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Figure 1. Identification of transcription factors predominantly expressed in articular cartilage cells. (A) Total RNA was isolated from SFZ, CC, LB, and C3H10T1/2 (10T1/2) cells. <t>Gdf5</t> and Prg4 expression was analyzed by RT-qPCR in triplicate. (B) Total RNA from SFZ cells and CC was analyzed by microarray. Genes with expression exceeding 100 in terms of the SFZ raw values were counted as genes expressed in SFZ cells. The number indicates the number of genes expressed more than two-fold in SFZ cells compared with the level in CC. The panel on the right shows 11 transcription factors among the genes with expression in SFZ cells more than two-fold that in CC. (C) Total RNA was isolated from SFZ, CC, LB, and 10T1/2 cells. Indicated gene expression was analyzed by RT-qPCR in triplicate. Data are the mean ± SEM.

Gdf5, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 92 stars, based on 8 article reviews

gdf5 - by Bioz Stars, 2026-06

92/100 stars

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1) Product Images from "HOXA10 promotes Gdf5 expression in articular chondrocytes."

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

Journal: Scientific reports

doi: 10.1038/s41598-023-50318-7

Figure Legend Snippet: Figure 1. Identification of transcription factors predominantly expressed in articular cartilage cells. (A) Total RNA was isolated from SFZ, CC, LB, and C3H10T1/2 (10T1/2) cells. Gdf5 and Prg4 expression was analyzed by RT-qPCR in triplicate. (B) Total RNA from SFZ cells and CC was analyzed by microarray. Genes with expression exceeding 100 in terms of the SFZ raw values were counted as genes expressed in SFZ cells. The number indicates the number of genes expressed more than two-fold in SFZ cells compared with the level in CC. The panel on the right shows 11 transcription factors among the genes with expression in SFZ cells more than two-fold that in CC. (C) Total RNA was isolated from SFZ, CC, LB, and 10T1/2 cells. Indicated gene expression was analyzed by RT-qPCR in triplicate. Data are the mean ± SEM.

Techniques Used: Isolation, Expressing, Quantitative RT-PCR, Microarray, Gene Expression

Figure 2. Generation of Gdf5-HiBiT screening system. (A) Schematic diagram of Gdf5-HiBiT allele. The stop codon of Gdf5 and the knocked-in position of HiBiT tag are indicated. (B) Gdf5-HiBiT KI mice were generated using the Technique for Animal Knockout system by Electroporation (TAKE) method based on CRISPR/Cas9. Genomic DNA sequence analysis of the Gdf5 gene was performed, which confirmed that the genome of Gdf5- HiBiT KI allele had been edited correctly. Two possible off-target sites for the CRISPR gRNA (target sequence: TCGTGGAATCTTGTGGCTGC) are shown. Genomic DNA sequence analysis of two off-target sites was performed, which confirmed that the sequences around off-target sites were intact. (C) Genomic PCR analysis of WT and Gdf5-HiBiT KI mice was performed. A representative result is shown. The original gel is presented in Supplementary Fig. 1. (D) SFZ cells were isolated from WT and Gdf5-HiBiT KI mice. The cells were cultured for 2 days and then the supernatants were collected. DMEM, 10% FBS DMEM, and supernatants of WT and Gdf5-HiBiT KI SFZ cells were subjected to HiBiT measurement (n = 3). RLU: relative light unit. Data are the mean ± SEM (****: P < 0.0001).

Figure Legend Snippet: Figure 2. Generation of Gdf5-HiBiT screening system. (A) Schematic diagram of Gdf5-HiBiT allele. The stop codon of Gdf5 and the knocked-in position of HiBiT tag are indicated. (B) Gdf5-HiBiT KI mice were generated using the Technique for Animal Knockout system by Electroporation (TAKE) method based on CRISPR/Cas9. Genomic DNA sequence analysis of the Gdf5 gene was performed, which confirmed that the genome of Gdf5- HiBiT KI allele had been edited correctly. Two possible off-target sites for the CRISPR gRNA (target sequence: TCGTGGAATCTTGTGGCTGC) are shown. Genomic DNA sequence analysis of two off-target sites was performed, which confirmed that the sequences around off-target sites were intact. (C) Genomic PCR analysis of WT and Gdf5-HiBiT KI mice was performed. A representative result is shown. The original gel is presented in Supplementary Fig. 1. (D) SFZ cells were isolated from WT and Gdf5-HiBiT KI mice. The cells were cultured for 2 days and then the supernatants were collected. DMEM, 10% FBS DMEM, and supernatants of WT and Gdf5-HiBiT KI SFZ cells were subjected to HiBiT measurement (n = 3). RLU: relative light unit. Data are the mean ± SEM (****: P < 0.0001).

Techniques Used: Generated, Knock-Out, Electroporation, CRISPR, Sequencing, Isolation, Cell Culture

Figure 3. Role of HOXA10 in Gdf5 expression in articular cartilage cells. (A) SFZ cells isolated from WT mice were infected with empty (control) or indicated lentiviruses. Indicated gene expression was analyzed by RT-qPCR in duplicate. (B) Schematic diagram of Gdf5-HiBiT screening system. (C) SFZ cells were isolated from Gdf5-HiBiT KI mice and were plated on a 96-well plate. Gdf5-HiBiT KI SFZ cells were infected with the indicated lentiviruses. One day later, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). (E) SFZ cells isolated from WT mice were infected with empty (control), shHoxa10-1, or shHoxa10-2 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Figure Legend Snippet: Figure 3. Role of HOXA10 in Gdf5 expression in articular cartilage cells. (A) SFZ cells isolated from WT mice were infected with empty (control) or indicated lentiviruses. Indicated gene expression was analyzed by RT-qPCR in duplicate. (B) Schematic diagram of Gdf5-HiBiT screening system. (C) SFZ cells were isolated from Gdf5-HiBiT KI mice and were plated on a 96-well plate. Gdf5-HiBiT KI SFZ cells were infected with the indicated lentiviruses. One day later, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). (E) SFZ cells isolated from WT mice were infected with empty (control), shHoxa10-1, or shHoxa10-2 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Techniques Used: Expressing, Isolation, Infection, Control, Gene Expression, Quantitative RT-PCR, Cell Culture

Figure 5. Effect of HOXA10 on the Gdf5 gene promoter. (A) The ATAC-Seq database (articular chondrocytes: SRX13791211, costal chondrocytes: SRX11156876) from ChIP-Atlas was analyzed by integrative genomics viewer IGV2.8.6. A Gdf5 gene promoter (1393 bp) exhibits an open chromatin region on the Gdf5 gene in articular chondrocytes. (B) Schematic diagram of luciferase reporter construction with mouse Gdf5 gene promoter (− 1081 to + 312). A putative HOXA10 binding motif (− 538 to − 529) is shown with reference to previous study30. (C) HEK293T cells were transfected with empty or FLAG-Hoxa10 plasmids as well as luciferase reporter plasmids with mouse Gdf5 gene promoter. Cell lysates were subjected to luciferase measurement (n = 4). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Gdf5 gene promoter fragments collected by ChIP using anti-FLAG antibody were analyzed by real-time qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Figure Legend Snippet: Figure 5. Effect of HOXA10 on the Gdf5 gene promoter. (A) The ATAC-Seq database (articular chondrocytes: SRX13791211, costal chondrocytes: SRX11156876) from ChIP-Atlas was analyzed by integrative genomics viewer IGV2.8.6. A Gdf5 gene promoter (1393 bp) exhibits an open chromatin region on the Gdf5 gene in articular chondrocytes. (B) Schematic diagram of luciferase reporter construction with mouse Gdf5 gene promoter (− 1081 to + 312). A putative HOXA10 binding motif (− 538 to − 529) is shown with reference to previous study30. (C) HEK293T cells were transfected with empty or FLAG-Hoxa10 plasmids as well as luciferase reporter plasmids with mouse Gdf5 gene promoter. Cell lysates were subjected to luciferase measurement (n = 4). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Gdf5 gene promoter fragments collected by ChIP using anti-FLAG antibody were analyzed by real-time qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Techniques Used: Luciferase, Binding Assay, Transfection, Isolation, Infection, Control

Figure 4. Role of HOXA10 in Gdf5 expression in LB cells. (A) LB cells were isolated from Gdf5-HiBiT KI mice and plated on a 96-well plate. The cells were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. One day after infection, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (B) LB cells isolated from WT mice were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, **: P < 0.01).

Figure Legend Snippet: Figure 4. Role of HOXA10 in Gdf5 expression in LB cells. (A) LB cells were isolated from Gdf5-HiBiT KI mice and plated on a 96-well plate. The cells were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. One day after infection, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (B) LB cells isolated from WT mice were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, **: P < 0.01).

Techniques Used: Expressing, Isolation, Infection, Control, Cell Culture, Quantitative RT-PCR

Figure 6. Immunofluorescent analysis of HOXA10 and GDF5 in articular cartilage. Tibial sections from 3-month-old mice were subjected to co-immunostaining with anti-HOXA10 and anti-GDF5 antibodies. DAPI indicates nucleus. Scale bar, 50 μm.

Figure Legend Snippet: Figure 6. Immunofluorescent analysis of HOXA10 and GDF5 in articular cartilage. Tibial sections from 3-month-old mice were subjected to co-immunostaining with anti-HOXA10 and anti-GDF5 antibodies. DAPI indicates nucleus. Scale bar, 50 μm.

Techniques Used: Immunostaining

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Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 1. Identification of transcription factors predominantly expressed in articular cartilage cells. (A) Total RNA was isolated from SFZ, CC, LB, and C3H10T1/2 (10T1/2) cells. Gdf5 and Prg4 expression was analyzed by RT-qPCR in triplicate. (B) Total RNA from SFZ cells and CC was analyzed by microarray. Genes with expression exceeding 100 in terms of the SFZ raw values were counted as genes expressed in SFZ cells. The number indicates the number of genes expressed more than two-fold in SFZ cells compared with the level in CC. The panel on the right shows 11 transcription factors among the genes with expression in SFZ cells more than two-fold that in CC. (C) Total RNA was isolated from SFZ, CC, LB, and 10T1/2 cells. Indicated gene expression was analyzed by RT-qPCR in triplicate. Data are the mean ± SEM.

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Isolation, Expressing, Quantitative RT-PCR, Microarray, Gene Expression

Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 2. Generation of Gdf5-HiBiT screening system. (A) Schematic diagram of Gdf5-HiBiT allele. The stop codon of Gdf5 and the knocked-in position of HiBiT tag are indicated. (B) Gdf5-HiBiT KI mice were generated using the Technique for Animal Knockout system by Electroporation (TAKE) method based on CRISPR/Cas9. Genomic DNA sequence analysis of the Gdf5 gene was performed, which confirmed that the genome of Gdf5- HiBiT KI allele had been edited correctly. Two possible off-target sites for the CRISPR gRNA (target sequence: TCGTGGAATCTTGTGGCTGC) are shown. Genomic DNA sequence analysis of two off-target sites was performed, which confirmed that the sequences around off-target sites were intact. (C) Genomic PCR analysis of WT and Gdf5-HiBiT KI mice was performed. A representative result is shown. The original gel is presented in Supplementary Fig. 1. (D) SFZ cells were isolated from WT and Gdf5-HiBiT KI mice. The cells were cultured for 2 days and then the supernatants were collected. DMEM, 10% FBS DMEM, and supernatants of WT and Gdf5-HiBiT KI SFZ cells were subjected to HiBiT measurement (n = 3). RLU: relative light unit. Data are the mean ± SEM (****: P < 0.0001).

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Generated, Knock-Out, Electroporation, CRISPR, Sequencing, Isolation, Cell Culture

Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 3. Role of HOXA10 in Gdf5 expression in articular cartilage cells. (A) SFZ cells isolated from WT mice were infected with empty (control) or indicated lentiviruses. Indicated gene expression was analyzed by RT-qPCR in duplicate. (B) Schematic diagram of Gdf5-HiBiT screening system. (C) SFZ cells were isolated from Gdf5-HiBiT KI mice and were plated on a 96-well plate. Gdf5-HiBiT KI SFZ cells were infected with the indicated lentiviruses. One day later, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). (E) SFZ cells isolated from WT mice were infected with empty (control), shHoxa10-1, or shHoxa10-2 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Expressing, Isolation, Infection, Control, Gene Expression, Quantitative RT-PCR, Cell Culture

Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 5. Effect of HOXA10 on the Gdf5 gene promoter. (A) The ATAC-Seq database (articular chondrocytes: SRX13791211, costal chondrocytes: SRX11156876) from ChIP-Atlas was analyzed by integrative genomics viewer IGV2.8.6. A Gdf5 gene promoter (1393 bp) exhibits an open chromatin region on the Gdf5 gene in articular chondrocytes. (B) Schematic diagram of luciferase reporter construction with mouse Gdf5 gene promoter (− 1081 to + 312). A putative HOXA10 binding motif (− 538 to − 529) is shown with reference to previous study30. (C) HEK293T cells were transfected with empty or FLAG-Hoxa10 plasmids as well as luciferase reporter plasmids with mouse Gdf5 gene promoter. Cell lysates were subjected to luciferase measurement (n = 4). RLU: relative light unit. (D) SFZ cells isolated from WT mice were infected with empty (control) or FLAG-Hoxa10 lentiviruses. Gdf5 gene promoter fragments collected by ChIP using anti-FLAG antibody were analyzed by real-time qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, ****: P < 0.0001).

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Luciferase, Binding Assay, Transfection, Isolation, Infection, Control

Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 4. Role of HOXA10 in Gdf5 expression in LB cells. (A) LB cells were isolated from Gdf5-HiBiT KI mice and plated on a 96-well plate. The cells were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. One day after infection, the medium was changed. The cells were cultured for 2 days and then the supernatants were collected. The supernatants were subjected to HiBiT measurement (n = 3). RLU: relative light unit. (B) LB cells isolated from WT mice were infected with empty (control), Venus, or FLAG-Hoxa10 lentiviruses. Hoxa10, Gdf5, and Prg4 expression was analyzed by RT-qPCR (n = 3). Data are the mean ± SEM (*: P < 0.05, **: P < 0.01).

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Expressing, Isolation, Infection, Control, Cell Culture, Quantitative RT-PCR

Journal: Scientific reports

Article Title: HOXA10 promotes Gdf5 expression in articular chondrocytes.

doi: 10.1038/s41598-023-50318-7

Figure Lengend Snippet: Figure 6. Immunofluorescent analysis of HOXA10 and GDF5 in articular cartilage. Tibial sections from 3-month-old mice were subjected to co-immunostaining with anti-HOXA10 and anti-GDF5 antibodies. DAPI indicates nucleus. Scale bar, 50 μm.

Article Snippet: Slides were incubated overnight with primary antibodies against HOXA10 (GTX37412, GeneTex, Irvine, CA, USA; 1:100) and GDF5 (AF853, R&D Systems; 1:100).

Techniques: Immunostaining

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