Arthritis & Rheumatism, Volume 60,
October 2009 Abstract Supplement
The 2009 ACR/ARHP Annual Scientific Meeting
Philadelphia October 16-21, 2009.
Lentiviral-Based Bioluminescent Imaging of Synovial Inflammatory Subtypes in Experimental Arthritis and Human Synovial Fibroblasts
Geurts1, Jeroen, Vermeij1, Eline A., Arntz1, Onno J., Kinne2, Raimund W., van den Berg1, Wim B., van de Loo1, Fons A.
Chronic synovial inflammation is a hallmark of rheumatoid arthritis and gene expression analyses of RA synovial tissues have revealed molecularly distinct, high and low inflammatory subtypes. We evaluated the applicability of lentiviral-based (LV) promoter-luciferase reporters for assessing differential inflammation subtypes in experimental arthritis and primary OA and RA synovial fibroblasts (SF).
Murine promoters of Saa3 and Cxcl1 genes, which are strongly regulated in experimental arthritis, were cloned upstream of the luciferase (Luc) cDNA in LV vectors. Murine synovium was transduced in vivo and two consecutive flares of acute arthritis were induced by intra-articular injection with streptococcal cell wall (SCW) material. Kinetics of synovial inflammation were assessed using ex vivo luciferase assays and 99mTechnetium (Tc) uptake. Classification of SF (n=12) inflammatory subtypes was accomplished using hierarchical clustering of gene expression profiles. SF subtypes were transduced with LV-Saa3/Cxcl1-Luc and stimulated with IL-1b, TNFa or toll-like receptor (TLR) 2/4 agonists. Promoter activity was measured using luciferase assays.
Luciferase expression was strongly upregulated (50100 fold) during the first flare and Saa3, but not Cxcl1 promoter activity correlated (P<0.0005, Pearson r 0.82) with inflammation as measured by 99mTc uptake. While the Saa3 promoter could be reactivated in a second flare (500 fold), the Cxcl1 promoter remained stably activated. Interestingly, using lower SCW dosages the bioluminescent reporters, but not 99mTc measurements were able to assess synovial activation, which indicated a superior sensitivity of our novel method. Hierarchical clustering of SF gene expression profiles discriminated between two subtypes. The high inflammatory subtype was characterized by predominantly increased expression of chemokines (IL8, MCP1) and matrix-degrading enzymes (MMP1/3). Relative Saa3 promoter responses to cytokines and a TLR4 agonist were significantly (P<0.001) increased in OA/RA SF from the high inflammatory subtype (n=5) compared to the low inflammatory subtype (n=3). Relative Cxcl1 promoter responses did not discriminate between inflammatory subtypes.
Saa3 promoter activity is a more sensitive read-out for synovial inflammation than 99mTc uptake in experimental arthritis. The Saa3 promoter response in stimulated primary OA/RA SF can distinguish between molecularly distinct high or low inflammatory subtypes. These results demonstrate the sensitivity and versatility of genetic bioluminescent reporters for quantitative imaging of heterogeneous synovial inflammation.
To cite this abstract, please use the following information:
Geurts, Jeroen, Vermeij, Eline A., Arntz, Onno J., Kinne, Raimund W., van den Berg, Wim B., van de Loo, Fons A.; Lentiviral-Based Bioluminescent Imaging of Synovial Inflammatory Subtypes in Experimental Arthritis and Human Synovial Fibroblasts [abstract]. Arthritis Rheum 2009;60 Suppl 10 :2000