Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: ELF5 regulate casein synthesis in GMECs. (A, B) The expression of αS1-casein, αS2-casein, β-casein, κ-casein, phosphorylated JAK2 and STAT5 after transfection with pcDNA3.1-ELF5 or pcDNA3.1-NC for 48 h. (C, D) The expression of αS1-casein, αS2-casein, β-casein, κ-casein, p-JAK2, and p-STAT5 after transfection with siRNA-ELF5 or siRNA-NC (100 nM) for 48 h. The relative protein abundance of αS1-casein, αS2-casein, β-casein, and κ-casein was normalized to β-tubulin. The relative protein abundance of p-JAK2 and p-STAT5 was normalized to total JAK2 and STAT5, respectively. Values are presented as mean±SEM. * p<0.05, ** p<0.01. siRNA, small interfering ribonucleic acid; NC, negative control; GMECs, goat mammary epithelial cells; SEM, standard error of the mean.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Expressing, Transfection, Quantitative Proteomics, Negative Control

Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: The protein abundance of caseins after JAK2 and STAT5 inhibition in GMECs. (A) The expression of αS1-casein, αS2-casein, β-casein, κ-casein, and phosphorylated JAK2 and STAT5 after JAK2 inhibitor Tyrphosting AG490 (30 μM) treatment for 48 h. (B) The expression of αS1-casein, αS2-casein, β-casein, κ-casein, and p-STAT5 after STAT5 inhibitor STAT5-IN-1 (50 μM) treatment for 48 h. The relative protein abundance of αS1-casein, αS2-casein, β-casein, and κ-casein was normalized to β-tubulin. The relative protein abundance of p-JAK2 and p-STAT5 was normalized to total JAK2 and STAT5, respectively. Values are presented as mean±SEM. ** p<0.01. GMECs, goat mammary epithelial cells; SEM, standard error of the mean.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Quantitative Proteomics, Inhibition, Expressing

Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: ELF5 mediates casein synthesis by STAT5 activity in GMECs. (A) Cells were treated with STAT5-IN-1 (50 μM) or DMSO, followed by pcDNA3.1-ELF5 or pcDNA3.1-NC transfection for 48 h. (B) Cells were treated with STAT5-IN-1 (50 μM) or DMSO, followed by siRNA-ELF5 or siRNA-NC (100 nM) transfection for 48 h. The protein abundances of αS1-casein, αS2-casein, β-casein, κ-casein, and p-STAT5 were detected. The relative protein abundance of αS1-casein, αS2-casein, β-casein, and κ-casein was normalized to β-tubulin. The relative protein abundance of p-STAT5 was normalized to total STAT5. Values are presented as mean±SEM. a–d Different lowercase letters represent significant differences (* p<0.05). siRNA, small interfering ribonucleic acid; NC, negative control; GMECs, goat mammary epithelial cells; SEM, standard error of the mean.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Activity Assay, Transfection, Quantitative Proteomics, Negative Control

Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: ELF5 mediates casein synthesis by JAK2 and STAT5 activity in GMECs. (A) Cells were treated with Tyrphosting AG490 (30 μM) or DMSO, followed by pcDNA3.1-ELF5 or pcDNA3.1-NC transfection for 48 h. (B) Cells were treated with Tyrphosting AG490 (30 μM) or DMSO, followed by siRNA-ELF5 or siRNA-NC (100 nM) transfection for 48 h. The protein abundances of αS1-casein, αS2-casein, β-casein, κ-casein, p-JAK2, and p-STAT5 were detected. The relative protein abundance of αS1-casein, αS2-casein, β-casein, and κ-casein was normalized to β-tubulin. The relative protein abundance of p-JAK2 and p-STAT5 was normalized to total JAK2 and STAT5, respectively. Values are presented as mean±SEM. a–d Different lowercase letters represent significant differences (* p<0.05). NC, negative control; siRNA, small interfering ribonucleic acid; GMECs, goat mammary epithelial cells; SEM, standard error of the mean.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Activity Assay, Transfection, Quantitative Proteomics, Negative Control

Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: The interaction between ELF5 and STAT5 in GMECs. (A) Cells were transfected with ELF5-MYC and STAT5-FLAG for 48 h. Then, total protein of cells was extracted and incubated with MYC (or FLAG) tag antibody and protein G magnetic beads at 4°C overnight. The protein abundance of ELF5 and STAT5 was detected. (B) The mRNA expression of STAT5a and ELF5 gene was measured after pCMV-STAT5a and pCMV-NC transfection for 48 h. Values are presented as mean±SEM. ** p<0.01. GMECs, goat mammary epithelial cells; NC, negative control; SEM, standard error of the mean.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Transfection, Incubation, FLAG-tag, Magnetic Beads, Quantitative Proteomics, Expressing, Negative Control

Journal: Animal Bioscience

Article Title: ELF5 modulates casein synthesis in goat mammary epithelial cells via JAK2/STAT5 signaling pathway

doi: 10.5713/ab.25.0181

Figure Lengend Snippet: Molecular mechanism of ELF5 promotes casein synthesis by enhancing the activity of JAK2/STAT5 signaling pathway in goat mammary epithelial cells. PRLR stands for prolactin receptor.

Article Snippet: For the inhibition of JAK2/STAT5 pathway, JAK2 inhibitor AG490 (30 μM, MedChemExpress) or STAT5 inhibitor STAT5-IN-1 (50 μM, MedChemExpress) were treated with cells.

Techniques: Activity Assay

Journal: Aging Cell

Article Title: The Proinflammatory Secretome of Senescent Cells Can Be Controlled by a HIF2A ‐Dependent Upregulation and a FURIN ‐Dependent Cleavage of the ANGPTL4 Secreted Factor

doi: 10.1111/acel.70307

Figure Lengend Snippet: Knockdown of ANGPTL4 inhibits the ability of the SASP to induce activation of human neutrophils, a mark of inflammation. (A, B) Supernatants of MRC5/RAF:ER cells transfected with control (siCTRL), ANGPTL4 (siANGPTL4) or RELA (siRELA) siRNA and treated (+) or not (−) with 4‐OHT to induce senescence (OIS) were applied to primary human neutrophils. (A) Analysis of CD63 neutrophils degranulation marker. A representative flow cytometry profile is shown. Histograms on the right panel are the quantification of MFI. Mean ± SEM of n = 3 independent experiments. Paired one‐way ANOVA test results are shown. (B) Analysis of STAT5 pathway activation according to phosphorylation of STAT5. The left panel displays a representative flow cytometry profile. Histograms on the right panel are the quantification of MFI (Mean Fluorescence Intensity). Mean ± SEM of n = 3 independent experiments. Paired one‐way ANOVA test results are shown.

Article Snippet: Phosphorylation of STAT5, using pSTAT5 antibody (STAT5(PY694), BD), was evaluated using a panel of antibodies to identify neutrophils [CD45+ (HI30, BD), CD66b + (G10F5, BD), CD15high (HI98, BD), CD14‐ (M5E2, Biolegend), Siglec8‐ (REA1045, Miltenyi)].

Techniques: Knockdown, Activation Assay, Transfection, Control, Marker, Flow Cytometry, Phospho-proteomics, Fluorescence