Fig. 1

Accumulation and incorporation of vessels and connective tissue at the tumor surface. A, B Subcutaneously growing C38 tumor. The accumulation of collagen I (red)- and smooth muscle actin (SMA)-expressing myofibroblasts (blue) is present at the tumor surface (arrows). Black areas represent viable tumor mass (T) beneath the tumor surface. For clarity, B shows the blue (SMA) channel separately. Numerous vessels (CD31, green) are being incorporated (together with connective tissue) by the tumor (large arrowheads). Smaller connective tissue columns separated from the incorporated connective tissue are visible within the tumor tissue (small arrowheads). N necrosis. C, D C38 tumor (T) growing in the liver. In the left upper part of c, the peritumoral liver parenchyma—with the dense CD31-positive network of sinusoids (green)—is visible. The tumor-parenchyma interface is indicated by arrows. Here, accumulation of SMA-positive cells can be observed (see also D, for the separate blue channel). These cells are also present in the invaginations (large arrowheads) and in the incorporated host connective tissue deep within the tumor (small arrowheads). The invaginations and the incorporated host tissue pieces are delineated by the laminin (red) deposited by the tumor cells. Inset: high-power micrograph of the surface of a C38 liver metastasis, demonstrating the accumulation of SMA-positive cells (blue). Tumor tissue is present on the left side (black area). On the right side, pan-cytokeratin-expressing hepatocytes (green) and CD31-positive vessels (red) are visible. Between the hepatocytes and the tumor tissue, SMA-positive cells (blue) can be observed. E, F A C38 brain metastasis (E) is highlighted and bordered by laminin (red), which is deposited by tumor cells. CD31-positive vessels (green) appear yellowish because of their close proximity of the basement membrane (red) (E). No SMA accumulation is present at the periphery of the tumor, as can be seen on the separate blue (SMA) channel (F). The peritumoral vessels are smaller than the intratumoral vessels, which are surrounded by SMA-positive pericytes (blue). SMA-positive cells are not found around the peritumoral vessels, excluding the arterioles. N necrosis, T tumor tissue