Bloggers: Nathalie Niyonzima and Eivind Samstad

Atherosclerosis is a progressive disease that was once believed to be a disease of cholesterol storage. Today, it is well acknowledged that atherosclerosis is an inflammatory disease, and that cells and signalling molecules from the innate immune system shape the course of the disease in various ways. The defence of the normal artery depends on innate immune responses provided by endothelial cells. When challenged by inflammation, macrophages and other cells of the immune responses are recruited to the artery wall. The macrophage is an integral component to the pathogenesis of atherosclerosis, functioning at the intersection of inflammation and cholesterol homeostasis. Atherosclerotic plaque formation is driven by the persistence of lipid-laden macrophages in the artery wall. The mechanisms by which these cells become trapped, and thereby establishing chronic inflammation, remain unknown (Peter Libby, Nature 2011).

Atherosclerosis. Illustration: iStock

There has long been a focus on finding therapeutic methods to reduce the levels of cholesterol in the arterial wall. Studies have shown that high HDL levels are associated with reduced cardiovascular risk. This is mainly due to HDL’s ability to transport excess cholesterol in arterial macrophages to the liver for excretion (i.e., reverse cholesterol transport). Despite considerable understanding of HDL and its metabolism, therapies that aim to increase HDL levels have not been successful. Because of the heterogeneity in HDL particles, just increasing HDL levels has not been beneficial, reflecting the qualitative changes in the particles. HDL has also been shown to have other functions beyond cholesterol transport – several studies have shown that HDL is anti-inflammatory, but the mechanisms behind this are not well understood (Xuewei Zhu, Ann.Rev.Nutr 2012).

In a recent study published in the prestigious journal Nature Immunology, our research partners have taken a closer look at the anti-inflammatory effects of HDL (De Nardo, et.al, Nature 2013). They identified that HDL’s anti-inflammatory effects are mediated through the induction of ATF3. ATF3 is a key transcriptional regulator of innate immune response genes, which is induced by TLR stimulation and other stimuli, and acts as negative regulator of proinflammatory cytokines (Elisabeth S.Gold, JEM 2012).

Using mouse model and human bone marrow dendritic cells (BMDMs) treated with native HDL or reconstituted HDL prior to TLR stimuli; they showed that HDL regulates inflammation in macrophages by inhibiting transcription of proinflammatory genes such as IL-1β, IL-6 and IL-12.

In order to confirm that the anti-inflammatory effects of HDL were mediated through an inflammatory repressor, they performed microarray analysis on resting BMDMs and HDL-pretreated BMDMs subsequently stimulated with TLR ligand. ATF3 was the most induced transcription factor in the presence of HDL, and it was shown to bind to the promoters of several proinflammatory genes, thereby regulating inflammation.

They demonstrated the relevance of these findings by using Apoe-deficient mice fed a high-fat diet and injected with HDL. They observed that mice treated with HDL had lesser inflammation than the untreated ones, and that the induction of ATF3 correlated with the downregulation of proinflammatory cytokines.

Besides promoting cholesterol efflux from macrophages, HDL has been shown to protect endothelial integrity by promoting endothelial repair mechanisms. Using a model of vascular repair, they showed that the protective effects of HDL on endothelial repair are for the most part driven by ATF3.

These results provide us a link between HDL and its anti-inflammatory properties that has been a puzzle over long time. The fact that ATF3 is required for the anti-inflammatory effects of HDL shows that ATF3 is a key point for endothelial damage and inflammation. The current study has laid the foundation for understanding the regulatory mechanisms that control inflammation in atherosclerosis and other chronic inflammatory diseases.

One of the best things about being a researcher is that you can work on completely new ideas and concepts without giving much thought to what is actually possible or not. Often it turns out that what you thought was impossible is possible after all if you a) develop a new technology, b) let enough researchers work on it for long enough, or c) have a bit of luck. Preferably all three.

One of the areas where we are still at the starting line is nanomedicine. Even though we are capable of making particles at nanoscale today, we know little about what role they will play in tomorrow’s medicine. In the meantime we can enjoy ourselves with hatching clever, and less clever, ideas and evaluate how far they could contribute to improved treatments in the future.

Electron microscope image of nanoparticles. (Photo: Yrr Mørch/SINTEF)

This is the status at NTNU today: In cooperation with SINTEF, we can under controlled conditions produce particles with a diameter of a few hundred nanometres (which is about as much as your fingernails grow while you read this…). These particles are made from the same plastic materials that are used in superglue. But at SINTEF they let the glue “dry” in such a way that we maintain a whole different level of control over shape, size and characteristics, than if you glue a broken cup together. In this way they produce tiny little plastic particles. They can also put things inside them or decorate them with all sorts on the outside. If you place them in water together with some proteins and give them a shake, they can even join up and produce air-filled bubbles.

You can read more about nanotechnology at SINTEF here.

Gas bubbles made from nanoparticles seen through a confocal microscope. (Photo: Yrr Mørch/SINTEF)

Nanoparticles can be used for many things. At NTNU there are now about 20 researchers looking into how these tiny superglue particles can be used to improve cancer treatment. And so some of us privileged researchers are allowed to be creative.

Could we use these particles to detect cancer? Perhaps by placing a contrast agent inside and decorating the outside with something that recognises cancer cells? Or could it perhaps be smarter to use them to treat cancer? One of the problems with breast cancer treatment is that patients have to stop chemotherapy due to side effects. But if we could make sure that more chemo ends up in the tumour and less ends up in places where it causes side effects, this problem could largely be solved.

Researchers at NTNU, for example, work on filling the superglue droplets with chemo, making them into ‘large’ bubbles (about as much as a fingernail grows in an hour…), so that the superglue-droplet chemo can safely be transported around in the blood stream. And here comes the great trick: By targeting the superglue-droplet chemo-bubbles with ultrasound waves, we could make them burst where we want them to (for example in a tumour) so that the chemotherapy is released just there and nowhere else. This sounds difficult, but NTNU has for many years been a world leader in ultrasound technology. If we gain control over the behaviour of the nanoparticles, this should absolutely be possible.

Superglue, nanoparticles and breast cancer. (Illustration: iStock)

Finally, my own personal dream for the future: Nanoparticles containing chemo which are decorated with 1) structures that recognise cancer cells, and 2) structures making them ‘invisible’ to the body. Such particles could circulate in the blood over longer periods of time, acting as secret agents. When they find a unsuspecting cancer cell in the blood stream, they can attach themselves and deliver a suitable dose of chemo. The application? Well, you could imagine that such particles could help prevent a relapse in breast cancer patients after surgery, because often there will remain a few cancerous cells in the body that could cause a relapse several years after an operation.

Is any of this possible? 10 years ago many would have said a resounding “NO”, but now we can stretch to a “perhaps” or “why not”. The research area is growing, both at NTNU and in other places, thanks to research funding from, for example, the Norwegian Cancer Society and the Research Council of Norway. There is a steep learning curve, and we learn something new about these particles literally every day – a dream existence for a researcher.