Journal: Journal of Cellular and Molecular Medicine

Article Title: Hsa‐miR ‐409‐3p regulates endothelial progenitor senescence via PP2A‐P38 and is a potential ageing marker in humans

doi: 10.1111/jcmm.17691

Figure Lengend Snippet: Prediction of binding site of hsa‐miR‐409‐3p for protein phosphatase 2 catalytic subunit alpha (PPP2CA) gene and examination of its product protein phosphatase 2A (PP2A). (A) Diagram showing the seed sequences of hsa‐miR‐409‐3p and its predicted binding site in the 3’‐UTR of PPP2CA gene by prediction methods including TargetScan and miRanda. (B) Luciferase reporter assays of human embryonic kidney cells 293 co‐transfected with vector, wild‐type 3’‐UTR of PPP2CA (PPP2CA‐3’UTR‐WT) or mutant 3’‐UTR of PPP2CA (PPP2CA‐3’UTR‐MUT), along with mimic negative control (NC) or miR‐409‐3p mimics showed reduced activity only in the group co‐transfected with PPP2CA‐3’UTR‐WT and miR‐409‐3p mimics. Concordantly, in senescent cells, PP2A expression was downregulated at both (C) transcript and (D and E) protein levels, with activation of (F) phosphorylated p38 (p‐p38) and (G) phosphorylated JNK (p‐JNK). n = 4 for each bar. Data are mean ± SEM and analysed using unpaired Student's t test for comparison between groups. *, p < 0.05, ***, p < 0.001 vs. corresponding control. p38, p38 mitogen‐activated protein kinase. JNK, c‐Jun N‐terminal kinase

Article Snippet: The antibodies included phosphorylated p38 mitogen‐activated protein kinase (p‐p38), total p38, phosphorylated c‐Jun N‐terminal kinase (p‐JNK), total JNK, phosphorylated extracellular signal‐regulated kinase (p‐ERK) and total ERK from Cell Signalling Technology.

Techniques: Binding Assay, Luciferase, Transfection, Plasmid Preparation, Mutagenesis, Negative Control, Activity Assay, Expressing, Activation Assay

Journal: BMC Immunology

Article Title: A role for the Tec family kinase ITK in regulating SEB induced Interleukin-2 production in vivo via c-jun phosphorylation

doi: 10.1186/1471-2172-6-19

Figure Lengend Snippet: SEB induces activation of the JNK pathway specifically in responding T cells in vivo . WT mice were injected with 50 μg SEB or PBS i.v. and sacrificed after 1 hr. Spleen and lymph nodes were harvested and analyzed for the presence of phosphorylated c-jun in the Vβ8 + and Vβ6 + T cell populations. a) Spleen: (i) phosphorylation of c-jun in Vβ6 + T cells from PBS injected mice. (ii) phosphorylation of c-jun in Vβ6 + T cells from SEB injected mice. (iii) phosphorylation of c-jun in Vβ8 + T cells from the same PBS injected mice in (i). (iv) phosphorylation c-jun in Vβ8 + T cells from the same SEB injected mice in (ii). b) Lymph Node: (i) phosphorylation of c-jun in Vβ6 + T cells from PBS injected mice. (ii) phosphorylation of c-jun in Vβ6 + T cells from SEB injected mice. (iii) phosphorylation of c-jun in Vβ8 + T cells from the same PBS injected mice in (i). (iv) phosphorylation c-jun in Vβ8 + T cells from the same SEB injected mice in (ii). Arrow points to the population of cells responding to SEB stimulation by activation of the JNK pathway by inducing c-jun phosphorylation.

Article Snippet: Cells were then stained for intracellular phosphorylated-c-jun with a monoclonal antibody against phosphorylated c-jun (IgG 1 , Cell Signaling, Beverly, MA), followed by biotinylated rabbit anti-mouse IgG 1 and streptavidin conjugated to FITC.

Techniques: Activation Assay, In Vivo, Injection

Journal: BMC Immunology

Article Title: A role for the Tec family kinase ITK in regulating SEB induced Interleukin-2 production in vivo via c-jun phosphorylation

doi: 10.1186/1471-2172-6-19

Figure Lengend Snippet: ITK is required for SEB mediated induction of c-jun phosphorylation in responding T cells in vivo . WT (top panels) or ITK null (bottom panels) mice were injected with 50 μg SEB or PBS i.v. and sacrificed after 1 hr. Splenocytes were then analyzed for the presence of phosphorylated c-jun in the Vβ8 + and Vβ6 + T cell populations. a) Vβ6 + T cells from WT mice; b) Vβ8 + T cells from WT mice; c) Vβ6 + T cells from ITK null mice; d) Vβ8 + T cells from ITK null mice.

Article Snippet: Cells were then stained for intracellular phosphorylated-c-jun with a monoclonal antibody against phosphorylated c-jun (IgG 1 , Cell Signaling, Beverly, MA), followed by biotinylated rabbit anti-mouse IgG 1 and streptavidin conjugated to FITC.

Techniques: In Vivo, Injection