Journal: International journal of molecular sciences

Article Title: Focal Adhesion Kinase Alleviates Simulated Microgravity-Induced Inhibition of Osteoblast Differentiation by Activating Transcriptional Wnt/β-Catenin-BMP2-COL1 and Metabolic SIRT1-PGC-1α-CPT1A Pathways.

doi: 10.3390/ijms26041669

Figure Lengend Snippet: Figure 2. SMG attenuates OBD by inhibiting the Wnt/β-catenin signaling pathway. Western blot analysis of the steady-state levels of pFAK (Y397), COL1, β-catenin, ALP and BMP2 in MC3T3-E1 cells cultured under 1 g, SMG or SMG + CNF1 conditions. The histogram bars represent protein abundance normalized to the β-actin control and expressed relative to MC3T3-E1 cells cultured under the 1 g condition. Data from three independent experiments are presented as the mean ± SD. Significance analysis of the experimental data for each group was performed using Student’s t test (* p < 0.05; ** p < 0.01).

Article Snippet: Primary antibodies against SIRT1 (9475), β-catenin (8480), pFAK (pFAK-Y397) (3283), vinculin (13901) and COL1 (72026) were obtained from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Western Blot, Cell Culture, Quantitative Proteomics, Control

Journal: bioRxiv

Article Title: Peripheral glia and neurons jointly regulate activity-induced synaptic remodeling at the Drosophila neuromuscular junction

doi: 10.1101/2024.06.27.600908

Figure Lengend Snippet: (A) Representative images showing that Shv can be detected extracellularly when expressed using glial-specific driver. Extracellular Shv-HA (Shv extra )is monitored using an antibody against HA under detergent-free staining condition. (B) Expression of Shv using the glial-specific driver does not rescue pFAK levels during unstimulated and stimulated conditions, revealing that glial Shv does not activate integrin. (C) Knockdown of shv in glia upregulated pFAK, whereas upregulation of shv in glia blocked the activity-dependent increase normally seen in control. (D) Expression of shv in shv 1 mutants show activity-dependent release of Shv by neurons, but not by glia. Shv extra indicates extracellular Shv stained using detergent-free conditions. (E) Knockdown of endogenous Shv-eGFP in neurons or glia did not diminish extracellular presence of Shv-eGFP at the NMJ, suggesting homeostatic regulation of Shv protein level. (F) Staining of the larval brain confirms that the RNAi approach effectively reduces Shv-eGFP in the selective cell types. Yellow arrows highlight neurons (Elav positive), magenta arrows point to glial cells (Repo positive). Scale bar = 2 μm for all panels, except scale bar = 10 μm in (F). All values are normalized to unstimulated control and presented as mean ± S.E.M. One-way Anova followed by Tukey’s multiple comparison test was used to compare between unstimulated control and unstimulated NMJs across genotypes. Student’s t-test was used to compare between unstimulated and stimulated NMJs of the same genotype. # p ≤ 0.05; ** p ≤ 0.01; ### p ≤ 0.001 when comparing unstimulated samples to unstimulated control. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001 when comparing stimulated to unstimulated NMJs.

Article Snippet: Primary antibodies used were: Rabbit anti-pFAK, 1:250 (Invitrogen, 44-624G), Rabbit anit-HA,1:1000 (Sigma, H6908), Rabbit anti-GluRIIC, 1:1000 , Cy3/Alexa-647 conjugated anti-HRP, 1:100 for non-permeabilized staining or 1:250 for permeabilized staining (Jackson ImmunoResearch); rat anti-Elav, 1:500 (7E8A10, Developmental Studies Hybridoma Bank at the University of Iowa (DSHB)); mouse anti-Repo, 1:50 (8D12, DSHB); mouse anti-GFP, 1:100 (4C9, DSHB).

Techniques: Staining, Expressing, Knockdown, Activity Assay, Control, Comparison