Research Assistant/ PhD Candidate University at Buffalo Buffalo, New York, United States
Introduction:: Salivary gland disorders, such as xerostomia and salivary gland hypofunction, are common and can lead to significant difficulties in speaking, swallowing, and eating, causing a substantial impact on the quality of life of affected individuals. Current treatment options for these conditions are limited and often have undesirable side effects. As a result, there is an urgent need for the development of new therapeutic approaches for salivary gland regeneration.
Human induced pluripotent stem cells (iPSCs) have emerged as a promising candidate for this purpose. They have the ability to differentiate into all cell types in the human body, including salivary gland cells, and can be generated from easily accessible adult somatic cells. Our research shows the potential of iPSC-derived salivary gland epithelial progenitor (SGEP) cells for salivary gland regeneration. These cells are a subpopulation of salivary gland cells that can differentiate into mature glandular lineages, the acinar and ductal cells. Furthermore, iPSC-derived SGEP cells have the potential to generate salivary gland organoids, three-dimensional structures that mimic the architecture and function of the gland, providing an avenue for drug testing and the development of new therapies. Additionally, upon orthotopic transplantation these cells are able to survive and engraft into the tissue, as well as differentiate into the acinar and ductal lineages.
Here we show the potential of iPSC-derived SGEP cells for autologous salivary gland regeneration as it provides a promising avenue for the development of new therapies for salivary gland disorders.
Materials and Methods:: We developed a stepwise protocol for the development of salivary gland epithelial progenitors and later acinar cell-specific organoids from human Induced Pluripotent Stem Cells (IPSC) for biomedical applications such as in tissue engineering, regenerative medicine, and drug testing. The procedure employs a novel differentiation technique that guides IPSC through the various glandular developmental stages with each progenitor lineage being induced via treatment with specific cocktails of growth factors, small molecules (agonist/inhibitors) and extracellular matrix (ECM) to mimic the developmental microenvironment necessary for the specific stage.
This procedure has no use of virus induced gene overexpression, microdissection of progenitor buds protruding from embryoid bodies or sorting techniques for progenitors during any differentiation stage, hence making it readily usable. This invention is also directed to the development of functional and biologically accurate acinar spheroids that express the correct mature markers in their specific cellular compartment along with the spheroids being polarized and lumenized to mimic the acinar phenotype in vivo, hence providing a platform for drug testing. This disclosure also includes the method for implementing the differentiation protocol to achieve the various stages along with the delivery of progenitor cells into a wounded salivary gland model via an engineered hydrogel to promote regeneration and show its potential to achieve mature glandular cell phenotypes.
Results, Conclusions, and Discussions:: Organoids are three-dimensional cell constructs that can recapture the complicated tissue microarchitecture that imparts function to specialized epithelial tissues. Therefore, these organoids hold great promise in applications such as tissue engineering, drug testing and developmental biology. Many successful drug candidates tested in rodent models fail to achieve the same level of success in human trials due to the innate biological differences in human organs as compared to rodent tissue. Significant progress in human IPSC based tissue engineering has led to organoids being able to truly mimic the in vivo complexities of organs and their functions. Previous research in salivary gland organoid development, as mentioned above, has been able to recapture the glandular phenotype through viral overexpression of transcription factors and subsequent microdissection of three-dimensional buds originating from the embryoid body. These methods are inefficient and laborious to perform with low reproducibility being a major concern.
Our invention addresses these challenges by: 1) providing a strategy to create glandular epithelial progenitors that express all the initiation markers of the primordial epithelial prebud that develops into the gland. We can do this with the use of commercially available growth factors and inhibitors and hence avoid the use of any viral overexpression system or of three-dimensional cultures at this stage. 2)providing a strategy to build complex lumenized organoids that mimic the adult acinar compartment with the ability to express mature acinar markers in their correct subcellular compartments. We are also able to mimic human acinar cell development through its various developmental stages through commercially available growth factors, inhibitors and extracellular matrices. These organoids are clinically relevant models for drug testing and hence limit the use of rodent models of xerostomia or Sjogren’s syndrome. 3) providing a strategy to implant these cells and cell constructs as a means for cell therapy to restore glandular function and rescue epithelial tissue from fibrosis caused by head and neck proton therapy and autoimmune responses of Sjogren’s syndrome.
Overall, the ease of attaining glandular epithelial progenitors and the subsequent ease of construction of organoids that mimic the glandular microarchitecture show the superiority and help differentiate our method.
