In vitro cytokine licensing induces persistent permissive chromatin at the Indoleamine 2,3-dioxygenase promoter

Yazmin I. Rovira Gonzalez, Patrick J. Lynch, Elaine E. Thompson, Brian G. Stultz, Deborah A. Hursh
2016 Cytotherapy  
Background-Mesenchymal stromal cells (MSCs) are being investigated as therapies for inflammatory diseases due to their immunosuppressive capacity. IFN-γ treatment primes MSC immunosuppression partially through induction of Indoleamine 2,3-dioxygenase (IDO1), which depletes tryptophan necessary to support proliferation of activated T-cells. We investigated the role of histone modifications in the timing and maintenance of induced IDO1 expression in MSCs under clinical manufacturing conditions,
more » ... ch as cryopreservation. Methods-We used chromatin immunoprecipitation and quantitative polymerase chain reaction (PCR) to assay levels of transcriptionally permissive acetylated H3K9 and repressive trimethylated H3K9 histone modifications surrounding the transcriptional start site for IDO1, and reverse transcriptase PCR and immunoblotting to detect mRNA and protein. Results-MSCs derived from three donors approached maximum IDO1 mRNA levels following 24 hours of in vitro cytokine treatment. Induction of IDO1 expression correlated with increased acetylation of H3K9 concomitant with reduction of trimethylated H3K9 modifications at the promoter. Examination of two additional donors confirmed this result. While induced IDO1 levels declined within two days after cytokine removal and freeze thawing, the activated chromatin state was maintained. Upon re-exposure to cytokines, previously primed MSCs accumulated nearmaximum IDO1 mRNA levels within four to eight hours. Discussion-Our data indicate that in vitro priming of MSCs causes chromatin remodeling at the IDO1 promoter, that this alteration is maintained during processing commonly used to prepare MSCs for clinical use, and that once primed, MSCs are poised for IDO1 expression even in the absence of cytokines.
doi:10.1016/j.jcyt.2016.05.017 pmid:27421739 pmcid:PMC4983509 fatcat:6woj5gzfgfgptmcrjiyqbl7vn4