Understanding the miRNA response to opioid withdrawal and their uses as potential biomarkers for neonatal abstinence syndrome

点击翻译按钮获取中文摘要

Project Summary/Abstract Neonatal abstinence syndrome (NAS) is characterized by central nervous system hyperactivity that occurs when an infant experiences withdrawal from maternal opioid use at birth. Rates of NAS in the United States have skyrocketed amidst the opioid epidemic. Assessment and clinical management of NAS rely on subjective symptom scales, which may contribute to the prolonged hospital stays and poor neuro- developmental outcomes associated with NAS. Enhancing our knowledge about the molecular factors that regulate the biologic response to opioid withdrawal in developing infants will provide an opportunity to create objective clinical tests and novel therapies for this significant medical problem. To date, there is no biologic tool to determine the necessary morphine dose in withdrawing infants, or to predict which infants will experience neurodevelopmental delays. This is partly due to the fact that prior studies have largely focused on the molecular response to opioid administration in adults, rather than the response to opioid withdrawal in infants. Serum levels of certain micro-ribonucleic acids (miRNAs) are impacted by opioid administration in adults (e.g., let-7a, miR-146a, miR-192). These short, non-coding nucleic acids also regulate neurogenesis, neuronal progenitor cell (NPC) maturation, and cell survival by repressing target messenger RNAs in the Argonaute complex. My sponsor, Dr. Steve Hicks, MD, PhD has pioneered the use of salivary miRNAs as non-invasive markers for pediatric neurodevelopmental conditions. Most salivary miRNAs are derived from exosomes, which can arise from the cranial nerves that densely innervate the oropharynx. My preliminary data shows that several “opioid-responsive” miRNAs are perturbed in the saliva of infants with NAS relative to healthy infants. Further, I have developed an in vitro system of opioid withdrawal utilizing human NPCs that displays dose- related perturbations in miR-146a along with disruptions in NPC fate. Based on these findings, the central hypothesis of my fellowship application is that morphine withdrawal alters miRNA expression in the developing brain, which impairs neuronal maturation by mRNA translation via Argonaute binding. I also hypothesize that salivary levels of exosomal miRNAs from infants with NAS will be directly related to both the maximal dose of morphine required for symptom control, and neurodevelopmental outcomes at six months. I will test these hypotheses in two specific aims. First, I will perform a longitudinal cohort study of 50 infants with NAS to determine whether salivary miRNA levels within brain-related exosomes can be measured at admission to predict the maximum morphine dose required for symptom control, and again at discharge to predict neurodevelopmental outcomes at six months (Aim 1). Second, I will transfect miRNA mimics in my in vitro NPC design of opioid withdrawal, and assess the mechanism by which miRNAs impact the response to opioid withdrawal with immunocytochemistry, single-cell RNA sequencing, and an Argonaute pull-down assay (Aim 2).