes mitochondria to be less able to retain calcium and become more susceptible to oxidative injury. By correcting this defect with an antioxidant, the mitochondria of dietarily-restricted animals become the same as the control animals”, said Julian David Cualcialpud Serna, first author of the article. The study, published in the American Journal of Physiology-Renal Physiology, was part of Serna’s Ph.D.

The kidneys need a lot of energy to filter the blood, and since they have great metabolic activity, they have many mitochondria. However, according to Kowaltowski, there is little research on caloric restriction effects on this organ. “Our group investigates caloric restriction and what it does to mitochondria in terms of free radical production and also in terms of their core functions, such as oxidative phosphorylation, which generate ATP, and calcium transport. Some years ago, we saw that caloric restriction changes calcium transport in the brain and liver. Now we decided to look at the kidney and were surprised by the results”.

Calcium: far beyond the bones

What surprised the scientists was that, contrary to what happens in other tissues, the kidney mitochondria of lean animals generate more free radicals. In these mitochondria, the faster uptake of calcium was accompanied by increased production of hydrogen peroxide (H₂O₂), a potent oxidant. According to the researchers, higher rates of H₂O₂ liberation increase the propensity to calcium-induced permeability transition, as the process is associated with the oxidation of thiol membrane proteins. The effect was reversed with the use of the antioxidant dithiothreitol.

Known mainly for forming our bone structure, calcium (Ca²⁺, calcium ion) has several functions and is essential for our body, being present in the soluble form in body fluids, inside and outside cells. Calcium signals control processes such as muscle contraction, cell differentiation, and inflammation, among others. Furthermore, it is a central regulator of cellular functions, controlling metabolism in several aspects, for example, by regulating ATP production, glycogen breakdown, and the glycolytic pathway. Mitochondria capture and store calcium, maintaining its intracellular concentration at physiological levels.

“Mitochondria can respond to what is happening around them in several ways and one of them is by calcium signals. Taken up in small amounts and for short periods, calcium activates mitochondria. But, in excess, it leads to mitochondrial damage. And if the mitochondria are damaged, they stop working, stop producing energy for the cell, and the cells are damaged”, explains Serna.

In mitochondria, calcium uptake into the mitochondrial matrix occurs through the mitochondrial uniporter channel (MCU) and is driven by the inner mitochondrial membrane potential, which attracts positively charged species. The MCU is a complex formed by several proteins. One of them is MICU2, which behaves as a negative regulator. In the absence of this protein, calcium enters the mitochondria faster. “What we observed is that in animals under caloric restriction this protein is decreased and there is a higher calcium uptake rate. This is new, as we’ve never seen a diet change this protein in a physiological context”, said the researcher.

Thus, the renal mitochondria of caloric-restricted animals take up calcium faster than those of control animals but are damaged first, as their ability to store these ions is reduced. That means that animals under caloric restriction live longer and are healthier in many ways, but if they are prone to kidney damage for other reasons, having a slightly higher weight may be more favorable.

Calorie restriction and normal weight

The main point of caloric restriction, for Kowaltowski, is to prevent obesity, because in humans it is quite clear that obesity and overweight decrease life expectancy and increase age-associated diseases. But “caloric restriction is not an ultra-thin restriction, it is not an extreme. Having a normal weight is healthy, but a normal weight, not a low weight. The body mass index with the highest life expectancy is around 24, which is the upper limit of normality”, she says.

In the study, the researchers compared the kidney mitochondria of rats submitted for six months to a 40% reduced-calorie diet, without malnutrition, to a control group, which was fed ad libitum, i.e., rats that ate freely. The animals in the control group are sedentary and eat uncontrollably, becoming obese. “This 40% reduction seems a lot for a normal human being, but it is a reduction in relation to laboratory animals, which eat without limits and are really obese”, says the researcher.

Caloric restriction is a diet that reduces daily calorie consumption. It causes changes that impact various aspects of cellular and organism health, extending the lifespan of animals ranging from worms and flies to mice, rats, and primates, and preventing diseases associated with aging.

Previous studies

Kowaltowski’s group has been investigating the molecular mechanisms by which caloric restriction affects mitochondria and causes metabolic changes in the body for several years.

In 2016, in an article in the journal Aging Cell, the researchers showed that, in the brain, caloric restriction favors the mitochondrial calcium retention capacity, which results in a protection against excitotoxic damage related to the loss of neurons in pathologies such as stroke, Parkinson’s, and Alzheimer’s. In an article published in 2017 in the journal Free Radical Biology and Medicine, they showed that the diet protects mice’s liver against the mitochondrial permeability transition. And in another article in the journal Mechanisms of Aging and Development, they described how the diet promotes mitochondrial membrane remodeling, decreasing oxidized lipids and increasing levels and redistributing cardiolipin.

The article Regulation of Kidney Mitochondrial Function by Caloric Restriction, by Julian D. C. Serna, Andressa G. Amaral, Camille C. Caldeira da Silva, Ana C. Munhoz, Eloisa A. Vilas-Boas, Sergio L. Menezes-Filho, and Alicia J. Kowaltowski can be accessed here.