Artikel
Simulating the pathogenesis of arthritis in vitro by developing a human-based multicomponent 3D joint model
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Autoren
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Alexandra Damerau
- Charité – Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und Klinische Immunologie, Berlin
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Annemarie Lang
- Berlin-Brandenburg School of Regenerative Therapies, Charité University Hospital, Berlin
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Lisa Ehlers
- Charité – Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und Klinische Immunologie, Berlin
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Moritz Pfeiffenberger
- Charité – Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und Klinische Immunologie, Berlin
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Timo Gaber
- Charité – Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und Klinische Immunologie, Berlin
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Frank Buttgereit
- Charité – Universitätsmedizin Berlin, Medizinische Klinik mit Schwerpunkt Rheumatologie und Klinische Immunologie, Berlin
| Veröffentlicht: | 5. Februar 2019 |
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Gliederung
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Background: Our ultimate goal is to develop a valid human in vitro 3D joint model to simulate the pathogenesis of arthritis. The in vitro 3D joint model consists of different components including an (1) osteogenic and (2) chondrogenic part, (3) the joint space with synovial fluid and (4) the synovial membrane and contains all involved cell types. As an alternative experimental setup for animal models, our in vitro 3D joint model will enable us to study the influence and efficacy of drug treatment.
Methods: For the osteogenic component, we populated β-tricalcium phosphate (TCP) – mimicking the mineral bony part – with human bone marrow-derived mesenchymal stromal cells (hMSC). Adhesion, survival, structural integrity and osteogenic differentiation of the cells were evaluated by SEM, viability assays, qPCR and µCT, respectively. To mimic the chondrogenic part, a scaffold-free 3D cartilage construct was generated using hMSC. Constructs were analyzed by histology and qRT-PCR. Simulating the synovial fluid, hyaluronic acid was applied to the osteochondral model. To model the synovial membrane, a confluent monolayer of hMSC was formed on a polycarbonate membrane and visualized by hemacolor staining.
Results: We developed an in vitro 3D bone model by successfully seeding hMSC on a β-TCP scaffold. Cells consistently adhere onto the scaffold as observed by SEM. mRNA expression of bone-related genes as well as µCT analysis confirmed the osteogenic phenotype of hMSC. Mimicking the articular cartilage component, we verified its chondrogenic phenotype by HE and Alcian Blue staining as well as by the reduced mRNA expression of COL1A1 and an abundant expression of COL2A1. Co-cultivation of the osteogenic and chondrogenic part for up to 3 weeks demonstrated successful colonization, connectivity and initial calcification implying a functional transitional bridging area. Modelling the synovial membrane, we successfully and reproducibly created a confluent monolayer of hMSC, which is easily transferable to the model.
Conclusion: To finalize the development of healthy joint model, we will combine the established parts to provide a suitable 3D multi-component joint model, which enables us to study the efficacy of drug treatment <in vitro.
