Fig. 1: Negative regulation of bone formation by TAM receptor MERTK.

a mRNA expression analysis of TAM receptor family Mertk, Tyro3, Axl, Gas6, and Pros1 in primary murine osteoblast cultures after 0, 7, 14, and 21 days (n = 3 biological replicates) (Mertk expression d0 vs. d7: p = 0.0002; d0 vs. d14: p = 0.0042; d0 vs. d21: p = 0.0172), (Tyro3 expression d0 vs. d7: p = 0.009; d0 vs. d14: p = 0.9604; d0 vs. d21: p = 0.2928), (Axl expression d0 vs. d7: p = 0.0161; d0 vs. d14: p = 0.0152; d0 vs. d21: p = 0.0331), (Gas6 expression d0 vs. d7: p = 0.0011; d0 vs. d14: p = 0.0002; d0 vs. d21: p = 0.001), (Pros1 expression d0 vs. d7: p = 0.0157; d0 vs. d14: p = 0.0181 d0 vs. d21: p = 0.054). b, c Microcomputed tomography (μCT) of the metaphyseal proximal region of tibias from 8-week-old Mertk^flox/flox and Col1a1-cre⁺;Mertk^flox/flox female mice (top, longitudinal view of cortical and cancellous bone; bottom, longitudinal view of cancellous bone) (b). Quantification of bone volume (BV/TV) of cancellous bone determined by μCT analysis (c) (Mertk^flox/flox, n = 9; Col1a1-cre⁺;Mertk^flox/flox, n = 14). d, e Representative pictures of Calcein Demeclocycline labeling of Mertk^flox/flox and Col1a1-cre⁺;Mertk^flox/flox female mice (d). Bone formation rate of Col1a1-cre⁺;Mertk^flox/flox mice after 8 weeks (e) (Mertk^flox/flox, n = 5; Col1a1-cre⁺;Mertk^flox/flox, n = 8). f–h Representative pictures (f) and histomorphometric analysis of osteoblast (g), and osteoclast number (h) by TRAP/Hematoxylin staining in femur from Col1a1-cre⁺;Mertk^flox/flox mice. Green arrows pointing to osteoblasts visible as cuboidal or polygonal mononuclear cells on the endosteal bone surface (Mertk^flox/flox, n = 5; Col1a1-cre⁺;Mertk^flox/flox, n = 8). i Alizarin Red staining of ex vivo calvarial cell osteoblast culture from Col1a1-cre⁺;Mertk^flox/flox mice. j Analysis of MERTK protein in osteoblast cultures from Mertk^flox/flox mice treated with recombinant CRE recombinase. ß-ACTIN run on a separate gel. k Alizarin Red staining of MERTK KO calvarial cell cultures treated with PROS1 (100 nM). l, m RT-qPCR analysis of Alpl (l) and Bglap (m) mRNA expression in MERTK KO osteoblasts (n = 3 biological replicates). n Alizarin Red staining of wild-type calvarial cell cultures treated with PROS1 (100 nM) and MERTK-inhibitor R992 (200 nM). o, p RT-qPCR analysis of Alpl (o) and Bglap (p) mRNA expression (n = 3 biological replicates). Data were means ± SEMs. Statistical significance was determined by a two-tailed unpaired t-test.