Journal: Molecular Therapy Oncolytics

Article Title: Exosomal circSHKBP1 participates in non-small cell lung cancer progression through PKM2-mediated glycolysis

doi: 10.1016/j.omto.2022.01.012

Figure Lengend Snippet: Exosomal circSHKBP1 promotes M2 polarization of macrophages (A) Representative images by confocal microscopy of the internalization of PKH67-labeled A549 exosomes (red) by THP-1 macrophages. (B) Internalization of PKH67-labeled exosomes by macrophages was assessed by flow cytometry after coculture for 24 h. (C) The mRNA expression of the M2 polarization markers CD206, IL-10, IL-4, and Arg1 in macrophages cocultured with exosomes for 48 h. (D) The mRNA expression of the M1 polarization markers CD86, IL-12, TNF-α, and INF-γ in macrophages cocultured with exosomes for 48 h. (E and F) Flow cytometry was used to explore the surface expression of CD206 and CD86 in macrophages cocultured with exosomes for 48 h. (G) The levels of the respective inflammatory cytokines in cell culture supernatants of macrophages cocultured with exosomes assessed using a ProcartaPlex combinable panel. (H) miR-1294 levels in macrophages cocultured with exosomes for 48 h. (I) Protein levels of PKM2, HK2, and GLUT1 in macrophages cocultured with exosomes for 48 h. (J) The glucose consumption and lactate excretion were detected in macrophages cocultured with exosomes for 48 h. (K) THP-1 macrophage cells were added to the upper wells of a Boyden chamber for assessment of chemotaxis toward exosomes released by A549 cells added to the lower chamber. The cells were allowed to migrate for 24 h at 37°C before staining with crystal violet and quantification. Data represent mean ± SD of triplicate experiments. ∗∗p < 0.01 compared with control; ##p < 0.01 compared with EXO.

Article Snippet: ELISA was used to evaluate the IL-10, IL-12, IL-4, TNF-α, and INF-γ (Enzo Life Sciences, New York, NY, USA) contents in culture medium derived from macrophages.

Techniques: Confocal Microscopy, Labeling, Flow Cytometry, Expressing, Cell Culture, Chemotaxis Assay, Staining

Journal: Frontiers in Pharmacology

Article Title: Dose Dependent Antimicrobial Cellular Cytotoxicity—Implications for ex vivo Diagnostics

doi: 10.3389/fphar.2021.640012

Figure Lengend Snippet: Rifampicin DRESS patients. IFN-γ ELISpot graphical illustration of (A) Australian patient and (B) south African patient. Positivity is defined by ≥ 50 spot forming unit (SFU)/million cells after background (unstimulated control) removal (dotted line on the corner right figures). Positive controls were done using CD3 and the unstimulated wells represent media alone. The stimulations with the different drug concentrations (µg/ml) were done in triplicates. The numbers adjacent to the wells represent the number of spots measured with the ELISpot reader. ELISpot graphical illustration for Rifampicin 25, 250, and 2500 μg/ml. Percentage of cytotoxicity assessed by (C) LDH assay, (D) flow cytometry with 7-AAD viability staining in lymphocytes and (E) flow cytometry with 7-AAD viability staining in CD4 + and CD8 + T cells.

Article Snippet: The INF-γ ELISpot assay was performed using a negative (unstimulated, media alone) and a positive control (anti-human CD3 antibody, 1-D1K, Mabtech®).

Techniques: Enzyme-linked Immunospot, Control, Lactate Dehydrogenase Assay, Flow Cytometry, Staining