Characterizing Regulatory T Cells in the Intestine
Dr. Krausgruber worked in Dr. Fiona Powrie's lab investigating Treg adaptation and specialisation in different tissues. The Powrie lab studies the complex relationship between our immune system and the resident community of commensal bacteria in the gut. Specifically, their research focuses on dissecting the cellular and molecular components of these interactions, and how they become perturbed during the onset and progression of chronic conditions such as ulcerative colitis and Crohn's disease; commonly referred to as inflammatory bowel disease (IBD). The lab uses a combination of intestinal inflammation model systems and human samples to study the innate and adaptive immune responses involved in IBD with a dual emphasis on basic and translational science.
- Role of Tregs in maintaining intestinal homeostasis
- Influence of microbiome in host immunity
1. What made you choose scientific research as a career path?
I was interested in biology from very early on, and I really enjoyed watching all kinds of documentaries as a child, particularly those with David Attenborough. Furthermore, I always wanted to understand and find out how things work on a mechanistic level.
This interest was further fostered during graduate studies; I obtained my MSc in Genetics and Molecular Biology from University of Salzburg, Austria, and then applied for a PhD position with Professor Irina Udalova at Imperial College London. My PhD thesis was dedicated to human macrophage polarization, and I discovered that the transcription factor IRF5 (interferon regulatory factor 5) plays a pivotal role in promoting a pro-inflammatory macrophage phenotype. During my PhD I realised that I really enjoy research and that I want to follow a research-related career.
2. Who is your scientific idol?
Although I am now working on aspects of adaptive immunity, my “upbringing" in the macrophage field has largely influenced me. One scientist with a huge impact on this area is Siamon Gordon, Professor Emeritus of Cellular Pathology at the University of Oxford. He is best known for his work on the phenotypic and functional diversity of macrophages including the identification of the pan-macrophage marker F4/80 as well as other surface receptors such as Dectin-1.
3. What was your role in the Powrie lab?
Previous work in the Powrie lab has shown that functionally specialised populations of Treg cells play a non-redundant role in maintaining intestinal homeostasis. Although these cells have been extensively studied in various organs and experimental settings, there are still questions regarding how intestinal Tregs carry out their function in the challenging intestinal microenvironment.
I was interested in cell plasticity, specifically which environmental cues induce Treg adaptation to different tissues and which transcription factors ultimately enforce these processes. Based on a whole genome gene expression analysis comparing Tregs from secondary lymphoid organs to colonic Tregs, we identified several differentially expressed effector proteins. One of these proteins was the interleukin-33 receptor (IL-33R). My project focused on the characterisation of IL-33R-expressing Tregs in the colon and how downstream signals might be important for cell adaptation and specialisation of these cells in the intestine.
1. In your opinion, what has been the most important advancement in the field in the last five years?
I think studies relating to microbiome research have attracted a lot of interest recently, both academically as well as in the mainstream media. Usually the microbiome co-exists in a symbiotic manner with the host but a breakdown of this mutual relationship is thought to be linked to severity of many diseases, including chronic intestinal inflammation. The identification of a range of bacterial strains that preferentially promote the accumulation of Tregs in the colon but also of those bacteria that are linked to disease progression, if allowed to grow abnormally, have opened up new ways of assessing as well as potentially treating intestinal inflammation.
Furthermore, there is now a new wave of drugs at various stages of clinical trials (e.g. Mongersen from Celgene or Ustekinumab from Janssen Biotech) that could add to the current standards therapies (broad immune-suppressive and anti-TNFɑ), leading to the broadening of therapeutic targets and ultimately enhancing the chances of treatment success.
2. Where is mucosal research heading?
I think a lot of effort will be put into patient stratification and developing a more tailored/personalised treatment regime. This will of course heavily depend on the identification of robust, reliable and easy-to-test biomarkers. For example, there are a relatively high number of non-responders to standard IBD biologics and it would be very beneficial if we could predict whether an individual is likely to respond to the current standard or if alternative treatment would be a better option. This would ensure that patients receive the most suitable medication early on and furthermore, the burden on health care systems would be reduced. The Powrie lab actually has a very promising project that has identified a potential novel IBD biomarker and it will hopefully be tested in a clinical setting soon.
3. What are the technical challenges currently facing the mucosal immunology field?
Microbiome research is a very hot topic at the moment. However, most of the studies investigating the microbiome use mice as a model system, and projects comparing human healthy individuals to those with various diseases are only starting to reveal differences. The big challenge will be moving from basic observations (composition of microbiota linked to person's disease development/progression) to unraveling how bacteria influence host immunity on a mechanistic level, and ultimately use this knowledge to develop new therapeutic interventions. Hopefully, initiatives such as the Human Microbiome Project and MetaHit (Metagenomics of the Human Intestinal Tract) will be successful in this regard.
1. Why does your lab use EasySep™?
The Powrie lab has been very satisfied with the quality of the reagents. We adopted for our cell isolation protocols primarily because of the speed but we also benefit from time-savings when FACS-sorting cells thanks to the high purity of cells isolated with EasySep™. Additionally, the protocol is really straight-forward so it is easy for new group members to use the EasySep™ system.
2. How has EasySep™ enabled your lab’s research?
Before , we used in-house purified antibody cocktails for the enrichment of mouse CD4+ T cells, which is a routine process in the lab and is usually followed by FACS sorting of specific T cell subsets. However, we started looking into alternatives due to batch-to-batch variability of these antibodies and a rather long cell isolation protocol. Cell isolation with the EasySep™ system is really quick. Especially when working with intestinal Tregs, which are known to be fragile, time is a key parameter.
We initially only used the mouse CD4+ T cell isolation kit, but we started to use kits for the isolation of various cell types of both mouse and human origin. This allows the lab to save time and take advantage of purer cell populations in a wider variety of research projects. Also, the system is very flexible and allows for easy up-scaling when setting up larger experiments.
On the role of Tregs in maintaining intestinal homeostasis
Regulatory T cells (Tregs) are a subset of immune cells that suppresses inflammation and autoimmunity. Due to their potent, antigen-specific suppressive effects, Tregs have been proposed as a candidate for immune therapy in inflammatory diseases, including inflammatory bowel disease (IBD)1. In this field, the Powrie lab has been a pioneer who first reported the ability of Tregs to cure colitis in mice. The Powrie lab recently discovered that a large proportion of Tregs in the colon express ST2 (IL-33R), and that IL-33 promotes their stability and function2. Other studies have also elucidated the molecular mechanisms of Treg function in the maintenance of intestinal homeostasis. For example, the expression of IL-103 and OX404 by Tregs have been identified as important factors in controlling colitis. By manipulating Tregs in vivo, these studies have continually contributed to discoveries of new potential therapeutic targets for IBD. A recent study by Antignano et al suggested that one potential therapy may be the inhibition of G9a, a methyltransferase which negatively regulates Treg differentiation5.
On the influence of microbiome in host immunity
The microbiota is composed of ~100 trillion cells that reside mainly in the intestine. These commensals have been suggested to contribute to the development and regulation of host immunity, as well as to the pathogenesis or protection against various diseases, including IBD, cancer and diabetes6. In the past few years, several studies have proposed mechanisms behind the ability of the microbiome to promote immune regulation. Atarashi et al. suggested that Clostridium species in the microbiome promote the accumulation of suppressive Tregs7. Interestingly, a few studies have suggested that the induction of intestinal Tregs by the microbiome is mediated by commensal-produced short-chain fatty acids, such as butyrate8,9. Although these findings propose the use of microbial components for potential therapies against diseases such as IBD, much work remains to be done to investigate whether these observations can be translated to human disease.
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- Himmel ME et al. (2012) Regulatory T-cell therapy for inflammatory bowel disease: more questions than answers. Immunology 136(2): 115–122.
- Schiering C et al. (2014) The alarmin IL-33 promotes regulatory T-cell function in the intestine. Nature 513(7519): 564–568.
- Schmitt EG et al. (2012) IL-10 produced by induced regulatory T cells (iTregs) controls colitis and pathogenic ex-iTregs during immunotherapy. J Immunol 189 (12 ): 5638–5648.
- Griseri T et al. (2010) OX40 is required for regulatory T cell-mediated control of colitis. J Exp Med 207 (4 ): 699–709.
- Antignano F et al. (2014) Methyltransferase G9A regulates T cell differentiation during murine intestinal inflammation. J Clin Invest 124(5): 1945–1955.
- Belkaid Y & Hand TW. (2015) Role of the microbiota in immunity and inflammation. Cell 157(1): 121–141.
- Atarashi K et al. (2011) Induction of colonic regulatory T cells by indigenous Clostridium species. Sci 331 (6015): 337–341.
- Furusawa Y et al. (2013) Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells. Nature 504(7480): 446–450.
- Smith PM et al. (2013) The microbial metabolites, short-chain fatty acids, regulate colonic Treg cell homeostasis. Science (80- ) 341(6145): 569–573.
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Thomas Krausgruber, PhD