Investigating the unconventional cytokine, interleukin-32 in chronic inflammation

This is a summer student position supervised by Rose Hodgson in Zaeem Cader's lab.

Introduction to the science

The Cader lab is interested in the interplay between the immune system and metabolism in inflammatory disease. The incidence of autoimmune diseases such as Inflammatory Bowel Disease is rising worldwide, with no curative treatments currently available for IBD. An improved understanding of disease processes is urgently required to help identify more effective therapies that actively interrupt disease pathogenesis.

Chronic inflammatory diseases such as IBD are driven by a dysregulation of the immune system leading to excess inflammation and tissue damage. Immune homeostasis is maintained through the concerted action of numerous immune cell populations; for example, intraepithelial lymphocytes and tissue resident macrophages interact to maintain tolerance to foreign material across tissue environments. Such functions are energetically expensive and thus are critically dependent on cellular nutrient utilisation and metabolism. An over-looked aspect is how such immune cells sustain necessary energy metabolism, and consequently maintain immune homeostasis, under the relative hypoxic conditions of the intestine.

About the project

All cells, including immune cells, must sense and adapt to changing oxygen levels, such as the hypoxic environment of the gut. Recent genetic studies have highlighted that failure to appropriately respond to hypoxia potentially contributes to the development of chronic inflammatory conditions such as Inflammatory Bowel Disease. However, the mechanisms accounting for this are yet to be elucidated. Our lab is studying a novel, disease-linked oxygen sensing pathway that could explain the relationship between oxygen adaptation and immunoregulation. One of the targets of this pathway is the unconventional cytokine, IL-32. However, this has been extremely poorly characterised to date and barely anything is known regarding its mechanism of action.

Interleukin-32 (IL32) is an unconventional cytokine that is primarily produced by NK and T cells, however induced in many cell types after stimulation. Altered expression of IL32 is reported to be associated with inflammatory diseases including rheumatoid arthritis, dermatitis, and inflammatory bowel disease, as well as in cancer. Regulated by hypoxia, IL32 is implicated in proinflammatory signalling, macrophage differentiation and also in regulation of metabolism in plasma cells. There are multiple isoforms of IL32, however their regulation and action is unclear, and it is also unknown how IL32 is secreted, or which receptor is targeted. This project would contribute to addressing the research gap in understanding the role of IL32 in immune homeostasis, inflammation and disease, information which could provide value in bettering therapies that target disease pathogenesis rather than the symptoms.

Specifically, we plan to explore the different isoforms of IL32 in these different cell types and inflammatory conditions. To do this, the student would culture relevant cell lines and primary cells in vitro and use qPCR and western blots to characterise the expression of IL32 isoforms under different stimuli. Such stimuli could include hypoxia, LPS, and activating or tolerogenic cytokine conditions. The project aims to characterise the function of individual IL32 isoforms using genetic modification by CRISPR-Cas9 to knockout the IL32 gene, followed by subsequent addition of individual isoforms in vitro. Here the student may learn how to use genetic modification techniques such as CRISPR, sanger sequencing and molecular cloning. The student may need to explore co-cultures of different cell types from these genetic models to characterise the action of individual IL32 isoforms produced by specific cell types in an intrinsic and extrinsic manner. Finally, based on project progress, we may want to use other techniques to consider the function of IL32, which may include flow cytometric cell sorting, RNASeq and metabolic profiling assays. Ultimately, this project will shed light on the role of IL32 isoforms in inflammatory signalling in immune mediated diseases.

Candidate background