Fig. 7: ISTA treatment improves bone mechano-responsiveness in high-fat diet/streptozotocin-induced T2D mice via osteocytes-mediated regulation of osteoblasts and osteoclasts. | Nature Communications

Fig. 7: ISTA treatment improves bone mechano-responsiveness in high-fat diet/streptozotocin-induced T2D mice via osteocytes-mediated regulation of osteoblasts and osteoclasts.

From: Rescuing SERCA2 pump deficiency improves bone mechano-responsiveness in type 2 diabetes by shaping osteocyte calcium dynamics

Fig. 7

a The experimental protocol of ISTA treatment (intravenous infusion of 73 μg/kg body weight per min for 15 min every other day with three times per week), and subsequent application of cyclic compressive loading (1200 cycles/day) on the tibiae of high-fat diet/streptozotocin-treated (HFD + STZ) mice. b, c Micro-CT data of cortical and trabecular bone architecture in proximal tibiae. d Three-point bending testing showing tibial whole-bone mechanical properties. e, f Immunohistochemical staining of the RANKL and OPG expression of osteocytes within tibial cortical bone matrix. g Toluidine blue staining to label osteoblasts on tibial trabecular bone surface. h The experimental protocol of the conditioned medium collection from high-glucose and high-fat (HGHF)-exposed MLO-Y4 osteocytic cells treated with ISTA (1 μM) in response to FSS stimulation (2 Pa), and its incubation in HGHF-exposed primary osteoblasts and RAW264.7 cells. i Western blotting assays showing the effects of the conditioned medium collected from HGHF-exposed MLO-Y4 cells treated with ISTA in response to FSS on the expression of osteogenic differentiation-related markers (including Col1a1, Osx, and Runx2) in HGHF-exposed primary osteoblasts. j, k The ALP activity assays, ALP staining, and Alizarin red staining in primary osteoblasts. l The gene expression of osteoclastogenesis-related markers, including cathepsin K, TRAP, NFATc1, and calcitonin receptor in HGHF-exposed RAW264.7 cells. m Western blotting assays of osteoclasts-related markers, including cathepsin K, TRAP, NFATc1, and calcitonin receptor. Graphs represent mean ± SD (b–g: n = 8 mice per group; i–m: n = 6 biologically independent replicates). b–g *P < 0.05, **P < 0.01 and ***P < 0.001 by two-way ANOVA with Bonferroni’s post test. j–l *P < 0.05, **P < 0.01 and ***P < 0.001 by one-way ANOVA with Bonferroni’s post test. Specific P values are provided in the Source Data file. Scale bars: e–g 20 μm; k 50 μm.

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