Journal: Frontiers in Immunology

Article Title: Inhibition of NKCC1 Modulates Alveolar Fluid Clearance and Inflammation in Ischemia-Reperfusion Lung Injury via TRAF6-Mediated Pathways

doi: 10.3389/fimmu.2018.02049

Figure Lengend Snippet: Expressions of MAPKs and α-ENaC in MLE-12 cells. (A) Total Erk MAPK (T-Erk) and phosphorylated Erk MAPK (p-Erk), (B) total JNK MAPK (T-JNK) and phosphorylated JNK MAPK (p-JNK), (C) total p38 MAPK (T-p38) and phosphorylated p38 MAPK (p-p38) after HR treated with BMT. (D) α-ENaC expression after HR treated by p38 MAPK inhibitor, BIRB-796 10 μM. BMT, bumetanide 20-μM; CTRL, control. Data are expressed as the mean ± SD ( n = 5 per group). * P < 0.05 compared with the control group; # P < 0.05 compared with the HR group.

Article Snippet: After 24 h of incubation, the cells were pretreated with vehicle, bumetanide (10, 20, 40-μM), or p38 MAPK inhibitor BIRB-796 (10-μM; Enzo Life Sciences, Inc., San Diego, CA, USA).

Techniques: Expressing

Journal: Frontiers in Immunology

Article Title: Inhibition of NKCC1 Modulates Alveolar Fluid Clearance and Inflammation in Ischemia-Reperfusion Lung Injury via TRAF6-Mediated Pathways

doi: 10.3389/fimmu.2018.02049

Figure Lengend Snippet: The mechanisms of NKCC1 modulating alveolar fluid clearance and inflammation in IR-ALI. IR stress causes phosphorylation of NKCC1 and activation of TRAF6, which result in cell swelling and inflammation of alveolar epithelium. Inhibition of NKCC1 by bumetanide reciprocally modulates epithelial TRAF6 expression. This interaction suppresses downstream p38 MAPK and NF-κB pathways, which attenuates the reduction of AFC via upregulating α-ENaC expression and reduces the alveolar inflammation.

Article Snippet: After 24 h of incubation, the cells were pretreated with vehicle, bumetanide (10, 20, 40-μM), or p38 MAPK inhibitor BIRB-796 (10-μM; Enzo Life Sciences, Inc., San Diego, CA, USA).

Techniques: Activation Assay, Inhibition, Expressing

Journal: Annals of the rheumatic diseases

Article Title: MicroRNA-199a* regulates the expression of cyclooxygenase-2 in human chondrocytes

doi: 10.1136/annrheumdis-2011-200519

Figure Lengend Snippet: miR-199a* directly inhibits the expression of cyclooxygenase-2 (COX-2) in osteoarthritis chondrocytes. (A) Luciferase activity in chondrocytes transfected with the reporter vector and premiR-199a* (*p<0.05). (B, C) Expression of COX-2 protein in premiR-199a* or antimiR-199a* transfected chondrocytes after 72 h of stimulation with interleukin-1β (IL-1β). (D, E) Expression of COX-2 protein in negative control premiRNA or negative control-antimiRNA-transfected chondrocytes after 72 h of stimulation with IL-1β. (F, G) Expression levels of miR-199a* and COX-2 in chondrocytes treated with the p38-MAPK inhibitor (SB202190) and stimulated with IL-1β were determined by TaqMan assays. Gene expression in unstimulated chondrocytes was used as a control and expression of RNU6B/glyceraldehyde 3-phosphate dehydrogenase was used as an endogenous control. COX-2 protein expression was determined by western immunoblotting. Each experiment was performed in duplicate with samples from the indicated number of patients. *p<0.05 vs control; #p<0.05 vs IL-1β-stimulated chondrocytes.

Article Snippet: OA chondrocytes were serum-starved overnight and then stimulated with IL-1β (5 ng/ml; R&D Systems, St Paul, Minnesota, USA) or p38-MAPK inhibitor (SB202190 100 μM; A G Scientific Inc, San Diego, California, USA) for the indicated time and total RNA was prepared using the TRIZOL reagent (Invitrogen, Carlsbad, California, USA).

Techniques: Expressing, Luciferase, Activity Assay, Transfection, Plasmid Preparation, Negative Control, Gene Expression, Control, Western Blot

Journal: Bioengineered

Article Title: Coordination of PRKCA/PRKCA-AS1 interplay facilitates DNA methyltransferase 1 recruitment on DNA methylation to affect protein kinase C alpha transcription in mitral valve of rheumatic heart disease

doi: 10.1080/21655979.2021.1971482

Figure Lengend Snippet: The role of p38/MAPK in regulating PRKCA-AS1 expression in RHD. (a) the RNA levels of PRKCA-AS1 and PRKCA in TNF-α-induced AC16 cells treated with inhibition of different signaling pathways. (b) Smad2 occupancy on PRKCA-AS1 in TNF-α-induced AC16 cells treated with inhibition of different signaling pathways. (c) Smad2 occupancy on PRKCA-AS1 in mitral valve of RHD. the given data from triplicate experiments was processed as mean ± standard error and compared by student’s t-test. ‘FC’: fold change; ‘BK’: blocking

Article Snippet: The conditions of 100 ng/mL tumor necrosis factor-α (TNF-α) (APExBIO, Houston, TX, USA) for 16 h [ ], 20 ng/mL 5-Azacytidine (APExBIO) for 24 h [ ], 1 μM p38/MAPK inhibitor (2-(4-chlorophenyl)-4-(4-fluorophenyl)-5-(pyridin-4-yl)-1H-pyrazol-3(2 H)-one) (APExBIO) for 4 h, 50 mM transforming growth factor β1 (TGFB1) inhibitor (4-[2-(6-methylpyridin-2-yl)-5,6-dihydro-4 H-yrrolo[1,2-b]pyrazol-3-yl]quinoline-6-carboxamide) (APExBIO) for 4 h, 0.1 μM Rho associated coiled-coil containing protein kinase 1/2 (ROCK1/2) (4-[(1 R)-1-aminoethyl]-N-pyridin-4-ylcyclohexane-1-carboxamide) (APExBIO) for 6 h, 10 μM peroxisome proliferator activated receptor gamma/delta (PPARγ/δ) (2,5-dichloro-N-(2-methyl-4-nitrophenyl) benzenesulfonamide) (APExBIO) for 4 h, and 100 μM signal transducer and activator of transcription 3 (STAT3) (5,15-diphenyl-21 H,23 H-porphine) (APExBIO) for 6 h were adopted.

Techniques: Expressing, Inhibition, Blocking Assay

Journal: International Journal of Molecular Sciences

Article Title: Recent Advances in Fragment-Based QSAR and Multi-Dimensional QSAR Methods

doi: 10.3390/ijms11103846

Figure Lengend Snippet: Summary of different QSAR methods and source information.

Article Snippet: 4D-QSAR , 4D , 20 DHFR inhibitors; 42 PGF 2 a analogs; 40 2-substituted dipyridodiazepione inhibitors 33 p38-MAPK inhibitors , PLS GL-PLS , r 2 = [0.90, 0.95]; r 2 = [0.73, 0.86]; r 2 = [0.67, 0.76] [ ] q 2 = [0.67, 0.85] [64] , [ ] http://www.seascapelearning.com/4DsgiSW/ [64] .

Techniques: Pesticides, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: The Soluble Fms-like Tyrosine Kinase-1 Contributes to Structural and Functional Changes in Endothelial Cells in Chronic Kidney Disease

doi: 10.3390/ijms232416059

Figure Lengend Snippet: Chronic kidney disease patient sera and excess sFlt-1 stiffen endothelial cell cortex. ( A ) Endothelial stiffness of EA.hy926 cells was measured by atomic force microscopy (AFM) after 24 h of incubation with controls’ and patients’ sera. The values obtained (20–50 values/patient) were grouped according to the stage of CKD (stages three–five), plotted as box and whiskers, and were used for comparison among all three stages and controls (* p < 0.0001). Mean values obtained from each patient (dots) were used for comparison between Total (all patients) and controls (** p = 0.0020). ( B ) The graphic represents the effect of sFlt-1 neutralization by incubation with a specific antibody followed by immunoprecipitation in protecting cells against patient serum-induced stiffness. Values represent 15–30 measurements/treatments based on two independent experiments. Fetal calf serum (FCS) was used as a reference value (control). ( C ) Aortae from mice exposed to continuous in vivo administration of recombinant sFlt-1 (300 ng/h; N = 3) or control protein (IgG-Fc, 300 ng/h; N = 3) for three days were isolated and analyzed ex vivo by AFM. All values obtained (20–29 values/mouse) were plotted as box and whiskers and used for comparison between groups. Mean values obtained from individual mice are represented as dots. ( D ) Dose-response curve of human recombinant sFlt-1 (0.5–2 µg/mL) incubated with EA.hy926 cells for 24 h. Control cells were treated with control protein (2 µg/mL). ( E ) Endothelial stiffness of primary HUVECs was measured by AFM 24 h after incubation with sFlt-1 or control protein (2 µg/mL). Coincubation with the SB203580 (10 µM), a specific p38 MAPK inhibitor, protected cells from stiffening upon sFlt-1 treatment. Results are expressed as mean ± SEM.

Article Snippet: Blockers and inhibitors were pre-incubated for 30 min before stimulation or as indicated in Figure legends: B203580 (10 μM, p38 MAPK-inhibitor; LC Laboratories, Woburn, MA, USA), VEGF (recombinant human VEGF 165 , 50 ng/mL; R&D Systems, Minneapolis, MN, USA).

Techniques: Microscopy, Incubation, Comparison, Neutralization, Immunoprecipitation, Control, In Vivo, Recombinant, Isolation, Ex Vivo

Journal: International Journal of Molecular Sciences

Article Title: The Soluble Fms-like Tyrosine Kinase-1 Contributes to Structural and Functional Changes in Endothelial Cells in Chronic Kidney Disease

doi: 10.3390/ijms232416059

Figure Lengend Snippet: Activation of p38 MAPK and ROS generation upon sFlt-1 treatment. ( A ) Representative Western blotting image of phosphorylated (phospho) and total p38 MAPK 10 min after sFlt-1 treatment. ( B ) Relative densitometric analysis based on two independent experiments. ( C ) ROS was detected by measuring the fluorescence of CellROX Deep Red, a cell-permeant ROS sensor, in cells treated with sFlt-1 for 10 and ~30 min in the presence or not of SB203580 (10 µM). Data are given as fold-change relative to control and expressed as mean ± SEM. MAPK, mitogen-activated protein kinase; ns, not significant; ROS, reactive oxygen species.

Article Snippet: Blockers and inhibitors were pre-incubated for 30 min before stimulation or as indicated in Figure legends: B203580 (10 μM, p38 MAPK-inhibitor; LC Laboratories, Woburn, MA, USA), VEGF (recombinant human VEGF 165 , 50 ng/mL; R&D Systems, Minneapolis, MN, USA).

Techniques: Activation Assay, Western Blot, Fluorescence, Control

Journal: International Journal of Molecular Sciences

Article Title: The Soluble Fms-like Tyrosine Kinase-1 Contributes to Structural and Functional Changes in Endothelial Cells in Chronic Kidney Disease

doi: 10.3390/ijms232416059

Figure Lengend Snippet: Excess sFlt-1 affects the actin cytoskeleton and disturbs the endothelial barrier function. Primary HUVECs were treated with sFlt-1 or control protein (2 µg/mL) for 24 h in the presence or not of SB203580 (10 µM). ( A ) F-actin was quantitatively assessed by measuring phalloidin fluorescence intensity using a microplate reader. ( B ) Changes in F-actin distribution in cells treated with sFlt-1 for 30 min and 24 h were assessed by immunofluorescence microscopy. Scare bar = 20 µm ( C ) Endothelial cell permeability, as a measure of barrier function, was determined by the passage of streptavidin-HRP through the endothelial monolayer seeded on transwell units (0.4 μm pore). Data are given as fold-change relative to control and expressed as mean ± SEM. HRP, horseradish peroxidase; SB203580, specific p38 MAPK inhibitor.

Article Snippet: Blockers and inhibitors were pre-incubated for 30 min before stimulation or as indicated in Figure legends: B203580 (10 μM, p38 MAPK-inhibitor; LC Laboratories, Woburn, MA, USA), VEGF (recombinant human VEGF 165 , 50 ng/mL; R&D Systems, Minneapolis, MN, USA).

Techniques: Control, Fluorescence, Immunofluorescence, Microscopy, Permeability