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Phosphorylation of ERK and p38 MAPK signaling pathways was markedly impaired in Bgn/Dcn double KO mice. Whole bone lysates extracted from the cortical bones of WT, Dcn HO, Bgn HO, and Bgn HO/Dcn HT mice were analyzed by western blot using antibodies against (a) phosphorylated ERK (p-ERK), total ERK, and β-actin, and (b) phosphorylated p38 MAPK (p-p38 MAPK), total MAPK, and β-actin. c ERK phosphorylation was determined by normalizing the intensity of p-ERK protein bands to total ERK. d Total ERK was determined by normalizing the intensity of ERK protein bands to β-actin. e p38 MAPK phosphorylation was quantified by normalizing the intensity of p-p38 MAPK protein bands to total p38 MAPK. n = 3 mice/group. f Total p38 MAPK was determined by normalizing the intensity of p38 MAPK protein bands to β-actin. g Schematic summary of the roles of Bgn and Dcn in water retention and the activation of ERK and p38 MAPK signaling pathways (modified from13,24). Dcn and Bgn are characterized by one or two chondroitin sulfate/dermatan sulfate chains attached to horseshoe-shaped core proteins, respectively. The GAG chains are highly negatively charged, which play a pivotal role in retaining bound water, thus conferring ductility and plasticity to bone. Bgn and Dcn have been reported to interact with various cytokines, growth factors, and cell surface receptors to form stable complexes, thereby activating downstream pathways that regulate bone homeostasis.13,26,27 Specifically, our data suggest that Bgn and Dcn are indispensable for the activation of ERK and p38 MAPK signaling pathways, which may explain the compensatory response observed between these two PGs. Figure created using BioRender (https://biorender.com/). Data are presented as mean ± SEM. One-way ANOVA with Tukey test was used for statistical analysis. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.000 1.