It seems that an enzyme is capable of increasing the size of the heart, in some cases benefiting the organism and in others damaging it. This may be good when the effect is temporary, as happens with those who exercise frequently, since this enzyme prepares the heart to send oxygen and nutrients to the body’s tissues even faster. However, this effect, if continuous, may weaken the heart and reduce its capacity to pump blood (the case of people who have chronic arterial hypertension) and lead to cardiac insufficiency, one of the main causes of death in Brazil.
By different biochemical paths, the enzyme FAK (focal adhesion kinase) proved to be necessary and sufficient for making the heart as much as double in volume, according to studies carried out by cardiologist Kleber Gomes Franchini and his teams at the State University of Campinas (Unicamp) and at the National Laboratory of Biosciences (LNBio) over the last 10 years. Two articles published in October in scientific journals, Nature Chemical Biology and Journal of Molecular and Cellular Cardiology, detail the action mechanisms of FAK and confirm these conclusions.
According to these studies, FAK makes the cardiac muscle cells, called cardiomyocytes, increase in size. “Initially, this effect results in a harmonic increase in the cardiomyocytes, which contract more efficiently. This is seen as an advantage in the heart response process, particularly in people with hypertension or some other problem,” says Franchini. “However, excessive activation of FAK may result in damage to the cardiomyocytes, even leading e to cell death.”
The researchers saw that this enzyme may also induce the multiplication of another type of heart cell, fibroblasts, which are smaller and more numerous than muscle cells. According to the studies carried out so far, the new fibroblasts migrate to the places left by the dead muscle cells. Subsequently, the fibroblasts form less elastic fibers, which may make the heart stiffen and harm its functioning. Franchini emphasizes that it may be decades before the negative effects of FAK are noticed.
“Kleber is a clinical investigator with a molecular view of diseases. He manages to explore the physiological, biochemical and molecular aspects. This is rare,” comments Mário Saad, a professor at the School of Medical Sciences at Unicamp, with whom Franchini has worked.
Now, in addition to studying the mechanisms of the action of FAK, the teams from Campinas are developing compounds that can hinder the action of this enzyme, particularly in fibroblasts. If they identify some substance that gets through all the tests carried out on laboratory animals and if it proves capable of acting effectively and with low toxicity in human beings, perhaps the researchers will help reduce the risk of cardiac insufficiency that results from heart hypertrophy. People with cardiac insufficiency suffer from weakness, shortage of breath during routine activities and a swelling of the body. In the most serious cases, even getting out of bed becomes difficult.
“The death rate within five years after the first symptom of cardiac insufficiency is around 40%, even with maximum and improved use of the best medication and therapeutic procedures,” says Franchini. “That’s why there is the expectation that increased knowledge of the mechanisms involved in the alterations that cause cardiac insufficiency may result in new drugs that alleviate the suffering and risk of death.” According to the Ministry of Health, cardiovascular diseases kill 300,000 people a year on average, i.e., 30% of the deaths caused by health problems in Brazil.
The heart of athletes
Expansion of the heart of an athlete and that of someone with hypertension may have the same beginnings (probably FAK) and intensify the production of various proteins in common, but the differences are now clearer. By measuring the expression of a type of RNA that blocks the action of proteins responsible for the increase in heart volume, Edilamar Oliveira and other researchers from the School of Physical Education and Sports of the University of São Paulo (USP) saw that the muscle cells of the heart of a person with hypertension accentuate the activity of proteins like beta-type myosin heavy chain, the atrial natriuretic factor and skeletal alpha actin, which do not appear in the hearts of trained rats.
Another difference is what is called physiological hypertrophy, which is typical of sportspeople, is generally reversible (the size of the heart of sportspeople returns to normal after some weeks without exercise) and is not associated with cardiac insufficiency; in other words, it is not harmful. Hypertrophy defined as pathological, on the other hand, which is seen in those who have some organic imbalance, is permanent.
As the limits between the two types of hypertrophy are not always clear, cardiologists frequently face delicate situations when they are examining the hearts of athletes. One of the first exams, the echocardiogram, detects the thickness of the walls and the functioning of the heart. A heart is considered normal when the thickness of its walls is between 9 and 12 mm; between 13 and 14 mm. lies a gray area, difficult to diagnose, and 15mm and above is even more difficult, because it indicates hypertrophy that may or may not be reversible.
To check, Patrícia Alves de Oliveira and other doctors from the cardiovascular and exercise physiology rehabilitation unit of the Heart Institute (InCor) of USP insist on athletes temporarily interrupting their training as a way of clarifying the possible cause of the hypertrophy.
“After four months without training, there is normally a reversal in the hypertrophy caused by excess exercise, but in pathological hypertrophy, there isn’t,” she says. Carlos Eduardo Negrão, director of the School of Physical Education and a researcher at InCor, observes: “Some sportspeople don’t accept the diagnosis and can’t stop, despite the risk they’re running.”
Today, Franchini believes that FAK acts as a heart sensor, preparing the muscle cells for situations of intense energy expenditure. This conclusion matured slowly. Discovered in 1992 simultaneously by three research groups in the US, FAK is a protein considered to be average in size, weighing 125 kilodaltons (dalton is the unit used for measuring protein mass).
It was initially believed to favor cell multiplication, but subsequent evidence indicated that it could do much more. In 1996, Franchini wanted to find out how the cells of hollow organs, such as the heart, detected those moments when they should expand to compensate for the increase in strain generated by intense physical exercise and by the hardening of blood vessels.
Perhaps there were sensors, but where could they be? This was when he read the article “Architecture of life” in the Scientific American magazine. Its author, US doctor Donald Ingber, currently at Harvard University, suggested that these sensors could be in areas in the cells where there was contact with the extracellular environment, called focal adhesion points. Franchini looked for and found a high concentration of FAK in the heart. The next step was to find out what the enzyme was doing there.
Franchini suspected that FAK could be the sensor he was looking for, but like a good son of Minas Gerais State, born in Uberaba, he kept quiet. He and another native of Minas Gerais, Mario Saad, head of a laboratory at the School of Medical Sciences at Unicamp, saw that the quantity of FAK at the focal points increased in response to a rise in blood pressure induced by an adjustable ring placed around the aorta of a rat. Afterwards, says Franchini, they created a device that stretched the muscle cells of the heart of rats, and the quantity of FAK increased by 100% after 12 continuous hours of stretching. In another experiment, they showed that without FAK, which was inhibited using specific compounds, the heart did not expand, even when it should have.
Little by little, the team from Unicamp brought together further evidence of the relevance of FAK in the muscle cells and fibroblasts of the heart. In 2007, Carolina Clemente, a researcher at LNBio, revealed the double role of this enzyme, which makes myocytes expand and fibroblasts multiply. The group showed that it was sufficient for causing these phenomena when, in a collaboration with José Xavier Neto, now at LNBio, they bred a mouse that expresses FAK only in the muscle cells of the heart.
The cardiac cells of these animals, with two or three copies more than normal of the gene that induces production of this enzyme, gained as much as 20% extra mass, a situation similar to that of the heart of an athlete, in which the enzyme exhibits the beneficial effect only on the muscle cells and not on the fibroblasts. “The prolonged action of FAK on fibroblasts may be harmful and irreversible,” says Ana Paula Dalla Costa, a researcher from Franchini’s group at Unicamp.
Within the muscle cells of the heart, FAKs remain inactive while they are anchored to myosin molecules, which function like columns that help to make muscle cells rigid. Muscular contraction, as a result of physical effort, makes the myosins stretch and release FAKs, which migrate to the cell extremities and activate proteins that act upon some 40 genes. These, in turn, induce the formation of proteins that make the cell more robust for facing up to situations that demand more effort.
In a recent study, published in October in Nature Chemical Biology, Aline Santos, Franchini and other researchers from LNBio, Unicamp and USP describe the stretch of FAK that activates it, making it break free from the myosin. Furthermore, they show that just this stretch, a fragment of protein, can act like the whole of the FAK, thus unleashing the processes that make the heart increase in volume.
The results of the experiments led the researchers to ask the question: if it were possible to turn off FAK or at least to reduce the amount, would it be good for the organism? “In principle,” says Franchini, “interfering in the process of pathological hypertrophy may be beneficial because it reduces expansion of the muscle cells and the fibrosis generated by the multiplication of fibroblasts.”
In 2010, the researchers from LNBio began to look for molecules that could block the action of FAK. They assessed around 40 and found one, identified by the code D5, which seems to have this effect. Franchini believes that D5 or another equivalent molecule, if they act only on fibroblasts and pass the subsequent effectiveness and toxicity tests, open up prospects of their future use for eliminating fibrosis in diseases such as hepatic and pulmonary cirrhosis and schistosomiasis.
Finding a way to deter FAK without damaging the organism is a delicate task. This enzyme is linked to dozens of proteins in many types of cells, in addition to those of the heart, and it has many functions, such as being involved in the proliferation, migration and survival of the cells, not only of healthy ones, but of tumoral ones as well. “These properties mean that FAK is very attractive as a potential therapeutic target,” says cardiologist, Wilson Nadruz Jr., a professor at the School of Medical Sciences at Unicamp, who took part in the studies.
Compounds are already beginning to appear that can deter the action of this enzyme, which is abundant in various types of tumor, such as those of the brain, breast, prostate and liver. A compound called TAE226, presented in 2007 by researchers from the United States, has proved capable of inhibiting FAK and, therefore, of checking the growth of tumor cells in the brains of mice. In August, in the International Journal of Cancer, researchers from Taiwan presented the first evidence that another molecule, SK228, curtailed the growth of tumors in cell culture and in animals, also because it inhibited the action of FAK.
1. Pathogenesis of cardiac hypertrophy and insufficiency: mechanisms activated by mechanical stimulus (nº 2006/54878-3); Modality Thematic Projects; Coordinator Kleber Gomes Franchini – Unicamp / LNBio; Investment R$ 1,158,498.59 (FAPESP)
2. Cell and functional bases of physical exercise in cardiovascular disease (nº 2010/50048-1); Modality Thematic Projects; Coordinator Carlos Eduardo Negrão – USP; Investment R$ 2,153,787.14 (FAPESP)
CLEMENTE, C.F. et al. Focal adhesion kinase governs cardiac concentric hypertrophic growth by activating the AKT and mTOR pathways. Journal of Molecular and Cellular Cardiology. Oct. 2011 (online).
FERNANDES, T. et al. Eccentric and concentric cardiac hypertrophy induced by exercise training: microRNAs and molecular determinants. Brazilian Journal of Medical and Biological Research 44, 9. p 836-84. Sep. 2011.
SANTOS, A.M. et al. FERM domain interaction with myosin negatively regulates FAK in cardiomyocyte hypertrophy. Nature Chemical Biology. Oct. 2011 (online).