Intravital 2-photon imaging of leukocyte trafficking in beating heart.
Two-photon intravital microscopy has substantially broadened our understanding of tissue- and organ-specific differences in the regulation of inflammatory responses. However, little is known about the dynamic regulation of leukocyte recruitment into inflamed heart tissue, largely due to technical difficulties inherent in imaging moving tissue. Here, we report a method for imaging beating murine hearts using intravital 2-photon microscopy. Using this method, we visualized neutrophil trafficking at baseline and during inflammation. Ischemia reperfusion injury induced by transplantation or transient coronary artery ligation led to recruitment of neutrophils to the heart, their extravasation from coronary veins, and infiltration of the myocardium where they formed large clusters. Grafting hearts containing mutant ICAM-1, a ligand important for neutrophil recruitment, reduced the crawling velocities of neutrophils within vessels, and markedly inhibited their extravasation. Similar impairment was seen with the inhibition of Mac-1, a receptor for ICAM-1. Blockade of LFA-1, another ICAM-1 receptor, prevented neutrophil adherence to endothelium and extravasation in heart grafts. As inflammatory responses in the heart are of great relevance to public health, this imaging approach holds promise for studying cardiac-specific mechanisms of leukocyte recruitment and identifying novel therapeutic targets for treating heart disease.
Department of Surgery, Washington University School of Medicine, St. Louis, MO, USA.
Wenjun
W
Li
Ruben G
RG
Nava
Alejandro C
AC
Bribriesco
Bernd H
BH
Zinselmeyer
Jessica H
JH
Spahn
Andrew E
AE
Gelman
Alexander S
AS
Krupnick
Mark J
MJ
Miller
Daniel
D
Kreisel
D000818:Animals; D057791:Cardiac Imaging Techniques / Q000379:methods; D003331:Coronary Vessels / Q000166:cytology; D049452:Green Fluorescent Proteins / Q000096:biosynthesis; D016027:Heart Transplantation / Q000276:immunology / Q000502:physiology; D036904:Leukocyte Rolling; D051379:Mice; D008810:Mice, Inbred C57BL; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods; D009200:Myocardial Contraction; D009206:Myocardium / Q000166:cytology* / Q000276:immunology; D020556:Neutrophil Infiltration; D045663:Quantum Dots; D011994:Recombinant Proteins / Q000096:biosynthesis; D059008:Time-Lapse Imaging
D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins