Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models

Ganokon Urkasemsin; Phoebe Castillo; Sasitorn Rungarunlert; Nuttha Klincumhom; Joao N Ferreira

doi:10.3390/biom9110657

Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models

Biomolecules. 2019 Oct 25;9(11):657. doi: 10.3390/biom9110657.

Authors

Ganokon Urkasemsin¹, Phoebe Castillo², Sasitorn Rungarunlert¹, Nuttha Klincumhom^{3

4}, Joao N Ferreira^{3

4}

Affiliations

¹ Department of Preclinical and Applied Animal Science, Faculty of Veterinary Science, Mahidol University, Nakhon Pathom 73170, Thailand.
² Faculty of Dentistry, National University of Singapore, Singapore 119085, Singapore.
³ Exocrine Gland Biology and Regeneration Research Group, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.
⁴ Center of Excellence in Regenerative Dentistry, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, Thailand.

Abstract

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.

Keywords: neurons; porcine; primary cells; progenitor cells; salivary glands; secretory epithelia; xerostomia.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cell Differentiation
Epithelium*
Salivary Glands*
Swine
Tissue Engineering*