Lupus Research: TLR Signaling Pathways and Their Role in Disease Expression

New systemic lupus animal model research by Jeremy Tilstra, MD, PhD, in the laboratory of Mark Shlomchik, MD, PhD, UPMC endowed professor and chair of the Department of Immunology, is working to better understand the pathogenesis of lupus and lupus nephritis and the roles ofToll-like receptor (TLR) signaling, specifically the TLR9 and TLR7 pathways, on the disease.

Dr. Tilstra completed both his MD and PhD training at the University of Pittsburgh, followed by a clinical fellowship in Rheumatology that he completed in 2016 before subsequently joining the Division as assistant professor. "I've always had an interest in autoimmune and inflammatory diseases, and actually started out my research investigating inflammatory signaling pathways and macrophage signaling in inflammatory bowel disease in the lab of Scott Tabe. As my clinical training progressed in medical school and residency, I began to gravitate toward rheumatology, the pathogenesis questions with many of the rheumatic conditions, and specifically with lupus," says Dr. Tilstra.

As the timing would have it, Dr. Tilstra's interests and work on the rheumatology and lupus front, and his fellowship in rheumatology, coincided with the arrival of Dr. Shlomchik at the University of Pittsburgh, and he began to work in Dr. Shlomchick's lab. "Having a scientist of the caliber of Dr. Shlomchik become my mentor in studying a disease that he also had an interest in made it an easy choice to continue my research into lupus."

TLR Signaling in Lupus: Murine Models

One of the major features of lupus is the formation of autoantibodies that work against cell proteins. The vast majority of these antibodies can work against either DNA or RNA. "Work that had been done far prior to when I started in Dr. Shlomchik's lab showed that the antibodies were mediated by toll-like receptors. Interestingly, in our lupus murine model, TLR7 recognizes single-stranded RNA and is actually responsible for the anti-RNA antibodies. Conversely, TLR9 is responsible for double-stranded DNA anti-DNA antibodies."

TLRs are implicated as a central factor in the pathogenesis of lupus. In prior years and experiments in the field, it appeared that both TLR9 and TLR7 signaling pathways behaved in a similar manner when looked at through in vitro cell cultures. However, murine models showed completely dichotomous results. Murine lupus models deficient in TLR9 manifested a much worse disease indicating a mediating effect of TLR7. Conversely, "When TLR7 is knocked out in the model, we saw a slight improvement in disease. However, when both TLR9 and TLR7 are knocked out, the mouse models became much better in terms of their disease. The complex interaction between TLR7 and TLR9 was unexpected and not seen before in previous models of the disease," says Dr. Tilstra.

Dr. Tilstra explains that they have several hypotheses for why there is a beneficial effect when both TLR9 and TLR7 are knocked out in the animal model. One hypothesis is that the signaling pathways act in different cell types, or that there may be cell-specific effects. This led Dr. Tilstra to explore how TLR9 functions in different cell types. In order to test this approach, Dr. Tilstra and colleges analyzed cell specific knockouts of TLR9 in several cell types in the lupus murine model. This experiment showed that only in B cells was there a noticeable acceleration of disease.

The research team then conducted a unique experiment with a mouse model that over-expressed TLR9. These mice actually saw an improved disease state, suggesting that a response can be mediated in both ways with TLR9: with it knocked out you see worse disease, and with over-expression you see improved disease.

Using these different genetic models to tease apart how toll-like receptor signaling is implicated in disease, Dr. Tilstra and his colleagues hope to find to what degree cell specificity matters, and the different kinetics of that pathway. Ultimately, we want to know if these are something we can target through one method or another to suppress or reverse aspects of lupus development and progression," says Dr. Tilstra.

Research Into Tissue and Cellular Interactions in Lupus

Another research interest of Dr. Tilstra that is currently in progress seeks to better understand the interaction between the tissue parenchyma and infiltrating cells in lupus murine models. The science related to how cells act in the periphery is becoming better understood. However, there is currently much less knowledge about how cells behave when they enter into tissues. Recently completed research by Dr. Tilstra and his study collaborators has uncovered several novel, and unexpected, findings in this area of research. Their work is currently in submission for publication.

Lupus and the Microbiome

Dr. Tilstra is part of a University of Pittsburgh Department of Medicine-wide project that is collecting prospective tissue samples and data on patients with a variety of conditions and subpopulations of these patients. He currently heads up the lupus registries initiative that is part of this project which is collecting both genetic and microbiome data to be used in disease-state comparisons in an effort to find any associations between the microbiome and various conditions.

"This work is somewhat analogous to my research into how cells act in the tissues that lupus is causing damage and disease in. It's really trying to understand how the body acts in the natural environment, and what these upper level associations may be between the billions of bacterial cells in, on, and around us." Other members of the Division of Rheumatology and Clinical Immunology are contributing to the broader project with other diseases, including rheumatoid arthritis, myositis, and scleroderma.

References and Further Reading

1. Tilstra J, Gordon R, John S, Marburger B, Bastacky S, Nickerson K, Shlomchik M. B Cell Specific TLR9 Suppresses Disease in Murine Lupus [abstract]. Arthritis Rheumatol. 2017; 69 (suppl 10). http://acrabstracts.org/abstract/b-cell-specific-tlr9-suppresses-disease-in-murine-lupus/.