A proteomic atlas of the human umbilical cord across gestation reveals fetal immune maturation.

INTRODUCTION

Prematurity remains a leading cause of neonatal morbidity and mortality, yet the molecular mechanisms underlying functional immaturity at birth are poorly understood. To our knowledge, no study has conducted a proteomic analysis of whole umbilical cords spanning a full range of gestational ages, including infants born at the periviable gestational age (22-25 weeks). Identifying protein expression patterns that correlate with gestational maturity may offer key insights into the molecular drivers of vulnerability in preterm neonates, particularly in the domains of immune and organ development. In this study, we applied data-independent acquisition (DIA)-based mass spectrometry to profile the proteomes of umbilical cords from neonates born between 22 and 40 weeks of gestation. Our aims were to establish the feasibility of this approach and to characterize gestational age-dependent protein expression patterns with relevance to neonatal development and immune maturation.

METHODS

Umbilical cord samples were obtained from neonates admitted to the Neonatal Intensive Care Unit of the University of Tokyo Hospital. Fresh umbilical cord segments approximately 5 cm in length were collected immediately after delivery. Subsequently, the cords were sectioned into 1 cm fragments, snap-frozen at -80°C, and stored until DIA proteomic analysis. For quantitative proteomics data obtained by DIA, gene symbols were assigned to each protein, and the expression values were transformed into z-scores within each sample. To evaluate temporal changes across four time points, we performed time-course analysis. The Jonckheere-Terpstra test was used to assess the overall monotonic trend across the four time points within each gene set. Gene Ontology (GO) enrichment analysis was performed using Metascape for each expression pattern obtained above.

RESULTS

A total of 6,801 proteins were identified from umbilical cord tissues collected from 15 neonates divided into four gestational age groups: 22-25 weeks (n = 4, median birth weight 634 grams (interquartile range [IQR]: 468.5-690)), 26-30 weeks (n = 4, median birth weight 1008 g (IQR: 678.5-1140.8)), 31-34 weeks (n = 3, median birth weight 1521 g (IQR: 1389-1600)), and 35-40 weeks (n = 4, median birth weight 2477 g (IQR: 1858-2677.3)). Proteins involved in immune responses, especially those mediating B cell function, were significantly upregulated with advancing gestational age, indicating progressive immune system maturation (22-25 weeks vs 35-40 weeks, p < 0.01). In contrast, proteins related to mRNA processing, cytoplasmic translation, and ribonucleoprotein complex biogenesis showed inverse trends, suggesting elevated biosynthetic activity in extremely preterm neonates (22-25 weeks vs 35-40 weeks, p < 0.01). Protein markers associated with organ development exhibited heterogeneous trajectories.

CONCLUSION

These findings demonstrate that umbilical cord proteomics can serve as a powerful platform for assessing functional maturity at birth-particularly of the immune system-and may facilitate the discovery of biomarkers and therapeutic targets relevant to neonatal care. The study's limitations include the small sample size and the use of whole umbilical cord rather than isolated tissue compartments. This work establishes the first proteomic atlas of human umbilical cord tissue across gestation and provides a foundation for molecularly informed strategies in the management of preterm infants.