Abstract 1360: Reovirus-1 (a dsRNA Virus) and Poly (I:C) Induce High Expression of BAFF mRNA and Protein by Salivary Epithelial Cells Through TLR3 / Type I-interferon - Dependant and - Independent Pathways
Thomas Grader-Beck
Authors
M. Ittah, C. Miceli-Richard, J. Gottenberg, J. Sellam, P. Eid, C. Lepajolec, P. Lebon, X. Mariette.
Background
B-cell activating factor (BAFF) plays a key role for promoting B-lymphocyte activation and survival in primary Sjögren’s syndrome (pSS). We have previously reported that salivary gland epithelial cells (SGEC) have the property to express and secrete BAFF after stimulation by type I interferon (IFN), suggesting the pivotal role of SGEC and IFN in the pathophysiology of pSS.
Objective
To investigate if viral infection could induce BAFF expression in SGEC. If so, define if BAFF induction pathways were Toll-like receptors (TLRs) and/or type I IFN dependant.
Methods
Cultures of SGEC were established from minor salivary glands obtained from 7 patients with sicca symptoms . Expression of TLRs was investigated by RT-PCR. BAFF mRNA and protein expression was analyzed by Q-PCR and ELISA after stimulation of the different TLRs by agonists or viruses. The endosomal pathway was inhibited by chloroquine. Type I IFN induction pathway was inhibited by an anti-IFNAR1 antibody. The proinflammatory cytokines IL-6, IL-8, and RANTES were analyzed by ELISA.
Results
We detected expression of TLR-2, -3, and -7, but not TLR-9 mRNA. Chemical agonists of TLR-2 (PGN), TLR-7 (R837) and TLR-9 (CpG) did not induce BAFF expression. HSV1 (a dsDNA virus) induced BAFF mRNA but no protein. Reovirus (a dsRNA virus) and Poly (I:C) (a TLR-3 agonist) induced high expression of BAFF mRNA (ratio BAFF/actin: 81.37 ± 15.89 (SEM) and 100.45 ± 19 respectively). Likewise BAFF protein was detected in the supernatants of SGEC after reovirus infection and poly (I:C) stimulation: 76.32 pg/mL ± 7.27 and 62.09 ± 5.83 respectively. After reovirus infection, the inhibitions of endosomal pathway by chloroquine and of IFN pathway by anti-IFNAR antibody partially down-regulated BAFF mRNA by 37 % and 40 %, respectively (p=0.029 and p=0.03) and BAFF protein by 31 % and 24 % (p= 0.03 and p=0.08). After poly (I:C) stimulation, the inhibition of endosomal pathway was weaker than with reovirus-1 and IFNAR1 blockade had no effect on BAFF mRNA and protein. IL-6, IL-8 and RANTES were significantly up-regulated after reovirus-1 infection and poly (I:C) stimulation (p<0.01). Chloroquine inhibited reovirus-induced IL-6 and IL-8 secretion (p<0.01).
Conclusions
dsRNA virus infection and poly (I:C) stimulation directly induce high expression of BAFF mRNA and protein by SGEC probably through two different mechanisms. The first one implicates TLR-3 and type I IFN dependent pathways, the second one is TLR-3 and type I IFN independent and could involve NF-kB through the RIG-I/MDA5 pathway, two intracellular receptors for dsRNA. This high induction of BAFF by SGEC after viral infection sheds some new lights on the mechanisms of induction of autoimmunity by innate immunity.
Editorial Comment
BAFF (B cell activating factor) is a member of the TNF superfamily that has been implicated in the pathogenesis of Sjögren’s syndrome. BAFF promotes proliferation, differentiation and survival of B lymphocytes and BAFF levels have been found to be elevated in serum and salivary gland tissue of patients with Sjögren’s syndrome. Hypergammaglobulinemia and germinal center formation in Sjögren’s disease has been linked to elevated BAFF expression. In this study, Miceli-Richard et al. provide evidence that exposure of SGEC to reovirus leads to upregulation of BAFF protein and BAFF secretion. Furthermore, the group demonstrates that upregulation of BAFF by reovirus is partly dependent on type I interferon. This implies that salivary epithelial cells may themselves become a source of Interferon alpha and play an important role in the propagation of disease through two distinct mechanisms that stimulate B lymphocytes, production of BAFF and type I interferon. Interestingly, the group of seven patients included 3 patients with primary Sjögren’s syndrome and 4 patients with sicca symptoms due to non-Sjögren’s sialadenitis. No difference was noted between these two groups in terms of BAFF upregulation after viral infection and blockade by chloroquine and anti-IFN I antibodies. This suggests that an intrinsic abnormality of Sjögren’s salivary gland epithelial cells in response to viral stimulation appears unlikely
