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Heart muscle cell 1.0 and 2.0 – two epigenetic programmes in one cell

All the cells in an organism have to adapt to changing requirements as they develop and grow - including muscle cells in the heart. Crucial to this process are the cells’ growth in size and epigenetic factors that play a role in modulating the expression of various genes. The role of epigenetics in cancer development has been the focus of research for quite some time. The question is, what role do epigenetic factors play in the development of the heart? Prof. Dr. Lutz Hein from the Institute of Pharmacology and Toxicology at the University of Freiburg studies the maturation of cardiac muscle cells and the epigenetic programmes involved. One of his goals is to better understand cardiac diseases.

The heart is the first organ to form in the growing embryo. It beats continuously from as early as the fifth week of pregnancy until death to supply the body with oxygen and nutrients. The relatively large muscle cells of the heart develop from precursor cells that initially, as the heart grows rapidly, still have the ability to divide. However, shortly after birth, most heart muscle cells lose this ability and have to adapt differently to changing physiological conditions. This means that heart muscle cells lose an important repair mechanism that cells in other body tissues maintain for life.

"After a heart attack, only a few heart cells are able to regenerate through cell division," explains Prof. Dr. Lutz Hein from the University of Freiburg. “Following disease, most of the heart cells have to find other ways to adapt to changing conditions and repair damage. Conceptually, this is a particular challenge as damage needs to be repaired while the heart is beating. “Although many more people survive myocardial infarction than was previously the case due to progress in medical technology, it leaves many survivors with chronic heart failure or high blood pressure. Hein believes that therapeutic improvements can only be achieved on the basis of fundamental insights into how cardiac muscle cells work and how they regenerate.

Schematic showing the development of a cardiac muscle cell into an adult organ, and later into a diseased heart. The schematic also shows how methylation changes.
During embryonic development, heart muscle cells undergo a maturation process (top) which is epigenetically controlled by DNA (bottom). As the cells mature, methyl groups (CH3) are removed from the DNA, thereby enabling transcription factors (TF) to access and transcribe the genes. In people with chronic heart failure, pathological growth is governed by proteins that recognize DNA methylation (e.g. MeCP2). © Prof. Dr. Lutz Hein, University of Freiburg

Epigenetics affects function

All cell types in the human body have the same DNA. “However, an immune cell chooses a different programme from a liver cell, or to put it differently, it chooses a different chapter in the maturation book,” says Hein. Some DNA areas are of no interest at a specific point in time and must not be transcribed. Which areas happen to be active at a certain point in time also depends on factors closely related to the DNA, such as histone modifications and DNA methylations. These epigenetic factors determine the activity of a certain gene, and hence cell function, and have become a major focus of scientific attention. However, the role of epigenetics in the development of diseases such as cardiac heart failure is still poorly understood. Before birth, a heart beats under different oxygen and nutrition conditions than after birth. The transition from pre- to postnatal life requires adjustments that allow the heart to grow while contracting optimally. Before birth, cells produce energy from glucose; after birth, energy is mainly produced from fatty acids. This requires cardiac muscle cells to carry out different tasks simultaneously, and they effectively do so by activating different epigenetic programmes. A metabolic shift generates different metabolites and different components are incorporated into the contractile cell apparatus consisting of actin and myosin filaments.
Schematic showing how protein composition of the contraction apparatus changes from foetal to adult cardiac cell.<br />
Different protein composition of the contraction fibres of a heart muscle cell before and after birth, enabling the heart to adapt optimally to different oxygen and nutrient conditions. The proteins that are used until birth (left) are switched off (red) after birth because the underlying DNA (i.e. genes) is methylated (i.e. genes). Proteins that are needed after birth (blue) start functioning as a result of DNA demethylation. © Prof. Dr. Lutz Hein, University of Freiburg

DNA methylation is a dynamic process

Hein and his team are investigating the epigenetic switches that trigger the cardiac gene programme during birth. They found that the birth process is not responsible for the switch in cardiac programme; the observed changes can occur earlier in some people and later in others. In order to understand why this is so, Hein and his team focused specifically on the cellular control centre, i. e. the nucleus. Hein has developed a method to isolate cardiac muscle cell nuclei from murine heart tissue and map the DNA methylation of cardiac muscle cells. The researchers initially focused on the methylation of DNA, which is one of the most important epigenetic mechanisms for the regulation of gene activity. They found large sections of DNA that were methylated differently during the development and maturation of cardiac muscle cells. The promoter regions of specific genes have methyl groups that prevent the transcription of such genes, thereby preventing the DNA from being transcribed into RNA and the protein being synthesized. The gene is in effect switched off. As Hein says, “the genes only function if they are cleaned.” The scientists found that an increasing number of methyl groups were removed as the cardiac muscle cells of mice mature. “The methyl groups are removed from the DNA after birth,” says Hein. This leads to the formation of isoforms of the prenatal proteins, which have a similar function but a slightly different metabolic effect. At the same time, the expression of foetal proteins decreases as the relevant genes are methylated. Hein has already discovered the mRNAs of these protein isoforms. “We could say that there is a 1.0 heart muscle cell and a 2.0 heart muscle cell,” he jokes.

Role of the methyl transferase enzymes

Cell nuclei isolated from heart biospies, labelled with fluorescent antibodies and purified by sorting. DNA and RNA are isolated and sequenced. © Prof. Dr. Lutz Hein, University of Freiburg

In the mouse model, Hein was able to switch off two enzymes that mediate the methylation of the genes coding for foetal proteins and found that two cardiac muscle protein isoforms were in fact present when methylation was prevented. “When methylation is prevented, some of the foetal cardiac muscle genes remain functional. With this model, we showed that in foetal proteins there was a direct link between methylation and gene expression.” Hein is still looking for the molecular factors that remove methyl groups from genes after birth, activating the adult genes. 

It is noteworthy that the epigenetic programme also changes in the cardiac muscle cells of diseased hearts, in the case of chronic heart failure, for example. The expression of proteins that have not reached full power and are thus involved in the reduced performance of the organ is a marker of chronic heart failure. Hein calls this a return to the foetal gene programme, but he is still looking for explanations as to why it occurs. Maybe being able to form other protein isoforms under stress benefits cardiac muscle cells. Rather than removing methyl groups, cells up- or downregulate a protein that recognizes the methyl groups, thereby affecting the expression of the relevant genes. “This allows the cells to temporarily fine-tune gene expression, while the long-term switch, i.e. methylation, remains,” said Hein. In mouse models used to examine the protein MeCP2, Hein and his team were able to substantiate the plastic situation that they had assumed existed. 

Chronic heart failure is characterized by a huge increase in cell size, similar to that which is observed in newborns. The diseased heart needs to cope somehow with increased stress and does so getting bigger. If the heart recovers, these changes can be reversed. The researchers are already thinking about ways to diagnose heart disease very early on. They have developed a method to separate cardiac muscle cells from tiny frozen tissue samples that uses probes and antibodies to map the epigenome of cardiac muscle cells. The researchers want to refine the method in order to conduct epigenetic analysis on miniscule tissue biopsies. “I am sure that this will keep us busy for quite some time to come,” says Hein. 

Further information:
Prof. Dr. Lutz Hein
Institute of Pharmacology and Toxicology
University of Freiburg
Albertstr. 25
79104 Freiburg
Tel.: +49 (0)761 / 203 - 5313
Fax: +49 (0)761/ 203 - 5318
E-mail: lutz.hein(at)pharmakol.uni-freiburg.de

Website address: https://www.gesundheitsindustrie-bw.de/en/article/news/heart-muscle-cell-1-0-and-2-0-two-epigenetic-programmes-in-one-cell