Systemic sclerosis (SSc), or scleroderma, is a complex, rare disease of unknown etiology. Macrophages constitute a large portion of the immune cell infiltrate in the skin of patients with SSc, and are an important target of study. Particularly, the M2 macrophage has been implicated scleroderma and other fibrotic diseases as a key contributor to fibrotic processes. However, the definition of an M2 macrophage appears to change with context, and is poorly elucidated in different species. With varying characterizations between species and disease models, there is a need to establish some consensus on how to identify this macrophage in an uniform manner across species. We used a bioinformatic approach to identify a unique gene signature for the M2 macrophage phenotype, which is shared between human and mouse systems. We were able to confirm a 7-gene subset of this theorized signature using human and mouse in vitro systems. In addition, we selected one of the identified genes, Clec7a, and characterized its expression at the protein level on different macrophage phenotypes, across several human and mouse models. Our data show that Clec7a is a more selective marker of murine M2 macrophages than current reference markers, and is useful in human models as well. Using our M2-specific gene signature, we also identified a potential inhibitor of the signature and showed its effects on M2 marker expression. Finally, we showed some preliminary work into Clec7a expression in skin tissue from patients with scleroderma. Overall, our data suggest that Clec7a may be a valuable addition to the panel of markers used to characterize M2 macrophages and distinguish between macrophage phenotypes, and perhaps provide clarity into the development and function of the M2 macrophage. Better understanding of the M2 macrophage would ultimately be useful to the study of fibrotic diseases such as scleroderma, wherein this macrophage phenotype may be a viable target for antifibrotic therapy. / Thesis / Master of Science (MSc)

An estimated 45% of all deaths can be attributed to various chronic fibroproliferative diseases. Idiopathic pulmonary fibrosis (IPF) is the most common form of interstitial lung disease which is characterized by progressive decline in lung function. While the pathogenesis of IPF is not fully understood, alternatively activated macrophages (M2) have been implicated as a key contributor to the fibrotic process. The plasticity of macrophages in vivo challenges the ability to specifically target the M2 macrophage phenotype across species. Previous bioinformatic analysis from our lab identified Dectin-1/Clec7a as a unique marker of M2 macrophages in both human and murine model systems. The expression of the transmembrane receptor Dectin-1 has not been elucidated in the context of pulmonary fibrosis. To prevent the progression of fibrosis by targeting alternatively activated macrophages, we investigated the expression of Dectin-1 in IPF and an experimental model of fibrotic lung disease. Our data demonstrated that while protein expression of Dectin-1 was increased in archived lung tissues of patients with IPF, mRNA expression of this receptor was downregulated in the tissues of these IPF patients. Gene expression of Dectin-1 was shown to be increased in monocyte-derived macrophages, further suggesting a circulatory component contributing to lung fibrosis. As expected, we confirmed that Dectin-1 was highly expressed past the injury phase of the bleomycin-model of induced pulmonary fibrosis which aligns with the increased immune infiltrates at this time point. Preliminary work into the time dependency of the resolution phase of the bleomycin-induced model of lung fibrosis was shown. All in all, our data suggests that Dectin-1 may be a useful marker in characterizing and differentiating phenotypes of macrophages implicated in the fibrotic process. Future efforts aim to gain insight into the functional requirement of Dectin-1 in the alternative activation of profibrotic macrophages to identify novel therapeutic targets for fibrotic lung disease. / Thesis / Master of Science (MSc)