Microstructural densification and alignment by aspiration-ejection influence cancer cell interactions with three-dimensional collagen networks.

Extracellular matrix microstructure and mechanics are crucial to breast cancer progression and invasion into surrounding tissues. The peritumor collagen network is often dense and aligned, features which in vitro models lack. Aspiration of collagen hydrogels led to densification and alignment of microstructure surrounding embedded cancer cells. Two metastasis-derived breast cancer cell lines, MDA-MB-231 and MCF-7, were cultured in initially 4 mg/ml collagen gels for 3 days after aspiration, as well as in unaspirated control hydrogels. Videomicroscopy during aspiration, and at 0, 1, and 3 days after aspiration, epifluorescence microscopy of phalloidin-stained F-actin cytoskeleton, histological sections, and soluble metabolic byproducts from constructs were collected to characterize effects on the embedded cell morphology, the collagen network microstructure, and proliferation. Breast cancer cells remained viable after aspiration-ejection, proliferating slightly less than in unaspirated gels. Furthermore, MDA-MB-231 cells appear to partially relax the collagen network and lose alignment 3 days after aspiration. Aspiration-ejection generated aligned, compact collagen network microstructure with immediate cell co-orientation and higher cell number density apparently through purely physical means, though cell-collagen contact guidance and network remodeling influence cell organization and collagen network microstructure during subsequent culture. This study establishes a platform to determine the effects of collagen density and alignment on cancer cell behavior, with translational potential for anticancer drug screening in a biomimetic three-dimensional matrix microenvironment, or implantation in preclinical models.