human hif-1 alpha elisa kit, 1x96tests  (Advisains)

Journal: Nutrients

Article Title: Hydroxysafflor Yellow A Alleviates Acute Myocardial Ischemia/Reperfusion Injury in Mice by Inhibiting Ferroptosis via the Activation of the HIF-1α/SLC7A11/GPX4 Signaling Pathway

doi: 10.3390/nu15153411

Figure Lengend Snippet: Hydroxysafflor yellow A alleviated ferroptosis by activating the HIF-1α/SLC7A11/GPX4 signaling pathway. ( A ) The levels of ferroptosis-related proteins were assessed by Western blotting ( n = 3). ( B ) The total iron content in cardiac tissue was determined using a kit ( n = 6). ## p < 0.01 vs. the sham group; * p < 0.05, ** p < 0.01 vs. the MI/RI group.

Article Snippet: Anti-GPX4 (ab125066), anti-HIF-1α (ab179483), and Abcam. anti-SLC7A11 (ab216876) fluorescent antibodies were purchased from the USA.

Techniques: Western Blot

Journal: Nutrients

Article Title: Hydroxysafflor Yellow A Alleviates Acute Myocardial Ischemia/Reperfusion Injury in Mice by Inhibiting Ferroptosis via the Activation of the HIF-1α/SLC7A11/GPX4 Signaling Pathway

doi: 10.3390/nu15153411

Figure Lengend Snippet: Hydroxysafflor yellow A inhibited oxygen–glucose deprivation/reoxygenation (OGD/R)-induced ferroptosis in H9C2 cells. ( A ) The Fe 2+ level was detected by flow cytometry ( n = 3). ( B ) The total iron content in cells was detected using a kit ( n = 6). ( C ) The expression of GPX4 was evaluated by ELISA ( n = 6). ( D ) The expression of the ferroptosis-related protein GPX4 was measured by Western blot ( n = 3). ## p < 0.01 vs. the control group; * p < 0.05, ** p < 0.01 vs. the OGD/R group.

Article Snippet: Anti-GPX4 (ab125066), anti-HIF-1α (ab179483), and Abcam. anti-SLC7A11 (ab216876) fluorescent antibodies were purchased from the USA.

Techniques: Flow Cytometry, Expressing, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration assessed by ELISA in the brain compartment of the blood–brain barrier model in different glycemic conditions (hypo-, normo-, and hyperglycemia, resulting from applying 2.2 mM, 5 mM, and 25 mM glucose to the microvascular compartment, respectively). Measurements started 24 h after the establishment of different glycemic conditions in the microvascular compartment (MC) and continued for 36 h. Samples were taken from the brain compartment (BC) every 12 h (time points: 0 h, 12 h, 24 h, and 36 h). Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. Asterisks denote significant differences between glycemic groups (ANOVA and t -tests where appropriate, p value < 0.05). * p < 0.05; ** p < 0.002.

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration was assessed by ELISA upon LPS-induced neuroinflammation in the brain compartment of the blood–brain barrier model in different glycemic conditions (hypo-, normo-, and hyperglycemia, resulting from applying 2.2 mM, 5 mM, and 25 mM glucose to the microvascular compartment, respectively). Neuroinflammation was induced by the addition of lipopolysaccharide (LPS) at time point 0 h for 12 h to the brain compartment (BC) of the blood–brain barrier (BBB) to reach a final concentration of 0.2 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. Asterisks denote a significant difference between glycemic groups (ANOVA and t -tests where appropriate, p value < 0.05). *** p <0.001.

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration assessed by ELISA in hypoglycemic conditions (resulting from applying 2.2 mM glucose to the microvascular compartment) upon lipopolysaccharide and resveratrol treatment. Neuroinflammation was induced by the addition of lipopolysaccharide (LPS) at time point 0 h for 12 h in the brain compartment (BC) of the blood–brain barrier (BBB) to reach a final concentration of 0.2 μM. Resveratrol (RSV) solution was added at time point 12 h for 24 h to the microvascular compartment (MC) of the BBB to reach a final concentration of 50 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. Asterisks denote significant differences between groups (ANOVA and t -tests where appropriate, p value < 0.05). * p < 0.05; *** p < 0.001.

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration assessed by ELISA in normoglycemic conditions (resulting from applying 5 mM glucose to the microvascular compartment) upon lipopolysaccharide and resveratrol treatment. Neuroinflammation was induced by the addition of lipopolysaccharide (LPS) at time point 0 h for 12 h in the brain compartment (BC) of the blood–brain barrier (BBB) to reach a final concentration of 0.2 μM. Resveratrol (RSV) solution was added at time point 12 h for 24 h to the microvascular compartment (MC) of the BBB to reach a final concentration of 50 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. Asterisks denote significant differences between groups (ANOVA and t -tests where appropriate, p value < 0.05). ** p < 0.01.

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration assessed by ELISA in hyperglycemic conditions (resulting from applying 25 mM glucose to the microvascular compartment) upon lipopolysaccharide and resveratrol treatment. Neuroinflammation was induced by the addition of lipopolysaccharide (LPS) at time point 0 h for 12 h in the brain compartment (BC) of the blood–brain barrier (BBB) to reach a final concentration of 0.2 μM. Resveratrol (RSV) solution was added at time point 12 h for 24 h to the microvascular compartment (MC) of the BBB to reach a final concentration of 50 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. Asterisks denote significant differences between groups (ANOVA and t -tests where appropriate, p value < 0.05). * p < 0.05.

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration assessed by ELISA upon resveratrol administration to the microvascular compartment of the blood–brain barrier model in different glycemic conditions (hypo-, normo-, and hyperglycemia, resulting from applying 2.2 mM, 5 mM, and 25 mM glucose to the microvascular compartment, respectively). Resveratrol (RSV) solution was added at time point 12 h for 24 h to the microvascular compartment (MC) of the blood–brain barrier (BBB) to reach a final concentration of 50 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. There were no significant differences between glycemic groups at any analyzed time point (ANOVA, p value > 0.05).

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: International Journal of Molecular Sciences

Article Title: Interplay between Systemic Glycemia and Neuroprotective Activity of Resveratrol in Modulating Astrocyte SIRT1 Response to Neuroinflammation

doi: 10.3390/ijms241411640

Figure Lengend Snippet: Astrocyte-secreted sirtuin 1 concentration was assessed by ELISA upon resveratrol treatment of LPS-induced neuroinflammation in the brain compartment of the blood–brain barrier model in different glycemic conditions (hypo-, normo-, and hyperglycemia, resulting from applying 2.2 mM, 5 mM, and 25 mM glucose to the microvascular compartment, respectively). Neuroinflammation was induced by the addition of lipopolysaccharide (LPS) at time point 0 h for 12 h to the brain compartment (BC) of the blood–brain barrier (BBB) to reach a final concentration of 0.2 μM. Resveratrol (RSV) solution was added at time point 12 h for 24 h to the microvascular compartment (MC) of the BBB to reach a final concentration of 50 μM. Mean values of sirtuin 1 (SIRT1) concentration and standard errors are presented. There were no significant differences between glycemic groups at any analyzed time point (ANOVA, p value > 0.05).

Article Snippet: The level of astrocyte-secreted SIRT1 in the brain compartment (BC) was assessed using the Human SIRT1 ELISA Kit (Abcam, Cambridge, UK, cat. no. ab171573), following the manufacturer’s instructions.

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Acta Pharmaceutica Sinica. B

Article Title: GPRC5B protects osteoarthritis by regulation of autophagy signaling

doi: 10.1016/j.apsb.2023.05.014

Figure Lengend Snippet: Gprc5b -deficiency enhances cartilage degradation and inhibits ECM generation. (A) Representative IF or IHC images for COL10A1, MMP13, ACAN and COL2A1 expression in articular cartilage of mice following DMM surgery or sham control operation after 8 weeks. Scale bar, 100 μm. (B–E) Percentage of COL10A1 + (B), MMP13 + (C), and ACAN + (D), COL2A1 + (E) chondrocytes in articular of samples shown in (A). All data are presented as mean ± SD; n = 6; ∗∗ P < 0.01, ∗∗∗ P < 0.001. Two-way ANOVA followed by Dunnett's test was performed for multiple comparisons test.

Article Snippet: For immunofluorescence (IF) staining, the sections were incubated overnight at 4 °C with primary antibodies for Collagen Type X Alpha 1 (COL10A1, 1:100, Abcam, ab260040), Aggrecan (ACAN, 1:100, Proteintech, 13880-1-AP) and microtubule-associated protein light chain 3 (LC3, 1:100, Proteintech, 14600-1-AP).

Techniques: Expressing

Journal: Acta Pharmaceutica Sinica. B

Article Title: GPRC5B protects osteoarthritis by regulation of autophagy signaling

doi: 10.1016/j.apsb.2023.05.014

Figure Lengend Snippet: Overexpression of Gprc5b ameliorates OA pathogenesis in mice. (A) Experimental pattern for intra-articular injections of Gprc5b overexpression lentivirus vectors or vector controls. (B) Representative images of safranin O/Fast green staining showing cartilage destruction in Gprc5b -overexpression lentivirus injected mice and vector controls after DMM surgery. Scale bar, 100 μm. (C–E) The OARSI scoring of the medial tibia plateau (MTP) (C), medial femoral condyle (MFC) (D), total scoring (MTP + MFC) (E). All data are presented as mean ± SD; n = 6; ∗∗ P < 0.01. Two-tailed Student's t -test was performed. (F) OA-associated pain was measured by the hot-plate assay. All data are presented as mean ± SD; n = 6; ∗∗ P < 0.01. Two-tailed Student's t -test was performed. (G) Representative IF or IHC images for COL10A1, MMP13 and ACAN expression in articular cartilage of mice injected with Gprc5b- overexpression lentivirus vectors or vector controls after DMM surgery. Scale bar, 100 μm. (H–J) Percentage of COL10A1 + (H), MMP13 + (I), and ACAN + (J) chondrocytes in articular of samples shown in (G). All data are presented as mean ± SD; n = 6; ∗∗ P < 0.01, ∗∗∗ P < 0.001. Two-tailed Student's t -test was performed with experiments for two groups comparisons test.

Article Snippet: For immunofluorescence (IF) staining, the sections were incubated overnight at 4 °C with primary antibodies for Collagen Type X Alpha 1 (COL10A1, 1:100, Abcam, ab260040), Aggrecan (ACAN, 1:100, Proteintech, 13880-1-AP) and microtubule-associated protein light chain 3 (LC3, 1:100, Proteintech, 14600-1-AP).

Techniques: Over Expression, Plasmid Preparation, Staining, Injection, Two Tailed Test, Expressing

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: A PFKFB4 expression was elevated in NSCLC tissue samples, as analyzed by UALCAN database. LUSC: lung squamous cell carcinoma; LUAD: lung adenocarcinoma. B Kaplan–Meier analysis showing the negative correlation between PFKFB4 and the overall survival rate of NSCLC patients. Red: high expression of PFKFB4; black: low expression of PFKFB4. C Co-IP analysis of the interaction of endogenous FBXL7 with PFKFB4 in 293T cells. D Co-IP analysis of the interaction of exogenous FBXL7 with PFKFB4 in 293T cells. E GST-pull down assay analysis of the direct interaction between FBXL7 and PFKFB4 in 293T cells. F PFKFB4 mRNA expression in A549 cells wasn’t changed in response to oe-FBXL7, as determined by RT-qPCR. G Western blot analysis showing inhibited PFKFB4 protein expression in A549 cells in response to oe-FBXL7. H Western blot analysis showing enhanced degradation of PFKFB4 protein in 293T cells in response to oe-FBXL7 and CHX. I Ubiquitination level of PFKFB4 protein was reduced in 293T cells in the presence of si-FBXL7, as determined by IP assay. J Ubiquitination level of PFKFB4 protein was elevated in 293T cells in the presence of oe-FBXL7, as determined by IP assay. ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns p > 0.05. The cell experiment was repeated three times independently.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Expressing, Co-Immunoprecipitation Assay, Pull Down Assay, Quantitative RT-PCR, Western Blot

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: A Western blot analysis of FBXL7 and PFKFB4 proteins in A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. B Glucose uptake measurement in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. C Pyruvate level measurement in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. D Lactate production measurement in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. E ATP level measurement in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. F ROS content measurement in A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. G ROS content measurement in mitochondrion in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG. H Glycolysis of A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG as detected by ECAR assay ( # p < 0.05 vs. the oe-FBXL7 group; @ p < 0.05 vs. the oe-FBXL7 + oe-PFKFB4 group). I Viability of A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG, as analyzed by CCK-8 assay. J Migration and invasion of A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG, as analyzed by Transwell assay. K Apoptosis of A549 cells in response to oe-FBXL7, oe-FBXL7 + oe-PFKFB4, or oe-FBXL7 + oe-PFKFB4 + 2-DG, as analyzed by flow cytometry. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns p > 0.05. The cell experiment was repeated three times independently.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Western Blot, ECAR Assay, CCK-8 Assay, Migration, Transwell Assay, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: A Western blot analysis showing elevated expression of EZH2 and HIF-1α proteins in hypoxic A549 cells. B Verification of the binding of HIF-1α to the EZH2 promoter region, as evaluated by dual-luciferase reporter assay. C Enrichment of HIF-1α in the promoter region of EZH2, as observed by ChIP assay. D mRNA expression of EZH2, FBXL7, and PFKFB4 in normoxic or hypoxic A549 cells in the presence of si-EZH2. E Western blot analysis showing the expression of EZH2, FBXL7, and PFKFB4 proteins in normoxic or hypoxic A549 cells in the presence of si-EZH2. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns p > 0.05. The cell experiment was repeated three times independently.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Western Blot, Expressing, Binding Assay, Luciferase, Reporter Assay

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: A The mRNA expression of EZH2, FBXL7 and PFKFB4 in hypoxia-exposed A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7 or si-EZH2 + si-FBXL7 + 2-DG. B Western blot analysis showing the expression of EZH2, FBXL7, and PFKFB4 proteins in hypoxia-exposed A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7 or si-EZH2 + si-FBXL7 + 2-DG. C Glucose uptake measurement in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG. D Pyruvate level measurement in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG. E Lactate production measurement in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG. F ATP level measurement in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG. G ROS content measurement in A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7 or si-EZH2 + si-FBXL7 + 2-DG. H ROS content measurement in mitochondrion in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG. I Glycolysis of A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG, as detected by ECAR assay ( # p < 0.05 vs. the Hypoxia + si-EZH2 group; @ p < 0.05 vs. the Hypoxia + si-NC + si-FBXL7 group). J Viability of A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7 or si-EZH2 + si-FBXL7 + 2-DG, as analyzed by CCK-8 assay. K Migration and invasion of A549 cells, as analyzed by Transwell assay. L Apoptosis of A549 cells in response to si-EZH2, si-EZH2 + si-FBXL7, or si-EZH2 + si-FBXL7 + 2-DG, as analyzed by flow cytometry. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. ns p > 0.05. The cell experiment was repeated three times independently.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Expressing, Western Blot, ECAR Assay, CCK-8 Assay, Migration, Transwell Assay, Flow Cytometry

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: A Quantitative analysis for tumor volume of nude mice subcutaneously inoculated with cells transduced with sh-EZH2 alone or combined with oe-PFKFB4. B Quantitative analysis for tumor weight of nude mice subcutaneously inoculated with cells transduced with sh-EZH2 alone or combined with oe-PFKFB4. C HE staining analysis of the tumor tissues of nude mice subcutaneously inoculated with cells transduced with sh-EZH2 alone or combined with oe-PFKFB4. D Immunohistochemical staining analysis of Ki-67, EZH2, PFKFB4, and FBXL7 proteins in the tumor tissues of nude mice subcutaneously inoculated with cells transduced with sh-EZH2 alone or combined with oe-PFKFB4. E TUNEL-positive cells in the tumor tissues of nude mice subcutaneously inoculated with cells transduced with sh-EZH2 alone or combined with oe-PFKFB4. *** p < 0.001, **** p < 0.0001. ns p > 0.05. n = 6 for mice in each group.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Transduction, Staining, Immunohistochemical staining, TUNEL Assay

Journal: Cell Death & Disease

Article Title: Hypoxia-mediated promotion of glucose metabolism in non-small cell lung cancer correlates with activation of the EZH2/FBXL7/PFKFB4 axis

doi: 10.1038/s41419-023-05795-z

Figure Lengend Snippet: Hypoxia induces the expression of HIF-1α in NSCLC cells. HIF-1α elevates expression of EZH2 which in turn reduces expression of FBXL7, and inhibits the ubiquitination of PFKFB4 by FBXL7, resulting in upregulation of PFKFB4 protein expression. By this mechanism, NSCLC cell glycolysis and malignant phenotypes are accelerated while cell apoptosis was decelerated.

Article Snippet: After being blocked using 5% BSA, membranes underwent overnight incubation at 4 °C with the diluted primary antibodies against EZH2 (5246, 1:1000, Cell Signaling Technologies [CST], Beverly, MA), FBXL7 (sc-374319, 1:1000, Santa Cruz Biotechnology, Santa Cruz, CA), HA (ab9110, 1:1000, Abcam, Cambridge, UK), PFKFB4 (ab137785, 1:1000, Abcam), HIF-1α (36169, 1:1000, CST), Myc (ab32, 1:1000, Abcam) and Actin (ab8226, 1:2000, Abcam).

Techniques: Expressing