Clockwork: Scientists Tinker with the Inner Workings of Circadian Rhythm

By Paul Nicolaus

December 9, 2019 | Living organisms are run by an internal, biological clock that anticipates the natural rhythm of a day. Made up of genes and proteins that interact during 24-hour cycles, this timekeeper controls everything from sleep and heart rate to metabolic and immune systems. But the human clock is increasingly being thrown off-kilter by our modern lifestyles—TVs in bedrooms, electronic books in bed, smartphones sitting on nightstands.

These types of technologies, and other disruptors like jet lag or third shift work, can wreak havoc on our bodily rhythm and, along with it, bodily processes. Circadian dysfunction has been linked to a higher risk of health issues like obesity, diabetes, and certain forms of cancer. Enter circadian rhythm research, a field of study that received heightened attention in 2017 when the Nobel Prize in Physiology or Medicine was awarded to a trio of American researchers for discoveries of the molecular mechanisms regulating this clock.

The hope is that an improved understanding of its inner workings could pave new pathways for repairing broken rhythms and improving health. For some scientists, this means targeting proteins that directly regulate the circadian rhythm. For others, it's figuring out the best time of day to deliver existing medications to improve outcomes and reduce side effects. In recent months alone, new findings from across the globe have added to the collective understanding of the body's day-to-day pattern and its relationship to the heart, gut, and brain.

The Role of Hormones and Diet

Different parts of the nervous system normally communicate by nerve tracts carrying electric potentials, but in the suprachiasmatic nucleus— a tiny region of the brain where clock genes are especially active—circadian information is sent to the cerebellum without a neuronal connection. It's a phenomenon that piqued the interest of researchers at the University of Copenhagen in Denmark.

To explore further, they removed the suprachiasmatic nucleus from rats, which removed their circadian rhythm. The researchers were then able to restore that rhythm by implanting a micropump and delivering doses of corticosterone (known as cortisol in humans) at times of day and night resembling the rats' natural pattern. Their findings, published Sept. 27 in Neuroendocrinology (doi: 10.1159/000503720), reveal that this stress hormone found in the blood helps control the circadian rhythm of brain cells.

"We normally regard the central nervous system as the master in the circadian regulatory hierarchy with the hormonal system acting in response to the nervous system to control body function," Martin Fredensborg Rath, a professor in the department of neuroscience, told Bio-IT World in an email interview. "However, in this case we show that the hormonal system can control daily rhythms in the brain."

Although rats are nocturnal creatures, humans essentially have the same hormonal system, and the discovery reveals that we have two systems—the nervous and the hormonal—that communicate and influence one another throughout a daily cycle. Furthermore, Rath points out that both stress and the medical use of glucocorticoids could interfere with the circadian system.

Meanwhile, other researchers have shown that our diet could alter our internal clocks and hormone responses. Led by Henriette Uhlenhaut, a team of researchers from Helmholtz Zentrum München, the German Center for Diabetes Research, Max Planck Institute of Biochemistry, Technical University Munich, and Northwestern University set out to understand the relevance of the daily peaks of stress hormone secretion and the role they play in the day-to-day cycles of metabolism.

Glucocorticoid stress hormones are produced each morning by the adrenal gland. The secretion of these hormones peaks before we wake, nudging the body to use fatty acids and sugar as energy to begin daily activities. The researchers used genomic, proteomic, and bioinformatic technologies to gain insight into the fluctuation of glucocorticoid levels during the daily cycle of liver metabolism by examining mice, some of which were given a high-fat diet.

The researchers were able to see how glucocorticoids—a group of natural and synthetic steroid hormones such as cortisol—control metabolism at different times of the day. Because glucocorticoids can control immune system activity, they are commonly targeted for the treatment of inflammatory diseases. A notable drawback, however, is that glucocorticoids can cause issues like obesity, fatty liver, hypertension, or type 2 diabetes.

To learn more, the team looked into the genomic effects that follow the injection of dexamethasone, a synthetic glucocorticoid. They discovered that the drug response differed in obese and lean mice, revealing that diet can alter hormonal and drug responses in metabolic tissues.

The findings, published Nov. 6 in Molecular Cell (doi: 10.1016/j.molcel.2019.10.007), suggest that lean and obese patients may respond differently to daily hormone secretion or steroid therapy. If the findings can be confirmed in humans, Uhlenhaut told Bio-IT World in an email interview, clinicians would need to consider this while treating patients.

The timing of glucocorticoid therapy taken by human patients, she added, "presumably has a different effect depending on the time of administration."

Daily Rhythm and Gut Immunity

Another group has zeroed in on the impact of circadian rhythm on the function of immune cells in the gut, which may help explain some of the gastrointestinal health issues tied to jet lag, shift work, and chronic sleep deprivation. The findings, published Oct. 4 in Science Immunology (doi: 10.1126/sciimmunol.aay7501) suggest that targeting clock genes could help fight off the harmful effects of intestinal illnesses associated with disruptions to the circadian rhythm.

The researchers identified an immune cell known as type 3 innate lymphoid cells (ILC3) that helps keep time within the digestive system and keep the intestine functioning in a healthy manner. Found throughout the intestines, the cells produce a cytokine called Interleukin-22 (IL-22), which stimulates the production of mucus, lipid transporters, and antimicrobial peptide. The cells also produce Interleukin-17 (IL-17), which protects against pathogenic bacteria in the gut.

This immune cell, ILC3, has large amounts of REV-ERB alpha, a clock regulating protein. Using genetically modified mice that lack this protein and healthy mice for comparison, the researchers studied the response to bacterial infection. The mice without REV-ERB alpha, they found, struggled to put up an effective defense in the gut. Knowing that REV-ERB has a circadian pattern of function, the researchers also performed experiments that put some of the mice on the type of schedule faced by a shift worker and observed reduced function in the ILC3 cells.

"There is less production of IL-22. There is less production of IL-17. And this can predispose to bacterial infection and fungal infection," Marco Colonna, a professor of pathology and immunology at Washington University School of Medicine in St. Louis, told Bio-IT World. It can also reduce the efficiency of absorption processes in the gut.

These cells are not just passive, according to Colonna. They are, rather, part of a system in which everything is coordinated. We know that the gut is tied to a daily cycle. "What we find is that these cells are also attuned to a daily cycle," he said, and they express molecules known to control the circadian rhythm. "So they basically coordinate their activity with the activity of the gut."

From Colonna's vantage point, circadian rhythms of the gut should be taken into consideration when choosing optimal timing for nutritional and pharmacological interventions for diseases of the gut. This temporal aspect, he said, creates a whole new dimension.

"We have identified one particular gene that is important in regulating the circadian function and the function in general of these cells," Colonna explained, but many others are waiting to be discovered and studied. Moving forward, his team plans to explore the impact of chronic disruption of the circadian rhythm and study the effect of drugs on the regulation of the gut's immune system.

Heart Healing Tied to Circadian Clock

Meanwhile, the circadian clock has been linked to heart health and healing from heart disease. In a paper published Oct. 3 in Nature Communications Biology (doi: 10.1038/s42003-019-0595-z), researchers reveal their efforts to target a component of the cellular clock mechanism, using medication to disrupt the expression of genes that lead to adverse immune responses following a heart attack.

During a heart attack, there is a limit of oxygen to a certain part of the heart. That portion of the heart begins to die, which causes an inflammatory response. That response leads to further cell death, additional damage, and scarring. Over time this can lead to incurable heart failure.

The researchers found that as little as one dose of the potential drug, called SR9009, limited the severity of damage following experimentally induced heart attacks in mice. The discovery could open the door to the use of circadian medicine therapies to heal heart attacks and prevent the later development of heart failure.

"We were amazed to see how quickly it worked, and how effective it was at curing heart attacks and preventing heart failure in our mouse models of the disease," Tami Martino, a professor in the department of biomedical sciences at the University of Guelph in Ontario, Canada, said in a news release. The discovery could wind up helping with other heart therapies that involve early adverse inflammatory response like valve replacement or transplant.

More broadly, it could be useful with the treatment of other events that spark adverse inflammatory responses, the researchers pointed out, such as strokes or traumatic brain injury.

Implications for Drug Delivery and Discovery

"Since we've learned how to target some of these circadian components that are responsible for what we call our clock proteins—since we can regulate some of those with drugs—we can kind of hijack the system" to suppress it when it's overactive, explained Thomas Burris, a professor in the departments of anesthesiology and genetics at Washington University School of Medicine in St. Louis and a professor and vice president of research at St. Louis College of Pharmacy.

This applies to heart attacks, but there are all kinds of problems with overactivity concerning autoimmune diseases, too, according to Burris, who is a co-author of both the heart and gut immunity papers detailed above. The compound that worked in the heart model also works in models of multiple sclerosis, for example, "so you can calm down the immune system from when it's set too high based on targeting pieces of it that normally regulate time of day," he told Bio-IT World.

The compound published in the heart attack paper is a first-generation treatment targeting a particular receptor, but the next step is to design compounds that are much more drug-like. The advantage of having the current compound in place is that it has allowed the researchers to explore what diseases they ought to focus on moving forward. Therapeutics to reduce damage after a heart attack would be valuable, he pointed out, but there are other areas of possibility to consider, too.

"We're looking at neuroinflammatory models where there's a lot of inflammation in the brain in diseases like Alzheimer's," he said. The drugs have promise, but "we just don't know quite the right fit." There is a need to balance potential side effects with benefits, and that balance changes according to the disease. "So that's where we're headed," he added. "We're trying to design better compounds that may actually be able to become drugs."

Paul Nicolaus is a freelance writer specializing in science, nature, and health. Learn more at www.nicolauswriting.com.