Supporting Online Material


Intravascular Danger Signals Guide Neutrophils to Sites of Sterile Inflammation
Braedon McDonald, Keir Pittman, Gustavo B. Menezes, Simon A. Hirota, Ingrid Slaba, Christopher C. M. Waterhouse, Paul L. Beck, Daniel A. Muruve, Paul Kubes

Supporting Online Material

This supplement contains:
Materials and Methods
Figs. S1 to S14
References

This file is in Adobe Acrobat PDF format.

Other Supporting Online Material for this manuscript includes the following:

Movie S1. Response of circulating neutrophils to focal sterile injury in vivo. The response of eGFP-expressing neutrophils (green) to a focal necrotic lesion in the liver (red, propidium iodide) in an untreated lysM-eGFP mouse was visualized by SD-IVM (x4 objective) for four hours after injury. Elapsed time shown in top right corner. Scale bar: 200 µm.

Movie S2. Rapid chemotaxis of neutrophils to sites of sterile injury through the intravascular channels. Migratory behavior of eGFP-expressing neutrophils (green) in a lysM-eGFP mouse visualized by SD-IVM (x10 objective) between 2 and 3 hours after induction of a focal necrotic lesion in the liver (red, propidium iodide). Hepatic microvasculature is visualized by circulating Alexa-fluor-647 labeled BSA (blue). Elapsed time shown in top right corner. Scale bar: 100 µm.

Movie S3. Chemotaxing neutrophils remain intravascular as they migrate to sites of sterile injury. Representative 3-dimentional volume rendering of the liver microvasculature in a lysM-eGFP mouse 2 hours after induction of a focal necrotic lesion. eGFP-expressing neutrophils (green) remain within the vascular channels (blue; Alexa-fluor-647 labeled BSA) en route to foci of sterile tissue injury (red, propidium iodide).

Movie S4. Intravascular chemotaxis of neutrophils to sites of sterile injury requires functional Mac1. Migratory behavior of eGFP-expressing neutrophils (green) in a lysMeGFP mouse treated with a Mac1-blocking antibody visualized by SD-IVM (x10 objective) after induction of a focal necrotic lesion in the liver (red, propidium iodide). Hepatic microvasculature is visualized by circulating Alexa-fluor-647 labeled BSA (blue). Elapsed time shown in top right corner. Scale bar: 100 µm.

Movie S5. Apyrase administration reduces neutrophil infiltration to sites of sterile injury in the liver. The response of eGFP-expressing neutrophils (green) in a lysMeGFP mouse treated with apyrase was visualized by SD-IVM (x4 objective) for four hours after induction of a focal necrotic lesion in the liver (red, propidium iodide). Elapsed time shown in top right corner. Scale bar: 240 µm.