Journal: Rsc Advances

Article Title: Detecting hypoxia in vitro using 18 F-pretargeted IEDDA “click” chemistry in live cells †

doi: 10.1039/d1ra02482e

Figure Lengend Snippet: The in vitro uptake of [ 18 F]FB-Tz in (A) EMT6 and (B) HCT116 cells. Cells were treated with 8 (0.01–10 μM) and cultured under normoxia or hypoxia (0.1% O 2 ) conditions for 24 h prior to radioactive uptake experiment: 0.74 MBq per well of [ 18 F]FB-Tz was added to cells and incubated for 60 min. (C) Time course of [ 18 F]FB-Tz uptake in EMT6 cells in vitro . Cells were treated with 8 (10 μM) for 24 h under hypoxia before incubation with 0.74 MBq per well of [ 18 F]FB-Tz for 5, 10, 15, 30, 45 and 60 min. (D) Radioactive uptake of [ 18 F]FMISO (1.48 MBq per well) for 0 min (pulse) or 3 h under normoxia or hypoxia conditions in EMT6 and HCT116. Data were expressed as a percentage of total radioactivity incorporated into cells, normalized for total cellular protein. Mean values ± SD ( n = 6; * P < 0.05, ** P < 0.01 and **** P < 0.0001).

Article Snippet: EMT6 murine breast cancer cells were a kind gift from Prof. Sofia Pascu (University of Bath, UK) and were grown in Waymouth medium (Thermo Fisher Scientific); HCT116 human colorectal cancer cells (ATCC) were grown in Dulbecco Modified Eagle Medium (DMEM, Sigma-Aldrich).

Techniques: In Vitro, Cell Culture, Incubation, Radioactivity

Journal: Journal of Advanced Research

Article Title: Innate tumor-targeted nanozyme overcoming tumor hypoxia for cancer theranostic use

doi: 10.1016/j.jare.2021.02.004

Figure Lengend Snippet: In vitro study. (A) T1-weighted MRI in vitro with various MnO 2 concentrations under pH 6.0 and pH 7.4. (B) Concentration-dependent longitudinal relaxation time T1, based on readings from A. (C) Cytotoxicity of HFn carrier toward HeLa, 4 T1, and SKOV3 cancer cells. (D-F) Cell viability of three cell lines under hypoxic conditions determined by MTT assay exposed to various concentrations of nanomaterials for 48 h. The Dox concentration axis represents the quantity of Dox in each group for Dox contained formulations as well as the MnO 2 concentration axis for MnO 2 contained nanomaterials. (G) IC 50 values of various cancer cells based on Dox concentration (*p < 0.05, **p < 0.01 vs SKOV3). Data are presented as means ± SD (n = 3).

Article Snippet: The human cervical cancer cells (HeLa), human ovarian cancer cells (SKOV3) and 4 T1 murine breast cancer cells (4 T1) were originally obtained from the American Type Culture Collection (ATCC) and cultured with the recommended medium at 37 °C within 5% CO 2 atmosphere.

Techniques: In Vitro, Concentration Assay, MTT Assay

Journal: Journal of Advanced Research

Article Title: Innate tumor-targeted nanozyme overcoming tumor hypoxia for cancer theranostic use

doi: 10.1016/j.jare.2021.02.004

Figure Lengend Snippet: T1-MR images of 4 T1 tumor-bearing mice through (A) local and (C) intravenous injection of MnO 2 -Dox@HFn. (B) and (D) Quantification of T1-MRI signals via intratumorally and intravenously injection according to A and C, respectively (n = 3; **p < 0.01, ***p < 0.001 vs before). (E) Tumor growth in various treated groups (n = 3). (F) Tumor weight at the endpoint of the study (**p < 0.01). (G) Representative tumor images were harvested from mice after the treatment. (H) Tumor growth inhibition rate.

Article Snippet: The human cervical cancer cells (HeLa), human ovarian cancer cells (SKOV3) and 4 T1 murine breast cancer cells (4 T1) were originally obtained from the American Type Culture Collection (ATCC) and cultured with the recommended medium at 37 °C within 5% CO 2 atmosphere.

Techniques: Injection, Inhibition

Journal: The Journal of biological chemistry

Article Title: Formation of STAT5-containing DNA binding complexes in response to colony-stimulating factor-1 and platelet-derived growth factor.

doi: 10.1074/jbc.271.31.18350

Figure Lengend Snippet: FIG. 1. SIF-A complex contains STAT3 and an anti-STAT5A reactive protein. A, BAC1.2F5 cells were cultured in the absence of CSF-1 for 16 h. The cells were then incubated for 5 min in the presence of 10,000 units/ml recombinant human CSF-1 prior to the preparation of protein extracts. The protein extracts were used for electrophoretic mobility shift assays with the m67 high affinity SIE probe. Where indicated, the protein extract was pretreated for 15 min with antibodies to STAT1, STAT3, STAT5A, or STAT5B prior to the addition of the probe. B, FDwtfms cells, after incubation for 16 h in the absence of IL-3, were incubated with 10,000 units/ml CSF-1 prior to the preparation of the protein extracts. The assay was performed as in A. C, Same as A but using extracts of T56 cells after incubation with CSF-1. D, Same as C but using extracts of T56 cells after incubation with PDGF.

Article Snippet: Three different anti-STAT5 antibodies were used for these experiments, i.e. anti-STAT5 (Transduction Laboratories); anti-STAT5A, raised against a C-terminal peptide, DQDGEFDLDESMDVA, of the murine STAT5A protein, which does not cross-react with STAT5B or STAT1; and anti-STAT5B (Santa Cruz Laboratories), raised against a C-terminal peptide, DVARRVEELLGRPMDSQ, of the murine STAT5B protein.

Techniques: Cell Culture, Incubation, Recombinant, Electrophoretic Mobility Shift Assay

Journal: The Journal of biological chemistry

Article Title: Formation of STAT5-containing DNA binding complexes in response to colony-stimulating factor-1 and platelet-derived growth factor.

doi: 10.1074/jbc.271.31.18350

Figure Lengend Snippet: FIG. 2. The anti-STAT5A antibody does not cross-react with STAT3. Proteins of T56 cells were immunoprecipitated with antibodies to STAT3 or STAT5A, and the Western blot was probed first with anti-STAT5 and then reprobed with anti-STAT3.

Article Snippet: Three different anti-STAT5 antibodies were used for these experiments, i.e. anti-STAT5 (Transduction Laboratories); anti-STAT5A, raised against a C-terminal peptide, DQDGEFDLDESMDVA, of the murine STAT5A protein, which does not cross-react with STAT5B or STAT1; and anti-STAT5B (Santa Cruz Laboratories), raised against a C-terminal peptide, DVARRVEELLGRPMDSQ, of the murine STAT5B protein.

Techniques: Immunoprecipitation, Western Blot

Journal: The Journal of biological chemistry

Article Title: Formation of STAT5-containing DNA binding complexes in response to colony-stimulating factor-1 and platelet-derived growth factor.

doi: 10.1074/jbc.271.31.18350

Figure Lengend Snippet: FIG. 3. CSF-1- and PDGF-activated PIE-binding complexes containing STAT5. A, protein extracts of T56 cells, incubated with CSF-1, were used for elec- trophoretic mobility shift assays with the PIE probe. B, same as in A but using extracts of T56 cells after incubation with PDGF. C, same as in A but using extracts of FDwtfms cells. D, same as in A but using proteins of BAC1.2F5 cells after treatment with CSF-1. Where indicated, the protein extracts were pretreated for 15 min with antibodies to STAT1, STAT3, STAT5A, or STAT5B.

Article Snippet: Three different anti-STAT5 antibodies were used for these experiments, i.e. anti-STAT5 (Transduction Laboratories); anti-STAT5A, raised against a C-terminal peptide, DQDGEFDLDESMDVA, of the murine STAT5A protein, which does not cross-react with STAT5B or STAT1; and anti-STAT5B (Santa Cruz Laboratories), raised against a C-terminal peptide, DVARRVEELLGRPMDSQ, of the murine STAT5B protein.

Techniques: Binding Assay, Incubation, Mobility Shift

Journal: The Journal of biological chemistry

Article Title: Formation of STAT5-containing DNA binding complexes in response to colony-stimulating factor-1 and platelet-derived growth factor.

doi: 10.1074/jbc.271.31.18350

Figure Lengend Snippet: FIG. 4. IL-3-activated STAT5A and STAT5B of FDwtfms cells formed distinct PIE-binding complexes. FDwtfms cells were cul- tured for 16 h in the absence of IL-3, followed by incubation with IL-3 for 10 min. Protein extracts from these cells were used for electro- phoretic mobility shift assays using the PIE probe. Where indicated, the extracts were pretreated with antibodies to STAT3, STAT5A, or STAT5B.

Article Snippet: Three different anti-STAT5 antibodies were used for these experiments, i.e. anti-STAT5 (Transduction Laboratories); anti-STAT5A, raised against a C-terminal peptide, DQDGEFDLDESMDVA, of the murine STAT5A protein, which does not cross-react with STAT5B or STAT1; and anti-STAT5B (Santa Cruz Laboratories), raised against a C-terminal peptide, DVARRVEELLGRPMDSQ, of the murine STAT5B protein.

Techniques: Binding Assay, Incubation, Mobility Shift