The purpose of our research program is to elucidate the molecular and immunologic mechanisms regulating fibrosos in schistosomiasis and other chronic fibrotic diseases. Transgenic and knockout mice are employed in these experimental studies so that basic pathogenic processes can be investigated. Key findings from our murine studies are then extended to patients suffering from various debilitating forms of fibrosis, including schistosomiasis, a major cause of chronic liver fibrosis. The ultimate goal of our research is to understand how fibrosis is regulated by the immune response so that an immunologically based strategy might be developed to combat fibrosis. Tissue fibrosis or scarring is a leading cause of morbidity and mortality worldwide. Current statistics suggest that almost 45% of all deaths can be attributed to some type of chronic fibrotic disorder. Progress was achieved in the following areas during the year: ? ? 1) Studies on the role of IL-5 and eosinophils: In mice, S. mansoni worms induce an early Th1-type cytokine response. However, pathological fibrosis is due to the chronic production of Th2 cytokines in response to the parasite eggs that are produced in large numbers. Interleukin-4 (IL-4) is important for the initiation of Th2-type cytokine responses during schistosomiasis, while IL-13 is the key mediator of fibrosis. It is well established that IL-5 is an important mediator of eosinophil progenitor expansion in the bone marrow and survival in the periphery. They are also the most abundant cell type within S. mansoni-induced granulomas. Surprisingly, despite their abundance, the contribution of eosinophils and IL-5 to the progression of S. mansoni-induced liver fibrosis was unclear. We investigated whether IL-5 and/or eosinophils regulate the development of egg-induced pathology in the liver. To do this, we exposed IL-5-deficient mice to S. mansoni cercariae and examined their immune responses and pathologies at both acute and chronic time points postinfection. We also performed granuloma studies in the lung to determine whether there were any anatomical or model-specific differences in the activity of IL-5. We found that while infection intensities were similar in C57BL/6 and IL-5 KO mice, the average size of granulomas was significantly smaller in both acutely and chronically infected IL-5 KO mice. Their granulomas were also completely devoid of eosinophils. In addition, the knockout mice displayed over a 40% reduction in hepatic fibrosis by week 16 postinfection. The reduced fibrosis was associated with increased production of the antifibrotic cytokine gamma interferon. Although IL-13 production did not decrease consistently in the absence of IL-5, IL-13-triggered responses were substantially reduced in the granulomatous tissues. This was confirmed by analyzing the expression of several genes associated with alternative macrophage activation, including arginase 1, Fizz-1, and YM-1. Importantly, all of these IL-13-regulated genes have been linked with the mechanisms of wound healing and fibrosis. In addition to IL-5 polarizing the antigen-specific CD4+ Th2 cell response, we found that granuloma eosinophils were themselves a significant source of IL-13. Similar results were obtained with chronically infected eosinophil-lineage-ablated mice (DeltadblGATA and TgPHIL). Thus, by producing profibrotic mediators and polarizing the Th2 response, these findings illustrate both direct and indirect roles for eosinophils and IL-5 in the pathogenesis of schistosomiasis-induced liver fibrosis. Thus, inhibiting the activity of IL-5 or eosinophils may prove effective for a variety of chronic fibrotic diseases. ? ? 2) Studies on the role of IL-21/IL-21 receptor signaling: Th2 cytokines play an indispensable role in the pathogenesis of fibrosis. Indeed, IL-4/IL-13?, IL-4R?, and Stat6-deficient mice all show significantly impaired granuloma formation and liver fibrosis following infection with Schistosoma mansoni. Given the recent classification of IL-21 as a Th2 cytokine, the striking similarities between the IL-4 and IL-21 receptors, and the critical role of the related IL-4R/Stat6 signaling pathway in this disease as well as in other Th2 cytokine?driven inflammatory disorders, an important question evolving from these studies was whether IL-21R signaling was playing a significant role in the initiation and/or maintenance of Th2 immunity. To investigate the regulation and function of the IL-21R in vivo, several different experimental systems of Th2-dependent inflammation were examined, including models of pulmonary and hepatic inflammation as well as an experimental model of nematode infection. In each case, the immune responses of WT animals were compared with IL-21R?deficient mice. We showed that granulomatous inflammation and liver fibrosis are significantly reduced in S. mansoni?infected IL-21R?/? mice and in IL-21R+/+ mice treated with soluble IL-21R?Fc (sIL-21R?Fc). The impaired granulomatous response was also associated with a marked reduction in Th2 cytokine expression and function, as evidenced by the attenuated IL-4, IL-13, AMCase, Ym1, and FIZZ1 responses in the tissues. A similarly impaired Th2 response was observed following N. brasiliensis infection. In vitro, IL-21 significantly augmented IL-4R and IL-13R1 expression in macrophages, resulting in increased FIZZ1 mRNA and arginase-1 activity following stimulation with IL-4 and IL-13. As such, these data identify the IL-21R as an important amplifier of alternative macrophage activation. Collectively, these results illustrate an essential function for the IL-21R in the development of pathogen-induced Th2 responses, which may have relevance in therapies for both inflammatory and chronic fibrotic diseases.
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