Research

Type 1 Diabetes:

It is now commonly understood that type 1 diabetes (T1D) is the result of a T cell-dependent autoimmune process that destroys the insulin-producing beta-cells in the pancreatic islets of Langerhans. The specificity of this destruction suggests that autoimmunity is directed at one or more beta-cell specific targets. In addition, several recent findings by members of our group support the contention that beta-cell destruction is the result of direct contact between the target beta-cell and specific effector cytotoxic T lymphocytes (CTL's). These findings include the following: (a) a large fraction of all islet associated CD8+ T-cells in non-obese diabetic (NOD) mice use highly homologous TCR-alpha chains on their T cell-receptors (TCR's); (b) these cells are a significant component of the earliest islet infiltrates (c) their levels in blood are detectable and predictive of subsequent diabetes development (d) they recognize a peptide from an islet specific protein, islet glucose-6-phosphatase catalytic subunit related protein (IGRP). Type 1 diabetes (T1D) results from complex interactions of many organs, cell types, and soluble mediators. To effectively prevent or interrupt the development of T1D we must understand how these systems function in a coordinated manner in vivo. Our team has been working for over two years, combining mathematical and experimental expertise to improve understanding of the dynamic, interacting complex systems involved in the initiation, progression, and cure of T1D.
At the core of T1D pathogenesis is the body's own immune system. The immune system is designed to detect and destroy foreign antigen, but in autoimmune (type 1) diabetes, this defense mechanism becomes destructive. By a complex set of interactions, a battalion of specialized immune system cells called (CD8+) effector T cells is created to target the insulin-producing pancreatic beta-cells. (These T cells recognize a specific feature of the beta-cells called an epitope.) Diabetes results when a large fraction of the beta-cells have been killed. Once this has progressed beyond some level, it is an irreversible condition. Fundamental questions in T1D include: (A) what mechanism leads to the priming of T cells? (If this mechanism could be blocked, it may be possible to envision a cure.) (B) What other effects (cytokines, macrophages) promote autoimmune conditions? (C)Why do some self-proteins become auto-antigens while others do not? (How does the type of protein, its location in the cell, its tendency to get misfolded implicate its role?)

(1) Peptide Therapy: (Maree, Santamaria, Keshet, others) The gradual amplification of increasingly effective killer T cells that bind more avidly to the beta-cell epitope is termed "avidity maturation". Our models have been aimed at understanding (1) what leads to this faulty immune response and avidity maturation, and (2) under what conditions peptide therapy could subdue the deadly T cells. A potential therapy involves injection of artificial peptides that mimic the beta-cell-specific epitope to modulate the observed avidity maturation. (Carried out by Pere Santamaria, U Calgary, using non-obese diabetic (NOD) mice, animals that spontaneously develop diabetes). A carefully selected dose and avidity for the injected peptide leads to magnification of harmless ("low avidity") T cells at the expense of the ("high avidity") killer T cells. Former MITACS PDF, Stan Maree, who continues to be an international team member, has modeled the process, using biologically relevant values of rates and parameters. His model reproduces the experimental data well, and discriminates between several underlying hypotheses, thus providing insight about what causes disease progression and what conflicting effects could result from peptide at the wrong dose or avidity. The model suggests under what conditions APL therapy is promising, and when it can actually be dangerous. The close relationship between experimental and theoretical scientists that characterizes this project would not have been possible without MITACS funding. (This work has been submitted for publication.)

(2) Macrophage clearance: (Maree, Komba, Keshet, Finegood, Dyck) A second model focuses on defective clearance of dead (apoptotic) pancreatic beta-cells as a possible cause of immune system triggering and inflammation leading to T1D. Clearance is usually handled by macrophages, cells that ingest and remove debris to avoid buildup of antigen stimulus. Finegood's group has observed that macrophages derived from diabetes-prone (NOD) mice are poor at clearance. She has also shown in previous work that in early life, there is a naturally occurring wave of programmed cell death of beta cells in experimental animals. The idea of a causative link between the death of beta cells and the buildup of inflammation and autoimmunity is being tested in our models.

By analyzing data for the numbers of macrophages that have ingested 1, 2, .. N apoptotic cells (in vitro), and using a simple mathematical model for this stage-structured population of macrophages, we were able to quantify the rates of engulfment and the rates of digestion of dead cells by the macrophages. We found that macrophages from diabetes resistant animals were able to engulf apoptotic cells significantly faster than those derived from diabetes-prone animals; digestion rates were relatively similar. We were able to quantify the differences that had been observed. Our model and analysis helped to improve the design of the experiment, so that a second round of data collected was much cleaner and more easily interpreted. New MITACS graduate student Mitsu Komba (started in Finegood's lab at SFU, Sept 2003) will be investigating the consequence of defective macrophage engulfment experimentally, while Stan Maree and Leah Keshet continue modeling work.

(3) Inflammed versus normal states: (Kublik, Komba, Maree, Keshet, Finegood, others) New MSc student Richard Kublik is studying the way that inflammation can be amplified by the wave of apoptosis of beta cells. Starting with the "Copenhagen model", he is determining under what conditions this (normally harmless) process can lead to runaway inflammation and overt disease conditions. His work is based on projects initiated by former PDF Labecki, and is being carried out in close consultation with Finegood's lab. Mitsu Komba's work on the role of defective macrophages described above in (2), and its implications on danger signals and dendritic cell response should provide insight the Copenhagen model extension.

(4) The epitope IGRP and T-Cell priming: (Pere Santamaria, Daniel Coombs, L Keshet) Santamaria's group has discovered an important protein found in beta cells which acts as autoantigen in T1D. Called IGRP, this transmembrane protein resides in a cell compartment in which antigen-MHC complexes are constructed (the endoplasmic reticulum). IGRP is highly hydrophobic, frequently misfolded, and thus often degraded into peptides by the proteosome. This could be one reason why it tends to be presented as a stimulus that primes T-cells. Santamaria, Coombs, and Keshet, plan to investigate the trafficking of IGRP from synthesis (by the ribosomes) through processing, and to rates of presentation. Experimental techniques of Pinciotta et al, Immunity 18:343-354, will be used in the Santamaria lab to provide data from which we will identify the key rate constants governing the major steps of antigen production and presentation, from ribosome to cell surface. Quantitative modeling at this level of detail has not previously been undertaken for a protein relevant to actual disease.

(5) Other potential future work: (Pere Santamaria, Maree, others?) Santamaria is interested in understanding how T-cells compete with one another in vivo. He can experiment with an in vivo reductionist 3-clonal population which he has developed, and we could base future detailed models on the observations. Pur previous models could be modified to include other effects, such as antigen sharing. For example, mature dendritic cells (DC's) are not able to make new peptide/MHC complexes, and cannot down-regulate pre-made complexes. They can also share exogenous antigens with other DC's by packaging them in exosomes. It is interesting to investigate how the small amounts of antigen captured by dendritic cells stimulate so many T-cells (in a TCR-transgenic mouse, for example).
Another possible project concerns cyclic wave of tetramer positive cells seen in peripheral blood in the pre-diabetic state (Trudeau et al, Clin Invest 111:217-23, 2003). Where do these cells come from? Do they represent cells that arise from the pancreatic lymph nodes? This is hard to believe given the few tetramer positive cells that are present there. Or do they represent memory T-cells that are hiding elsewhere and are then induced to migrate through the bloodstream in response to hypothetical stimuli? A mathematical analysis of the problem, together with carefully designed experiments might provide us with clues as to how this works.

Type 2 Diabetes:

Stem Cells:
Project C: Investigating Gene Expression of Stem Cells in Culture

Stem Cells:
Project D: Treatment Strategies for Acute Myelogenous Leukemia and Diseases of the Regulatory Systems for White Blood Cells and Platelets

Amplitude and period of the oscillations during periodic myelogenous leukaemia

Actin Dynamics and Cell Motility