Date of Award

Spring 2023

Project Type

Dissertation

Program or Major

Chemical Engineering

Degree Name

Doctor of Philosophy

First Advisor

Kyung Jae Jeong

Second Advisor

John Tsavalas

Third Advisor

Kang Wu

Abstract

In this dissertation, I developed and investigated gelatin-based microporous injectable hydrogels for the encapsulation of stem cells for multiple applications in cell delivery. Utilizing microgels composed from a mixture of gelatin and modified gelatin, I demonstrated the utility of a dual crosslinking mechanism, which enabled rapid gelation and tissue adhesion with improved cytocompatibility. Mesenchymal stem cells (MSCs) encapsulated in this hydrogel proliferated at a more rapid rate than in a nonporous counterpart, and showed increased immunomodulatory potential. Then, I investigated gelatin microporous hydrogel for the encapsulation of MSCs for bone tissue regeneration. Encapsulated cells more readily differentiated into osteoblasts (i.e. bone-forming cells) in the microporous environment observed by morphological changes and quantitative assays. This is believed to be due to enhanced cell spreading and cell-cell communication in the unique 3D environment provided to the cells by the microporous hydrogel. Transcriptomic analysis was performed by mRNA sequencing (RNA-seq) of MSCs encapsulated in the differing 3D microenvironments. Results indicated that the 3D environment influenced the expression of genes that are related to cell adhesions, cell-cell interactions, cytoskeletal organization, and matrix remodeling, in addition to MSC differentiation. Because neuronal development is highly dependent on cell-cell communication, I encapsulated an established neural stem cell line (ReNcell) in gelatin microporous hydrogel to investigate neuronal differentiation in comparison to a nonporous analog. Laminin was chemically conjugated to microgel surfaces, which controlled the organization of encapsulated cells in the hydrogel environment. Cell differentiation was examined by immunofluorescence staining, and JC-1 assay was utilized to examine mitochondrial membrane polarization. The microporous hydrogel induced substantially greater cell spreading, morphological changes and cell-cell connections than nonporous hydrogel. The majority of the cells in the microporous hydrogel differentiated into neural lineages, evidenced by immunostaining by MAP2 and GFAP. In summary, this work demonstrates the utility of gelatin microporous injectable hydrogels for applications in in situ cell encapsulation and stem cell delivery for tissue regeneration.

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