The epithelial–mesenchymal transition (EMT) is an important biological process that is physiologically observed during development, wound healing, and cancer invasion. During EMT induction, cancer cells lose their epithelial properties owing to various tumor microenvironmental factors and begin to exhibit mesenchymal properties, such as loss of apical–basal polarity, weakened intercellular adhesion, and promotion of single cell migration. Several factors, including growth factor stimulation and adhesion to type I collagen (Col-I), induce EMT in cancer cells. Cells adhere to Col-I via specific receptors and induce EMT by activating outside-in signals. In vivo, Col-I molecules often form fibrils, which then assemble into supramolecular structures (gel form). Col-I also self-assembles in vitro under physiological conditions. Notably, Col-I can be used as a culture substrate in both gel and non-gel forms, and the gel formation state of Col-I affects cell fate. Although EMT can be induced in both forms of Col-I, the effects of gel formation on EMT induction remain unclear and somewhat inconsistent. Therefore, this study reviews the relationship between Col-I gel-forming states and EMT induction in cancer cells.

• Established a mouse model for the basement membrane live imaging. • Recombinant nidogen-1, tagged with a fluorescent protein EGFP or mCherry, labeled the basement membranes both in vitro and in vivo. • The nidogen-based fluorescent probes enable monitoring the dynamics of basement membranes in mouse tissues/organoids.

SUMMARY: The fibrous structures of collagen provide physical strength and stability to tissues and organs. Abnormalities in their orientation, growth, and remodeling cause morphogenetic defects and diseases including fibrosis, highlighting the importance of understanding how collagen fibers are organized within tissues. However, this process remains difficult to study, as methods for fluorescently labeling collagen fibers with simple protocols and visualizing their three-dimensional structure are still limited. Here we describe a convenient method for fluorescent labeling of collagen fibers in vertebrate tissues. Using DAF-FM, a probe originally developed for nitric oxide detection, premature collagen fibers can be visualized through covalent binding to allysine residues, which serve as precursors of collagen cross-linking. We further show that combining two probes with different emission spectra, DAF-FM and DAR-4M, enables pulse–chase labeling of newly synthesized collagen fibers. This approach provides a practical tool for investigating collagen dynamics during tissue development and remodeling.