Date of Award

2016

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Pharmacology and Experimental Therapeutics

Abstract

Changes in gene expression and protein levels are an important aspect of cardioprotection in which short non-coding RNA known as miRNA may play a key regulatory role. We investigated the functions of several miRNAs in the context of two cardioprotective stimuli, ischemic preconditioning (IPC) and mesenchymal stem cell (MSC) paracrine effects. We hypothesized that downregulation of a set of miRNAs (miR-148a/b, miR-30b, and let-7a*) augments expression of protective heat shock proteins during IPC, and that MSC exosomes transfer miR-21 to cardiomyocytes, resulting in downregulation of pro-apoptotic genes and reduction of infarct size.

IPC increased the level of Hsp70, Hsp90, and Hsp40 family members within 6 hours as measured by qPCR and Western blot. Luciferase reporter assays and miRNA mimic transfection and knockdown were used to confirm effects of miR-148a/b, miR-30b, and let-7a* on translation. Combinations of miRNAs had more pronounced effects than single miRNAs alone. Pretreatment with wild type exosomes, but not those lacking miR-21, reduced cell death in vitro, and decreased infarct size in mice. The wild type exosomes, and miR-21 mimic, decreased protein levels of the miR-21 target genes Fas Ligand, Programmed Cell Death 4, Phosphatase and Tensin Homolog, and Pellino1.

In conclusion, a small set of miRNAs may act synergistically as regulatory nodes in a heat shock protein expression network after IPC. Future studies will test whether manipulation of this set of miRNAs can induce cardioprotection. miR-21 plays a key role in pro-survival paracrine effects mediated by MSC exosomes. Future studies will test whether MSC exosomes mediate regeneration as well as cardioprotection.

Creative Commons License

Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.

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