RT Journal Article
SR Electronic
T1 Unlabeled salivary gland organoids have distinct Raman signatures following FGF2-induced proacinar cell differentiation
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2021.09.16.460651
DO 10.1101/2021.09.16.460651
A1 Kate Tubbesing
A1 Nicholas Moskwa
A1 Ting Chean Khoo
A1 Deirdre A. Nelson
A1 Anna Sharikova
A1 Melinda Larsen
A1 Alexander Khmaladze
YR 2021
UL http://biorxiv.org/content/early/2021/09/16/2021.09.16.460651.abstract
AB Organoids are self-organized three-dimensional (3D) tissue cultures that model the structure and function of organs to provide insights into signaling during organ formation and have translational applications in disease modeling and assessing drug responses. Due to their heterogeneity, there is a need for non-destructive methods to identify the differentiation state, or the phenotype, of organoids. As organoids often contain complex mixtures of basement membrane and/or extracellular matrix proteins, which are often highly auto-fluorescent, it typically makes low-resolution Raman measurements a challenge. We developed Raman confocal micro-spectroscopy methods to avoid and minimize the matrix signal and define specific Raman signatures for growth factor-differentiated and non-differentiated organoids. In complex, branched salivary gland organoids derived from mouse embryonic epithelial and stromal cells embedded within the laminin-rich basement membrane matrix, Matrigel, we identified specific Raman spectral signatures for organoids in different differentiation states. We report that either comparison of spectral signatures or multivariate SVD analysis can be used to distinguish between organoids treated with FGF2, organoids treated with EGF, and non-treated controls. Raman spectral signatures can be used to non-invasively distinguish between different phenotypes in the 3D context of unlabeled organoids.HighlightsFGF2-dependent proacinar cell differentiation in salivary organoids have unique Raman signatures detected with a novel confocal-based Raman imaging approach.These signatures can be used in unlabeled salivary organoids to monitor proacinar cell differentiation.Confocal-based Raman imaging may be applicable to monitoring differentiation state of other types of organoids.Competing Interest StatementThe authors have declared no competing interest.AQP5Aquaporin-5, used to identify proacinar cellsDAPI4′,6-diamidino-2-phenylindole, a nucleic acid dyeDMEM/F12Dulbecco’s Modified Eagle Medium /Nutrient Mixture F12EGFEpidermal growth factor, used to promote growth not proacinar differentiationEpCAMEpithelial cell adhesion molecule, epithelial cell markerFBSfetal bovine serumFGF2Fibroblast growth factor 2, used to promote proacinar cell differentiationFITCFluorescein isothiocyanateK7Keratin-7, used to identify ductal cellsPen-Streppenicillin and streptomycin