Session Title: Vasculitis
Session Type: Abstract Submissions (ACR)
Background/Purpose: Giant cell arteritis (GCA) is the most common form of primary vasculitis. The pathogenesis of this disease is characterised by disregulated angiogenesis and inflammatory infiltration, however the causative mechanisms involved in regulating these processes have yet to be elucidated. This study examines the role of acute serum amyloid A (A-SAA), TLR2 activation and the downstream Notch signalling pathway in mediating the pro-inflammatory response in GCA.
Methods: A-SAA expression in Temporal artery (TA) sections from GCA patients was assessed by immunohistology. Ex vivo TA explant cultures were established from GCA patients, and the effect of pro-inflammatory stimuli A-SAA (10μg/ml) and a TLR2 agonist Pam3CYSK4 (1μg/ml) on Ang2, MMP2, MMP9, IL-6 and IL-8 was quantified by ELISA and gelatin zymography. Spontaneous release of A-SAA from ex vivo TA explant cultures was quantified by ELISA. Furthermore, myofibroblast outgrowth from TA explants was assessed using matrigel assays over a time course of 1-15 days. A-SAA induced TA explant and microvascular endothelial cell function in the presence of Notch-1 inhibition (γ-secretase inhibitor (DAPT)) or anti-TLR2 was assessed by gelatine zymography, angiogenic assays and ELISA. The effect of conditioned media from TA explants on Human embryonic kidney (HEK) -TLR2 cells was quantified by NFkb luciferase reporter assays.
Results: A-SAA expression was demonstrated in the advential and intimal regions, particularly localised to endothelial cells. High A-SAA levels (386 +/- 191 pg/ml) were spontaneously released from TA explants cultures. Ex-vivo TA explant cultures spontaneously released pro-inflammatory mediators, maintained cell viability and histological morphology, reflecting the in vivo microenvironment. A-SAA and Pam3CSK4 induced expression of instability growth factor Ang2 (p<0.05), IL-8 (p<0.05), IL-6 (p<0.05) and MMP2 and MMP 9. A-SAA and Pam3CSK4 induced myofibroblast outgrowths from TA explants embedded in matrigel over a time course of 1-15 days. In parallel, Ang2, MMP2/9 and pro-inflammatory chemokine IL-8 were induced in myofibroblast outgrowth cultures in response to A-SAA, and to a lesser extent Pam3CSK4. A-SAA induced Ang2, IL-8, IL-6 were significantly decreased in the presence of Notch inhibitor DAPT. Furthermore anti-TLR2 inhibited the effects of A-SAA on endothelial cell function. Finally conditioned media from TA explants significantly induced TLR2 activation through induction of NFκb activation, suggesting the presence of a TLR2 ligand in the inflamed microenvironment.
Conclusion: A-SAA-induced pro-inflammatory events in GAC are mediated through Notch signaling and the TLR2 activation. A better understanding of A-SAA/TLR2-mediated inflammatory pathways may lead to novel treatment strategies for RA.
S. M. Miggin,
D. J. Veale,
Abbott Immunology Pharmaceuticals,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tlr2-activation-by-acute-serum-amyloid-a-induces-pro-inflammatory-mechanisms-in-a-novel-ex-vivo-temporal-artery-explant-culturemodel-of-giant-cell-arteritis/