Journal: Journal of nanobiotechnology

Article Title: ALPL regulates pro-angiogenic capacity of mesenchymal stem cells through ATP-P2X7 axis controlled exosomes secretion.

doi: 10.1186/s12951-024-02396-6

Figure Lengend Snippet: Fig. 5 ALPL mutation increases hBMMSCs exosomes secretion through ATP axis. (A) HPP hBMMSCs was treated with 2 U/mL ATP-apyrase (HPP + Apy) and the exosomes markers were analyzed. Exosomes proteins from equal volumes of culture supernatant of Nor, HPP and HPP + Apy hBMMSCs were loaded for Western Blot. (B) Exosomes volumes derived from each groups were detected by exosomes ELISA complete kit. (C-D) The intracellular expression of CD9 and CD81 were analyzed by Western Blot. (E) Immunofluorescent staining showed CD9 and CD81-positive labeled exosomal proteins localized in Nor, HPP and HPP + Apy hBMMSCs. Scale bar, 10 μm. (F) Nor hBMMSCs was treated with 10µmol/L ATP and the exosomes markers were analyzed. Exo somal proteins from equal volumes of culture supernatant of Nor and Nor + ATP hBMMSCs were loaded for Western Blot. (G) Exosomes volumes derived from Nor and Nor + ATP groups were detected by exosomes ELSA complete kit. (H-I) The intracellular expression of CD9 and CD81 were analyzed by Western Blot. (J) CD9 and CD81-positive labeled exosomal proteins localized in Nor and Nor + ATP hBMMSCs, as assessed by immunofluorescent staining. Scale bar, 10 μm. All results were generated in three independent experiments. Data were shown as mean ± standard deviation (SD); *P < 0.05; **P < 0.01; ***P < 0.001

Article Snippet: The secreted VEGF, PDGFBB, Angiostatin and Endostatin in the samples were quantified using human VEGF ELISA Kit (Proteintech, KE00216, China), human PDGFBB ELISA Kit (Proteintech, KE00161, China), human Angiostatin ELISA kit (Neobioscience Technology, E-EL-H6165, China) and human Endostatin ELISA Kit (Proteintech, KE00259, China) according to manufacture’s instruction.

Techniques: Mutagenesis, Western Blot, Derivative Assay, Enzyme-linked Immunosorbent Assay, Expressing, Staining, Labeling, Generated, Standard Deviation

Journal: Oncology Letters

Article Title: Endostatin reverses immunosuppression of the tumor microenvironment in lung carcinoma

doi: 10.3892/ol.2017.7455

Figure Lengend Snippet: Antitumor and anti-vascular effect of Endostatin on established LLC tumors. (A and B) The tumors were dissected and the tumor growth curves are shown. (C and D) The microvessel density (MVD) is determined by blinded measurement of CD31 expression. The data shown are representative results of independent experiments in the figure, n=7 per group for each experiment. *P<0.05; **P<0.01.

Article Snippet: Antibodies and reagents Recombinant human endostatin (rhEndostatin; Simcere Pharm, Nanjing, China); EZ-SepTM mouse percollase (Amresco, Dallas, TX, USA); RPMI-1640 medium, FBS (Gibco, Grand Island, NY, USA); ConA, DMEM medium, phosphate-buffered saline (PBS) buffer (Sigma, St. Louis, MO, USA); fluorescently-labeled antibody CD3, CD4, CD8, CD11c, CD86, major histocompatibility complex (MHC) II, CD11b, Gr-1, CD206, CD68 and NOS2 and their isotype controls (eBioscience, Inc., San Diego, CA, USA); mouse lymphocyte factor ELISA kit (Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China); BCA Protein Assay kit (Beyotime, Shanghai, China); anti-mouse hypoxia-inducible factor-1α (HIF-1α), VEGF antibody (Abcam, Cambridge, MA, USA); rmGM-CSF, rmIL-4 (Peprotech), rmIL-2, rmTNF-α (Biological); anti-mouse CD31 nonlabeled immunohistochemical monoclonal antibody (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA); MCO-15AC CO2 incubator (Sanyo, Osaka, Japan); sterile 1.5 laminar flow bechtop (Thermo Fisher Scientific); FACSCalibur flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA); NanoDrop ND-1000 ultraviolet spectrophotometer (Agilent Technologies, Inc., Santa Clara, CA, USA); Model 680 Microplate Reader (Bio-Rad, Hercules, CA, USA).

Techniques: Expressing

Journal: Oncology Letters

Article Title: Endostatin reverses immunosuppression of the tumor microenvironment in lung carcinoma

doi: 10.3892/ol.2017.7455

Figure Lengend Snippet: Endostatin efficiently descreased multiple proangiogenic factors, and augmented hypoxia in the tumors. (A and B) The VEGF and HIF-1α expressions in the tumors were showed. The expression of β-actin served as an internal control. (C) The expression of VEGF and HIF-1α in the tumors. (D) Expression of the pro/anti-antigiogenic cytokines IL-6, IL-17 and IFN-γ. Data are expressed as the mean ± SD of three independent experiments. *P<0.05.

Article Snippet: Antibodies and reagents Recombinant human endostatin (rhEndostatin; Simcere Pharm, Nanjing, China); EZ-SepTM mouse percollase (Amresco, Dallas, TX, USA); RPMI-1640 medium, FBS (Gibco, Grand Island, NY, USA); ConA, DMEM medium, phosphate-buffered saline (PBS) buffer (Sigma, St. Louis, MO, USA); fluorescently-labeled antibody CD3, CD4, CD8, CD11c, CD86, major histocompatibility complex (MHC) II, CD11b, Gr-1, CD206, CD68 and NOS2 and their isotype controls (eBioscience, Inc., San Diego, CA, USA); mouse lymphocyte factor ELISA kit (Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China); BCA Protein Assay kit (Beyotime, Shanghai, China); anti-mouse hypoxia-inducible factor-1α (HIF-1α), VEGF antibody (Abcam, Cambridge, MA, USA); rmGM-CSF, rmIL-4 (Peprotech), rmIL-2, rmTNF-α (Biological); anti-mouse CD31 nonlabeled immunohistochemical monoclonal antibody (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA); MCO-15AC CO2 incubator (Sanyo, Osaka, Japan); sterile 1.5 laminar flow bechtop (Thermo Fisher Scientific); FACSCalibur flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA); NanoDrop ND-1000 ultraviolet spectrophotometer (Agilent Technologies, Inc., Santa Clara, CA, USA); Model 680 Microplate Reader (Bio-Rad, Hercules, CA, USA).

Techniques: Expressing, Control

Journal: Oncology Letters

Article Title: Endostatin reverses immunosuppression of the tumor microenvironment in lung carcinoma

doi: 10.3892/ol.2017.7455

Figure Lengend Snippet: Endostatin discreased inhibitory immune cells in the tumor microenvironment and increased tumor-infiltrated mDCs and CD8+T cells. (A) The immunosuppressive MDSC and TAM are significantly lower. (B) The mDCs are upregulated, and cytotoxic CD8+T cells are recruited to infiltrate the tumors by endostatin. Data expressed as the mean ± SD are representative of three independent experiments, using tumor pooled from seven animals per group. *P<0.05.

Article Snippet: Antibodies and reagents Recombinant human endostatin (rhEndostatin; Simcere Pharm, Nanjing, China); EZ-SepTM mouse percollase (Amresco, Dallas, TX, USA); RPMI-1640 medium, FBS (Gibco, Grand Island, NY, USA); ConA, DMEM medium, phosphate-buffered saline (PBS) buffer (Sigma, St. Louis, MO, USA); fluorescently-labeled antibody CD3, CD4, CD8, CD11c, CD86, major histocompatibility complex (MHC) II, CD11b, Gr-1, CD206, CD68 and NOS2 and their isotype controls (eBioscience, Inc., San Diego, CA, USA); mouse lymphocyte factor ELISA kit (Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China); BCA Protein Assay kit (Beyotime, Shanghai, China); anti-mouse hypoxia-inducible factor-1α (HIF-1α), VEGF antibody (Abcam, Cambridge, MA, USA); rmGM-CSF, rmIL-4 (Peprotech), rmIL-2, rmTNF-α (Biological); anti-mouse CD31 nonlabeled immunohistochemical monoclonal antibody (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA); MCO-15AC CO2 incubator (Sanyo, Osaka, Japan); sterile 1.5 laminar flow bechtop (Thermo Fisher Scientific); FACSCalibur flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA); NanoDrop ND-1000 ultraviolet spectrophotometer (Agilent Technologies, Inc., Santa Clara, CA, USA); Model 680 Microplate Reader (Bio-Rad, Hercules, CA, USA).

Techniques:

Journal: Oncology Letters

Article Title: Endostatin reverses immunosuppression of the tumor microenvironment in lung carcinoma

doi: 10.3892/ol.2017.7455

Figure Lengend Snippet: Endostatin decreased immune inhibitory cytokines in the tumor microvironment. Endostatin significantly reduced the expression of IL6, IL-10, TGF-β and IL-17 in the tumor tissue, and concurrently elevated IFN-γ in comparison with control group. Data expressed as the mean ± SD are representative of three independent experiments, using tumor pooled from seven animals per group. *P<0.05.

Article Snippet: Antibodies and reagents Recombinant human endostatin (rhEndostatin; Simcere Pharm, Nanjing, China); EZ-SepTM mouse percollase (Amresco, Dallas, TX, USA); RPMI-1640 medium, FBS (Gibco, Grand Island, NY, USA); ConA, DMEM medium, phosphate-buffered saline (PBS) buffer (Sigma, St. Louis, MO, USA); fluorescently-labeled antibody CD3, CD4, CD8, CD11c, CD86, major histocompatibility complex (MHC) II, CD11b, Gr-1, CD206, CD68 and NOS2 and their isotype controls (eBioscience, Inc., San Diego, CA, USA); mouse lymphocyte factor ELISA kit (Shanghai Enzyme-linked Biotechnology Co., Ltd., Shanghai, China); BCA Protein Assay kit (Beyotime, Shanghai, China); anti-mouse hypoxia-inducible factor-1α (HIF-1α), VEGF antibody (Abcam, Cambridge, MA, USA); rmGM-CSF, rmIL-4 (Peprotech), rmIL-2, rmTNF-α (Biological); anti-mouse CD31 nonlabeled immunohistochemical monoclonal antibody (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA); MCO-15AC CO2 incubator (Sanyo, Osaka, Japan); sterile 1.5 laminar flow bechtop (Thermo Fisher Scientific); FACSCalibur flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA); NanoDrop ND-1000 ultraviolet spectrophotometer (Agilent Technologies, Inc., Santa Clara, CA, USA); Model 680 Microplate Reader (Bio-Rad, Hercules, CA, USA).

Techniques: Expressing, Comparison, Control