domain inhibitor  (Novus Biologicals)

Journal: Stem Cells Translational Medicine

Article Title: Extracellular vesicles conjugated with c(RGDyk) peptide targeting integrin αVβ3 repair optic nerve injury through YAP/TAZ and Smad2/3 signaling

doi: 10.1093/stcltm/szag006

Figure Lengend Snippet: Validation of target genes obtained via single-cell RNA sequencing in vivo/vitro models. Levels of interested proteins in (A) the retina and (B) optic nerve of ONC were presented. During recovery process by SC_EVs, protein levels involved in signal pathway were confirmed in (C) retina of ONC models and (D) R28 cells (* P < .05, ** P < .05 vs the age-matched sham; # P < .05, ## P < .01 vs EVs). At 9 h prior to SC_EVs treatment, R28 cells were exposed to 200 µM of CoCl 2 and 1 µM of TM2 TEAD inhibitor. Then, by (D) immunoblot and (E) immunofluorescence were performed for expression of proteins. The results are expressed as the mean ± standard error of the mean (SEM; scale bar: 10 μm). Statistical significance was determined using a nonparametric statistical test, followed by the Mann–Whitney U test (* P < .05, ** P < .01 vs control; # P < .05, ## P < .01vs CoCl 2 ; ++ P < .01 vs CoCl 2 +SC_EVs).

Article Snippet: Aliquots of 2 × 10 5 R28 cells were seeded onto glass coverslips and treated with 200 μM CoCl 2 and 1 μM of the TEA/TEF-domain (TEAD) transcription factor inhibitor TM2 (Tocris Bioscience, Bristol, UK) for 9 h. The hypoxia-damaged R28 cells were then incubated with EVs at a concentration of 24 μg/mL.

Techniques: Biomarker Discovery, Single Cell, RNA Sequencing, In Vivo, Western Blot, Immunofluorescence, Expressing, MANN-WHITNEY, Control

Journal: Liver Research

Article Title: Aluminum adjuvant promotes liver inflammation and fibrosis in mice: A novel approach to establish a liver fibrosis animal model

doi: 10.1016/j.livres.2025.05.001

Figure Lengend Snippet: Imject Alum increased the level of pro-inflammatory factors and inflammatory cells in the liver. (A) qRT-PCR analysis of specific leukocyte markers in the liver of Imject Alum (Al(OH) 3 160 mg/kg) D54 and control (CTR) groups. CTR group, n = 8; Imject Alum (Al(OH) 3 160 mg/kg) D54 group, n = 10. ∗ P < 0.05, compared with the CTR group. ns, no significant. (B) Representative immunohistochemical staining with F4/80 for macrophages in the liver from different groups. (C) Representative immunohistochemical staining with Cd3e for T lymphocytes in the liver from different groups. (D) Representative immunofluorescence staining of hepatic Cd11b. (E) Quantitative analysis of hepatic F4/80 and Cd3e immunohistochemical staining. CTR group, n = 8; Imject Alum (Al(OH) 3 160 mg/kg) D54 group, n = 10. ∗ P < 0.05, compared with the CTR group. (F) Hepatic mRNA expression of cytokines, chemokines, and cell adhesion molecules (CAMs) significantly increased in Imject Alum (Al(OH) 3 160 mg/kg) D54 group. CTR group, n = 8; Imject Alum (Al(OH) 3 160 mg/kg) D54 group, n = 10. ∗ P < 0.05, compared with the CTR group. (G) Relative levels of pyroptosis protein markers in the CTR and Imject Alum (Al(OH) 3 160 mg/kg) D54 groups. (H) Primary mouse hepatocyte mRNA levels of pyroptosis markers were significantly increased after treating with Imject Alum. n = 3. ∗ P < 0.05, compared with PBS group. # P < 0.05, compared with Imject Alum (Al(OH) 3 0.5 mg/mL) group. (I) The NLRP3 inhibitor MCC950 can inhibit the activation of primary mouse hepatocyte pyroptosis induced by Imject Alum. n = 3. ∗ P < 0.05, compared with PBS group. # P < 0.05, compared with Imject Alum (Al(OH) 3 1 mg/mL) group. Abbreviations: Asc, apoptosis-associated speck-like protein containing a caspase recruitment domain; Ccl2, C–C motif ligand 2; Cxcl, CXC motif chemokine ligand; Gsdmd, gasdermin D; Icam1, intercellular adhesion molecule 1; Il, interleukin; MOD, mean optical density; Nlrp3, nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3; PBS, phosphate-buffered saline; qRT-PCR, quantitative real-time polymerase chain reaction; Tnf-α, tumor necrosis factor-alpha; Vcam1, vascular cell adhesion molecule 1.

Article Snippet: Primary mouse hepatocytes were isolated from 8 to 10-week-old wild-type C57BL/6J mice, following a previously described protocol, then plated into collagen-coated 6-well plates (Corning, New York, USA), cultured in William's E Medium (Gibco, Thermo scientific, Rockford, IL, USA), supplemented with 5% fetal bovine serum (FBS; Lonsera, Suzhou, China), and subsequently treated with 0 μg/mL, 500 μg/mL, or 1 mg/mL of Imject Alum (Thermo Scientific), with or without prior administration of the nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3) inhibitor MCC950 (1 μmol/L; MedChemExpress, Monmouth Junction, NJ, USA) for 1 h. Total RNA was extracted after 12 h of Imject Alum (Thermo Scientific) treatment and subsequently used for quantitative real-time polymerase chain reaction (qRT-PCR) analysis.

Techniques: Quantitative RT-PCR, Control, Immunohistochemical staining, Staining, Immunofluorescence, Expressing, Activation Assay, Binding Assay, Saline, Real-time Polymerase Chain Reaction