Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models.
Ganokon UrkasemsinPhoebe CastilloSasitorn RungarunlertNuttha KlincumhomJoão N FerreiraPublished in: Biomolecules (2019)
Research efforts have been made to develop human salivary gland (SG) secretory epithelia for transplantation in patients with SG hypofunction and dry mouth (xerostomia). However, the limited availability of human biopsies hinders the generation of sufficient cell numbers for epithelia formation and regeneration. Porcine SG have several similarities to their human counterparts, hence could replace human cells in SG modelling studies in vitro. Our study aims to establish porcine SG explant outgrowth models to generate functional secretory epithelia for regeneration purposes to rescue hyposalivation. Cells were isolated and expanded from porcine submandibular and parotid gland explants. Flow cytometry, immunocytochemistry, and gene arrays were performed to assess proliferation, standard mesenchymal stem cell, and putative SG epithelial stem/progenitor cell markers. Epithelial differentiation was induced and different SG-specific markers investigated. Functional assays upon neurostimulation determined α-amylase activity, trans-epithelial electrical resistance, and calcium influx. Primary cells exhibited SG epithelial progenitors and proliferation markers. After differentiation, SG markers were abundantly expressed resembling epithelial lineages (E-cadherin, Krt5, Krt14), and myoepithelial (α-smooth muscle actin) and neuronal (β3-tubulin, Chrm3) compartments. Differentiated cells from submandibular gland explant models displayed significantly greater proliferation, number of epithelial progenitors, amylase activity, and epithelial barrier function when compared to parotid gland models. Intracellular calcium was mobilized upon cholinergic and adrenergic neurostimulation. In summary, this study highlights new strategies to develop secretory epithelia from porcine SG explants, suitable for future proof-of-concept SG regeneration studies, as well as for testing novel muscarinic agonists and other biomolecules for dry mouth.
Keyphrases
- endothelial cells
- stem cells
- smooth muscle
- induced apoptosis
- signaling pathway
- flow cytometry
- mesenchymal stem cells
- induced pluripotent stem cells
- pluripotent stem cells
- cell therapy
- cell cycle arrest
- single cell
- gene expression
- cell death
- oxidative stress
- genome wide
- bone marrow
- cell proliferation
- mass spectrometry
- current status
- copy number
- high resolution
- subarachnoid hemorrhage
- quality improvement
- cerebral ischemia
- high density
- blood brain barrier