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GABAFG ameliorates lysosomal autophagy disorders and apoptotic in vivo and in vitro . (A-B)Western blotting analysis for Atg3, Atg5, Atg7, LC3II/I and p62 in pancreas tissues. Mice were experimented with HFD/STZ, HFD/STZ+GABAFG (40 mg/kg) or HFD/STZ+GABAFG (80 mg/kg). Data are expressed as mean ± SEM. (n = 3). (C-D) INS-1cells were treated with 40 mM HG or 40 mM HG with 2 μmol/L, 4 μmol/L, and 8 μmol/L GABAFG. Western blotting analysis of the expression of Atg3, Atg5, Atg7, LC3II/I and p62 in INS-1cells. INS-1cells were treated with 8 μmol/L GABAFG, 40 mmol/L HG, 40 mmol/L HG with 2 μmol/L GABAFG, 40 mmol/L HG with 4 μmol/L GABAFG or 40 mmol/L HG with 8 μmol/L GABAFG. (C) Western blotting analysis of the protein levels of Lamp2, CTSB, TFEB, and p-TFEB in pancreas tissues. (D) Western blotting analysis of the expression level of Lamp2, CTSB, TFEB, and p-TFEB proteins in untreated and treated INS-1 cells. (E) Western blotting analysis for nuclear and cytosolic TFEB in pancreas tissues from the Control group, HFD/STZ mice and T2DM mice with 80 mg/kg GABAFG. (F) Western blotting analysis of the nuclear and cytosolic expression levels of TFEB in INS-1 cells in the Control cells, 40 mmol/L HG cells and 40 mmol/L HG with 8 μmol/L GABAFG cells. (G) INS-1 cells in the presence or absence of 8 μmol/L GABAFG were treated or untreated with 50 μmol/L CQ under normal or HG conditions before collecting proteins. Western blotting analysis for autophagy-lysosome-related protein. (H) Western blotting analysis for Bcl-2, Bax, cleaved-caspase 3, and caspase 3 in pancreas tissues. (I) Western blotting analysis of the protein expression level of apoptosis-related proteins in INS-1 cells. (J) INS-1 cells in the presence or absence of 8 μm GABAFG were treated or untreated with 50 μmol/L CQ under normal or HG conditions before collecting proteins. (Q) INS-1 cells from the Control, HG (40 mmol/L), and HG (40 mmol/L)+GABAFG (8 μmol/L) groups were subjected to the Annexin V/PI test experiments in the presence or absence of CQ. (Z) INS-1 cells were transfected with si-NC or si-TFEB, then treated with or without 8 μmol/L GABAFG for 24 h under HG conditions.

Journal: Journal of Advanced Research

Article Title: Ginseng-derived GABAFG ameliorates type 2 diabetes mellitus by modulating autophagy-lysosome pathway and gut microbiota

doi: 10.1016/j.jare.2025.01.003

Figure Lengend Snippet: GABAFG ameliorates lysosomal autophagy disorders and apoptotic in vivo and in vitro . (A-B)Western blotting analysis for Atg3, Atg5, Atg7, LC3II/I and p62 in pancreas tissues. Mice were experimented with HFD/STZ, HFD/STZ+GABAFG (40 mg/kg) or HFD/STZ+GABAFG (80 mg/kg). Data are expressed as mean ± SEM. (n = 3). (C-D) INS-1cells were treated with 40 mM HG or 40 mM HG with 2 μmol/L, 4 μmol/L, and 8 μmol/L GABAFG. Western blotting analysis of the expression of Atg3, Atg5, Atg7, LC3II/I and p62 in INS-1cells. INS-1cells were treated with 8 μmol/L GABAFG, 40 mmol/L HG, 40 mmol/L HG with 2 μmol/L GABAFG, 40 mmol/L HG with 4 μmol/L GABAFG or 40 mmol/L HG with 8 μmol/L GABAFG. (C) Western blotting analysis of the protein levels of Lamp2, CTSB, TFEB, and p-TFEB in pancreas tissues. (D) Western blotting analysis of the expression level of Lamp2, CTSB, TFEB, and p-TFEB proteins in untreated and treated INS-1 cells. (E) Western blotting analysis for nuclear and cytosolic TFEB in pancreas tissues from the Control group, HFD/STZ mice and T2DM mice with 80 mg/kg GABAFG. (F) Western blotting analysis of the nuclear and cytosolic expression levels of TFEB in INS-1 cells in the Control cells, 40 mmol/L HG cells and 40 mmol/L HG with 8 μmol/L GABAFG cells. (G) INS-1 cells in the presence or absence of 8 μmol/L GABAFG were treated or untreated with 50 μmol/L CQ under normal or HG conditions before collecting proteins. Western blotting analysis for autophagy-lysosome-related protein. (H) Western blotting analysis for Bcl-2, Bax, cleaved-caspase 3, and caspase 3 in pancreas tissues. (I) Western blotting analysis of the protein expression level of apoptosis-related proteins in INS-1 cells. (J) INS-1 cells in the presence or absence of 8 μm GABAFG were treated or untreated with 50 μmol/L CQ under normal or HG conditions before collecting proteins. (Q) INS-1 cells from the Control, HG (40 mmol/L), and HG (40 mmol/L)+GABAFG (8 μmol/L) groups were subjected to the Annexin V/PI test experiments in the presence or absence of CQ. (Z) INS-1 cells were transfected with si-NC or si-TFEB, then treated with or without 8 μmol/L GABAFG for 24 h under HG conditions.

Article Snippet: In terms of cells, the primary antibody against p62 (1:200) was incubated with the treated INS-1 cells for an entire night at 4 °C, followed by the secondary antibody DyLight594 (1:400) (BOSTER, Wuhan, China).

Techniques: In Vivo, In Vitro, Western Blot, Expressing, Control, Transfection