Session Type: ACR Poster Session C
Session Time: 9:00AM-11:00AM
Background/Purpose: The strong genetic association between the HLA-B27 gene and ankylosing spondylitis (AS) has been known for over 40 years. Among AS patients, HLA-B27 positivity is associated with worse disease outcome. The long-term outcome of AS is determined by both inflammation and structural damage. The latter is characterized by new cartilage and bone formation, often leading to ankylosis. In this study, we investigated the in vitro and in vivo impact of HLA-B27 (over)expression in model systems of cartilage and bone formation.
We used different in vitro systems mimicking endochondral bone formation. We set up micromasses with ATDC5 cells and human periosteal derived cells (hPDCs) transduced with a HLA-B27 or HLA-B7 (as a control) lentiviral vector. Micromasses were kept for 21 (ATDC5) or 14 (hPDCs) days in culture. RNA extraction and colorimetric tests were performed at different time points (d1, d7, d14, d21). For the ATDC5 cells, after 14 days the chondrogenic medium was switched to mineralization medium. To specifically study osteogenesis in hPDCs, cells were kept in monolayer for 28 days and stimulated with osteogenic media. We also set up micromasses from limb bud cells derived from HLA-B27 transgenic and wild-type (WT) C57BL/6 mice embryos at day 12.5E. These cells are also known to undergo endochondral bone formation in vitro. Samples were taken for RNA extraction at d1, d7 and d14. Chondrogenesis (Col II, Aggrecan, Col X) and osteogenesis associated markers (OSC, ALP, Runx2) were studied by quantitative RT-PCR. Stainings with alcian blue, alizarin red, picosirius red and safranin O were performed to measure proteoglycans, mineralization and collagens. For the in vivostudy, we induced collagen antibody induced arthritis (CAIA) in HLA-B27 transgenic and WT C57BL/6 mice (n=10 in each group). Clinical scoring was performed twice a week. µCT scans were performed at baseline and 3 and 6 weeks after disease induction to study new bone formation. Statistical analyses were performed by 2 way ANOVA taking into account repeated measurements as appropriate.
Results: There was no difference in expression of Col II, Aggrecan or Col X in the HLA-B27 ATDC5 micromasses compared to the HLA-B7 micromasses. There was no difference between the stainings at the different time points. For the hPDC micromasses, there was no difference in expression of the chondrogenesis markers between HLA-B27 and HLA-B7 micromasses. Stainings with alcian blue could not show any difference between the two conditions. Expression of osteogenesis markers en alizarin red staining was comparable in the HLA-B27 and the HLA-B7 hPDCs in monolayer. The limb bud micromasses showed no difference in expression of chondrogenesis associated genes between HLA-B27 transgenic and WT embryos. In the CAIA experiment, the HLA-B27 transgenic mice showed more severe arthritis compared to WT mice. µCT analysis showed no increased bone formation in the post-inflammatory phase when HLA-B27 transgenic mice were compared to WT.
Conclusion: The presence of HLA-B27 seems to enhance joint inflammation in the CAIA model in C57BL/6 mice. These in vivo and the in vitrodata do not support a direct role for HLA-B27 in new cartilage or bone formation leading to ankylosis.
To cite this abstract in AMA style:Neerinckx B, Heritage K, Shaw J, Kollnberger S, Bowness P, Lories R. The Presence of HLA-B27 Increases Severity of Arthritis but Does Not Appear to Influence New Bone Formation in Different in Vitro and in Vivo Models [abstract]. Arthritis Rheumatol. 2015; 67 (suppl 10). https://acrabstracts.org/abstract/the-presence-of-hla-b27-increases-severity-of-arthritis-but-does-not-appear-to-influence-new-bone-formation-in-different-in-vitro-and-in-vivo-models/. Accessed September 19, 2019.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-presence-of-hla-b27-increases-severity-of-arthritis-but-does-not-appear-to-influence-new-bone-formation-in-different-in-vitro-and-in-vivo-models/