Paper: Increased CD68+ Macrophages Are Associated with Liver Fibrosis in a Humanized Chronic Graft-Versus-Host Disease Mouse Model

Poster Abstracts

Grand Hall CD (Manchester Grand Hyatt)

Hisaki Fujii, MD, PhD , Haematology/Oncology, The Hospital for Sick Children, Toronto, ON, Canada

Zhijuan Luo, MD, PhD , Developmental & Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada

Hye Jin Kim, MSc , Developmental & Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada

Susan Newbigging, MSc, DVM, DVSc , Pathology Core Centre for Modeling Human Disease, Toronto Centre for Phenogenomics, Toronto, ON, Canada

Armand Keating, MD , Department of Medical Oncology/Hematology, Princess Margaret Hospital, University of Toronto, Toronto, ON, Canada

R. Maarten Egeler, MD, PhD , Haematology/Oncology, The Hospital for Sick Children, Toronto, ON, Canada

Muhammad Ali, MD , Haematology/Oncology, The Hospital for Sick Children, Toronto, ON, Canada

Presentation recording not available for download or distribution as requested by the presenting author.

Introduction

Chronic Graft-versus-Host Disease (cGvHD) occurs in 40%-70% of allogeneic hematopoietic stem cell transplantation. Their quality of life is severely affected, and the two-year overall survival is about 60%. cGvHD displays autoimmune-like and fibrotic symptoms.B cell activation appears to be involved in this process, however, little is known about the role of the macrophage in cGvHD. Previously, we reported a humanized cGvHD model with lung fibrosis induced by G-CSF mobilized human PBMCs (G-hPBMCs) 8 weeks post transplantation. We further developed the model with liver fibrosis.

Materials and Methods

NSG mice were treated with 20mg/kg cyclophosphamide (CTX) at day -3 and -2 combined with 200cGy total body irradiation (TBI) at day -1. This was followed by injection of 1x10⁶ G-hPBMCs or 1x10⁵ CD34⁺ cells at day 0. Engraftment was assessed in PB and in specific target organs by either flow cytometry or immunohistochemistry (IHC). Liver samples were taken 8 weeks post transplantation, and fixed and stained with hematoxylin and eosin and Masson's trichrome. The specimens were evaluated by a pathologist who was blinded to the treatment group. Serum was collected 8 weeks post transplantation and analysed for human cytokine/chemokine array.

Results

The percentage of hCD45⁺ cells in PB showed a significant increase at 8 weeks compared to 4 weeks (p=0.01). Mice that received 1x10⁶ G-hPBMCs had hCD45⁺cells in PB at a median of 18.0% (1.6-79.4%), and comprized more than 95% of hCD3⁺ T cells. Histology showed peri-portal and bile duct inflammation and fibrosis, with aggregation of hCD68⁺ macrophages with lymphocytes (Figure 1). There was a correlation between liver pathology score and percentage of hCD68⁺cells (linear regression; r²=0.70, p<0.01). Macrophage derived cytokines/chemokines; human IL-6, IL-8, CCL2, CXCL10, sCD40L were significantly increased in the serum from mice with liver fibrosis. Flowcytometry analysis showed that the CD68⁺ cells were CD14^lowCD16⁺CCR6⁺HLADR⁺ positive.

Conclusion

We show that the combination of cyclophosphamide and 200cGy TBI with a low number of G-CSF-mobilized human PBMCs (1x10⁶) leads to portal inflammation and bile duct damage which mimic liver pathology in patients with human cGvHD. Human CD68⁺macrophages appear to cause liver fibrosis in this model. This may serve as an important pre-clinical model of the severe form of cGvHD.

FIGURES

Disclosures:

Nothing To Disclose

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