Feature: Uncovering the genetics of heart development

By Susan Williamson

This feature appeared in the September/October 2010 issue of Australian Life Scientist. To subscribe to the magazine, go here.

When Richard Harvey began his PhD on histone genes from chickens in the late 1970s, recombinant DNA technology was considered controversial, and the ethical debate around stem cell research was far in the future.

At the time of his PhD, there was a voluntary moratorium on using recombinant DNA technology because of safety issues. Endorsed by scientists internationally, the moratorium nonetheless led to the rapid development of the technology as well as modification of the host bacteria used to harbour recombinant plasmids, rendering them safe and unlikely to colonise humans.

Having obtained his PhD in 1982 from the University of Adelaide under Professor Julian Wells, a pioneer of molecular biology in Australia, Harvey went looking for a more biological context to apply his skills. A brief stint in the commercial world in France, where he was involved in cloning an anti-thrombin factor from medicinal leeches, made him realise he did not fit the mould of industry.

A subsequent post-doc in Professor Doug Melton’s lab at Harvard University, Massachusetts, was where he found his biological home – in embryology and developmental biology. Melton is a leader in developmental and stem cell biology and Harvey spent three years in his lab looking at issues of pattern formation and lineage specification in the frog embryo.

This is where he began working with the homeobox genes, or Hox genes, which are a group of related genes that encode a class of transcriptional regulators known to specify the position of cells within the embryo.

“Understanding how the germ layers and body axes are formed is essential if you’re working with embryos,” says Harvey. “My time at Harvard set things in train for the rest of my career – dealing with key patterning issues or the geometry of the embryo, how the location of organs is specified and or how cell lineages arise and specialise.”

Australia called and Harvey returned to a position at the Walter and Eliza Hall Institute of Medical Research in Melbourne in 1988 and stayed there for 10 years. The publication of Dr Mario Capecchi’s pioneering work on producing knockout mice consolidated Harvey’s decision to move to working with a mammalian system – the mouse.

Harvey was one of the first to develop this technique in Australia and, with his own lab established and with independent funding, he began looking for interesting developmental genes in the mouse to study.

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“This research has taken off world wide,” says Harvey. “So much so that now every gene in the mouse genome is being knocked out in embryonic stem cells and individuals can import these from repositories around the world to make their own mutant strains. In developmental biology the mouse serves as a good model for humans because the fundamental rules of patterning in the embryo are conserved.”

The discovery of the homeobox gene, NKX2-5, which is expressed very early in heart progenitor cells, was a defining moment in his research, and since then his main experimental focus has been on heart development.

The NKX2-5 gene was one of the first transcription factors to be identified in the developing heart. “This finding opened doors for us to pursue the molecular dissection of heart development,” says Harvey. “And it turns out that the NKX2-5 gene is the most commonly mutated single gene in CHD in children.”

Congenital heart disease genes

Harvey is now the head of the Developmental Biology Division and Co-Deputy Director at the Victor Chang Cardiac Research Institute in Sydney, where he heads up a team of about 15 people conducting medical molecular biology research.

The team’s current research involves a candidate gene approach to identify genes that may be causative in congenital heart disease (CHD) as well as revealing the basic biology of adult stem cells with the aim of taking these into regenerative medicine.

Harvey’s work has involved striking a balance between contributing to understanding the basic mechanisms of developmental biology and promoting its relevance to disease mechanisms and therapies. One of his long-term goals is to find simple treatments that render the regulatory pathways of the heart more robust to genetic or environmental insult.

The heart is the first organ to function in the mammalian embryo and is highly susceptible to genetic perturbations that lead to congenital malformations. Babies born with severe heart defects are commonly known as ‘blue babies’ because of inadequate oxygenation of the blood.

These babies have a strong enough heart to get them through foetal life but, once they are born and the pulmonary system kicks in, this increases stress on the heart, and within the first few days of life these babies require surgery and critical management for their survival.

About 10 per cent of CHD is hereditary, being caused by dominant single gene mutations. The majority, however, are due to the combined effects of many mutant genes combined with influences from the foetal environment. Mutations in the NKX2-5 gene occur in about 1 to 3 per cent of patients with CHD, which is as common as any single gene defects in that category.

Harvey’s team has used a candidate gene approach to identify genes that may be causative in human disease. A key focus of this approach has been the identification of transcription factors that regulate cardiogenesis.

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In addition to NKX2-5, work in mouse embryos and human patients has identified two other genes, one from the T-box family of transcription factors, Tbx20, and a cardiac myosin gene, both of which are involved in CHD. They have gone on to show that these genes play fundamental roles in formation and patterning of the heart in the embryo.

“This candidate gene approach is time consuming but is satisfying because it involves interfacing with clinicians to obtain DNA from patients, as well as recruitment of patients and controls, and screening for gene mutations,” says Harvey. “We collaborate with Associate Professor David Winlaw at The Children’s Hospital at Westmead, as well as collaborators overseas, and look at upwards of 300 patients in each study.”

Genes and strokes

About 10 years ago it was thought that the adult heart could not repair itself because it had no regenerative reserve. However, recent research suggests that replacement of cardiomyocytes does occur over an individual’s life, and the heart does attempt to repair itself in injury.

Harvey entered the stem cell field about eight years ago because of his work on heart progenitor cells and his interest in defining their characteristics, as well as to maintain a contemporary edge with his research. He is currently receiving funding from the Australian Stem Cell Centre for this work.

His team has been working with a population of adult cardiac stem cells that they believe maintain the vascular and stromal component of the heart, including smooth muscle cells and myofibroblasts.

So far this work has involved basic research on defining, quantifying and assessing how these cells behave in relation to disease and aging, as well as looking at how they participate in repairing the heart after injury.

The main aim of the work is to provide for regenerative medicine – repair of the coronary circulation and replacement of lost muscle is fundamental to repairing the heart, especially in ischemic injury.

“The field has very much relied on the dream that if stem cells were identified and grown in a dish, then they could be put into the heart and would know what to do to regenerate and create new tissue,” Harvey says.

“This is far from being reality. There are benefits from this approach, for example in cell therapy to repair the heart after a heart attack, but the mechanism is unlikely to harness the full potential of stem-cells, or even be stem cell related at all.

“So the field is now entering a new era in which more basic science will be needed to work out how to facilitate stem cell growth and differentiation in the adult heart.”

This feature appeared in the September/October 2010 issue of Australian Life Scientist. To subscribe to the magazine, go here.