Journal: Research

Article Title: Imbalanced Skeletal Muscle Mitochondrial Proteostasis Causes Bone Loss

doi: 10.34133/research.0465

Figure Lengend Snippet: Skeletal muscle-specific ablation of LONP1 causes bone loss and mechanical impairments. (A to D) Scheme (A) for proteomic analysis using muscle samples from HU and control mice. Data were obtained from published dataset PXD041190. (B) Heatmap showing differentially regulated proteins in HU muscles ( n = 5). (C) GO analysis from up-regulated proteins in HU muscles ( n = 5). (D) Bar graphs for comparison of normalized protein expression of LONP1 in muscle tissues from HU muscles and controls ( n = 5). (E to N) WT and LONP1 mKO male mice were harvested at 8 weeks old. (E and F) μCT pictures (E) and quantification (F) of trabecular bone parameters in the distal femur metaphysis ( n = 6). (G) Bending test analysis of elastic modulus, maximum force, compressive ultimate stress, and post-yield displacement of femurs ( n = 6). (H and I) μCT pictures (H) and quantification (I) of trabecular bone parameters in the fifth LVs ( n = 6). (J) Pictures of calcein double labeling in femur sections (scale bar, 50 μm) and quantification of MAR ( n = 5). (K) Images of ALP staining in femur sections (scale bar, 50 μm) and quantification of ALP + cell surface per bone surface (ALP + .s/BS) in femurs ( n = 5). (L) TRAP staining pictures in femur sections (scale bar, 50 μm) and quantification of TRAP + cell surface per bone surface (TRAP + .s/BS) in femurs ( n = 5). (M and N) Serum osteocalcin levels (M) and CTX-1 levels (N) ( n = 8). Data are shown as the mean ± SEM. * P < 0.05 versus corresponding controls. P values were determined by an unpaired 2-tailed Student’s t test.

Article Snippet: Serum osteocalcin (Novus,NBP2-68151), CTX-1 (Cloud-Clone, CEA665Mu), and FGF21 (Proteintech, KE10042) in male LONP1 mKO, LONP1 HSA-MCM , MCK-ΔOTC, and LONP1/ATF4 DmKO mice and serum FGF21 (Mlbio, ml058174) in humans were conducted using enzyme-linked immunosorbent assay (ELISA) kits.

Techniques: Control, Muscles, Comparison, Expressing, Labeling, Staining

Journal: Research

Article Title: Imbalanced Skeletal Muscle Mitochondrial Proteostasis Causes Bone Loss

doi: 10.34133/research.0465

Figure Lengend Snippet: The sudden loss of LONP1 in mature muscles results in bone loss. (A) Diagrammatic drawing demonstrating the generation of tamoxifen-inducible muscle-specific deletion of Lonp1 (LONP1 HSA-MCM ). (B to I) LONP1 f/f and LONP1 HSA-MCM male mice were fed CD and injected by tamoxifen at 6 weeks old and harvested at 15 weeks old. (B) Representative quantification of LONP1 protein expression in skeletal muscles, eWAT, liver, and heart from LONP1 f/f and LONP1 HSA-MCM male mice. Quantification of the LONP1/β-actin ratio was determined by ImageJ ( n = 3). (C and D) μCT pictures (C) and quantification (D) of trabecular bone parameters in the distal femur metaphysis ( n = 5). (E) Bending test analysis of elastic modulus, maximum force, compressive ultimate stress, and post-yield displacement of femurs from LONP1 f/f and LONP1 HSA-MCM mice ( n = 5). (F and H) Pictures of ALP staining and TRAP staining in femur sections (scale bar, 50 μm). (G and I) Serum osteocalcin levels (G) and CTX-1 levels (I) of LONP1 f/f and LONP1 HSA-MCM mice ( n = 8). Data are shown as the mean ± SEM. * P < 0.05 versus corresponding controls. P values were determined by an unpaired 2-tailed Student’s t test. Ud, undetectable.

Article Snippet: Serum osteocalcin (Novus,NBP2-68151), CTX-1 (Cloud-Clone, CEA665Mu), and FGF21 (Proteintech, KE10042) in male LONP1 mKO, LONP1 HSA-MCM , MCK-ΔOTC, and LONP1/ATF4 DmKO mice and serum FGF21 (Mlbio, ml058174) in humans were conducted using enzyme-linked immunosorbent assay (ELISA) kits.

Techniques: Muscles, Injection, Expressing, Staining

Journal: Research

Article Title: Imbalanced Skeletal Muscle Mitochondrial Proteostasis Causes Bone Loss

doi: 10.34133/research.0465

Figure Lengend Snippet: Skeletal muscle-specific overexpression of mitochondrial-retained ΔOTC protein induces bone loss. (A to J) NTG and MCK-ΔOTC male mice were harvested at 8 weeks old. (A and B) μCT pictures (A) and quantification (B) of trabecular bone parameters in the distal femur metaphysis ( n = 5). (C) Bending test analysis of elastic modulus, maximum force, compressive ultimate stress, and post-yield displacement of femurs from NTG and MCK-ΔOTC mKO. (D and E) μCT pictures (D) and quantification (E) of trabecular bone parameters in the fifth LVs ( n = 5). (F) Pictures of calcein double labeling in femur sections (scale bar, 50 μm) and quantitative analysis of MAR ( n = 5). (G) Images of ALP staining in femur sections (scale bar, 50 μm) and quantification of ALP + .s/BS ( n = 5). (H) Pictures of TRAP staining in femur sections (scale bar, 50 μm) and quantification of TRAP + .s/BS ( n = 5). (I and J) Serum osteocalcin levels (I) and CTX-1 levels (J) ( n = 8). Data are shown as the mean ± SEM. * P < 0.05 versus corresponding controls. P values were determined by an unpaired 2-tailed Student’s t test.

Article Snippet: Serum osteocalcin (Novus,NBP2-68151), CTX-1 (Cloud-Clone, CEA665Mu), and FGF21 (Proteintech, KE10042) in male LONP1 mKO, LONP1 HSA-MCM , MCK-ΔOTC, and LONP1/ATF4 DmKO mice and serum FGF21 (Mlbio, ml058174) in humans were conducted using enzyme-linked immunosorbent assay (ELISA) kits.

Techniques: Over Expression, Labeling, Staining

Journal: Research

Article Title: Imbalanced Skeletal Muscle Mitochondrial Proteostasis Causes Bone Loss

doi: 10.34133/research.0465

Figure Lengend Snippet: Skeletal muscle mitochondrial proteostasis stress-induced bone loss is independent of ATF4. (A) Heatmap analysis of muscle-UPR mt myokines and Lonp1 or Atf4 genes in indicated mice (GSE192990; n = 2). (B to J) WT, LONP1 mKO, and LONP1/ATF4 DmKO male mice were harvested at 8 weeks old. (B and C) μCT pictures (B) and quantification (C) of trabecular bone parameters in the distal femur metaphysis ( n = 5 to 6). (D) Bending test analysis of elastic modulus, maximum force, compressive ultimate stress, and post-yield displacement of femurs ( n = 5). (E and F) μCT pictures (E) and quantification (F) of trabecular bone parameters in the fifth LVs ( n = 5 to 6). (G) Pictures of ALP staining in femur sections (scale bar, 50 μm) and quantification of ALP + .s/BS ( n = 5). (H) Pictures of TRAP staining in femur sections (scale bar, 50 μm) and quantification of TRAP + .s/BS ( n = 5). (I and J) Serum osteocalcin levels (I) and CTX-1 levels (J) ( n = 8). Data are shown as the mean ± SEM. * P < 0.05 versus corresponding controls. P values were determined by one-way ANOVA.

Article Snippet: Serum osteocalcin (Novus,NBP2-68151), CTX-1 (Cloud-Clone, CEA665Mu), and FGF21 (Proteintech, KE10042) in male LONP1 mKO, LONP1 HSA-MCM , MCK-ΔOTC, and LONP1/ATF4 DmKO mice and serum FGF21 (Mlbio, ml058174) in humans were conducted using enzyme-linked immunosorbent assay (ELISA) kits.

Techniques: Staining

Journal: bioRxiv

Article Title: Interleukin-3 as a Potential Bone Anabolic Agent in treating Postmenopausal Osteoporosis

doi: 10.1101/2025.07.12.664485

Figure Lengend Snippet: IL-3 alters OVX-induced changes in osteoclasts- and osteoblasts-specific gene expression. Marrow free bones from IL-3 treated animals from both Prophylactic and Therapeutic treatment strategy, as indicated, were subjected to mRNA expression profile and ( A, B ) Osteoclast-specific markers: integrinβ3, Dc-stamp, and Nfatc1 , Ctsk, Tnfrsf11a(RANK), Mmp9, Trap, and Ctr. (B, D) Osteoblast-specific markers: Runx2, Osx, Opn, Alp, Bmp2, Bmp4, Ocn, Bmp6, and Col1 were assessed by qPCR. Serum ( E, G ) CTX and ( F, H ) OCN were also measured by ELISA in both the strategies. Data is presented as mean ± SEM with n=3 mice/group for qPCR, and n=8 mice/group for ELISA.

Article Snippet: The serum level of CTX-I (CUSABIO; CSB-E12782m) and OCN (CUSABIO ; CSB-E06917m) was assessed by ELISA according to manufacturers’ instructions.

Techniques: Gene Expression, Expressing, Enzyme-linked Immunosorbent Assay

Journal: International journal of molecular sciences

Article Title: Methodology and Characterization of a 3D Bone Organoid Model Derived from Murine Cells.

doi: 10.3390/ijms25084225

Figure Lengend Snippet: Figure 2. Relative expression screening of key osteoblast and osteoclast markers by in response to differentiation media. (A) RUNX2. (B) TRAP. Relative fluorescence intensity associated with protein expression evaluated from MC3T3-E1 and RAW 264.7 which were grown in 2D culture with exposure to media containing mixtures of osteoblast (BMP2) or osteoclast (RANKL/M-CSF) differentiating factors as compared to complete culture media (negative).. Fluorescence intensity for each condition was assessed by flow cytometry. Raw fluorescence data were normalized to a cell line specific unstained control for each group. Statistical significance was evaluated by one-way ANOVA (A,B) or unpaired T test (** = p < 0.01; **** = p < 0.0001). (C) Qualitative assessment of relative gene expression (2−∆∆Ct) of osteoblastogenic (ALPL, Runx2, Sp7, Bglap, Col1a1) and osteoclastogenic (Trap) genes of undifferentiated cell lines (pooled) versus cells differentiated (pooled) in two-dimensional culture for (MC3T3-E1: day 14; RAW 264.7: day 5). Where appropriate, data are summarized as the sample mean, and error bars represent the sample standard deviation.

Article Snippet: Osteocalcin/Bglap (Bglap) ELISA assays (Novus Biologicals, Catalogue# NBP2-68151, Centennial, CO, USA) were performed per the vendor-supplied protocol.

Techniques: Expressing, Fluorescence, Flow Cytometry, Control, Gene Expression, Standard Deviation