human wildtype full-length sp7 coding sequence Search Results


human osx cdna  (OriGene)
93
OriGene human osx cdna
Figure 1. Protein levels of β-catenin (β-cat) and <t>osterix</t> <t>(OSX)</t> in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.
Human Osx Cdna, supplied by OriGene, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/human+wildtype+full-length+sp7+coding+sequence/SP7+(NM_001173467)+Human+Untagged+Clone/pm26496941-30-1-9
Average 93 stars, based on 1 article reviews
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anti p53  (Cell Signaling Technology Inc)
97
Cell Signaling Technology Inc anti p53
Figure 1. Protein levels of β-catenin (β-cat) and <t>osterix</t> <t>(OSX)</t> in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.
Anti P53, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/human+wildtype+full-length+sp7+coding+sequence/p53+Antibody/pmc09953509-72-39-41
Average 97 stars, based on 1 article reviews
anti p53 - by Bioz Stars, 2026-09
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pd-l1 rabbit  (Quartett GmbH)
90
Quartett GmbH pd-l1 rabbit
Figure 1. Protein levels of β-catenin (β-cat) and <t>osterix</t> <t>(OSX)</t> in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.
Pd L1 Rabbit, supplied by Quartett GmbH, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/human+wildtype+full-length+sp7+coding+sequence/qr1/pmc08317124__jitc___2021___002844supp001-62-258-257
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3873s  (Cell Signaling Technology Inc)
97
Cell Signaling Technology Inc 3873s
Figure 1. Protein levels of β-catenin (β-cat) and <t>osterix</t> <t>(OSX)</t> in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.
3873s, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 97/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/human+wildtype+full-length+sp7+coding+sequence/alpha-Tubulin+Mouse+mAb/pm31636113-498-175-173
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Image Search Results


Figure 1. Protein levels of β-catenin (β-cat) and osterix (OSX) in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 1. Protein levels of β-catenin (β-cat) and osterix (OSX) in human pre-osteoblastic and bone marrow stromal cells. In (A) HS-27A human bone marrow stromal cells and (B) MG-63 human pre-osteoblastic cells, the protein levels of cytoplasmic/soluble β-cat were measured by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h (lane 5), cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX-shRNA (active β-cat + OSX-shRNA, lane 6), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h (lane 7). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of the OSX and the cytoplasmic/soluble β-cat blots was normalized against that of the GAPDH blot to obtain a relative blot density. In cells overexpressing ∆N151/active β-cat, the relative density of ∆N151/active β-cat instead of that of wild-type soluble β-cat was calculated and is shown in the bar graph. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Western Blot, Control, Stable Transfection, Transfection, Plasmid Preparation, Transduction, shRNA

Figure 2. β-catenin (β-cat) signaling luciferase reporter activities and target gene mRNA levels in human pre-osteoblastic and bone marrow stromal cells. (A) HS-27A (left panel) and MG-63 (right panel) cells were transfected with TOPflash, a synthetic β-cat signaling luciferase reporter, or FOPflash, a nega tive control reporter. Thirty hours later, luciferase activity was determined in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h, cells stably transfected with constitutively active (∆N151) β-cat and stably trans duced with osterix (OSX)-shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h. Luciferase activity was measured 30 h after transfection and expressed as a fold change to that of NC (designated as 1). aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA. (B) The mRNA levels of OSX and established β-cat signaling target genes c-Myc and c-Jun were measured by RT-qPCR in the HS-27A (left panel) and MG-63 (right panel) cells. The mRNA levels of c-Myc, c-Jun and OSX were normalized against those of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). *P<0.05 vs. controls (NC, VC and SC).

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 2. β-catenin (β-cat) signaling luciferase reporter activities and target gene mRNA levels in human pre-osteoblastic and bone marrow stromal cells. (A) HS-27A (left panel) and MG-63 (right panel) cells were transfected with TOPflash, a synthetic β-cat signaling luciferase reporter, or FOPflash, a nega tive control reporter. Thirty hours later, luciferase activity was determined in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) for 30 h, cells stably transfected with constitutively active (∆N151) β-cat and stably trans duced with osterix (OSX)-shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) for 30 h. Luciferase activity was measured 30 h after transfection and expressed as a fold change to that of NC (designated as 1). aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA. (B) The mRNA levels of OSX and established β-cat signaling target genes c-Myc and c-Jun were measured by RT-qPCR in the HS-27A (left panel) and MG-63 (right panel) cells. The mRNA levels of c-Myc, c-Jun and OSX were normalized against those of glyceraldehyde-3-phosphate dehydrogenase (GAPDH). *P<0.05 vs. controls (NC, VC and SC).

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Luciferase, Transfection, Control, Activity Assay, Stable Transfection, Plasmid Preparation, Transduction, shRNA, Quantitative RT-PCR

Figure 3. Mutational analysis of putative c-Jun binding sites in the human osterix (OSX) gene promoter. (A) In the OSX gene promoter sequence in a commercial human OSX promoter/luciferase reporter (SwitchGear Genomics), the ATG translation start codon is marked as +1. A putative c-Jun binding site at -858/-852 and a putative AP-1/c-Jun/c-Fos binding site at -669/-657 (AP-1, -669/-661; c-Jun, -667/-661; c-Fos, -666/-657) are underlined and in boldface. The start sites of OSX transcript 1 (Accession no. NM_001173467.1) and transcript 2 (Accession no. NM_152860.1) are underlined in boldface at -574 and -107, respectively. (B) Schematic representation of the wild-type (WT) and mutant (Mut) human OSX promoter/luciferase reporter constructs. Mutated sequence at the -858/-852 (Mut858) and the -669/-657 (Mut669) putative transcription factor binding sites are represented in boldface. (C) WT and Mut human OSX promoter/luciferase reporter constructs were respectively transfected into the HS-27A (left panel) and MG-63 (right panel) cells, which were stably transfected with the empty pcDNA3.1 vector (VC) or constitutively active (∆N151) β-catenin (active β-cat). Luciferase activity was measured 30 h following transfection and expressed as a fold change to that of VC (designated as 1). *P<0.05 vs. VC.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 3. Mutational analysis of putative c-Jun binding sites in the human osterix (OSX) gene promoter. (A) In the OSX gene promoter sequence in a commercial human OSX promoter/luciferase reporter (SwitchGear Genomics), the ATG translation start codon is marked as +1. A putative c-Jun binding site at -858/-852 and a putative AP-1/c-Jun/c-Fos binding site at -669/-657 (AP-1, -669/-661; c-Jun, -667/-661; c-Fos, -666/-657) are underlined and in boldface. The start sites of OSX transcript 1 (Accession no. NM_001173467.1) and transcript 2 (Accession no. NM_152860.1) are underlined in boldface at -574 and -107, respectively. (B) Schematic representation of the wild-type (WT) and mutant (Mut) human OSX promoter/luciferase reporter constructs. Mutated sequence at the -858/-852 (Mut858) and the -669/-657 (Mut669) putative transcription factor binding sites are represented in boldface. (C) WT and Mut human OSX promoter/luciferase reporter constructs were respectively transfected into the HS-27A (left panel) and MG-63 (right panel) cells, which were stably transfected with the empty pcDNA3.1 vector (VC) or constitutively active (∆N151) β-catenin (active β-cat). Luciferase activity was measured 30 h following transfection and expressed as a fold change to that of VC (designated as 1). *P<0.05 vs. VC.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Binding Assay, Sequencing, Luciferase, Mutagenesis, Construct, Transfection, Stable Transfection, Plasmid Preparation, Activity Assay

Figure 4. Specific protein-binding activity at the putative c-Jun binding site in human osterix (OSX) gene promoter. (A) Oligonucleotide WT865/836 contained the human OSX gene promoter sequence from -865 to -836 encompassing the -858/-852 putative c-Jun binding site. Oligonucleotide Mut865/836 contained the same sequence as WT865/836 except for mutations at the -858/-852 putative c-Jun binding site (underlined and in boldface). (B) Electrophoretic mobility shift assays (EMSAs) were performed using WT865/836 as the radiolabeled probe in the presence of an equal amount of nuclear extract from HS-27A cells stably transfected with the empty pcDNA3.1 vector (lane 2) or HS-27A cells stably transfected with constitutively active (∆N151) β-cat (active β-cat) (lanes 3-8). Lane 1, radiolabeled probe only; lanes 2-3, control reaction; lane 4, a 100-fold molar excess of unlabeled oligonucleotide WT865/836 as competitor; lane 5, a 100-fold molar excess of unlabeled oligonucleotide Mut865/836 as competitor; lane 6, control serum; lane 7, anti-cJun antibody; lane 8, anti-cFos antibody. Major protein- DNA complexes and supershifted complexes are indicated.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 4. Specific protein-binding activity at the putative c-Jun binding site in human osterix (OSX) gene promoter. (A) Oligonucleotide WT865/836 contained the human OSX gene promoter sequence from -865 to -836 encompassing the -858/-852 putative c-Jun binding site. Oligonucleotide Mut865/836 contained the same sequence as WT865/836 except for mutations at the -858/-852 putative c-Jun binding site (underlined and in boldface). (B) Electrophoretic mobility shift assays (EMSAs) were performed using WT865/836 as the radiolabeled probe in the presence of an equal amount of nuclear extract from HS-27A cells stably transfected with the empty pcDNA3.1 vector (lane 2) or HS-27A cells stably transfected with constitutively active (∆N151) β-cat (active β-cat) (lanes 3-8). Lane 1, radiolabeled probe only; lanes 2-3, control reaction; lane 4, a 100-fold molar excess of unlabeled oligonucleotide WT865/836 as competitor; lane 5, a 100-fold molar excess of unlabeled oligonucleotide Mut865/836 as competitor; lane 6, control serum; lane 7, anti-cJun antibody; lane 8, anti-cFos antibody. Major protein- DNA complexes and supershifted complexes are indicated.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Protein Binding, Activity Assay, Binding Assay, Sequencing, Electrophoretic Mobility Shift Assay, Stable Transfection, Transfection, Plasmid Preparation, Control

Figure 5. Knockdown of c-Jun abolished the effect of β-catenin (β-cat) on the expression of osterix (OSX) in human pre-osteoblastic and bone marrow stromal cells. (A) The protein levels of c-Jun and OSX in HS-27A (left panel) and MG-63 (right panel) cells were measurd by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), and cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with lentiviral shRNA against c-Jun (active β-cat + cJun-shRNA, lane 5). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of (B) c-Jun and (C) OSX blots was normalized against that of the GAPDH blot to obtain a relative blot density, respectively. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. (D) The mRNA levels of OSX were measured by RT-qPCR assays in the HS-27A (left panel) and MG-63 (right panel) cells and normalized against those of GAPDH. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 5. Knockdown of c-Jun abolished the effect of β-catenin (β-cat) on the expression of osterix (OSX) in human pre-osteoblastic and bone marrow stromal cells. (A) The protein levels of c-Jun and OSX in HS-27A (left panel) and MG-63 (right panel) cells were measurd by western blot analyses in normal control cells (NC, lane 1), cells stably transfected with the empty pcDNA3.1 vector (VC, lane 2), cells stably transduced with scramble control shRNA (SC, lane 3), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat, lane 4), and cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with lentiviral shRNA against c-Jun (active β-cat + cJun-shRNA, lane 5). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) blotting was used as a loading control. Density of (B) c-Jun and (C) OSX blots was normalized against that of the GAPDH blot to obtain a relative blot density, respectively. Three independent experiments were performed for each western blot analysis. Data are expressed as the means + SD. (D) The mRNA levels of OSX were measured by RT-qPCR assays in the HS-27A (left panel) and MG-63 (right panel) cells and normalized against those of GAPDH. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Knockdown, Expressing, Western Blot, Control, Stable Transfection, Transfection, Plasmid Preparation, Transduction, shRNA, Quantitative RT-PCR

Figure 7. Effect of overexpression and inhibition of β-catenin (β-cat) and/or osterix (OSX) on calcium deposition in human pre-osteoblastic and bone marrow stromal cells. For osteoblast differentiation, (A) HS-27A and (B) MG-63 cells (5,000 cells/well) were cultured in osteoblastogenic medium. On day 14 in HS-27A cells and on day 28 in MG-63 cells, calcium deposition was measured using a calcium (CPC) liquicolor kit (Stanbio Laboratory) in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) during the entire osteoblastogenic culture period, cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with Osterix (OSX)-shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) during the entire osteoblastogenic culture period. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 7. Effect of overexpression and inhibition of β-catenin (β-cat) and/or osterix (OSX) on calcium deposition in human pre-osteoblastic and bone marrow stromal cells. For osteoblast differentiation, (A) HS-27A and (B) MG-63 cells (5,000 cells/well) were cultured in osteoblastogenic medium. On day 14 in HS-27A cells and on day 28 in MG-63 cells, calcium deposition was measured using a calcium (CPC) liquicolor kit (Stanbio Laboratory) in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) during the entire osteoblastogenic culture period, cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with Osterix (OSX)-shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) during the entire osteoblastogenic culture period. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Over Expression, Inhibition, Cell Culture, Control, Stable Transfection, Transfection, Plasmid Preparation, Transduction, shRNA

Figure 6. Effect of overexpression and inhibition of β-catenin (β-cat) and/or osterix (OSX) on alkaline phosphatase (ALP) activity in human pre-osteoblastic and bone marrow stromal cells. For osteoblast differentiation, (A) HS-27A and (B) MG-63 cells (5,000 cells/well) were cultured in osteoblastogenic medium. ALP activity was measured using a colorimetric ALP assay kit (Abcam) on day 14 of osteoblastogenic culture in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) during the entire osteoblastogenic culture period, cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) during the entire osteoblastogenic culture period. ALP activity was normalized against the total protein concentration in each sample. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Journal: International journal of molecular medicine

Article Title: β-catenin signaling induces the osteoblastogenic differentiation of human pre-osteoblastic and bone marrow stromal cells mainly through the upregulation of osterix expression.

doi: 10.3892/ijmm.2015.2382

Figure Lengend Snippet: Figure 6. Effect of overexpression and inhibition of β-catenin (β-cat) and/or osterix (OSX) on alkaline phosphatase (ALP) activity in human pre-osteoblastic and bone marrow stromal cells. For osteoblast differentiation, (A) HS-27A and (B) MG-63 cells (5,000 cells/well) were cultured in osteoblastogenic medium. ALP activity was measured using a colorimetric ALP assay kit (Abcam) on day 14 of osteoblastogenic culture in normal control cells (NC), cells stably transfected with the empty pcDNA3.1 vector (VC), cells stably transduced with scramble control shRNA (SC), cells stably transfected with constitutively active (∆N151) β-cat (active β-cat), cells treated with selective β-cat signaling inhibitor CCT031374 (50 µM) during the entire osteoblastogenic culture period, cells stably transfected with constitutively active (∆N151) β-cat and stably transduced with OSX shRNA (active β-cat + OSX-shRNA), and cells stably transfected with OSX and treated with CCT031374 (50 µM) during the entire osteoblastogenic culture period. ALP activity was normalized against the total protein concentration in each sample. aP<0.05 vs. controls (NC, VC and SC); bP<0.05 vs. active β-cat; cP<0.05 vs. CCT031374; dP<0.05 vs. active β-cat + OSX-shRNA.

Article Snippet: The human OSX cDNA clone (SC328709) was purchased from OriGene Technologies (Beijing, China), and the full-length OSX cDNA sequence was subcloned into the pcDNA 3.1 plasmid (Life Technologies).

Techniques: Over Expression, Inhibition, Activity Assay, Cell Culture, ALP Assay, Control, Stable Transfection, Transfection, Plasmid Preparation, Transduction, shRNA, Protein Concentration