Tumor MMP-1 Activates Endothelial PAR1 to Facilitate Vascular Intravasation and Metastatic Dissemination

Abstract

Intravasation, the active entry of primary tumor cells into the vasculature, remains the least studied step in the metastatic cascade. Protease-mediated escape and stromal invasion of tumor cells represent widely-accepted processes leading up to the intravasation step. However, molecular factors that contribute directly to tumor cell vascular penetration have not been identified. In this study, the in vivo role of the collagenolytic protease, MMP-1, in cancer cell intravasation and metastasis was analyzed by employing a highly-disseminating variant of human HEp3 epidermoid carcinoma, HEp3-hi/diss. Whereas naturally-acquired or experimentally-induced MMP-1 deficiency substantially suppressed HEp3-hi/diss intravasation, supplementation of recombinant MMP-1 to MMP-1-silenced primary tumors, restored their impaired vascular dissemination. Surprisingly, abrogation of MMP-1 production and activity did not affect significantly HEp3-hi/diss migration or matrix invasion, suggesting non-collagenolytic mechanisms underlying MMP-1-dependent cell intravasation. In support of such non-collagenolytic mechanisms, MMP-1 silencing in HEp3-hi/diss cells modulated the microarchitecture and integrity of the angiogenic vasculature in a novel microtumor model. Concomitantly, MMP-1 deficiency led to decreased levels of intratumoral vascular permeability, tumor cell intravasation and metastatic dissemination. Taking advantage of PAR1 deficiency of HEp3-hi/diss cells, we further demonstrate that endothelial PAR1 is a putative non-tumor-cell/non-matrix target, activation of which by carcinoma-produced MMP-1 regulates endothelial permeability and transendothelial migration. The inhibitory effects of specific PAR1 antagonists in live animals have also indicated that the mechanisms of MMP-1-dependent vascular permeability in tumors involve endothelial PAR1 activation. Together, our findings mechanistically underscore the contribution of a tumor MMP-1/endothelial PAR1 axis to actual intravasation events manifested by aggressive carcinoma cells.

Matrix metalloproteinases in the brain and blood–brain barrier: Versatile breakers and makers

“…2. Intravasation of metastatic cells into the blood circulation: Juncker-Jenson et al 257 found that MMP-1 participates in the intravasation of metastatic cells from a human HEp3 epidermoid carcinoma graft in chick embryos. The authors showed that MMP-1 regulating endothelial permeability and transendothelial migration supported tumor invasion by activating the endothelial non-tumor/nonmatrix receptor PAR1.…”

Pericellular proteolysis in cancer

“…For example, MMP17 facilitates breast cancer cell intravasation by promoting the detachment of pericytes from blood vessels, which also causes increased tumor vascular leakage (Chabottaux et al 2009). Tumor cellderived MMP1 cleaves and activates protease-activated receptor-1 (PAR-1) on endothelial cells, which leads to increased endothelial permeability and trans-endothelial migration of cancer cells (Juncker-Jensen et al 2013). MMP1 and MMP2 are also critical mediators of breast cancer cell extravasation through lung capillaries to seed lung metastases (Minn et al 2005;Gupta et al 2007).…”

An Improved In Vivo Methodology to Visualise Tumour Induced Changes in Vasculature Using the Chick Chorionic Allantoic Membrane Assay

“…Intravasation is considered to be the rate-limiting step in cancer metastasis however it is under-investigated (16), due to the requirement for sophisticated technical imaging equipment (e.g. time-lapse confocal microscopy) that is required to study in vivo tumour intravasation and metastasis (17).…”

“…This methodology was applicable because we did not use a cell carrier material unlike previous studies that have used Matrigel or a collagen matrix to implant tumour cells on the CAM (17). Although using a cell carrier has obvious advantage of keeping tumour cells together, our initial experiments demonstrated a moderate inflammatory reaction to the natural ECM based carriers especially after EDD 12 (30).…”