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High tumor interstitial fluid pressure (TIFP) is a characteristic of most solid tumors. TIFP may hamper adequate uptake of macromolecular therapeutics in tumor tissue. In addition, TIFP generates mechanical forces affecting the tumor cortex, which might influence the growth parameters of tumor cells. This seems likely as, in other tissues (namely, blood vessels or the skin), mechanical stretch is known to trigger proliferation. Therefore, we hypothesize that TIFP-induced stretch modulates proliferation-associated parameters. Solid epithelial tumors (A431 and A549) were grown in Naval Medical Research Institute nude mice, generating a TIFP of about 10 mm Hg (A431) or 5 mm Hg (A549). Tumor drainage of the central cystic area led to a rapid decline of TIFP, together with visible relaxation of the tumor cortex. It was found by sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot analysis that TIFP lowering yields a decreased phosphorylation of proliferation-associated p44/42 mitogen-activated protein kinase and tumor relaxation. In confirmation, immunohistochemical staining showed a decrease of tumor-associated proliferation marker Ki-67 after TIFP lowering. These data suggest that the mechanical stretch induced by TIFP is a positive modulator of tumor proliferation.

Spontaneous and experimental metastasis can be effectively inhibited by the widely used anticoagulant heparin in different tumor models. At the cellular level, many of the antimetastatic effects of heparin in vivo are due to its action on P-selectin-mediated binding. Whereas previous attention has focused on P-selectin-dependent tumor-cell-platelet interactions in blood-borne metastasis, we sought to address the potential contribution of endothelial P-selectin expression to adhesive events between the microvasculature and melanoma cells in vivo. Transplantation of bone marrow from P-selectin-deficient into wild-type mice conveyed inhibition of ex-perimental melanoma metastasis. However, the extent to which bone marrow-conferred lack of platelet P-selectin expression attenuated melanoma lung metastasis was significantly less than that seen in P-selectin-deficient mice, suggesting that endothelial P-selectin expression may additionally contribute to formation of hematogenous metastases. This assumption was supported by our intravital microscopy studies, in which a significant proportion of melanoma cells were capable of directly interacting with postcapillary venules of the murine ear in a P-selectin-dependent manner. Heparin not only inhibits P-selectin-mediated melanoma cell rolling but also attenuates melanoma metastasis formation in vivo, further supporting the concept that endothelial P-selectin expression may represent an additional target of heparin action in experimental melanoma lung metastasis.

Selectin-mediated tethering and rolling initiates the multi-step process of leukocyte extravasation which is crucial for the formation of an inflammatory infiltrate. We studied the impact of platelets on this process in the skin. Using intravital microscopy, we analyzed platelet interactions with cutaneous post-capillary venules of mouse ears and observed an increase in platelet rolling if platelets were activated (41.6+/-20.2% vs. 13.1+/-8.5% rolling of resting platelets). Experiments with P-selectin deficient mice and antibodies blocking either P-selectin, GPIIb/IIIa or GPIb showed that rolling depends on platelet PSGL-1 and GPIIb/IIIa on one hand, and endothelial P-selectin on the other. Next, formation of platelet-leukocyte aggregates was demonstrated by simultaneous observation of platelets and leukocytes in vivo utilizing a newly developed two-color technique. Aggregates increased overall leukocyte rolling (leukocytes alone: 27.4+/-11.2%, leukocytes with resting platelets: 25.3+/-10.2%, leukocytes with activated platelets 38.1+/-11.8%). To investigate if activated platelets may contribute to the pathogenesis of chronic cutaneous inflammation, platelet P-selectin expression was studied in 8 patients with psoriasis. A correlation between platelet P-selectin expression and disease severity was established. In summary, we show that activated, not resting, platelets increase leukocyte rolling in murine skin. This increased rolling is due to the aggregate formation of platelets with leukocytes. We also provide evidence for a potential role of this mechanism in the pathogenesis of chronic inflammatory skin diseases.