This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our overall objectives are to understand the role of microRNAs during pluripotency, differentiation and pathology in cynomolgus macaques as an accessible, important and feasible model. This pilot project complements other on-going studies on miRNA regulation in pluripotent and differentiated stem cells in vitro as well as parallel studies on miRNA antagonist administration into mouse lungs. The complexity of the lung suggests that lung development involves the expression of many genes. Today, one can use microarrays to quickly study gene expression in the both the lung as well as its associated pathologies. However, most of our information relies on rodent models. This study will attempt to bridge the gap between rodent and human development by utilizing macaques as a research model. Micro RNAs (miRNAs) are being shown to be involved in many cellular processes such as cell cycle regulation, development timing, signal transduction, tissue differentiation and disease. miRNAs have also been proposed to regulate pluripotency and differentiation in vitro and in vivo. One such miRNA, let-7d, is highly expressed in lung cells in patients with Idiopathic Pulmonary Fibrosis (IPF). Recently, we have conducted a number of in vivo studies in mice, in which we have injected a short RNA sequence (antagomir;purchased from Dharmacon - part of Thermo Fisher Scientific Inc.) consists of a short RNA sequence that binds to the miRNA and causes it to undergo degradation. The antagomir was injected intra tracheal to mice lungs. We found an increase in the levels of mesenchymal markers in the lung, indicating that the antagomirs not only entered the lung epithelia, but could cause the alteration of gene expression in the lung. In this study, we propose to administer this let-7 antagomir into one lung of a monkey with the other as a control in which a scrambled miRNA is delivered to determine if the same process occurs in the NHP model as in the mouse. The oligo is very small, and should regionalize to the small airways very quickly. Although definitive studies have not been done to determine exactly how quickly the oligo is regionalized, this is not an anticipated problem. Additionally, we propose to explore the intravenous administration (IV) vs. intra-bronchial. The significance of such experiments is not only to study the ontology of disease in a model closely related to the human, but also as a step in developing treatment for an incurable disease and the molecular mechanisms of normal development.
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