Andrew Shubin receives commendation for PhD Dissertation

May 23, 2017

efAndrew Shubin, a 2017 PhD graduate in the lab of Professor Danielle Benoit, has been selected to receive commendation in the Outstanding Dissertation Award Competition for Engineering. Andrew's PhD research project is titled, "Poly(ethylene glycol) Hydrogels for Salivary Gland Regeneration.”

Over 500,000 people worldwide are diagnosed with head and neck cancers yearly.  Radiation, a mainstay of curative therapy, causes irreparable damage to the salivary gland acinar cells, which results in chronic dry mouth or xerostomia.  Xerostomia negatively affects patient quality of life for which no current treatments can ameliorate.  Recently, regenerative therapies utilizing the direct injection of primary submandibular gland (SMG) cells into irradiated salivary glands have shown promise in regenerating salivary gland tissues and partially restoring gland function.  However, the amount of regeneration is variable and the little is known about the mechanism of healing and the characteristics of primary SMG cells.  To address these limitations we propose the use of poly(ethylene glycol) (PEG) hydrogels to enhance in vitro culture conditions and in vivo cellular transplantation.  PEG is a “bio-inert” polymer which provides “blank-slate” to control and study specific cell-material interactions.  Initial work focused on designing cytocompatible methods to encapsulate and culture primary SMG cells within PEG hydrogels.  The minimization of radicals using thiol-ene versus methacrylate-based chain polymerizations of hydrogel macromers and the inclusion of cell-cell interactions through the formation of multicellular “spheres” supported the survival and proliferation of primary SMG cells within PEG hydrogels for a 14 day culture period.   Genetic lineage tracing was employed to determine the in vivo origin of encapsulated cells and showed that the majority (>80%) of encapsulated cells come from acinar and duct populations.  To improve PEG hydrogels as a tissue engineering platform, the effects of laminin incorporation and different forms of hydrogel degradation on the proliferation, development of acinar cell phenotypes, and epithelial morphogenesis of encapsulated SMG cells were explored.  The encapsulation of SMG cells within enzymatically (e.g., via cell-dictated processes) degradable hydrogels resulted in increased expression of the acinar water channel Aquaporin-5 (AQP5) and cellular organization reminiscent of native gland tissue compared to SMG cells encapsulated in hydrogels with bulk-hydrolytic degradability.  Degradable hydrogels (either hydrolytic or enzymatic) supported the greatest amount of cellular proliferation versus non-degradable gels.  PEG hydrogels were further utilized to study cells originating from acinar tissues in primary culture.  Despite previous studies suggesting otherwise, genetic lineage tracing showed that acinar lineage cells compose a significant portion of primary SMG cells in culture.  However, acinar lineage cells underwent significant changes in morphology and exhibited a ~100-fold decrease in expression of the acinar cell markers AQP5 and Mist1.  Furthermore, many acinar-lineage cells express cytokeratins and proliferate, indicating a transition to a duct-cell like phenotype. This acinar cell plasticity has important underpinnings for salivary gland biology and tissue engineering as acinar cells are the most prevalent cell type in the salivary gland.  Finally, in vivo methods of hydrogel mediated cell transplantation were developed, but caused sufficient mortality with irradiation.  Similar in vivo methods were well tolerated with minimal mortality in non-irradiated mice, suggesting that irradiation substantially affects surgical survival.  Taken together, this work demonstrates the utility of PEG hydrogels in characterization primary SMG cells and highlights several future areas of inquiry.