Journal: American journal of physiology. Lung cellular and molecular physiology

Article Title: Paramyxovirus replication induces the hexosamine biosynthetic pathway and mesenchymal transition via the IRE1α-XBP1s arm of the unfolded protein response.

doi: 10.1152/ajplung.00127.2021

Figure Lengend Snippet: Figure 4. IRE1a-XBP1 pathway regulates HBP and EMT gene networks: IRE1a kinase inhibitor. hSAECs were infected with RSV (MOI = 1.0) for 24h in the absence (DMSO, solvent carrier) or presence of the IRE1a endoribonuclease inhibitor KIRA8 (KIRA) or the ATF6 inhibitor ceapin-A7 (A7) at 10mM. Shown is fold change of mRNA relative to mock infection determined by Q-RT-PCR. Bars represent means ± ranges plus all data points of three independent experiments. GFPT2 (A); IL6 (B); SNAI1, ZEB1, VIM, and FN1 (C); MMP9 (D); and effect on RSV transcription (E). Note the equivalent expression of RSV N transcript between treatments indicates that RSV replication was not significantly affected by either KIRA8 or ceapin-A7. F: effect on RSV infectivity. Shown are focus forming units (FFU) determined by colorimetric assay using polyclonal anti-RSV antibodies. P < 0.05, P < 0.01, post hoc Tukey’s pairwise comparison. EMT, epithelial mesenchymal transition; HBP, hexosamine biosynthetic pathway; hSAECs, human small airway epithelial cells; IRE1a-XBP1, inositol-requiring enzyme 1a-X-box binding protein 1; Q-RT-PCR, quantitative reverse transcription polymerase chain reaction; RSV, respira- tory syncytial virus; ZEB1, zinc finger E-box binding homeobox 1.

Article Snippet: After treatment, cells were fixed with 4% paraformaldehyde (10min), permeablized with 0.1% Triton X-100 (10min), AJP-Lung Cell Mol Physiol doi:10.1152/ajplung.00127.2021 www.ajplung.org L577 Downloaded from journals.physiology.org/journal/ajplung (2405:4803:DB54:6CA0:48BC:7FD6:D8FE:BCC7) on March 11, 2025. blocked with 5% goat serum in PBS (2h), and incubated with primary antibody in blocking buffer overnight at 4 C. Primary antibodies used were anti-ATF6 (Cat. No. 24169-1-AP at VWR, 1:50 dilution), SNAI1 (Cat. No. 3895S at Cell Signaling, 1:50 dilution), glutamine-fructose-6-phosphate transaminase 2 (GFPT2) (Abcam, Cat. No. ab190966 at Abcam, 1:100 dilution), VIM (Cell Signaling, Cat. No. 5741S, 1:100 dilution), IRE1a (Abcam Cat. No. ab37073, 1:50 dilution), Calnexin (Abcam Cat. No. ab22595, 1:1,000 dilution), and GOLG2a/GM130 (Abcam Cat. No. ab52649, 1:200 dilution).

Techniques: Infection, Solvent, Reverse Transcription Polymerase Chain Reaction, Expressing, Colorimetric Assay, Comparison, Binding Assay, Reverse Transcription, Polymerase Chain Reaction, Virus

Journal: American journal of physiology. Lung cellular and molecular physiology

Article Title: Paramyxovirus replication induces the hexosamine biosynthetic pathway and mesenchymal transition via the IRE1α-XBP1s arm of the unfolded protein response.

doi: 10.1152/ajplung.00127.2021

Figure Lengend Snippet: Figure 5. IRE1a-XBP1 pathway regulates HBP and EMT gene networks: shRNA silencing. Q-RT-PCR analysis of hSAECs stably expressing nontargeting shRNA (Luc), IRE1a-targeting shRNA (IRE1), or XBP1-targeting shRNA (XBP1). Cells were mock or RSV-infected (MOI= 1, 24 h). Shown is fold change of mRNA relative to mock infection determined by Q-RT-PCR. Bars represent means ± ranges plus all data points of three independent experiments. P < 0.01. A: XBP1-Total; B: IRE1a; C: XBP1s; D: GFPT2; E: SNAI1; F: IL6; G: FN1; H: VIM; I: MMP9; J: RSV N transcription; K: RSV infectivity by colorimetric mea- surement of FFUs. EMT, epithelial mesenchymal transition; HBP, hexosamine biosynthetic pathway; hSAECs, human small airway epithelial cells; IRE1a inositol-requiring enzyme 1a; MOI, multiplicity of infection; Q-RT-PCR, quantitative reverse transcription polymerase chain reaction; RSV, respiratory syn- cytial virus; SNAI1, Snail family transcriptional repressor 1; XBP1, X-box binding protein 1.

Article Snippet: After treatment, cells were fixed with 4% paraformaldehyde (10min), permeablized with 0.1% Triton X-100 (10min), AJP-Lung Cell Mol Physiol doi:10.1152/ajplung.00127.2021 www.ajplung.org L577 Downloaded from journals.physiology.org/journal/ajplung (2405:4803:DB54:6CA0:48BC:7FD6:D8FE:BCC7) on March 11, 2025. blocked with 5% goat serum in PBS (2h), and incubated with primary antibody in blocking buffer overnight at 4 C. Primary antibodies used were anti-ATF6 (Cat. No. 24169-1-AP at VWR, 1:50 dilution), SNAI1 (Cat. No. 3895S at Cell Signaling, 1:50 dilution), glutamine-fructose-6-phosphate transaminase 2 (GFPT2) (Abcam, Cat. No. ab190966 at Abcam, 1:100 dilution), VIM (Cell Signaling, Cat. No. 5741S, 1:100 dilution), IRE1a (Abcam Cat. No. ab37073, 1:50 dilution), Calnexin (Abcam Cat. No. ab22595, 1:1,000 dilution), and GOLG2a/GM130 (Abcam Cat. No. ab52649, 1:200 dilution).

Techniques: shRNA, Reverse Transcription Polymerase Chain Reaction, Stable Transfection, Expressing, Infection, Reverse Transcription, Polymerase Chain Reaction, Virus, Binding Assay

Journal: Frontiers in pharmacology

Article Title: Autophagic Inhibition of Caveolin-1 by Compound Phyllanthus urinaria L. Activates Ubiquitination and Proteasome Degradation of β-catenin to Suppress Metastasis of Hepatitis B-Associated Hepatocellular Carcinoma.

doi: 10.3389/fphar.2021.659325

Figure Lengend Snippet: FIGURE 2 | CP inhibited migration and invasion of HBV-associated HCC. (A) HBV-associated cells including HepG2-HBx and HepG2-URG11 following CP administration and sorafenib treatment exhibited a significant delay in wound healing compared with that of control group. (B) CP treatment remarkably decreased the number of HBV-associated cells that migrated across matrigel and transwell chamber. (C) Western blot analysis showed that CP dramatically decreased expression levels of Vimentin and N-cadherin and increased E-cadherin level in both HepG2-HBx and HepG2-URG11 cells in a time-concentration dependent manner. All values represented the means ± SD (n 3, *p < 0.05, **p < 0.01, ***p < 0.001).

Article Snippet: Primary antibodies used in our study contained Cav1(16447-1-AP, Proteintech, Chicago, United States), N-cadherin (22018-1-AP, Proteintech, Chicago, United States), E-cadherin (20874-1-AP, Proteintech, Chicago, United States), Vimentin (10366-1-AP, Proteintech, Chicago, United States), URG11 (BS2170R, Beijing, China), β-catenin (66379-1-Ig, Proteintech, Chicago, United States), P-Akt (AF0016, Affinity, United States), Akt (60302-2-Ig, Proteintech, Chicago, United States), P-GSK3β (AB11002, AbSci, Baltimore, United States), GSK3β (BF0695, Affinity, United States) and β-actin (No.3700S, Cell Signaling Technology, CST, Boston, MA, United States).

Techniques: Migration, Control, Western Blot, Expressing, Concentration Assay

Journal: Frontiers in pharmacology

Article Title: Autophagic Inhibition of Caveolin-1 by Compound Phyllanthus urinaria L. Activates Ubiquitination and Proteasome Degradation of β-catenin to Suppress Metastasis of Hepatitis B-Associated Hepatocellular Carcinoma.

doi: 10.3389/fphar.2021.659325

Figure Lengend Snippet: FIGURE 8 | CP inhibited proliferation and migration of HBV-associated HCC ex vivo and in vivo. (A–C) HepG2-null and HepG2-HBx cells were grafted to the flank of nude mice by injecting the cells subcutaneously to observe tumor development, and volumes of tumors were determined at 3-day intervals. (D) Mice model with lung metastasis was established by tail vein injection of cancer cells. Upper panel, metastatic nodules in lungs. Lower panel, H&E staining of lung tissues. (E) Western blot analysis of tumor tissue showed that CP treatment decreased the expression levels of β-catenin and Cav-1, and enhanced the EMT process. (F) CP at concentrations from 60 to 120 μg/ml significantly decreased the primary as well as metastatic lesions in the zebrafish body when compared with control group. (G) H&E staining showed that CP suppressed the tumor growth in zebrafish. (H) Immunohistochemical staining of tumor in zebrafish showed CP treatment dramatically downregulated the expression levels of β-catenin, Cav-1 and Vimentin, and upregulated the expression level of E-cadherin. All values represented the means ± SD (n 6, *p < 0.05, **p < 0.01, ***p < 0.001).

Article Snippet: Primary antibodies used in our study contained Cav1(16447-1-AP, Proteintech, Chicago, United States), N-cadherin (22018-1-AP, Proteintech, Chicago, United States), E-cadherin (20874-1-AP, Proteintech, Chicago, United States), Vimentin (10366-1-AP, Proteintech, Chicago, United States), URG11 (BS2170R, Beijing, China), β-catenin (66379-1-Ig, Proteintech, Chicago, United States), P-Akt (AF0016, Affinity, United States), Akt (60302-2-Ig, Proteintech, Chicago, United States), P-GSK3β (AB11002, AbSci, Baltimore, United States), GSK3β (BF0695, Affinity, United States) and β-actin (No.3700S, Cell Signaling Technology, CST, Boston, MA, United States).

Techniques: Migration, Ex Vivo, In Vivo, Injection, Staining, Western Blot, Expressing, Control, Immunohistochemical staining