Journal: Biology

Article Title: 1,2-Dichloroethane Induces Neuronal Apoptosis Through AMPK-mTOR-ULK1 Pathway Activation and Autophagic Flux Impairment

doi: 10.3390/biology15181580

Figure Lengend Snippet: 1,2-DCE exposure activates the AMPK-mTOR-ULK1 pathway and impairs autophagic flux in the mouse hippocampus. ( A ) Representative western blot bands of LC3-II/I, Beclin 1, p62, LAMP1 and CTSD. ( B ) Quantitative analysis of LC3-II/I expression. ( C ) Quantitative analysis of Beclin 1 expression. ( D ) Quantitative analysis of p62 expression. ( E ) Quantitative analysis of LAMP1 expression. ( F ) Quantitative analysis of CTSD expression. ( G ) Representative western blot bands of p-AMPK, AMPK, p-mTOR, mTOR, p-ULK1 and ULK1. ( H ) Quantitative analysis of the p-AMPK/AMPK ratio. ( I ) Quantitative analysis of the p-mTOR/mTOR ratio. ( J ) Quantitative analysis of the p-ULK1/ULK1 ratio. n = 5. *, vs. control; &, vs. 0.225 g/m 3 ; #, vs. 0.45 g/m 3 , p < 0.05.

Article Snippet: The primary antibodies used in this study were as follows: Bax (1:1000, A20227, ABclonal, Wuhan, China), Bcl-2 (1:1000, A19693, ABclonal), cleaved caspase 3 (1:1000, YM343, Immunoway, San Jose, CA, USA), LC3B (1:1000, A19665, ABclonal), Beclin 1 (1:1000, A21191, ABclonal), p62 (1:1000, WL02385, Wanleibio, Shenyang, China), lysosome-associated membrane protein 1 (LAMP1, 1:1000, A23947, ABclonal), cathepsin D (CTSD, 1:1000, 74089, Cell Signaling Technology, Danvers, MA, USA), p-AMPK (Thr172, 1:1000, WL05103, Wanleibio), AMPK (1:1000, WL02254, Wanleibio), p-mTOR (Ser2448, 1:1000, AP0115, ABclonal), mTOR (1:1000, WL02477, Wanleibio), p-ULK1 (Ser555, 1:1000, YP1542, Immunoway), ULK1 (1:1000, WL03067, Wanleibio), GAPDH (1:2000, sc-32233, Santa Cruz, Dallas, TX, USA), and β-actin (1:2000, sc-47778, Santa Cruz).

Techniques: Western Blot, Analysis, Expressing, Control

Journal: Biology

Article Title: 1,2-Dichloroethane Induces Neuronal Apoptosis Through AMPK-mTOR-ULK1 Pathway Activation and Autophagic Flux Impairment

doi: 10.3390/biology15181580

Figure Lengend Snippet: 2-CE exposure induces AMPK-mTOR-ULK1 pathway activation in PC12 cells. ( A ) Representative western blot bands of p-AMPK, AMPK, p-mTOR, mTOR, p-ULK1 and ULK1. ( B ) Quantitative analysis of p-AMPK/AMPK ratio. ( C ) Quantitative analysis of p-mTOR/mTOR ratio. ( D ) Quantitative analysis of p-ULK1/ULK1 ratio. n = 3. *, vs. control; &, vs. 22.5 mM, p < 0.05.

Article Snippet: The primary antibodies used in this study were as follows: Bax (1:1000, A20227, ABclonal, Wuhan, China), Bcl-2 (1:1000, A19693, ABclonal), cleaved caspase 3 (1:1000, YM343, Immunoway, San Jose, CA, USA), LC3B (1:1000, A19665, ABclonal), Beclin 1 (1:1000, A21191, ABclonal), p62 (1:1000, WL02385, Wanleibio, Shenyang, China), lysosome-associated membrane protein 1 (LAMP1, 1:1000, A23947, ABclonal), cathepsin D (CTSD, 1:1000, 74089, Cell Signaling Technology, Danvers, MA, USA), p-AMPK (Thr172, 1:1000, WL05103, Wanleibio), AMPK (1:1000, WL02254, Wanleibio), p-mTOR (Ser2448, 1:1000, AP0115, ABclonal), mTOR (1:1000, WL02477, Wanleibio), p-ULK1 (Ser555, 1:1000, YP1542, Immunoway), ULK1 (1:1000, WL03067, Wanleibio), GAPDH (1:2000, sc-32233, Santa Cruz, Dallas, TX, USA), and β-actin (1:2000, sc-47778, Santa Cruz).

Techniques: Activation Assay, Western Blot, Analysis, Control

Journal: Biology

Article Title: 1,2-Dichloroethane Induces Neuronal Apoptosis Through AMPK-mTOR-ULK1 Pathway Activation and Autophagic Flux Impairment

doi: 10.3390/biology15181580

Figure Lengend Snippet: AMPK-mTOR-ULK1 signaling mediates 2-CE-induced autophagy and subsequent apoptosis in PC12 cells. ( A ) Representative western blot bands of p-AMPK, AMPK, p-mTOR, mTOR, p-ULK1 and ULK1. ( B ) Quantitative analysis of the p-AMPK/AMPK ratio. ( C ) Quantitative analysis of the p-mTOR/mTOR ratio. ( D ) Quantitative analysis of the p-ULK1/ULK1 ratio. ( E ) Representative western blot bands of LC3-II/I, Beclin 1, p62, LAMP1 and CTSD. ( F ) Quantitative analysis of the LC3-II/I ratio. ( G ) Quantitative analysis of Beclin 1 expression. ( H ) Quantitative analysis of p62 expression. ( I ) Quantitative analysis of LAMP1 expression. ( J ) Quantitative analysis of CTSD expression. ( K ) Representative fluorescence images of autophagic flux. ( L ) Representative western blot bands of BAX and Bcl-2. ( M ) Quantitative analysis of the Bcl-2/BAX ratio. n = 3. *, vs. control; Δ, vs. 90 mM, p < 0.05.

Article Snippet: The primary antibodies used in this study were as follows: Bax (1:1000, A20227, ABclonal, Wuhan, China), Bcl-2 (1:1000, A19693, ABclonal), cleaved caspase 3 (1:1000, YM343, Immunoway, San Jose, CA, USA), LC3B (1:1000, A19665, ABclonal), Beclin 1 (1:1000, A21191, ABclonal), p62 (1:1000, WL02385, Wanleibio, Shenyang, China), lysosome-associated membrane protein 1 (LAMP1, 1:1000, A23947, ABclonal), cathepsin D (CTSD, 1:1000, 74089, Cell Signaling Technology, Danvers, MA, USA), p-AMPK (Thr172, 1:1000, WL05103, Wanleibio), AMPK (1:1000, WL02254, Wanleibio), p-mTOR (Ser2448, 1:1000, AP0115, ABclonal), mTOR (1:1000, WL02477, Wanleibio), p-ULK1 (Ser555, 1:1000, YP1542, Immunoway), ULK1 (1:1000, WL03067, Wanleibio), GAPDH (1:2000, sc-32233, Santa Cruz, Dallas, TX, USA), and β-actin (1:2000, sc-47778, Santa Cruz).

Techniques: Western Blot, Analysis, Expressing, Fluorescence, Control

Journal: International Journal of Molecular Sciences

Article Title: 1-Hydroxypyrene Promotes Bladder Cancer Progression and Tumor-Associated Immunosuppression via MIF

doi: 10.3390/ijms27177778

Figure Lengend Snippet: MIF mediates 1-OHP-induced tumor-associated immunosuppression and PI3K/AKT/mTOR pathway activation. ( A , B ) Correlation analysis showing the associations of MIF expression with CD74 and CD44, respectively. ( C , D ) Flow cytometric analysis of CD74 and CD44 expression in EJ cells after the indicated treatments. ( E , F ) Schematic illustration of PBMC isolation, CD8 + T-cell sorting, and co-culture with bladder cancer cells. ( G – L ) Flow cytometric analysis of Ki67, GZMB, and IFNG expression in CD8 + T cells after co-culture with differently treated EJ cells. ( M ) GSEA plot showing enrichment of the HALLMARK_MTORC1_SIGNALING pathway. ( N ) Correlation analysis showing a positive association between MIF expression and PI3K/AKT pathway activity. ( O , P ) Western blot analysis of PI3K/AKT/mTOR pathway activation in EJ and T24 cells under the indicated treatments. ns, not significant; * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: The antibodies included ACTIN (Abclonal, Cat. No. AC026), mTOR (Abclonal, Cat. No. A2445), p-mTOR (Abclonal, Cat. No. AP0094), AKT (Abclonal, Cat. No. A29131), p-AKT (Abclonal, Cat. No. AP1208), PI3K (Abclonal, Cat. No. A22730), and p-PI3K (Abclonal, Cat. No. AP1463).

Techniques: Activation Assay, Expressing, Isolation, FACS, Co-Culture Assay, Activity Assay, Western Blot