Identifying a gene expression signature of cluster headache in blood

Abstract

Cluster headache is a relatively rare headache disorder, typically characterized by multiple daily, short-lasting attacks of excruciating, unilateral (peri-)orbital or temporal pain associated with autonomic symptoms and restlessness. To better understand the pathophysiology of cluster headache, we used RNA sequencing to identify differentially expressed genes and pathways in whole blood of patients with episodic (n = 19) or chronic (n = 20) cluster headache in comparison with headache-free controls (n = 20). Gene expression data were analysed by gene and by module of co-expressed genes with particular attention to previously implicated disease pathways including hypocretin dysregulation. Only moderate gene expression differences were identified and no associations were found with previously reported pathogenic mechanisms. At the level of functional gene sets, associations were observed for genes involved in several brain-related mechanisms such as GABA receptor function and voltage-gated channels. In addition, genes and modules of co-expressed genes showed a role for intracellular signalling cascades, mitochondria and inflammation. Although larger study samples may be required to identify the full range of involved pathways, these results indicate a role for mitochondria, intracellular signalling and inflammation in cluster headache.

Figure 1. RT-qPCR validation of genes differentially expressed between cluster headache patients and controls.

Four genes were chosen from the top 15 differentially expressed genes between participants with cluster headache and controls. Data were normalized to TBP mRNA expression. (a) Gene expression measured by RT-qPCR in the cluster headache group relative to the expression in the control group. (b) Comparison of fold-change of gene expression in participants with cluster headache relative to controls measured by RNA-seq and RT-qPCR. Means ± SD, *P < 0.05; **P < 0.01.

Figure 2. Gene co-expression modules in whole blood associated with cluster headache.

Detection of co-expression modules in RNA-seq data of whole blood samples of cluster headache patients and controls using weighted gene co-expression network analysis (WGCNA). (a) The cluster dendrogram that gave rise to 40 modules, indicated by different colours. (b–g) Bar graphs of the module eigengenes in controls, episodic (ECH) and chronic cluster headache (CCH) of the six modules associated with cluster headache. The module eigengene can be considered the optimal summary of the module-specific gene expression pattern. Top three pathways significantly overrepresented in each module are listed below each bar graph. P = P-value, C = count of genes in the module belonging to the pathway.