Microscopes Yield New Insight into Life and Death of Blood Vessels

By Jeffrey Norris

Donald McDonald, MD, PhD

Donald McDonald, MD, PhD

Developing drugs that stop blood vessel growth in cancer — causing tumors to shrink — is a hot area in pharmaceuticals. Still, researchers and clinicians don’t know as much as they would like about how these drugs work and why they work better in some tumor types than in others. Sometimes the drugs are effective by themselves, but in other tumors they must be used with chemotherapy or radiation. Working with approved and experimental drugs, Donald McDonald, MD, PhD, a UCSF professor of anatomy and member of the UCSF Comprehensive Cancer Center and the Cardiovascular Research Institute, has discovered clues to help understand these unusual features. By studying mice that spontaneously develop tumors, he is learning how newer drugs can destroy the blood vessels of tumors. Clinical trial results are the bottom line in drug testing. But drug trials don’t necessarily explain the biology underlying the clinical response. Moving from the scale of the whole organism to the single molecule requires increasingly powerful laboratory assays to detect telltale gene activity, enzymes and other molecules in prepared lab samples. Tests and experiments based on these methods can shed light on clinically important biochemical processes and events. McDonald uses these assays, but he also knows that a microscope lens trained on cells of intact tissue can be a powerful tool.

How Do Cells Change with Disease?

For three decades, McDonald’s lab group has worked on techniques for better visualizing the cellular components of blood vessels in three dimensions. The goal is to see how cells change as disease progresses, or how they change after treatment. A tumor-prone strain of mice, developed by Doug Hanahan, PhD, of the UCSF Diabetes Center, makes it possible for tumors and their blood vessels to be studied at all stages of cancer growth.
An image obtained with a scanning electron microscope

An image obtained with a scanning electron microscope shows a translucent blood vessel in a tumor, with disk-shaped red blood cells within the vessel. Cancerous cells are visible at the center of the image, above the blood vessel.

Lab members Peter Baluk, PhD, and Hiroya Hashizume, MD, PhD, have refined both light and electron microscopy techniques to permit imaging of the cells of small blood vessels and their supporting scaffolding in normal organs and in tumors in unprecedented detail. The inner wall of a blood vessel is formed from a single layer of cells known as endothelial cells. The endothelial cells are joined by tight junctions to prevent leaks. Pictures of blood vessels often represent the wall as made of cobblestone-shaped endothelial cells. But in reality, the cells are amazingly thin. McDonald’s research team observed that tumor blood vessels have a single layer of endothelial cells. But the cells are even thinner than normal — so thin that the vessels are translucent. Individual red blood cells are clearly visible within.

Tumor Vessels Are Surprisingly Fragile

“We were surprised at how thin and fragile the blood vessels are in tumors,” McDonald says. “The endothelium of tumor vessels is like plastic wrap, with holes that make it leaky.” Microscopic images showed that vessels in tumors close off before disappearing during the course of treatment. That’s why there is no hemorrhaging when the vessels die. McDonald notes that drugs that target growing blood vessels — called angiogenesis inhibitors — were designed to stop new blood vessel formation. But in tumors, they also destroy some of the existing blood vessels. The first approved anti-angiogenesis drugs target a receptor for a blood vessel growth factor called VEGF. Although it was unknown until after the drugs were developed, tumor blood vessels are especially reliant on VEGF to survive. That’s why the drugs not only prevent new blood vessel growth, but also kill preexisting tumor blood vessels. But even after the endothelial cells lining the tumor vessels vanish, other components of the vessels remain, McDonald found. Specifically, cells known as pericytes survive. Pericytes are peculiar cells that look a bit like little octopuses tightly embracing the surface of blood vessels. In tumors, they are abnormal. Their cell bodies are only loosely associated with the blood vessel surface, and they do not provide the normal support for endothelial cells.

Scaffolding Persists

The scaffolding that supports blood vessels also remains in tumors after blood vessels are destroyed, McDonald and colleagues discovered. McDonald’s lab team found that when treatment stops, the blood vessels grow back along the same scaffolding — within days.
An image obtained with a fluorescent confocal light microscope

An image obtained with a fluorescent confocal light microscope shows abnormal blood vessels within a tumor in green and yellow. The scaffolding that supports the blood vessels appears red. The right image is the same tissue after treatment with an angiogenesis inhibitor. Few blood vessels remain. However, the scaffolding is largely intact. Blood vessels quickly grow back along this scaffolding when treatment ends.

McDonald likes to use a railroad analogy. It is as if the trains are removed, but the tracks remain. If the trains are allowed to run again, they already have an established distribution system of tracks. Just as the endothelial cells that line blood vessels in tumors require VEGF, researchers believe that pericytes require a different growth factor, called PDGF. With tumor-bearing mice, McDonald is investigating whether surviving pericytes might support the railroad tracks that allow blood vessels to grow back quickly. He is exploring this by using experimental drugs that target the action of PDGF on pericytes. Through the microscope, it will be clear to the researchers whether pericytes can be destroyed, and whether regrowth of blood vessels in tumors can thereby be slowed or prevented after anti-VEGF treatment stops. Tumor blood vessels are clearly “bizarre” compared with normal blood vessels, McDonald says. One might think that all blood vessels are very much alike, like PVC pipes in a house. But capillaries and other small blood vessels differ from organ to organ — which piques the researchers’ interest. “Every organ has its own unique microcirculation, which is adapted to the specialized functions of the organ,” McDonald explains. “And the microcirculation of tumors is unlike that of any normal organ, making it a promising target for destruction by novel drugs.”