Capillaries are the most abundant vessels in the vast vascular network that supports our tissue and organs. These underappreciated mighty microvessels, which enable the exchange of nutrients and waste, also carry blood from the arteries to the veins, fueling the metabolic processes that keep us balanced and alive. Yet, these tiny tubes are so narrow that red blood cells must pass through in a single file line, making them challenging to study and prohibitively difficult to engineer.
Right: Engineered capillaries showing 3D structure and open lumens. Credit: Caitlin Howard
Disorders that affect the vascular system can disrupt almost every aspect of normal functioning, often fatally. Consider malaria. In this deadly disease, parasites enter the blood, causing swelling and distortion of red blood cells. The result is a catastrophic traffic jam at rush hour. Abnormal or infected red blood cells become trapped in the tiny capillary spaces, stick to the vessel walls, and – like an oversized truck in a tunnel – turn order into chaos.
The miniscule nature of capillaries – about 10-50 times thinner than a strand of human hair – has been a major obstacle for researchers studying the mechanisms of malaria and other small vessel diseases, like sickle cell anemia. And tissue engineers, who are growing more skilled at building models that enable them to recreate diseases, have so far been unable to functionally recapitulate the capillaries in those models.
Now, a team led by Ying Zheng, PhD, Associate Professor of Bioengineering and a faculty member at the Institute for Stem Cell and Regenerative Medicine (ISCRM), has developed a technique to finally break the capillary barrier. The breakthrough is detailed in a new paper, co-authored by Cole DeForest, PhD, Assistant Professor, Bioengineering and Chemical Engineering, and Joseph Smith, PhD, Affiliate Professor, Global Health and Seattle Children’s Research Institute. The findings were published this week in the journal Science Advances.
Malaria was the inspiration – and the context – for attempting the previously impossible. The key to success, though, was the multidisciplinary approach that brought together expertise in biology, pathology, and engineering.
Zheng attributes the advances in capillary construction to a collaborative design strategy that merged principles of technology and vascular biology. There were two major steps. First, the researchers used traditional soft lithography to generate 100 micrometer vessels with sturdy endothelium (the razor thin walls of the capillaries that must precisely control the two-way flow of molecules and gasses between tissues and the vascular network). Previous work from the Zheng lab had established that while these larger vessels were stable, it remains technically difficult to achieve robust lumen structure and endothelium when the caliber is smaller than 50 micrometers. To get down to capillary-size they would need to get 10-fold smaller.
To accomplish this, the group used multiphoton technology developed by the DeForest Lab to carve tiny channels into the collagen from preformed vessels. “By focusing high-energy pulsed laser light at precise locations within the collagen gels, we are able to carve out capillary-sized vessels with single-micron resolution and full 3D control between two larger sized vessels,” says DeForest. “When combined with Zheng’s soft-lithography techniques to generate larger vessels with high throughput, this strategy provides a uniquely powerful approach to create synthetic microvasculature of physiological relevance and probe fundamental questions in vascular biology.”
As Zheng relates, the big breakthrough came when the engineering team discovered that they could create stable capillaries by disturbing, or provoking, the larger endothelium vessels by causing an injury. “We found that creating the disturbance triggered the endothelial cells to leave their comfort zone and migrate into the tiny channels where they formed robust capillaries. Remarkably, in the smallest regions of the connecting channels, the circumference of the capillary’s wall is formed from single endothelial cells – just as it is within your body.”
Crucially, the research team was able to accurately engineer the endothelium. The result, says Zheng, is the best opportunity yet to study and explain the two-step process that causes red blood cells to become trapped in the capillaries, leading to dangerous blockages. “In the past, biologists hoping to understand adhesion did not have an adequate capillary model, while engineers did not have a sufficient endothelium model. Now we can look at how individual proteins and cells interact with the cell walls. There are no limits to what we can do anymore.”
The implications for disease modeling and tissue engineering are profound, beginning with small vessel diseases like malaria and extending to other blood disorders. “The ability to engineer capillaries opens the door to more detailed study of malaria and all small vessel diseases,” says Smith. “For the first time, we can visualize how deformed red blood cells become trapped within the microcirculation and this approach provides a new experimental platform to test novel therapeutic treatments to prevent microvessel clogging in malaria and sickle cell disease.”
Already powerful tools like organoids, a technology used to study tissue formation and function in 3D, will become even more useful as flow dynamics are mimicked more faithfully at the capillary level. Right now, the main limitation of organoids is the lack of blood vessels. The presence of small capillary beds within organoids will notably advance research in this field, facilitating the growth of tissue-like structures in the lab. This will benefit both basic research studies and the development of new tissue regenerative strategies.
This work was primarily supported by National Institutes of Health (NIH) grant RO1 HL130488-01, and partially by R01HL141570, UG3TR002158 and UH2/UH3 DK107343, as well as a predoctoral fellowship F30HL134298 and a Postdoctoral Fellowship from the American Heart Association