Fig 1: Anti-inflammatory activities of seven Wet Tropics plant extracts at 100 μg/mL. Selected extracts inhibited the production of proinflammatory cytokines in the supernatant of LPS-stimulated PBMCs: (A) tumour necrosis factor (TNF); (B) interleukin-23 (IL-23). The soluble proinflammatory cytokines were determined in the PBMCs supernatant of HBD (n = 4). PBMCs (1 × 106 cells per well at a final volume of 1 mL) were cultured for 20 h at 37 °C and 5% CO2: PBMCs were stimulated with 10 ng/mL LPS. Dexamethasone was used as a positive control. Data provided in mean ± SD (one-way ANOVA was performed in GraphPad prism to test significance, where **** p < 0.0001, *** p < 0.0002, ** p < 0.0021, and * p = 0.0332). LW—Leptospermum wooroonooran; AO—Alyxia orophila; LA—Linospadix apetiolatus; CH—Ceratopetalum hylandii; GB—Garcinia brassii; LG—Litsea granitica; PW—Polyscias willmottii; Dex—dexamethasone (a positive drug control); NS-Media—non stimulated media; S-Media—stimulated media.
Fig 2: mDia2 promotes protein secretion from HDFs. A, Quantification of total proteins secreted by sh-EV and sh-mDia2 HDFs. n = 3. B, Representative Western blots of CM of sh-EV and -mDia2 fibroblasts for fibronectin 1 (FN1), collagen (COL)4A1, and INHBA under reducing conditions. Ponceau S staining of the membrane served as loading control. C and D, Representative immunofluorescence images of sh-EV and sh-mDia2 fibroblasts stained for INHBA, FN1, and ELN (C), and quantification of intracellular fluorescence intensity (D). n = 3. E, Western blot for FN1, COL4A1, elastin (ELN), and vinculin (VIN, loading control) using total lysates of sh-EV or sh-mDia2 fibroblasts. n = 3. F, Representative FN1, COL1A1, and ELN immunofluorescence images and quantification of the stained area in the decellularized matrix from sh-EV and sh-mDia2 fibroblasts cultured in glucose-containing medium. n = 3. G, IFNα2, IL1β, and IL6 concentrations (ng/mL) in supernatants from sh-EV and sh-mDia2 fibroblasts. n = 3. H, Quantification of the SCC13 tumor spheroid area in hanging drop including CM from sh-mDia2 and sh-EV fibroblasts and representative images (left). The area of the spheroid formed in one control sample was set to 1. n = 10–11. I, Western blots of total lysate from sh-EV or sh-mDia2 fibroblasts transfected with expression vectors encoding wild-type (WT) mDia2, an actin-polymerization–deficient mDia2 mutant (IA) or empty vector (pMX) for mDia2 and GAPDH. J and K, Percentage of total cells with actin stress fibers (J) and scratch closure (K) of sh-EV and sh-mDia2 fibroblasts transfected with expression vectors encoding WT or IA mDia2 or pMX. n = 3 (J) and 6 (K). L and M, sh-EV or -mDia2 fibroblasts were transfected with expression vectors encoding WT or IA mDia2 or pMX. Quantification of FN1 immunofluorescence staining of the decellularized ECM (L), and representative Western blot of total cell lysate and data quantification (M). n = 3. Graphs show mean ± SEM. ns, P > 0.05; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 [one-way ANOVA with Bonferroni post hoc test (D, E, J, and K–M), or unpaired Student t test (A and F–H)]. Scale bars, 50 μm (C), 100 μm (F).
Fig 3: Mice demonstrate transient increases in multiple pro-inflammatory cytokines at the 24 h mark after all three models; however, this increase nearly always abates at the 48 h time point in L-CLP mice. (A) IL-23; (B) IL-1α; (C) IFN-γ; (D) TNF-α; (E) MCP-1; (F) IL12p70; (G) IL-1β; (H) IL-10; (I) IL-6; (J) IL-27; (K) IL-17A; (L) GM-CSF. (A–L) Summary graphs show mean ± SEM all after Hem/CLP, S-CLP, and L-CLP treatments [Sham Hem/CLP n = 11, Hem/CLP n = 11, Sham S-CLP n = 10, S-CLP n = 10, Sham L-CLP n = 7, 24 h L-CLP n = 6, 48 h L-CLP n = 6]; significance *p < 0.05; **p < 0.01.
Fig 4: PVOs derived from placenta of preeclampsia patients resemble the pathological features. A) Scheme depicting the culture of PE/Control placenta villi organoids. B) Representative bright‐field and H&E staining images of PE placenta villi, PE‐PVOs at D7 and D14. Scale Bar, 50 µm. C) Representative confocal microscopy images of EpCAM, Ki67, Vimentin, CD31, CK7, CD45, NKG7, and CD14 in PE placenta and PE‐PVOs cultured for 7 days. Experiments were performed in six biological replicates. Scale Bar, 50 µm. D) Representative images of H&E and immunochemistry staining for CK7 in Control‐derived and PE‐derived PVOs (cultured for 7 days). Scale Bar, 50 µm. E) Relative expression of GCM1, ERVW‐1 and ERVFRD‐1 in Ctrl‐PVOs and PE‐PVOs. ** p < 0.01, * p < 0.05. F) The ratio of sFLT/PLGF levels in the supernatant of Ctrl‐PVOs and PE‐PVOs at day 7 of culture. *** p < 0.001. G) Relative levels of IL‐1β, IFN‐γ, TNF‐α, IL‐10, and IL‐18 in the culture medium supernatant of PE‐PVOs compared to control‐PVOs using flow multifactor detection assay. * p < 0.05. Experiments were performed in 4 biological replicates.
Fig 5: Inflammatory cytokines changes in critical illness, including notable decreases in TNF-α and increases in IL-6 when compared to healthy controls. (A) IL-1β. (B) IFN-α2. (C) IFN-γ. (D) TNF-α. (E) MCP-1. (F) IL-6. (G) IL-8. (H) IL-10. (I) IL-12p70. (J) IL-17A. (K) IL-18. (L) IL-23. (A–L) Summary graphs show mean ± SEM all in critically ill patients as compared to healthy human controls [Control n = 14, Patients n = 23]; significance *p < 0.05; **p < 0.01.
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