How does the circadian clock change when we get older? This is the question that has driven research associate Guiomar Solanas and “la Caixa” PhD student Francisca Oliveira Peixoto, both at IRB Barcelona, to study the daily pattern of gene activity. The answer, as laid out in two studies headed by Salvador Aznar Benitah and published in the journal Cell, rejects the widely held belief that stem cells lose their circadian rhythm with age. The studies demonstrate that the daily pattern of gene activity is not lost with age but is rather reprogrammed for new functions and that a low calorie diet delays alterations in the rhythm of stem cells.
Francisca’s pursuit of circadian rhythms brought her to the Stem Cells and Cancer Lab at IRB Barcelona. “I had read two articles the group had published before I arrived and I was fascinated by the circadian clock. By the time I got here, they were already thinking of tackling a project on ageing. They had previously studied the circadian clock in the basal state and then in skin stem cells. The next step was to give a fresh perspective to this clock. Circadian rhythms are involved in ageing and it had always been believed that they are altered during this process, but nobody had described how. We followed this line of reasoning,” Guiomar says. The missing link was diet. “It is known that calorie restriction extends life and delays the development of diseases during ageing and we wanted to see what effect calorie restriction had on the circadian clock”.
“It was known that ageing interferes with oscillations in the electrical activity of neurons in the suprachiasmatic nucleus—the brain region responsible for regulating the circadian clock—making these oscillations lose amplitude, and it was assumed that other tissues would also lose their rhythms,” explains Guiomar. But was this actually the case? By studying the stem cells in two types of tissue, namely skin, muscle, and in liver, in young (three months old) and aged (18 to 22 months old) mice every four hours, “we observed that the core components of the circadian clock had no changes in their rhythm with age”. How could that be? “The circadian clock was not regulating the functions associated with tissue homeostasis but rather functions linked to the specific stress experienced by each kind of tissue,” Francisca explains.
And what about the diet? In other experiments, Guiomar and Francisca gave mice a low-calorie diet for six months and compared their circadian clock with that of counterparts on a normal diet. The animals on the low-calorie diet conserved most of the rhythmic functions of their youth. The researchers thus concluded that such a diet delays alterations in the rhythmic functions of stem cells and slows down ageing.
“These results were totally unexpected,” recalls Francisca. “When we started the first experiments we thought we would find some kind of loss of rhythm. It was surprising to see that there were in fact many oscillations and that some of them were specific to aged animals,” explains Francisca.
And why had this not been discovered until now? “These are very complex studies that require a lot of time and resources, and we have been working on them for more than five years. The experiments go on for a long time and over many hours because you have to collect samples six times over the course of a day”. A project this size calls for perseverance. Francisca goes on to say, “You have to accept that things don’t always go as planned and not let this affect your enjoyment of what you are doing. Sometimes you don’t lose four samples but fifty. And this is the big difference, because we spend months collecting samples without knowing what results we are going to get or whether things are going to work or not.”
In the near future, “The project will go on to tackle metabolism: how metabolism instructs the circadian clock to regulate which genes. We want to unravel why the same components of the circadian clock in an adult regulate one set of genes while in an aged adult they regulate another set. This change fascinates us. We know that metabolism is involved and we want to determine the exact role of this process and how it triggers this change,” says Guiomar.
A project like this calls for a great deal of organisation and the capacity to foresee problems, to plan, and to know exactly what will be needed in a month’s time. The key to success in this case, has been a true joint effort of the entire lab and the support of IRB Barcelona’s Core Facilities, as well as the active involvement of researchers at the Pompeu Fabra University and Irvine University, in California.