Journal: Journal of Innate Immunity

Article Title: Neutrophil Killing of Peptoanaerobacter stomatis Requires TAK1-Driven Degranulation

doi: 10.1159/000553300

Figure Lengend Snippet: Activation of kinases in P. stomatis (Ps)- challenged neutrophils: Human neutrophils (10 × 10 6 /mL) were either left untreated (basal) or challenged with Ps at an MOI 10 for 5, 15, 30, 60, and 90 min. Cell lysates were analyzed by immunoblotting using phospho-specific antibodies and subsequently reprobed for total protein or β-actin as loading controls. Representative immunoblots showing phosphorylation kinetics ( a , e ) and densitometric analysis of ERK1/2, p38, AKT, and Src and Syk ( b , c , d , f , g ) are shown. Densitometric analyses of phosphorylated kinases were normalized to their respective total protein levels, except for p-TAK1, which was normalized to β-actin. Data represents the mean ± SEM of three independent experiments, and statistical significance was determined by one-way ANOVA (* p < 0.05, ** p < 0.01, *** p < 0.001). MOI, multiplicity of infection.

Article Snippet: The following inhibitors were used: diphenyleneiodonium (DPI; 10 μ m ; NOX2 inhibitor; Sigma-Aldrich), 5Z-7-oxozeaenol (1 μ m ; TAK1 inhibitor), SB203580 (10 μ m ; p38 MAPK inhibitor; MedChemExpress), U0126 (10 μ m ; ERK inhibitor; Calbiochem), LY294002 (10 μ m ; PI3K inhibitor; Calbiochem), piceatannol (10 μ m ; Syk inhibitor; Cayman Chemical), Src-1 (10 μ m ; Src family kinase inhibitor; Cayman Chemical), and GSK484 (10 μ m ; PAD4 inhibitor; Cayman Chemical).

Techniques: Activation Assay, Western Blot, Phospho-proteomics, Infection

Journal: Journal of Innate Immunity

Article Title: Neutrophil Killing of Peptoanaerobacter stomatis Requires TAK1-Driven Degranulation

doi: 10.1159/000553300

Figure Lengend Snippet: TAK1 acts as a critical regulator for kinase activation during P. stomatis (Ps) challenge: Human neutrophils (10 × 10 6 /mL) were either left untreated (basal) or challenged with Ps for 30 min after the indicated inhibitor pre-treatment. Cell lysates were analyzed by immunoblotting using phospho-specific antibodies and subsequently reprobed for total protein or β-actin as loading controls. Representative immunoblots and densitometric analysis of ERK1/2 ( a ), p38 ( b ), AKT ( c ), Src ( d ), and Syk ( e ) are shown. Densitometric analyses of phosphorylated kinases were normalized to β-actin. Data represent the mean ± SEM of three independent experiments, and statistical significance was determined by one-way ANOVA (* p < 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: The following inhibitors were used: diphenyleneiodonium (DPI; 10 μ m ; NOX2 inhibitor; Sigma-Aldrich), 5Z-7-oxozeaenol (1 μ m ; TAK1 inhibitor), SB203580 (10 μ m ; p38 MAPK inhibitor; MedChemExpress), U0126 (10 μ m ; ERK inhibitor; Calbiochem), LY294002 (10 μ m ; PI3K inhibitor; Calbiochem), piceatannol (10 μ m ; Syk inhibitor; Cayman Chemical), Src-1 (10 μ m ; Src family kinase inhibitor; Cayman Chemical), and GSK484 (10 μ m ; PAD4 inhibitor; Cayman Chemical).

Techniques: Activation Assay, Western Blot

Journal: Journal of Innate Immunity

Article Title: Neutrophil Killing of Peptoanaerobacter stomatis Requires TAK1-Driven Degranulation

doi: 10.1159/000553300

Figure Lengend Snippet: P. stomatis evades TAK1-dependent ROS- and NET-mediated killing but not degranulation. For the killing assay, neutrophils were pretreated for 15 min with inhibitors followed by P. stomatis incubation for 0 and 3 h. Then, neutrophil cell pellets (for cell-associated) and supernatant (for extracellular) were used to determine and quantify the CFU, which was quantified for percent killing after 1 µ m of 5Z-7-oxozeaenol ( a ), 10 µ m of DPI ( b ), GSK484 ( c ), or 10 µ m SB203580 pre-treatment. d P. stomatis was exposed to conditioned media collected from either unstimulated (US) or latrunculin +fMLF (L+F) stimulated neutrophils for 3 h and CFU enumeration was performed. Percent killing was determined by quantifying P. stomatis exposed to stimulated conditioned media over unstimulated condition. Data represents the mean ± SEM of three independent experiments, and statistical significance was determined by unpaired t -test for each pair (* p < 0.05, ** p < 0.01). ns, non-significant.

Article Snippet: The following inhibitors were used: diphenyleneiodonium (DPI; 10 μ m ; NOX2 inhibitor; Sigma-Aldrich), 5Z-7-oxozeaenol (1 μ m ; TAK1 inhibitor), SB203580 (10 μ m ; p38 MAPK inhibitor; MedChemExpress), U0126 (10 μ m ; ERK inhibitor; Calbiochem), LY294002 (10 μ m ; PI3K inhibitor; Calbiochem), piceatannol (10 μ m ; Syk inhibitor; Cayman Chemical), Src-1 (10 μ m ; Src family kinase inhibitor; Cayman Chemical), and GSK484 (10 μ m ; PAD4 inhibitor; Cayman Chemical).

Techniques: Incubation

Journal: Poultry Science

Article Title: Pathogenic mechanism of Eimeria tenella Et MIC2 promotes Eimeria tenella invasion and inhibits host cell apoptosis through binding to the ITGAV receptor

doi: 10.1016/j.psj.2026.106922

Figure Lengend Snippet: The mRNA expression of ITGAV, FAK, PLC, PKC, p65, ERK, JNK, p38, PI3K, Akt, Bax, Bcl2 , and Caspase 3 in E.tenella host cells.

Article Snippet: Membranes were blocked with 5 % BSA at 37°C for 2 h and incubated overnight at 4°C with the following primary antibodies: ITGAV Rabbit Ab, PLC Rabbit Ab, p-PLC Rabbit Ab, p-p65 Rabbit Ab, and Bcl2 Rabbit Ab were purchased from Bioss (Beijing, China); FAK Rabbit Ab, p-FAK Rabbit Ab, ERK Rabbit Ab, p-ERK Rabbit Ab, JNK Rabbit Ab, p-JNK Rabbit Ab, p38 MAPK Rabbit Ab, p-p38 MAPK Rabbit Ab, PI3K Rabbit Ab, p-PI3K Rabbit Ab, AKT Rabbit Ab, p-AKT Rabbit Ab, PKC Rabbit Ab, p-PKC Rabbit Ab, Bax Rabbit Ab, and Caspase 3 Rabbit Ab were purchased from Abmart (Shanghai, China); p65 Rabbit Ab (Proteintech, Wuhan, China).

Techniques: Expressing

Journal: Poultry Science

Article Title: Pathogenic mechanism of Eimeria tenella Et MIC2 promotes Eimeria tenella invasion and inhibits host cell apoptosis through binding to the ITGAV receptor

doi: 10.1016/j.psj.2026.106922

Figure Lengend Snippet: The protein activity changes of ITGAV, FAK, PLC, PKC, p65, ERK, JNK, p38, PI3K, Akt, Bax, Bcl2, and Caspase 3 in E.tenella host cells.

Article Snippet: Membranes were blocked with 5 % BSA at 37°C for 2 h and incubated overnight at 4°C with the following primary antibodies: ITGAV Rabbit Ab, PLC Rabbit Ab, p-PLC Rabbit Ab, p-p65 Rabbit Ab, and Bcl2 Rabbit Ab were purchased from Bioss (Beijing, China); FAK Rabbit Ab, p-FAK Rabbit Ab, ERK Rabbit Ab, p-ERK Rabbit Ab, JNK Rabbit Ab, p-JNK Rabbit Ab, p38 MAPK Rabbit Ab, p-p38 MAPK Rabbit Ab, PI3K Rabbit Ab, p-PI3K Rabbit Ab, AKT Rabbit Ab, p-AKT Rabbit Ab, PKC Rabbit Ab, p-PKC Rabbit Ab, Bax Rabbit Ab, and Caspase 3 Rabbit Ab were purchased from Abmart (Shanghai, China); p65 Rabbit Ab (Proteintech, Wuhan, China).

Techniques: Activity Assay