acta 1 cat no e2972hu  (Shanghai Korain Biotech Co Ltd)

Journal: Journal of the American Heart Association

Article Title: Methyltransferase‐Like 3–Mediated N6‐Methyladenosine RNA Methylation Regulates Hypoxia‐Induced Pulmonary Arterial Smooth Muscle Cell Pyroptosis by Targeting PTEN

doi: 10.1161/jaha.124.034470

Figure Lengend Snippet: Figure 3. PTEN facilitates METTL3-mediated m6A modification. A, CatRAPID analysis prediction of the interaction between METTL3 and PTEN. B, Potential sites and regions of m6A modification in the PTEN gene sequence (http://www.cuilab.cn/sramp). C, D, The protein and mRNA levels of PTEN in METTL3-overexpressing PASMCs were measured by western blot and qRT-PCR (Nor, n=6–7; Hyp, n=6–7; HYP+METTL3, n=6–7; HYP+Vector, n=6–7). E, F, Repressive effects of METTL3 siRNA on PTEN expression at both the protein and mRNA levels (Nor, n=6; HYP, n=6; HYP+si- METTL3-1, n=6; HYP+si-METTL3-2, n=6; HYP+si-nitrocellulose, n=6). G, The expression of METTL3 was positively correlated with the expression of PTEN in hypoxic mice. H, Colocalization of METTL3 and PTEN mRNA in PASMCs as determined by the confocal immunofluorescence assay. Scale bars=100 μm. I, J, RIP analysis with an anti-m6A antibody was employed to detect METTL3- mediated PTEN m6A modifications. The m6A modification on PTEN was enhanced by METTL3 overexpression (Vector, n=4; METTL3, n=4), while it was depleted upon METTL3 knockdown (nitrocellulose-METTL3, n=4; Si-METTL3-1, n=4; Si-METTL3-2, n=4). K, L, At the indicated time points, the mRNA levels of PTEN with (Vector, n=4; METTL3, n=4) or without METTL3 (nitrocellulose-METTL3, n=4; Si-METTL3-1, n=4; Si-METTL3-2, n=4) depletion in the presence of the transcription inhibitor actinomycin D were measured by qRT-PCR. Each data point in the figure represents a unique biological replicate. Statistical analysis was performed with one-way ANOVA. The data are presented as the mean±SD. *P<0.05, **P<0.01, ***P<0.001. ActD indicates actinomycin D; Hyp, hypoxia; m6A, N6-methyladenosine; METTL, methyltransferase-like; NC, nitrocellulose; Nor, normal; PASMC, pulmonary artery smooth muscle cell; PTEN, phosphate and tension homology deleted on chromosome 10; qRT-PCR, quantitative real-time polymerase chain reaction; RIP, RNA immunoprecipitation; and si-METTL, small interfering methyltransferase-like.

Article Snippet: The cells were incubated with an anti–α- smooth muscle actin antibody (1:100; Boster,Wuhan, China ), an anti- METTL3 antibody (1:100; Proteintech,IL,USA), and an anti- PTEN antibody (1:100; Proteintech,IL,USA) overnight at 4 °C.

Techniques: Modification, Sequencing, Western Blot, Quantitative RT-PCR, Plasmid Preparation, Expressing, Immunofluorescence, Over Expression, Knockdown, Real-time Polymerase Chain Reaction, RNA Immunoprecipitation

Journal: Aging and disease

Article Title: The NRF2/ID2 Axis in Vascular Smooth Muscle Cells: Novel Insights into the Interplay between Vascular Calcification and Aging

doi: 10.14336/ad.2024.0075

Figure Lengend Snippet: Figure 1. Age Exacerbated VC and Reduced NRF2 Signal in Mice. (A) Representative micrographs depict von Kossa staining in serial sections of aged and young mice aortas under normal or high-phosphate (High-Pi) conditions (Scale bars represent 250μm and 50μm). (B) Quantification of calcium deposition in the aortas of aged and young mice with or without High-Pi stimulation (n=6, 10, 6, and 10, respectively). (C) Immunofluorescence co-staining illustrates P16 (red) and α-SMA (α-smooth muscle actin; green) in serial sections of aged and young mice aortas under both conditions (Scale bars represent 100μm, 50μm and 20μm). Arrows indicate areas of intranuclear P16 upregulation. (D-E) Representative micrographs of TUNEL assay (Scale

Article Snippet: The primary antibodies were: α-smooth muscle actin (α-SMA) (BM0002, 1:200, BOSTER, China), ID2 (3431, 1:70, CST, USA), p16 (ab54210, 1:100, Abcam, USA), and Nrf2 (GTX103322, 1:500, GeneTex, USA).

Techniques: Staining, Immunofluorescence, TUNEL Assay

Journal: iScience

Article Title: TRAIL-dependent apoptosis of peritoneal mesothelial cells by NK cells promotes ovarian cancer invasion

doi: 10.1016/j.isci.2023.108401

Figure Lengend Snippet:

Article Snippet: Mouse monoclonal anti-human alpha-smooth muscle actin (aSMA) (clone ASM-1) , Progen , Cat# 61001; RRID: AB_2920672.

Techniques: Control, Recombinant, Polymer, Software