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. 2020 Sep 24;18:157. doi: 10.1186/s12964-020-00570-5

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© The Author(s) 2020

Open AccessThis 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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

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Fig. 4 — Attenuated MMP-9 expression in the infiltrating TAMs in tumor-bearing mice infected with Plasmodium parasites led to inhibition of tumor angiogenesis. a-b Tumor-bearing mice were sacrificed on day 17 after infection with Plasmodium parasites. TAMs were sorted, RNA was extracted, and the gene expression in TAMs was then detected through a gene expression (Roche NimbleGen) analysis. Genes with a ≥ 2-fold difference in expression and a q-value ≤0.05 were selected and mapped (a). A molecular annotation system was used to analyze the functions of these differentially expressed genes and the biological processes involved. Genes associated with tumor angiogenesis and that had a ≥ 2-fold difference in expression and a q-value ≤0.05 were selected for further analysis (b). c-f The relative MMP-9 and IGF-1 mRNA levels in the sorted TAMs (c) and tumor tissue (e) were quantified by qRT-PCR. A Western blotting analysis was performed to visualize the expression of MMP-9 and IGF-1 in the sorted TAMs (d, left panel) and tumor tissue (f, left panel). A quantitative analysis was performed using ImageJ software to quantify the endogenous MMP-9 and IGF-1 levels in the sorted TAMs (d, right panel) and tumor tissue (f, right panel). g A FACS-like tissue cytometer analysis system was used to analyze MMP-9 expression in the cells in the tumor margin by immunohistochemical staining with an antibody reactive to MMP-9 on day 17 after infection with Plasmodium parasites. The double-positive scatter plots represent the MMP-9-positive cells in scattergrams (left panel). The percentages represent the ratio of MMP-9-positive cells to total cells in the selected area. The mean intensity (MI) represents the average DAB intensity (right panel). h Representative images of MMP-9-positive cells identified by immunohistochemical staining with an antibody reactive to MMP-9 on day 17 after infection with Plasmodium parasites (original magnification: 400×). The insets show representative cells expressing MMP-9 (left panel) and the number of cells expressing MMP-9 (right panel). i A comprehensive analysis of the correlation among MMP-9 expression, MVD, and macrophage infiltration in the tumors of tumor-bearing mice on day 17 after Plasmodium infection compared with that in the tumors from uninfected tumor-bearing mice. p < 0.05 was considered significant