Journal: International Journal of Nanomedicine

Article Title: Sequential Release of mRNA Complex and T Cells by a Double-Layered Implantable Scaffold for Combination Therapy of Head and Neck Squamous Cell Carcinoma

doi: 10.2147/IJN.S582078

Figure Lengend Snippet: Characteristics of DMP nanoparticles and the proliferation-inhibiting effect of the DMP-mBim complex in vitro. ( A ) Size distribution and zeta potential of DMP nanoparticles. ( B ) Transmission electron microscopy (TEM) photomicrographs of DMP stained with phosphotungstic acid solution (scale bar: 50 nm). ( C ) Cell viability MTT assay of DMP and PEI25K in 293T cells. ( D ) Hemolysis test of DMP nanoparticles, Triton X-100 was used as positive control, and normal saline was used as negative control (***P<0.001, ****P<0.0001). ( E ) Critical micelle concentration evaluation of DMP using pyrene as the fluorescent dye. ( F ) Murine Bim mRNA (mBim) obtained by the in vitro transcription method was detected by electrophoresis. ( G ) Gel-retardation assay of mRNA and DMP at different weight ratios. ( H ) RNase protection electrophoresis assay of the DMP-mRNA complex. ( I ) Transfection efficiency of DMP-EGFP in SCC-VII cells (scale bar: 100 μm). ( J ) Bim levels in SCC-VII cells after DMP-mBim complex transfection (***P<0.001). ( K ) MTT assay of SCC-VII cells after DMP-mBim transfection and representative images. The cell viability and inhibition rate were also calculated (scale bar: 100 μm, ****P<0.0001). ( L ) Proliferation-inhibition ability of the DMP-mBim complex on SCC-VII cells evaluated by clonogenic assay (****P<0.0001). ( M ) Apoptosis-inducing ability of DMP-mBim complex in SCC-VII cells using Annexin V/PI staining by flow cytometry (****P<0.0001). ( N ) Live/dead staining evaluation of SCC-VII cells treated with the DMP-mBim complex (scale bar: 100 μm, ****P<0.0001).

Article Snippet: Cells were dual-stained for the plasma membrane and tubulin using DiI (Cell Plasma Membrane Staining Kit; Beyotime, China) and Tubulin-Tracker Green (Live Cell Staining Kit; Beyotime, China).

Techniques: In Vitro, Zeta Potential Analyzer, Transmission Assay, Electron Microscopy, Staining, MTT Assay, Positive Control, Saline, Negative Control, Concentration Assay, Electrophoresis, Electrophoretic Mobility Shift Assay, Transfection, Inhibition, Clonogenic Assay, Flow Cytometry

Journal: International Journal of Nanomedicine

Article Title: Sequential Release of mRNA Complex and T Cells by a Double-Layered Implantable Scaffold for Combination Therapy of Head and Neck Squamous Cell Carcinoma

doi: 10.2147/IJN.S582078

Figure Lengend Snippet: Transfection efficiency and treatment effect of DMP-mBim complex after release from scaffold. ( A and B ) Representative fluorescence images of SCC-VII cells transfected with supernatants of the outer layer of the scaffold containing the DMP-EGFP complex in short-term release (scale bar: 50 μm) and detection of efficiency by flow cytometry. ( C and D ) Representative fluorescence images of SCC-VII cells transfected with supernatants of the outer layer of the scaffold containing the DMP-EGFP complex in long-term release (scale bar: 100 μm) and the calculated transfection efficiency. ( E ) Cumulative RNA-release rate in the outer layer scaffold. ( F ) Bim levels in SCC-VII cells after treatment with supernatants of the outer layer of the scaffold containing DMP-mBim complex (***P<0.001). ( G and H ) MTT assay of SCC-VII cells after treatment with the supernatants of the outer layer of the scaffold containing DMP-mBim complex (scale bar: 100 μm, ****P<0.0001, **P<0.01). ( I ) Live/dead staining evaluation of SCC-VII cells treated with the supernatants of the outer layer of the scaffold (scale bar: 100 μm, ****P<0.0001).

Article Snippet: Cells were dual-stained for the plasma membrane and tubulin using DiI (Cell Plasma Membrane Staining Kit; Beyotime, China) and Tubulin-Tracker Green (Live Cell Staining Kit; Beyotime, China).

Techniques: Transfection, Fluorescence, Flow Cytometry, MTT Assay, Staining