In her PhD, Violeta Heras studied the biological mechanisms behind the link between early-onset puberty and infantile obesity in female rats.

Animals have always fascinated Violeta Heras. As a child, questions like “how do they interact with the environment?”, “how do they adapt to different conditions?” started to pop up in her mind. Later on, when she was about to decide what to study in university, this early fascination kicked in again. But she recognized what interested her wasn’t the macroscopic part of the animals. The microscopic world, meaning the cells and molecules inside the animals, interested her the most. She then decided to study Biotechnology and Molecular Biology. During her Bachelor’s studies, she had the opportunity to do an academic exchange in Belgium. There she worked for the first time in a lab. This experience made her realize that she liked doing science. And after another lab internship during her Master’s, the decision to do a Ph.D. in Biomedicine came naturally.

In her doctoral studies at the University of Cordoba, in Spain, the scientist specialized in the field of neuroendocrinology. This branch of biology studies how the brain influences the action of hormones throughout the body. One example of this is how puberty and sexual maturity are controlled in mammals. During this time, the brain increases the production of a hormone called gonadotropin-releasing hormone (GnRH). This hormone acts in a gland located at the base of the brain, the pituitary. Hormones produced by this structure then lead the ovaries in girls and the testicles in boys to produce steroid hormones: estrogen and progesterone, and testosterone, respectively. These will ultimately lead to sexual maturation. Such process is affected by external stimuli. One example of that is what one eats and the impact of that on metabolism. “It is well established that pubertal activation of the reproductive axis and maintenance of fertility are critically dependent on the metabolic state of the organism,” explains Violeta.

On average, puberty happens between the ages of 8 and 13 in girls, and 9 and 14 in boys. Signs of sexual maturity, e.g. pubic hair, breast and genital development, that occur before the ages of 8 in girls and 9 in boys fall in the definition of precocious puberty. And this situation is becoming more and more common. For example, comparing data from 1977 until 2013, a study observed that girls started to develop breast tissue, a sign of puberty, one year earlier in 2013, compared to the 70s. But why is this happening? Why are children biologically becoming adults earlier?

The scientific literature has pointed to obesity as one of the contributing causes. According to the World Health Organization (WHO), in 1975, under 1% of children and teenagers between 5 and 19 year old were obese. In 2016, 6% of girls and 8% of boys were diagnosed with the condition. And both issues, childhood obesity, and precocious puberty are associated with health problems later in life. “The escalating prevalence of child obesity parallels that of alterations in pubertal timing, also linked to deleterious consequences in our health such as cardiovascular, cancer, or physiological risk later on in life,” explains Violeta. Thus, studying how early-onset puberty and childhood obesity relate is a matter of young and adult public health.

One way to study this association is to do lab experiments with rodent models that mimic obesity. Both rodents and humans share brain regions and hormones that regulate feeding and metabolism. And, indeed, obesity correlates with advanced puberty not only in humans but in laboratory rodents as well. Animal research, though, has the advantage of performing invasive experiments that wouldn’t be possible in humans. Also, it allows better control of different parameters that influence the results. Thus, using rats as her animal model, Violeta studied what could explain the relationship between infantile obesity and early onset puberty.

As mentioned before, puberty is largely regulated by hormones produced by the brain and different glands. But the Spanish scientist was interested in an alternative type of molecule, that hadn’t been explored before: ceramides. In summary, ceramides are lipid molecules present all over the body. They are involved in many biological processes, working as a signal from one cell to another. When she started her project, Violeta already knew that ceramides could work in the control of energy balance. Back then, recent reports suggested that ceramides could interfere with hormones involved with metabolism. Also, increased levels of ceramides were observed after overfeeding. But all of these were only investigated in adult rodents. What are the levels of ceramide in younger animals? Do they also correlate with obesity/overfeeding? Do they interfere with puberty? These were the questions that Violeta’s study addressed.

To answer these questions, first, Violeta and her colleagues fed female rats with a high-fat diet. When the animals were close to the puberty age, they started to look for signs of puberty, until the expected pubertal age. Once they reached puberty, they also measured the amounts of ceramides in a brain region called hypothalamus. More specifically, the paraventricular nucleus of the hypothalamus (PVN), which is related to the control of feeding and also has a connection with the ovaries. “As we expected, most of the overfed animals showed advanced puberty. Importantly, when we analyzed the hypothalamic levels of ceramides in these animals, we observed a significant increase in the concentration of total ceramides”, says Violeta. Thus, as observed in adults by other studies, the amounts of ceramide correlated with obesity in puberty. In addition, overfeeding came together with an early onset of puberty.

But what happens if ceramides are increased or decreased in the brain, without overfeeding the rats? Do they also affect puberty onset? To answer that, the team injected two drugs into the brain, one at each time. One of them increased ceramides levels, the other decreased. Interestingly, increased ceramide amounts correlated with early signs of puberty. But this didn’t affect the body weight. By contrast, decreasing ceramide levels led to a significant delay in puberty onset, without changing body weight. So it seems like ceramides by themselves could affect puberty, but they are likely not the cause of obesity. Instead, the results suggest the increased amounts of ceramide are a consequence of obesity. And as such, they could lead to early-onset puberty.

Then how exactly ceramides affect puberty? Do they affect hormones or other molecules that regulate puberty? This was their next question. For this, they focused on one of the small proteins that induce the start of puberty, kisspeptin. Using that same drug that decreased ceramide levels together with kisspeptin, they saw a delay in puberty onset. It’s like without ceramides kisspeptin couldn’t act as usual (initiating puberty). What suggests an interplay between ceramides and kisspeptin. The same effect of the ceramide-blocking drug wasn’t observed on other pubertal hormones. Violeta and her colleagues then bet that ceramides, possibly with kisspeptin, act through an alternative pathway.

And, according to their results, they probably bet right. As mentioned before, hormones are critical for the control of sexual maturation. But, at least in females, hormones might not be the only ones affecting the production of steroids by the ovaries during puberty.  The sympathetic nervous system, the part of the nervous system that regulates automatic physiological responses, may also play a role. To cut a long story short, in this case, not hormones, but another type of molecule is the one in charge: the neurotransmitter norepinephrine. “We determined norepinephrine (NE) and its metabolite […] content as markers for this sympathetic activity in obese versus lean immature female rats […]. In line with our hypothesis, NE and its metabolite levels were significantly increased in […] the ovary of [obese] female rats [at puberty] […] compared to their control counterparts”, explains Violeta. Also, when the ceramide reducing drug was used, the ovarian levels of NE in young obese females came partially back to normal. Thus, these results could suggest that ceramides may influence the ovaries by sympathetic activation. But the study didn’t measure the sympathetic neuronal activity itself. Neither evaluated the sympathetic ovarian innervation. Thus, further studies are needed to establish this association.

Thus, based on the article published last year by Violeta and her colleagues, ceramides produced by the PVN of the hypothalamus may be one of the links that connect early-onset obesity and precocious puberty in female rats. And that ceramides possibly do this via interactions with the puberty activator, kisspeptin, and with the sympathetic nervous system. The study, however, was mostly performed in females, so the effect of ceramides in males should be addressed in the future to check if the findings apply to both sexes. Also, more studies are needed to understand if the results obtained in rats can be extrapolated to humans.

Given the growing concerns on childhood obesity and precocious puberty, and their impact later in health, studies like Violeta’s are very important nowadays. First of all, we need to understand what is going on in the body, to later come up with preventive or therapeutical solutions for these issues. “This role of central ceramides and its alternative pathway could explain different forms of advanced puberty in childhood obesity not associated with high levels of gonadotropins. Our results may help to define better strategies for the management of pubertal disorders associated with metabolic disease, specifically in girls”, completes Violeta. Thus, hopefully, more and more studies like hers will help understand all the variables that regulate puberty and becoming an adult.

Today, Violeta is a postdoc researcher at Santiago de Compostela University, in Spain. And she studies the role of one type of cell present in the hypothalamus, called tanycytes, in metabolic and neurodegenerative disorders. Besides science, in her free time, Violeta loves spending time with her friend, cooking, watching movies, and traveling around the world.

Thank you very much, Violeta, for sharing a bit of your story and your work. All the best in your career. Keep up the good work!