Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 2. The GLUT3 protein is upregulated in hypoxic immortalized prostate epithelial cells. (Upper and Middle panels) RWPE1 cells were cultured in either normoxic or under chronic hypoxic (1% O2) conditions for 48 h and then processed for immunofluorescence microscopy. Cells were stained for either CA9, GLUT1, or GLUT3 (green, Upper and Middle panels). DNA, blue. Scale, 25 µm. (Lower panels) Comparison of flow cytometry profiles of CA9, GLUT1, or GLUT3 in cells cultured in normoxia (red) versus hypoxia (yellow). Cells stained with only secondary antibody is shown (dark red). The fold increase in level of the indicated proteins is denoted. PE, phycoerythrin.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Cell Culture, Microscopy, Staining, Comparison, Cytometry

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 3. GLUT3 co-localizes with pimonidazole (Hypoxyprobe) in mouse xenograft tumors formed from prostate epithelial CTN-2-2 cells. Non-tumorigenic immortalized prostate epithelial PrEC-Hahn cells were transformed by transiently inducing chromosomal instability. The resulting tumorigenic line was named CTN-9, which produced malignant solid xenograft tumors in subcutaneously in- jected NSG mice [52]. Cell lines isolated from these tumors were named CTN1-2 and 2-2 lines [52]. (Upper panel) Single tumor section stained for pimonidazole (PIMO) (green) and GLUT3 (red). DNA, blue. Boxed regions are shown at higher magnification in the lower panels. (Lower panels) Higher magnification images from boxed regions 1 or 2 in upper panel. Serial tissue sections were stained for hematoxylin and eosin (H&E), PIMO (green), and GLUT3, GLUT1 or CA9 (red) as indicated. Concentrations of GLUT3 (yellow arrows) and CA9 (white arrows) that juxtapose with PIMO are marked. Dashed white boxes show GLUT3 concentrated at the plasma membrane in tumor cells (insets). Scale, 100 µm.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Transformation Assay, Produced, Isolation, Staining, Clinical Proteomics, Membrane

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 4. GLUT3 co-localizes with pimonidazole (Hypoxyprobe) in mouse xenograft tumors formed from prostate cancer DU145 cells. (Upper panel) Single tumor section stained for pimonidazole (PIMO) (green) and GLUT3 (red). DNA, blue. Boxed regions are shown at higher magnification in the lower panels. (Lower panels) Higher magnification images from boxed regions 1, 2, or 3 in upper panel. Serial tissue sections were stained for hematoxylin and eosin (H&E), PIMO (green), and GLUT3, GLUT1 or CA9 (red) as indicated. Concentrations of GLUT3 (yellow arrows) and CA9 (white arrows) that juxtapose with PIMO are marked. Dashed white boxes show GLUT3 concentrated at the plasma membrane in tumor cells (insets). Scale, 100 µm.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Staining, Clinical Proteomics, Membrane

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 5. Measurements of the co-localization of hypoxia markers in xenograft tumors show a high degree of association between pimonidazole and GLUT3. (a) Single section from a CTN2-2 tumor immunostained for pimonidazole (PIMO) (green) and GLUT3 (red). DNA, blue. Scale, 100 µm. (b) To quantify co-localization of hypoxia markers GLUT1, GLUT3, and CA9 with PIMO, binary tracings were placed around the PIMO signal (green lines) defined a threshold of intensity specific to each tumor. Binaries were then dilated >0–15 µm (orange lines) and >15–30 µm (red lines) to capture the adjacent signal of each hypoxia marker. Same image as in (a). (c) Graphs show fold change in mean fluorescence intensities of hypoxia markers GLUT1, GLUT3 and CA9 within three PIMO-positive or adjacent regions captured within the expanded binary masks: 0 µm where the PIMO signal resides, >0–15 µm (orange bars) and >15–30 µm (red bars). Mean fluorescence intensities within these three regions was divided by ‘background’ signal from regions >30 µm from the PIMO signal. A dashed line indicates no change in signal relative to the background. Four optical fields in tissue sections from two different DU145, CTN1-2 and CTN2-2 tumors were analyzed (six different tumors examined in total).

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Marker

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 6. Patient-derived xenograph (PDX) model of primary prostate cancer contains distinct pockets of GLUT3 staining. (a) Patient-derived xenograft (PDX) from a primary prostate tumor stained with H&E. Scale, 1.5 mm. (b) Section of PDX tumor immunostained for E-Cadherin (green) to mark cell borders and GLUT3 (red). DNA, blue. Boxed region is shown at higher magnification in (c). Scale, 250 µm. (c) Serial tissue sections were immunostained for either GLUT3 (red, left panel), a cocktail containing anti-GLUT1 and anti-CA9 antibodes (red, right panel), or antibodies against all three proteins GLUT1, GLUT3, and CA9 (red, middle panel) as indicated. Dashed white boxes show GLUT3 concentrated at the plasma membrane in tumor cells (insets). Scale, 100 µm.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Derivative Assay, Staining, Clinical Proteomics, Membrane

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 7. Patient-derived xenograph (PDX) model of bone metastatic prostate cancer contains distinct pockets of GLUT3 staining. (Upper panel) Section of PDX tumor immunostained for E-Cadherin (green) to mark cell borders and GLUT3 (red). DNA, blue. Boxed regions are shown at higher magnification (Lower panels). Scale, 500 µm. (Lower panels) Higher magnification images from boxed regions 1 or 2 in upper panel. Serial tissue sections were stained with H&E (column 1) or immunostained for either GLUT3 (red, column 2), a cocktail containing anti-GLUT1, GLUT3, and CA9, antibodies (red, column 3), or antibodies against GLUT1 and CA9 (red, column 4) as indicated. Dashed white boxes show GLUT3 concentrated at the plasma membrane in tumor cells (insets). Scale, 100 µm.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Derivative Assay, Staining, Clinical Proteomics, Membrane

Journal: Diagnostics (Basel, Switzerland)

Article Title: GLUT3/SLC2A3 Is an Endogenous Marker of Hypoxia in Prostate Cancer Cell Lines and Patient-Derived Xenograft Tumors.

doi: 10.3390/diagnostics12030676

Figure Lengend Snippet: Figure 8. Compared to hypoxia biomarkers GLUT1 and CA9, GLUT3 labels larger areas within PDX tumor sections. (a) Patient-derived xenograft (PDX) from a primary prostate tumor immunostained with the triple anti-CA9, GLUT1, and GLUT3 antibody cocktail. Scale, 100 µm. (b) Same image as in (a) with binary mask to outline regions of the triple stain. (c) Graph shows measurements of total area of each stain within 3 different PDX prostate tumors. Three sequential sections were obtained from each tumor, stained for either CA9/GLUT1, GLUT3, or the triple stain CA9/GLUT1/GLUT3, and imaged. Total area of staining was quantified by masking regions of high intensity staining using Nikon Elements image analysis software. Note, in the metastatic prostate PDX tumor samples, the vast majority of the signal observed in the triple stain is due to the GLUT3 stain. However, in the primary prostate PDX, GLUT3 staining alone is not as effective at revealing putative hypoxic regions compared to the triple stain.

Article Snippet: Antibodies used include GLUT1 (Atlas, Fort Collins, CO, USA, HPA058494), GLUT3 (Proteintech, Rosemont, IL, USA, 20403-IAP), and CA-9 (Novus, Littleton, CO, USA, NB100-417SS) at 1:1000 dilution.

Techniques: Derivative Assay, Staining, Software

Journal: Journal of Functional Foods

Article Title: Fucoidan ameliorates diabetic skeletal muscle atrophy through PI3K/Akt pathway

doi: 10.1016/j.jff.2024.106076

Figure Lengend Snippet: Fig. 7. Fucoidan ameliorates glucose metabolism in diabetic skeletal muscle and PA treated C2C12 myotubes through activating PI3K/Akt. (A) Muscle glycogen content of gastrocnemius muscle. (B) PAS staining in the gastrocnemius muscle. Magnification, ×200. Scale bar, 50 μm. (C) Muscle glycogen area in the gastroc nemius muscle. (D) Relative mRNA level of GLUT4, GSK-3β and GS. (E–F) Western blot and quantification of GLUT4, GSK-3β and GS in gastrocnemius muscle. (G) Glycogen content in C2C12 cells. (H) The different concentrations of PA treated glucose consumption of C2C12 myotubes. (I) Glucose consumption in C2C12 cells. (J) Relative mRNA level of GLUT4, GSK-3β and GS in C2C12 cells. (K–L) Western blot and quantification of GLUT4, GSK-3β and GS in gastrocnemius muscle. Data are expressed as means ± SD (n = 3). *Р < 0.05, as compared to the NC group, #Р < 0.05, as compared to the DM or PA group. &Р < 0.05, as compared to the PA + Fu group.

Article Snippet: The membrane was incubated with primary antibody for 1.5 h, including PI3K p85 (dilution 1:4000; cat# 60225-1-Ig; Proteintech, Wuhan, China), Akt (dilution 1:5000; cat# 60203-2-Ig; Proteintech, Wuhan, China), p-Akt (dilution 1:5000; cat# 60225-1-Ig; Proteintech, Wuhan, China), mTOR (dilution 1:2000; cat# ab32028; Abcam, Cambridge, MA, United States), p-mTOR (dilution 1:1000; cat# ab109268; Abcam, Cambridge, MA, United States), p70S6K (dilution 1:1000; cat# ER31205; Huabio, Hangzhou, China), p-p70S6K (dilution 1:1000; cat# 34475S; Cell Signaling Technology, Danvers, MA, United States), GLUT4 (dilution 1:2000; cat# 66846-1-Ig; Proteintech, Wuhan, China), GSK-3β (dilution 1:2000; cat# ET1607-71; Huabio, Hangzhou, China), p-GSK-3β (dilution 1:2000; cat# ET1607-60; Huabio, Hangzhou, China), GS (dilution 1:1000; cat# ET1611-59; Huabio, Hangzhou, China), FoxO1 (dilution 1:3000; cat# 18592-1-AP; Proteintech, Wuhan, China), Atrogin-1 (dilution 1:1000; cat# ET7109-25; Huabio, Hangzhou, China), MuRF1 (dilution 1:2000; cat# HA500057; Huabio, Hangzhou, China) and β-actin (dilution 1:50000; cat# ET1701-80; Huabio, Hangzhou, China).

Techniques: Staining, Western Blot

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 4. Angptl2 knockout improved renal UA excretion in HUA mice model (A) Angptl2 knockout and identification protocol (left panel), and genotype identification results by DNA agarose gel electrophoresis (right panel); (B) HUA mice model flow chart. (C) Fasting body weight before sacrifice. (D–G) Serum UA, CREA, BUN and XOD levels in WT, HUA, Angptl2 ko-HUA groups. (H) Urinary UA level in WT, HUA, Angptl2 ko-HUA groups. (I) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in the kidney. (J) Representative data of ABCG2 and GLUT9 in the kidney by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; CREA, creatinine; BUN, blood urea nitrogen; UA, uric acid; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Knock-Out, Agarose Gel Electrophoresis, Western Blot, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 5. Adipocyte-specific overexpression of Angptl2 significantly increased serum UA levels in mice induced by adenine and potassium oxonate (A) HUA mice model flow chart. (B) Fasting body weight before sacrifice. (C) Serum ANGPTL2 levels after adipocyte-specific Angptl2 overexpression. (D–F) Serum CREA, BUN and UA levels after adipocyte-specific Angptl2 overexpression. (G) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in the kidney. (H) Representative data of ABCG2 and GLUT9 in the kidney by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; CREA, creatinine; BUN, blood urea nitrogen; HUA, hyperuricemia; UA, uric acid; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Over Expression, Western Blot, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 6. ANGPTL2 decreased ABCG2 by suppressing AKT activity in HK2 cells (A) Flow chart of HK2 treatment groups. (B) UA levels in HK2 cell supernatants. (C) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in HK2 cells. (D) Representative data of ABCG2 and GLUT9 in HK2 cells by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; rhA2, human recombinant proteins ANGPTL2; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Activity Assay, Western Blot, Recombinant, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 7. ANGPTL2 decreased ABCG2 by suppressing AKT activity in primary renal tubular epithelial cells. (A) Flow chart of RTECs treatment groups. (B) UA levels in RTECs supernatants. (C) Relative mRNA level of Abcg2, Slc22a8, Slc2a9 and Slc22a12 in RTECs. (D) Representative data of ABCG2 and GLUT9 in RTECs by Western blotting (left panel), and statistic data (right panel). ANGPTL2, angiopoietin-like protein 2; rhA2, human recombinant proteins ANGPTL2; HUA, hyperuricemia; ABCG2, ATP binding cassette subfamily G member 2; OAT3, organic anion transporter 3; GLUT9, glucose transporter 9; URAT1, urate transporter 1. *p < 0.05, **p < 0.01, ****p < 0.0001.

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Activity Assay, Western Blot, Recombinant, Binding Assay

Journal: Free radical biology & medicine

Article Title: Adipocyte-secreted ANGPTL2 promotes hyperuricemia through inhibiting AKT/ABCG2 signaling.

doi: 10.1016/j.freeradbiomed.2025.03.048

Figure Lengend Snippet: Fig. 9. Adipocyte-secreted ANGPTL2 promotes UA level through AKT/ABCG2 signaling in HK2 cells. (A) Flow diagram of C3H10 induction and treatment with FFA, as well as oil red O staining. (B) Representative data of ANGPTL2 after FFA treated C3H10 cells differentiated adipocytes by Western blotting (left panel), and statistic data (right panel). (C) ANGPTL2 levels in FFA-treated C3H10 cells differentiated adipocytes. (D) Flow chart of conditioned medium collected from C3H10 differ entiated into adipocytes treated with HK2 cells. (E) UA levels in HK2 cell supernatants after conditioned medium treatment. (F) Representative data of ABCG2 in HK2 cells by Western blotting (left panel), and statistic data (right panel). (G) Graphic summary of the study. Adipocyte-secreted ANGPTL2 decreased ABCG2 through inhibited AKT activation, then increased UA levels and led to HUA. ANGPTL2, angiopoietin-like protein 2; FFA, free fatty acid; HUA, hyperuricemia; ABCG2, ATP binding cassette G member 2; GLUT9, glucose transporter 9. *p < 0.05, **p < 0.01, ****p < 0.0001. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: After being deparaffinized, 0.5uM kidney sections were incubated overnight with the primary antibody ABCG2 (Abcam, USA) and GLUT9 (Proteintech, China) and stained the next day with the IHC kit (MXB Biotechnologies, China) and DAB (Proteintech, China) kit.

Techniques: Staining, Western Blot, Activation Assay, Binding Assay

Journal: PLoS ONE

Article Title: Glucose-coated superparamagnetic iron oxide nanoparticles prepared by metal vapor synthesis can target GLUT1 overexpressing tumors: In vitro tests and in vivo preliminary assessment

doi: 10.1371/journal.pone.0269603

Figure Lengend Snippet: GLUT1 levels estimated by ELISA in PSN-1, BCPAP and HEK293 cells (at the top). Uptake studies: PSN-1 and BCPAP cells were incubated with 0.1 mg/mL of Glc-SPIONs for 30 min, 1, 3, 6, and 12 h. The Fe cellular content was estimated by GF-AAS analysis. Uptake studies after treating the cells with GLUT1 inhibitors: PSN1 and BCPAP cells were pre-incubated for 1 h with anti-GLUT1 polyclonal antibody, WZB117, Fasentin, BAY-876, and STF-31. Subsequently, cells were treated for 3 or 6 h with 0.1 mg/mL of Glc-SPIONs.

Article Snippet: Expression levels in cancer cells were evaluated by means of the GLUT1 Colorimetric Cell-Based ELISA kit (Boster Biological Technology, Pleasanton CA, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Incubation