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

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...

Full description

Bibliographic Details
Main Authors: Ganokon Urkasemsin, Phoebe Castillo, Sasitorn Rungarunlert, Nuttha Klincumhom, Joao N. Ferreira
Format: Article
Language:English
Published: MDPI AG 2019-10-01
Series:Biomolecules
Subjects:
Online Access:https://www.mdpi.com/2218-273X/9/11/657
id doaj-460b2f55419a408fbbba7608aacb39e0
record_format Article
spelling doaj-460b2f55419a408fbbba7608aacb39e02020-11-25T02:16:07ZengMDPI AGBiomolecules2218-273X2019-10-0191165710.3390/biom9110657biom9110657Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture ModelsGanokon Urkasemsin0Phoebe Castillo1Sasitorn Rungarunlert2Nuttha Klincumhom3Joao N. Ferreira4Department of Preclinical and Applied Animal Science, Faculty of Veterinary Science, Mahidol University, Nakhon Pathom 73170, ThailandFaculty of Dentistry, National University of Singapore, Singapore 119085, SingaporeDepartment of Preclinical and Applied Animal Science, Faculty of Veterinary Science, Mahidol University, Nakhon Pathom 73170, ThailandExocrine Gland Biology and Regeneration Research Group, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, ThailandExocrine Gland Biology and Regeneration Research Group, Faculty of Dentistry, Chulalongkorn University, Bangkok 10330, ThailandResearch 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.https://www.mdpi.com/2218-273X/9/11/657xerostomiasalivary glandsprimary cellsprogenitor cellssecretory epithelianeuronsporcine
collection DOAJ
language English
format Article
sources DOAJ
author Ganokon Urkasemsin
Phoebe Castillo
Sasitorn Rungarunlert
Nuttha Klincumhom
Joao N. Ferreira
spellingShingle Ganokon Urkasemsin
Phoebe Castillo
Sasitorn Rungarunlert
Nuttha Klincumhom
Joao N. Ferreira
Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
Biomolecules
xerostomia
salivary glands
primary cells
progenitor cells
secretory epithelia
neurons
porcine
author_facet Ganokon Urkasemsin
Phoebe Castillo
Sasitorn Rungarunlert
Nuttha Klincumhom
Joao N. Ferreira
author_sort Ganokon Urkasemsin
title Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
title_short Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
title_full Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
title_fullStr Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
title_full_unstemmed Strategies for Developing Functional Secretory Epithelia from Porcine Salivary Gland Explant Outgrowth Culture Models
title_sort strategies for developing functional secretory epithelia from porcine salivary gland explant outgrowth culture models
publisher MDPI AG
series Biomolecules
issn 2218-273X
publishDate 2019-10-01
description 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.
topic xerostomia
salivary glands
primary cells
progenitor cells
secretory epithelia
neurons
porcine
url https://www.mdpi.com/2218-273X/9/11/657
work_keys_str_mv AT ganokonurkasemsin strategiesfordevelopingfunctionalsecretoryepitheliafromporcinesalivaryglandexplantoutgrowthculturemodels
AT phoebecastillo strategiesfordevelopingfunctionalsecretoryepitheliafromporcinesalivaryglandexplantoutgrowthculturemodels
AT sasitornrungarunlert strategiesfordevelopingfunctionalsecretoryepitheliafromporcinesalivaryglandexplantoutgrowthculturemodels
AT nutthaklincumhom strategiesfordevelopingfunctionalsecretoryepitheliafromporcinesalivaryglandexplantoutgrowthculturemodels
AT joaonferreira strategiesfordevelopingfunctionalsecretoryepitheliafromporcinesalivaryglandexplantoutgrowthculturemodels
_version_ 1724892648980873216