Key Components of the Complement Lectin Pathway Are Not Only Required for the Development of Inflammatory Arthritis but Also Regulate the Transcription of Factor D.

Abstract

The complement system plays an important role in the pathogenesis of rheumatoid arthritis (RA). Besides driving lectin pathway (LP) activation, the mannan-binding lectin (MBL)-associated serine proteases (MASPs) also play a key role in regulating the alternative pathway (AP). We evaluated the effects of N-acetylgalactosamine (GalNAc)-conjugated MASP-1 and MASP-2 duplexes in vitro and in mice with and without arthritis to examine whether knockdown of MASP-1 and MASP-2 expression affects the development of arthritis. GalNAc-siRNAs for MASP-1 and MASP-2 demonstrated robust silencing of MASP-1 or MASP-2 at pM concentrations in vitro. To evaluate the impact of silencing in arthritic mice, we used the collagen antibody-induced arthritis (CAIA) mouse model of RA. Mice were injected a 10 mg/kg dose of GalNAc-siRNAs 3x s.q. prior to the induction of CAIA. Liver gene expression was examined using qRT-PCR, and protein levels were confirmed in the circulation by sandwich immunoassays and Western blot. At day 10, CAIA mice separately treated with MASP-1 and MASP-2 duplexes had a specific reduction in expression of liver MASP-1 (70-95%, p < 0.05) and MASP-2 (90%, p < 0.05) mRNA, respectively. MASP-1-siRNA treatment resulted in a 95% reduction in levels of MASP-1 protein in circulation with no effect on MASP-2 levels and clinical disease activity (CDA). In mice injected with MASP-2 duplex, there was a significant (p < 0.05) 90% decrease in ex vivo C4b deposition on mannan, with nearly complete elimination of MASP-2 in the circulation. MASP-2 silencing initially significantly decreased CDA by 60% but subsequently changed to a 40% decrease vs. control. Unexpectedly, GalNAc-siRNA-mediated knockdown of MASP-1 and MASP-2 revealed a marked effect of these proteins on the transcription of FD under normal physiological conditions, whereas LPS-induced inflammatory conditions reversed this effect on FD levels. LPS is recognized by Toll-like receptor 4 (TLR4), we found MBL not only binds to TLR4 an interaction with a Kd of 907 nM but also upregulated FD expression in differentiated adipocytes. We show that MASP-2 knockdown impairs the development of RA and that the interrelationship between proteins of the LP and the AP may extend to the transcriptional modulation of the FD gene.

• Authors: V Michael Holers, Anna Borodovsky, Robert I Scheinman, Nhu Ho, Joseline Ramos Ramirez, József Dobó, Péter Gál, Jared Lindenberger, Annette G Hansen, Dhruv Desai, Rasmus Pihl, Steffen Thiel, Nirmal K Banda
• Journal: Frontiers in immunology (Front Immunol) Vol. 11 Pg. 201 ( 2020) ISSN: 1664-3224 [Electronic] Switzerland
• PMID: 32153567 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2020 Holers, Borodovsky, Scheinman, Ho, Ramirez, Dobó, Gál, Lindenberger, Hansen, Desai, Pihl, Thiel and Banda.

Chemical References

• Lipopolysaccharides
• RNA, Small Interfering
• MASP-1 protein, mouse
• MASP-2 protein, mouse
• Mannose-Binding Protein-Associated Serine Proteases
• Complement Factor D

Topics

• Animals
• Arthritis, Experimental (immunology, metabolism)
• Arthritis, Rheumatoid (immunology, metabolism)
• Complement Factor D (genetics, metabolism)
• Complement Pathway, Alternative (genetics)
• Complement Pathway, Mannose-Binding Lectin (genetics)
• Gene Expression
• Lipopolysaccharides (pharmacology)
• Liver (metabolism)
• Mannose-Binding Protein-Associated Serine Proteases (genetics, metabolism)
• Mice
• Mice, Inbred C57BL
• Mice, Knockout
• RNA Interference
• RNA, Small Interfering (administration & dosage, genetics)
• Signal Transduction (drug effects, genetics)
• Transcription, Genetic (genetics)
• Transfection