Download Platelet Role in Leukocyte Extravasation

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Rachel Andrus
Evelyn Galvez
Biology 1615
7/19/16
Platelet Role in Leukocyte Extravasation
Introduction
When body tissue is injured, the immune response to the injury involves cells called leukocytes,
of which neutrophils and monocytes are two examples. Leukocytes "roll" or move along the
inside of the blood vessel until they find a good site for "extravasation" or exiting the blood
vessel. How the leukocytes actually extravasate to the tissues is unknown. What is known is that
immune cells gather at endothelial cell junctions, which are places where endothelial cells join
together. Scientists wanted to know how leukocytes find these points of extravasation, how they
actually leave the blood vessels, and what role platelets play in both of these processes. The
scientists hypothesized that platelets "guide leukocytes to their site of extravasation" by helping
them find endothelial cell junctions (Zuchtriegel, 2016). Because inflammation plays a role in
many different diseases, this research is pertinent to many conditions such as atherosclerosis and
auto-immune diseases. It also furthers our knowledge of how the immune system and body work.
Materials and Methods
To test their hypothesis, the scientists acquired mice that had fluorescence-labeled immune cells
that could be viewed using in vivo microscopy (viewing cultures in real time still in the animal).
Using this technique, they were able to see where the immune cells went and in what order. They
then induced inflammation in a muscle of the mouse scrotum and observed the responding
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immune response. They chose several small vessels, at least 150 micrometers from each other
and, after inducing inflammation, they measured the number of rolling leukocytes (those rolling
along the vessel) and adhered leukoctyes (those attaching to the vessel wall). Then they depleted
the platelets in the area and repeated the experiment. They performed this experiment one more
time, this time blocking integrins (proteins that allow cells to attach to other cells). After the
microscopy, blood was collected from the mouse and the leukocytes and platelets in the blood
were counted by a Coulter ACT Counter.
The scientists also used a peritonitis assay, which involved inducing inflammation similarly to
the mice used for the microscopy, and then washing the part of the mouse they affected with
saline. The saline used to wash the mouse was then measured for leukocytes by flow cytometry.
Results
Platelets were found in two forms in the post capillary vessels of the control animals. Rolling
platelets were defined as platelets “moving slower than the associated blood flow,” while
adhered platelets were those “resting in the associated blood flow for 30s and related to the
luminal surface” (Zuchtriegel, 2016). The luminal surface is the inside surface of the blood
vessel. In the normal, uninflamed animal, some rolling platelets and very few adhered platelets
were found in post capillary venules (small vessels between capillaries and veins), however once
the nearby tissue was inflamed, the platelet composition changed immediately. The number of
rolling platelets decreased, while the number of adhered platelets increased and 95% of these
were found near endothelial cell junctions. Immune cells (neutrophils and monocytes) were
found to adhere at the same location, but after the platelets. 85% of neutrophils, and most of the
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monocytes at endothelial cell junctions were found to be near adhered platelets. When platelets
were depleted by antibodies, immune cells failed to adhere to the vessel walls. Surprisingly, this
mostly happened in small vessels (diameter of 40 micrometers or less). In larger vessels, the
count of rolling leukoctyes increased, but the ratio between of rolling leukocytes and adhered
leukocytes remained the same when platelets were depleted. Platelets adhered almost not at all in
arterioles. Also, in the experiment where integrins were blocked, adherence of neutrophils was
found to be significantly reduced, and extravasation decreased significantly as well.
Discussion
This study suggests an order of cell adhesion during leukocyte extravasation: “…upon onset of
inflammation, platelets are the first cellular blood components adhering to the endothelial surface
of post capillary venules…" (Zuchtriegel, 2016). Leukocytes adhere immediately after platelets.
Therefore, the scientists’ hypothesis was supported, and they found that platelets gather around
areas of endothelial cell junctions and act as "pathfinders" to help guide leukocytes to endothelial
cell junctions. Once there, they can participate in extravasation and move to injured tissue.
Platelets adhere to the inside of blood vessel walls first, followed by neutrophils and monocytes.
Integrins are important in this process. The platelets may also be instrumental in activating
leukocytes and helping them in the process of extravasation.
The study did not list any limitations to their study or areas where further study would be useful.
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Works Cited
Zuchtriegel G, Uhl B, Puhr-Westerheide D, Pörnbacher M, Lauber K, Krombach F, et al. (2016)
Platelets Guide Leukocytes to Their Sites of Extravasation. PLoS Biol 14(5): e1002459.
doi:10.1371/journal.pbio.1002459