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ABSTRACT: IL-10-producing B cells, also known as regulatory B cells (Bregs), play a key role in controlling autoimmunity. In this study, we report that chimeric mice specifically lacking IL-10-producing B cells (IL-10(-/-)B cell) developed an exacerbated arthritis compared with chimeric wild-type (WT) B cell mice. A significant decrease in the absolute numbers of Foxp3 regulatory T cells (Tregs), in their expression level of Foxp3, and a marked increase in inflammatory Th1 and Th17 cells were detected in IL-10(-/-) B cell mice compared with WT B cell mice. Reconstitution of arthritic B cell deficient (μMT) mice with different B cell subsets revealed that the ability to modulate Treg frequencies in vivo is exclusively restricted to transitional 2 marginal zone precursor Bregs. Moreover, transfer of WT transitional 2 marginal zone precursor Bregs to arthritic IL-10(-/-) mice increased Foxp3(+) Tregs and reduced Th1 and Th17 cell frequencies to levels measured in arthritic WT mice and inhibited inflammation. In vitro, IL-10(+/+) B cells established longer contact times with arthritogenic CD4(+)CD25(-) T cells compared with IL-10(-/-) B cells in response to Ag stimulation, and using the same culture conditions, we observed upregulation of Foxp3 on CD4(+) T cells. Thus, IL-10-producing B cells restrain inflammation by promoting differentiation of immunoregulatory over proinflammatory T cells.
The Journal of Immunology 04/2011; 186(10):5569-79. · 5.79 Impact Factor
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ABSTRACT: Merozoite Surface Protein 1 is expressed on the surface of malaria merozoites and is important for invasion of the malaria parasite into erythrocytes. MSP1-specific CD4 T cell responses and antibody can confer protective immunity in experimental models of malaria. In this study we explore the contributions of cathepsins D and E, two aspartic proteinases previously implicated in antigen processing, to generating MSP1 CD4 T-cell epitopes for presentation. The absence of cathepsin D, a late endosome/lysosomal enzyme, is associated with a reduced presentation of MSP1 both following in vitro processing of the epitope MSP1 from infected erythrocytes by bone marrow-derived dendritic cells, and following in vivo processing by splenic CD11c+ dendritic cells. By contrast, processing and presentation of the soluble recombinant protein fragment of MSP1 is unaffected by the absence of cathepsin D, but is inhibited when both cathepsin D and E are absent. The role of different proteinases in generating the CD4 T cell repertoire, therefore, depends on the context in which an antigen is introduced to the immune system.
PLoS ONE 01/2011; 6(10):e24886. · 4.09 Impact Factor
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ABSTRACT: The bactericidal function of macrophages against pneumococci is enhanced by their apoptotic demise, which is controlled by the anti-apoptotic protein Mcl-1. Here, we show that lysosomal membrane permeabilization (LMP) and cytosolic translocation of activated cathepsin D occur prior to activation of a mitochondrial pathway of macrophage apoptosis. Pharmacological inhibition or knockout of cathepsin D during pneumococcal infection blocked macrophage apoptosis. As a result of cathepsin D activation, Mcl-1 interacted with its ubiquitin ligase Mule and expression declined. Inhibition of cathepsin D had no effect on early bacterial killing but inhibited the late phase of apoptosis-associated killing of pneumococci in vitro. Mice bearing a cathepsin D(-/-) hematopoietic system demonstrated reduced macrophage apoptosis in vivo, with decreased clearance of pneumococci and enhanced recruitment of neutrophils to control pulmonary infection. These findings establish an unexpected role for a cathepsin D-mediated lysosomal pathway of apoptosis in pulmonary host defense and underscore the importance of apoptosis-associated microbial killing to macrophage function.
PLoS Pathogens 01/2011; 7(1):e1001262. · 9.13 Impact Factor
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ABSTRACT: Improved chemical inhibitors are required to dissect the role of specific antigen processing enzymes and to complement genetic models. In this study we explore the in vitro and in vivo properties of a novel class of targeted inhibitor of aspartic proteinases, in which pepstatin is coupled to mannosylated albumin (MPC6), creating an inhibitor with improved solubility and the potential for selective cell tropism. Using these compounds, we have demonstrated that MPC6 is taken up via mannose receptor facilitated endocytosis, leading to a slow but continuous accumulation of inhibitor within large endocytic vesicles within dendritic cells and a parallel inhibition of intracellular aspartic proteinase activity. Inhibition of intracellular proteinase activity is associated with reduction in antigen processing activity, but this is epitope-specific, preferentially inhibiting processing of T cell epitopes buried within compact proteinase-resistant protein domains. Unexpectedly, we have also demonstrated, using quenched fluorescent substrates, that little or no cleavage of the disulfide linker takes place within dendritic cells. This does not appear to affect the activity of MPC6 as an inhibitor of cathepsins D and E in vitro and in vivo. Finally, we have shown that MPC6 selectively targets dendritic cells and macrophages in spleen in vivo. Preliminary results suggest that access to nonlymphoid tissues is very limited in the steady state but is strongly enhanced at local sites of inflammation. The strategy adopted for MPC6 synthesis may therefore represent a more general way to deliver chemical inhibitors to cells of the innate immune system, especially at sites of inflammation.
ACS Chemical Biology 03/2010; 5(5):461-76. · 6.45 Impact Factor
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ABSTRACT: Cathepsin E is an aspartic endosomal proteinase, expressed at high levels in some epithelial and haemopoetic cells. The enzyme has been implicated in a variety of functions, including antigen processing. This study documents strain-specific variation in expression of cathepsin E in mice. The levels of cathepsin E protein and message are profoundly decreased in haemopoetic cells from C57BL/6J mice, compared to levels in 129S2/Sv or Balb/c. The deficiency is cell-type-specific, as protein levels in gut are not affected. Deficiency affects B cell, T cells, macrophages and dendritic cells. The low cathepsin E phenotype cosegregates with the C57BL/6J genotype in a panel of C57BL/6J x 129S2/Sv F2 mice. Analysis of the promoter region of cathepsin E reveals a polymorphism which destroys a previously described functional PU.1 transcription binding consensus sequence in the C57BL/6J genome. Antigen processing of ovalbumin by dendritic cells, which has previously been shown to require cathepsin E, is impaired in C57BL/6J-derived dendritic cells. C57BL/6J mice thus exhibit a profound tissue-specific deficiency in cathepsin E expression, which may have important implications for the immune phenotype of this mouse strain.
Immunogenetics 01/2008; 59(12):927-35. · 2.93 Impact Factor
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ABSTRACT: Cathepsin D is a well-characterized aspartic protease expressed ubiquitously in lysosomes. Cathepsin D deficiency is associated with a spectrum of pathologies leading ultimately to death. Cathepsin D is expressed at high levels in many cells of the immune system, but its role in immune function is not well understood. This study examines the reconstitution and function of the immune system in the absence of cathepsin D, using bone marrow radiation chimaeras in which all haematopoietic cells are derived from cathepsin D deficient mice.
Cathepsin D deficient bone marrow cells fully reconstitute the major cellular components of both the adaptive and innate immune systems. Spleen cells from cathepsin D deficient chimaeric mice contained an increased number of autofluorescent granules characteristic of lipofuscin positive lysosomal storage diseases. Biochemical and ultrastructural changes in cathepsin D deficient spleen are consistent with increased autolysosomal activity. Chimaeric mice were immunised with either soluble (dinitrophenylated bovine gamma globulin) or particulate (sheep red blood cells) antigens. Both antigens induced equivalent immune responses in wild type or cathepsin D deficient chimaeras.
All the parameters of haematopoietic reconstitution and adaptive immunity which were measured in this study were found to be normal in the absence of cathepsin D, even though cathepsin D deficiency leads to dysregulation of lysosomal function.
BMC Immunology 02/2007; 8:22. · 2.53 Impact Factor
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ABSTRACT: Abstract
Background
Cathepsin D is a well-characterized aspartic protease expressed ubiquitously in lysosomes. Cathepsin D deficiency is associated with a spectrum of pathologies leading ultimately to death. Cathepsin D is expressed at high levels in many cells of the immune system, but its role in immune function is not well understood. This study examines the reconstitution and function of the immune system in the absence of cathepsin D, using bone marrow radiation chimaeras in which all haematopoietic cells are derived from cathepsin D deficient mice.
Results
Cathepsin D deficient bone marrow cells fully reconstitute the major cellular components of both the adaptive and innate immune systems. Spleen cells from cathepsin D deficient chimaeric mice contained an increased number of autofluorescent granules characteristic of lipofuscin positive lysosomal storage diseases. Biochemical and ultrastructural changes in cathepsin D deficient spleen are consistent with increased autolysosomal activity. Chimaeric mice were immunised with either soluble (dinitrophenylated bovine gamma globulin) or particulate (sheep red blood cells) antigens. Both antigens induced equivalent immune responses in wild type or cathepsin D deficient chimaeras.
Conclusion
All the parameters of haematopoietic reconstitution and adaptive immunity which were measured in this study were found to be normal in the absence of cathepsin D, even though cathepsin D deficiency leads to dysregulation of lysosomal function.
BMC Immunology. 01/2007;
