Expression of Mitochondrial Stress Protein (Cpn60) in in vitro Cultured Neonatal Porcine Islet Cells
Munif, F. (2006). Expression of Mitochondrial Stress Protein (Cpn60) in in vitro Cultured Neonatal Porcine Islet Cells (Thesis, Master of Science (MSc)). The University of Waikato, Hamilton, New Zealand. Retrieved from http://hdl.handle.net/10289/2510
Permanent Research Commons link: http://hdl.handle.net/10289/2510
Xenotransplantation of neonatal porcine islets have been demonstrated to be a viable alternative to exogenous insulin therapy for diabetes mellitus. The use of liberase has gained much success in islet isolation but factors such as batch-to-batch variation and deterioration of a batch with storage time have hampered the quality and reproducibility of tissue dissociation. Islet culture aims to optimise islet survival and insulin release in response to glucose challenge. However, it is difficult to recover and preserve islets in vitro. Mitochondria play a key role in the secretion of insulin from pancreatic islet cells in response to glucose stimulation. Mitochondrial dysfunction results in the induction (at mRNA and protein levels) of a molecular stress protein/heat chock protein called Cpn60. Since mitochondrial impairment will have a significant effect on the ability of in vitro cultured islet cells to function properly (i.e. release insulin in response to glucose stimulation), the expression of Cpn60 was investigated as a function of exposing neonatal porcine islet cells to various growth conditions. The best choice of media to culture neonatal porcine islet cells was found to be not heated activated serum which showed the least levels of Cpn60 expression at mRNA levels suggesting that the cells had low levels of mitochondrial stress. Neonatal porcine islet cells would be best digested in cells digested with new liberase (QC 1050) while in 2% not heat inactivated porcine serum (NPS) as this gave the lowest levels of Cpn60 expression suggesting low levels of mitochondrial stress. Although expression of Cpn60 at mRNA levels seems to be modulated during the growth of the porcine islet cells in media supplemented with different serum, heat treatment of serum and liberase content, no firm conclusion can be made with regard to the effect of the different treatments on mitochondrial health status until the porcine Cpn60 protein can be unequivocally identified.
The University of Waikato
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