Journal: Pharmaceutics

Article Title: Amino Acid Nanofibers Improve Glycemia and Confer Cognitive Therapeutic Efficacy to Bound Insulin

doi: 10.3390/pharmaceutics14010081

Figure Lengend Snippet: AAC2-hINS increases glucose uptake in 3T3-L1 preadipocytes and human BBB endothelial cells via GLUT4 and GLUT1. ( a ) FD-glucose uptake in mouse 3T3-L1 preadipocytes ( n = 24 per treatment) treated with vehicle (Veh; PBS, open bar) or hINS (1.7 µM, red bar) or AAC2 (0.03 and 3 µM black bar) or AAC2-hINS (purple bar) with the respective concentration of AAC2 and a constant hINS concentration. After 2 h 15 min starvation, FD glucose and treatment reagents were added for an additional 85 min of incubation. Data (%) were normalized to FD-glucose uptake in control (Veh) 3T3-L1 cells (100%). Student’s independent t -test for all tests in figure. ( b ) AAC2 dose-dependent uptake FD-glucose in 3T3-L1 preadipocytes. Data are shown as % of increase compared to non-stimulated cells. ( c ) Non-differentiated NIH-3T3 cells were transiently transfected with GLUT4-GFP. Translocation of GLUT4-GFP (white, yellow arrow) cytosol to the membrane after stimulation with Veh (PBS), hINS (10 µg/mL), AAC2 (10 µM), or AAC2-hINS was demonstrated using confocal microscopy (60× magnification). ( d ) FD-glucose uptake in human brain endothelial cells ( n = 7 per treatment) treated with vehicle (Veh; PBS, open bar) or hINS (1.7 µM, red bar) or AAC2 (0.1 µM black bar), or AAC2-hINS (purple bar) in the presence and absence of GLUT1 inhibitor (BAY-876; 10 nM in DMSO). Cells were pre-treated with or without BAY-876 for 40 min and then FD glucose and treatment reagents were added for an additional 50 min of incubation. Data (%) were normalized to FD-glucose uptake in control (Veh without inhibitor, 100%).

Article Snippet: GLUT1 inhibitor (BAY-876, Selleckchem, Houston, TX S8452, USA)

Techniques: Concentration Assay, Incubation, Control, Transfection, Translocation Assay, Membrane, Confocal Microscopy

Journal: MedComm

Article Title: Deletion of Trp53 and Rb1 in Ctsk‐expressing cells drives osteosarcoma progression by activating glucose metabolism and YAP signaling

doi: 10.1002/mco2.131

Figure Lengend Snippet: Loss of Trp53 and Rb1 promotes osteosarcoma progression by overactivating energy metabolism. (A) Volcano plot of transcriptome profiles between human osteosarcoma samples and controls from BioProject database under the accession code: PRJNA539828; 3996 genes upregulated; 2953 genes downregulated. (B) KEGG analyses of the osteosarcoma RNA‐seq data showing the top 20 enriched metabolism‐related pathways. Red boxes direct to glucose‐involved metabolic pathways. (C) Glut1 expression between human osteosarcoma samples and controls (BioProject database, PRJNA539828). OS: osteosarcoma. (D) Glut1 expression in the cortical bones of the Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls as indicated. (E–H) The cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls as indicated are seeded in 12‐well plate. After incubation for 48 h, the glucose consumption (E), lactate production (F), and ATP level (H) were identified as described in Materials and Methods section. After incubation with 100 μM 2‐NBDG of 8 h, the glucose uptake was determined by a fluorescence microscope at 485/540 nm (G). (I–L) Analyses of the cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice treated with or without 1 μM Glut1 inhibitor Bay‐876 as indicated. The corresponding quantification was identified at right. (I) Migration. (J) Invasion. (K) Soft agar. (L) Tumorsphere. Scale bar: 100 μm. Error bars are the means ± SEM from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Glut1 inhibitor Bay‐876 was obtained from MCE Med Chem Express.

Techniques: RNA Sequencing, Expressing, Incubation, Fluorescence, Microscopy, Migration

Journal: MedComm

Article Title: Deletion of Trp53 and Rb1 in Ctsk‐expressing cells drives osteosarcoma progression by activating glucose metabolism and YAP signaling

doi: 10.1002/mco2.131

Figure Lengend Snippet: Trp53 and Rb1 orchestrate energy metabolism through Glut1 in osteosarcoma. (A) Diagram of the key enzymes in the glucose metabolism such as Glut1, Hk2, Pfkfb3/4 and Ldha. (B–F) mRNA expressions of Glut1, Hk2, Pfkfb3/4 and Ldha were identified after transfection for 48 h with Ad‐Cre or Ad‐GFP in the cortical bone cells from Trp53 f/f /Rb1 f/f mice and controls as indicated. (G–I) The cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls as indicated are seeded in 12‐well plate. After incubation for 48 h, the glucose consumption (G), lactate production (H), and ATP level (I) were identified as described in Materials and Methods section. (J–L) The glucose consumption (J), lactate production (K), and ATP level (L) were identified in cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice after treatment of 48 h with or without 1 μM Glut1 inhibitor Bay‐876. Error bars are the means ± SEM from three independent experiments. * p < 0.05, ** p < 0.01

Article Snippet: Glut1 inhibitor Bay‐876 was obtained from MCE Med Chem Express.

Techniques: Transfection, Incubation

Journal: MedComm

Article Title: Deletion of Trp53 and Rb1 in Ctsk‐expressing cells drives osteosarcoma progression by activating glucose metabolism and YAP signaling

doi: 10.1002/mco2.131

Figure Lengend Snippet: Enhanced energy metabolism in osteosarcoma depends on increased YAP activity and its mediated‐Glut1 expression. (A) qRT‐PCR analysis of YAP in cortical bone of Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls. (B) Representative images of immunofluorescent staining of YAP in the cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls. Scale bars: 25 μm. (C) Western blot as indicated. (D) The cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls were co‐transfected with luciferase reporter and pRL‐TK plasmids (internal control) as indicated, respectively. After transfection of 48 h, the luciferase activities were identified by the Dual‐Luciferase Assay Kit. (E) Schematic binding diagram of YAP/TEAD1 complex in Glut1 promoter. BS and TSS represented binding site and transcription start site, respectively. (F and G) ChIP assay. Co‐occupation of YAP/TEAD1 complex in Glut1 promoter in cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice and controls (F). Co‐occupation of YAP/TEAD1 in Glut1 promoter after silence of TEAD1 (G). (H and I) qRT‐PCR analysis of Glut1 after overexpression (H) or silence (I) of YAP. (J–L) After silence of YAP in cortical bone cells from Ctsk‐Cre;Trp53 f/f /Rb1 f/f mice for 48 h, the glucose consumption (J), lactate production (K), and ATP level (L) were identified as described in Materials and Methods section. Error bars are the means ± SEM from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Glut1 inhibitor Bay‐876 was obtained from MCE Med Chem Express.

Techniques: Activity Assay, Expressing, Quantitative RT-PCR, Staining, Western Blot, Transfection, Luciferase, Control, Binding Assay, Over Expression

Journal: MedComm

Article Title: Deletion of Trp53 and Rb1 in Ctsk‐expressing cells drives osteosarcoma progression by activating glucose metabolism and YAP signaling

doi: 10.1002/mco2.131

Figure Lengend Snippet: Deletion of Trp53 and Rb1 in Ctsk‐expressing cells drives osteosarcoma progression by activating YAP‐mediated energy metabolism. Glut1‐mediated glucose metabolism was enhanced after loss of Trp53 and Rb1 in Ctsk‐expressing cells, which was dependent on YAP activity and directly regulated by YAP/TEAD1 complex

Article Snippet: Glut1 inhibitor Bay‐876 was obtained from MCE Med Chem Express.

Techniques: Expressing, Activity Assay