Journal: Biomarker Research

Article Title: Single-cell and spatial transcriptomics reveal a high glycolysis B cell and tumor-associated macrophages cluster correlated with poor prognosis and exhausted immune microenvironment in diffuse large B-cell lymphoma

doi: 10.1186/s40364-024-00605-w

Figure Lengend Snippet: Reagents and tools table

Article Snippet: FFPE samples were incubated with primary rabbit anti-human IgG antibodies specific to stathmin 1 (STMN1, ab52630, Abcam), enolase 1 (ENO1, ab227978, Abcam), peptidylprolyl isomerase A (PPIA, ab126738, Abcam), pyruvate kinase M (PKM, ab137791, Abcam), and cyclin dependent kinase 1 (CDK1, ab133327, Abcam) at dilutions of 1:1000, 1:1000, 1:50, 1:500, and 1:250. respectively, following blocking with rabbit serum.

Techniques: RNA Sequencing Assay, Sequencing, Expressing, Formalin-fixed Paraffin-Embedded, Immunohistochemistry, Microscopy, Immunofluorescence, Software

Journal: Biomarker Research

Article Title: Single-cell and spatial transcriptomics reveal a high glycolysis B cell and tumor-associated macrophages cluster correlated with poor prognosis and exhausted immune microenvironment in diffuse large B-cell lymphoma

doi: 10.1186/s40364-024-00605-w

Figure Lengend Snippet: Identifcation of glycolysis / gluconeogenesis maker genes in high malignant B cells. A Volcano plot of differential genes among the benign B cells, low malignant B cells, and high malignant B cells. B. Functional analysis of highly expressed genes in high malignant B cells by Metascape. C. Identification of eight commonly genes overlapped across three groups and glycolysis hallmark genes. D. Module trait correlation showed the relationships between modules, CNV score, and Glycolysis score. E. Network visualization of 10 modules of high maligant B cells.( The modules highlighted in red and underlined are modules associated with CNV score and Glycolysis score. ) F. The first 25 eigengenes of each module. G. Trajectory of different malignant B subclusters predicted by monocle. H. Genes expression level in single spot ordered along the pseudotime for MKI67 and seven glycolysis / gluconeogenesis gene markers ( STMN1, ENO1, LDHA, TPI1, CDK1, PKM , and PPIA ). ( Abbreviation : HMB: high malignant B cells; CNV: copy number variation; UMAP: uniform manifold approximation and projection. *** p < 0.001.)

Article Snippet: FFPE samples were incubated with primary rabbit anti-human IgG antibodies specific to stathmin 1 (STMN1, ab52630, Abcam), enolase 1 (ENO1, ab227978, Abcam), peptidylprolyl isomerase A (PPIA, ab126738, Abcam), pyruvate kinase M (PKM, ab137791, Abcam), and cyclin dependent kinase 1 (CDK1, ab133327, Abcam) at dilutions of 1:1000, 1:1000, 1:50, 1:500, and 1:250. respectively, following blocking with rabbit serum.

Techniques: Functional Assay, Expressing

Journal: Biomarker Research

Article Title: Single-cell and spatial transcriptomics reveal a high glycolysis B cell and tumor-associated macrophages cluster correlated with poor prognosis and exhausted immune microenvironment in diffuse large B-cell lymphoma

doi: 10.1186/s40364-024-00605-w

Figure Lengend Snippet: Prognostic value of four glycolysis / gluconeogenesis (STMN1, ENO1, CDK1, PKM, and PPIA) proteins in IHC cohort ( n = 34, 100X) and IFN_TAMs (CD68 + CXCL10 + PD-L1 + ) in mIF cohort ( n = 20, 10X). A-B. Kaplan–Meier curves of OS and PFS according to STMN1, CDK1, ENO1 and PKM proteins expression. C. Representative IHC staining of STMN1, CDK1, ENO1 and PKM in patient 1 (PFS = 7 months, OS = 9 months) and patient 2 (PFS = 135 months, OS = 135 months). D. Kaplan-Meier analysis for PFS based on IFN_TAMs intensity. E. Comparison of IFN_TAMs intensity in relapse and non_relapse groups. F. Correlation of IFN_TAMs’ intensity, CD8 + T cells’ intensity, and TGFβ1 intensity. G. Representative mIF staining of IFN_TAMs and CD8 + T cells in patient #1 (PFS = 2.7 months) and patient #2 (PFS = 90 months). ( Abbreviation : IHC: immunohistochemistry; IFN_TAMs: interferon-primed tumor-associated macrophages; mIF: multiple immunofluorescence; OS: overall survival; PFS: progression -free survival, Relapse: relapsed patients, patients without EFS24; Non_Relapse: non-relapsed patients, patients with EFS24. Mann-Whitney test was performed between groups.)

Article Snippet: FFPE samples were incubated with primary rabbit anti-human IgG antibodies specific to stathmin 1 (STMN1, ab52630, Abcam), enolase 1 (ENO1, ab227978, Abcam), peptidylprolyl isomerase A (PPIA, ab126738, Abcam), pyruvate kinase M (PKM, ab137791, Abcam), and cyclin dependent kinase 1 (CDK1, ab133327, Abcam) at dilutions of 1:1000, 1:1000, 1:50, 1:500, and 1:250. respectively, following blocking with rabbit serum.

Techniques: Expressing, Immunohistochemistry, Comparison, Staining, Immunofluorescence, MANN-WHITNEY

Journal: BMC Medicine

Article Title: Glucose competition between endothelial cells in the blood-spinal cord barrier and infiltrating regulatory T cells is linked to sleep restriction-induced hyperalgesia

doi: 10.1186/s12916-024-03413-z

Figure Lengend Snippet: Dysfunctional ECs in low-glucose intracellular microenvironment. A Protein levels of ZO-1, Occludin, Claudin-5, Glut1, Eno1 and LDHα detected by Western blot (left) and their quantitative analyses (right). n =3 per group. * p <0.05, ** p <0.01. B Tube formation by HUVECs in five random fields. n =5 per group. ** p <0.01. C Proliferative rate of HUVECs at different time points detected by CCK-8 assay. n =5 per group. *** p <0.001, **** p <0.0001. D The standard curve of glycolysis levels in HUVECs (left), optical density (middle) and the level of L-Lactate by conversion (right). n =3 per group. ** p <0.01

Article Snippet: The rabbit monoclonal antibodies used for Western blotting were anti-Glut1, anti-lactate dehydrogenase α (LDHα), anti-Enolase 1 (ENO1), anti-ZO-1, anti-Occludin, anti-Claudin-5 (all from Affinity Bioscience, Jiangsu, China), anti-phospho-mTOR, anti-phospho-p70S6K, anti-phospho-AMPK, anti-phospho-4E-BP1, and anti-β-actin antibodies (Cell Signaling Technology, BO, USA).

Techniques: Western Blot, CCK-8 Assay

Journal: BMC Medicine

Article Title: Glucose competition between endothelial cells in the blood-spinal cord barrier and infiltrating regulatory T cells is linked to sleep restriction-induced hyperalgesia

doi: 10.1186/s12916-024-03413-z

Figure Lengend Snippet: Infiltrating Tregs in the spinal cord influence functions and glycolysis levels of ECs. A The number of CD4 + CD25 + T cells in mice spinal cord (left) and the proportion on Day 1 (right). n =8 per group. * p <0.05. B Protein levels of ZO-1, Occludin, Claudin-5, Glut1, Eno1 and LDHα detected by Western blot (left) and their quantitative analyses (right). n =3 per group. * p <0.05, ** p <0.01, *** p <0.001, **** p <0.0001. C Tube formation by HUVECs in five random fields. n =5 per group. **** p <0.0001. D Proliferative rate of HUVECs at different time points detected by CCK-8 assay. n =5 per group. *** p <0.001, **** p <0.0001. E The standard curve of glycolysis levels in HUVECs (left), optical density (middle) and the level of L-Lactate by conversion (right). n =3 per group. **** p <0.0001

Article Snippet: The rabbit monoclonal antibodies used for Western blotting were anti-Glut1, anti-lactate dehydrogenase α (LDHα), anti-Enolase 1 (ENO1), anti-ZO-1, anti-Occludin, anti-Claudin-5 (all from Affinity Bioscience, Jiangsu, China), anti-phospho-mTOR, anti-phospho-p70S6K, anti-phospho-AMPK, anti-phospho-4E-BP1, and anti-β-actin antibodies (Cell Signaling Technology, BO, USA).

Techniques: Western Blot, CCK-8 Assay

Journal: BMC Medicine

Article Title: Glucose competition between endothelial cells in the blood-spinal cord barrier and infiltrating regulatory T cells is linked to sleep restriction-induced hyperalgesia

doi: 10.1186/s12916-024-03413-z

Figure Lengend Snippet: Lactate administration rescues functions of ECs and relieves SR-induced hyperalgesia. A Concentration of NaLA in mice spinal cord detected by colorimetry. n =3 per group. * p <0.05. B Time-dependent changes in PWMT after continuous 48-h SR (Day 0) and a week of lactate administration (Day 1, 2, 3, 5, 7, and 9) in mice (left) and those on the day of incision after SR (Day 0) and post-incision with lactate administration (Day 1, 3, 5, 7, and 9) (right). n =8 per group. SR + PBS vs. SR + NaLA: * p <0.05, ** p <0.01; SI + PBS vs. SI + NaLA: # p <0.05, ## p <0.01, ### p <0.001. C NaFlu distribution in mice taken by in vivo imaging system (left) and the quantitative analysis (right). n =3 per group. * p <0.05. D Tight junctions in BSCB after lactate administration visualized by TEM. Scale bar=2.0 μm. n =6 per group. E Protein levels of ZO-1, Occludin, Claudin-5, Glut1, Eno1 and LDHα detected by Western blot (left) and their quantitative analyses (right). Samples of SR + PBS and SR + NaLA groups were collected on Day 1 and those of SI + PBS and SI + NaLA groups were collected on Day 7. n =3 per group. * p <0.05, ** p <0.01, *** p <0.001, ns: no significance

Article Snippet: The rabbit monoclonal antibodies used for Western blotting were anti-Glut1, anti-lactate dehydrogenase α (LDHα), anti-Enolase 1 (ENO1), anti-ZO-1, anti-Occludin, anti-Claudin-5 (all from Affinity Bioscience, Jiangsu, China), anti-phospho-mTOR, anti-phospho-p70S6K, anti-phospho-AMPK, anti-phospho-4E-BP1, and anti-β-actin antibodies (Cell Signaling Technology, BO, USA).

Techniques: Concentration Assay, Colorimetric Assay, In Vivo Imaging, Western Blot

Journal: BMC Medicine

Article Title: Glucose competition between endothelial cells in the blood-spinal cord barrier and infiltrating regulatory T cells is linked to sleep restriction-induced hyperalgesia

doi: 10.1186/s12916-024-03413-z

Figure Lengend Snippet: Lactate administration rescues functions and glycolysis of HUVECs. A Proliferative rate of HUVECs at different time points detected by CCK-8 assay. n =5 per group. Control vs. Low glucose: *** p <0.001, **** p <0.0001; Low glucose vs. Low glucose + NaLA: ### p <0.001, #### p <0.0001. B Tube formation by HUVECs in five random fields. n =5 per group. **** p <0.0001. C Protein levels of ZO-1, Occludin, Claudin-5, Glut1, Eno1 and LDHα detected by Western blot (left) and their quantitative analyses (right). n =3 per group. * p <0.05, ** p <0.01, *** p <0.001

Article Snippet: The rabbit monoclonal antibodies used for Western blotting were anti-Glut1, anti-lactate dehydrogenase α (LDHα), anti-Enolase 1 (ENO1), anti-ZO-1, anti-Occludin, anti-Claudin-5 (all from Affinity Bioscience, Jiangsu, China), anti-phospho-mTOR, anti-phospho-p70S6K, anti-phospho-AMPK, anti-phospho-4E-BP1, and anti-β-actin antibodies (Cell Signaling Technology, BO, USA).

Techniques: CCK-8 Assay, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Steady Laminar Flow Decreases Endothelial Glycolytic Flux While Enhancing Proteoglycan Synthesis and Antioxidant Pathways

doi: 10.3390/ijms25052485

Figure Lengend Snippet: Glycolytic enzyme expression and protein levels did not change in HUVECs exposed to steady laminar versus oscillating disturbed flow. Relative ( A ) mRNA levels measured via RT-PCR, and ( B ) protein levels measured via Western blot of glycolytic enzymes in HUVEC exposed to static culture (S), steady laminar flow (L), or oscillating disturbed flow (D). GLUT1: glucose transporter 1, GLUT3: glucose transporter 3, HK1: hexokinase 1, HK2: hexokinase 2, GPI: glucose-6-phosphate isomerase, PFK-1: phosphofructokinase-1, PFKFB3: 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3, ALDO-A: aldolase A, ALDO-C: aldolase C, TPI-1: Triosephosphate isomerase-1, GAPDH: Glyceraldehyde 3-phosphate dehydrogenase, PGK-1: phosphoglycerate kinase-1, PGK-2: phosphoglycerate kinase-2, PGM-1: phosphoglucomutase-1, PGM-5: phosphoglucomutase-5, ENO-1: enolase-1, ENO-3: enolase-3, LDH-A: lactate dehydrogenase A, LDH-B: lactate dehydrogenase B, PKM-1: pyruvate kinase-1, PFKM-2: pyruvate kinase-2. n = 3 samples per condition.

Article Snippet: After blocking in 5% BSA, membranes were incubated with primary antibodies for hexokinase 1 (46695), hexokinase 2 (374091), phosphofructokinase-1 (166722), aldolase A (390733), phosphoglycerate kinase (48342), phosphoglucomutase-1 (373796), lactate dehydrogenase (133123), glucose-6-phosphate dehydrogenase (373886), and enolase-1 (100812, all from Santa Cruz), as well as PFKFB3 (13123S) and GFAT1 (5322, from Cell Signaling, Danvers, MA, USA) at a 1:1000 dilution overnight at 4 °C followed by the appropriate secondary horseradish peroxidase-conjugated antibody (1:2000; Promega, Madison, WI, USA) for 1 h at room temperature. β-actin (SC47778-C4, Santa Cruz, Dallas, TX, USA) was used as the loading control.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: Steady Laminar Flow Decreases Endothelial Glycolytic Flux While Enhancing Proteoglycan Synthesis and Antioxidant Pathways

doi: 10.3390/ijms25052485

Figure Lengend Snippet: PCR primer sequences.

Article Snippet: After blocking in 5% BSA, membranes were incubated with primary antibodies for hexokinase 1 (46695), hexokinase 2 (374091), phosphofructokinase-1 (166722), aldolase A (390733), phosphoglycerate kinase (48342), phosphoglucomutase-1 (373796), lactate dehydrogenase (133123), glucose-6-phosphate dehydrogenase (373886), and enolase-1 (100812, all from Santa Cruz), as well as PFKFB3 (13123S) and GFAT1 (5322, from Cell Signaling, Danvers, MA, USA) at a 1:1000 dilution overnight at 4 °C followed by the appropriate secondary horseradish peroxidase-conjugated antibody (1:2000; Promega, Madison, WI, USA) for 1 h at room temperature. β-actin (SC47778-C4, Santa Cruz, Dallas, TX, USA) was used as the loading control.

Techniques: Sequencing