Clonal Proliferation Within SMCs in Atherosclerotic Lesions

Atherosclerosis is a disease of blood vessels that causes thickening of the arteries. Build-up of fatty deposits (fat and cholesterols) known as plaques that form on the arterial wall cause narrowing or total occlusion of the lumen, thereby blocking efficient blood flow to vital organs such as the heart and brain. Complications such as acute myocardial infarction (heart attack), stroke, and peripheral vascular disease (plaque buildup in the leg arteries) are commonly seen as individuals age.

In their recent study, Kenji Kawai and his colleagues at CVPath Institute wanted to better understand the complex inner workings of atherosclerotic cells. Although healthy practices like weight loss, exercise, quitting smoking, etc., can successfully diminish the progression of heart disease, Kawai et al. emphasize that studying plaque cellular constituents at the molecular level allows for the design of more precise treatments, rather than just reducing cholesterol.

The study focused on smooth muscle cells (SMCs), which are fundamental in building components of an atherosclerotic lesion in a diseased artery. Because multiple smooth muscle cells comprise atherosclerotic plaque, the investigators wanted to understand the origin of these cells. Do they originate from just one progenitor that proliferates (similar to cancers) or from multiple smooth muscle cells that proliferate? To find this answer, Dr. Kawai and his colleagues used a strategy that relied on the identification of clones by locating a gene on the X chromosome. Females carry 2 X chromosomes, and in every cell, one of those chromosomes is randomly inactivated. If an individual carrier has one copy of a mutated gene on one X chromosome and a normal copy of the gene on the other chromosome, it is possible to study the distribution of the mutated gene and the normal gene within cells. Cells coming from a common progenitor that carries the mutated gene would all carry copies of the mutant gene, whereas if many progenitor cells existed, one would expect cells within a specific tissue to have a 50/50 split in the mutant to normal gene ratio. Such techniques allowed the researchers to track the clonality of cells.

Work done over 50 years ago first suggested smooth muscle cells in atherosclerotic plaques may be clonal in origin based on zymography of homogenized plaque material. Dr. Kawai used a detection method called RNA in situ hybridization (BaseScope), which identified a naturally occurring mutation of the biglycan gene, a protein encoded by a gene on the X chromosome and secreted by SMCs. The detection of either a 24-nucleotide deletion in the mRNA of the biglycan gene or the normal variant was used as an indicator to measure the spatial distribution of the clonal cells. They determined the arrangement and distribution of SMC clonal proliferation in atherosclerotic lesions which had never been done before until now.

Using these techniques, Kawai et al. found that a majority of cells in the plaque did not have the same progenitor and thus were not of clonal origin. Ultimately, the concept of an atherosclerotic stem cell posited by researchers over 50 years ago does not adequately explain the mechanisms by which SMCs populate atherosclerotic plaques. Indeed, atherosclerotic plaques predominantly display a substantial proportion of non-clonal SMC regions, suggesting SMCs proliferate in a heterogeneous manner. This may explain why therapies targeting mechanisms such as lipids and inflammation, which drive SMC proliferation, have been shown to be successful targets for anti-atherosclerotic therapies. “Nobody has ever shown the precise location of the SMC clones in atherosclerotic plaques like we have shown,” Dr. Kawai said. “This study will be a fundamental driver in developing a new drug to treat atherosclerosis.”

You can access the paper here if you would like to read more about Kenji et al.’s recent findings.

Written by Aloke Finn and Gina Miller