Journal: International Journal of Molecular Sciences

Article Title: Lysophosphatidic Acid Is a Proinflammatory Stimulus of Renal Tubular Epithelial Cells

doi: 10.3390/ijms23137452

Figure Lengend Snippet: Pharmacologic dissection of LPA-induced cellular signaling pathways. HKC-8 cells were pretreated for 1 h with 666-15 (CREB1 inhibitor) 10 μM in ( A ), JSH23 (NFκΒ inhibitor) 100 μM in ( B ), PD98059 (MEK/ERK inhibitor) 50 μM in ( C ), or SP600125 (JNK inhibitor) 50 μM in ( D ) and then activated with LPA at a final concentration of 10 μΜ for 4 h. mRNA-expression levels of the indicated secreted factors were quantified with RT-qPCR. The Cq values of each gene were normalized against the Cq values of B2M . Statistical analysis was performed with unpaired t -test or Welch’s test in the case of normal data and with Mann–Whitney in the case of non-normal data. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. See also .

Article Snippet: LPAR1/3 inhibitor (Ki16425, Cat. no: HY-13285, MedChemExpress, Monmouth Junction, NJ, USA) was added at 10 μM; LPAR2 inhibitor (H2L5186303, Cat. no: 10-1452, Focus Biomolecules, Plymouth Meeting, PA, USA) was added at 10 μM; CREB inhibitor (666-15, Cat. no: A616443, Toronto Research Chemicals, North York, ON, Canada) was added at 10 μM; JNK inhibitor (SP600125, Cat. no: 420119, Calbiochem, San Diego, CA, USA) was added at 50 μM; NFκB inhibitor (JSH-23, Cat. no: HY-13982, MedChemExpress, Monmouth Junction, NJ, USA) was added at 100 μM; and MEK/ERK inhibitor (PD98059, Cat. no: 513000, Calbiochem, San Diego, CA, USA) was added at 50 μM.

Techniques: Dissection, Protein-Protein interactions, Concentration Assay, Expressing, Quantitative RT-PCR, MANN-WHITNEY

Journal: The Journal of Clinical Investigation

Article Title: Activin A activation of Smad3 mitigates innate inflammation in mouse models of psoriasis and sepsis

doi: 10.1172/JCI187063

Figure Lengend Snippet: ( A ) Heatmap representing significantly downregulated genes in macrophages from Smad3 -KO mice (compared with WT macrophages) stimulated with LPS for 24 hours. ( B and C ) MitoTracker staining in macrophages stimulated with LPS for 24 hours and isolated from Smad3 -KO mice ( B ) or Acvr1b -Lyz2cre mice ( C ). ( D and E ) ATP production (intracellular) in macrophages stimulated with LPS for 24 hours and isolated from Smad3 -KO mice ( D ) ( n = 6) or Acvr1b -Lyz2cre mice ( E ). ( F and G ) RT-qPCR analysis of Arg1 and Tgfbi expression in macrophages stimulated with LPS for 24 hours in combination (or not) with 20 μM of ATP and isolated from Smad3 -KO mice ( F ) or Acvr1b -Lyz2cre mice ( G ). ( H ) RT-qPCR analysis of Nt5e (encoding CD73) expression in macrophages stimulated with LPS for 24 hours and isolated from Smad3 -KO or Acvr1b -Lyz2cre mice. ( I and J ) RT-qPCR analysis of Tgfbi and Arg1 expression in macrophages stimulated with LPS for 24 hours in combination (or not) with ATP and a CD73 inhibitor or a CREB inhibitor and isolated from Smad3 -KO mice ( I ) or Acvr1b -Lyz2cre mice ( J ). ( F – J , n = 4.) Pooled or representative of at least 2 independent experiments. * P < 0.05, ** P < 0.01, *** P < 0.005, **** P < 0.001 by Student’s t test ( B – G ) or 1-way ANOVA ( H – J ).

Article Snippet: Macrophages were pretreated for 1 hour before these treatments with follistatin (0.5 μg/mL; BioLegend), cycloheximide (5 μM; Cayman Chemical), anti–TGF-β antibody (50 μg/mL; Bio X Cell), anti–activin A antibody (2 μg/mL; R&D Systems), Smad3 inhibitor (SIS3, 2 μM; Cayman Chemical), TAK1 inhibitor (5 nM; Sigma-Aldrich), MEK inhibitor (10 μM; Cayman Chemical), ERK inhibitor (1 μM; Cayman Chemical), STAT5 inhibitor (100 μM; Cayman Chemical), CD73 inhibitor (10 μM; Cayman Chemical), and CREB inhibitor (1 μM; Cayman Chemical).

Techniques: Staining, Isolation, Quantitative RT-PCR, Expressing