integrin α v hdr plasmid  (Santa Cruz Biotechnology)

Journal: Nature Communications

Article Title: Integrin-α V -mediated activation of TGF-β regulates anti-tumour CD8 T cell immunity and response to PD-1 blockade

doi: 10.1038/s41467-021-25322-y

Figure Lengend Snippet: a Representative IHC images of tumour samples from patients with low and high α V expression in tumour cells. Objective: 20×. b Kaplan−Meier curve of OS for stage I treatment-naïve lung cancer patients according to the α V expression by IHC analysis of FFPE tumours. c Kaplan−Meier curve of PFS of PD-1 blockade-treated patients with tumours harbouring low and high expression of α V integrin. d Percentages of anti-PD-(L)1-treated patients displaying α V high tumours among long-responders (LR: PFS > 6 months and OS > 12 months) or fast progressors (FP: defined by “early death” occurring within 12 weeks of treatment initiation). e Representative digital mark-up image of fluorescent IHC of CD8 (green), cytokeratin (turquoise), and dapi (blue) staining in α V low and α V high tumour sections. d = CD8 + cell density. Left, the density of CD8 + TIL in α V low and α V high tumours. The numbers of tumours in each group are indicated (* p = 0.046). Scale bar, 2 cm. f Representative digital mark-up image of CD8 + CD103 neg (green), CD8 + CD103 + (orange), CD8 - CD103 + (red), cytokeratin (turquoise) and dapi (blue) staining in α V low and α V high tumour sections. d = CD8 + CD103 + cell density. Left, the density of CD8 + CD103 + (* p = 0.016) and CD8 + CD103 neg ( p = 0.120) cells in tumour regions of α V low and α V high tumours. Scale bar, 2 cm. Each symbol represents an individual cell type from tumour samples; horizontal lines correspond to mean ± standard error of the mean (SEM) ( e , f ). Data were calculated with the log-rank test ( b , c ) and Welch’s two-sided t -test ( e , f ). Source data are provided as a Source Data file.

Article Snippet: The cells were double transfected with integrin α V CRISPR-Cas9 KO plasmid (Santa Cruz Biotechnology, sc-400506) and integrin α V HDR plasmid (Santa Cruz Biotechnology, sc-400506-HDR).

Techniques: Expressing, Staining

Journal: Nature Communications

Article Title: Integrin-α V -mediated activation of TGF-β regulates anti-tumour CD8 T cell immunity and response to PD-1 blockade

doi: 10.1038/s41467-021-25322-y

Figure Lengend Snippet: a Representative flow cytometry plots (bi-exponential scale) of α V expression in EpCAM + E-cadherin + and EpCAM neg E-cadherin neg cells from a lung tumour. Right, percentage of α V expression in EpCAM + E-cadherin + and EpCAM neg E-cadherin neg cells ( n = 18, *** p = 0.0002). b Representative flow cytometry plots of β 6 subunit expression in EpCAM + E-cadherin + α V + and EpCAM neg E-cadherin neg α V + cells from a tumour sample. Right, expression of β 6 integrin in EpCAM + E-cadherin + α V + and EpCAM neg E-cadherin neg α V + cells ( n = 16), * p = 0.013. c Surface expression of α V , β 6 , and β 8 subunits in the IGR-B2 cell line. d Concentration of total TGF-β in CM from IGR-B2, IGR-B2T, and IGR-B2T-KO cells measured by ELISA (*** p = 0.0004). Results are presented as mean ± SEM of six independent experiments. Right, relative luciferase activity in the Mu.1LV cell line transfected with (CAGA)9-Lux reporter plasmid and treated with CM from IGR-B2, IGR-B2T, and IGR-B2T-KO cells, normalized to luciferase activity in Mu.1LV cell treated with CM from IGR-B2. Results are presented as mean ± SEM of six independent experiments (* p = 0.011, **** p < 0.0001). e Expression of α V integrin on IGR-B2T and IGR-B2T-KO cells. An isotype control was included. f Representative photos of the morphology of IGR-B2T and IGR-B2T-KO cells by phase-contrast light microscope from one experiment out of five. Objective: 20×. Each symbol represents the individual cell type from tumour samples ( a , b ); horizontal lines correspond to mean ± SEM ( a , b , d ). Data were calculated with paired Student t -tests ( a , b ) and one-way ANOVA with Tukey’s correction ( d ). ns: non-significant. Source data are provided as a Source Data file.

Article Snippet: The cells were double transfected with integrin α V CRISPR-Cas9 KO plasmid (Santa Cruz Biotechnology, sc-400506) and integrin α V HDR plasmid (Santa Cruz Biotechnology, sc-400506-HDR).

Techniques: Flow Cytometry, Expressing, Concentration Assay, Enzyme-linked Immunosorbent Assay, Luciferase, Activity Assay, Transfection, Plasmid Preparation, Control, Light Microscopy