Journal: Biochemistry and Biophysics Reports

Article Title: Effects of 1α,25-dihydroxyvitamin D 3 and tacalcitol on cell signaling and anchorage-independent growth in T98G and U251 glioblastoma cells

doi: 10.1016/j.bbrep.2022.101313

Figure Lengend Snippet: ( A ) Western blot analysis of pY783-PLCγ, PLCγ, pT389-p70-S6 kinase, p70-S6 kinase, pY705-STAT3 and STAT3 after treating T98G cells with 1 μM of 1α,25-dihydroxyvitamin D 3 (1α,25), 1 μM of tacalcitol (Tac) or vehicle (EtOH) for 24 h. ( B ), ( C ) and ( D ) Quantitative analysis of the levels of pY783-PLCγ, pT389-p70-S6 kinase and pY705-STAT3 inT98G cell lysates in the treatment groups as compared to vehicle. β-Actin was used as loading control for pY783-PLCγ and pT389-p70-S6. pY705-STAT3 was normalized to total STAT3. The values are presented as mean ± standard deviation of at least four independent determinations and shown as fold difference compared to ethanol-treated cells. ( E ) Analysis of STAT3 mRNA in T98G cells treated with tacalcitol (1 μM, 24h) compared to vehicle-treated cells. The graph shows the means ± standard deviation of four independent experiments. ( F ) Determination of STAT3 activation in T98G cells using a STAT3 assay kit. STAT3 activation was measured in cells cultured for 24 h in the absence (EtOH) and presence of 1α,25-dihydroxyvitamin D 3 (1α,25) or tacalcitol (Tac) in the indicated concentrations. The values are presented as mean ± standard deviation of five independent determinations. * = p < 0.05.

Article Snippet: The following primary antibodies were used: rabbit anti-pY783-PLCγ1 (#2821, 1:1000, Cell Signaling Technology), mouse anti-PLCγ1 (H00005335-M01, 1:1000, Novus Biologicals), mouse anti-pThr389-p70-S6 Kinase (#7053, 1:1000, Cell Signaling Technology), rabbit anti-p70-S6 Kinase (#7053, 1:1000, Cell Signaling Technology), rabbit anti-pY705-STAT3 (#9145, 1:500, Cell Signaling Technology), mouse anti-STAT3 (#9139, 1:1000, Cell Signaling Technology), rabbit anti-pS473-AKT (#4060, 1:500, Cell Signaling Technologies), mouse anti-AKT (#2920, 1:500 Cell Signaling Technologies), rabbit anti-pT202/Y204-ERK1/2 (#9101, 1:1000, Cell Signaling Technology), rabbit anti-ERK1/2 (#4695, 1:1000, Cell Signaling Technology), mouse anti-β-Actin (sc-47778, 1:1000, Santa Cruz Biotechnology), mouse anti-pT180/182-p38 (#9216, 1:1000, Cell Signaling Technology), rabbit anti-p38 (#9212, 1:1000, Cell Signaling Technology) and rabbit anti-β-Actin (ab8227, 1:1000, Abcam).

Techniques: Western Blot, Control, Standard Deviation, Activation Assay, Cell Culture

Journal: Toxics

Article Title: Ferulic Acid Derivatives Ameliorate Intestine Barrier Destruction by Alleviating Inflammatory Responses in Dextran Sulfate Sodium-Induced Inflammatory Bowel Disease

doi: 10.3390/toxics12040268

Figure Lengend Snippet: Anti-inflammatory effect of FA derivatives C1 and C1a in macrophages: ( a ) the cytotoxicity of FA, C1, and C1a was determined on J774A.1 cells and mouse peritoneal macrophages; ( b ) LPS-stimulated nitric oxide production was determined using a nitric oxide detection kit on J774A.1 cells and mouse peritoneal macrophages; ( c ) the LPS-stimulated expression of pro-inflammatory cytokines and mediators including IL-1β, IL-6, MCP-1, iNOS, and COX-2 in J774A.1 cells was analyzed by qPCR; ( d ) the LPS-stimulated activation of intracellular mediators, including iNOS and COX-2, was confirmed by Western blot. ( e ) the LPS-stimulated phosphorylation of NF-κB was assessed by ELISA. Data are means ± SEM of three independent experiments. Note: # p < 0.05 is a significant difference compared with control group; * p < 0.05 is a significant difference compared with LPS-treated group.

Article Snippet: The phosphorylation of nuclear factor (NF)-κB in LPS-stimulated J774A.1 cells was determined with an ELISA kit (#7834S; Cell Signaling Technology) according to the manufacturer’s protocol.

Techniques: Expressing, Activation Assay, Western Blot, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Control