Journal: International journal of molecular sciences

Article Title: miR-212-5p Regulates PM 2.5 -Induced Apoptosis by Targeting LAMC2 and LAMA3.

doi: 10.3390/ijms26041761

Figure Lengend Snippet: Figure 3. miR-212-5p negatively regulates LAMC2 and LAMA3: (A) miRanda (v3.3a) and TargetScan (v5.0) projected miR-212-5p target genes displayed; (B) similarity of rno-miR-212-5p and has-miR- 212-5p genes shown; (C) schematic representation of LAMC2-3′ UTR and LAMA3-3′ UTR with miR- 212-5p target prediction and reporter plasmid construction, where red characters indicate binding sites and ”*” characters indicate mutation sites; (D) luciferase activity assessed in 293T cells after co- transfection of LAMC2-WT, LAMC2-MUT, LAMA3-WT, and LAMA3-MUT with miR-212-5p mimics or miR-mimics NC for 48 h; (E) LAMC2-mVenus-WT, LAMC2-mVenus-MUT, LAMA3-mVenus-WT, and LAMA3-mVenus-MUT co-transfected with miR-mimics NC or miR-212-5p in 293T cells (scale bar: 300 µm); (F) RT-PCR assay for miR-212-5p mimics and miR-212-5p inhibitors on LAMC2 and LAMA3 expression after transfection; (G) Western blotting used to identify alterations in the LAMC2 and LAMA3 proteins. The mean ± standard deviation (n = 3) is used to express the data. ns, no significance; *, p < 0.05; **, p < 0.01; and ***, p < 0.001 indicate significance.

Article Snippet: Next, 5% skimmed milk (Beyotime Biotechnology, Shanghai, China) was blocked for 2 h, and the membranes were incubated with specific primary antibodies overnight at 4 ◦C: BAD, BCL-2, caspase-3, LAMC2, p85, AKT, p-AKT, IκBα, β-action (1:1000 dilution, Proteintech, Rosemont, IL, USA), p-IκBα, p65, p-p65 (1:500 dilution, Proteintech, IL, USA), LAMA3, p-p85 (1:500 dilution, Abways, Shanghai, China), and GAPDH (1:7000 dilution, Abways, Shanghai, China).

Techniques: Plasmid Preparation, Binding Assay, Mutagenesis, Luciferase, Activity Assay, Cotransfection, Transfection, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot, Standard Deviation

Journal: International journal of molecular sciences

Article Title: miR-212-5p Regulates PM 2.5 -Induced Apoptosis by Targeting LAMC2 and LAMA3.

doi: 10.3390/ijms26041761

Figure Lengend Snippet: Figure 4. PM2.5-induced changes in LAMC2 and LAMA3 expression: (A) LAMC2 and LAMA3 gene expression detected with RT-PCR after PM2.5 stimulation; (B) LAMC2 and LAMA3 protein changes detected with Western blotting; (C) silencing efficiency of LAMC2 and LAMA3 after transfection with siRNA−LAMC2 or siRNA−LAMA3 detected with RT-PCR; (D) LAMC2 and LAMA3 protein changes detected with Western blotting. The mean ± standard deviation (n = 3) is used to express the data. ns, no significance; *, p < 0.05; **, p < 0.01; and ***, p < 0.001 indicate significance.

Article Snippet: Next, 5% skimmed milk (Beyotime Biotechnology, Shanghai, China) was blocked for 2 h, and the membranes were incubated with specific primary antibodies overnight at 4 ◦C: BAD, BCL-2, caspase-3, LAMC2, p85, AKT, p-AKT, IκBα, β-action (1:1000 dilution, Proteintech, Rosemont, IL, USA), p-IκBα, p65, p-p65 (1:500 dilution, Proteintech, IL, USA), LAMA3, p-p85 (1:500 dilution, Abways, Shanghai, China), and GAPDH (1:7000 dilution, Abways, Shanghai, China).

Techniques: Expressing, Gene Expression, Reverse Transcription Polymerase Chain Reaction, Western Blot, Transfection, Standard Deviation

Journal: International journal of molecular sciences

Article Title: miR-212-5p Regulates PM 2.5 -Induced Apoptosis by Targeting LAMC2 and LAMA3.

doi: 10.3390/ijms26041761

Figure Lengend Snippet: Figure 5. LAMC2 and LAMA3 inhibit apoptosis through the PI3K/AKT/NF-κB pathway: (A) PM2.5- stimulated (180 µg/mL for 24 h) cells transfected with siRNA−LAMC2 or siRNA−LAMA3, and p-IκBα/IκBα and p-p65/p65 protein changes; (B) p-AKT/AKT and p-p85/p85 protein changes; (C) BCL-2, BAD, and caspase-3 protein changes detected with Western blotting; (D) apoptosis rate detected with flow cytometry. The mean ± standard deviation (n = 3) is used to express the data. ns, no significance; *, p < 0.05; **, p < 0.01; and ***, p < 0.001 indicate significance.

Article Snippet: Next, 5% skimmed milk (Beyotime Biotechnology, Shanghai, China) was blocked for 2 h, and the membranes were incubated with specific primary antibodies overnight at 4 ◦C: BAD, BCL-2, caspase-3, LAMC2, p85, AKT, p-AKT, IκBα, β-action (1:1000 dilution, Proteintech, Rosemont, IL, USA), p-IκBα, p65, p-p65 (1:500 dilution, Proteintech, IL, USA), LAMA3, p-p85 (1:500 dilution, Abways, Shanghai, China), and GAPDH (1:7000 dilution, Abways, Shanghai, China).

Techniques: Transfection, Western Blot, Flow Cytometry, Standard Deviation