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. 2016 Oct;14(10):953-965.
doi: 10.1158/1541-7786.MCR-16-0153. Epub 2016 Aug 17.

High-Fat Diet-Induced Complement Activation Mediates Intestinal Inflammation and Neoplasia, Independent of Obesity

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Free PMC article

High-Fat Diet-Induced Complement Activation Mediates Intestinal Inflammation and Neoplasia, Independent of Obesity

Stephanie K Doerner et al. Mol Cancer Res. .
Free PMC article

Abstract

Obesity and related metabolic disturbances are closely associated with pathologies that represent a significant burden to global health. Epidemiological and molecular evidence links obesity and metabolic status with inflammation and increased risk of cancer. Here, using a mouse model of intestinal neoplasia and strains that are susceptible or resistant to diet-induced obesity, it is demonstrated that high-fat diet-induced inflammation, rather than obesity or metabolic status, is associated with increased intestinal neoplasia. The complement fragment C5a acts as the trigger for inflammation and intestinal tumorigenesis. High-fat diet induces complement activation and generation of C5a, which in turn induces the production of proinflammatory cytokines and expression of proto-oncogenes. Pharmacological and genetic targeting of the C5a receptor reduced both inflammation and intestinal polyposis, suggesting the use of complement inhibitors for preventing diet-induced neoplasia.

Implications: This study characterizes the relations between diet and metabolic conditions on risk for a common cancer and identifies complement activation as a novel target for cancer prevention. Mol Cancer Res; 14(10); 953-65. ©2016 AACR.

Conflict of interest statement

Competing financial interests: S.K.D., N.A.B., J.D.L. and J.H.N. are co-inventors in a patent application titled “A method to treat polyp formation”.

Figures

Figure 1
Figure 1. Diet and genetic background determine obesity and insulin resistance
(A) Study design for diet studies. Male mice were maintained on a stock 5010 (grey) diet until 30 days of age (d.o.a.) before being randomly assigned to a HFD (purple) or LFD (blue). Mice remained on a diet study for 3 days (33 d.o.a.), 30 days (60 d.o.a.) or 60 days (90 d.o.a.). Wild-type B6.Apc+/+, A2.Apc+/+, A9.Apc+/+, A7.Apc+/+, and A17.Apc+/+ strains and the equivalent ApcMin/+ (B6.ApcMin/+, A2.ApcMin/+, A9.ApcMin/+, A7.ApcMin/+, and A17.ApcMin/+) strains were fed the LFDCoco or HFDCoco for 60 days. (B) Body weight curves from B6.Apc+/+ and B6.ApcMin/+. Final body weight (C) and, HOMA-IR (D) were measured at Day 60 of treatment. (EF) The total number of polyps (E) and polyp mass (F) were measured in the intestine of each mouse at Day 60 of treatment. Values represent means ± s.e.m. (n=5–54 males/group). *P<0.05, **P<0.01, ***P<0.001 in relation to the corresponding LFD-fed group. #####P<0.001 in relation to B6.ApcMin/+ fed HFD.
Figure 2
Figure 2. Diet, but not obesity or metabolic status, promotes intestinal tumorigenesis
B6.ApcMin/+ mice were fed the LFD or HFD with coconut (purple), corn (green), or olive (blue) oil as a source of fat for 60 days. The final body weight (A), HOMA-IR (B), total number of polyps (C) and polyp mass (D) were assessed after Day 60 of diet treatment. Values represent means ± s.e.m. (n=7–54 males/group). *P<0.05, **P<0.01, ***P<0.001 in relation to equivalent LFD-fed group. ###P<0.001 in relation to HFDCoco- or HFDCorn-fed groups.
Figure 3
Figure 3. Intestinal neoplasia is associated with diet-induced inflammation
B6.Apc+/+ and B6.ApcMin/+ mice were fed the LFDCoco or HFDCoco for 30 days. Levels of anti- and pro-inflammatory mediators implicated in metabolic syndrome and cancer were measured in the blood (AF), adipose tissue (G, H), and intestinal tissue (IL) at day 30 of treatment. Values represent means ± s.e.m. (n=5–7 males/group). *P<0.05, **P<0.01, ***P<0.001 in relation to the same strain fed the LFDCoco. ##P<0.01 in relation to B6.Apc+/+ fed HFDCoco.
Figure 4
Figure 4. Inflammation and intestinal neoplasia are triggered soon after exposure to HFD- the 3 days study
B6.Apc+/+ and B6.ApcMin/+ mice were fed the LFDCoco or HFDCoco for 3 days. Final body weight (A) and HOMA-IR (B) were measured at Day 3 of treatment. Intestinal polyp number (C) and mass (D) were analyzed. Circulating levels of C5a (E) and adiponectin (F) were measured in plasma. Levels of anti- and pro-inflammatory mediators implicated in inflammation and cancer- C5a (G), IL-6 (H), VEGF (I), IL-23 (J), TNF-α (K), and IL-10 (L) were measured in the intestine of B6.Apc+/+ or B6.ApcMin/+ at Day 3 of treatment. Values represent means ± s.e.m. (A–F, n=6–9 males/group; G–L, n=3–5). *P<0.05, **P<0.01, ***P<0.001 in relation to the corresponding LFD-fed group.
Figure 5
Figure 5. Source of dietary fat determines local pro-inflammatory environment and intestinal polyposis
B6.Apc+/+ and B6.ApcMin/+ mice were fed diets supplemented with coconut (purple), corn (green) or olive (blue) oil for 30 days. Levels of anti- and pro-inflammatory mediators implicated in inflammation and cancer such as VEGF (A), TNF-α (B), IL-23 (C), IL-1β (D) and IL-10 (E) were measured in intestinal tissue at day 30 of treatment. Values represent means ± s.e.m. (n=3–5 males/group). *P<0.05, **P<0.01 in relation to the same strain fed the LFDCoco. #P<0.05, ##P<0.01 in relation to the same strain fed the corresponding LFDCoco or HFDCoco.
Figure 6
Figure 6. Complement C5a mediates diet-induced inflammation and intestinal neoplasia
B6.Apc+/+ and B6.ApcMin/+ mice were fed the LFD or HFD with coconut (purple), corn (green), or olive oil (blue) as a source of fat for 30 days. Circulating levels of the complement activation fragment C5a were measured in the plasma at Day 30 of treatment (A). (BF) B6.ApcMin/+ (+/+; Min/+) and the complement-deficient C5aR+/−; ApcMin/+ (C5aR+/−; Min/+), and C5aR−/−; ApcMin/+ (C5aR−/−; Min/+) strains were fed HFDCoco or LFDCoco for 30 days. The total number of polyps (B) and polyp mass (C) were measured in the intestine of each mouse at Day 30 of treatment. (n=5–20/group). Intestinal inflammation was measured in mice genetically deficient in C5aR (DF). Protein levels of VEGF (D), TNFα (E), and IL-23 (F) were measured in the intestine (n=3–5/group) B6.Apc+/+ and B6.ApcMin/+ mice were fed HFDCoco for 30 days, and concomitantly treated with the C5aR antagonist peptide (PMX53) or a scrambled inactive peptide (control) (2 mg/kg s.c.) every other day (B, C). (D, E) Intestinal inflammation was measured in males treated with PMX53 or control inhibitor (n=3–5). Total polyp number (B) and polyp mass (C) were measured. Protein levels of VEGF (G), TNF-α (H), and IL-23 (I) and expression of IL-1β (J) and Myc (K) were measured in the intestine of males treated with PMX53 or the control inhibitor (n=3–6/group). (L) The percentage of viable CD11b+/GR-1+ neutrophils was measured with flow cytometry in the intestinal lamina propria of B6.Apc+/+ and B6.ApcMin/+ males treated with PMX53 or the control peptide (n=4/group). Levels of phosphorylated AKT (pS473), IKK (pS176) and NFκB (pS536) were assessed with Western blotting (M) in polyp tissue of B6.ApcMin/+ males or normal intestinal tissue of B6.Apc+/+ males treated with PMX53 or the control peptide. Levels of AKT and NFκB were measured as a control for the protein amount loaded. Results are representative of 3 independent experiments. *P<0.05, **P<0.01, ***P<0.001 compared to same strain fed corresponding LFD. #P<0.05, ##P<0.01, ###P<0.001 compared to B6.Apc+/+ or B6.ApcMin/+ fed the same diet. $P<0.05, $$P<0.01, $$$P<0.001 compared to same strain treated with the control inhibitor.
Figure 7
Figure 7. Molecular mechanisms determining HFD-induced intestinal polyp progression
Diets high in specific dietary fats induce an inflammatory environment in the intestine that promotes intestinal tumorigenesis in genetically susceptible APCmin/+mice, independent of obesity and related metabolic conditions. HFD containing coconut or corn oil as a source of saturated fat induce complement activation with subsequent release of C5a. C5a in turn induces the infiltration of inflammatory neutrophils to the intestinal lamina propria and activate the AKT/IKK/NFκB signaling pathway in intestinal cells with consequent production of pro-inflammatory mediators such as IL-1β, IL-6, TNF-α and the proto-oncogene c-myc. Conversely, mice fed a HFD containing olive oil as a source of fat gained significant body weight, but did not promote a pro-inflammatory environment or development of polyps in the intestine.

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