Journal: Artificial cells, nanomedicine, and biotechnology

Article Title: Silencing MEG3 protects PC12 cells from hypoxic injury by targeting miR-21.

doi: 10.1080/21691401.2020.1725533

Figure Lengend Snippet: Figure 6. Silence of MEG3 activates PI3K/AKT pathway while deactivates NF-jB pathway via targeting miR-21. PC12 cells were co-transfected with si-MEG3/si-NC and miR-21 inhibitor/NC inhibitor, and then incubated in hypoxia for 24 h. The cells without transfection in normoxic condition served as control (CTRL). The expres- sion of proteins involved in (A,B) PI3K/AKT and (C,D) NF-jB pathways was tested by western blot. n ¼ 3.p < .05 represents a comparison of marked groups.

Article Snippet: Proteins were carried out by using primary antibodies specific for CyclinD1 (orb77046), p53 (orb178524), p21 (orb38089), Bcl-2 (orb378569), Bax (orb378566), pro-caspase-3 (orb13278), cleaved-caspase-3 (orb310032), PTEN (orb39110), PI3K (orb395443), p-PI3K (orb106105), AKT (orb29949), p-AKT (orb344403), IjBa (orb13487), p-IjBa (orb338946), p65 (orb344389), p-p65 (orb106211), and b-actin (orb378579, all from Biorbyt, San Francisco, CA).

Techniques: Transfection, Incubation, Control, Western Blot, Comparison

Journal: Molecular Medicine Reports

Article Title: SDF-1α/CXCR4 signaling promotes capillary tube formation of human retinal vascular endothelial cells by activating ERK1/2 and PI3K pathways in vitro

doi: 10.3892/mmr.2022.12821

Figure Lengend Snippet: Effect of SDF-1α/CXCR4 signaling on the protein expression of ERK1/2 and PI3K pathway. (A) Representative results of ERK1/2, p-ERK1/2, PI3K and p-PI3K expression of HRECs were determined using western blotting. SDF-1α/CXCR4 signaling significantly increased the protein expression of ERK1/2 and PI3K pathway. (B) Statistical analysis of ratios of p-ERK1/2 and p-PI3K to GAPDH protein. (C) Statistical analysis of ratios of p-ERK1/2/total ERK1/2 and p-PI3K/total PI3K. All values represent mean ± standard error of the mean (n=3). *P<0.05 vs. control. SDF, stromal cell-derived factor; CXCR, chemokine (C-X-C motif) receptor; p-phosphorylated; HRECs, human retinal endothelial cells.

Article Snippet: Rabbit anti-Erk 1,2 monoclonal antibody (cat. no. orb178404; Clone B20-U; 1:5,000), Rabbit anti-phosphorylated (p-) ERK 1,2 monoclonal antibody (cat. no. orb178405; Clone G15-B; 1:5,000), rabbit anti-PI3K P85 (phospho-Tyr467) polyclonal antibody (cat. no. orb14998, 1:5,000) and rabbit anti PI3K polyclonal antibody (cat. no. orb1089274, 1:1,000) were purchased from Biorbyt Ltd. Human retinal vascular endothelial cells (HRECs; cat. no. YS0884) were purchased from Yaji Biological Technology Co., Ltd.

Techniques: Expressing, Western Blot, Control, Derivative Assay

Journal: Journal of Ovarian Research

Article Title: Gut microbiota-derived Tryptophanol driven by N-Carbamylglutamate alleviates premature ovarian failure through inhibiting oxidative stress

doi: 10.1186/s13048-026-02152-4

Figure Lengend Snippet: Tryptophanol treatment alleviated POF symptoms by inhibiting PI3K/AKT and MAPK pathways. A Experimental scheme. B Estrus status of mice ( n = 8). C Body weights of mice ( n = 8). D Ovary weights of mice ( n = 8). E Representative Ovary from the Control, the POF and the NCG groups mice with H&E and IHC staining. H&E scale bar = 200 μm; IHC scale bar = 100 μm. F Quantification of primordial, primary, secondary, antral follicles, corpora lutea and corpora atretica per ovary. n = 3 ovaries per each condition. Levels of FSH ( G ), LH ( H ), E 2 ( I ) in mouse serum ( n = 6). Levels of ROS ( J ), MDA ( K ), SOD ( L ), GSH-Px ( M ) in mouse ovaries ( n = 3–4). N Western blotting ( a ) and quantitative ( b ) analysis of proteins ( n = 3). O Litter sizes of mice ( n = 3). Data presented as the mean ± SD, statistical significance was determined using one-way or two-way ANOVA. * p < 0.05, ** p < 0.01. *** p < 0.001. Different lowercase letters indicate significant differences ( p < 0.05)

Article Snippet: The membrane was blocked with 5% bovine serum albumin (BSA; Sigma) for 1 h at room temperature, followed by overnight incubation at 4 °C with the primary antibodies: rabbit Cleaved caspase 3 (MCE, HY- P80623 , 1:800), rabbit BAX (Bioworld, BS61098, 1:800), rabbit BCL2 (Bioworld, BS1511, 1:800), rabbit ERK1/2 (Bioworld, BS90472, 1:1,000), rabbit p-ERK1/2 (Bioworld, BS4621P, 1:1,000), rabbit p38 (Bioworld, BS4635, 1:1,000), rabbit p-p38 (Bioworld, BS4766, 1:1,000), rabbit PI3K (MCE, HY- P80867 , 1:1,000); rabbit p-PI3K (MCE, HY- P80846 , 1:1,000), rabbit AKT (Bioworld, MB3288 1:1,000); rabbit p-AKT (Bioworld, BS4007 1:1,000); rabbit IL-6 (Abclonal, A0286, 1:800), rabbit IL-1β (MCE, HY- P80720 , 1:1,000), rabbit Occludin (Abclonal, A2601, 1:1,000); rabbit ZO-1 (Bioworld, BS71522, 1:1,000); rabbit GPX4 (MCE, HY- P80450 , 1:1,000); mouse GAPDH (Bioworld, MB001, 1:1,000); rabbit SOD2 (Bioworld, BS6734, 1:1,000).

Techniques: Control, Immunohistochemistry, Western Blot

Journal: Journal of Ovarian Research

Article Title: Gut microbiota-derived Tryptophanol driven by N-Carbamylglutamate alleviates premature ovarian failure through inhibiting oxidative stress

doi: 10.1186/s13048-026-02152-4

Figure Lengend Snippet: Tryptophanol treatment alleviated POF by inhibiting PI3K/AKT and MAPK pathways in vitro . A Experimental scheme. B Different concentrations of Cy on GCs viability was assessed by CCK8 assay. C and E Different concentrations of Trp on GCs viability was assessed by CCK8 assay. D The effect of Trp on GCs cell viability following Cy treatment. The control and treatment groups were stained with DCFH-DA, the cells were washed and examined by fluorescence microscopy. Scale bar = 50 μm. F The fluorescence intensity of ROS in mouse GCs. G The level of MDA in mouse GCs. H Flow cytometry apoptosis analysis graph and cell death rate in different groups. The level of SOD ( I ), GSH-Px ( J ), and E 2 ( K ) in mouse GCs. ( L ) Western blotting ( a ) and quantitative ( b ) analysis of proteins ( n = 3). All values are presented as the mean ± SD ( n = 3). Different lowercase letters indicate significant differences compared to the control group ( p < 0.05)

Article Snippet: The membrane was blocked with 5% bovine serum albumin (BSA; Sigma) for 1 h at room temperature, followed by overnight incubation at 4 °C with the primary antibodies: rabbit Cleaved caspase 3 (MCE, HY- P80623 , 1:800), rabbit BAX (Bioworld, BS61098, 1:800), rabbit BCL2 (Bioworld, BS1511, 1:800), rabbit ERK1/2 (Bioworld, BS90472, 1:1,000), rabbit p-ERK1/2 (Bioworld, BS4621P, 1:1,000), rabbit p38 (Bioworld, BS4635, 1:1,000), rabbit p-p38 (Bioworld, BS4766, 1:1,000), rabbit PI3K (MCE, HY- P80867 , 1:1,000); rabbit p-PI3K (MCE, HY- P80846 , 1:1,000), rabbit AKT (Bioworld, MB3288 1:1,000); rabbit p-AKT (Bioworld, BS4007 1:1,000); rabbit IL-6 (Abclonal, A0286, 1:800), rabbit IL-1β (MCE, HY- P80720 , 1:1,000), rabbit Occludin (Abclonal, A2601, 1:1,000); rabbit ZO-1 (Bioworld, BS71522, 1:1,000); rabbit GPX4 (MCE, HY- P80450 , 1:1,000); mouse GAPDH (Bioworld, MB001, 1:1,000); rabbit SOD2 (Bioworld, BS6734, 1:1,000).

Techniques: In Vitro, CCK-8 Assay, Control, Staining, Fluorescence, Microscopy, Flow Cytometry, Western Blot

Journal: Cellular and molecular biology (Noisy-le-Grand, France)

Article Title: Pro-inflammatory GPR75 and anti-apoptotic phospholipase signaling pathways contribute to the ameliorating effect of soluble epoxide hydrolase inhibition on chronic experimental autoimmune encephalomyelitis in mice.

doi: 10.14715/cmb/2023.69.10.2

Figure Lengend Snippet: Figure 3. TPPU increased the PLC β4, PI3K p85α, Akt1, MEK1/2, ERK1/2, CREB1, and Bcl-2 expression and/or activity induced by MOG35-55 peptide/PT in the (a, b, c, d, e, f, and g) brain and (h, i, j, k, l, m, and n) spinal cord of PBS-, DMSO-, or TPPU-injected mice. The expression of unphosphorylated and phosphorylated pro teins for PLC β4, PI3K p85α, Akt1, MEK1/2, ERK1/2, CREB1, and Bcl-2 in tissues was measured by immunoblotting technique. Data are expressed as means ± SEM of 3-4 animals. + P<0.05 versus PBS- injected mice (EAE control group); # P<0.05 versus DMSO-injected mice (vehicle control group).

Article Snippet: Nitrocellulose membranes containing transferred proteins from the gel were blocked with nonfat dry milk in Tris-buffered saline and then incubated with specific antibodies against (1) sEH (13560; Cayman), (2) GPR75 (GTX55193; GeneTex, Hsinchu City, Taiwan), (3) Gαq/11 (sc-515689; Santa Cruz Biotechnology, Santa Cruz, CA, USA), (4) c-Jun (sc-1694; Santa Cruz), (5) phosphorylated c-Jun (p-c-Jun) (sc-16312; Santa Cruz), (6) PLC β4 (sc-16631; Santa Cruz); (7) PI3K p85α (OAAF05915; Aviva Systems Biology, San Diego, CA, USA), (8) phosphorylated PI3K p85α (p-PI3K p85α) (OAEC00301; Aviva Systems Biology), (9) Akt1 (sc-271149; Santa Cruz), (10) phosphorylated Akt1 (p-Akt1) (sc-52940; Santa Cruz), (11) MEK1/2 (sc-81504; Santa Cruz), (12) phosphorylated MEK1/2 (p-MEK1/2) (sc-81503; Santa Cruz), (13) ERK1/2 (sc-514302; Santa Cruz), (14) phosphorylated ERK1/2 (p-ERK1/2) (sc-136521; Santa Cruz), (15) CREB1 (sc-271; Santa Cruz), (16) phosphorylated CREB1 (p-CREB1) (sc-81486; Santa Cruz), (17) Bcl-2 (sc-7382; Santa Cruz), (18) myelin PLP (bs-11093R-HRP; Bioss Antibodies Inc., Woburn, MA, USA), and (19) SEMA3A (sc-74554; Santa Cruz) in bovine serum albumin overnight at 4°C.

Techniques: Expressing, Activity Assay, Injection, Western Blot, Control