Integrated transcriptome and methylome analysis in youth at high risk for bipolar disorder: a preliminary analysis

Abstract

First-degree relatives of patients with bipolar disorder (BD), particularly their offspring, have a higher risk of developing BD and other mental illnesses than the general population. However, the biological mechanisms underlying this increased risk are still unknown, particularly because most of the studies so far have been conducted in chronically ill adults and not in unaffected youth at high risk. In this preliminary study we analyzed genome-wide expression and methylation levels in peripheral blood mononuclear cells from children and adolescents from three matched groups: BD patients, unaffected offspring of bipolar parents (high risk) and controls (low risk). By integrating gene expression and DNA methylation and comparing the lists of differentially expressed genes and differentially methylated probes between groups, we were able to identify 43 risk genes that discriminate patients and high-risk youth from controls. Pathway analysis showed an enrichment of the glucocorticoid receptor (GR) pathway with the genes MED1, HSPA1L, GTF2A1 and TAF15, which might underlie the previously reported role of stress response in the risk for BD in vulnerable populations. Cell-based assays indicate a GR hyporesponsiveness in cells from adult BD patients compared to controls and suggest that these GR-related genes can be modulated by DNA methylation, which poses the theoretical possibility of manipulating their expression as a means to counteract the familial risk presented by those subjects. Although preliminary, our results suggest the utility of peripheral measures in the identification of biomarkers of risk in high-risk populations and further emphasize the potential role of stress and DNA methylation in the risk for BD in youth.

Figure 1

‘Risk genes' identified by the differential expression and methylation analyses. The lists of DEGs (a) and DMPs (b) between controls versus BD patients and between controls versus offspring were compared to identify concordant genes. Thirty-three genes were concordant in the gene expression analysis, while 18 probes were concordant in the methylation analysis (annotated to 10 genes). (c) Correlation between gene expression and methylation at the MED1 and TAF15 genes. (d) Expression levels of GR signaling pathway risk genes. (e) Correlation between expression levels and family cohesion scores for the GR signaling pathway risk genes. (f) Correlation between expression levels and family conflict scores for the GR signaling pathway risk genes. **P<0.01; ***P<0.001 when compared to controls. Blue spots represent values for controls, green for offspring and red for patients. BD, bipolar disorder; DEG, differentially expressed gene; DMP, differentially methylated probes; GR, glucocorticoid receptor.

Figure 2

Connections between ‘risk genes' identified in our analysis and molecules previously reported to be associated with bipolar disorder. Molecules painted in blue are ‘risk genes' identified in our analysis, whereas those in grey are known to have been associated with bipolar disorder (available at IPA Knowledge Base). Relationships are based on expression, protein–protein binding, protein–DNA binding, microRNA targeting, activation or transcription. Dashed lines represent indirect relationships (where the two molecules do not need to physically interact). IPA, Ingenuity Pathway Analysis.

Figure 3

Effects of 5-aza-2'-deoxycytidine treatment in lymphoblastoid cells from adult patients with bipolar disorder and controls. Cells were treated for 96 h (1 or 5 μM). (a) Cell viability assessed by MTT assay; (b) 5-methylcytosine (%) level; (c–f) mRNA levels for MED1, TAF15, HSPA1L and GTF2A1. *P<0.05, **P<0.01, and ***P<0.001 when compared to vehicle treatment in the same group. ^#P<0.05 when compared to the same treatment in the control group. MTT, (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium.

Figure 4

Glucocorticoid receptor-responsive gene expression after stimulation with dexamethasone. Lymphoblastoid cells from adult patients with bipolar disorder and controls were treated with 10⁻⁷ M for 4 h (a–c) or 48 h (d–f) and the expression of glucocorticoid receptor-responsive genes was measured (TSC22D3, PER1 and FKBP5). The levels of these genes were also measured by real-time quantitative PCR in PBMCs from children and adolescents that are healthy controls, unaffected offspring of parents with bipolar disorder and patients with pediatric bipolar disorder. *P<0.05 compared to vehicle in the same group. DEX, dexamethasone; PBMC, peripheral blood mononuclear cell; PCR, polymerase chain reaction.