Intraventricular
hemorrhage (IVH) represents a high risk of neonatal mortality and later neurodevelopmental impairment in prematurity. IVH is accompanied with
inflammation,
hemolysis, and extracellular
hemoglobin (Hb) oxidation. However,
microRNA (
miRNA) expression in cerebrospinal fluid (CSF) of preterm infants with IVH has been unknown. Therefore, in the present study, candidate pro-inflammatory cell-free
miRNAs were analyzed in CSF samples from 47 preterm infants with grade III or IV IVH vs. clinical controls (n = 14).
miRNAs were quantified by RT-qPCR, normalized to "spike-in" cel-miR-39. Oxidized Hb and total
heme levels were determined by spectrophotometry as well as
IL-8,
VCAM-1,
ICAM-1, and
E-selectin concentrations by ELISA. To reveal the origin of the investigated
miRNAs, controlled
hemolysis experiments were performed in vitro; in addition, human choroid plexus epithelial cell (HCPEpiC) cultures were treated with metHb,
ferrylHb,
heme, or TNF-α to replicate IVH-triggered cellular conditions. Levels of miR-223, miR-155, miR-181b, and miR-126 as well as Hb metabolites along with
IL-8 were elevated in CSF after the onset of IVH vs. controls. Significant correlations were observed among the
miRNAs, oxidized Hb forms, and the soluble adhesion molecules. During the post-IVH follow-up, attenuated expression of
miRNAs and
protein biomarkers in CSF was observed upon elimination of Hb metabolites. These
miRNAs remained unaffected by a series of artificially induced
hemolysis, which excluded red blood cells as their origin, while stimulation of HCPEpiCs with oxidized Hb fractions and
heme resulted in increased extracellular
miRNA levels in the cell culture supernatant. Overall, the
hemorrhage-induced CSF
miRNAs reflected inflammatory conditions as potential
biomarkers in preterm IVH.