It is well known that the BBB becomes dysfunctional when genetic defects disrupt pericyte-endothelial signaling from birth. However, very little is known about how normally developed pericytes in the adult brain respond during acute ischemia, and only one or two studies have investigated this in vivo.

"Pericytes have a lot of potential functions--they seem to be a sort of a jack-of-all-trades," said Shih. "We've had an idea of what these cells do, but we haven't been able to visualize and track them in vivo until recently."


The team was also intrigued by a handful of published studies showing that various inflammatory signaling cascades can induce pericyte MMP-9 expression.

"The problem is that, like pericytes, MMPs are hard to study in vivo--most of what we know about them is from studies on cultured cells or extracted brains," said Underly. "We wanted to probe this process in live animals so we could see the spatiotemporal relationship between pericytes and MMP activation in vivo--in the acute stroke time frame."

To do this, the team combined several novel tools to create a unique study protocol using transgenic mice, two-photon fluorescent microscopy, and a fluorescent gelatin-based reporter of MMPs that only one other research group had ever used to study the intact brain.

The researchers also used photothrombosis to block blood flow in a small area of the capillary bed and imaged transgenic mouse lines expressing bright fluorescent reporters specifically in the pericytes to clearly identify them.

"The successful combination of technologies is certainly one of the innovations of this project," said Shih. "It's the first study to combine these tools to look at the relationship among pericytes, MMP activity, and BBB leakage in ischemia."

Their findings revealed that ischemia resulted in extremely rapid (within tens of minutes), localized activation of MMP-9 and plasma leakage. Furthermore, plasma leakage occurred preferentially where the pericyte somata adjoined the capillary wall--not homogeneously along the length of the capillaries as previously imagined by the group.

These results provide strong evidence that pericytes--normally protectors of the BBB--contribute to early BBB degradation during ischemic stroke.

Using the new technology, the team did not have to extract and cut the brain and so did not lose the structure of the vasculature and blood flow.

"We had an intact system and could see where things were coming from and we were very surprised," said Shih. "I thought, 'I've been looking at this for years and I never knew that there was this beautiful co-localization.' It told us we were looking at something really interesting. The pericytes seem to nurture or damage the BBB depending on the conditions they're put in."

Medical Research Homepage