Cross-tolerance: embryonic heat conditioning induces inflammatory resilience by affecting different layers of epigenetic mechanisms regulating IL6 expression later in life

ABSTRACT

A stressor can induce resilience in another, different stressor, a phenomenon known as cross-tolerance. To learn if cross-tolerance is governed by epigenetic regulation, we used embryonic heat conditioning (EHC) in chicks, during the development of the hypothalamus, to increase the immunization response. Indeed, EHC induced a lifelong systemic antibody response to immunization, in addition to reduced hypothalamic IL6 inflammatory expression following LPS challenge. Since the outcome of EHC was long-term cross-tolerance with the immune system, we studied possible epigenetic mechanisms. We first analysed the methylation and hydroxymethylation patterns of IL6. We found reduced hydroxymethylation on IL6 intron 1 in the EHC group, a segment enriched with CpGs and NFkB-binding sites. Luciferase assay in cell lines expressing NFkB showed that IL6 intron 1 is indeed an enhancer. ChiP in the same segment against NFkB in the hypothalamus presented reduced binding to IL6 intron 1 in the EHC group, before and during LPS challenge. In parallel, EHC chicks’ IL6 intron 1 presented increased H3K27me3, a repressive translational modification mediated by EZH2. This histone modification occurred during embryonic conditioning and persisted later in life. Moreover, we showed reduced expression of miR-26a, which inhibits EZH2 transcription, during conditioning along with increased EZH2 expression. We demonstrate that stress cross-tolerance, which was indicated by EHC-induced inflammatory resilience and displayed by attenuated inflammatory expression of IL6, is regulated by different epigenetic layers.

KEYWORDS:

• Cross-tolerance
• epigenetic
• IL6
• H3K27
• EZH2
• miR-26a

Acknowledgments

We are grateful to the Volcani Center’s chicken farm staff for their dedicated work. We would also like to thank Dr. Yuval Cinnamon for his instruction and guidance regarding the extraction of chick embryo midbrain. Additional thanks to the staff of the Crown Institute for Genomics of the Nancy and Stephen Grand Israel National Center for Personalized Medicine, Weizmann Institute of Science, Israel, and especially to Gilgi Friedlander for her skilled bioinformatics work.

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

This work was supported by the Israel Science Foundation [1646/15].