An investigation of the translocator protein as a biomarker of disease activity in rheumatoid arthritis

It is now widely accepted that musculoskeletal imaging has a useful role to play in the early diagnosis of inflammatory arthritis, as well as providing a rapid and sensitive means of assessing treatment response, not only for individual patients in clinical practice, but also in clinical trials. However, the most frequently employed imaging modalities assess morphological changes in inflamed tissues. In contrast, Positron Emission Tomography (PET) generates images based on the detection of a desired molecular target of interest in vivo, and has been proposed to have potential as a highly sensitive and specific means of imaging inflammatory arthritis.
The major goal of the work documented in this thesis was to undertake a proof of principle pilot study to assess the ability of the purportedly macrophage-specific second generation translocator protein (TSPO) targeted PET tracer [11C]PBR28, to detect and quantify synovitis in a small cohort of patients with inflammatory arthritis. Measures of tracer uptake were compared with clinical examination and ultrasound imaging, as standard means of assessment for evidence of synovitis. Additionally, the relationship between TSPO PET tracer signal and immunohistochemical staining for TSPO on synovial tissue samples was assessed. The positive correlation between TSPO tracer signal in the joint compartment and synovial tissue staining for TSPO, provided strong evidence that the tracer signal was the result of target expression, and not due to confounding factors.
In order to ascertain which cell groups in inflamed synovium were likely to be significant contributors to the TSPO PET signal observed in RA, an expression profiling of the tracer target (i.e. TSPO), in cell types comprising human pannus was conducted. Using immunofluorescence studies of human RA synovial tissue, ex vivo expression of TSPO on macrophages as well as fibroblast-like synoviocytes and CD4+ T cells was noted. By measurement of mRNA expression by real-time PCR, and TSPO protein quantification, using [3H]PBR28 radioligand binding saturation assays, I have provided the first data comparing TSPO expression in major cellular components of human pannus. Collectively, data presented herein demonstrated that TSPO was not solely a macrophage specific marker in synovium. Rather, the most significant cellular contributors to TSPO PET signal were activated 'M2 phenotype' macrophages, and activated fibroblast-like synoviocytes. These findings establish the likely cellular correlates of the TSPO PET signal generated on imaging of inflamed joints in human RA.
Finally, this thesis also provided preliminary data regarding the expression of TSPO in macrophages of different phenotypes, and demonstrated a consistent downregulation of TSPO in macrophages polarised to the pro-inflammatory 'M1' end of a spectrum of macrophage phenotypes, whereas TSPO expression remained unchanged in macrophages of ‘M2 phenotype’. These novel findings suggest a role for TSPO in the negative regulation of inflammation, which we hypothesise is secondary to a function of TSPO in cholesterol transport within macrophages.

Advisor: Sabokbar, A
Taylor, P