The Role of Nitric Oxide and Peroxynitrite in Human Embryonic Stem Cell Differentiation

Human embryonic stem cells (hESCs) have the potential to grow into almost all types of cells in the human body. This special property makes hESC invaluable for the restoration of tissue function after damage from different diseases like Parkinson's disease, Alzheimer’s disease, stroke, and diabetes. The potential therapeutic applications of hESC have been hindered by a lack of knowledge of the differentiation mechanisms. It is not clear what kind of message has to be delivered to hESC to start differentiation, and what process leads to the selected cell differentiation directions. At this stage of research, it is impossible to direct the differentiation of hESC to functional specific cells for the restoration of damaged tissues. This study focused on the role of small, diffusible molecules on hESC differentiation. We hypothesized that nitric oxide (NO) and/or peroxynitrite (ONOO-) may play a role as redox mediators during early embryogenesis. After having working cell banks of mouse fibroblasts and hESCs, an eighteen-day hESC early differentiation system was established through the formation of embryoid bodies (EBs). The differentiation status of cells was evaluated through flow cytometry, proteome profiler arrays, and immunocytostaining. A nanomedical approach was used in this study. The bioavailable NO and ONOO- concentration was measured with selective electrochemical nanosensors during the differentiation process. A detailed profile of the NO and ONOO- concentrations as a function of time during the differentiation process was established. Also, several compounds which can influence the NO or ONOO- production were used to treat the cells during the differentiation process. We found that in the established differentiation system, the differentiation of hESC towards all three germ layers occurred. NO and ONOO- were both available in the microenvironment during hESC differentiation. We proved that both NO and ONOO- were crucial to the differentiation process and had a facilitating effect on the production of neuroectoderm cells. NO has a profound inhibitory effect on the production of mesoderm cells. This study is the first to demonstrate on the molecular level that both NO and ONOO- play a fundamental redox regulatory role in the early differentiation of hESCs.

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Chemistry and Biochemistry (Arts and Sciences)

Grantor dc:publisher

Ohio University

Year dc:date

2013

Author dc:creator

• Wang, Han

Contributors dc:contributor

• Malinski, Tadeusz

• Biochemistry
• Nitric Oxide
• Stem Cell
• Peroxynitrite
• Differentiation

Statement dc:rights

• unrestricted
• This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.

Language dc:language

English

Repository record dc:identifier

http://rave.ohiolink.edu/etdc/view?acc_num=ohiou1363779566

OAI identifier oai:identifier

oai:etd.ohiolink.edu:ohiou1363779566