MCF10CA Breast Cancer Cells Utilize Hyaluronan-Coated EV-Rich Trails for Coordinated Migration

Abstract

Invasion of tumor cells through the stroma is coordinated in response to migratory cues provided by the extracellular environment. One of the most abundant molecules in the tumor microenvironment is hyaluronan, a glycosaminoglycan known to promote many hallmarks of tumor progression, including the migratory potential of tumor cells. Strikingly, hyaluronan is also often found to coat extracellular vesicles (EVs) that originate from plasma membrane tentacles of tumor cells crucial for migration, such as filopodia, and are abundant in tumor niches. Thus, it is possible that hyaluronan and hyaluronan-coated EVs have a cooperative role in promoting migration. In this work, we compared the hyaluronan synthesis, EV secretion and migratory behavior of normal and aggressive breast cell lines from MCF10 series. Single live cell confocal imaging, electron microscopy and correlative light and electron microscopy experiments revealed that migrating tumor cells form EV-rich and hyaluronan -coated trails. These trails promote the pathfinding behavior of follower cells, which is dependent on hyaluronan. Specifically, we demonstrated that plasma membrane protrusions and EVs left behind by tumor cells during migration are strongly positive for CD9. Single cell tracking demonstrated a leader-follower behavior, which was significantly decreased upon removal of pericellular hyaluronan, indicating that hyaluronan promotes the pathfinding behavior of follower cells. Chick chorioallantoic membrane assays in ovo suggest that tumor cells behave similarly in 3D conditions. This study strengthens the important role of extracellular matrix production and architecture in coordinated tumor cell movements and validates the role of EVs as important components and regulators of tumor matrix. The results suggest that tumor cells can modify the extracellular niche by forming trails, which they subsequently follow coordinatively. Future studies will clarify in more detail the orchestrated role of hyaluronan, EVs and other extracellular cues in coordinated migration and pathfinding behavior of follower cells.

Evaluation of cell disruption technologies on magnetosome chain length and aggregation behaviour from Magnetospirillum gryphiswaldense MSR-1

“…CellMask™ Deep Reed has been previously used to stain both cells and extracellular vesicle lipidic membranes ( Shentu et al, 2017 ; Aaltonen et al, 2022 ). In this study, we tested a range of different fluorophore concentrations to determine the optimal magnetosome membrane staining conditions using nFCM technology.…”

Centrosome amplification promotes cell invasion via cell–cell contact disruption and Rap-1 activation

“…The chorioallantoic membrane assay (CAM assay), is a rapid, cost effective in vivo model for the study of tumorigenesis, angiogenesis, invasion and metastases (reviewed in DeBord et al, 2018 ; Komatsu et al, 2019 ; Miebach et al, 2022 ; Ribatti, 2014 ). The CAM assay has been used to study angiogenesis and tumour cell invasion in breast, pancreatic, colorectal, prostate, brain and ovarian cancers using both human tumour cell line-derived xenografts as well as patient-derived xenografts ( Aaltonen et al, 2022 ; Harper et al, 2021 ; Lokman et al, 2012 ). The highly vascularised chicken chorioallantoic membrane of the fertilised chicken egg (onto which the MCF10A cells are grafted) consists of the chorionic epithelium (the outer layer) with a superficial vascular plexus, and the intermediate mesoderm layer and the allantoic epithelium (inner layer).…”

“Centrosome Amplification promotes cell invasion via cell-cell contact disruption and Rap-1 activation”

“…The Chorioallantoic membrane assay (CAM assay), is a rapid, cost effective in vivo model for the study of tumourigenesis, angiogenesis, invasion and metastases (reviewed in (DeBord, 2018; Komatsu et al , 2019; Miebach et al , 2022; Ribatti, 2014). The CAM assay has been used to study angiogenesis and tumour cell invasion in breast, pancreatic, colorectal, prostate, brain and ovarian cancers using both human tumour cell line-derived xenografts as well as patient-derived xenografts (Aaltonen et al , 2022; Harper et al , 2021; Lokman et al , 2012). The highly vascularised chicken chorioallantoic membrane of the fertilised chicken egg (onto which the MCF10A cells are grafted) consists of the chorionic epithelium (the outer layer) with a superficial vascular plexus, and the intermediate mesoderm layer and the allantoic epithelium (inner layer).…”