Journal: Frontiers in Pharmacology

Article Title: The Protective Effects of Ivabradine in Preventing Progression from Viral Myocarditis to Dilated Cardiomyopathy

doi: 10.3389/fphar.2016.00408

Figure Lengend Snippet: Expression of p38 MAPK in the myocardial tissues of mice on days 35 and 65. (A) Western blot analysis for phospho-p38 MAPK (P-p38) and total-p38 MAPK (T-p38). (B) Densitometric analysis of relative protein levels for P-p38/ T-p38. ∗∗ P < 0.001, versus CON-35; ∗∗∗ P < 0.001 versus untreated CVMC- 65; ∗ P < 0.001 versus CON-65.

Article Snippet: For the Western blot, proteins were separated on polyacrylamide gels and transferred to a PVDF membrane for detection with various antibodies, including primary monoclonal antibodies for tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), phospho-p38 MAPK (P-p38) and total-p38 MAPK (T-p38) (Cell Signaling Technology Corporation, USA) and a polyclonal antibody for collagen I, collagen III (Biorbyt Corporation, USA).

Techniques: Expressing, Western Blot

Journal: Scientific Reports

Article Title: DC-SIGN and Toll-like receptor 4 mediate oxidized low-density lipoprotein-induced inflammatory responses in macrophages

doi: 10.1038/s41598-017-03740-7

Figure Lengend Snippet: DC-SIGN participated in the TLR4-NFκB pathway. Negative control (NC) or DC-SIGN siRNA was transfected into macrophages treated or not treated with oxLDL (50 μg/ml) or LPS (62.5 ng/ml) for 60 min. Western blot analysis detected the knockdown efficiency of DC-SIGN and the phosphorylation of p38, JNK, IKKε and NFκB ( A and B ), which was quantified by densitometry in 3 independent experiments and presented as relative units (DC-SIGN/α-tubulin, p38, JNK, IKKε and NFκB phosphorylated protein/total protein). The data are expressed as the mean ± SD from 3 independent tests. * P < 0.05, ** P < 0.01 compared with the macrophages not treated with oxLDL or LPS, ## P < 0.01 compared with the NC in the same group. ( C ) Negative control (NC) or DC-SIGN siRNA was transfected into macrophages treated or not treated with oxLDL (50 μg/ml) or LPS (62.5 ng/ml) for 60 min. Nuclear extracts were then prepared and assayed for p65 activation by EMSA.

Article Snippet: Primary antibodies for DC-SIGN, TLR4, α-tubulin, anti-FLAG, anti-His (Abcam, USA), p65 (t-p65), phosphorylated-p65 (p-p65), IKKε (t- IKKε), phosphorylated-IKKε (p- IKKε), p38 MAPK (t-p38), phosphorylated-p38 MAPK (p-p38), c-Jun N-terminal kinase (t-JNK), and phosphorylated-JNK (p-JNK) were purchased from Cell Signaling Technology (MA, USA).

Techniques: Negative Control, Transfection, Western Blot, Knockdown, Phospho-proteomics, Activation Assay

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: β-HB treatment reverses sorafenib resistance by shifting glycolysis-lactate metabolism in HCC.

doi: 10.1016/j.biopha.2023.115293

Figure Lengend Snippet: Fig. 5. β-HB treatment reversed sorafenib resistance and enhanced regorafenib sensitivity by inhibiting the B-raf/MAPK pathway in Huh7-SR and Sk-Hep-1-SR cells. (A) Huh7-SR and SK-Hep-1-SR cells were treated with sorafenib (0 and 10 μM) and β-HB (0, 2.5, and 5 mM) for 48 h, and cell viability was then analyzed using an MTT assay. (B) Huh7-SR and SK-Hep-1-SR cells were treated with regorafenib (0 and 5 μM) and β-HB (0, 2.5, 5, and 10 mM) for 48 h, and cell viability was then analyzed using an MTT assay. (C) Western blotting was conducted to analyze the protein expression of phosphorylated (p)- and total (t)-MAPK-related signaling cascades; α-tubulin was used as an internal control. (D) Western blot images from Fig. 4 C were quantified using ImageJ software. (E) Representative contour plots of apoptosis were detected by flow cytometry stained with Annexin V-FITC and PI. (F) Apoptosis rates were assessed in dose-dependent regorafenib with or without β-HB treatment in Huh7-SR and Sk-Hep-1-SR with Annexin V/PI staining. * p < 0.05; * * p < 0.01; * ** p < 0.001 vs. 0 mM β-HB cells. Data are presented as mean ± SD.

Article Snippet: The following antibodies were used in the experiments: HMGCS2 (1:1000, #ab137043, Abcam, Cambridge, UK), hexokinase I (1:1000, #2024, Cell Signaling, Danvers, MA, USA), hexokinase II (1:1000, #2867, Cell Signaling), phosphofructokinase (1:1000, #8164, Cell Signaling), LDHA (1:1000, #3582, Cell Signaling), PDH (1:1000, #3205, Cell Signaling), IDH (1:1000, #8137, Cell Signaling), phosphorylated (p)- and total (t)-B-raf (1:1000, #2696 and #9433, Cell Signaling), p- and t-MAPK kinase (MEK; 1:1000, #2338 and #9122, Cell Signaling), ERK (1:1000, #9101 and #4695, Cell Signaling), β-catenin (1:1000, #8480, Cell Signaling), ZO-1 (1:1000, #8193, Cell Signaling), Vimentin (1:1000, #5741, Cell Signaling), N-cadherin (1:1000, #13116, Cell Signaling) and antiα-tubulin (1:5000 dilution, #T9026, Sigma-Aldrich).

Techniques: MTT Assay, Western Blot, Expressing, Control, Software, Flow Cytometry, Staining