Inhibiting CXCL12 blocks fibrocyte migration and differentiation and attenuates bronchiolitis obliterans in a murine heterotopic tracheal transplant model  David A. Harris, BS, Yunge Zhao, MD, PhD, Damien J. LaPar, MD, MSc, Abbas Emaminia, MD, John F. Steidle, BA, Mark Stoler, MD, Joel Linden, MD, Irving L. Kron, MD, Christine L. Lau, MD  The Journal of Thoracic and Cardiovascular Surgery  Volume 145, Issue 3, Pages 854-861 (March 2013) DOI: 10.1016/j.jtcvs.2012.03.079 Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions

Figure 1 Box and whisker plots depicting total fibrocyte populations in both anti-CXCL12 F(ab′)2–treated and control goat IgG F(ab′)2–treated mice after heterotopic transplant of allograft tracheas. Box and whiskers represent 25th to 75th and 10th to 90th percentiles, respectively; transverse lines represent the median. Each box and whisker plot at a given point represents 5 mice. A, C, E, Undifferentiated, CD45+collagen 1 (Col1)+CXCR4+ fibrocyte populations over time in buffy coat (BC), bone marrow (BM), and tracheal allograft isolations (Trach), respectively. B, D, F, Differentiated, CD45+Col1+smooth muscle actin (SMA)+ fibrocyte populations in BC, bone marrow, and trachea allograft isolations, respectively. *P < .05, Mann-Whitney U test. The Journal of Thoracic and Cardiovascular Surgery 2013 145, 854-861DOI: (10.1016/j.jtcvs.2012.03.079) Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions

Figure 2 Representative images of hematoxylin and eosin–stained histopathologic sections of (left), goat IgG F(ab′)2–treated or (right), anti-CXCL12 F(ab′)2–treated mice at 3, 7, 12, 21, and 28 days after heterotopic transplantation of allograft trachea. The Journal of Thoracic and Cardiovascular Surgery 2013 145, 854-861DOI: (10.1016/j.jtcvs.2012.03.079) Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions

Figure 3 Box and whisker plots depicting percentage of luminal fibrosis in tracheal allograft sections in both anti-CXCL12 F(ab′)2–treated and control goat IgG F(ab′)2–treated mice as assessed by a lung pathologist in a blinded fashion. Box and whiskers represent 25th to 75th and 10th to 90th percentiles, respectively; transverse lines represent the median. *P < .05, Mann-Whitney U test. The Journal of Thoracic and Cardiovascular Surgery 2013 145, 854-861DOI: (10.1016/j.jtcvs.2012.03.079) Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions

Figure 4 Box and whisker plots depicting percentage of tracheal luminal fibrosis as measured using Direct Red 80 collagen stain at 21 and 28 days after transplantation. Box and whiskers represent 25th to 75th and 10th to 90th percentiles, respectively; transverse lines represent the median. *P < .05, Mann-Whitney U test. The Journal of Thoracic and Cardiovascular Surgery 2013 145, 854-861DOI: (10.1016/j.jtcvs.2012.03.079) Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions

Figure 5 Combined kinetics of both undifferentiated, CD45+collagen 1 (Col1)+CXCR4+ fibrocytes in tracheal allografts, as determined by flow cytometry, and tracheal tissue collagen deposition, as determined by Direct Red 80 densitometry of tracheal allograft sections. A, Kinetics of control, goat IgG F(ab′)2–treated mice showed more gradual and less complete decrease in tracheal allograft undifferentiated fibrocyte number with a concomitant more rapid and greater increase in tracheal collagen deposition compared with (B) anti-CXCL12 F(ab′)2–treated mice. Kruskal-Wallis analysis demonstrated that both control and anti-CXCL12 fibrocyte populations (P < .001) and tracheal collagen deposition (P < .001) were significantly different. The Journal of Thoracic and Cardiovascular Surgery 2013 145, 854-861DOI: (10.1016/j.jtcvs.2012.03.079) Copyright © 2013 The American Association for Thoracic Surgery Terms and Conditions