Theodore Steiner
Relevant Thesis-Based Degree Programs
Graduate Student Supervision
Doctoral Student Supervision
Dissertations completed in 2010 or later are listed below. Please note that there is a 6-12 month delay to add the latest dissertations.
Inflammatory bowel disease (IBD) consists of two diseases, ulcerative colitis (UC) and Crohn’s disease (CD), both characterized by chronic inflammation in the gastrointestinal tract. Some of the strongest risk alleles, such as NOD2, ATG16L1, and XBP1, affect the functionality of intestinal epithelial cells (IECs). Recent evidence suggests that IECs from IBD patients have long lasting molecular rewiring of the intestinal cellular compartment, with inflammatory gene signatures persisting once inflammation is resolved. Due to the high metabolic burden of their physiologic requirements, IECs are prone to endoplasmic reticulum (ER) stress, which leads to the unfolded protein response (UPR)—a cellular protection pathway that restores translational homeostasis and removes irreversibly stressed cells. Since ER stress has been shown to contribute to the pathogenesis of IBD, I sought to understand how ER stress can drive anti-microbial immune responses, as well as its ability to alter intestinal stem cell (ISC) self-renewal and IEC de-differentiation to recover from ER stress induced damage. I found that ER stress alters the ability of IECs to respond to TLR5 agonist, bacterial flagellin, ultimately leading to the maturation of monocyte derived dendritic cells (moDCs). This mechanism would ultimately link IEC dysfunction to anti-commensal adaptive immunity, which is what is observed in IBD. I also found that IBD-derived IECs display persistent ER stress, which alters their ability to respond to FliC. To understand how ER stressed induced chronic damage alters IEC function, I created a new chronic damage model using repeated cycles of air-liquid interface (ALI) and submergence injury of human derived colon organoid monolayers (colonoids). Through repeated damage by submerging colonoids in culture media, I show significant changes in epithelial cell lineages and gene expression that correlate with epigenetic modifications similar to that described for inflammation-associated GI diseases such as IBD and cancer.
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Inflammatory Bowel Disease (IBD) is life-changing because of recurrent intestinal inflammation. Current therapies are associated with mild to severe side effects, and none provide a cure. Recent research has provided pre-clinical and clinical data on cell-based therapy using the two best-characterized types of T regulatory cells, Foxp3⁺ Tregs and Foxp3⁻ Type 1 regulatory (Tr1) cells. However, major hurdles exist. The ability of Tregs to regulate innate immunity is not well understood, and while high numbers of antigen-specific Tregs are needed, these cells are scarce. Studies showed that engineered chimeric antigen receptor-expressing Tregs (CAR-Treg) could curb intestinal inflammation, but these CARs were not relevant to human disease. CAR-Treg therapy is logistically and conceptually complex and patients might perceive an unacceptable risk to be associated with this therapy. Finally, people living with IBD have an increased risk of Clostridioides difficile infection (CDI), the most prevalent cause of nosocomial infectious diarrhea in Canada, and patient experiences with CDI have not been researched. The purpose of this thesis was to elucidate how Tr1 cells and/or Tregs regulate innate immunity, which I addressed by investigating the suppressive effect of Tr1 and Tregs on the inflammasome. I then developed and tested a new CAR-Treg relevant to human IBD and conducted a survey to investigate patients’ willingness to try CAR-Treg. Finally, I analyzed a survey to describe the impact of CDI on patients in Canada. My results demonstrate that Tr1 cells may have unique therapeutic effects in reducing inflammasome activation via Interleukin 10. I provide evidence that CAR-Tregs suppress proinflammatory T cells. People with IBD indicated high willingness to try CAR-Treg therapy in both a clinical trial and as a new treatment. Willingness to try was not correlated with disease state or medication history. Finally, CDI patients highlighted the symptom-related impact and long-lasting effect on quality of life. Patient priorities to attenuate impact include reducing time to diagnosis and improving patient education. My research has implications on future development of Treg-based therapy for IBD and other immune-mediated diseases and demonstrates the promise of moving this therapy into clinical practice, as most patients indicated willingness to try.
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During chronic inflammation and tissue injuries, various danger-associated molecules can be released and are able to potentiate inflammation and T cell responses. Among the many possible danger signals, I focused on studying high concentrations of extracellular ATP since it has been implicated in a variety of autoinflammatory diseases. ATP activates the inflammasome in macrophages, stimulates dendritic cell (DC) maturation, and inhibits regulatory T cell (Treg) function. However, how ATP regulates Toll-like receptor (TLR) responses in intestinal epithelial cells (IECs), which represent the front line of enteric defense, remains unclear. Therefore, I examined how ATP modulates TLR responses in IECs and found that it enhanced the response of IECs to a TLR1/2 ligand Pam₃CSK₄ primarily through the P2X7 purinergic receptor, leading to increased DC maturation and antigen-specific T cell proliferation. Furthermore, intra-rectal delivery of ATP lowered the activation threshold of epithelial cells to endogenous TLR ligands, making IECs more prone to immune activation. Since ATP is an important molecule that can potentiate inflammatory responses, the second aim of the study was to investigate if Tregs, including Foxp3+ Tregs and IL-10 producing Tr1 cells, can regulate ATP induced inflammasome activation and IL-1β production. I found that Tr1 cells inhibited the production of Il1b mRNA, inflammasome-mediated activation of caspase-1, and secretion of mature IL-1β, in an IL-10 dependent manner. Surprisingly, Foxp3+ Tregs, despite the production of IL-10, failed to inhibit IL-1β production. The important role of IL-10 in regulating inflammasome activation was further illustrated in the monosodium urate induced peritonitis model, where IL-10R-deficient mice had an increased influx of peritoneal neutrophils compared to wild type mice. Moreover, IL-1β production from macrophages derived from Nlrp3A350V knock-in mice, which carry a mutation found in cryopyrin associated periodic syndrome patients, was suppressed by Tr1 cells, but not Foxp3+ Tregs. Using an adoptive transfer model, I found Tr1 cells can protect against weight loss in mice expressing a gain-of-function mutation in NLRP3. Collectively, these data demonstrated the complex regulation of host response to cellular stress signal ATP, and that IL-10 producing Tr1 cells may have unique therapeutic effects in controlling ATP-mediated inflammasome activation via IL-10-mediated suppression.
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Master's Student Supervision
Theses completed in 2010 or later are listed below. Please note that there is a 6-12 month delay to add the latest theses.
Inflammatory bowel disease (IBD), including Crohn’s disease (CD) and ulcerative colitis (UC), is characterized by intestinal inflammation. Intestinal epithelial cells play a critical role in mucosal homeostasis and dysregulation of pro-inflammatory epithelial cell function could lead to the intestinal inflammation that characterizes IBD. However, we do not know the events that initiate inflammation or the cell types involved. One type of cell that may play a role is the tuft cell. Tuft cells are the only epithelial cells in the uninflamed intestine that express cyclooxygenase (COX)1 and COX2, the rate-limiting enzymes required for production of prostaglandins, like PGE2 and PGD2 which play important roles in immunity. In our research investigating the lipid phosphatase SHIP, it was discovered that tuft cells express SHIP. SHIP deficiency leads to increased PI3-kinase activity in cells resulting in increased cell proliferation, reduced apoptosis, and increased cell activation. SHIP expression is currently believed to be restricted to hematopoietic cells. However, using bone marrow transplantation, our laboratory found that tuft cells were not radiosensitive, suggesting that they are not bone-marrow derived and are not hematopoietic in origin.In addition, SHIP-deficient mice develop spontaneous Crohn’s disease-like intestinal inflammation. The onset of inflammation coincides with the developmental appearance of tuft cells. In wild type mice, tuft cells are found in the lung and ileum, both locations where SHIP-deficient mice develop spontaneous inflammation, and I found that tuft cell numbers were increased 6-fold in the inflamed ileum of SHIP-deficient mice. Based on this, I hypothesized that SHIP-deficient tuft cells may initiate or contribute to inflammation in the SHIP-deficient mouse. I found that SHIP-deficient mice had more COX1 positive cells in the ileum, more COX activity, and more PGD2 and PGE2 in full thickness ileal tissue homogenates, compared to their wild-type littermates. Finally, prophylactic inhibition of COX activity with piroxicam reduced the development of intestinal inflammation in SHIP-deficient mice whereas therapeutic treatment had little effect. This suggests that tuft cells may be critical in the initiation of spontaneous intestinal inflammation in SHIP-deficient mice and help elucidate some of the basic biology involved in the inflammation present in patients with CD.
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