1. Background:
• Neutrophilic infiltration is prominent during acute lung injury
(ALI)1.
• Pericytes (peri-microvascular stromal cells), contribute to
neutrophil accumulation in non-pulmonary tissues2.
Pericyte activation and expression of pro-inflammatory markers in
acute lung injury
Bonnie L. Hastings, Kristen L. Mittelsteadt, Chi F. Hung, William A. Altemeier
Center for Lung Biology, University of Washington, Seattle, WA, USA
Conclusions:
• The lung contains putative pericytes that are derived from
Foxd1-expressing mesenchyme.
• Lung pericytes upregulate pro-inflammatory transcripts,
including adhesion molecules and chemokines, with injury.
• Lung pericytes increase surface expression of ICAM-1 in
response to injury.
Methods:
• Sterile lung injury is induced by intratracheal rhFasL instillation,
followed by mechanical ventilation.
• Primary putative pericytes are identified in whole lung digest as
CD45-, CD31-, and either PDGFRβ+ or derived from Foxd1+
progenitors.
• Expression of relevant genes are identified by qRT-PCR and flow
cytometry.
Citations
1. Ware, L. B. (2006). Pathophysiology of acute lung injury and the acute
respiratory distress syndrome. Seminars in Respiratory and Critical
Care Medicine, 27(4), 337–349.
2. Proebstl, D., Voisin, M.-B., Woodfin, A., Whiteford, J., D'Acquisto, F.,
Jones, G. E., et al. (2012). Pericytes support neutrophil subendothelial
cell crawling and breaching of venular walls in vivo. Journal of
Experimental Medicine, 209(6), 1219–1234.
3. Hung, C., Linn, G., Chow, Y.-H., Kobayashi, A., Mittelsteadt, K.,
Altemeier, W. A., et al. (2013). Role of lung pericytes and resident
fibroblasts in the pathogenesis of pulmonary fibrosis. American Journal
of Respiratory and Critical Care Medicine, 188(7), 820–830.
ICAM-1 expression
Meanfluorescenceintensity
U
ninjured
Injured
0
2000
4000
6000
Figure 2:
1. Alveolar damage causes release of damage-
associated molecular patterns (DAMPs).
2. Pericytes are activated and express adhesion
molecules.
3. Circulating leukocytes enter interstitium via
activated pericytes.
Figure 5: After sterile lung injury, TdTomato+(Foxd1-derived), CD45-, CD31- pericytes
upregulate ICAM-1 surface expression.
• Pericytes upregulate surface ICAM-1 expression in response to
sterile lung injury.
Results
• Pulmonary microvascular pericytes are derived from embryonic
Foxd1+ mesenchymal cells, express PDGFRβ, and are in close
proximity to CD31+ endothelial cells.
Figure 3: Foxd1 expression is restricted to embryonic development. Foxd1-Cre; R26-tdTomato
mice express TdTomato in Foxd1-derived cells. In these mice, TdTomato co-localizes with
PDGFRβ-expressing cells, and are in close proximity to CD31-expressing cells.
Figure 4: Primary lung pericytes were isolated from intact and injured lungs by enzymatic digestion
and serial magnetic cell sorting, using depletion of CD45+ and CD31+ cells followed by
enrichment of PDGFRβ+ cells. Expression of relevant genes was determined by qRT-PCR.
Hypothesis:
Pericytes regulate leukocyte trafficking in the lung during acute
lung injury.
• Pericytes increase pro-inflammatory transcripts relevant to neutrophil
recruitment during sterile lung injury.
Funding: American Heart Association, Western Affiliates, University of Washington Molecular Medicine Training Program (NIH T32 GM95421)
Figure 1: Pericytes associate closely with microvascular endothelial cells and instruct neutrophils
to enter interstitial space in cremaster muscle2.
ICAM-1
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
RelativeExpression
p=0.0974
Cxcl1/KC
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
RelativeExpression
p=0.0504
SELE
RelativeExpression
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
p=0.0490
Cxcl2/Mip2
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
1.5
2.0
RelativeExpression
p=0.0036
IL1a
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
1.5
RelativeExpression
p=0.0004
Ccl2/MCP-1
RelativeExpression
PBS
rhFasL+M
V
-0.5
0.0
0.5
1.0
1.5
2.0
p<0.0001
• Lung pericytes are enriched for immune-related transcripts as
compared to other lung stromal cells at baseline3.