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. 2020 Sep 23;11:4810. doi: 10.1038/s41467-020-18444-2

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a Specific cytotoxicity of primary CD147-CAR-T cells was measured by FFLuc reporter assays, compared to κ-CAR-T cells (control group). Data represent the mean ± SEM (n = 6). b, c Significantly decreased cytotoxicity of CD147-CAR-T cells using knockout-CD147 FFLuc-GFP-SK-Hep1 cell line and HepG2 cell line by FFLuc reporter assays. Data represent the mean ± SEM from three independent experiments. d Cytotoxicity of primary CD147-CAR-NK cells was measured by the 4-h standard ⁵¹Cr release assays, compared to κ-CAR-T cells (control group). Data represent the mean ± SEM (n = 6). e, f Significantly decreased cytotoxicity of primary CD147-CAR-NK cells using knockout-CD147 FFLuc-GFP-SK-Hep1 cell line and HepG2 cell line by the 4-h standard ⁵¹Cr release assays. Data represent the mean ± SEM from three independent experiments. g Anti-NKG2D antibody blocks primary CD147-CAR-NK naturally killing to FFLuc-GFP-SK-Hep1. Data represent the mean ± SEM from three independent experiments. h Cytotoxicity of CD147-CAR-NK-92MI to the SK-Hep1 was measured by a standard 4-h ⁵¹Cr release assay. Data represent the mean ± SEM (n = 6). i, j FFLuc reporter system assay for specific killing of FFLuc-GFP-SK-Hep1 and FFLuc-GFP-HepG2 cell lines by CD147-CAR-NK-92MI. Effector cells (CD147-CAR-NK-92MI and NK-92MI) were cocultured with 1 × 10⁴ FFLuc-GFP-SK-Hep1 (i) or FFLuc-GFP-HepG2 (j) target cells per well in a 96-well optical-bottom microplate for 6 h. The control groups (blue) used were wild-type NK-92MI incubated with CD147⁺ FFLuc-GFP-SK-Hep1 or CD147⁺ FFLuc-GFP-HepG2. Data represent the mean ± SEM (n = 6). k, l Decreased cytotoxicity of CD147-CAR-NK-92MI cells using knockout-CD147 FFLuc-GFP-SK-Hep1 (k) and knockout-CD147-FFLuc-GFP-HepG2 (l) cell lines by FFLuc reporter system assay. Data represent the mean ± SEM (n = 3, 6 for SK-Hep1 and HepG2 groups, respectively). m, n Anti-CD147 (clone, HIM6) inhibits the CD147-CAR-NK-92MI-specific lysis effect against and FFLuc-GFP-SK-Hep1 (m) and FFLuc-GFP-HepG2 (n). Data represent the mean ± SEM from three independent experiments. Two-tailed unpaired Student’s t test was employed for all the panels. ***p < 0.001; n.s. not significant.