Fig. 3

“Processing” of the invaginations. A A C38 tumor growing in the cecal wall. Incorporated connective tissue, which contains myofibroblasts (SMA, blue) and numerous vessels (CD31, green) (arrows) is visible throughout the tumor mass. All blue areas represent the columns, excluding necrotic areas (marked by N). Necrotic tissue is also highlighted by the nonspecific binding of the anti-mouse secondary antibody (used to detect the mouse monoclonal primary antibody against SMA) to these areas. Note the high number of advanced-stage connective tissue columns that contain only one single vessel (arrowheads). The columns are delineated by laminin (red) containing basement membrane produced by the tumor (T). B C38 metastasis in the liver. The early-stage incorporations contain numerous vessels (CD31, green) (arrows). Advanced-stage connective tissue columns, which contain single central vessels, are present intratumorally (arrowheads). Laminin (red) deposited by the tumor cells borders the incorporated tissue and the columns. The inset shows the fine structure of a longitudinally sectioned column containing a single central vessel (CD31, green). The basement membrane of the vessel is marked by small arrowheads. The basement membrane marked by small arrows is deposited by the tumor (T) at the surface of the column. C Subcutaneous C38 tumor. In addition to columns filled with myofibroblasts (SMA, blue) (small arrowheads) and containing a single central vessel (CD31, green), there are areas containing connective tissue columns with numerous vessels (arrows). Note the columns (large arrowheads) separated only partially from a larger connective tissue mass during the final steps of the maturation process. The incorporated connective tissue and the columns are bordered by laminin (red) deposited by the tumor cells. N marks areas of necrotic tissue that also appear blue, although this staining is due to the nonspecific binding of the anti-mouse secondary antibody used to detect the anti-SMA antibody. D C38 lung metastasis. The section is labeled with anti-CD31 (green), anti-laminin (red) and anti-SMA antibodies (blue). There are no SMA-positive cells (blue) in the alveolar walls of the normal lung tissue far from the metastasis (right edge of the micrograph). SMA-positive cells begin to appear in the peritumoral lung tissue (small arrowheads), but the number of these cells only notably increases intratumorally. Large early-stage connective tissue columns are present at the tumor periphery (large arrows), from which smaller tissue pieces containing different numbers of vessels are detaching (small arrows). Large arrowheads indicate mature columns with single vessels. N marks areas of necrotic tissue that also appear blue, although this staining is due to the nonspecific binding of the anti-mouse secondary antibody used to detect the anti-SMA antibody