Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were exposed to 10 nM of estradiol for 1, 6, 12, and 24 h. Cell morphology was observed with a light microscope (A) . Cell proliferation was analyzed using a trypan blue exclusion assay (B) . After drug administration, cellular proteins were prepared for immunoblot analyses. Levels of ERα and ERβ were immunodetected (C and E, top panels) . Amounts of β-actin were analyzed as the internal standard (C and E, bottom panels) . These protein bands were quantified and statistically analyzed (D and F) . Each value represents the mean ± SEM for n = 6. The symbol * indicates that the values significantly ( p < 0.05) differed from the respective control group, p < 0.05.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Light Microscopy, Trypan Blue Exclusion Assay, Western Blot, Control

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were exposed to 10 nM of estradiol for 1, 6, 12, and 24 h. Distribution of the ERα protein in human osteoblasts was immunodetected using an antibody with Cy3-conjugated streptavidin ( A , top panel). Cellular nuclei were stained with 4’,6-diamidino-2-phenylindole (DAPI) (middle panel). The merged signals indicated that the ERα protein had been translocated into nuclei (bottom panel). These merged fluorescent signals were quantified and statistically analyzed (B) . Each value represents the mean ± SEM for n = 6. The symbol * indicates that the value significantly differed from the respective control group, p < 0.05.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Staining, Control

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were treated with 10 nM of estradiol for 48 h. Total RNA were isolated for analysis of mitochondrial energy metabolism genes using a PCR array, containing 84 genomic genes encoding certain mitochondrial enzymes for ATP synthesis and 12 loading controls (A) . Differential expressions of these genes were measured and shown as a hot map in the order of genes indicated in panel A (B) . Percentages of upregulated, downregulated, and unchanged expressions of these genes were further statistically analyzed (C) . Also, the major genomic complex genes upregulated by estradiol in human osteoblasts were summarized (D) .

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Isolation

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were exposed to 10 nM of estradiol for 1, 6, 12, and 24 h. Distribution of the ERα protein in human osteoblasts was immunodetected using an antibody with Cy3-conjugated streptavidin ( A , top panel). Mitochondria of human osteoblasts were stained with 3,3′-dihexyloxacarbocyanine (DiOC6), a positively charged dye (middle panel). Merged signals indicated that the ERα protein had been translocated into mitochondria (bottom panels). These fluorescent signals were quantified and statistically analyzed (B) . Each value represents the mean ± SEM for n = 6. The symbol * indicates that the value significantly differed from the respective control group, p < 0.05.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Staining, Control

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were exposed to 10 nM of estradiol for 1, 3, 6, 12, 18, and 24 h. Levels of COX I and II mRNA were analyzed using an RT-PCR ( A and C , top panels). Amounts of β-actin mRNA were assayed as the internal standard (bottom panel). These bands were quantified and statistically analyzed (B and D) . A quantitative real-time PCR analysis was carried out to confirm expression of COX I mRNA in U2OS cells and rat calvarial osteoblasts (E) . Each value represents the mean ± SEM for n = 6. The symbol * indicates that the value significantly differed from the respective control group, p < 0.05.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Expressing, Control

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were exposed to 10 nM of estradiol for 1, 6, 12, and 24 h. Levels of COX I and II proteins were immunodetected (A and C, top panels) . Amounts of β-actin were analyzed as the internal standard (A and C, bottom panels) . These protein bands were quantified and statistically analyzed ( B and D ). Mitochondria of human osteoblasts were stained with 3,3′-dihexyloxacarbocyanine (DiOC6), a positively charged dye. The mitochondrial membrane potential (MMP) of human osteoblasts was determined by quantifying DiOC6-positive signals (E) . The mitochondrial enzyme activity was assayed using a colorimetric method (F) . Cellular ATP levels were quantified using a bioluminescence assay (G) . Each value represents the mean ± SEM for n = 6. The symbol * indicates that the value significantly differed from the respective control group, p < 0.05. FI, fluorescent intensity.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Staining, Membrane, Activity Assay, ATP Bioluminescent Assay, Control

Journal: Oncotarget

Article Title: Estrogen/ERα signaling axis participates in osteoblast maturation via upregulating chromosomal and mitochondrial complex gene expressions

doi: 10.18632/oncotarget.23453

Figure Lengend Snippet: Human osteoblast-like U2OS cells were treated with ERα siRNA for 24 and 48 h. Scrambled siRNA was administered to control cells as the negative standard. Levels of ERα were immunodetected ( A , top panel). Amounts of β-actin were analyzed as the internal standard (bottom panel). These protein bands were quantified and statistically analyzed (B) . After knocking-down ERα translation for 24 h, human osteoblasts were treated with estradiol for another 6 h. A quantitative PCR analysis was conducted to determine COX I mRNA expression (C) . Each value represents the mean ± SEM, n = 3. The symbols * and # indicate that a value significantly ( p < 0.05) differed from the control and estradiol-treated groups, respectively.

Article Snippet: Briefly, human osteoblast-like U2OS cells, derived from a female osteosarcoma patient, were purchase from Bioresource Collection and Research Center (Hsinchu, Taiwan).

Techniques: Control, Real-time Polymerase Chain Reaction, Expressing

Journal: Materials

Article Title: The Biocompatibility and Self-Healing Effect of a Biopolymer’s Coating on Zn Alloy for Biomedical Applications

doi: 10.3390/ma16237486

Figure Lengend Snippet: Confocal images of U2OS cells with DAPI-stained nucleus after 7 days of incubation on casein-based layer (2cas1h) on tested materials: ZnMg3.2 alloy ( a ); casein coating (2cas1h) ( b ). Additional 3D visualization from Immaris software (Bitplane Scientific Software; version 10.0.0; Olympus, Zurich, Switzerland) for ZnMg3.2 alloy ( c ), casein coating (2cas1h) ( d ).

Article Snippet: Biological characterization was based on a human osteosarcoma cell line U2OS (cat. No. 92022711; Merck, Rahway, NJ, USA).

Techniques: Staining, Incubation, Software

Journal: Cell Death & Disease

Article Title: Rho-GEF Trio regulates osteosarcoma progression and osteogenic differentiation through Rac1 and RhoA

doi: 10.1038/s41419-021-04448-3

Figure Lengend Snippet: A The Trio mRNA levels in different cancer type cell lines from the CCLE database. B The expression levels of Trio in sarcoma and normal control tissues were achieved from the TCGA database. C and D Kaplan–Meier curves show the disease-free survival and the overall survival of patients with sarcoma (TCGA database) in terms of Trio gene expression. E and F Kaplan–Meier curves show the disease-free survival and the overall survival of patients with osteosarcoma (TARGET database) in terms of Trio gene expression. G – I The expression levels of Trio in hFOB1.19, U2OS,143B were measured by Western blot and qRT-PCR. J The expression levels of Trio in osteosarcoma and normal control tissues were detected by IHC. Representative images are shown (magnification at ×200). Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01. Scale bars: 100 μm.

Article Snippet: Normal human osteoblast cell line hFOB1.19, as well as human OS cell line U2OS, were purchased from GeneChem (Shanghai, China).

Techniques: Expressing, Control, Gene Expression, Western Blot, Quantitative RT-PCR

Journal: Cell Death & Disease

Article Title: Rho-GEF Trio regulates osteosarcoma progression and osteogenic differentiation through Rac1 and RhoA

doi: 10.1038/s41419-021-04448-3

Figure Lengend Snippet: A – C Efficiency of siRNA-mediated Trio knockdown in U2OS and 143B cells was measured by qRT-PCR and Western blot. D SiRNA-mediated Trio knockdown suppressed OS cell growth, as determined by the CCK-8 assay. E Efficiency of shRNA-mediated Trio knockdown in U2OS and 143B cells was detected by GFP. F and G Efficiency of shRNA-mediated Trio knockdown in U2OS and 143B cells was measured by qRT-PCR and Western blot. H and I Trio knockdown suppressed the colony formation of U2OS and 143B cells. J Cell apoptosis was measured by cytometry. Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01. Scale bars: 100 μm.

Article Snippet: Normal human osteoblast cell line hFOB1.19, as well as human OS cell line U2OS, were purchased from GeneChem (Shanghai, China).

Techniques: Knockdown, Quantitative RT-PCR, Western Blot, CCK-8 Assay, shRNA, Cytometry

Journal: Cell Death & Disease

Article Title: Rho-GEF Trio regulates osteosarcoma progression and osteogenic differentiation through Rac1 and RhoA

doi: 10.1038/s41419-021-04448-3

Figure Lengend Snippet: A and B Wound-healing assay was used to measure the migration capacity of U2OS and 143B cells. Representative images are shown (magnification at ×100). C and D Transwell migration and Matrigel invasion assays were used to measure the migration and invasion ability of U2OS and 143B cells. Representative images are shown (magnification at ×200). E Cells of the siRNA-Trio group displayed more stress fibers and stable actin structures compared with the control group. Representative images are shown (magnification at ×400). Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01. Scale bars: 100 μm.

Article Snippet: Normal human osteoblast cell line hFOB1.19, as well as human OS cell line U2OS, were purchased from GeneChem (Shanghai, China).

Techniques: Wound Healing Assay, Migration, Control

Journal: Cell Death & Disease

Article Title: Rho-GEF Trio regulates osteosarcoma progression and osteogenic differentiation through Rac1 and RhoA

doi: 10.1038/s41419-021-04448-3

Figure Lengend Snippet: A and B EMT markers and Snail were measured by western blotting in U2OS and 143B cells after transfection. C EMT markers and transcription factors (MMP9 and Snail) were measured by qRT-PCR. D U2OS and 143B cells were subjected to immunofluorescence with E-Cadherin and N-Cadherin antibodies. Local enlarged images showed the membrane localization of E-cad. Representative images are shown (magnification at ×200). Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01. Scale bars: 100 μm.

Article Snippet: Normal human osteoblast cell line hFOB1.19, as well as human OS cell line U2OS, were purchased from GeneChem (Shanghai, China).

Techniques: Western Blot, Transfection, Quantitative RT-PCR, Immunofluorescence, Membrane

Journal: Cell Death & Disease

Article Title: Rho-GEF Trio regulates osteosarcoma progression and osteogenic differentiation through Rac1 and RhoA

doi: 10.1038/s41419-021-04448-3

Figure Lengend Snippet: A and B osteogenic markers were measured by western blotting in U2OS and 143B cells after transfection. C osteogenic markers and transcription factors were measured by qRT-PCR. D , E U2OS and 143B cells were stained with the ALP kit. Representative images are shown (magnification at ×200). F , G U2OS and 143B cells were stained with ARS solution. Representative images are shown (magnification at 200×). Data are shown as the mean ± SD. * p < 0.05, ** p < 0.01. Scale bars: 100 μm.

Article Snippet: Normal human osteoblast cell line hFOB1.19, as well as human OS cell line U2OS, were purchased from GeneChem (Shanghai, China).

Techniques: Western Blot, Transfection, Quantitative RT-PCR, Staining