TRANSCRIPTOMIC PROFILING REVEALS A SEVERITY-ASSOCIATED SIGNATURE IN HYPOXIC-ISCHAEMIC ENCEPHALOPATHY

Despite hypoxic ischaemic encephalopathy (HIE) being a major cause of neonatal morbidity and mortality, mild cases remain difficult to clinically classify, risking misclassification and under- or overtreatment. Clarifying its molecular basis may improve diagnostic accuracy and allow for targeted neuroprotective therapies.

Next-generation RNA Sequencing was used to identify an early whole-blood transcriptomic signature of HIE severity. Neonates with mild (n=39), moderate/severe (n=35) HIE defined using Expanded Modified Sarnat staging, and healthy controls (n=40) (≥36 weeks’ gestation, ≥1.8 kg) were enrolled. Blood samples were collected within 6 hours of birth and prior to whole-body hypothermia for moderate/severe HIE (mild HIE were not cooled). Differential expression used quasi-likelihood negative binomial models adjusted for batch, sex, birthweight and gestational age (FDR q<0.05 and
|log₂FC|>1). A novel Disease Risk Score algorithm removed highly correlated transcripts to simplify classification and functional enrichment was conducted via Database for Annotation, Visualization and Integrated Discovery.

Mean (SD) birthweight was 3.45 (0.42) kg and gestational age 39.0 (1.48) weeks. 1259 differentially expressed genes in mild HIE and 852 in moderate/severe HIE were identified, with minimal overlap (47). Mild HIE enriched ER protein processing p=5.18e-4) and TNF signalling (p=1.95e-3); moderate/severe HIE enriched neutrophil extracellular trap formation (p=1.01e-7) and cell-cycle pathways (p=1.95e-5). A 35-gene set from 469 Bonferroni-corrected genes defined an early transcriptional signature of severity (Figure 1), supported by Disease Risk Scores increasing with clinical severity. Early postnatal gene expression profiling distinguishes mild HIE from controls and moderate/severe HIE and could improve clinical stratification and aetiology prediction.