Journal: Biomedical Journal

Article Title: Lidocaine reinforces the anti-inflammatory action of dexamethasone on myeloid and epithelial cells activated by inflammatory cytokines or SARS-CoV-2 infection

doi: 10.1016/j.bj.2022.07.008

Figure Lengend Snippet: Lidocaine in combination with dexamethasone has an additive modulatory effect on the innate inflammatory pathway of inflammasome activation. THP1-ASC-GFP cells were primed with EcLPS for 3 h, stimulated ON with transfected pcDNA and modulated or not with lidocaine and/or dexamethasone. Cells were prepared for fluorescent microscopy observation (A, 20×), intense green specks and cells were counted and ASC + cells/total cells ratio calculated (B) and hIL-1β was measured in culture supernatants (C). ∗ ( p < 0.05), ∗∗ ( p < 0.01), ∗∗∗ ( p < 0.001), ∗∗∗∗ ( p < 0.0001), a result significantly different from the stimulated unmodulated condition. # ( p < 0.05), ## ( p < 0.01), ### ( p < 0.001), a result significantly different from its counterpart stimulated and modulated with only lidocaine at the same concentration. Data are represented as mean ± SEM of three replicates. Graphs are representative of at least two independent assays.

Article Snippet: Inflammasome activators: Ultra-pure EcLPS (InvitroGen®), transfected pcDNA (generated by PCR cloning of a cDNA fragment of a random not related gene) with LipofectamineTM LTX Reagent with PLUSTM Reagent (Invitrogen®) following the manufacturer instructions.

Techniques: Activation Assay, Transfection, Microscopy, Concentration Assay

Journal: Blood Advances

Article Title: Clinical characteristics, management, and hematopoietic cell transplantation of patients with TLR8 gain-of-function

doi: 10.1182/bloodadvances.2025016338

Figure Lengend Snippet: TLR8 variants associated with disease. (A) Schematic representation of TLR8 protein structure showing the localization of TLR8 variants associated with disease, including 2 published variants resulting in disease phenotypes other than INFLTR8 (p.G572V results in TLR8 protein loss, dysregulation of TLR7/8 signaling, and anemia and autoinflammation and p.V434L in inherited anemia ). (B) NF-κB reporter cells (HEK-Blue Null1 cells) that do not express endogenous TLR8 were transfected with WT TLR8, patient TLR8 variants (encoding p.P432L, p.P432Q, p.P432R), or a loss-of-function TLR8 variant (encoding p.D543A) and stimulated with the indicated doses of the TLR8-specific agonist TL8-506. The newly identified patient TLR8 variants lead to GOF in TLR8 activity as measured by NF-κB transcriptional activity. Data are represented as mean ± standard deviation (SD) of biological replicates and representative of 2 independent experiments, each performed in duplicate. ∗∗∗ P < .001, by 2-way analysis of variance (ANOVA) test. (C) VAF in the peripheral blood by age of disease onset for patients with mosaic variants in TLR8 . (D) Number of patients with the most commonly reported disease manifestations. LOF, loss-of-function; SEAP, secreted alkaline phosphatase.

Article Snippet: New variants identified in patients 7 and 8 (p.P432Q and p.P432R) or reported in a family with anemia (p.V434L) were tested using HEK-Blue Null1 NF-κB reporter cells (InvivoGen) lacking TLR8 expression as previously described.

Techniques: Transfection, Variant Assay, Activity Assay, Standard Deviation

Journal: Life

Article Title: Casomorphine-10 (CM-10) Peptide Orchestrates Circadian and Neurodevelopmental Gene Clusters via δ-Opioid Receptor Signaling: Insights from Transcriptome Analysis with δ-Opioid Receptor-Expressing HEK293 Cells

doi: 10.3390/life15101636

Figure Lengend Snippet: δ-opioid receptor (DOR)-expressing HEK293 cells and HEK293 cells stained with anti-DOR antibody and followed with FITC-conjugated anti-mouse IgG ( A ). Volcano plot of identified differentially expressed genes (DEGs) in DOR-HEK293 cells after 1 h treatment with CM-10 ( B ). Red boxes show changed gene expression over 1.4-time higher and less than 0.71 with p -value predicted by EdgeR of less than 0.05.

Article Snippet: The fixed DOR-HEK293 and HEK293 cells were then incubated with a rabbit anti-DOR antibody (GeneTex, Irvine, CA, USA) after dilution with 1% casein in PBS (1/1000) and stained with a secondary antibody (Cy3-conjugated anti-rabbit IgG, Rockland Inc., PA, USA).

Techniques: Expressing, Staining, Gene Expression

Journal: Life

Article Title: Casomorphine-10 (CM-10) Peptide Orchestrates Circadian and Neurodevelopmental Gene Clusters via δ-Opioid Receptor Signaling: Insights from Transcriptome Analysis with δ-Opioid Receptor-Expressing HEK293 Cells

doi: 10.3390/life15101636

Figure Lengend Snippet: Predicted DOR agonistic signaling networks in DOR-HEK293 cells after 1 h treatment with CM-10. Fourteen genes with suggested involvement in cAMP-dependent protein kinases, transcriptional regulators in response to cAMP, circadian rhythm, stress and depression are shown in and were applied for network analysis by STRING. Red: circadian rhythm, Green: regulation of transcription of Notch receptor target, Yellow: PKA activation in glucagon signalling.

Article Snippet: The fixed DOR-HEK293 and HEK293 cells were then incubated with a rabbit anti-DOR antibody (GeneTex, Irvine, CA, USA) after dilution with 1% casein in PBS (1/1000) and stained with a secondary antibody (Cy3-conjugated anti-rabbit IgG, Rockland Inc., PA, USA).

Techniques: Activation Assay