Journal: Disease Markers

Article Title: Leptin Promotes HTR-8/SVneo Cell Invasion via the Crosstalk between MTA1/WNT and PI3K/AKT Pathways

doi: 10.1155/2022/7052176

Figure Lengend Snippet: Leptin mediates β -catenin activation through the crosstalk between MTA1/WNT and PI3K/AKT pathways in HTR-8/SVneo cells. (a) HTR-8/SVneo cells were treated with exogenous leptin (0 and 200 ng/ml) for 24 h, and MTA1, WNT1, p-GSK3 β (Ser9), and p-AKT (Ser473) levels were detected by Western blot. Data are shown as mean ± SD; ∗ P < 0.01 vs. leptin (0 ng/ml). (b) The knockdown efficiencies of MTA1 and WNT1 were analyzed by Western blot analysis and qRT-PCR. ∗ P < 0.05 vs. control. (c) HTR-8/SVneo cells were transfected with MTA1 siRNA or scramble siRNA (Scr) in the presence or absence of 200 ng/ml leptin for 24 h, and Western blot analysis was performed to detect the expression of MTA1, WNT1, p-GSK3 β (Ser9), and nuclear β -catenin. ∗ P < 0.01 vs. control and # P < 0.01 vs. leptin. (d) HTR-8/SVneo cells were transfected with WNT1 siRNA or scramble siRNA (Scr) in the presence or absence of 200 ng/ml leptin for 24 h, and Western blot analysis was performed to detect the expression of WNT1, p-GSK3 β (Ser9), and nuclear β -catenin. ∗ P < 0.01 vs. control and # P < 0.01 vs. leptin. (e) The knockdown efficiencies of AKT and β -catenin were analyzed by Western blot analysis and qRT-PCR. ∗ P < 0.05 vs. control. (f) HTR-8/SVneo cells were transfected with AKT siRNA or scramble siRNA (Scr) in the presence or absence of 200 ng/ml leptin for 24 h, and Western blot analysis was performed to detect the expression of AKT, p-GSK3 β (Ser9), and nuclear β -catenin. ∗ P < 0.01 vs. control and # P < 0.01 vs. leptin. All experiments were performed in triplicate.

Article Snippet: The following primary antibodies were used: MMP9 (ab76003), MTA1 (ab71153), WNT1 (ab15251), AKT (ab179463), GSK3 β (ab32391), β -catenin (ab32572), histone 3 (ab1791), β -actin (ab6276) (Abcam, USA); p-AKT (Ser473) (#4060), p-GSK3 β (Ser9) (#9322) (Cell Signaling Technology, USA).

Techniques: Activation Assay, Western Blot, Quantitative RT-PCR, Transfection, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Preparation and Multitarget Anti-AD Activity Study of Chondroitin Sulfate Lithium in AD Mice Induced by Combination of D-Gal/AlCl 3

doi: 10.1155/2022/9466166

Figure Lengend Snippet: Effects of CS-Li on tau phosphorylation in the hippocampi of mice. Western blotting of GSK-3 β , p-GSK-3 β (Ser9), p-tau (Ser396), p-tau (Ser404), and tau, GAPDH was used as the internal control; Western blotting of PP2A, β -actin was used as the internal control. # p < 0.05, ## p < 0.01, and ### p < 0.001 vs. the control group; ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001 vs. the model group.

Article Snippet: Rabbit anti-TNF- α , anti-IL-6, anti-IL-1 β , anti-p-p38MAPK, anti-p-Erk1/2, anti-p-NF- κ B, anti-p-GSK3 β (Ser9), and anti-PP2A antibodies were purchased from Cell Signaling Technology Inc. (Danvers, MA).

Techniques: Western Blot

Journal: BioMed Research International

Article Title: The Effects of Omega-3 Fatty Acid Supplementation on Dexamethasone-Induced Muscle Atrophy

doi: 10.1155/2014/961438

Figure Lengend Snippet: Protein expression in GA muscle samples from rats treated with n-3 PUFA or vehicle for 40 days and dexamethasone or vehicle (last 10 days). Relative optical density of total Akt (tAkt) and phosphorylated Akt (P-Akt) ((a) and (c)), total GSK3 β (tGSK3 β ) and phosphorylated GSK3 β (P-GSK3 β ) ((b) and (d)), and tAkt/P-Akt and tGSK3 β /P-GSK3 β ratios ((e) and (f), resp.). Bands representing tAkt, P-Akt, tGSK3, and P-GSK3 β are illustrated in (g). α -Tubulin (55 kDa) was used as control. The numbers represent the mean ± S.E.M. CT = control group ( n = 6), DX = dexamethasone group ( n = 5), n-3 = n-3 PUFA group ( n = 6), and DX + n-3 = dexamethasone + n-3 PUFA group ( n = 6). Values represent means ± S.E.M., analyzed by Student's t -test, # P < 0.05.

Article Snippet: The primary antibodies used were Akt (1 : 2,000, Cell Signaling #4691), P-Akt (1 : 2,000, Cell Signaling #4060, Ser473); mTOR (1 : 500, Cell Signaling #2972), P-mTOR (1 : 500, Cell Signaling #2971, Ser2448), GSK3 β (1 : 2,000, Cell Signaling #9315), P-GSK3 β (1 : 2,000, Cell Signaling #9322, Ser9), FoXO3a (1 : 500, Cell Signaling #2497), P-FoXO3a (1 : 500, Cell Signaling #9466, Ser253), and α -tubulin (1 : 30,000, Hybridoma bank) in 5% BSA/TBS-T.

Techniques: Expressing

Journal: International Journal of Endocrinology

Article Title: Ghrelin Increases Beta-Catenin Level through Protein Kinase A Activation and Regulates OPG Expression in Rat Primary Osteoblasts

doi: 10.1155/2015/547473

Figure Lengend Snippet: Representative Western blot (a) and quantification (b) of timecourse experiments performed on rat osteoblast-like cells treated with ghrelin (AG, 10 −10 M) for 15, 30, 60, and 90 min. Phosphorylated-GSK-3 β (P-GSK-3 β ) levels were measured in whole cell lysates and normalized to total GSK-3 β levels (Tot-GSK3 β ). Results are shown as ratio of P-GSK3 β measured in treated versus untreated cells. Data are the mean ± SEM of 5 experiments * P < 0.05, versus untreated cells.

Article Snippet: Western blots were performed using a specific antibody against rat β -catenin diluted 1 : 5000 in 5% milk in Tris–HCl with 0.1% Tween-20 (TBST); rat P-GSK3 β (1 : 2000 5% milk in TBST); rat total GSK3 β (1 : 2000 5% milk in TBST) (Cell Signaling Technology, Boston, MA) or against rat β -actin (1 : 2000 5% milk in TBST) (Santa Cruz Biotechnology, Inc., Heidelberg, Germany).

Techniques: Western Blot

Journal: International Journal of Endocrinology

Article Title: Ghrelin Increases Beta-Catenin Level through Protein Kinase A Activation and Regulates OPG Expression in Rat Primary Osteoblasts

doi: 10.1155/2015/547473

Figure Lengend Snippet: Effect of the pretreatment with the PKA inhibitor H89 (2 × 10 −6 M, 30 min before) on (a) β -catenin and (b) phosphorylated-GSK3 β (P-GSK3 β ) levels measured in whole cell lysates by Western blot on rat osteoblast-like cells (rOB) treated with ghrelin (AG, 10 −10 M for 1 h). β -catenin levels were normalized to β -actin levels; P-GSK3 β was normalized to total GSK3 β levels (Tot-GSK3 β ). Results are expressed as ratio of β -catenin or P-GSK3 β measured in treated cells versus untreated cells. Insets : typical gels obtained from rOB cell lysates after treatment with ghrelin alone or together with H89. Data are the mean ± SEM of 5 experiments * P < 0.05 versus untreated cells; § P < 0.05 versus AG treated cells.

Article Snippet: Western blots were performed using a specific antibody against rat β -catenin diluted 1 : 5000 in 5% milk in Tris–HCl with 0.1% Tween-20 (TBST); rat P-GSK3 β (1 : 2000 5% milk in TBST); rat total GSK3 β (1 : 2000 5% milk in TBST) (Cell Signaling Technology, Boston, MA) or against rat β -actin (1 : 2000 5% milk in TBST) (Santa Cruz Biotechnology, Inc., Heidelberg, Germany).

Techniques: Western Blot