Analysis of Single-Cell Expression Data of Liver Sinusoidal Endothelial Cells Reveals Strong Variability of F8 Expression Associated with Specific Expression Profile

Introduction Hemophilia A (HA) is an X-linked genetic disorder caused by reduced or no activity of coagulation FVIII gene (F8). In the early 1990s, F8 was known to be expressed/secreted by hepatocytes, whereas now it has been well established that FVIII is mainly produced by liver sinusoidal endothelial cells (LSECs). The LSECs are cells spread across the different liver lobules and show heterogeneity in their functional characteristics.

Aim To understand the molecular basis of variable F8 expression in different population of LSECs.

Method To understand the mechanism (genes and pathway involved) underlying F8 expression in LSECs, we analyzed single-cell (SC) data of human liver downloaded from NCBI GEO “GSE124395” data set. The LSECs were filtered out from the whole liver data. Data were analyzed using RaceID package in R. Pathway analyses were carried out using ingenuity pathway analysis (IPA) and gene ontology analyses were carried out using Bingo and EnrichmentMap in Cytoscape v3.7.1.

Results Single-cell data showed that F8 expressed in a range from 0 to 12 (expression level units) in LSECs. The majority of LSECs showed no F8 expression (NoF8) and only 85 out of 260 LSECs showed F8 expression. Clustering of SC data resulted in two major clusters of F8 expressing LSECs and non-expressing LSECs. The F8 expressing LSECs cluster showed variability in F8 expression levels. Differential gene expression analysis of NoF8—low expressing F8 (LowF8) and high expressing F8 (HighF8)—showed 2,567 and 523 genes, respectively, at p < 0.05. The NoF8 LSECs were found to be enriched in immune response–related genes, whereas F8 expressing LSECs were enriched in genes related to biological regulation and hematopoiesis. Similarly IPA analysis showed enrichment of signaling pathways-related genes in HighF8 expressing LSECs when compared to NoF8 or LowF8 LSECs. We also found 757 genes significantly correlated with F8 expression at 1% FDR (false discovery rate). Regulatory networks of these genes showed inhibition of bleeding, activation of hematopoiesis, endocytosis, and homeostasis-related functions.

Conclusion We conclude that LSECs are heterogeneous cell populations and not all LSECs express F8 at same level. Further follow-up studies directed by our outcomes are required to understand the mechanism of F8 expression.

Publication History

Article published online:
13 November 2020