3D Organoid Culture From Adult Salivary Gland Tissues as an ex vivo Modeling of Salivary Gland Morphogenesis

Abstract

Lumen formation of salivary glands has been investigated using in vivo or ex vivo rudiment culture models. In this study, we used a three-dimensional (3D) salivary gland organoid culture system and demonstrated that lumen formation could be recapitulated in mouse SMG organoids. In our organoid culture system, lumen formation was induced by vasoactive intestinal peptide and accelerated by treatment with RA. Furthermore, lumen formation was observed in branching duct-like structure when cultured in combination of fibroblast growth factors (FGF) in the presence of retinoic acid (RA). We suggest RA signaling-mediated regulation of VIPR1 and KRT7 as the underlying mechanism for lumen formation, rather than apoptosis in the organoid culture system. Collectively, our results support a fundamental role for RA in lumen formation and demonstrate the feasibility of 3D organoid culture as a tool for studying salivary gland morphogenesis.

Encapsulation of Primary Salivary Gland Acinar Cell Clusters and Intercalated Ducts (AIDUCs) within Matrix Metalloproteinase (MMP)‐Degradable Hydrogels to Maintain Tissue Structure and Function

“…Previous studies by other groups have also observed dead cells and positive caspase-3 staining in the center of the acini-like structures when cultur-ing salivary gland progenitor cells in various hydrogels. [20,27,[40][41][42][43] Cavities with varying number and size were observed by day 14 (Figure 3D,E), but they cannot be designated as lumens, as it is unclear how or when they are formed and whether they are polarized differently from the outer cell layer. We speculate that cells on the inside of the spheres may undergo apoptosis because they are excluded from cell-cell contacts found on the outside of the sphere.…”

Emerging organoid-based platforms to study salivary gland hypofunction

“…Several attempts have been conducted to fulfill these criteria (Table 2) [29,[35][36][37][38][39][40][41][42][43]. Specifically, the regulation of signaling pathways associated with SG organogenesis is one of the common strategies to improve basic culture conditions.…”

“…In addition, chemical inhibition of activin receptor-like kinase signaling or treatment of morphogenic proteins such as the Fgf family can induce the development of acini-containing budding structure in SGOs [30,36,37]. In addition to [38] focused on recapitulating the lumen formation process during the maturation of SGOs. According to that study, the Fgf2/Fgf10 combination initiates the budding and branching elongation process, and the addition of retinoic acid (RA) enhances lumen formation within this duct-like compartment.…”

“…Thus, the ECM and ECM-mimicking substances in SGO culture not only function as a physical scaffold, but also provide biological interactions facilitating the generation and maturation of SGOs. Matrigel is the most commonly used hydrogel at present, but potential benefits of other natural-and synthetic biomaterials have been explored for organoid culture [38,44]. The natural ECM obtained from naïve SG tissue can be utilized as hydrogel or scaffold for SG-derived cell culture after decellularization [39,45].…”

A Mammary Organoid Model to Study Branching Morphogenesis

“…In parallel to emerging imaging technologies, efforts have been concentrated on in vitro organotypic systems of the mammary gland ( Simian et al, 2001 ; Debnath et al, 2003 ; Ewald et al, 2008 , 2012 ). Although multiple organoid models derived from different organs have achieved high levels of complexity and morphological reproducibility of in vivo structures, the complexity of murine mammary organoids is still limited ( Clevers, 2016 ; Kim et al, 2021 ; Vazquez-Armendariz and Herold, 2021 ). Current mammary organoid culture protocols result in either bi-layered sphere-shaped organoids (referred to as cysts) or budding structures upon supplementation with different growth factors, such as FGF2, FGF7, FGF10, and Neuregulin1 ( Fata et al, 2007 ; Nguyen-Ngoc et al, 2015 ; Jardé et al, 2016 ; Sumbal et al, 2020 ).…”