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

Journal: bioRxiv

Article Title: Inducing oncolytic cell death in human cancer cells by the long non-coding RNA let-A

doi: 10.1101/2021.07.16.452707

Figure Lengend Snippet: ( A ) Cell viability of ph 505 cells treated with inhibitors against Toll signaling components TLR3, MyD88, and TBK1, as well as with two NF-κB inhibitors. Cells were first incubated with different inhibitors for one hour, then treated with purified RNAs from mCherry/medium or let-A/medium for three hours. ( B ) Cell viability of HEK393T cells treated with inhibitors against Toll signaling components. Cells were treated by the same inhibitors and processes as ( A ). ( C ) SEAP activity in HEK-Dual hTLR3 cells treated with purified RNA from let-A/medium, mCherry/medium, or poly(I:C), with or without Toll signaling inhibitors. ( D ) Cell viability of ph 505 cells with or without pre-treatment with LPS before let-A (blue) or mCherry (red) expression was induced. Note that let-A induction could kill the LPS pre-treated cells even faster. ( E ) Cell viability of HEK393T cells with or without pre-treated with poly(I:C), before treated with purified RNA from mCherry/medium or let-A/medium. let-A/medium purified RNA could kill the poly(I:C) pre-treated cells much faster.

Article Snippet: The following inhibitors were used: TLR3/dsRNA Complex Inhibitor (Sigma) 30 μM; MyD88 inhibitor T6167923 (Anawa) 50 μM; TBK1 inhibitor MRT67307 (Sigma) 2 μM; and the NF-kB inhibitors Rolipram (Abcam) 1 μM; pyrrolidine dithiocarbonate (PDTC, Sigma) 25 μM.

Techniques: Incubation, Purification, Activity Assay, Expressing

Journal: Molecules

Article Title: Aminoclay Nanoparticles Induce Anti-Inflammatory Dendritic Cells to Attenuate LPS-Elicited Pro-Inflammatory Immune Responses

doi: 10.3390/molecules27248743

Figure Lengend Snippet: ACN treatment down-regulates basal AKT/mTOR/HIF1α signaling in DCs in a MyD88-independent fashion in vitro. ( A – C ) Splenic CD11c + DCs were purified from B6 mice using a MACS system, and DCs were cultured with ACNs (125 and 500 μg/mL) for 16 h. ( A ) Intracellular expression of P-AKT, P-mTOR, and HIF1α in DCs was assessed via flow cytometry. ( B ) Splenic DCs were cultured with ACNs (125 and 500 μg/mL). The percentage of three subsets (pDCs (CD11b − B220 + ), mDCs (CD11b + B220 − ), and cDCs (CD11b − B220 − )) among splenic DCs was evaluated using flow cytometry. ( C ) Intracellular expression of P-AKT, P-mTOR, and HIF1α was determined in DC subpopulations (pDCs, mDCs, and cDCs) via flow cytometry. Two-way ANOVA (ACN × MyD88 inhibitor) showed an interaction between these two factors ( # p < 0.05, ## p < 0.01). ( D ) Splenic DCs were cultured for 16 h with either vehicle (Veh) or MyD88 inhibitor (T6167923; 500 μM) in the absence or presence of ACNs (125 and 500 μg/mL). Intracellular expression of P-mTOR in DCs was assessed via flow cytometry. Two-way ANOVA (subpopulation × ACN) showed an interaction between these two factors. The mean values ± SD ( n = 3; per group in the experiment; Student’s t -test; * p < 0.05, ** p < 0.01, *** p < 0.001) are shown. One representative experiment of three experiments is shown. ns, not significant.

Article Snippet: MyD88 inhibitor (T6167923) was purchased from Toronto Research Chemicals (Martin Ross Ave, Toronto, Canada).

Techniques: In Vitro, Purification, Cell Culture, Expressing, Flow Cytometry

Journal: International Journal of Molecular Sciences

Article Title: Targeting Myeloid Differentiation Primary Response Protein 88 (MyD88) and Galectin-3 to Develop Broad-Spectrum Host-Mediated Therapeutics against SARS-CoV-2

doi: 10.3390/ijms25158421

Figure Lengend Snippet: Schematic representation of MyD88 primary and tertiary structures. The 3D model of the MyD88 TIR domain and the BB loop region (in dark blue) was adapted from Refs. [ , ]. DD, death domain; ID, intermediary domain; TIR, Toll-interleukin-1 receptor.

Article Snippet: GlycoMantra owns a patent (pending) on the combination of MyD88 inhibitor and Gal3 inhibitor for the therapy of respiratory viral infections, including SARS-CoV-2.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: Targeting Myeloid Differentiation Primary Response Protein 88 (MyD88) and Galectin-3 to Develop Broad-Spectrum Host-Mediated Therapeutics against SARS-CoV-2

doi: 10.3390/ijms25158421

Figure Lengend Snippet: Schematic representation showing MyD88-mediated pro-inflammatory response and plausible mechanism of MyD88 inhibition in restoring host-mediated immune responses.

Article Snippet: GlycoMantra owns a patent (pending) on the combination of MyD88 inhibitor and Gal3 inhibitor for the therapy of respiratory viral infections, including SARS-CoV-2.

Techniques: Inhibition

Journal: International Journal of Molecular Sciences

Article Title: Targeting Myeloid Differentiation Primary Response Protein 88 (MyD88) and Galectin-3 to Develop Broad-Spectrum Host-Mediated Therapeutics against SARS-CoV-2

doi: 10.3390/ijms25158421

Figure Lengend Snippet: Chemical structures and compositions of MyD88 and Gal3 inhibitors. Structures of compound 1, EM163, and compound 4210 were adapted from Ref. . Structure of T6167923 was adapted from Ref. . Structures of TD139 and GB1211 were adapted from Ref. .

Article Snippet: GlycoMantra owns a patent (pending) on the combination of MyD88 inhibitor and Gal3 inhibitor for the therapy of respiratory viral infections, including SARS-CoV-2.

Techniques:

Journal: NPJ Regenerative Medicine

Article Title: Proteoglycan 4 (PRG4) treatment enhances wound closure and tissue regeneration

doi: 10.1038/s41536-022-00228-5

Figure Lengend Snippet: In primary MEFs, rhPRG4 induced VEGF protein expression in a dose dependent manner with a plateau at 100 µg/ml a , b . This effect was confirmed at the mRNA level c . Vegf and Hif1α mRNA expression was quantified under normoxic and hypoxic conditions in the presence or absence of rhPGR4 and/or Hif1α inhibition (BAY 87-2243) to demonstrate that rhPRG4 induction of Vegf was independent of Hif1α d , e . To determine if Vegf activation was downstream of Toll-like receptors (TLRs), HEK cells lacking all TLRs or expressing a single TLR (2, 4 or 5) were treated with rhPRG4 or their specific agonist f . TLR4 expressing cells demonstrated the maximal Vegf activation (f) and this activation was independent of MYD88 activity since the MYD88 inhibitor (ST2825) had no effect on rhPRG4 Vegf activation g . All experiments were undertaken on at least 3 biological and 3 technical replicates unless otherwise stated. n.s. = not significant. Error bars equal mean ± SD ( a , c , d – 1 way ANNOVA, f , g – t-test).

Article Snippet: TLR Null, TLR Null-2, TLR-2, −4, and −5 cell lines (Invivogen) were exposed to either positive controls for the TLRs (Heat-killed Listeria Monocytogenes:HKLM for TLR-2 (108 cells/mL), LPS for TLR-4 (100 ng/mL), and FLA for TLR-5 (100 ng/mL)) or rhPRG4 (100 µg/mL) in the presence of absence of the MyD88 inhibitor ST2835 (10 µM, Cedarlane).

Techniques: Expressing, Inhibition, Activation Assay, Activity Assay