IL-1 is required for tumor invasiveness and angiogenesis

Abstract

Here, we describe that microenvironmental IL-1 beta and, to a lesser extent, IL-1 alpha are required for in vivo angiogenesis and invasiveness of different tumor cells. In IL-1 beta knockout (KO) mice, local tumor or lung metastases of B16 melanoma cells were not observed compared with WT mice. Angiogenesis was assessed by the recruitment of blood vessel networks into Matrigel plugs containing B16 melanoma cells; vascularization of the plugs was present in WT mice, but was absent in IL-1 beta KO mice. The addition of exogenous IL-1 into B16-containing Matrigel plugs in IL-1 beta KO mice partially restored the angiogenic response. Moreover, the incorporation of IL-1 receptor antagonist to B16-containing plugs in WT mice inhibited the ingrowth of blood vessel networks into Matrigel plugs. In IL-1 alpha KO mice, local tumor development and induction of an angiogenic response in Matrigel plugs was less pronounced than in WT mice, but significantly higher than in IL-1 beta KO mice. These effects of host-derived IL-1 alpha and IL-1 beta were not restricted to the melanoma model, but were also observed in DA/3 mammary and prostate cancer cell models. In addition to the in vivo findings, IL-1 contributed to the production of vascular endothelial cell growth factor and tumor necrosis factor in cocultures of peritoneal macrophages and tumor cells. Host-derived IL-1 seems to control tumor angiogenesis and invasiveness. Furthermore, the anti-angiogenic effects of IL-1 receptor antagonist, shown here, suggest a possible therapeutic role in cancer, in addition to its current use in rheumatoid arthritis.

Figure 1

Increased survival rate and decreased tumor growth in IL-1β KO mice. (A) Survival after i.v. injection of B16 melanoma into IL-1β KO (n = 10) compared with WT mice (n = 10). (B) Frequency of lung micrometastases as detected in histological sections on day 14 after tumor cell injection (n = 10). (C) Tumor size of B16 melanoma after i.f.p. injection (2 × 10⁵ cells per mouse; n = 5 per group). Results shown are from one experiment of six performed.

Figure 2

Blood vessel growth in B16 cells embedded in Matrigel plugs. (Upper) Section through a B16-embedded Matrigel plug injected s.c. in WT mice and removed on day 7 (×400). (Inset) Gross morphology of Matrigel plugs. (Lower) Similarly stained sections through Matrigel plugs injected into IL-1β KO mice. Results shown are from one of four experiments performed.

Figure 3

MVD in B16 melanoma embedded in Matrigel plugs. (A) MVD ± SEM in B16 melanoma-embedded Matrigel plugs in WT or IL-1β KO mice on day 7. IL-1β KO mice were also implanted with Matrigel plugs embedded with a mixture of B16 cells and 100 ng of murine IL-1α per 0.4 ml of Matrigel (n = 5 in each group). (B) Day 7 MVD of Matrigel plugs embedded into WT mice containing a mixture of B16 melanoma cells plus saline, 50 or 100 μg of recombinant human IL-1Ra per 0.4 ml of Matrigel (n = 5 in each group).

Figure 4

Growth and MVD of B16 melanoma cells in WT, IL-1α, and IL-1β KO mice. (A) Tumor growth in mice injected i.f.p. with B16 cells, as indicated in legend to Fig. 1C. The results represent one of three experiments. Mean ± SEM for each (n = 8 per group) are shown. (B) MVD ± SEM in B16 melanoma-embedded Matrigel plugs (2 × 10⁵) in WT, IL-1β, or IL-1α KO mice (n = 5 in each group).

Figure 5

Growth and MVD of DA/3 mammary cancer cells in WT BALB/c, IL-1α, or IL-1β KO mice. (A) Mice were injected i.f.p. with DA/3 cells (5 × 10⁵ cells per mouse). The results represent one of three experiments. Mean ± SEM from each group (n = 8 per group) are shown. (B) MVD ± SEM in DA/3-embedded Matrigel plugs (2 × 10⁵) in WT, IL-1β, or IL-1α KO mice (n = 5 in each group).

Figure 6

VEGF secretion in cocultures of B16 melanoma cells and macrophages from WT, IL-1α, or IL-1β KO mice. PEC were obtained and cocultured with B16 melanoma cells, as indicated in Materials and Methods. Mean ± SEM VEGF levels in 48-h supernatants were measured (n = 5 experiments).

Figure 7

TNFα secretion in cocultures of B16 melanoma cells and macrophages from WT and IL-1β KO mice. PEC were obtained and stimulated with melanoma cells in the presence or absence of IL-1Ra (10 μg/ml). Mean ± SEM TNFα levels in 48-h supernatants are shown (n = 5 experiments).