VPB0250 - Heme regulates inflammatory signaling pathways in human choroid plexus epithelial cells after preterm intraventricular hemorrhage

Sunday, July 10, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: Virtual

Presenting Author(s)

Zsolt Fejes, PhD

research fellow
Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary

Co-Author(s)

MP

Marianna Pócsi

PhD student
Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
�G

Ágnes Gyetvai

Research fellow
Department of Internal Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
SP

Szilárd Póliska

Research fellow
Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
AT

Andrea Tóth

PhD student
University of Debrecen
Debrecen, Hajdu-Bihar, Hungary
EB

Enikő Balogh

Junior research fellow
Research Centre for Molecular Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
�R

Ágnes Rusznyák

Junior research fellow
Department of Pharmaceutical Technologies, Faculty of Pharmacy, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
FF

Ferenc Fenyvesi

associate professor
Department of Pharmaceutical Technologies, Faculty of Pharmacy, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
VJ

Viktória Jeney

senior research fellow
Research Centre for Molecular Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
JK

János Kappelmayer

professor
Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary
BN

Béla Nagy Jr., Jr., MD, PhD

associate professor
Department of Laboratory Medicine, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
Debrecen, Hajdu-Bihar, Hungary

Background: Upon preterm intraventricular hemorrhage (IVH), released heme can induce inflammation, cell activation, and dysfunction of choroid plexus epithelial cells of the ventricles. We have recently reported that coculturing of cerebrospinal fluid samples derived from these patients with human choroid plexus epithelial cells (HCPEpiCs) provoked inflammatory cellular response in vitro causing altered expression of some selected miRNAs.

Aims: In this study, our aim was to further investigate the genetic background of heme-induced inflammatory mechanisms in HCPEpiCs.

Methods: To observe the direct effect of hemorrhage, HCPEpiCs were exposed to heme (25 μM) or TNF-α (100 ng/mL) as a classic pro-inflammatory agonist for 24 hours. The consequence of heme treatment on mRNA expression was investigated by RNA-seq. In parallel, we profiled miRNAs using TaqMan OpenArray panels and Cytoscape-ClueGO was used for functional enrichment analysis. Concentrations of released proteins in cell cultures were measured by ELISA, while the activation of NF-κB pathway was observed by fluorescence microscopy.

Results: Similarly to TNF-α, heme induced the NF-κB pathway via nuclear translocation of the p65 subunit. As a result, there were 231 significantly upregulated, and 106 attenuated mRNAs in heme treated HCPEpiCs compared to untreated cells. Among of the top 50 differentially expressed genes HMOX1, IL8, IL1B and ICAM1 mRNAs were upregulated in association with several mechanisms, such as cell response to cytokine activity and chemical stress, TLR4 mediated signaling and regulation of apoptosis based on ClueGO analysis. Furthermore, significantly elevated concentrations of IL-8 and ICAM-1 were detected in cell culture supernatants. Out of 358 detected miRNAs, 25 significantly decreased (e.g. miR-223, miR-215 and miR-9), and 10 were upregulated (e.g. miR-181c) in stimulated cells, which participate in the development of oxidative stress-induced cell death.

Conclusion(s): In preterm IVH, heme has important regulatory functions in choroid plexus epithelial cells that may contribute to the development of clinical complications in affected infants.