Journal: Molecules

Article Title: In Vitro Neuroprotective and Anti-Inflammatory Activities of Natural and Semi-Synthetic Spirosteroid Analogues

doi: 10.3390/molecules21080992

Figure Lengend Snippet: Anti-excitotoxic activity of compounds in glutamate-induced cell death. ( A ) PC12 cell cultures were exposed to an established excitotoxic concentration of glutamate (Glu) (20 mM), and some were also treated with each compound ( S9 and S15 semi-synthetic derivatives; E3, estriol; SL, solasodine; E1, estrone; T, testosterone; TDCA, taurodeoxycholic acid) or solvent control (DMSO). After 24 h of exposition to the damage and treatment, cellular viability was determined by the MTT assay; ( B ) Dose-response of Compound S15 in the same model of glutamate-induced excitotoxicity. Data are represented as the percent with respect to the untreated cells, representing the highest cell viability. * p < 0.05, ** p < 0.01 and *** p < 0.001, according to one-way ANOVA and Tukey’s post-hoc test.

Article Snippet: The amount of IL-1β released into the culture medium was measured using a sandwich ELISA system with antibodies specifics for rat IL-1β (Polyclonal Rat IL-1β/IL-1F2 Catalog Number AF-501-NA and Polyclonal Rat IL-1β/IL-1F2 Biotinylated Catalog Number BAF501, R&D Systems, Minneapolis, MN, USA), following the manufacturer’s instructions.

Techniques: Activity Assay, Concentration Assay, Solvent, Control, MTT Assay

Journal: Molecules

Article Title: In Vitro Neuroprotective and Anti-Inflammatory Activities of Natural and Semi-Synthetic Spirosteroid Analogues

doi: 10.3390/molecules21080992

Figure Lengend Snippet: Neuroprotective activity of compounds against cell damage induced by chemical hypoxia and glucose deprivation. PC12 cells were cultured in oxygen and glucose deprivation conditions (OGD), specifically in the absence of glucose and in the presence of 600 µM of potassium cyanide (KCN) for 1 h, and then, normal culture medium was reperfused to the cells. After 4 h of the reperfusion start, some PC12 cells were treated with solvent control (DMSO), solasodine (SL) or different concentrations of the S15 derivative. After 20 h of exposition to the reperfusion damage and treatment, cellular viability was determined by the MTT assay. Data are represented as the percent with respect to the untreated cells, representing the highest cell viability. Asterisks indicate statistical differences between the treatment and respective control condition (*** p < 0.001) according to one-way ANOVA and Tukey’s post-hoc test.

Article Snippet: The amount of IL-1β released into the culture medium was measured using a sandwich ELISA system with antibodies specifics for rat IL-1β (Polyclonal Rat IL-1β/IL-1F2 Catalog Number AF-501-NA and Polyclonal Rat IL-1β/IL-1F2 Biotinylated Catalog Number BAF501, R&D Systems, Minneapolis, MN, USA), following the manufacturer’s instructions.

Techniques: Activity Assay, Cell Culture, Solvent, Control, MTT Assay

Journal: Molecules

Article Title: In Vitro Neuroprotective and Anti-Inflammatory Activities of Natural and Semi-Synthetic Spirosteroid Analogues

doi: 10.3390/molecules21080992

Figure Lengend Snippet: In vitro anti-inflammatory activity of the S15 derivative in activated primary microglia. Microglia-enriched primary cultures were obtained from brains of adult male rats. Different concentrations of the S15 derivative, solvent control (DMSO) or 10 µM of dexamethasone (DX) as the positive control of inhibition were applied to some microglia cultures, 30 min before the activation of microglia cells with lipopolysaccharide (LPS, 1 µg/mL) and murine recombinant interferon gamma (IFNγ, 20 U/mL). Untreated (UT) microglia cells were included as the negative control of inhibition. After 24 h of exposition or not to the stimulus and treatment, the IL-1β level was quantified by ELISA ( A ); and the content of nitrite was determined by the Griess assay ( B ). Asterisks indicate statistical differences between the treatment and respective control condition (* p < 0.05, ** p < 0.01 and *** p < 0.001) according to one-way ANOVA and Tukey’s post-hoc test.

Article Snippet: The amount of IL-1β released into the culture medium was measured using a sandwich ELISA system with antibodies specifics for rat IL-1β (Polyclonal Rat IL-1β/IL-1F2 Catalog Number AF-501-NA and Polyclonal Rat IL-1β/IL-1F2 Biotinylated Catalog Number BAF501, R&D Systems, Minneapolis, MN, USA), following the manufacturer’s instructions.

Techniques: In Vitro, Activity Assay, Solvent, Control, Positive Control, Inhibition, Activation Assay, Recombinant, Negative Control, Enzyme-linked Immunosorbent Assay, Griess Assay

Journal:

Article Title: Cytokine response to vitamin E supplementation is dependent on pre-supplementation cytokine levels

doi:

Figure Lengend Snippet: Description of SNPs, primers and probes

Article Snippet: IL-1β was detected using mouse anti-human IL-1β MAb and biotinylated anti-human IL-1β Ab (R&D Systems, Minneapolis, MN).

Techniques:

Journal:

Article Title: Cytokine response to vitamin E supplementation is dependent on pre-supplementation cytokine levels

doi:

Figure Lengend Snippet: Genotype frequencies

Article Snippet: IL-1β was detected using mouse anti-human IL-1β MAb and biotinylated anti-human IL-1β Ab (R&D Systems, Minneapolis, MN).

Techniques:

Journal:

Article Title: Cytokine response to vitamin E supplementation is dependent on pre-supplementation cytokine levels

doi:

Figure Lengend Snippet: The effect of vitamin E supplementation on cytokine production depends on baseline cytokine production. Cytokine production was measured from whole blood at the beginning and end of a one year vitamin E supplementation in the elderly. TNF-α, IL-1β, and IL-6 was measured from whole blood elicited for 24 hours with lipopolysacchride (LPS; 1.0 µg/mL). IFN-γ was measured from whole blood elicited for 48 hours with phytohemagluttinin (PHA; 20 µg/mL) or concalavinA (ConA; 40 µg/mL). Cytokines production was corrected for the number of monocytes and lymphocytes in the blood (pg/ 106 lymphocyte and monocyte). P values (P) for the interaction are unadjusted.

Article Snippet: IL-1β was detected using mouse anti-human IL-1β MAb and biotinylated anti-human IL-1β Ab (R&D Systems, Minneapolis, MN).

Techniques:

Journal: Cardiovascular drugs and therapy

Article Title: Inhibition of platelet activation by clopidogrel prevents hypertension-induced cardiac inflammation and fibrosis.

doi: 10.1007/s10557-013-6471-z

Figure Lengend Snippet: Fig. 4 Clopidogrel treatment decreases inflammatory cytokine production in Ang II infused mice hearts. a Immunohistochemical analysis of cytokine expression was detected by anti-IL-1β and anti-TGF-β in Ang II or vehicle infused heart sections with clopidogrel or vehicle treatment (Bar=50 μm, n=8). b Histogram shows less IL-1β+ and TGF-β+ area in clopidogrel

Article Snippet: For immunostaining, deparaffinized heart sections (5 μm) were incubated with avidin, biotin (Maixin, Fuzhou, China) for 15 min. After the inactivation of endogenous peroxides, they were incubated with anti-mouse monoclonal antibodies against Mac2(1:400 dilution), interleukin-1β (IL-1β) (1:200 dilution), transforming growth factor-β (TGF-β) (1:300 dilution) and α-smooth muscle actin (α-SMA) (1:200 dilution, Santa Cruz Biotechnology, Santa Cruz, CA, USA) at 4 °C overnight.

Techniques: Immunohistochemical staining, Expressing

Journal: Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology

Article Title: P2X7 Receptor Antagonist A804598 Inhibits Inflammation in Brain and Liver in C57BL/6J Mice Exposed to Chronic Ethanol and High Fat Diet

doi: 10.1007/s11481-018-9816-3

Figure Lengend Snippet: Primer sequences for marker genes used in PCR reactions

Article Snippet: The following antibodies and dilutions were used: biotinylated anti-mouse IL-1β antibody (1:1000, overnight at 4 °C, R&D Systems, Biotechne, Minneapolis, MN), rabbit polyclonal p65 NF-kB1 (1:1000, overnight at 4 °C, Cell Signaling Technology, Danvers, MA), mouse monoclonal β-Actin (1:10000, 10–15 min room temperature, Sigma).

Techniques: Marker, Sequencing

Journal: Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology

Article Title: P2X7 Receptor Antagonist A804598 Inhibits Inflammation in Brain and Liver in C57BL/6J Mice Exposed to Chronic Ethanol and High Fat Diet

doi: 10.1007/s11481-018-9816-3

Figure Lengend Snippet: A804598 treatment reversed significant increases in the protein levels of pro-IL1β, the precursor of IL-1β, and p65 NF-kB1, a component of NF-kB1 complex. Upper panels - Western blots of representative bands for the proteins. Lower panels – densitometry analyses of the pro-IL-1β (a) and NF-kB1 (b) levels normalized to their corresponding actin levels. Data are presented as fold differences from the Control values, which were averaged and set as “1”. Data are reported as mean ± SEM. * P < 0.05, ** P < 0.01 compared to Control, # P < 0.05, # P < 0.01 compared to Hybrid, (n = 5/Control, n = 4 for Hybrid and Hybrid + A804598)

Article Snippet: The following antibodies and dilutions were used: biotinylated anti-mouse IL-1β antibody (1:1000, overnight at 4 °C, R&D Systems, Biotechne, Minneapolis, MN), rabbit polyclonal p65 NF-kB1 (1:1000, overnight at 4 °C, Cell Signaling Technology, Danvers, MA), mouse monoclonal β-Actin (1:10000, 10–15 min room temperature, Sigma).

Techniques: Western Blot, Control

Journal: Frontiers in Immunology

Article Title: The monocyte-derived cytokine response in whole blood from preterm newborns against sepsis-related bacteria is similar to term newborns and adults

doi: 10.3389/fimmu.2024.1353039

Figure Lengend Snippet: The production of TNF-α and IL-6 by term neonatal and adult WB shows a high degree of inter-individual variation. WB from term cord blood (term, N=11-17) and healthy adults (adult, N=11-18) was incubated with (A, B) LPS, Pam3Cys4K or R848 or with (C, D) E. coli , S. agalactiae (GBS), S. aureus or S. epidermidis (CNS) bacteria for 5h and the production of TNF-α and IL-6 was measured. (E) The inter-individual variation in production of TNF-α and IL-6 between individuals was determined for WB from term cord blood and healthy adults (N=11-18) for LPS (100 ng/mL), Pam3Cys4K (10 μg/mL) or R848 (10 μM) or bacteria (10 5 CFU/well). Cytokine concentrations are shown in pg/mL. CFU, colony-forming units.

Article Snippet: Biotin Mouse anti-Human TNF-α detection antibody (BD PharmingenTM), Biotin Mouse Anti-Human IL-6 detection antibody (eBioscienceTM) or Biotin mouse anti-Human IL-1β detection antibody (R&D Systems) was added as secondary antibody.

Techniques: Incubation, Bacteria

Journal: Frontiers in Immunology

Article Title: The monocyte-derived cytokine response in whole blood from preterm newborns against sepsis-related bacteria is similar to term newborns and adults

doi: 10.3389/fimmu.2024.1353039

Figure Lengend Snippet: Preterm neonatal WB produces lower levels of IL-6 and similar levels of TNF-α compared to term neonatal WB. Whole blood from preterm cord blood (preterm, N=8-10), term cord blood (term, N=17) and healthy adults (adult, N=17-21) was incubated with TLR ligands LPS (100 ng/mL), Pam3Cys4K (10 μg/mL) or R848 (10 μM) or with E. coli , S. agalactiae (GBS), S. aureus or S. epidermidis (CNS) bacteria (10 5 CFU/well) for 5h and the production of (A, B) TNF-α, (C, D) IL-6 and (E, F) IL-1β was measured. (G-I) The inter-individual variation in production of (G) TNF-α, (H) IL-6 and (I) IL-1β between individuals was determined for preterm cord blood (N=8-10). Cytokine concentrations are shown in log-transformed pg/mL. The dotted line indicates the limit of detection. CV = coefficiency of variation. * = P<0.05. ** = P<0.01. *** = P<0.001.

Article Snippet: Biotin Mouse anti-Human TNF-α detection antibody (BD PharmingenTM), Biotin Mouse Anti-Human IL-6 detection antibody (eBioscienceTM) or Biotin mouse anti-Human IL-1β detection antibody (R&D Systems) was added as secondary antibody.

Techniques: Incubation, Bacteria, Transformation Assay