primary rabbit anti mouse il 6  (Abcam)

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: Establishment of mesenchymal stem cells (MSCs) stably expressing NIS regulated by the IL-6 promoter (A–C) Stimulation with cytokines IL-1β (0.5–5 ng/mL), IFN-γ (40–60 ng/mL), and TNF-α (1–15 ng/mL) showed NIS-specific 125 I-uptake that was reduced to background levels using perchlorate as NIS-specific inhibitor. (D) Treatment of IL-6-NIS-MSCs with IL-1β (1 ng/mL), IFN-γ (50 ng/mL), and TNF-α (10 ng/mL) revealed an increased radioiodide uptake as compared with single stimulation studies. (E) Stimulation with GL261-CM led to a significant increase of radioiodide uptake as compared with unstimulated IL-6-NIS-MSCs and was further increased by combined treatment with IL-1β (1 ng/mL), IFN-γ (50 ng/mL), and TNF-α (10 ng/mL). In contrast, stimulation with 4T1-CM was significantly lower as compared with stimulation with GL261-CM, but treatment with 4T1-CM and IL-1β (1 ng/mL), IFN-γ (50 ng/mL), and TNF-α (10 ng/mL) led to similar radioiodide uptake activity of IL-6-NIS-MSCs as treatment with GL261-CM and combined factors. Results are expressed as mean ± SEM of three independent experiments (two-tailed Student’s t test ∗p < 0.05, ∗∗p < 0.01; ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001). CM = conditioned medium.

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Stable Transfection, Expressing, Activity Assay, Two Tailed Test

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: Impact of GL261-CM on IL-6-NIS-MSC migratory behavior Chemotactic behavior of MSCs subjected to a gradient of GL261-CM was evaluated using 3D live-cell imaging migration assay over a period of 24 h. (A) Negative control is shown without the influence of a gradient on MSCs. (B) Migration of IL-6-NIS-MSCs along a gradient of GL261-CM resulted in strong migration toward the GL261-CM gradient. (C) Quantification of the chemotaxis parameters mean forward migration index (yFMI) and center of mass (yCoM, displayed by red dotes). Results are expressed as mean ± SEM from two independent experiments. Two-tailed Student’s t test was used for statistical analysis (∗p < 0.05; ∗∗p < 0.01). CM = conditioned medium.

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Live Cell Imaging, Migration, Negative Control, Chemotaxis Assay, Two Tailed Test

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: 18 F-TFB uptake of brain tumors is elevated after systemic IL-6-NIS-MSC delivery Representative images of 18 F-TFB-PET imaging (horizontal planes; SUV bw = 1.6) are shown 1 h after tracer injection of GBM-bearing mice after (A) single IL-6-NIS-MSC injection (n = 5), (B) two IL-6-NIS-MSC injections (n = 5), after (C) WT-MSC injection (n = 3), which served as a control, and (D) no MSC injection (n = 1). Brain area is encircled in red dotted lines and tumor is marked with a white arrow. (E) Tumoral radionuclide accumulation showed significantly higher levels after IL-6-NIS-MSC delivery as compared with WT-MSC control mice 1 h post 18 F-TFB injection. (F–I) Exemplary images of co-registration of anatomical MRI and PET imaging of the brain of the same mice as demonstrated in (A)–(D) showing tumor-selective tracer accumulation. A strong tumoral 18 F-TFB accumulation is revealed after systemic IL-6-targeted MSC-mediated NIS gene transfer (F–G), while tumors after WT-MSC injection did not show levels above background level (H) as well as compared with non-treated tumor (I). Tumors are encircled in yellow dotted lines. Results are expressed as % of the injected dose per mL (volume assessed by MRI) and mean ± SEM is given (two-tailed Student’s t test at each time point; ∗p < 0.05). sg = salivary glands.

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Imaging, Injection, Two Tailed Test

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: Ex vivo analysis of brain tumors and control organs show tumor-specific IL-6 and NIS protein expression (A, upper and lower panel) IL-6 protein expression was confirmed in GL261 tumors and throughout the tumor stroma (DAB-positive stain is shown in brown). (B) NIS immunofluorescence staining is shown with ×1 magnification to demonstrate tumor mass and the implantation site in the right caudate putamen. Tumor is encircled in yellow dotted lines. (C) No detectable NIS protein expression is observed in non-tumor brain tissue (close-up view: pink window in B). (D, upper and lower panel) Close-up view (indicated by the white window in B) shows NIS-specific immunoreactivity within the tumor stroma, with areas of high and low NIS protein expression, after systemic application of IL-6-NIS-MSCs, while tumors of mice that had received (E, upper and lower panel) WT-MSCs did not show detectable NIS expression. (F–I) No NIS protein expression was detected in control organs such as liver, lung, spleen, and kidney after IL-6-NIS-MSC administration. A representative image is shown in each.

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Ex Vivo, Expressing, Staining, Immunofluorescence

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: IL-6-targeted MSC-mediated NIS gene 131 I therapy of GBM-bearing mice led to reduced tumor growth and improved survival Three cycles of a single i.v. IL-6-NIS-MSC injection was applied followed by an i.p. administration of 131 I 48 h later (MSCs on days 1/5/9 and 131 I on days 3/7/11, respectively). Representative MR images 10 days after therapy start are shown after treatment with (A) IL-6-NIS-MSCs + 131 I, (B) IL-6-NIS-MSCs + NaCl, and (C) NaCl + NaCl. (D) Tumor growth was monitored using MRI showing a significantly reduced tumor mass of IL-6-NIS-MSCs + 131 I as compared with controls after completion of two therapy cycles (day 7 after treatment start, n = 7 each, ∗p < 0.05). At day 10 after therapy start, tumor growth of mice from the therapy group (n = 7/7) was delayed as compared with the NaCl-only group (n = 6/7, ∗p < 0.05) and the IL-6-NIS-MSC + NaCl-treated mice (n = 5/7, ns). Fourteen days after therapy start, all mice from the therapy schedule were included in the tumor measurement (n = 7/7), but n = 3 had to be euthanized the same day, while in the NaCl-only group only n = 1 of 7 and the IL-6-NIS-MSC + NaCl n = 2 of 7 of the mice were still alive and included in the measurement. (E) The survival of IL-6-NIS-MSCs + 131 I-treated mice was significantly extended as compared with the controls treated with NaCl + NaCl (∗∗∗p < 0.001) and IL-6-NIS-MSCs + NaCl (∗∗p < 0.01). Tumors are encircled in yellow dotted lines and results are expressed as mean ± SEM. One-way ANOVA using Tukey’s post hoc test was performed for tumor growth analysis and log rank test for comparison of survival curves.

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Injection, Comparison

Journal: Molecular Therapy Oncolytics

Article Title: Interleukin-6-controlled, mesenchymal stem cell-based sodium/iodide symporter gene therapy improves survival of glioblastoma-bearing mice

doi: 10.1016/j.omto.2023.08.004

Figure Lengend Snippet: Ex vivo analysis of brain tumors after IL-6 promoter-induced MSC-mediated NIS gene therapy At the end of the therapy study, frozen brain tissue sections derived from mice that were treated with IL-6-NIS-MSCs + 131 I (A), IL-6-NIS-MSCs + NaCl (B), and NaCl only (C) were subjected to Ki67 (proliferation index; green) and CD31 (blood vessels; red) immunofluorescence staining. Nuclei were counterstained with Hoechst (blue). A representative picture is shown in each at ×40 magnification (scale bar, 40 μm). Quantification of the proliferation index (D) and blood vessel density of IL-6-NIS-MSCs + 131 I in comparison with both control groups (E). Results are expressed as mean ± SEM (two-tailed Student’s t test ∗p < 0.05; ∗∗p < 0.01).

Article Snippet: Immunohistochemical staining for mouse IL-6 of FFPE brain sections was performed on a Bond RXm autostainer (Leica Biosystems, Nussloch, Germany) using primary rabbit anti-mouse IL-6 (ab290735, abcam, Cambridge, UK; 1:50 dilution) antibody and polymer refined detection kit (Leica Biosystems).

Techniques: Ex Vivo, Derivative Assay, Immunofluorescence, Staining, Comparison, Two Tailed Test