The role of innate immunity in the protection conferred by a bacterial infection against cancer: study of an invertebrate model

Abstract

All multicellular organisms are exposed to a diversity of infectious agents and to the emergence and proliferation of malignant cells. The protection conferred by some infections against cancer has been recently linked to the production of acquired immunity effectors such as antibodies. However, the evolution of innate immunity as a mechanism to prevent cancer and how it is jeopardized by infections remain poorly investigated. Here, we explored this question by performing experimental infections in two genetically modified invertebrate models (Drosophila melanogaster) that develop invasive or non-invasive neoplastic brain tumors. After quantifying tumor size and antimicrobial peptide gene expression, we found that Drosophila larvae infected with a naturally occurring bacterium had smaller tumors compared to controls and to fungus-infected larvae. This was associated with the upregulation of genes encoding two antimicrobial peptides-diptericin and drosomycin-that are known to be important mediators of tumor cell death. We further confirmed that tumor regression upon infection was associated with an increase in tumor cell death. Thus, our study suggests that infection could have a protective role through the production of antimicrobial peptides that increase tumor cell death. Finally, our study highlights the need to understand the role of innate immune effectors in the complex interactions between infections and cancer cell communities in order to develop innovative cancer treatment strategies.

Figure 1

Tumor visualization in vivo in (A) non-cancerous and (B) cancerous larvae characterized by GFP-labeled cells (in white) in the eye-antennal disc region. (C) Fold induction of the three immune genes in cancerous larvae (yellow; n = 10 pools) without infection relative to non-cancerous larvae (dark blue; n = 10 pools). The error bars represent standard deviation (SD) of two independent experiments (***p < 0.001; **p < 0.01; *p < 0.05; Sidak’s test). Effect of infection by the bacterium Pectobacterium carotovorum carotovorum (Pcc, red; n = 96 total) and the fungi Beauvaria bassiana (Bb, orange; n = 102 total) compared to uninfected flies (blue; n = 97 total) on individual tumor size with a low intensity threshold of detection (S2 threshold: 75% of maximum intensity value) in three independent experiments. The error bars represent standard deviation (***p < 0.001; **p < 0.01; *p < 0.05; NS > 0.05; Tukey’s test). (E) Impact of body size, approximated by mouth hook size, on tumor size depending on infectious treatment (n = 20 by infection group; error bars are SD).

Figure 2

(A) mRNA levels of expression for the three AMP genes, dpt, drs, and upd3, in cancerous larvae in response to infection by the bacterium Pcc (red; n = 15 pools) and the fungi Bb (orange; n = 14 pools) compared to uninfected larvae (Ctl, blue; n = 13 pools). Data shown here were pooled from five independent experiments (***p < 0.001; **p < 0.01; *p < 0.05; Kruskal-Wallis test). Gene expression was assessed by qRT PCR with the primers described in Table S2. (B) mRNA levels of expression for the three AMP genes (dpt, drs, upd3) in non-cancerous larvae following infections (n = 10 pools by infection group). The error bars represent standard deviation and data were pooled from two independent experiments (****p < 0.0001; ***p < 0.001; **<0.01; *p < 0.05; Kruskal-Wallis test).

Figure 3

(A,B) Representative immunofluorescence images of the tissues quantified in both conditions, stained with DAPI (white) to measure tumor volume. Wing discs are highlighted by the yellow dotted lines. Scale bars = 50 mm. (C) Quantification of wing disc tumor volumes from uninfected dlg mutant larvae (Uninfected, n = 33) or from dlg mutant larvae infected with Pcc (n = 28). (D,E) Representative immunofluorescence images of the quantified tissues that have been stained with the anti-Dcp1 antibody (red) to measure tumor cell death. F) Quantification of wing disc tumor cell death  in dlg mutant larvae uninfected (Uninfected, n = 33) or infected with Pcc (n = 28). Results are presented as a pool of two independent replicates (****p < 0.0001; Student’s t-test).