Alison G Paquette1, Oksana Shynlova2, Mark Kibschull3, Nathan D Price1, Stephen J Lye4. 1. Institute for Systems Biology, Seattle, WA. 2. Program in Development and Fetal Health, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto, Ontario, Canada; Department of Obstetrics and Gynecology, University of Toronto, Toronto, Ontario, Canada. 3. Program in Development and Fetal Health, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto, Ontario, Canada. 4. Program in Development and Fetal Health, Lunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto, Ontario, Canada; Department of Obstetrics and Gynecology, University of Toronto, Toronto, Ontario, Canada. Electronic address: lye@lunenfeld.ca.
Abstract
BACKGROUND: Preterm birth is the leading cause of newborn death worldwide, and is associated with significant cognitive and physiological challenges in later life. There is a pressing need to define the mechanisms that initiate spontaneous preterm labor, and for development of novel clinical biomarkers to identify high-risk pregnancies. Most preterm birth studies utilize fetal tissues, and there is limited understanding of the transcriptional changes that occur in mothers undergoing spontaneous preterm labor. Earlier work revealed that a specific population of maternal peripheral leukocytes (macrophages/monocytes) play an active role in the initiation of labor. Thus, we hypothesized that there are dynamic gene expression changes in maternal blood leukocytes during preterm labor. OBJECTIVE: Using next-generation sequencing we aim to characterize the transcriptome in whole blood leukocytes and peripheral monocytes of women undergoing spontaneous preterm labor compared to healthy pregnant women who subsequently delivered at full term. STUDY DESIGN: RNA sequencing was performed in both whole blood and peripheral monocytes from women who underwent preterm labor (24-34 weeks of gestation, N = 20) matched for gestational age to healthy pregnant controls (N = 30). All participants were a part of the Ontario Birth Study cohort (Toronto, Ontario, Canada). RESULTS: We identified significant differences in expression of 262 genes in peripheral monocytes and 184 genes in whole blood of women who were in active spontaneous preterm labor compared to pregnant women of the same gestational age not undergoing labor, with 43 of these genes differentially expressed in both whole blood and peripheral monocytes. ADAMTS2 expression was significantly increased in women actively undergoing spontaneous preterm labor, which we validated through digital droplet reverse transcriptase polymerase chain reaction. Intriguingly, we have also identified a number of gene sets including signaling by stem cell factor-KIT, nucleotide metabolism, and trans-Golgi network vesicle budding, which exhibited changes in relative gene expression that was predictive of preterm labor status in both maternal whole blood and peripheral monocytes. CONCLUSION: This study is the first to investigate changes in both whole blood leukocytes and peripheral monocytes of women actively undergoing spontaneous preterm labor through robust transcript measurements from RNA sequencing. Our unique study design overcame confounding based on gestational age by collecting blood samples from women matched by gestational age, allowing us to study transcriptomic changes directly related to the active preterm parturition. We performed RNA profiling using whole genome sequencing, which is highly sensitive and allowed us to identify subtle changes in specific genes. ADAMTS2 expression emerged as a marker of prematurity within peripheral blood leukocytes, an accessible tissue that plays a functional role in signaling during the onset of labor. We identified changes in relative gene expression in a number of gene sets related to signaling in monocytes and whole blood of women undergoing spontaneous preterm labor compared to controls. These genes and pathways may help identify potential targets for the development of novel drugs for preterm birth prevention.
BACKGROUND: Preterm birth is the leading cause of newborn death worldwide, and is associated with significant cognitive and physiological challenges in later life. There is a pressing need to define the mechanisms that initiate spontaneous preterm labor, and for development of novel clinical biomarkers to identify high-risk pregnancies. Most preterm birth studies utilize fetal tissues, and there is limited understanding of the transcriptional changes that occur in mothers undergoing spontaneous preterm labor. Earlier work revealed that a specific population of maternal peripheral leukocytes (macrophages/monocytes) play an active role in the initiation of labor. Thus, we hypothesized that there are dynamic gene expression changes in maternal blood leukocytes during preterm labor. OBJECTIVE: Using next-generation sequencing we aim to characterize the transcriptome in whole blood leukocytes and peripheral monocytes of women undergoing spontaneous preterm labor compared to healthy pregnant women who subsequently delivered at full term. STUDY DESIGN: RNA sequencing was performed in both whole blood and peripheral monocytes from women who underwent preterm labor (24-34 weeks of gestation, N = 20) matched for gestational age to healthy pregnant controls (N = 30). All participants were a part of the Ontario Birth Study cohort (Toronto, Ontario, Canada). RESULTS: We identified significant differences in expression of 262 genes in peripheral monocytes and 184 genes in whole blood of women who were in active spontaneous preterm labor compared to pregnant women of the same gestational age not undergoing labor, with 43 of these genes differentially expressed in both whole blood and peripheral monocytes. ADAMTS2 expression was significantly increased in women actively undergoing spontaneous preterm labor, which we validated through digital droplet reverse transcriptase polymerase chain reaction. Intriguingly, we have also identified a number of gene sets including signaling by stem cell factor-KIT, nucleotide metabolism, and trans-Golgi network vesicle budding, which exhibited changes in relative gene expression that was predictive of preterm labor status in both maternal whole blood and peripheral monocytes. CONCLUSION: This study is the first to investigate changes in both whole blood leukocytes and peripheral monocytes of women actively undergoing spontaneous preterm labor through robust transcript measurements from RNA sequencing. Our unique study design overcame confounding based on gestational age by collecting blood samples from women matched by gestational age, allowing us to study transcriptomic changes directly related to the active preterm parturition. We performed RNA profiling using whole genome sequencing, which is highly sensitive and allowed us to identify subtle changes in specific genes. ADAMTS2 expression emerged as a marker of prematurity within peripheral blood leukocytes, an accessible tissue that plays a functional role in signaling during the onset of labor. We identified changes in relative gene expression in a number of gene sets related to signaling in monocytes and whole blood of women undergoing spontaneous preterm labor compared to controls. These genes and pathways may help identify potential targets for the development of novel drugs for preterm birth prevention.