jg98  (MedChemExpress)

Journal: Toxicological sciences : an official journal of the Society of Toxicology

Article Title: Benzene metabolites increase vascular permeability by activating heat shock proteins and Rho GTPases.

doi: 10.1093/toxsci/kfaf055

Figure Lengend Snippet: Fig. 3. Muconaldehyde-induced stimulation of HSPs in endothelial cells. (A) MCMVEC were incubated with 10 μM MA for 6 h and gene transcription was measured by RNA sequencing. (i) Volcano plot of differentially expressed genes. (ii) Reactome pathway enrichment analysis of MA-treated MCMVEC. (B) HAECs were incubated with MA (10 µM) for 2, 6, or 24 h, and gene transcription was measured by RNA sequencing. (i) Venn diagram of differentially expressed genes at indicated time points (padj < 0.05; log2FoldChange ≥j2j). (ii) Pathway enrichment analysis of MA-treated HAEC. (iii) HSP70 and HSP90 induction in MA-treated HAEC. (iv) Gene network of HSP-related pathways activated after 6 h exposure to MA. (C) MA (10 µM)-induced expression of Hspa1b gene (i) and upregulation of HSPA1B protein in MCMVEC (ii). (D) Immunofluorescent staining of Hsp70 and in MCMVEC treated with MA (10 µM) for 24 h. (E) Colocalization of Hsp70 with F-actin in MA-treated MCMVEC.

Article Snippet: HSP70 inhibitor JG98, HSP90 inhibitor 17-AAG, and ROCK1 (p160ROCK) inhibitor Y-27632 were purchased from the Selleckchem.com.

Techniques: Incubation, RNA Sequencing, Expressing, Staining

Journal: Toxicological sciences : an official journal of the Society of Toxicology

Article Title: Benzene metabolites increase vascular permeability by activating heat shock proteins and Rho GTPases.

doi: 10.1093/toxsci/kfaf055

Figure Lengend Snippet: Fig. 4. Contribution of HSPa in MA-induced changes in endothelial permeability. The endothelial monolayer was exposed to a medium containing 0.1% DMSO in PBS or 10 µM MA for the indicated times. Fluorescent dextran concentrations were measured in the lower chambers as described in the Materials and Methods section. Cells were co-exposed to the HSP90 inhibitor 17-AAG (A, 2 μM) or the HSP70 inhibitor JG98 (C, 2 μM). Data are shown as mean±SE. P < 0.05, P < 0.01, P < 0.0001 for MA versus Control; # #P < 0.01, # # # #P < 0.0001 for inhibitor/activator/MA versus MA, 2-way ANOVA with Tukey posttest. The AUC was calculated for the effects of 17-AAG (B) and JG98 (D) on MA-induced permeability. Data are shown as mean±SE. P < 0.05, P < 0.01, P < 0.001, P < 0.0001, 1-way ANOVA with Bonferroni posttest. (E) Characterization of HSPA1B-TGEC mice. (i) Schematic of transgene construct used for HSPA1B-TGEC mice generation. Mice that were homozygous for Hspa1b were crossed with mice expressing Cre recombinase under the control of the endothelial-specific Cdh5 promoter. (ii) Genotyping of transgenic mice. (iii) Detection of HSPA1b levels in isolated lung endothelial cells (EC) using western blot. (iv) The Hspa1bþ/þ/Creþ/− (HSPA1B-TGEC) and the corresponding littermate controls (Hspa1bþ/þ/Cre−/−) mice were injected (i.v.) with Evans blue and then treated with intradermal injections of MA (400 pmol per injection) or the vehicle. Group data on vascular leakage in skin extracts 30 min after MA treatment. Data are mean±SEM. P < 0.05, P < 0.001,P < 0.0001 following 2-way ANOVA with Tukey posttest. (v) MA-induced fold change in the vascular leakage detected in HSPA1B-TGEC versus littermate controls. Data are mean±SEM. P < 0.05 as per 2-tailed t-test (unpaired).

Article Snippet: HSP70 inhibitor JG98, HSP90 inhibitor 17-AAG, and ROCK1 (p160ROCK) inhibitor Y-27632 were purchased from the Selleckchem.com.

Techniques: Permeability, Control, Construct, Expressing, Transgenic Assay, Isolation, Western Blot, Injection