Journal: Environmental Health Perspectives

Article Title: Prolonged Cadmium Exposure and Osteoclastogenesis: A Mechanistic Mouse and in Vitro Study

doi: 10.1289/EHP13849

Figure Lengend Snippet: The effects of cadmium (Cd) exposure on bone metabolic homeostasis in global Nrf2 knockout ( N r f 2 − / − ) and WT littermate ( N r f 2 + / + ) control mice. (A) Schematic illustration of the experimental design. Male mice (14 wk old) were exposed to cadmium chloride ( CdCl 2 ; 100 mg Cd/L) in drinking water for 16 wk. (B) Representative images of the left femur of mice examined by micro-CT. Scale bar: 0.5 mm . (C) Bone density estimations, including bone volume per tissue volume (BV/TV), trabecular separation (Tb.Sp), trabecular thickness (Tb.Th), trabecular number (Tb.N), total area (Tt.Ar), cortical area (Ct.Ar), structure thickness (St.Th) and cortical area per total area (Ct.Ar/Tt.Ar), were calculated using the Skyscan CT-Analyser software (version 1.1.7; Skyscan CTAn) based on micro-CT measurements. n = 6 – 9 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used. * p < 0.05 vs. N r f 2 + / + with the same treatment; # p < 0.05 vs. control (Cont) of the same genotype. (D) Results of the three-point bending test performed on the left femurs. The maximum load, breaking load, energy absorption, and stiffness were calculated using Bluehill Elements. n = 4 – 5 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used for the maximum load and breaking load; Kruskal–Wallis test with Dunn’s multiple comparisons posttest was used for the energy absorption and stiffness. * p < 0.05 vs. N r f 2 + / + with the same treatment. # p < 0.05 vs. Cont of the same genotype. (E) Plasma levels of TRAP5b and CTX-I. n = 5 – 6 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used. * p < 0.05 vs. N r f 2 + / + with the same treatment; # p < 0.05 vs. Cont of the same genotype. (F) Representative histological images of TRAP staining of the right femur of mice. Black arrow, osteoclasts. Scale bar: 1 mm . (G) Osteoclast number (OcN) and osteoclast surface (OcS) in the femur sections with TRAP staining normalized to bone perimeter (Bpm) and bone surface (BS), respectively. n = 4 – 6 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used. * p < 0.05 vs. N r f 2 + / + with the same treatment; # p < 0.05 vs. Cont of the same genotype. Summary data are provided in Tables S6–S21. Note: ANOVA, analysis of variance; CTX-I, C-terminal telopeptides of type I; micro-CT, micro-computed tomography; Nrf2, nuclear factor erythroid 2-related factor 2; SD, standard deviation; Trap, tartrate-resistant acid phosphatase; W, weeks; WT, wild type.

Article Snippet: Following centrifugation at 5,000 g at 4°C for 5 min to remove cells, the plasma was collected and stored at − 80 ° C . The activity of TRAP5b was measured using the TRAP Activity Assay Kit (E-BC-K871-M, Elabscience).

Techniques: Knock-Out, Control, Micro-CT, Software, Comparison, Clinical Proteomics, Staining, Standard Deviation

Journal: Environmental Health Perspectives

Article Title: Prolonged Cadmium Exposure and Osteoclastogenesis: A Mechanistic Mouse and in Vitro Study

doi: 10.1289/EHP13849

Figure Lengend Snippet: The impacts of cadmium (Cd) exposure on bone metabolic homeostasis in myeloid-specific Nrf2 knockout [ Nrf2 (M)-KO] and their littermate control (Cont) mice. (A) Schematic illustration of the experimental design. Male mice (14 wk old) were exposed to cadmium chloride ( CdCl 2 ; 100 mg Cd/L) in drinking water for 8 or 16 wk. (B) Representative images of the left femur of mice examined by micro-CT. Scale bar: 0.5 mm . (C) Bone density estimations, including bone volume per tissue volume (BV/TV), trabecular separation (Tb.Sp), trabecular thickness (Tb.Th), trabecular number (Tb.N), total area (Tt.Ar), cortical area (Ct.Ar), structure thickness (St.Th), and cortical area per total area (Ct.Ar/Tt.Ar), were calculated using the Skyscan CT-Analyser software (version 1.1.7; Skyscan CTAn) based on micro-CT measurements. n = 5 – 9 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used for BV/TV, Tb.Sp, Tb.Th, and Tb.N; Kruskal–Wallis test with Dunn’s multiple comparisons posttest was used for Tt.Ar, Ct.Ar, St.Th, and Ct.Ar/Tt.Ar. * p < 0.05 vs. KI with the same treatment; # p < 0.05 vs. Cont of the same genotype. (D) Representative histological images of TRAP staining of the right femur of mice. Black arrow, osteoclasts. Scale bar: 1 mm . (E) Osteoclast number (OcN) and osteoclast surface (OcS) in the femur sections with TRAP staining normalized to bone perimeter (Bpm) and bone surface (BS), respectively. n = 4 – 5 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used. * p < 0.05 vs. KI with the same treatment; # p < 0.05 vs. Cont of the same genotype. (F–H) Plasma levels of TRAP5b and CTX-I (F), P1NP and OCN (G), and RANKL and OPG (H). n = 5 – 7 . Values are expressed as mean ± SD . Two-way ANOVA with Bonferroni multiple comparison posttest was used for OcN/Bpm, OcS/BS, TRAP5b, CTX-I, P1NP, and OPG; Kruskal–Wallis test with Dunn’s multiple comparisons posttest was used for OCN and RANKL. * p < 0.05 vs. KI with the same treatment; # p < 0.05 vs. Cont of the same genotype. Summary data are provided in Tables S22–S37. Note: ANOVA, analysis of variance; CTX-I, C-terminal telopeptides of type I; KI, N f e 2 l 2 LoxP / LoxP ; KO, Nrf2 (M)-KO (knockout); micro-CT, micro-computed tomography; Nrf2, nuclear factor erythroid 2-related factor 2; OCN, osteocalcin; OPG, osteoprotegerin; P1NP, procollagen type I N-terminal propeptide; RANKL, receptor activator of nuclear factor kappa-B ligand; SD, standard deviation; Trap, tartrate-resistant acid phosphatase; W, week.

Article Snippet: Following centrifugation at 5,000 g at 4°C for 5 min to remove cells, the plasma was collected and stored at − 80 ° C . The activity of TRAP5b was measured using the TRAP Activity Assay Kit (E-BC-K871-M, Elabscience).

Techniques: Knock-Out, Control, Micro-CT, Software, Comparison, Staining, Clinical Proteomics, Standard Deviation

Journal: Nutrients

Article Title: Effects and Mechanisms of Rhus chinensis Mill. Fruits on Suppressing RANKL-Induced Osteoclastogenesis by Network Pharmacology and Validation in RAW264.7 Cells

doi: 10.3390/nu14051020

Figure Lengend Snippet: Workflow of the research. OP, osteoporosis; MTT, 3(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; TRAP, tartrate-resistant acid phosphatase; RANKL, nuclear factor-κB ligand; NF-κB, nuclear factor kappa-B; c-Fos, cellular oncogene Fos; NFATc1, nuclear factor of activated T-cells cytoplasmic 1.

Article Snippet: BCA protein assay kit, 3(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and tartrate-resistant acid phosphatase (TRAP) activity kits were supplied by Beyotime Biotechnology (Shanghai, China).

Techniques:

Journal: Nutrients

Article Title: Effects and Mechanisms of Rhus chinensis Mill. Fruits on Suppressing RANKL-Induced Osteoclastogenesis by Network Pharmacology and Validation in RAW264.7 Cells

doi: 10.3390/nu14051020

Figure Lengend Snippet: Effects of the ethanolic extract from R. chinensis fruits on RANKL-induced osteoclastogenesis in RAW264.7 cells. ( a ) TRAP staining; ( b ) Relative area of TRAP-positive signal, normalized with the K group; ( c ) Quantitative results of TRAP activity. All the values are expressed as mean ± SD ( n = 3). Groups with different letters are significantly different ( p < 0.05). The TRAP positive cells after TRAP staining were quantified using Image-Pro Plus software. The quantitative result is expressed in terms of density mean (density mean = density sum/area sum) and the TRAP positive cells were quantified by normalizing with group K. K, the Control group; M, 50 ng/mL RANKL; RL, 50 ng/mL RANKL and 50 µg/mL of ethanolic extract; RH, 50 ng/mL RANKL and 100 µg/mL of ethanolic extract; TRAP, tartrate-resistant acid phosphatase; RANKL, receptor activator of nuclear factor-κB ligand.

Article Snippet: BCA protein assay kit, 3(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and tartrate-resistant acid phosphatase (TRAP) activity kits were supplied by Beyotime Biotechnology (Shanghai, China).

Techniques: Staining, Activity Assay, Software, Control