Journal: Journal of Indian Society of Periodontology

Article Title: Biomarkers in biological fluids in adults with periodontitis and/or obesity: A meta-analysis

doi: 10.4103/jisp.jisp_512_19

Figure Lengend Snippet: Search Strategy from different databases

Article Snippet: 3. Balli et al ., 2013[ ] , Turkey, University clinic , Prospective; Interventional , 20 , 20 , O+P+: 40.56±4.11 O−P+: 39.67±3.87 , Standards of the international world workshop for classification of periodontal disease and conditions[ ] , GCF Chemerin: ELISA, Hangzhou Eastbiopharm (China);IL-6: ELISA, Boster Biological Technology (USA) , O+P + group showed higher GCF levels of chemerin and IL-6, compared to O−P + group ( P <0.05 and P <0.008 respectively) , Not included in meta-analysis (just Spearman’s rank correlation values of inflammatory markers are shown).

Techniques: Biomarker Discovery, Marker

Journal: PLoS ONE

Article Title: Inactivation of the WNT5A Alternative Promoter B Is Associated with DNA Methylation and Histone Modification in Osteosarcoma Cell Lines U2OS and SaOS-2

doi: 10.1371/journal.pone.0151392

Figure Lengend Snippet: A) Presumed enrichment of H3K4me3 in the promoter B region of normal osteoblast would block binding of the DNA methyltransferase enzyme complex, keeping the DNA unmethylated. In osteosarcoma cells, the level of H3K4me3 decreases in the promoter B region R6 and unmethylated H3 increases, allowing the Dnmt3L/Dnmt3a/b complex to bind to the H3 and de novo methylate the DNA, leading to reduced transcription of promoter B. B) NCBI Epigenomics search for CTCF binding in the WNT5A genomic region of human osteoblasts. Location of the CpG regions R3, R4, and R5 are indicated, along with promoter A exon 1 (box) and promoter B exon 1β (box at arrow). Reduced CTCF binding in osteosarcoma cells could lead to increased methylation of associated DNA regions.

Article Snippet: Human osteoblast genomic DNA was obtained from ScienCell Research Laboratories (Cat. #4609) (Carlsbad, CA).

Techniques: Blocking Assay, Binding Assay, Methylation

Journal: International Journal of Molecular Sciences

Article Title: The Species Effect: Differential Sphingosine-1-Phosphate Responses in the Bone in Human Versus Mouse

doi: 10.3390/ijms25105118

Figure Lengend Snippet: S1P reduced osteoclastogenesis of primary human monocytes. Human PBMC-isolated monocytes were treated with RANKL to induce osteoclastogenesis. ( A , B ) RANKL-treated cells were either (untreated, black bars, n = 10), further stimulated with S1P (green bars, n = 10). ( A ) Representative images of untreated or S1P-treated cells. ( B ) Osteoclast counts (TRAP-positive, multinucleated) in each condition and presented as number of osteoclasts per well. ( C , D ) RANKL-induced osteoclasts were cultured on dentine slices for 21 days in the presence of vehicle control (untreated, black, n = 6) or S1P (green, n = 3) prior to cell removal and staining of resorption pits. ( C ) Representative images of masks generated to show areas resorbed by osteoclasts. ( D ) The resorption area was recorded and displayed as a percentage of the total area. ( E – G ) Cells were untreated (grey, n = 5) or treated with RANKL only (black, n = 5) or treated with RANKL and S1P (green, n = 3 independent donors) for 8 days and RNA and gene expression of ( E ) ACP5 , ( F ) CTSK , and ( G ) MSR1 was normalised as 2 −ΔCt to β 2 M . Data are mean ± SEM from (B) n = 10 from 3 independent donors, ( D – G ) n = 3–5 independent donors. * = p < 0.05, ** = p < 0.01 and *** = p < 0.001 by paired t -test.

Article Snippet: Tubes were placed into the EasySep ‘The Big Easy’ magnet (cat: 18001, Stem Cell) for 5 min before all non-stuck cells (monocytes) were collected and 1 × 10 6 /mL were cultured overnight in human osteoclast differentiation media containing 50 ng/mL m-CSF (cat: 216-MC-025, R&D systems, Abingdon, UK).

Techniques: Isolation, Cell Culture, Control, Staining, Generated, Gene Expression

Journal: International Journal of Molecular Sciences

Article Title: The Species Effect: Differential Sphingosine-1-Phosphate Responses in the Bone in Human Versus Mouse

doi: 10.3390/ijms25105118

Figure Lengend Snippet: S1P increased osteoclastogenesis of primary murine monocytes. Murine bone marrow-derived monocytes (BMDM) were treated with M-CSF for 24 h before RANKL treatment. ( A , B ) RANKL-treated cells were either (untreated, black bars, n = 4), further stimulated with S1P (green bars, n = 4). ( A ) Representative images of untreated or S1P-treated wells stained for TRAP. ( B ) Osteoclast counts (TRAP-positive, multinucleated) in each condition and presented as number of osteoclasts per well. ( C , D ) RANKL-induced osteoclasts were cultured on hydroxyapatite coated plates for 8 days in the presence of vehicle control (untreated, black, n = 4) or S1P (green, n = 4) prior to cell removal and staining of resorption pits. ( C ) Representative images of masks generated to show areas resorbed by osteoclasts. ( D ) The resorption area was displayed as a percentage of the total area. ( E , G ) Cells were untreated (black, n = 5) or treated with RANKL only (black, n = 5) or treated with RANKL and S1P (green, n = 3 independent donors) for 8 days and gene expression for ( E ) Acp5 , ( F ) Ctsk , and ( G ) Msr1 were analysed and normalised as 2 −ΔCt to β 2 m . Data are mean ± SEM from ( B ) n = 10 from 3 independent donors, ( D – G ) n = 4 independent donors. * = p < 0.05 by unpaired t -test.

Article Snippet: Tubes were placed into the EasySep ‘The Big Easy’ magnet (cat: 18001, Stem Cell) for 5 min before all non-stuck cells (monocytes) were collected and 1 × 10 6 /mL were cultured overnight in human osteoclast differentiation media containing 50 ng/mL m-CSF (cat: 216-MC-025, R&D systems, Abingdon, UK).

Techniques: Derivative Assay, Staining, Cell Culture, Control, Generated, Gene Expression

Journal: International Journal of Molecular Sciences

Article Title: The Species Effect: Differential Sphingosine-1-Phosphate Responses in the Bone in Human Versus Mouse

doi: 10.3390/ijms25105118

Figure Lengend Snippet: S1P receptor expression changes over osteoclastogenesis but not between species. ( A , B ) Analysis of publicly available single cell sequencing datasets ( A ) GSE147174 (mouse) and ( E ) GSE162454 (human) and looking at S1P receptors (S1PR1–S1PR5). Dotplot of clustered ( A ) murine monocytes, macrophages, pre-osteoclasts, and osteoclasts or ( B ) human monocytes, macrophages, pre-osteoclasts, and osteoclasts, showing expression of S1PR1-5, where circle size represents percentage of cells expressing the gene and colour indicates average expression value. ( C – E ) BMDMs cultured (8 days) with or without M-CSF and RANKL. Undifferentiated monocytes (Mono, blue, n = 3) or osteoclasts (Oc, black, n = 3), analysed for ( C ) S1pr1 , ( D ) S1pr3 , and ( E ) S1pr4 . ( F – H ) Human monocytes derived from PBMCs cultured (8 days) with or without M-CSF and RANKL. Undifferentiated monocytes (Mono, blue, n = 5) or osteoclasts (Oc, black, n = 5), analysed for ( F ) S1PR1 , ( G ) S1PR3 , and ( H ) S1PR4 . Data are displayed as 2 −ΔCt , where ΔCt is the relative expression compared to β 2 m or β 2 M housekeeping gene. Data are mean ± SEM. *** = p < 0.001 by unpaired t -test.

Article Snippet: Tubes were placed into the EasySep ‘The Big Easy’ magnet (cat: 18001, Stem Cell) for 5 min before all non-stuck cells (monocytes) were collected and 1 × 10 6 /mL were cultured overnight in human osteoclast differentiation media containing 50 ng/mL m-CSF (cat: 216-MC-025, R&D systems, Abingdon, UK).

Techniques: Expressing, Sequencing, Cell Culture, Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: The Species Effect: Differential Sphingosine-1-Phosphate Responses in the Bone in Human Versus Mouse

doi: 10.3390/ijms25105118

Figure Lengend Snippet: Kinases involved in S1P production vary over osteoclast differentiation and species. ( A , B ) Analysis of publicly available single cell sequencing datasets GSE147174 (mouse) and GSE162454 (Human) and looking at S1P producer ( Sphk1 and Sphk2 ) and transporter (Spns2). ( A ) Dotplot of clustered murine monocytes, macrophages, pre-osteoclasts, and osteoclasts, showing expression of Sphk1 , Sphk2 , and Spns2 . ( B ) Dotplot of clustered human monocytes, macrophages, pre-osteoclasts, and osteoclasts, showing expression of SPHK1 , SPHK2 , and SPNS2 . ( C – F ) Primary mouse bone marrow-derived monocytes cultured (8 days) with or without M-CSF and RANKL. Undifferentiated monocytes (mono, blue) or osteoclasts (Oc, black), analysed for ( C ) Sphk1 (n = 3) and ( D ) Sphk2 (n = 3) gene expression and ( E ) mSPHK1 and ( F ) mSPHK2 protein expression. ( G – J ) Human monocytes derived from PBMCs cultured (8 days) with or without M-CSF and RANKL. Undifferentiated monocytes (blue) or osteoclasts (black), analysed for ( G ) SPHK1 (n = 5) and ( H ) SPHK2 (n = 5) gene expression and ( I ) hSPHK1 and ( J ) hSPHK2 protein expression. ( K , L ) Primary mouse bone marrow-derived osteoclasts ( K ) or primary human osteoclasts ( L ) were treated with or without S1P and analysed for ( K ) mSPHK2 and ( L ) hSPHK1 protein expression. Gene expression data are displayed as 2 −ΔCt normalised to β 2 m or β 2 M . Protein expression data normalised to human or mouse β-actin. Data are mean ± SEM. ** = p < 0.01 by unpaired t -test.

Article Snippet: Tubes were placed into the EasySep ‘The Big Easy’ magnet (cat: 18001, Stem Cell) for 5 min before all non-stuck cells (monocytes) were collected and 1 × 10 6 /mL were cultured overnight in human osteoclast differentiation media containing 50 ng/mL m-CSF (cat: 216-MC-025, R&D systems, Abingdon, UK).

Techniques: Sequencing, Expressing, Derivative Assay, Cell Culture, Gene Expression