Journal: Journal of Crohn's & Colitis

Article Title: Efficacy of JAK inhibitors in Ulcerative Colitis

doi: 10.1093/ecco-jcc/jjz202

Figure Lengend Snippet: JAK inhibitors in IBD.

Article Snippet: Itacitinib [INCB039110] , JAK1 , Phase 2 ongoing [NCT03627052] , No published data in UC yet , Incyte Corporation.

Techniques:

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Inhibition of STAT1 phosphorylation in monocytes by non-cytotoxic concentrations of jakinibs. Monocytes isolated by adherence to plastic were pre-treated with 12.5 μM AG126 ( A ), 25 μM AG490 ( B ), 200 nM bariticinib ( C ), and 25 μM itacitinib ( D ) for 1 h and then stimulated with IFN-γ for 10 min. STAT1 phosphorylation was assessed by flow cytometry using anti-STAT1 (pY701)-Alexa Fluor®647 mAb. p < 0.05; **p < 0.01; ***p < 0.001 correspond to differences when compared to IFN-γ-stimulated cells ( E ). Representative histograms of one experiment. The dot plot chart shows the median MFI ± IQR of STAT1 (pY701). n = 3 independent experiments.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Inhibition, Phospho-proteomics, Isolation, Flow Cytometry

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Inhibition of HLA-DR expression on monocytes by non-cytotoxic concentrations of jakinibs . Human PBMCs were pre-treated with 200 nM baricitinib, 12.5 μM AG126, 25 μM AG490 and 25 μM itacitinib for 1 h, and then stimulated with IFN-γ for 24 h. Cells were stained with PI, anti-HLA-DR-APC-CY7, anti-CD19-V450, and anti-CD14-RD1 mAbs . A. Gating strategy for CD14 + cells (monocytes), and representative histograms of HLA-DR expression in monocytes cultured with and without IFN-γ. B. The bar chart represents the mean % ± 95%CI of HLA-DR MFI on monocytes relative to HLA-DR MIF in DMSO controls. Overton subtractions were performed to evaluate differences between treatments C. The bar chart represents the mean % ± SEM of cell viability relative to DMSO controls after 24 h-incubation with or without IFN-γ evaluated by PI exclusion and flow cytometry. *p < 0.05; **p < 0.01; ***p < 0.001 differences when compared to control without IFN-γ. Two-way ANOVA and post hoc Bonferroni's tests n = 3 independent experiments.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Inhibition, Expressing, Staining, Cell Culture, Incubation, Flow Cytometry, Control

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Size of extracellular vesicles derived from Jurkat and U937 cells treated with jakinibs . Extracellular vesicles were isolated from Jurkat (A) and U937 (B) cells cultured in the presence of 200 nM baricitinib (BARI), 25 μM itacitinib (ITA), or 0.5 % DMSO (vehicle control) for 24 h. The absolute sizes of EVs were determined by dynamic light scattering. C . EVs were negatively stained and analyzed under TEM. Images captured by UltraScan ®1000XP camera. Particle diameters are shown; n = 1 independent experiment.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Derivative Assay, Isolation, Cell Culture, Control, Staining

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Effect of extracellular vesicles derived from lymphocytes and monocytes treated with jakinibs on platelet aggregation . Platelet rich plasma samples were exposed to A . J-EVs, B. L-EVs, C. M-EVs and D. U-EVs released in the presence of 0.5 % DMSO, 200 nM bariticinib, and 25 μM itacitinib (100 cells: 1250 EVs). After 8 m, percentages of light transmission were measured. Lines represent median ± IQR. *p < 0.05; **p < 0.01; ***p < 0.001; Kruskal Wallis and post-hoc Dunn's tests; n = 5 independent experiments. J: Jurkat, L: Lymphocytes, M: Monocytes, U: U937.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Derivative Assay, Clinical Proteomics, Transmission Assay

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Effect of extracellular vesicles derived from lymphocytes and monocytes treated with jakinibs on cytokine accumulation in PHA-stimulated PBMC cultures. 1 × 10 5 CFS-labeled PBMCs from healthy donors were stimulated with 4 % v/v PHA (0.1 mg/ml) in the presence of autologous L-EVs (n = 3) and M-EVs (n = 5) released under treatment with 0.5 % DMSO, 25 μM itacitinib, and 200 nM bariticinib (100 cells: 1250 EVs). Culture supernatants were collected at 6 h and levels of cytokine were quantified by Luminex®Multiplex Assay. A . Heat map representing the accumulation of cytokines in every experimental condition. B. Bar charts represent concentration (median ± IQR) of IL-8, C. MIP-1β, D. MCP-1, E. IP-10, F. IL-1Ra, G. IL-2, H. MIP-1α, and I. TNF-α. *p < 0.05; **p < 0.01; ***p < 0.001; two-way ANOVA and Bonferroni post-hoc tests. CN: negative control, CFSE: CFSE control without stimulus.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Derivative Assay, Labeling, Luminex, Multiplex Assay, Concentration Assay, Negative Control, Control

Journal: Heliyon

Article Title: Janus kinase inhibitors modify the fatty acid profile of extracellular vesicles and modulate the immune response

doi: 10.1016/j.heliyon.2024.e24710

Figure Lengend Snippet: Fatty acid profile of extracellular vesicles derived from lymphocytes and monocytes treated with jakinibs. Extracellular vesicles were isolated from sorted monocytes and lymphocytes cultured in the presence of 0.5 % DMSO, 200 nM bariticinib, 25 μM itacitinib. A. The heat map represents the fatty acid composition and the fatty acid saturation index of the EVs. Bar charts represent the median ± IQR percentages of myristate (B) , arachidonate (C) , arachidonate detected by ELISA (D) and the fatty acid saturation index of the EVs (E) . *p < 0.05; **p < 0.01; ***p < 0.001; Kruskal-Wallis and post-hoc Dunn's tests. L-EVs: n = 3 independent experiments. M-EVs: n = 5 independent experiments. L-EVs: lymhocyte-derived EVs M-EVs: monocyte-derived EVs.

Article Snippet: Jakinibs (itacitinib and baricitinib) were purchased from Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Derivative Assay, Isolation, Cell Culture, Enzyme-linked Immunosorbent Assay

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: Physicochemical properties of F127/LEC PNPs.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques:

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: SEM of ITA-F127/LEC PNPs. (A) Low magnification high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) image of PLGA-LEC-F127 NPs. (B) Electronic scanning micrography of PLGA-LEC/F127 NPs loaded with Itacitinib.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Transmission Assay, Electron Microscopy

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: Effect of ITA-F127/LEC PNPs on monocyte activation. Monocytes, cultured in 96-well U-bottom plates with RPMI supplemented with penicillin/streptomycin and iFBS at 37°C, were stimulated with 2 ng/ml IFN-γ for 24 h in the presence of ITA, empty FL127/LEC PNPs, or ITA-F127/LEC PNPs at a concentration equivalent to 1200 ng/ml of ITA. Cells were then stained with anti-CD69-PE-ECD and anti-CD86-FITC mAbs, washed, and analyzed by flow cytometry. (A) CD69 MFI and (B) CD86 MFI of IFN-γ-stimulated monocytes were compared with the respective MFI of IFN-γ-stimulated monocytes previously exposed to different treatments. * p < 0.05; one-tailed Wilcoxon test. n = 5 independent experiments. ITA: Itacitinib; PNP: Poly(lactic- co -glycolic acid) nanoparticle.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Activation Assay, Cell Culture, Concentration Assay, Staining, Flow Cytometry, One-tailed Test

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: Effect of ITA-F127/LEC PNPs on STAT1 phosphorylation of monocytes. Monocytes, cultured in 96-well U-bottom plates with RPMI supplemented with penicillin/streptomycin and iFBS at 37°C, were treated with ITA, empty F127/LEC PNPs, or ITA-loaded-F127/LEC PNPs for one hour before stimulation with IFN-γ for 15 min. Cells were then fixed with BD Phosflow™ Lyse/Fix Buffer, permeabilized with BD Phosflow™ Perm Buffer IV, stained with an anti-STAT1(pY701)-Alexa Fluor ® 647, and analyzed by flow cytometry. (A) Representative histograms showing the STAT1(pY701) MFI changes under different treatments. (B) STAT1(pY701) MFI of IFN-γ-stimulated monocytes was compared with the STAT1(pY701) MFI of IFN-γ-stimulated monocytes previously exposed to different treatments. * p < 0.05; one-tailed Wilcoxon test. n = 5 independent experiments. ITA: Itacitinib; PNP: Poly (lactic- co -glycolic acid) nanoparticle.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Phospho-proteomics, Cell Culture, Staining, Flow Cytometry, One-tailed Test

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: ITA-F127/LEC PNPs affect the monocyte's ability to promote T-cell proliferation. Monocytes, cultured in 96-well U-bottom plates, were treated for 1 h with 1200 ng/ml ITA, empty FL127/LEC PNPs, or ITA-F127/LEC PNPs at a concentration equivalent to 1200 ng/ml of ITA. After PBS washing for removing NPs, pretreated monocytes were co-cultured with CFSE-labeled autologous T cells and 2 μg/ml PHA in RPMI supplemented with penicillin/streptomycin and 5% iFBS at 37°C for 72 h. (A) Representative histograms showing the percentage of proliferating T cells under different treatments. (B) The percentage of proliferating T cells in PHA-stimulated co-cultures was compared with that of proliferating T cells under different treatments. * p < 0.05; one-tailed Wilcoxon test. n = 5 independent experiments. ITA: Itacitinib; PHA: Phytohemagglutinin; PNP: Poly (lactic-co-glycolic acid) nanoparticle.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Cell Culture, Concentration Assay, Labeling, One-tailed Test

Journal: Nanomedicine

Article Title: Pluronic F127/lecithin PLGA nanoparticles as carriers of monocyte-targeted jakinibs: a potential therapeutic platform

doi: 10.1080/17435889.2024.2415877

Figure Lengend Snippet: Free and encapsulated Itacitinib modulates cytokines production. Monocytes from healthy controls incubated for one-hour empty F127/LEC PNPs, free Itacitinib, or encapsulated Itacitinib NPs (concentration equivalent to 400 ng/ml ITA) and co-cultured with autologous T lymphocytes stimulated or not with PHA. After 72 h, culture supernatants were collected to measure levels of (A) IL-2, (B) IFN-γ and (C) IL-17 by a Luminex bead-based multiplex assay. Three independent experiments, Wilcoxon (one-tailed) * p < 0.05.

Article Snippet: FluorSaveTM reagent was provided by Calbiochem (San Diego, CA, USA), Optylise C lysis buffer by Beckman Coulter (Brea, CA, USA) and Itacitinib by Cayman Chemical (Ann Arbor, MI, USA).

Techniques: Incubation, Concentration Assay, Cell Culture, Luminex, Multiplex Assay, One-tailed Test