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. 2020 Aug 7;10:13343. doi: 10.1038/s41598-020-70231-7

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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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AdIL-17A transduction in 4T1 tumor induces potent immune suppression due to the changes in the quantity of suppressive myeloid cells. Splenic T cells were isolated from the spleen of DO11.10 mouse, labelled with proliferation dye eFluor 670, and co-cultured with bulk peripheral blood leukocytes (PBL) as the course of MDSCs from naïve mice or Ad-transduced tumor-bearing mice or sort-purified myeloid subpopulations. OVA-specific CD4 T cell proliferation in response to OVA peptide stimulation under different culture conditions was measured by flow cytometry based on eFluor 670 dilution. (A) Representative histograms of the CD4⁺ T cell proliferation in the presence of PBL recovered from 40 µL of peripheral blood; no peptide stimulation is shown in grey. (B) CD4 T cell proliferation in the presence of PBL recovered from 40, 20, and 10 µL of blood collected 14 days after the inoculation of AdIL-17A- or Addl-transduced 4T1 tumor cells (n = 6 per group). (C) CD4 T cell proliferation in the presence of graded doses of sort-purified granulocytic cells (CD11b⁺Ly6C^intLy6G^hi) or monocytic cells (CD11b⁺Ly6C^hiLy6G^lo) isolated at day 14 from the spleens of mice inoculated with AdIL-17A- or Addl-transduced tumor cells. *P < 0.05; ***P < 0.001 compared to Addl using unpaired Student’s t-test.