homodimerization inhibitory peptide inhibitor  (Novus Biologicals)

Journal: Molecular oncology

Article Title: RING finger protein TOPORS modulates the expression of tumor suppressor SMAR1 in colorectal cancer via the TLR4-TRIF pathway.

doi: 10.1002/1878-0261.13126

Figure Lengend Snippet: Fig. 2. SMAR1 expression upon LPS treatment after inhibiting TLR4 signaling and its downstream pathways. Western blot analysis showing SMAR1 expression upon (A) inhibiting TLR4 pathway using TAK-242 (5 lM) in the absence and presence of LPS (1 lgmL1) treatment in HCT116 cell line. Only LPS treatment condition was used as positive control. Total and phospho-p65 were used as positive control (B) Real- time analysis was carried out to check the transcript levels of SMAR1 in HCT116 cell line upon inhibiting TLR4 pathway. Next, MyD88- dependent pathway was inhibited using peptide inhibitor against MyD88 (Pepinh-MYD: 10 lM). (C) SMAR1 expression was checked by western blotting, where pJNK/JNK was used as positive control to track the activation of MyD88 pathway. (D) Real-time quantification of SMAR1 transcript was also done upon inhibition of MyD88 pathway. TRIF-dependent pathway was inhibited using BX-795 (10 lM). (E) Expression of SMAR1 was observed using western blot and (F) real-time PCR upon inhibiting TRIF-dependent pathway. TLR3 agonist Poly(I: C) also uses TRIF-dependent pathway to relay the downstream signals. (G) Western blot showing the expression of SMAR1 upon Poly(I:C) treatment, where the expression of p-IRF3 was used as positive control. (H) Transcript levels of SMAR1 after Poly(I:C) treatment in HCT116 cell line. b-Actin was used as an endogenous loading control in all the western blots. The values below the blot represent the fold change relative to control, which was calculated after normalization with b-actin. Relative fold change for all the real-time quantitation was calculated using 18S rRNA. Data shown are representative of three independent experiments. Error bars indicate that all the values are mean SD, where *P < 0.05 **P < 0.01(one-way ANOVA, Bonferroni post-tests).

Article Snippet: MyD88 homodimerization inhibitor peptide set (Cat. no.: NBP2-29328) was purchased from Novus Biologicals, Centennial, CO, USA.

Techniques: Expressing, Western Blot, Positive Control, Activation Assay, Inhibition, Real-time Polymerase Chain Reaction, Control, Quantitation Assay

Journal: Theranostics

Article Title: Membrane TLR9 Positive Neutrophil Mediated MPLA Protects Against Fatal Bacterial Sepsis

doi: 10.7150/thno.37139

Figure Lengend Snippet: The effect of MPLA mediated inflammatory preconditioning (InP) on cav-1 -/- mice. (A) Survival curves, n=8 mice per group. The mice were first given 0.1×10 8 CFUs E. coli , followed by the lethal dose of E. coli (3×10 8 CFUs) 2 h later,* vs. Saline. (B) Experimental procedures. (1) The WT mice were injected i.p. with 0.1×10 8 CFUs E. coli, and 2 h later were injected with 3×10 8 CFUs E. coli per mouse, and all 8 mice survived (InP). (2) C av-1 -/ - mice were injected i.p. with 0.1×10 8 E. coli, and 2 h later were injected with 3×10 8 CFUs E. coli, and all 8 mice died. (C) Intestine and liver tissues of mice stained with H&E. (D) ELISA of TNF-α/IL-6 in the supernatants of peripheral blood neutrophils in mice stimulated with 0.1×10 8 CFUs E. coli for 2 h. ** P <0.01 (Student's t test). (E) Western blot shows that InP did not promote translocation of TLR9 from cytosol to cell membrane in cav -/- mice. (F) Western blot shows significant reduction of endogenous Cav-1 by shRNA against Cav-1 (Cav-1 shRNA). (G) Immunoblot of TLR9, MyD88, TRAF3 and IRF3 in HL60 cells transfected Cav-1 shRNA and then stimulated with 0.1×10 8 CFUs E. coli for indicated times. Similar results were obtained in three independent experiments. (H) ELISA of TNF-α/IL-6 in the supernatants of HL60 cells transfected Cav-1 shRNA, stimulated with 0.1×10 8 CFUs E. coli for 2 h. ** P <0.01 (Student's t test).

Article Snippet: MyD88 homodimerization inhibitor peptide (NBP2-29328) was purchased from Novus Biologicals (Novus, USA).

Techniques: Saline, Injection, Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Translocation Assay, Membrane, shRNA, Transfection

Journal: Theranostics

Article Title: Membrane TLR9 Positive Neutrophil Mediated MPLA Protects Against Fatal Bacterial Sepsis

doi: 10.7150/thno.37139

Figure Lengend Snippet: Expression of TLR9-Cav-1 signaling proteins in the neutrophils of patients with sepsis. (A) Membrane TLR9 expression. (B) Cav-1 expression. (C) ROC curve for mTLR9. P <0.05. (D) ROC curve for Cav-1. P <0.05. (E) Association of Cav-1 with surface TLR9 expression in neutrophils. r2 =0.5791. (F) MyD88 expression. (G) TRAF3 expression. (H) IRF3 expression.

Article Snippet: MyD88 homodimerization inhibitor peptide (NBP2-29328) was purchased from Novus Biologicals (Novus, USA).

Techniques: Expressing, Membrane