Fig. 8: Bone erosion in arthritis leads to locally increased [Ca²⁺] and elevated IL-1β secretion.

a Measurement of [Ca²⁺] in synovial fluid of either RA or control patients with non-erosive arthritis or osteoarthritis (ctrl). b Calcium red staining of synovial membrane biopsies for Ca²⁺ with Glyoxal-bis(2-hydroxyanil (GBHA) was done before or after incubation with 20% EDTA and immunohistochemistry for CaSR (right image, brown color). Scale bar = 150 µm. Shown is one representative experiment out of three. c CaSR protein expression was analyzed in serial sections of synovial membranes using anti-CaSR antibody (red color). Scale bar = 200 µm. Sections were counterstained with hematoxylin. Shown is one representative experiment out of six (RA) and two (osteoarthritis, OA). d Arthritis score in DBA/1J mice with collagen-antibody-induced arthritis (CAIA) treated i.p. with either 33 µM R568 or sodium acetate (NaAc) as control. e IL-1β secretion after 16 h incubation with LPS and 2.5 mM added [Ca²⁺] of blood monocytes (blood), CD11b⁺ mononuclear bone marrow cells (bone marrow) and peritoneal macrophages (peritoneum) of mice sacrificed 40 days after induction of collagen-induced arthritis (CIA) and of control animals. f Calcium red staining with GBHA of cryostat sections of paws from collagen-induced arthritis (CIA) mice, black frame shows region of erosion zone. Shown is one representative experiment out of six with three different mice. Scale bar = 200 µm. g [Ca²⁺] measured in bone marrow flushes purged with 0.9% NaCl from the medullary cavity of mice sacrificed 40 days after induction of CIA or of control mice (ctrl). h Correlation between IL-1β release from bone marrow-derived CD11b⁺ mononuclear cells after stimulation with LPS and 2.5 mM added [Ca²⁺] for 16 h and [Ca²⁺] measured in the medullary cavity from CIA mice. i Correlation between IL-1β release from bone marrow-derived CD11b⁺ mononuclear cells after stimulation with LPS and 2.5 mM added [Ca²⁺] for 16 h and clinical score of disease severity from CIA mice. j Overview of the hypothesized mechanism of Ca²⁺/CPP-induced inflammasome activation in monocytes in RA. Data are shown as mean ± s.e.m. a, d, e, g Two-tailed Mann–Whitney U test (a, d, e, g) or Pearson correlation coefficient (h, i) was used. p-values are indicated as *p < 0.05, **p < 0.01, ***p < 0.001.