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Updated: Nov 24, 2025
Analyses of Proteinuria, Renal Infiltration of Leukocytes, and Renal Deposition of Proteins in Lupus-prone MRL/lpr Mice
Published on: June 8, 2022
Microvesicles in active lupus nephritis show Toll-like receptor 9-dependent co-expression of galectin-3 binding
N S Rasmussen1,2, C T Nielsen2, C H Nielsen1
1Institute for Inflammation Research, Center for Rheumatology and Spine Diseases, Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark.
Abstract:
Circulating microvesicles (MVs) from patients with systemic lupus erythematosus (SLE) express the type 1 interferon (IFN)-inducible protein galectin-3 binding protein (G3BP), which may enhance their deposition in the glomerular basement membrane. The release of G3BP-expressing MVs from normal peripheral blood mononuclear cells (PBMCs) is induced by Toll-like receptor 9 (TLR-9) ligands, and these vesicles contain autoantibody-accessible double-stranded DNA (dsDNA). This study compares the release of MVs expressing G3BP and dsDNA from PBMCs derived from SLE patients with or without active lupus nephritis (LN) and from healthy donors, and taps further into the potential dependency on IFN-α for their generation and impacts of TLR-7/TLR-9 co-stimulation. PBMCs from 10 healthy donors and 12 SLE patients, six of whom had active LN at study inclusion, were stimulated in-vitro with recombinant human IFN-α and the TLR-9 agonists oligodeoxynucleotide (ODN)2216 or ODN2395 alone or in combination with the TLR-7 agonist gardiquimod. MVs in the supernatants were subsequently isolated by differential centrifugation and their expression of G3BP and dsDNA was quantified by flow cytometry. Stimulation with ODN2395 significantly increased the release of MVs co-expressing G3BP and dsDNA from PBMCs isolated from healthy donors and SLE patients. The expression of G3BP on individual MVs and the proportion of G3BP and dsDNA double-positive MVs released were increased in active LN patients. Neither co-stimulation with gardiquimod nor with the IFN-α inhibitor IN-1 had any effect on the MV release induced by ODN2395. In conclusion, the TLR-9-mediated inducibility of MVs co-expressing G3BP and dsDNA is increased in SLE patients with active LN.
Insights
Systemic lupus erythematosus (SLE) patients with active lupus nephritis release more microvesicles (MVs) expressing galectin-3 binding protein (G3BP) and double-stranded DNA (dsDNA), triggered by Toll-like receptor 9 (TLR-9) activation.
Area of Science:
- Immunology
- Nephrology
- Molecular Biology
Background:
- Circulating microvesicles (MVs) in systemic lupus erythematosus (SLE) carry galectin-3 binding protein (G3BP) and may deposit in the kidney.
- Toll-like receptor 9 (TLR-9) ligands induce G3BP-expressing MVs containing double-stranded DNA (dsDNA) from normal cells.
Purpose of the Study:
- To compare MV release of G3BP and dsDNA from SLE patients with and without active lupus nephritis (LN) versus healthy donors.
- To investigate the role of type 1 interferon-alpha (IFN-α) and TLR-7/TLR-9 co-stimulation in MV generation.
Main Methods:
- Peripheral blood mononuclear cells (PBMCs) from 10 healthy donors and 12 SLE patients (6 with active LN) were stimulated in vitro.
- Stimulation involved recombinant human IFN-α, TLR-9 agonists (ODN2216 or ODN2395), and the TLR-7 agonist gardiquimod.
- MVs were isolated via differential centrifugation and analyzed for G3BP and dsDNA expression using flow cytometry.
Main Results:
- ODN2395 stimulation significantly increased the release of MVs co-expressing G3BP and dsDNA in both healthy donors and SLE patients.
- Active LN patients showed increased G3BP expression on individual MVs and a higher proportion of G3BP/dsDNA double-positive MVs.
- Neither TLR-7 co-stimulation nor IFN-α inhibition affected ODN2395-induced MV release.
Conclusions:
- TLR-9 activation enhances the release of MVs co-expressing G3BP and dsDNA.
- This TLR-9-mediated MV release is elevated in SLE patients with active lupus nephritis.
