Fig 1: Inhibitors for oxidative stress and NF-kB reduce inflammation in ECs exposed to oscillating glucose. Cells were exposed alternatively to 5 mM/25 mM glucose (OG) at every 3 h, for 72 h. N-acetyl-cysteine (NAC, 7.5 mM) or Bay11-7085 (Bay, 15 µM) were added to the OG experimental condition, for the whole period of incubation. (a,d,e) Protein expression of Ninj-1 (a), TNFR1 (d) and RAGE (e) in EC lysates relative to β-actin (representative blot and densitometric analysis); (b) monocyte adhesion to endothelial cells; (c) secreted MCP-1 in the culture media relative to total cell protein (representative blots and densitometric analysis). All data are expressed as fold change versus OG and presented as mean ± SD. & p < 0.05, && p < 0.01, &&& p < 0.001 vs. OG.
Fig 2: Oscillating glucose stimulates Ninj-1, MCP-1, and monocyte adhesion to ECs to a greater extent than HG. Cells were exposed to 5 mM glucose (NG), 25 mM glucose (HG) or alternatively to 5 mM/25 mM glucose (OG) at every 3 h, for 72 h. Cells exposed alternatively to L-glucose or mannitol (using the same protocol as for OG) was considered osmotic control (OC). (a,c) mRNA levels of Ninj-1 (a) and MCP-1 (c); (b) Ninj-1 protein expression in whole cell lysate relative to β-actin (representative blots and densitometric analysis); (d) secreted MCP-1 in the culture media relative to total cell protein (representative blots and densitometric analysis); (e) monocyte adhesion to endothelial cells. All data are expressed as fold change versus NG and presented as mean ± SD. ** p < 0.01, *** p < 0.001 vs. NG; # p < 0.05, ## p < 0.01, ### p < 0.001 vs. HG.
Fig 3: Schematic representation of the benefic effects induced by the transcriptional activation of the endogenous apoAI or PON1 in enterocytes. Transfection of Caco-2 cells with CRISPR/dCas9 activation plasmids for apoAI or PON1 increases their gene and protein expression, as well as their secretion in the culture medium. Overexpressed apoAI and PON1 exert a positive feedback regulation on their transcription factors PPARγ, LXRs, and SIRT1. In turn, these transcription factors upregulate the lipid transporters ABCA1, ABCG8, and SR-BI which are known to be anti-atherogenic. The apoAI and PON1 secreted by the transfected enterocytes in the culture medium are able to attenuate the TNFα-induced inflammatory and oxidative stress in EC, by decreasing TNFR1, MCP-1, and p22phox levels.
Fig 4: Immunohistochemical examination of FFPE tissue samples (lungs and heart) from one meningococcal septic shock patient and one control (acute non-infectious death). Immunohistochemical detection of CD68, MPO, MCP-1, and PAI-1 positive cells (labeled brown) in sections from lungs and heart from meningococcal septic shock patient and control patient with acute non-infectious death (Obj. x20) (#The control patient, elderly smoker).
Fig 5: ERβ-induced reduction in proinflammatory cytokine release via suppression of the NF-κB pathway. (A) Expression of NF-κB pathway-associated proteins in NC empty vector-transfected, ERβ expression vector-transfected and PHTPP-treated PC-3 and DU145 cells was determined via western blot analysis. (B) Production of inflammatory cytokines was determined using ELISA in NC, ERβ-transfected and PHTPP-treated PC-3 and DU145 cells. Data are presented as the mean ± standard deviation; *P<0.05, **P<0.01, ***P<0.001 vs. NC; #P<0.05, ##P<0.01, vs. transfection group. ERβ, estrogen receptor β; IL, interleukin; NF-κB, nuclear factor κ-light-chain-enhancer of activated B cells; IκBα, NF-κB inhibitor α; LPS, lipopolysaccharide; MCP-1, monocyte chemoattractant protein 1p, phosphorylated; TNF-α, tumor necrosis factor-α.
Supplier Page from Abcam for Anti-MCP1 antibody