LA JOLLA, Calif.—The metabolic protein AMPK has been described as a kind of “magic bullet” for health. Studies in animal models have shown that compounds which activate the protein have health-promoting effects to reverse diabetes, improve cardiovascular health, treat mitochondrial disease and even extend lifespan. But how much of the effects of these compounds can be fully attributed to AMPK versus other potential targets is unknown.
Now, Salk Institute researchers have developed a new system that lets them study in more detail than ever exactly how, where and when AMPK carries out its molecular and therapeutic functions. In the paper, published January 2 in Cell Reports, the Salk team uses the new model to activate AMPK in the livers of adult mice with fatty liver disease.
“This model will allow us to answer questions that scientists could not answer before,” said Salk professor and director of the Salk Cancer Center Dr. Reuben Shaw, who led the new work. “It really gives us a new way to define the health benefits of this specific enzyme in a wide variety of diseases.”
AMP-activated protein kinase, or AMPK, is known as a master regulator of metabolism. Cells activate AMPK when they are running low on energy, and AMPK is activated in tissues throughout the body following exercise or during calorie restriction. In response, AMPK alters the activity of many other genes and proteins, helping keep cells alive and functioning even when they’re running low on fuel. In different tissues throughout the body and at different time points in development, AMPK likely has varying effects. Until now, the only way to study the specific impact of genetically increasing AMPK activity was to change its activity in an organism for its entire life, starting at embryogenesis.
“When AMPK is overactivated from the very beginning of embryogenesis, we don’t know what effects it’s having on normal development,” said Daniel Garcia, a senior research associate at Salk and first author of the new paper.
Garcia, Shaw and their colleagues enabled a mouse to have a special version of AMPK that lets the researchers activate the gene by feeding the adult mouse an antibiotic. By double-engineering the inducible AMPK gene in the mice, researchers can also control where in the body this AMPK activation happens—everywhere, or just in a select tissue or tissues.
“The model we’ve developed is much more similar to what you would see in a clinic if you target AMPK with drugs,” Garcia noted.
According to Shaw, who holds the William R. Brody Chair, “This system allows for both tissue-restricted expression (as did previous models), but also complete inducibility and the ability to turn the genetically encoded activated AMPK back off at will. This allows one to keep activated AMPK for days—or weeks to months—at any point after adulthood. Previous models simply recombined on a different type of activated AMPK, but then one could not turn them off or even turn them on at a specific point after another treatment (e.g., after a high fat diet or before a high fat diet), etc. This is a major advance in the ability to study the predicted effects of turning on this beneficial enzyme for different amounts of time in different tissues, in disease models of all kinds of diseases in the mouse.”
“The tissue distribution is controlled via use of a Cre recombinase under the control of a tissue-specific promoter, which then drives an artificial transcription factor,” Shaw continues. “The DNA binding element for that artificial transcription factor (Tet activator or Tet repressor) is then placed in front of of activated AMPK cDNA placed at a ‘safe harbor’ locus in the mouse genome.”
To test the utility of the new model, the researchers developed mice that could have AMPK activated in the liver. Then they fed a subset of these mice high-fat diets leading to diet-induced obesity and an excess accumulation of fats in the liver. This condition is equivalent to nonalcoholic fatty liver disease (NAFLD) in humans, the leading form of chronic liver disease in American adults.
In both mice with and without NAFLD, levels of fats in the liver dropped when AMPK was activated—new fat production was slowed, and existing fats were metabolized. When AMPK was activated in mice that were fed a high-fat diet, the mice were protected against weight gain and obesity, and had fewer signs of liver inflammation.
“This paper confirms that AMPK is a good target for treating NAFLD,” added Garcia. “It’s further confirmation that AMPK activators should be tested clinically.”
In addition to the effects on liver fat, AMPK activation—even though it was limited to the liver—also lowered levels of fats elsewhere in the body, suggesting that hormones released by the liver into the rest of the body were affected.
“These results indicate that AMPK could potentially be a powerful treatment to a host of diseases in humans,” says Shaw.
The researchers next plan to study AMPK activation in a plethora of other tissues, including muscles, where scientists have hypothesized AMPK could have a dramatic effect.
“We are making these mice available through Jackson Labs so that researchers worldwide can breed them into their favorite disease models or text the effects in a wide range of diseases, from cancer to diabetes, as exercise mimetics, lifespan extension or treatments for mitochondrial disease or muscle degeneration,” mentions Shaw. “Such experiments coupled with recent advances in small molecules activating AMPK in an on-target manner with good rodent bioavailability should lead to new hypotheses about where and when AMPK activation may have its greatest therapeutic benefit.”
“We are excited to examine how systemic activation of AMPK in the whole body of adult mice might compare to liver or muscle restricted AMPK activation in specific disease models. We look forward to seeing how other researchers use the mice to explore new worlds of possibilities, from immune modulation to expansion of stem cell homeostasis. The future is wide open,” finishes Shaw.