Platelet aggregation elicits FasL expression and hepatocyte apoptosis in sinusoidal obstruction syndrome

Figures

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  Fig. 1. Immunostaining for SE-1, a LSEC marker, and CD42b, a platelet marker, in the liver of MCT-injected SOS model mice. (a) SE-1 staining for LSEC detection at 0 and 48 h after MCT injection. (b, c) HE (b) or CD42b (c) staining at 0, 12, 24, and 48 h after MCT injection.

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  Fig. 2. Immunostaining for cleaved caspase-3 and TUNEL assay in the liver of MCT-injected SOS model mice. (a) Cleaved caspase-3 staining at 0 and 48 h after MCT injection. +rTM, pretreatment of recombinant human soluble thrombomodulin (4 mg/kg). (b) TUNEL assay at 0 and 48 h after MCT injection.

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  Fig. 3. (a) Immunostaining for Fas in the liver at 0, 24 and 48 h after MCT injection. (b) The quantitative evaluation of the Fas-positive area. Data are expressed as mean ± SE (n = 3). **, p < 0.01. *, p < 0.05.

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  Fig. 4. Immunostaining for FasL in the liver of MCT-injected SOS model mice. (a) FasL staining at 0, 24 and 48 h after MCT injection. (b) Higher magnification images at 48 h after MCT injection. Red arrows indicate positive stain. (c) FasL staining at 48 h after MCT injection in combination with pretreatment of recombinant human soluble thrombomodulin (4 mg/kg). (d) The quantitative evaluation of the FasL-positive area. Data are expressed as mean ± SE (n = 3). *, p < 0.05. (e) Immunofluorescent staining for FasL and CD42b.

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  Fig. 5. FasL appearance in activated and aggregated platelets. (a) Western blotting for mouse FasL protein in platelets isolated from the peripheral blood and the liver of MCT-injected SOS model mice (SOS model) or non-injected controls (Control). (b) Immunofluorescent study for the detection of CD62P and FasL using human platelets with or without their activation and coagulation [Agg(+) or Agg(-), respectively]. Normal IgG used as a negative control. (c) The quantitative data of CD62P and FasL fluorescence intensity. Data are expressed as mean ± SE (n = 3). **, p < 0.01. (d) Western blotting for human FasL protein in platelets with or without their activation and coagulation [Agg(+) or Agg(-), respectively]. (e) The quantitative data of the band intensity of FASL and GAPDH. Data are expressed as mean ± SE (n = 3). **, p < 0.01. (f) qPCR analyses for FasL mRNA in human platelets with or without their activation and coagulation [Agg(+) or Agg(-), respectively]. CACO-2 cells, colorectal cancer cell line. PBMC, peripheral blood mononuclear cells as a positive control.

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  Fig. 6. Immunofluorescent study for the detection of cleaved caspase-3 in HepG2 cells (a-d) or primary mouse hepatocytes (f-i) at 2–24 h after co-incubation of activated and coagulated human or mouse platelets with or without the treatment of 300 µM of Kp7-6, a Fas/FasL antagonist. (e and j) The quantitative data of cleaved caspase-3 fluorescence intensity. Data are expressed as mean ± SE (n = 3). **, p < 0.01.

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  Fig. 7. A proposed model of hepatocyte apoptosis after the interaction between FasL on activated and coagulated platelets and Fas-expressed hepatocytes in the pathogenesis of SOS.