Student Theses, Dissertations and Capstones

Document Type

Thesis - NSU Access Only

Degree Name

Master of Science (M.S.) in Dentistry

Copyright Statement

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Department

College of Dental Medicine

First Advisor

Umadevi Kandalam

Date of original Performance / Presentation

January 2022

Publication Date / Copyright Date

1-1-2022

Abstract

Background: Stem cell-based bone regenerative therapy is the promising option for regeneration of critical-sized bone defects in craniofacial area. However, insufficient vascularization presents a clinical challenge in bone regenerative therapies. Vascularization is critical because newly formed blood vessels are important for continuous supply of oxygen, nutrients, and growth factors essential for initial bone growth. The initial phase of acute/transient inflammation is necessary for recruitment of several cell types that facilitate wound healing. A repertoire of cytokines, like TNF-α, IL-1β, IL-8, IL-6, IL-10, IL-4, IL-13 that govern pro-and anti-inflammatory responses are released at the site of injury. An acute phase inflammation at the site of injury creates a microenvironment for recruiting cells that facilitate wound healing, protect from invading pathogens, and induces angiogenesis and hence is considered a homeostatic response for tissue repair. Regulated cytokine expression is therefore necessary for resolving or repairing the damaged tissues. Objective: The overall objective of this study was to investigate the cytokine profiles during the guided differentiation of gingival mesenchymal stem cells (GMSCs) to angiogenic precursor cells. Methodology: Mesenchymal stem cells were isolated from human gingiva (GMSCs) obtained as discarded tissue from dental procedures. The cells were cultured using standard culture conditions and further expanded. GMSCs were guided to differentiate into endothelial lineage, inducing with VEGF (50ng/ml) for 7, 14 and 21 days. The cells with no VEGF treatment served as the control group. The gene expressions of various pro-inflammatory and anti-inflammatory cytokines were measured using quantitative PCR. Quantitative analysis of cytokines was measured using ELISA. Results: The flow cytometry analysis indicated that the gingival cells were characterized for MSCs markers. The multilineage differentiation potential of gingival cells was confirmed by in vitro differentiation assays and the tube formation assay demonstrated the angiogenic potential of differentiating GMSCs. The cytokine IL-1β showed gradual increase with increase in the days of induction, while IL-6 showed a decrease during week 1 and 2 and there was a significant increase compared to undifferentiated cells at week 3. TNF-α increased during week 1 while there was significant decrease in week 2 and 3. The mRNA expression of other cytokines IL-4, IL-10 and IL-13 showed increase in mRNA expression during the time course. IL-8 on the other hand, downregulated during week 1 and there was significant upregulation during week 2 and 3. The Elisa results showed that there was difference in the level of the cytokine expression by GMSCs treated with or without VEGF. Conclusion: The VEGF supported the endothelial differentiation of GMSCs. The results showed that VEGF has potential of regulating the cytokine expression profiles during bone tissue engineering. Modulating cytokine expression may enhance the vascularization in cell-based bone regenerative techniques.

Disciplines

Dentistry

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