Salivary gland cell differentiation and organization on micropatterned PLGA nanofiber craters

Abstract

There is a need for an artificial salivary gland as a long-term remedy for patients suffering from salivary hypofunction, a leading cause of chronic xerostomia (dry mouth). Current salivary gland tissue engineering approaches are limited in that they either lack sufficient physical cues and surface area needed to facilitate epithelial cell differentiation, or they fail to provide a mechanism for assembling an interconnected branched network of cells. We have developed highly-ordered arrays of curved hemispherical “craters” in polydimethylsiloxane (PDMS) using wafer-level integrated circuit (IC) fabrication processes, and lined them with electrospun poly-lactic-co-glycolic acid (PLGA) nanofibers, designed to mimic the three-dimensional (3-D) in vivo architecture of the basement membrane surrounding spherical acini of salivary gland epithelial cells. These micropatterned scaffolds provide a method for engineering increased surface area and were additionally investigated for their ability to promote cell polarization. Two immortalized salivary gland cell lines (SIMS, ductal and Par-C10, acinar) were cultured on fibrous crater arrays of various radii and compared with those grown on flat PLGA nanofiber substrates, and in 3-D Matrigel. It was found that by increasing crater curvature, the average height of the cell monolayer of SIMS cells and to a lesser extent, Par-C10 cells, increased to a maximum similar to that seen in cells grown in 3-D Matrigel. Increasing curvature resulted in higher expression levels of tight junction protein occludin in both cell lines, but did not induce a change in expression of adherens junction protein Ecadherin. Additionally, increasing curvature promoted polarity of both cell lines, as a greater apical localization of occludin was seen in cells on substrates of higher curvature. Lastly, substrate curvature increased expression of the water channel protein aquaporin-5 (Aqp-5) in Par-C10 cells, suggesting that curved nanofiber substrates are more suitable for promoting differentiation of salivary gland cells.

Development of Poly(Ethylene Glycol) Hydrogels for Salivary Gland Tissue Engineering Applications

“…10,13 It is generally acknowledged that the maintenance and expansion of functional salivary gland cells requires a biomaterial scaffold. 21,24,104,105 Here, we have examined the feasibility of using PEG hydrogels. We have established permissive encapsulation procedures for multicellular SMG microspheres using thiol-ene polymerization.…”

“…Numerous studies have focused on feasibility of using nanofibers or hydrogel-based scaffolds. [15][16][17][18][19][20][21][22][23][24][25] Although a few studies have translated their findings in vivo, 16,17,23 no study to date has demonstrated that the tissue developed on or within biomaterials contributes to restoration of gland function. To overcome these limitations, we propose use of poly(ethylene glycol) (PEG) hydrogels for salivary gland cell transplantation.…”

Double emulsion electrospun nanofibers as a growth factor delivery vehicle for salivary gland regeneration

“…Scanning electron microscopy (SEM) imaging of the scaffolds was carried out using a Zeiss 1550 field emission scanning electron microscope (Leo Electron Microscopy Ltd., Cambridge, UK; Carl Zeiss, Jena, Germany), as described previously 11 . Briefly, the scaffolds were mounted on 1 cm 2 stubs and coated with approximately 5 nm of gold-palladium to minimize sample charging.…”

Core/Shell Nanofiber Characterization by Raman Scanning Microscopy

“…PGS pre-polymer was dissolved in HFIP (16% w/w). The PLGA solution consisted of 1% NaCl, 10µL SRB dye, and 85:15 PLGA dissolved in HFIP, making an 8% w/w solution [27][28][29][30]. Two independent syringe pumps were used with the two polymeric solutions, connected to the coaxial spinneret by PTFE tubing.…”