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|Title: ||In vitro generation of human FOXP3+ regulatory T cells|
|Authors: ||Abdulrasool, Ghusoon Abdulhameed Ebrahim Salman|
|Issue Date: ||9-Jan-2014|
|Publisher: ||University of Sydney.
Western Clinical School|
|Abstract: ||Human Foxp3+ regulatory T cells, particularly in vitro induced Tregs (iTregs), have potential therapeutic applications in transplantation and autoimmune disease. Several protocols for generating functionally stable iTregs have been developed. However, information regarding Foxp3 methylation status, the most specific measure for functional stability, in these differently induced Tregs is lacking.
Here human Foxp3+ iTregs were generated using seven different protocols with 24 different culture conditions. The methylation status of the Foxp3 gene was used as a measure of Treg functional stability and the percentage of iTregs was used for quantification.
Results showed that only six culture conditions significantly enhanced Foxp3 promoter demethylation, the mean percentage ranged from 20% to 36%. TGFβ and tRA protocol produced the highest proportion of iTregs with demethylated Foxp3 promoter. It converted 31% ± 6 of naïve T cells into Foxp3+ iTregs and induced Foxp3 demethylation in 36% ± 3 of total DNA.
In summary, our data identify the methylation status of the Foxp3 gene in different types of iTregs and proved that all iTregs displays partial demethylation of the Foxp3 promoter, which is consistent with a transient Treg phenotype. In addition, this study identified the most promising protocol (TGF-β+tRA) for generating Tregs in vitro. These findings hold important implications for the in vitro methods of Treg generation and provide the basis for future development of more effective strategies.|
|Type of Work: ||Masters Thesis|
|Type of Publication: ||Master of Philosophy M.Phil|
|Appears in Collections:||Sydney Digital Theses (Open Access)|
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