Figure 4: Circulating and recombinant sST2 suppresses the malignant growth of LuM1 tumours.

(a) Injected CRC cell pairs (group A-F) and schematic procedure of the injection of tumour cells. BALB/c mice were subcutaneously inoculated with sST2 high-expressing cells (NM11 or LuM1-sST2) or low-expressing cells (LuM1 or NM11-shsST2) into the left flank (L) on Day 0, followed by the injection of LuM1 cells into the right flank (R) on Day 5. The mice (n=6 mice per group) were killed on Day 30, and the blood, tumours and lungs were collected. (b) sST2 serum levels. (c) Tumour volume. (d) Number of lung metastatic nodules. (e) Vessel density in the tumours examined by CD31 staining (n=15 fields). The data are shown as the mean±s.d. (f) Protocol used for the tumour inoculation and hydrodynamic injection of the sST2-Fc-expressing plasmid (n=6 mice per group). The mice were killed and the tumours and lungs were removed on Day 28. (g) Monitoring of sST2 serum concentrations by ELISA. Serum samples were collected at the time points indicated in (f). The dashed line indicates the mean value of sST2 serum concentration in non-tumour-bearing age-matched BALB/c mice (17.9 ng ml⁻¹) (n=5). (h) Tumour growth. Arrows denote the plasmid injection time points. (i) Pulmonary metastasis. Scale bar: 1 cm. (j) CD31 staining of cryostat sections of tumours (left) and tumour vessel density (right) (n=15 fields). Scale bar: 100 μm. (k) Protocol used for intratumoural (i.t.) administration of the sST2-Fc protein (100 μg) into NM11-shsST2 tumours (n=7 mice per group). (l) Tumour growth. Arrows denote the protein injection time points. (m) Pulmonary metastasis. (n) Angiogenesis. CD31 staining of cryostat sections of tumours (left) and tumour vessel density (right) (n=39∼40 fields). Scale bar: 100 μm. The data are shown as the mean±s.d. *P<0.05, **P<0.01, ***P<0.001 by Student’s t-test.