antibody against jarid1d kdm5d  (Cell Signaling Technology Inc)

Journal: Cells

Article Title: Human Sex Matters: Y-Linked Lysine Demethylase 5D Drives Accelerated Male Craniofacial Osteogenic Differentiation

doi: 10.3390/cells11050823

Figure Lengend Snippet: Hierarchical clustering of differentially expressed genes revealed a sexually dimorphic osteogenic regulator profile between male and female NCSCs undergoing differentiation. ( A,B ) Cluster of upregulated genes in male NCSCs comprising autosomal osteogenic regulator CYP7B1 as well as the Y-linked genes KDM5D and USP9Y. ( C,D ). Female NCSCs showed a cluster of upregulated genes including the pro-osteogenic regulators KDM6A and XIST as well as the negative regulator TXLNG.

Article Snippet: Blocking of membrane with 5% milk powder (Carl Roth GmbH, Karlsruhe, Germany) in 1× TBS with 0.05% Tween 20 (Sigma-Aldrich, Taufkirchen, Germany) was followed by incubation with the first antibody against JARID1D/KDM5D (1:500; Cell Signaling Technology Inc., Danvers, MA, USA) in 1× TBS with 5% milk powder and 0.05% Tween 20 while shaking at 4 °C overnight.

Techniques:

Journal: Cells

Article Title: Human Sex Matters: Y-Linked Lysine Demethylase 5D Drives Accelerated Male Craniofacial Osteogenic Differentiation

doi: 10.3390/cells11050823

Figure Lengend Snippet: ( A,B ) Ribosomal Protein S4 Y-Linked 1 (RPS4Y1) and KDM5D (depicted in cyan, arrows) were most significantly downregulated in female NCSCs after 14 days and 30 of differentiation compared their male counterparts. ( C ) Differentially expressed genes between male and female NCSCs belonged, among others, to the osteogenic-related KEGG pathways and GO terms (depicted in orange). * padj < 0.05 was considered significant.

Article Snippet: Blocking of membrane with 5% milk powder (Carl Roth GmbH, Karlsruhe, Germany) in 1× TBS with 0.05% Tween 20 (Sigma-Aldrich, Taufkirchen, Germany) was followed by incubation with the first antibody against JARID1D/KDM5D (1:500; Cell Signaling Technology Inc., Danvers, MA, USA) in 1× TBS with 5% milk powder and 0.05% Tween 20 while shaking at 4 °C overnight.

Techniques:

Journal: Cells

Article Title: Human Sex Matters: Y-Linked Lysine Demethylase 5D Drives Accelerated Male Craniofacial Osteogenic Differentiation

doi: 10.3390/cells11050823

Figure Lengend Snippet: Knockdown of KDM5 significantly impaired the osteogenic differentiation capacity of male NCSCs. ( A ) Schematic view of the siRNA design. ( B ) qPCR analysis validating the knockdown of KDM5D. ( C ) Validation of the knockdown of KDM5D at the protein level via Western Blot. ( C – E ) Knockdown of KDM5D strongly inhibited the capability of male NCSCs to deposit calcium after 30 days of culture on collagen type I fibers compared to untreated and GFP-transduced cells. ( D–F ) Quantification of Alizarin Red S confirmed the significant inhibition of osteogenic differentiation in male NCSCs after knockdown of KDM5D. Mann–Whitney test, * p < 0.05 was considered significant.

Article Snippet: Blocking of membrane with 5% milk powder (Carl Roth GmbH, Karlsruhe, Germany) in 1× TBS with 0.05% Tween 20 (Sigma-Aldrich, Taufkirchen, Germany) was followed by incubation with the first antibody against JARID1D/KDM5D (1:500; Cell Signaling Technology Inc., Danvers, MA, USA) in 1× TBS with 5% milk powder and 0.05% Tween 20 while shaking at 4 °C overnight.

Techniques: Western Blot, Inhibition, MANN-WHITNEY

Journal: Cells

Article Title: Human Sex Matters: Y-Linked Lysine Demethylase 5D Drives Accelerated Male Craniofacial Osteogenic Differentiation

doi: 10.3390/cells11050823

Figure Lengend Snippet: Pharmacological inhibition of KDM5D significantly reduced the capability of male NCSCs to undergo osteogenic differentiation. ( A,B ) Exposure of male NCSCs to increasing concentrations (1 µM–100 µM) of KDOAM-25 resulted in a significant reduction in calcium deposition after 30 days of differentiation compared to untreated control. ( C ) Quantification of Alizarin Red S confirmed the significant inhibition of male osteogenic differentiation upon inhibition of KDM5D by KDOAM-25-treatment. Mann–Whitney test, * p < 0.05 was considered significant. n.d.: not detectable.

Article Snippet: Blocking of membrane with 5% milk powder (Carl Roth GmbH, Karlsruhe, Germany) in 1× TBS with 0.05% Tween 20 (Sigma-Aldrich, Taufkirchen, Germany) was followed by incubation with the first antibody against JARID1D/KDM5D (1:500; Cell Signaling Technology Inc., Danvers, MA, USA) in 1× TBS with 5% milk powder and 0.05% Tween 20 while shaking at 4 °C overnight.

Techniques: Inhibition, MANN-WHITNEY