Abstract
Background: Sialic acid-binding Ig-like lectin 9 (Siglec-9) is constitutively expressed on neutrophils and monocytes. The expression and potential role of siglec-9 in ANCA-associated vasculitis (AAV) is yet to be determined. We aimed to examine the expression of siglec-9 in patients with AAV and explore the impact of siglec-9 engagement on ANCA-mediated neutrophil responses in vitro.
Methods: Leukocytes and serum were isolated from peripheral venous blood of AAV patients and siglec-9 expression was measured by flow cytometry and ELISA, respectively. Immunohistochemistry was performed on kidney biopsies of AAV patients with AAGN and stained for siglec-9, recombinant siglec-9 Fc chimera and leukocyte markers. Functional studies were done using healthy donor neutrophils in the presence of ANCA and siglec-9 mAb to investigate its role in apoptosis and ROS production.
Results: We found increased serum siglec-9 expression in active AAV compared to remission AAV and a positive correlation with disease activity. Neutrophils and intermediate (CD14 + /CD16 + ) monocytes from PR3-AAV patients displayed higher siglec-9 expression compared to MPO-AAV patients. Siglec-9 expression in AAGN was restricted to areas of active inflammation. Using recombinant siglec-9 Fc chimera protein in immunohistochemistry experiments, we identified glomerular endothelial cells to express siglec-9 ligands. ANCA stimulation of primed neutrophils was associated with siglec-9 shedding. Siglec-9 engagement using anti-siglec-9 mAb was associated with increased neutrophil apoptosis and reduced ROS production compared to isotype control and unstimulated neutrophils.
Conclusion: Siglec-9 expression correlates with disease activity in AAV. Surface of siglec-9 on key cell players in AAV pathogenesis, is biased towards PR3-AAV. The biologic relevance of this finding is unclear. Our in vitro experiments support a potential role for siglec-9 in attenuating ANCA-mediated neutrophil responses. Further evaluation is required to determine the relevance of these findings on neutrophil-endothelial interactions.
Methods: Leukocytes and serum were isolated from peripheral venous blood of AAV patients and siglec-9 expression was measured by flow cytometry and ELISA, respectively. Immunohistochemistry was performed on kidney biopsies of AAV patients with AAGN and stained for siglec-9, recombinant siglec-9 Fc chimera and leukocyte markers. Functional studies were done using healthy donor neutrophils in the presence of ANCA and siglec-9 mAb to investigate its role in apoptosis and ROS production.
Results: We found increased serum siglec-9 expression in active AAV compared to remission AAV and a positive correlation with disease activity. Neutrophils and intermediate (CD14 + /CD16 + ) monocytes from PR3-AAV patients displayed higher siglec-9 expression compared to MPO-AAV patients. Siglec-9 expression in AAGN was restricted to areas of active inflammation. Using recombinant siglec-9 Fc chimera protein in immunohistochemistry experiments, we identified glomerular endothelial cells to express siglec-9 ligands. ANCA stimulation of primed neutrophils was associated with siglec-9 shedding. Siglec-9 engagement using anti-siglec-9 mAb was associated with increased neutrophil apoptosis and reduced ROS production compared to isotype control and unstimulated neutrophils.
Conclusion: Siglec-9 expression correlates with disease activity in AAV. Surface of siglec-9 on key cell players in AAV pathogenesis, is biased towards PR3-AAV. The biologic relevance of this finding is unclear. Our in vitro experiments support a potential role for siglec-9 in attenuating ANCA-mediated neutrophil responses. Further evaluation is required to determine the relevance of these findings on neutrophil-endothelial interactions.
| Original language | English |
|---|---|
| Pages (from-to) | NP22 |
| Number of pages | 1 |
| Journal | Scottish Medical Journal |
| Volume | 62 |
| Issue number | 2 |
| Early online date | 4 Apr 2022 |
| DOIs | |
| Publication status | Published - May 2022 |
| Event | Scottish Renal Association : 2021 - Online event Duration: 11 Nov 2021 → 11 Nov 2021 http://scottishrenalassociation.org/ |