1. Volume 44, Issue 6, Pages 1132-1140 (June 2006)
Short-term homing assay reveals a critical role for lymphocyte function-associated antigen-1 in the hepatic recruitment of lymphocytes in graft-versus-host disease 
Tohru Sato, Aida Habtezion, Andreas Beilhack, Stephan Schulz, Eugene Butcher, Henrik Thorlacius 
Journal of Hepatology 
Volume 44, Issue 6, Pages (June 2006)
DOI: /j.jhep
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

2. Fig. 1
Lymphocyte accumulation in the liver. GvHD was induced in BDF1 mice by transfer of 50×106 splenocytes from C57BL/6 mice, and LILs were harvested on indicated days after cell transfer. (a) Absolute numbers of lymphocytes were determined up to 14 days after induction of GvHD. (b) The CD8/CD4 ratio of LILs was determined by flow cytometry. Data are shown as mean±SEM and n=3–6.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

3. Fig. 2
Short-term homing of donor/allogeneic (H-2Dd−) lymphocytes in the liver. GvHD was induced by transferring 50×106 splenocytes from C57BL/6 mice into BDF1 mice. On the indicated day after transfer, 20×106 in vivo activated splenocytes were CFSE labeled and transferred into secondary recipients with the identical phase (0, 5, 8, or 14 days) of GvHD. LILs were recovered 6h later. The efficiency of recruitment is expressed as a percent recruited of injected cells for each phenotypically defined subset: (a) short-term homing (H-2Dd−/CFSE+) CD8+ and CD4+ cells at different phases of GvHD (e.g. day 5 in vivo activated splenocytes transferred and subsequently recovered from day 5 secondary recipient's liver); (c) activated (H-2Dd−/CFSE+/CD44+) and naive (H-2Dd−/CFSE+/CD44−) CD8+ and CD4+ cells at day 8 of GvHD. (b) Percentage recruited (H-2Dd−/CFSE+) of total (H-2Dd−) donor CD8+ and CD4+ LILs in secondary recipients was also calculated. Data are shown as mean±SEM and n=4–5.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

4. Fig. 3
Expression of surface activation and adhesion molecules on donor LILs. Flow cytometric analysis of (a) activation markers and (b) adhesion molecules on naïve splenocytes (control, dashed line) and donor LILs (GvHD mice 8 days after GvHD-inducing allogeneic splenocyte transfer, solid line) compared to isotype control antibodies (tinted line).
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

5. Fig. 4
Role of adhesion molecules in GvHD hepatitis. The effects of blocking antibodies against adhesion molecules on the recruitment of (H-2Dd−/CFSE+) donor (a) CD8+ and (b) CD4+T cells in the liver of secondary recipients was assessed. Animals were treated intravenously with monoclonal antibodies directed against α4-integrins, LFA-1, PSGL-1, or a control antibody immediately prior to transfer of in vivo activated donor lymphocytes from GvHD mice into secondary recipients with the identical phase (day 8) of GvHD. Liver lymphocytes were isolated 6h later, and liver localization was calculated for different donor cell subsets (CD8+H-2Dd−CFSE+ and CD4+H-2Dd−CFSE+) in the liver—([cell subset]recruited/[cell subset]injected×100). Data are shown as mean±SEM. P<0.05 and n=4–17.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

6. Fig. 5
Anti-LFA-1 antibody treatment decreases periportal infiltrates in GvHD hepatitis. 50×106 splenocytes from C57BL/6 mice were transferred to BDF1 recipients to induce GvHD. GvHD mice were treated daily with either (a) isotype control (clone Hermes-1, 200μg) or (b) anti-LFA-1 (clone TIB213, 200μg) antibody. Hematoxylin and eosin staining of GvHD livers is shown following 2 weeks treatment with control antibody (a, isotype Ab) or anti-LFA-1 antibody (b, LFA-1 Ab). Anti-LFA-1 treated (b) GvHD mice have marked reduction in periportal infiltrate as compared with GvHD mice treated with control antibody (a, arrows). Bars=50μm. (n=4 per condition).
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

7. Fig. 6
Immunohistochemical staining of normal liver (a) or livers from mice with GvHD (b and c) following 2 weeks treatment with either (b) control or (c) anti LFA-1 antibody. Frozen liver sections from normal or GvHD mice were stained with directly fluorophore-conjugated anti-CD8 (FITC, green; arrows), and two-step staining with rat anti-mouse keratin 19 (Troma III) antibody followed by anti-rat Texas red secondary to identify biliary epithelium (Texas red, red). Reduction in periportal CD8+T cell infiltration (green, arrows) is seen in GvHD livers from mice treated with anti-LFA-1 antibody (b, LFA-1 Ab) as compared to those treated with control antibody (a, control Ab). Bar=50μm (n=4 per condition).
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions

8. Fig. 7
Hepatocellular injury in GvHD. Alanine aminotransferase (ALT) levels were determined 7 and 14 days after induction of GvHD. Mice were treated intravenously daily with an isotype-matched control antibody or a monoclonal antibody against LFA-1. GvHD was induced in BDF1 mice by transfer of 50×106 splenocytes from C57BL/6 mice, and ALT was determined spectrophotometrically. One of two experiments is shown. Data are shown as mean±SEM and n=4–5.
Journal of Hepatology  , DOI: ( /j.jhep )
Copyright © 2005 European Association for the Study of the Liver Terms and Conditions