Mechanisms of Basement Membrane Micro-Perforation during Cancer Cell Invasion into a 3D Collagen Gel

• Published in: Gels 2022-09, Vol.8 (9), p.567
• Main Authors: Nazari, Shayan S, Doyle, Andrew D, Yamada, Kenneth M
• Format: Article
• Language: English
• Subjects: 3D culture; Angiogenesis; Basements; Cancer; cancer invasion; Cell adhesion & migration; cell-matrix interactions; Collagen; Enzymes; Gels; Growth factors; hydrogel; imaging; Laboratories; Lymphatic system; Membranes; Metastasis; Microscopy; Morphogenesis; Muscle proteins; Myosin; Perforation; Polymerization; Protease inhibitors; Proteases; Proteolysis; Spheroids; Tumors
• ISSN: 2310-2861; 2310-2861
• DOI: 10.3390/gels8090567

Cancer invasion through basement membranes represents the initial step of tumor dissemination and metastasis. However, little is known about how human cancer cells breach basement membranes. Here, we used a three-dimensional in vitro invasion model consisting of cancer spheroids encapsulated by a basement membrane and embedded in 3D collagen gels to visualize the early events of cancer invasion by confocal microscopy and live-cell imaging. Human breast cancer cells generated large numbers of basement membrane perforations, or holes, of varying sizes that expanded over time during cell invasion. We used a wide variety of small molecule inhibitors to probe the mechanisms of basement membrane perforation and hole expansion. Protease inhibitor treatment (BB94), led to a 63% decrease in perforation size. After myosin II inhibition (blebbistatin), the basement membrane perforation area decreased by only 15%. These treatments produced correspondingly decreased cellular breaching events. Interestingly, inhibition of actin polymerization dramatically decreased basement membrane perforation by 80% and blocked invasion. Our findings suggest that human cancer cells can primarily use proteolysis and actin polymerization to perforate the BM and to expand perforations for basement membrane breaching with a relatively small contribution from myosin II contractility.