Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 1 | The expression of ENO1 on the cell-surface of a subpopulation of CSCs. (A) Representative FACS plots showing patterns of CD44, CD133, and surface ENO1 (sENO1) staining of primary prostate adenocarcinoma (PAC)-derived 22Rv-1 cells with the frequency of the boxed CD44+CD133+ cell population (representing CSCs in PAC; left) or sENO1+ cells in CD44+CD133+ CSCs (middle) or cells in the other subpopulations (representing non-CSCs; right) shown. (B) The percentages of sENO1+ cell subpopulation in CD44+CD133+ 22Rv-1 cells or cells in the other subpopulations (others). (C) The percentages of sENO1+ cell subpopulation in CD44+CD133+ PC-3 cells or cells in the other subpopulations. (D) The percentages of sENO1+ cell subpopulation in CD90+ gastric adenocarcinoma (GAC) AGS or NCI-N87 cells (representing CSCs in GAC) or CD90- cells (representing non-CSCs). Error bars represent mean ± SEM from three independent experiment (n = 3). Unpaired t-test was performed throughout where **p < 0.01; ***p < 0.001 in (B–D).

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Expressing, Staining, Derivative Assay

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 2 | sENO1 marks a subpopulation of mesenchymal-like and highly invasive CSCs. (A) The relative transcript levels of the mesenchymal- (left) or pluripotency- (right) associated genes in sENO1+ CSCs (represented by CD44+CD133+ 22Rv-1 cells), sENO1- CSCs, and non-CSCs (represented by cells in the other subpopulations) using qRT-PCR analysis. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed throughout where *p < 0.05 versus non-CSCs; †p < 0.05 versus sENO1- CSCs. (B) Immunoblotting analysis of the indicated markers selected from (A) in non- CSCs, sENO+, and sENO- CSCs. Protein levels were quantified by densitometric analysis of the bands, normalized to b-tubulin (loading control). (C) Limiting dilution assay (LDA) demonstrating the tumorsphere-forming efficacy of each subset of tumor cells. Three independent experiments were performed (n = 6). Shown are maximum likelihood estimates with a 95% confidence interval, where **p < 0.01. (D) The invasive capacities of freshly sorted sENO1+ CSCs (represented by CD44+CD133+ 22Rv-1 cells), sENO1- CSCs, and non-CSCs in 22Rv-1 cells in a dual-chamber invasion assay. Shown are representative immunofluorescence images of the invaded cells, with cell nuclei stained with SYTOX-green (green). Scale bars = 500 µm. Right, the number of invaded cells. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed throughout where **p < 0.01.

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Quantitative RT-PCR, Western Blot, Control, Limiting Dilution Assay, Invasion Assay, Staining

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 3 | sENO1+ CSCs are highly pro-metastatic. (A) Representative BLI of NOD/SCID mice receiving an intra-splenic injection of sENO1+ CSCs (represented by CD90+ NCI-N87 cells), sENO1- CSCs (CD90- NCI-N87 cells), and non-CSCs (represented by CD90- cells). at the indicated time following cell inoculation. (B) Tumor bulk quantified as BLI normalized photon counts as a function of time. Error bars represent mean ± SEM from one experiment (n = 8 mice per group). Unpaired t-test was performed throughout where **p < 0.01 versus non-CSCs. (C) Representative BLI of NOD/SCID mice receiving intra-splenic injection of sENO1+

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Injection

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 4 | ENO1 is expressed on the invadopodial surface of CSCs. (A) Confocal views of PAC CSCs (represented by CD44+CD133+ PC-3 cells) showing the cross-section of invadopodia structures (represented by cortactin+F-acin+ puncta) with the colocalized surface ENO1 (sENO1; green), cortactin (red), and F-actin (magenta) that penetrate into the underlying gelatin matrix. Nuclei were counterstained with 4’,6-diamidino-2-phenylindole (DAPI; blue). Scale, 10 µm. (B) Top, a pie chart showing the percentage of sENO1+ invadopodia per PC-3 CSC. Bottom, a pie chart showing the percentage of sENO1+ invadopodia per GAC AGS CSC (represented by CD90+ AGS cells). (C) Left, representative three-dimensional (3D) reconstructed confocal image of CD44+CD133+ PC-3 CSCs showing the co- localization of sENO1 (green) and cortactin (red) at the ventral side of cell. Scale, 8 µm. Right upper, digital zoom-in image from serial Z sections (yellow rectangle) showing the spatial colocalization of sENO1 (green) and cortactin (red) at invadopodia. Scale, 5 µm. Right lower, the orthogonal view of the magnified areas (yellow squares at top) shown the distribution and localization of sENO1 and cortactin at the base of invadopodia. 3D rendered images of the invadopodia (arrows) were processed by using Imaris software. Scale, 1 µm.

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Software

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 5 | sENO1+ CSCs generate more invadopodia than their sENO1- counterparts. (A) Confocal views of sENO1+ PC-3 CSCs (represented by CD44+CD133+

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques:

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 6 | sENO1 contributes to the invadopodial formation and the matrix-degradative function of CSCs. (A) Immunoblotting analysis showing the effect of lentivirus shRNA-mediated knockdown (KD) of ENO1 expression in PC-3 cells. Protein levels were quantified by densitometric analysis of the bands, normalized to b- tubulin (loading control). (B) Bar graph showing the percentage of sENO1+ PC-3 cells with KD of ENO1 expression or control KD. (C) Bar graph showing the density of invadopodia (represented by cortactin+F-actin+ puncta) per cells in PC-3 CSCs (represented by CD44+CD133+ cells) or non-CSCs (represented by cells in other subpopulations) with ENO1 KD or control KD. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed where **p < 0.01, ***p < 0.001 in (B, C). (D) PC-3 cells with KD of ENO1 expression or the control KD cells were seeded on top of a fluorescein-conjugated gelatin matrix and immunostained with cortactin (green) or phalloidin (F-actin; red). Nuclei were counterstained with DAPI (blue). Right, the fluorescence intensity of fluorescein- conjugated gelatin within the boundary (determined by F-actin staining) of PC-3 cells with ENO1 KD or control KD (n = 50 cells counted per sample). Unpaired t-test was performed where ***p < 0.001. (E) Bar graph showing the invasive capacity of PC-3 CSCs with ENO1 KD or control KD in a dual-chamber invasion assay. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed where **p < 0.01 versus non-CSCs. (F) Representative immunofluorescence images of CD44+CD133+ PC-3 cells (representing CSCs) that had invaded the type I collagen matrix in the presence of an increasing concentration (0.1-1.0 µg/ml) of the anti-ENO1 polyclonal antibody (pAb; a-ENO1) in a dual-chamber invasion assay. The nuclei of the invaded cells were stained with SYTOX-green. Scale bars, 500 µm. Right, the number of invaded cells. Cells in other subpopulations (representing non-CSCs) were included as a control. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed throughout where *p < 0.05, **p < 0.01 versus non-CSCs. (G) The invadopodia density per cell in PC-3 CSCs or non-CSCs exposed to an increasing concentration of a-ENO1. Error bars represent mean ± SEM from three independent experiments (n = 50 cells counted per sample). Unpaired t-test was performed where *p < 0.05, **p < 0.01, ***p < 0.001 versus non-CSCs. (H) PC-3 CSCs were seeded on top of a gelatin matrix in the presence or absence of a-ENO1 (20 µg/ml). Shown are the extent of matrix degradation as reflected by immunostaining with anti-Col1-3/4C (red). Right, the total cell fluorescence intensity of Col1-3/4C in PC-3 CSCs treated with a-ENO1 or a control IgG (n = 50 cells counted per sample). Unpaired t-test was performed where ***p < 0.001.

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Western Blot, shRNA, Knockdown, Expressing, Control, Staining, Invasion Assay, Concentration Assay, Immunostaining

Journal: Frontiers in oncology

Article Title: A Novel Invadopodia-Specific Marker for Invasive and Pro-Metastatic Cancer Stem Cells.

doi: 10.3389/fonc.2021.638311

Figure Lengend Snippet: FIGURE 7 | CAV1 is indispensable for the surface localization of sENO1 on CSCs and its pro-invadopodia and pro-invasive functions. (A) Immunoblotting analysis showing the effect of lentivirus shRNA-mediated knockdown (KD) of CAV1 (top) or HSP70 (bottom) expression in PC-3 cells. Protein levels were quantified by densitometric analysis of the bands, normalized to b-tubulin (loading control). (B) Bar graph showing the percentage of sENO1+ cells in PC-3 CSCs (represented by CD44+CD33+ cells) with KD of CAV1 or HSP70 expression or control-KD. Unpaired t-test was performed throughout where ***p < 0.001 versus control KD. (C) Bar graph showing the density of invadopodia (represented by coractin+F-actin+ puncta) per cell in PC-3 CSCs or non-CSCs (represented by cells in other subpopulations) with CAV1 KD or control KD. Unpaired t-test was performed throughout where ***p < 0.001. (D) Bar graph showing the invasive capacity of PC-3 CSCs or non-CSCs with CAV1 KD or control KD in a dual-chamber invasion assay. Error bars represent mean ± SEM from three independent experiments (n = 3). Unpaired t-test was performed throughout where *p < 0.05 versus non-CSCs; †p < 0.05 versus control KD.

Article Snippet: In other experiments, to profile the expression pattern of sENO1 on invadopodia, the cells seeded on gelatin were immunostained for sENO1 with rabbit polyclonal anti-ENO1 (OriGene, Rockville, MD), after which the cells were fixed with 4% formaldehyde and then immunostained for cortactin and F-actin as described above.

Techniques: Western Blot, shRNA, Knockdown, Expressing, Control, Invasion Assay

Journal: Acta dermato-venereologica

Article Title: Reactivation of anti-human alpha-enolase antibody-positive Behçet's disease by carbon dioxide laser treatment.

doi: 10.2340/00015555-0830

Figure Lengend Snippet: Fig. 2. (a–b) Biopsy specimen obtained from the skin lesions on the forearm demonstrating septal infiltration of lymphohistiocytes and neutrophils with vasculitis consistent with the findings of erythema nodosum (haematoxylin and eosin stain, original magnification, a; ×100, b; ×400). (c–d) Immunohistochemistry of a biopsy section with rabbit anti-human α-enolase antibody showing expression of α-enolase in the cytoplasm of lymphohistiocytes, neutrophils, and endothelial cells (original magnification, c; ×100, d; ×400).

Article Snippet: Immunohistochemical analysis of a biopsy specimen was also performed using a rabbit anti-human α-enolase antibody (Acris Antibodies, Acris Antibodies GmbH, Herford, Germany) in accordance with a previous report (3).

Techniques: H&E Stain, Immunohistochemistry, Expressing

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: ENO1 expression in MM tumors is elevated compared with normal bone marrow tissues. A human MM tissue microarray (10 MM cases and 11 normal bone marrow cases, with duplicate cores per case) was used for immunohistochemical staining of ENO1. (A) Representative images were shown at ×10 (upper panels) or ×40 (lower panels) magnification. (B) Quantification of the ENO1-positively stained area. Each dot represents the result from one tissue core. The P-value was calculated with a two-tailed unpaired Student's t-test. ENO1, enolase-1; MM, multiple myeloma.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Expressing, Microarray, Immunohistochemical staining, Staining, Two Tailed Test

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: ENO1 knockdown attenuates lactate production, cell migration, cell viability and surface ENO1 expression. RPMI-8226 and U266 cells were transfected with ENO1-targeting siRNA (si-ENO1 #1 or #2) or control siRNA (scramble sequence) for 72 h, and ENO1 depletion efficiency was confirmed by (A and G) western blotting. GAPDH served as the loading control. (B) ENO1-knockdown RPMI 8226 cells were cultured for an additional 48 h, and then the supernatant was collected for determination of lactate levels. (C) Transwell migration assay, (D and H) cell viability assays, and measurement of cell surface ENO1 by (E and I) flow cytometry and (F) an antibody labeling assay were performed. All results are presented as the mean ± SD of three independent experiments. P-values were calculated with one-way ANOVA (with Tukey's post hoc test). ENO1, enolase-1; siRNA, small interfering RNA.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Knockdown, Migration, Expressing, Transfection, Control, Sequencing, Western Blot, Cell Culture, Transwell Migration Assay, Flow Cytometry, Antibody Labeling, Small Interfering RNA

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: Extracellular ENO1 enhances glycolysis and pro-cancer activities. RPMI-8226 cells were treated with the indicated concentrations of recombinant ENO1-WT. The studies were conducted in the presence or absence of 100 µg/ml ENO1 mAb (also termed HuL001). (A) The lactate concentration in the culture medium (upper panel) and intracellular LDH activity (lower panel) were measured 48 h after ENO1-WT treatment. (B) The HIF1A, HK2, PFKFB3, GLUT1 and ENO1 mRNA levels were quantified by reverse transcription-quantitative PCR after 6 h of ENO1-WT treatment. (C) The HIF-1α, HK2, PFKFB3, GLUT1 and ENO1 protein levels were analyzed by immunoblotting after 24 h of ENO1-WT treatment. The amounts of studied proteins were first normalized with GAPDH, and the Rel was then calculated by comparing with the levels in the untreated cells, of which the value is set to 1.0. (D) Cell viability was measured using Cell Counting Kit-8. (E) Secretion of VEGF was measured by ELISA after 48 h of ENO1-WT treatment. (F) The enolase activity of ENO1-WT and two catalytically dead mutants, ENO1-S40A and ENO1-D245R, was measured. RPMI-8226 cells were treated with the indicated concentrations of ENO1-WT, ENO1-S40A and ENO1-D245R for 48 h. The (G) lactate and (H) VEGF concentrations in the culture medium were measured. All results are presented as the mean ± SD of three independent experiments. P-values were calculated with one-way ANOVA (with Tukey's post hoc test). ENO1, enolase-1; ENO1-WT, wild-type ENO1; GLUT1, glucose transporter 1; HIF1A or HIF-1α, hypoxia-inducible factor 1-α; HK2, hexokinase 2; LDH, lactate dehydrogenase; PFKFB3, 6-phosphofructo-2-kinase/fructose-2,6 biphosphatase 3; Rel., relative ratio; Ut, untreated; VEGF, vascular endothelial growth factor.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Recombinant, Concentration Assay, Activity Assay, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot, Cell Counting, Enzyme-linked Immunosorbent Assay

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: Extracellular ENO1 enhances glycolysis and pro-cancer activities through HIF-1α. RPMI-8226 cells were transfected with HIF-1α-targeting siRNA (si-HIF1A) or control siRNA (scramble sequence) for 96 h. (A) The HIF1A, HK2 and GLUT1 mRNA levels were quantified by reverse transcription-quantitative PCR after 6 h of treatment with or without 100 µg/ml ENO1-WT. (B) The HIF-1α, HK2, GLUT1 and IκBα protein levels and the (C) lactate, (D) IL-6 and (E) VEGF concentrations in the culture medium were measured 24 h after treatment with ENO1-WT. The amounts of studied proteins were first normalized with GAPDH, and then the Rel. was calculated by comparing with the scramble siRNA-treated cells, of which the value is set to 1.0. (F) Cell viability and (G) Transwell migration assays of RPMI-8226 cells with or without ENO1-WT treatment were performed following transfection with si-HIF1A or control siRNA. All results are presented as the mean ± SD of three independent experiments. P-values were calculated with one-way ANOVA (with Tukey's post hoc test). ENO1, enolase-1; ENO1-WT, wild-type ENO1; GLUT1, glucose transporter 1; HIF1A or HIF-1α, hypoxia-inducible factor 1-α; HK2, hexokinase 2; IL-6, interleukin 6; LDH, lactate dehydrogenase; Rel., relative ratio; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Transfection, Control, Sequencing, Reverse Transcription, Real-time Polymerase Chain Reaction, Migration, Small Interfering RNA

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: ENO1 mAb reduces glycolytic and pro-cancer activities. (A) The lactate concentrations in the culture medium, collected from ENO1 mAb-treated RPMI-8226 (left panel) and U266 (right panel) cells, were measured after 48 h of ENO1 mAb treatment. In all studies, hIgG1 at the indicated concentrations was included as a specificity control for ENO1 mAb. RPMI-8226 and U266 cells were treated with 100 µg/ml ENO1 mAb or 100 µg/ml hIgG1 for 48 h. (B) Enolase activity and (C) glucose uptake in lysates were further analyzed. Phloretin (a GLUT1 inhibitor) was added at 100 µM as a positive control to inhibit glucose uptake. (D) RPMI-8226 cells were treated with the indicated concentrations of ENO1 mAb or hIgG1 for 24 h. The HIF-1α, HK2, PFKFB3, GLUT1 and ENO1 protein levels were analyzed by immunoblotting. The amounts of studied proteins were first normalized with GAPDH, and then the Rel. was calculated by comparing with the untreated cells, of which the value was set to 1.0. (E) RPMI-8226 (upper panel) or U266 (lower panel) cells were treated with the indicated concentrations of ENO1 mAb or hIgG1. Cell viability was assessed 1, 2 or 3 days after treatment using the Cell Counting Kit-8. The OD 450 nm value was positively associated with the number of viable cells. (F) RPMI-8226 cells were treated with the indicated concentrations of ENO1 mAb or hIgG1. Cell migration was measured by Transwell assay after 18 h of treatment. (G) RPMI-8226 (upper panel) and U266 (lower panel) cells were treated with the indicated concentrations of ENO1 mAb or hIgG1 followed by measurement of the plasminogen receptor activity. (H) RPMI-8226 (left panels) or U266 (right panels) cells were treated with the indicated concentrations of ENO1 mAb or hIgG1 for 48 h. Supernatant was collected for determination of human VEGF (upper panel) and TGF-β (lower panel) levels by ELISA. All results are presented as the mean ± SD of three independent experiments. P-values were calculated with one-way ANOVA (with Tukey's post hoc test). ENO1, enolase-1; GLUT1, glucose transporter 1; HIF-1α, hypoxia-inducible factor 1-α; hIgG1, human IgG1; HK2, hexokinase 2; mAb, monoclonal antibody; PFKFB3, 6-phosphofructo-2-kinase/fructose-2,6 biphosphatase 3; Rel., relative ratio; TGF, transforming growth factor; Ut, untreated; VEGF, vascular endothelial growth factor.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Control, Activity Assay, Positive Control, Western Blot, Cell Counting, Migration, Transwell Assay, Enzyme-linked Immunosorbent Assay

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: ENO1 mAb reduces tumor growth and glycolysis in vivo in a RPMI-8226 subcutaneous xenograft model. Male nude mice were subcutaneously implanted with RPMI-8226 cells and randomized when the tumor size reached >100 mm 3 (n=6). ENO1 mAb (30 mg/kg) was intraperitoneally injected twice a week at the indicated time points. (A) Each data point represents the mean volume ± SD from the ENO1 mAb-treated, the withdrawing ENO1 mAb treated or vehicle control groups. Mice were sacrificed on day 35 and (B) representative images of excised tumors and the (C) tumor weight are shown. (D) Sera were collected for measurement of lactate concentration. (E) Mice body measurements were collected at the indicated time points. P-values were calculated with one-way ANOVA (with Tukey's post hoc test). ENO1, enolase-1; mAb, monoclonal antibody.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: In Vivo, Injection, Control, Concentration Assay

Journal: Oncology Reports

Article Title: Unrevealed roles of extracellular enolase‑1 (ENO1) in promoting glycolysis and pro‑cancer activities in multiple myeloma via hypoxia‑inducible factor 1α

doi: 10.3892/or.2023.8642

Figure Lengend Snippet: Schematic diagram summarizing the effects and possible mechanisms of extracellular ENO1 in regulating glycolysis and pro-cancer activities. Extracellular ENO1 may enhance glycolytic activity and glycolysis-related genes, including HK2 and GLUT1, through HIF-1α. Moreover, extracellular ENO1 also promoted HIF-1α-mediated pro-cancer activities, such as cell migration, cell viability and production of tumor-promoting cytokines. Consistently, these effects on cancer progression could be attenuated by the ENO1 antibody or ENO1 siRNA. ENO1, enolase-1; GLUT1, glucose transporter 1; HIF-1α, hypoxia-inducible factor 1-α; HK2, hexokinase 2; siRNA, small interfering RNA.

Article Snippet: The ENO1 gene was inserted into the expression vector, pLenti-C-Myc-DDK-P2A-Puro (cat. no. RC205494L3; OriGene Technologies, Inc.).

Techniques: Activity Assay, Migration, Small Interfering RNA

Journal: Oncology Letters

Article Title: α-enolase is highly expressed in liver cancer and promotes cancer cell invasion and metastasis

doi: 10.3892/ol.2020.12003

Figure Lengend Snippet: Comparison of the serum anti-ENO1 antibody levels among the three groups of participants [P50 (P25-P75)].

Article Snippet: The anti-ENO1 antibody ELISA detection reagent kit (cat. no. CSB-EQ027775HU) was purchased from CUSABIO Technology LLC.

Techniques: Comparison, Control