Journal: Frontiers in Pharmacology

Article Title: Oxytocin Protects Against Isoproterenol-Induced Cardiac Hypertrophy by Inhibiting PI3K/AKT Pathway via a lncRNA GAS5/miR-375-3p/KLF4-Dependent Mechanism

doi: 10.3389/fphar.2021.766024

Figure Lengend Snippet: Antihypertrophic effects of OT in vitro . Neonatal rat cardiomyocytes stimulated with ISO for 24 h in the presence or absence of OT. (A) The cell morphology was evaluated by H&E staining. (B) Statistical results of measurement of cell surface areas. (C,D) Effects of OT on the protein expressions of BNP and β-MHC. (E) Effects of OT on the expression of lncRNA GAS5. (F) Effects of OT on the expression of miR-375-3p. (G,H) Effects of OT on the mRNA and protein expressions of KLF4. (I) Effects of OT on the p-PI3K/PI3K ratio. (J) Effects of OT on the p-AKT1/AKT1 ratio. (K) Western blot images of BNP, β-MHC, KLF4, p-PI3K, PI3K, AKT1, p-AKT1, and β-actin levels. Data are shown as the mean ± sd of three independent experiments. *, p < 0.05; **, p < 0.01.

Article Snippet: The blots were blocked with 5% non-fat milk and incubated overnight with the primary antibodies against BNP (1:500, rabbit #ab19645 abcam), β-MHC (1:1,000, rabbit #ab172967 abcam), KLF4 (1:1,000, rabbit #bs-1064R BIOSS), PI3K (1:1,000, rabbit #ab182651 abcam), p-PI3K (1:1,000, rabbit #205841-1-AP Proteintech), AKT1 (1:1,000, rabbit # bs-0115M BIOSS), p-AKT1 (1:1,000, rabbit # bs-0876R BIOSS), β-actin (1:1,000, rabbit #abmart P30002) and GAPDH (1:2,000, rabbit #2118S CST).

Techniques: In Vitro, Staining, Expressing, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Oxytocin Protects Against Isoproterenol-Induced Cardiac Hypertrophy by Inhibiting PI3K/AKT Pathway via a lncRNA GAS5/miR-375-3p/KLF4-Dependent Mechanism

doi: 10.3389/fphar.2021.766024

Figure Lengend Snippet: Over-expression of miR-375-3p blunted anti-hypertrophic effects of oxytocin via KLF4 and modulated the PI3K/AKT signaling pathway. (A) Expression of miR-375-3p in cardiomyocytes following transfection of miR-375-3p mimics and miR-375-3p mimics NC. (B) Expression of miR-375-3p was detected by qRT-PCR. (C,D) Expressions of BNP and β-MHC proteins were measured by Western blot analysis. (E) Expression of KLF4 mRNA was detected by qRT-PCR. (F) Expression of KLF4 protein was measured by Western blot analysis. (G) p-PI3K/PI3K ratio. (H) p-AKT1/AKT1 ratio. (I) Statistical results of measurement of cell surface areas. (J) Representative western blot images of KLF4, PI3K, p-PI3K, AKT1, p-AKT1, BNP, β-MHC and GAPDH levels. (K) Cardiomyocyte surface areas were measured by immunofluorescent staining. Scale bars represent 20 µm. Images were captured at ×400 magnification. (a) ISO+OT+miR-375-3p mimics NC group. (b) ISO+OT+miR-375-3p mimics group. (c) ISO+OT+miR-375-3p mimics+pcDNA-NC group. (d) ISO+OT+miR-375-3p mimics+pcDNA-KLF4 group. Data are shown as the mean ± sd of three independent experiments. *, p < 0.05; **, p < 0.01.

Article Snippet: The blots were blocked with 5% non-fat milk and incubated overnight with the primary antibodies against BNP (1:500, rabbit #ab19645 abcam), β-MHC (1:1,000, rabbit #ab172967 abcam), KLF4 (1:1,000, rabbit #bs-1064R BIOSS), PI3K (1:1,000, rabbit #ab182651 abcam), p-PI3K (1:1,000, rabbit #205841-1-AP Proteintech), AKT1 (1:1,000, rabbit # bs-0115M BIOSS), p-AKT1 (1:1,000, rabbit # bs-0876R BIOSS), β-actin (1:1,000, rabbit #abmart P30002) and GAPDH (1:2,000, rabbit #2118S CST).

Techniques: Over Expression, Expressing, Transfection, Quantitative RT-PCR, Western Blot, Staining

Journal: Frontiers in Pharmacology

Article Title: Oxytocin Protects Against Isoproterenol-Induced Cardiac Hypertrophy by Inhibiting PI3K/AKT Pathway via a lncRNA GAS5/miR-375-3p/KLF4-Dependent Mechanism

doi: 10.3389/fphar.2021.766024

Figure Lengend Snippet: knock-down of KLF4 blunted anti-hypertrophic effects of oxytocin via PI3K/AKT pathway. (A–C) Detection of relative shRNA-KLF4 interference effects. The changes in KLF4 mRNA and protein levels were detected by qRT-PCR and Western blotting after transfection of shRNA-KLF4 #1, shRNA-KLF4 #2, shRNA-KLF4 #3 in primary cardiomyocytes. (D) Expression of KLF4 mRNA was determined by qRT-PCR. (E) Expression of KLF4 protein was measured by Western blot analysis. (F,G) Expressions of BNP, β-MHC proteins were measured by Western blot analysis. (H) p-PI3K/PI3K ratio. (I) p-AKT1/AKT1 ratio. (J) Statistical results of measurement of cell surface areas. (K) Cardiomyocyte surface areas were measured by immunofluorescent staining. Scale bars represent 20 µm. Images were captured at ×400 magnification. (L) Western blot images of KLF4, PI3K, p-PI3K, AKT1, p-AKT1, BNP, β-MHC and GAPDH levels. (a) ISO+OT+shRNA-NC group. (b) ISO+OT+shRNA-KLF4 group. (c) ISO+OT+shRNA-KLF4+LY194002 group. Data are shown as the mean ± sd of three independent experiments. *, p < 0.05; **, p < 0.01.

Article Snippet: The blots were blocked with 5% non-fat milk and incubated overnight with the primary antibodies against BNP (1:500, rabbit #ab19645 abcam), β-MHC (1:1,000, rabbit #ab172967 abcam), KLF4 (1:1,000, rabbit #bs-1064R BIOSS), PI3K (1:1,000, rabbit #ab182651 abcam), p-PI3K (1:1,000, rabbit #205841-1-AP Proteintech), AKT1 (1:1,000, rabbit # bs-0115M BIOSS), p-AKT1 (1:1,000, rabbit # bs-0876R BIOSS), β-actin (1:1,000, rabbit #abmart P30002) and GAPDH (1:2,000, rabbit #2118S CST).

Techniques: shRNA, Quantitative RT-PCR, Western Blot, Transfection, Expressing, Staining

Journal: International Journal of Molecular Sciences

Article Title: Improved Wound Healing and Skin Regeneration Ability of 3,2′-Dihydroxyflavone-Treated Mesenchymal Stem Cell-Derived Extracellular Vesicles

doi: 10.3390/ijms24086964

Figure Lengend Snippet: List of antibodies for Western blot.

Article Snippet: Akt1 polyclonal antibody , rabbit , Cusabio (CSB-PA000855).

Techniques: Western Blot, Recombinant

Journal: Poultry Science

Article Title: MiR-223 inhibits proliferation and steroid hormone synthesis of ovarian granulosa cell via the AKT signaling pathway by targeting CRIM1 in chicken

doi: 10.1016/j.psj.2024.103910

Figure Lengend Snippet: Antibody dilution concentration.

Article Snippet: anti-AKT , Bioss , Bs-0115R , 1:1000.

Techniques: Concentration Assay

Journal: Oncology Letters

Article Title: Chronic restraint stress promotes gastric epithelial malignant transformation by activating the Akt/p53 signaling pathway via ADRB2

doi: 10.3892/ol.2022.13420

Figure Lengend Snippet: Chronic restraint stress increases the effects induced by the detrimental diet via the Akt/p53 signaling pathway mediated via ADRB2. Protein expression levels in gastric tissues were detected via western blotting. (A) Representative western blotting bands and the semi-quantification of (B) ADRB2; (C) Akt; (D) p-Akt; (E) p-Akt/Akt; and (F) p-p53/p53. *P<0.05, **P<0.01 and ***P<0.001. ADRB2, adrenoceptor β2; p, phosphorylated; DD, detrimental diet group; DR, detrimental diet with chronic restraint group; DRP, detrimental diet with chronic restraint and propranolol treatment group.

Article Snippet: After blocking with 5% skimmed milk dissolved in TBS containing 0.5% Tween-20 (TBST) at room temperature for 1 h, the membrane was incubated with the following primary antibodies: Akt (rabbit polyclonal antibody; 1:300; cat. no. bs-6951R; BIOSS), phosphorylated (p)-Akt (mouse monoclonal antibody; 1:100; cat. no.sc-514032; Santa Cruz Biotechnology, Inc.), p53 (rabbit polyclonal antibody; 1:500; 10442-1-AP; ProteinTech Group, Inc.), p-p53 (Thr18; rabbit polyclonal antibody; 1:500; cat. no.bs-3710R; BIOSS), ADRB2 (rabbit polyclonal antibody; 1:500; cat. no.bs-0947R; BIOSS) and the loading control GAPDH (rabbit polyclonal antibody; 1:1,000; cat. no.bs-10900R; BIOSS) overnight at 4°C.

Techniques: Expressing, Western Blot

Journal: Oncology Letters

Article Title: Chronic restraint stress promotes gastric epithelial malignant transformation by activating the Akt/p53 signaling pathway via ADRB2

doi: 10.3892/ol.2022.13420

Figure Lengend Snippet: Putative mechanism via which chronic restraint stress affects gastric epithelial transformation. A detrimental diet resulted in DNA damage and activated the protein expression of p53. Restraint stress may act on ADRB2 receptors by promoting catecholamine released from the local nerve terminals of the gut, thus inducing the activation of Akt, which may contribute to the ubiquitination and degradation of p53 and thereby reduce the effects of p53. The apoptosis of epithelial cells was finely regulated by p53. ADRB2, adrenoceptor β2; MNNG, N-methyl-N'-nitro-N-nitrosoguanidine; p, phosphorylated.

Article Snippet: After blocking with 5% skimmed milk dissolved in TBS containing 0.5% Tween-20 (TBST) at room temperature for 1 h, the membrane was incubated with the following primary antibodies: Akt (rabbit polyclonal antibody; 1:300; cat. no. bs-6951R; BIOSS), phosphorylated (p)-Akt (mouse monoclonal antibody; 1:100; cat. no.sc-514032; Santa Cruz Biotechnology, Inc.), p53 (rabbit polyclonal antibody; 1:500; 10442-1-AP; ProteinTech Group, Inc.), p-p53 (Thr18; rabbit polyclonal antibody; 1:500; cat. no.bs-3710R; BIOSS), ADRB2 (rabbit polyclonal antibody; 1:500; cat. no.bs-0947R; BIOSS) and the loading control GAPDH (rabbit polyclonal antibody; 1:1,000; cat. no.bs-10900R; BIOSS) overnight at 4°C.

Techniques: Transformation Assay, Expressing, Activation Assay

Journal: Journal of Advanced Research

Article Title: Efficient improvement of the proliferation, differentiation, and anti-arthritic capacity of mesenchymal stem cells by simply culturing on the immobilized FGF2 derived peptide, 44-ERGVVSIKGV-53

doi: 10.1016/j.jare.2023.09.041

Figure Lengend Snippet: Antibody list.

Article Snippet: Membrane blocking was performed using 5 % skimmed milk in Tris-buffered saline for 1 h, followed by incubation with the appropriate primary antibodies against total ERK (Cat No. B7074, Assay Biotechnology, Fremont, CA, USA), phosphorylated ERK (Cat No. 9101 s, Cell Signaling Technology, Danvers, MA, USA), total AKT (Cat No. CSB-PA000855, CUSABIO, Houston, TX, USA), phosphorylated AKT (Cat No. CSB-PA008120, CUSABIO), IKKα (Cat No. 11930, Cell Signaling Technology), FGFR1 (Cat No. NBP2-33784, Novus Biologicals, Centennial, CO, USA), phosphorylated FGFR1 (Tyr653, Tyr654) (Cat No. 44-1140G, Invitrogen) polyclonal Antibody, FRS2 antibody (Cat No. 3836–100, BioVision, Waltham, MA, USA), phosphorylated FRS2-alpha (Cat No. 3864S, Cell Signaling) or ACTIN (Cat No. sc-8432, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) overnight at 4 °C.

Techniques: Staining, Plasmid Preparation