Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: miR-486-5p expression and methylation in osteosarcoma cell lines and patient samples. A . miR-486-5p expression level in normal bone ( n = 6), cell lines ( n = 17) and patient samples ( n = 9) using qRT-PCR. The expression level was quantified for the groups of samples. The values are shown relative to mean expression of bone. B . DNA methylation and miRNA expression levels in normal bone samples ( n = 4) and osteosarcoma cell lines ( n = 19) using arrays. The expression level of miRNAs (log2) and DNA methylation level (Beta, probe cg00176210) were obtained using Agilent miRNA array v2 and Illumina Infinium Methylation27 BeadChip technology, respectively. Values are given as mean (SD) with whiskers from min to max. P -values * < 0.01 and ** < 0.0001

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Expressing, Methylation, Quantitative RT-PCR, DNA Methylation Assay

Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: Expression and correlation of miR-486-5p and hsa-mir-486-1 (pri-mir486) in osteosarcoma cell lines upon 5-Aza-2′-deoxycytidine treatment. Relative expression level A. miR-485-5p and B. pri-mir486 after 72 h of 5-Aza treatment . The values are shown relative to treated cell lines (set to 1, horizontal line). Induction of > 30% shown as dotted line. Values are given as mean (SD). Correlation between expresion level of miR-485-5p and pri-mir486 in C. untreated and D. 5-Aza treated cell lines. The expression levels are quantified using qRT-PCR, and normalized against RNU44 for miR-486-5p and GAPDH for pri-mir486

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Expressing, Quantitative RT-PCR

Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: Expression and correlation of miR-486-5p and ANK1 before and after 5-Aza-2′-deoxycytidine treatment in osteosarcoma cell lines. A . Quantification of expression of miR-486-5p and ANK1 versions 1–4 in untreated cells. B . The induction of miR-486-5p and ANK1 versions 1–4 after 5-Aza treatment. The induction fold change is calculated as the ratio between expression of untreated and 5-Aza treated transcripts as quantified by qRT-PCR. The expression is normalized against RNU44 for miR-486-5p and GAPDH for ANK1 . Pearson’s Correlation r is calculated with and without the outlier sample MG-63

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Expressing, Quantitative RT-PCR

Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: Genome-wide methylation profile of the mir-486/ANK1 locus and miR-486-5p expression levels in osteosarcoma patient samples. A. Representation of average methylation profile across the mir-486 / ANK1 locus in osteosarcoma patient samples ( n = 10) and bone ( n = 4). Methylation level (Beta) is shown for the individual CpG sites (ticks on horizontal axis) from the HumanMethylation450 BeadChips. The probes are ordered along the locus and intervals are not in scale. CGIs are shown as grey boxes with number refering to CpG count (from UCSC Genome Browser NCBI, GRCh37/hg19 assembly). Horizontal lines below plot show representative transcript variants of the respective mRNA genes (RefSeq) with exons as vertical bars, not in scale. Shaded vertical box: CGI shown in detail in B. Black, osteosarcomas; Grey, bone. B. Probe level methylation for CGI CpG79 in osteosarcoma patient samples and bone. Methylation levels as determined by Infinium 450 k arrays are given in Beta values for the individual CpG sites (cg). The selected CpGs within CGI CpG79 are highlighted with a shaded vertical box in A. Methylated: Beta 0.7–1.0; partially methylated: Beta 0.3–0.7; unmethylated: Beta< 0.3. C. Expression level of miR-486-5p in osteosarcoma patient samples and bone based on qRT-PCR. The osteosarcoma samples are grouped based on methylation status for probe cg08194989 on Infinium 450 k arrays, while the bone samples are shown as one group. Horizontal line: mean value

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Genome Wide, Methylation, Expressing, Quantitative RT-PCR

Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: Methylation distributions in osteosarcoma assessed by quantitative methylation-specific PCR. The methylation was measured for the groups of normal bones and osteosarcoma cell lines, xenografts and patient samples. PMR, percent of methylated reference. Horizontal black line: mean values

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Methylation

Journal: BMC Genomics

Article Title: miR-486-5p expression is regulated by DNA methylation in osteosarcoma

doi: 10.1186/s12864-022-08346-6

Figure Lengend Snippet: Introduction of miR-486-5p in osteosarcoma cell lines. Osteosarcoma cells (OSA, OHS and U-2 OS) were transiently transfected with synthetic miR-486-5p mimics or a negative control. Cellular proliferation rates were determined by live cell imaging for 48 h using the IncuCyte, measuring cell confluence over time. One representative experiment of three is shown ( n = 3). Error bars represent the standard error of means of values for replicate wells ( n ≥ =5)

Article Snippet: A panel of human osteosarcoma cell lines ( n = 21) composed of 143B, CAL-72, G-292, HAL, HOS, IOR/MOS, IOR/OS9, IOR/OS10, IOR/OS14, IOR/OS15, IOR/OS18, IOR/SARG, KPD, MG-63, MHM, MNNG/HOS, OHS, OSA, Saos-2, U-2 OS and ZK-58 were obtained from ATCC ( www.lgcstandards-atcc.org ), DSMZ ( http://www.dsmz.de/ ) or research partners in the EU funded EuroBoNeT project [ ].

Techniques: Transfection, Negative Control, Live Cell Imaging

Journal:

Article Title: Modulation of Gamma Interferon-Induced Major Histocompatibility Complex Class II Gene Expression by Porphyromonas gingivalis Membrane Vesicles

doi: 10.1128/IAI.70.3.1185-1192.2002

Figure Lengend Snippet: Inhibition of class II HLA-DRα surface expression in IFN-γ-inducible cells by P. gingivalis membrane vesicles. HUVECs, 143B osteosarcoma cells, MRC-5 fibroblasts, and THP-1 monocytic cells were treated with vesicles (OMVs, 30 μg of protein/ml) alone or IFN-γ (250 U/ml) in the absence or presence of vesicles at 37°C for 3 days. Control cells received no treatment. Cells were stained with either phycoerythrin-conjugated mouse anti-HLA-DR MAb or irrelevant isotype-matched MAb. Stained cells were analyzed by flow cytometry. Averages and standard deviations were calculated from three separate experiments.

Article Snippet: Other cell lines used, obtained from ATCC, included human embryonic lung fibroblasts (MRC-5; ATCC CCL 171), 143B human osteosarcoma cells (ATCC CRL 8304), and THP-1 monocytic cells (ATCC TIB 202).

Techniques: Inhibition, Expressing, Membrane, Control, Staining, Flow Cytometry

Journal: Journal of Translational Medicine

Article Title: The integrin α2-osteoclast axis: a key driver of bone destruction and therapeutic target in osteosarcoma

doi: 10.1186/s12967-025-06906-4

Figure Lengend Snippet: ITGA2 was upregulated in the osteosarcoma tissues and cells. A ITGA2 expression in normal and tumor tissues in unpaired samples in the respective datasets ( GSE19276 , GSE126209 , GSE42352 ). B , C IHC images revealed differential expression of ITGA2 in osteosarcoma (top) and normal bone (bottom). Scale bar: 80 µm and 200 µm. Data with error bars are shown as mean ± SEM. D The expression of ITGA2 in osteosarcoma cell lines (143B, U2OS, MG63 and SAOS) and hFOB cells was measured by WB. Error bars represent Mean ± SD. No significance (ns), p > 0.05, ****p < 0.0001 by One-way ANOVA

Article Snippet: Herein, 143B and U2OS cells were maintained overnight at a density of 2000 cells/ well in a 96-well plate, followed by treatment with various concentrations of ITGA2 inhibitor (E7820, TargetMol Co., Ltd., China, Cas.

Techniques: Expressing, Quantitative Proteomics

Journal: Journal of Translational Medicine

Article Title: The integrin α2-osteoclast axis: a key driver of bone destruction and therapeutic target in osteosarcoma

doi: 10.1186/s12967-025-06906-4

Figure Lengend Snippet: Transcriptional characteristics of 143B cells after inhibition of ITGA2. A Expression levels of ITGA2 in control and E7820-treated 143B cells, showing a significant difference (p = 0.03). B The differentially expressed genes in E7820-treated 143B cells compared to untreated controls. Red dots indicate upregulated genes, blue dots indicate downregulated genes, and gray dots represent non-significant genes. C GSVA demonstrated the expression profiles of genes related to osteoclast differentiation and signaling pathways across control (C1-C4) and E7820 treated (T1-T4) groups. D GO enrichment analysis and ( E ) KEGG pathway enrichment analyses of the DEGs in E7820-treated versus untreated groups in OS cells. F Venn diagrams compared the overlap of ITGA2-related DEGs between different datasets

Article Snippet: Herein, 143B and U2OS cells were maintained overnight at a density of 2000 cells/ well in a 96-well plate, followed by treatment with various concentrations of ITGA2 inhibitor (E7820, TargetMol Co., Ltd., China, Cas.

Techniques: Inhibition, Expressing, Control, Protein-Protein interactions

Journal: Journal of Translational Medicine

Article Title: The integrin α2-osteoclast axis: a key driver of bone destruction and therapeutic target in osteosarcoma

doi: 10.1186/s12967-025-06906-4

Figure Lengend Snippet: In vitro and in vivo anti osteosarcoma effects of silencing ITGA2. A CCK-8 assay showed the inhibitory effects of E7820 on the viability of 143B and U2OS cell lines, with IC50 values calculated from dose–response curves. B , C Flow cytometric analysis of apoptosis in 143B and U2OS cells treated with E7820 for 48 h. D Colony formation assay illustrated the dose-dependent inhibitory effects of E7820 on colony formation in 143B and U2OS cell lines. E Transwell assay investigated migration and invasion abilities while ITGA2 was inhibited in 143B and U2OS cell lines. Scale bar: 50 µm. F Scratch assay showed the inhibitory effects of E7820 on cell migration in 143B and U2OS cell lines. Scale bar: 50 µm. G Representative images of tumor xenografts in a nude mouse model, compared normal controls with E7820-treated groups. H Quantitative analysis of tumor weight and volume in the xenograft model, showed significant reductions in both parameters in the E7820-treated group. Error bars represent Mean ± SD. No significance (ns), p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001

Article Snippet: Herein, 143B and U2OS cells were maintained overnight at a density of 2000 cells/ well in a 96-well plate, followed by treatment with various concentrations of ITGA2 inhibitor (E7820, TargetMol Co., Ltd., China, Cas.

Techniques: In Vitro, In Vivo, CCK-8 Assay, Colony Assay, Transwell Assay, Migration, Wound Healing Assay

Journal: Journal of Translational Medicine

Article Title: The integrin α2-osteoclast axis: a key driver of bone destruction and therapeutic target in osteosarcoma

doi: 10.1186/s12967-025-06906-4

Figure Lengend Snippet: Effects of ITGA2 inhibition or knockdown on osteosarcoma osteolysis. A Representative X-ray images on Day 28 at the end point of the experiment. The location of bone destruction indicated by the white arrow. B Representative μCT, H&E staining and TRAP staining images from the experiment. n = 3 per group. T: tumor; B: bone; BM: bone marrow; M: muscle; BV/TV: Bone Volume/Total Volume; Tb.Sp: Trabecular Separation/Spacing; BMD: Bone Mineral Density. Scale bars: 100 μm. C , D Western blot analysis of ITGA2, RANKL, MMP9, MMP13 and OPN expression in 143B and U2OS cell lines with ITGA2 inhibition or knockdown. E ELISA quantification of MMP9 in the culture supernatant of 143B and U2OS cells after ITGA2 inhibition or knockdown. Data presented as mean ± SEM. No significance (ns), p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 by One-way ANOVA

Article Snippet: Herein, 143B and U2OS cells were maintained overnight at a density of 2000 cells/ well in a 96-well plate, followed by treatment with various concentrations of ITGA2 inhibitor (E7820, TargetMol Co., Ltd., China, Cas.

Techniques: Inhibition, Knockdown, Staining, Western Blot, Expressing, Enzyme-linked Immunosorbent Assay