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Additionally, the researchers found that connective tissue cells play an important role in regenerating the heart by temporarily entering an activated state.
Zebrafish can repair heart tissue after injury, according to research by the MDC team led by Jan Philipp Junker and Daniela Panáková.
When a person has a heart attack and does not receive prompt treatment, heart muscle cells (cardiomyocytes) are damaged by a lack of oxygen and begin to die. Scar tissue grows, and because we can’t make new cardiomyocytes, the heart can’t pump as efficiently as it should. However, things are radically different for lower vertebrates like zebrafish, which can regenerate organs including its heart.
“We wanted to find out how this little fish does this, and if we could learn from it,” says Professor Jan Philipp Junker, head of the quantitative developmental biology laboratory at the Berlin Institute for Biology of Medical Systems (BIMSB). , which is part of the Max Delbrück Center for Molecular Medicine of the Helmholtz Association (MDC) in Berlin.
The researchers simulated myocardial infarction lesions in the heart of their zebrafish with the help of Dr. Daniela Panáková, who leads the Electrochemical Signaling Laboratory in Development and Disease at MDC. They monitored cardiomyocyte regeneration using single cell analyzes and cell lineage trees. Their findings were recently published in Natural genetics.
Right: Adult zebrafish under a brightfield microscope. Left: zebrafish heart 7 days after cryoinjury. Transiently activated fibroblasts localize to the area of injury. Credit: Panáková Laboratory, MDC
Human hearts stop before regeneration
The zebrafish’s one-millimetre heart was exposed to a cold needle for a few seconds by the researchers while they observed it under a microscope. Any tissue touched by the needle is dead. Similar to those who have had a heart attack, this leads to an inflammatory response, which is followed by scarring produced by fibroblasts.
“Amazingly, the immediate response to injury is very similar. But while the process in humans stops at this point, it continues in fish. They form new cardiomyocytes, capable of contracting,” explains Junker.
“We wanted to identify signals from other cells and help drive regeneration,” he continues. Junker’s team used single-cell genomics to search the injured heart for cells that don’t exist in a healthy zebrafish heart.
Three new types of fibroblasts that activate momentarily have been discovered by researchers. Although they share an appearance with other fibroblasts, these activated cells have the ability to read a variety of additional genes involved in protein formation, such as connective tissue factors like collagen 12.
Fibroblasts give the signal for regeneration
In humans, fibrosis, also known as scarring, is considered an obstacle to the regeneration of the heart. However, once activated, the fibroblasts seem to play a crucial role in the process. When Panáková used a genetic trick to disable collagen 12-expressing fibroblasts in zebrafish, it became clear how crucial they are. Result: no regeneration. Junker thinks it makes sense that fibroblasts are responsible for giving the repair signals: “They just form at the site of injury, after all,” he says.
To identify the source of these activated fibroblasts, Junker’s team produced cell lineage trees using a technique called LINNAEUS, which his lab developed in 2018. LINNAEUS works with genetic scars that collectively act as a barcode for the origin of each cell.
“We create this barcode using CRISPR-Cas9 genetic scissors. If, after an injury, two cells have the same barcode sequence, it means that they are related,” says Junker.
The researchers identified two sources of temporarily activated fibroblasts: the outer layer of the heart (epicardium) and the inner layer (endocardium). Collagen-12 producing cells were found exclusively in the epicardium.
Different disciplines worked closely together on the study
Several MDC researchers collaborated throughout the study – from experiments on the fish to genetic analyzes to bioinformatics interpretation of the results.
“For me, the most exciting thing was to see how well our disciplines complemented each other and how we could verify the results of bioinformatics on a living animal,” says Sara Lelek, who is one of the lead authors of the study and was responsible for the animal. trials. “It was a big project that allowed us all to contribute our expertise. I think that’s why the study is so comprehensive and useful for many researchers.
Dr Bastiaan Spanjaard, also lead author, agrees: “Because we had such different areas of expertise, we often had to explain our experiences and analyzes to each other. Cardiac regeneration is a complex process that is influenced by many different things. The experiments produced huge amounts of data. Filtering out the correct biological signals among them was extremely difficult.
It remains unclear whether damaged hearts in mammals like humans and mice lack the necessary signals or the ability to read the signals. If the signals are lacking, drugs could possibly be developed to simulate them. But, says Junker, finding a way to mimic the interpretation of the signal would be much more difficult.
Fibroblasts also help form new blood vessels
The researchers now want to take a closer look at the genes that the temporarily activated fibroblasts particularly often read. They know that many of the genes in question are important for the release of proteins into the surrounding area. And these could include factors that also influence cardiomyocytes. And early evidence suggests that activated fibroblasts don’t just promote heart regeneration; they also contribute to the formation of new blood vessels which supply the heart with oxygen.
Reference: “Origin and function of activated fibroblast states during zebrafish heart regeneration” by Bo Hu, Sara Lelek, Bastiaan Spanjaard, Hadil El-Sammak, Mariana Guedes Simões, Janita Mintcheva, Hananeh Aliee, Ronny Schäfer, Alexander M Meyer, Fabian Theis, Didier YR Stainier, Daniela Panáková and Jan Philipp Junker, July 21, 2022, Natural genetics.
DOI: 10.1038/s41588-022-01129-5