Engineered Hot Fat Implants Reduce Weight Gain in Mice

UC Berkeley scientists have discovered a new way to engineer the growth and expansion of energy-burning “good” fat. Subsequently they showed that this fat helped reduce weight gain and lower blood glucose levels in mice. According to Andreas Stahl, the senior author of this study, this technique could potentially lead to new approaches to combat obesity, diabetes and other metabolic disorders.

Stahl and his coworkers devised a specifically tailored hydrogel that acted as a scaffold for stem cells that are able to form brown fat. Implantation of this stem cell-laced scaffold could form a functional brown-fat-like tissue. White fat, which is associated with obesity, stores excess energy, but brown fat is a heat generator and burns calories in order to heat your body.

“What is truly exciting about this system is its potential to provide plentiful supplies of brown fat for therapeutic purposes,” said study lead author Kevin Tharp, a doctoral student in the Department of Nutritional Sciences and Toxicology. “The implant is made from the stem cells that reside in white fat, which could be made from tissue obtained through liposuction.”

“This is figuratively and literally a hot area of research right now,” said Stahl, who is an associate professor of nutritional sciences and toxicology. “We are the first to implant in mice an artificial brown-fat depot and show that it has the expected effects on body temperature and beneficial effects on metabolism.”

Several studies have shown that cold temperatures can increase the metabolic activity of brown fat. However, Stahl pointed out that exposure to cold usually leads to increases in food intake, as well, which potentially negating any calorie-burning benefits from brown-fat activity.

There are three basic types of fat tissue in our bodies. These include energy-storing white fat that many of us are most familiar with, and two kinds of energy-burning fat used to generate heat, which include brown fat, which arises during fetal development, and beige fat, which is brown-like fat that is formed within white fat tissue after exposure to cold and other stressful situations.

For this experiment, Stahl teamed up with Kevin Healy, a UC Berkeley professor of bioengineering. The goal was to increase brown-like beige fat without exposing the animals to cold temperatures. Stahl and Healy wanted to develop a system of physical cues to guide stem cell differentiation.

“It’s already known that for a number of organs, including the heart, the extracellular matrix in which a cell resides provides signals to guide growth and development,” said Dr. Healy. “We applied this concept to stem cells isolated from white-fat tissue.”

The specific matrix recipe for converting white-fat stem cells to brown fat was quite unclear but Stahl and Healy noted that previous studies suggested that stiffness of the surrounding environment was a factor. If white-fat stem cells are placed in a 3D environment that is soft, with little resistance, they become fat. However, if the surrounding environment is rather stiff, the stem cells grew into bone.
Healy and his postdoctoral research fellow Amit Jha generated a tightly knit 3D mesh that consisted of hydrogel, water, hyaluronic acid and short protein sequences associated with brown-fat growth and function. Hyaluronic acid is a naturally occurring acidic carbohydrate that helps make water thicker and gel-like (stiffer in other words). They then took white-fat stem cells (adipose tissue-derived multipotent stem cells or ADMSCs) from mice that had been genetically engineered to express an enzyme from fireflies that made the cells luminescent. Because these cells glowed when incubated with the right substrate, they could be easily traced.

These ADMSCs were then added to the hydrogel and, before the mixture thickened, injected them under the skin of genetically identical mice.

The gel polymerized after injection and stiffens up under the skin of the animal. These labeled cells were then monitored to determine how well they stayed put, how long they persisted in the body and whether they were metabolically functional.

Stahl and Healy and their colleagues noticed an increase in the core body temperature of the mice at ambient temperatures of 21 degrees Celsius and after 24 hours at 4 degrees Celsius. In both cases, the mice with the implanted cells were up to half a degree Celsius warmer than a control group of mice with no injection. The higher the concentration of cells, the larger the effect on temperature.

Next, they put these experimental mice on a high-fat diet. By the end of three weeks, the mice with injected beige fat gained half as much weight and had lower levels of blood glucose and circulating fatty acids compared with control mice.

“This is a feasibility study, but the results were very encouraging,” said Dr. Stahl. “It is the first time an optimized 3D environment has been created to stimulate the growth of brown-like fat. Given the negative health effects of obesity, research into the role of brown fat should continue to see if these findings would be effective in humans.”

The study was published Aug. 20 in Diabetes.

Spanish Team Develops Anti-Obesity Treatment in Animal Models

A research team from the Spanish National Cancer Research Center (CNIO) has shown that partial pharmacological inhibition of the PI3K enzyme in obese mice and monkeys reduces body weight and physiological manifestations of metabolic syndrome, specifically diabetes and hepatic steatosis (fatty liver disease), without any signs of side effects or toxicities. They published their work in the journal Cell Metabolism. This collaborative project between the Tumor Suppression Group headed by Manuel Serrano at the CNIO (Madrid, Spain) and the Translational Gerontology Branch headed by Rafael de Cabo at the U.S. National Institute on Aging, National Institutes of Health (NIH, Baltimore, MD, USA), included the participation of the NeurObesity group of CIMUS led by Miguel Lopez at the University of Santiago de Compostela (Santiago de Compostela, Spain).

PI3K (phosphatidylinositol-3-kinase) is the name of an enzyme that regulates the balance between the biosynthesis of cellular components and the burning of nutrients to make energy in cells. Specifically, PI3K promotes cellular growth and biosynthesis, which can lead to the induction of growth and multiplication of cells, and ultimately could lead to cancer.

For this reason, scientists who investigate cancer have has a long-standing interest in designing pharmacological inhibitors of PIK3. CNIO, in fact, has developed its own experimental inhibitor, CNIO-PI3Ki, which is being studied for applications as a cancer treatment in combination with other compounds. As part of the characterization of the PI3K inhibitor and to understand how it affects the balance between the use and storage of nutrients in the body, the Serrano team decided to study the effects of CNIO-PI3Ki on metabolism.

“At this point we have veered away from the original anticancer aspects of these inhibitors. In our previous studies, we had seen that one of the normal physiological functions of the PI3K enzyme is to promote the storage of nutrients. We found this to be of particular interest because it is precisely this type of manipulation, regulation of the balance between storage and use of nutrients, that is sought after in treating obesity,” explains Ana Ortega-Molina, the first author of the study, who is working at the Memorial Sloan-Kettering Cancer Center in New York.

To test the effect of their PI3K inhibitor on metabolism, CNIO scientists administered small doses of the CNIO-PI3Ki inhibitor to obese mice for 5 months while those mice were fed a high-fat diet. During the first 50 days, the obese animals lost 20% of their body weight, at which point their weight stabilized. The treatment was administered for 5 months and during the whole time, these mice maintained a stable weight (20% below the weight of non-treated obese mice), despite continuing feeding with a high-fat diet. They also improved their pathological symptoms of diabetes (high glucose levels in the blood) and hepatic steatosis (fatty liver).

“When it comes to obesity, constant weight loss can be extremely dangerous. The ideal solution is to alter the balance between the use and storage of nutrients, to strike a new balance in which there is greater use and less storage,” explains Elena López-Guadamillas who, in collaboration with Ana Ortega-Molina, carried out most of the experimental work. This study showed that the drug had no side-effects and did not produce irreversible effects on metabolism, which is also desirable for its possible future use as a treatment in humans.

In non-obese animals that were fed a standard diet, the administration of the drug had no effect, which is another indication of its safety. “This shows that the activity of the PI3K enzyme is only relevant when there is an excess of nutrients, that is, a high-calorie or high-fat diet,” adds López-Guadamillas.

CNIO scientists then collaborated with the U.S. National Institutes of Health (NIH) in order to test the CNIO-PI3Ki compound on obese monkeys (macaques).  They used a very low dose to ensure higher safety margins, but even with these very high doses, the daily treatment of these obese animals over a 3-month period reduced the total amount of fatty tissue by 7.5% and improved the symptoms of diabetes.

Obesity is one of the most important risk factors within the spectrum of serious diseases that constitute the metabolic syndrome. Many pharmacological agents have been discovered that lead to weight loss, but these drugs often have unacceptable toxic effects (partly due to the fact that these previous agents act on the brain centers that control appetite). In this respect, CNIO-PI3Ki seems to be the exception, at least in animal models thus far, as no such side-effects have been observed, even after long-term treatments (5 months in mice and 3 months in monkeys).

A series of safety characteristics that have been demonstrated in mice is shown below:
1) Selective: CNIO-PI3Ki only produces weight loss in mice that receive an excess of nutrients and not in mice that eat a normal balanced diet. This shows that PI3K plays an important role in the storage of nutrients when food intake is excessive, but is not so important under a normal diet.
2) Weight loss in the mice is due exclusively to loss of fatty tissue; no losses occur in other tissues such as liver, muscle or bone.
3) It does not affect the brain: CNIO-PI3Ki does not cross the blood-brain barrier.
4) It does not affect the hypothalamus: The hypothalamus is a specialized structure of the brain that is exceptional because it lacks a blood-brain barrier (a structure that controls the entrance of substances from the blood to the brain) and it controls many metabolic processes, including appetite and satiety. No effects on the main neuropeptides produced by the hypothalamus related to appetite and satiety have been noted in the mice. These last studies have been carried out in collaboration with the research group led by Miguel López at the University of Santiago de Compostela.
5) It works on a long-term basis: The effects of CNIO-PI3Ki were maintained over at least a 5-month period of treatment in mice, which suggests that resistance mechanisms are not developed. This is very important, as it is a common problem found in other compounds that affect metabolism.
6) Reversibility: The effects of CNIO-PI3Ki were reversible, which means that when the treatment was interrupted and a high-fat diet maintained, the mice regained weight. This indicates that CNIO-PI3Ki does not cause irreversible changes.

The next logical step, once the beneficial effects of CNIO-PI3Ki have been demonstrated in obese mice and monkeys, is to perform clinical trials on humans. “The leap from animals to humans is complex, expensive and full of uncertainties. Many treatments that are promising in animals turn out not to be effective in humans or toxicities appear that were not observed in animals. But, obviously, in spite of the uncertainties, we have to give it a try,” says Manuel Serrano. “Clinical trials require large investments and are undertaken with the aim of marketing a treatment. We are very optimistic about the possibility of entering into an agreement soon with a multinational pharmaceutical company interested in carrying out clinical trials with CNIO-PI3Ki to treat obesity and metabolic syndrome in humans,” says Serrano.