Sinusoidal tumor angiogenesis is a key component in hepatocellular carcinoma metastasis

Abstract

Hepatocellular carcinoma (HCC) has a tendency for intravascular dissemination leading to a poor prognosis. The importance of the sinusoidal structure of the tumor vasculature in HCC has been implicated in the metastasis formation. To clarify the role of tumor angiogenesis in HCC metastasis, we morphologically investigated the interaction of HCC cells with blood vessels during the sequential process of metastasis. Autopsy specimens of 80 patients with HCC were examined with immunohistochemistry using a specific antibody against CD31, a marker for endothelial cells. The most frequent sites of metastasis were the liver (82.5%) and lung (43.8%). In most cases, the metastatic process was initiated by vascular involvement where tumor nests surrounded by sinusoidal vessels extend into the portal and hepatic veins. Subsequently, these endothelial-coated tumor emboli enter the circulation, embolize at distant organs, proliferate within the blood vessel and ultimately form metastatic foci. These steps are indicative of an invasion-independent pathway. Our findings in animal models and now in human cases suggest that sinusoidal angiogenesis may represent a novel target for therapeutic strategies to limit HCC metastasis. In combination with primary tumor treatment, perturbation of tumor emboli may reduce dissemination of disease.

Figures

Fig. 1

Sinusoidal vascular architecture and cancer cell intravasation in primary liver tumor. (a) Basic structure of HCC forming thick trabecular pattern (hematoxylin-eosin stain, original magnification 100×). Immunostaining of CD31 (b) and laminin (c) shows sinusoidal tumor vasculature with basement membrane surrounding the tumor nests (original magnification, panel b, 100×; panel c, 400×). (d, e) Intravasation of HCC. (d) Multiple tumor nests move into an efferent vein (EV) from the primary tumor (PT) without destruction of the vascular wall (hematoxylin-eosin stain, original magnification 100×). (e) The tumor emboli conserve their trabecular architecture with endothelial coverage during the intravasation process (immunostaining of CD31, original magnification 100×)

Fig. 2

Circulating HCC cells. (a, b) A free embolus in the main trunk of the pulmonary artery. (a) The tumor embolus within blood coagulation is composed of multiple tumor nests. (b) The embolus keeps their tissue organization with sinusoidal tumor vasculature. (c, d) Tumor emboli in the esophageal varix. Well-organized tumor nests with endothelial lining are embolized in dilated veins. [(a, c) Hematoxylin eosin stain, original magnification 40×, (b, d) immunostaining of CD31, original magnification 100×]

Fig. 3

Pulmonary tumor emboli of HCC. (a) Large tumor mass composed of multiple tumor nests, and with central necrosis, are embolized in a pulmonary artery (hematoxylineosin stain, original magnification 20×). (b) A single tumor nest is arrested in an arteriole (hematoxylin-eosin stain, original magnification 200×). (c, d) Immunostaining of CD31 highlights tumor-associated blood vessels surrounding tumor emboli both in the pulmonary artery (c) and arterioles (d) (original magnification (c) 100×; (d) 200×). (e) The embolus conserves the trabecular architecture with the basement membrane (Immunostaining of laminin, original magnification 200×). (f) Intravascular growth and extravasation of HCC cells embolized in pulmonary artery (Elastica Masson stain, original magnification 20×)

Fig. 4

Lymphatic dissemination of HCC. Immunostaining with D2-40 antibody shows multicellular tumor emboli covered by lymphatic endothelial cells in a lymph vessel (a) and the marginal sinus in a lymph node (b) (original magnification 200×)