Journal: Frontiers in Immunology

Article Title: A flagellin-conjugate protein induces dual NLRC4- and NLRP3-inflammasome activation which modulates inflammatory cytokine secretion from macrophages

doi: 10.3389/fimmu.2023.1136669

Figure Lengend Snippet: Both NLRP3- and NLRC4-inflammasome activation contributes to rFlaA:Betv1 induced IL-1β and pro-inflammatory cytokine secretion from THP-1 macrophages. PMA-differentiated wild-type (WT), NLRP3-, NLRC4-, or ASC-deficient THP-1 macrophages were stimulated with either LPS as a positive control, or equimolar amounts of rFlaA + rBet v 1, rFlaA:Betv1, rFlaA *D1 :Betv1, or rFlaA ΔDC0 :Betv1 for 24 h (A) . Supernatants were collected and checked for the secretion of IL-1β, IL-6, IL-12, and TNF-α by ELISA (B) . Data are the mean results of three independent experiments ± SD, and statistical significances are indicated as ns: p-value > 0.05, *: p-value < 0.05, **: p-value < 0.01, ***: p-value < 0.001.

Article Snippet: PMA-differentiated, THP-1 macrophages were pre-incubated with the indicated amounts of either the IKK-β inhibitors TPCA-1 (Abcam, Cambridge, UK) or BMS-345541 (Abcam), the unspecific inflammasome inhibitor VX-765 ( Invivo Gen), which inhibits caspase-1 activity, the specific NLRP3-inflammasome inhibitor MCC950 ( Invivo Gen), or the MAPK inhibitors SP600125 (SAP/JNK MAPK inhibitor, Invivo Gen), SB202190 (p38α/β MAPK inhibitor, Invivo Gen), or U0126 (ERK MAPK inhibitor, Cell Signaling Technologies, Leiden, The Netherlands) for 90 min and subsequently stimulated with rFlaA:Betv1 for 24 h. The target molecules of the used inhibitors are summarized in .

Techniques: Activation Assay, Positive Control, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: A flagellin-conjugate protein induces dual NLRC4- and NLRP3-inflammasome activation which modulates inflammatory cytokine secretion from macrophages

doi: 10.3389/fimmu.2023.1136669

Figure Lengend Snippet: NFκB- and SAP/JNK MAP kinase-signaling pathways contribute to inflammasome activation by regulating pro-Caspase-1, pro-IL-1β-, and NLRP3-expression in rFlaA:Betv1-stimulated THP-1 macrophages. PMA-differentiated THP-1 macrophages were pre-treated with the indicated inhibitors (500 nM TPCA-1, 5 µM BMS-345541, 25 µM SP600125, or 10 µM SB202190) for 90 minutes, followed by stimulation with 27.4 µg/mL rFlaA:Betv1 for additional 24 h (A) . Proteins in supernatant (Sup) and cell lysate (Lysate) were examined by Western Blot (B) . The intensity of Western Blot bands from three independent experiments was analyzed, first normalized to the loading control β-Tubulin and then again normalized to the unstimulated group (shown as value of “1”) (C) . Data are either representative (B) or mean results of three independent experiments ± SD (C) . Statistical significances are indicated as ns: p-value > 0.05, *: p-value < 0.05, **: p-value < 0.01, ***: p-value < 0.001.

Article Snippet: PMA-differentiated, THP-1 macrophages were pre-incubated with the indicated amounts of either the IKK-β inhibitors TPCA-1 (Abcam, Cambridge, UK) or BMS-345541 (Abcam), the unspecific inflammasome inhibitor VX-765 ( Invivo Gen), which inhibits caspase-1 activity, the specific NLRP3-inflammasome inhibitor MCC950 ( Invivo Gen), or the MAPK inhibitors SP600125 (SAP/JNK MAPK inhibitor, Invivo Gen), SB202190 (p38α/β MAPK inhibitor, Invivo Gen), or U0126 (ERK MAPK inhibitor, Cell Signaling Technologies, Leiden, The Netherlands) for 90 min and subsequently stimulated with rFlaA:Betv1 for 24 h. The target molecules of the used inhibitors are summarized in .

Techniques: Activation Assay, Expressing, Western Blot

Journal: Biological Trace Element Research

Article Title: Effects of a Combination of Arginine Silicate Inositol Complex and a Novel Form of Biotin on Hair and Nail Growth in a Rodent Model

doi: 10.1007/s12011-022-03176-9

Figure Lengend Snippet: Effect of inositol-stabilized arginine silicate complex (ASI) and magnesium biotinate (MgB) on skin tissue representative protein expression of blots (acetyl-CoA carboxylase 1 (ACC1) A , acetyl-CoA carboxylase 2 (ACC2) B , pyruvate carboxylase (PC) C , propionyl-CoA carboxylase (PCC) D , and 3-methylcrotonyl-CoA carboxylase (MCC) E ) levels in rats. The densitometric analysis of the relative intensity according to the control group of the western blot bands was performed with β-actin normalization to ensure equal protein loading. Blots were repeated at least three times ( n = 3), and a representative blot ( F ) was shown. The bars point out the standard deviation of the mean. Different symbols (*, # , + ) indicate difference compared to the control, ASI, and ASI + MgB I groups, respectively) above the bars indicate statistical differences among the groups (ANOVA and Tukey’s post hoc test; * p < 0.05; ** p < 0.01, *** p < 0.001; **** p < 0.0001; ## p < 0.01; ### p < 0.001; #### p < 0.001 + p < 0.05, ++ p < 0.01). Control: shaved; ASI: shaved + ASI (4.14 mg/rat/day); ASI + MgB I: shaved + ASI (4.14 mg/rat/day) + MgB (48.7 μg/rat/day); ASI + MgB II: shaved + ASI (4.14 mg/rat/day) + MgB (325 μg/rat/day)

Article Snippet: The following primary antibodies were used: ACC1, ACC2, PC, PCC, MCC, FGF-2, KGF-2, VEGF, SIRT1, MMP-1, MMP-3, and β-actin (Santa Cruz Biotechnology, CA, USA).

Techniques: Expressing, Control, Western Blot, Standard Deviation