Effective treatment of the acute systemic inflammatory response associated with sepsis is lacking, but likely will require new ways to rebalance dysregulated immune responses. One challenge is that human sepsis often is diagnosed too late to reduce the hyperinflammation of early sepsis. Another is that the sequential response to sepsis inflammation rapidly generates an adaptive and immunosuppressive state, which by epigenetic imprint may last for months or years. Emerging data support that the immunosuppressive phase of sepsis can both directly reprogram gene expression of circulating and tissue cells, and disrupt development and differentiation of myeloid precursor cells into competent immunocytes. We recently reported that adoptive transfer of bone marrow CD34+ cells into mice after sepsis induction by cecal ligation and puncture significantly improves late-sepsis survival by enhancing bacterial clearance through improved neutrophil and macrophage phagocytosis. That study, however, did not examine whether CD34+ transfer can modify noninfectious acute systemic inflammatory responses. Here, we report that CD34+ cell transfer mice that have survived late sepsis also resist lethal lipopolysaccharide (LPS)-induced inflammatory shock (88% lived vs 0% of naive mice). The CD34+ cell-recipient survivor mice administered LPS had globally reduced levels of circulating inflammatory mediators compared with naive mice, but their peritoneal and bone marrow-derived macrophages (BMDMs), unlike those from naïve mice, remained LPS responsive ex vivo. We further found that CD34+ cell transfer into LPS-challenged naïve mice had diminished immunosuppression, as assessed by ex vivo responses of peritoneal and BMDMs to LPS challenge. We conclude that CD34+ cell adoptive transfer rebalances dysregulated immune responses associated with sepsis and endotoxin shock.

In the present study, we investigated the functional differences between cluster of differentiation (CD)14++CD16- and CD14+CD16+ monocytes during anti-Candida host defense. CD14++CD16- are the "classical" monocytes and represent the majority of circulating monocytes in humans, while CD14+CD16+ monocytes patrol the vasculature for maintenance of tissue integrity and repair. Both monocyte subsets inhibited the germination of live Candida albicans, and there was no difference in their capacity to phagocytose and kill Candida. Although production of IL-6 and IL-10 induced by C. albicans was found to be similar between monocyte subsets, IL-1β and prostaglandin E2 (PGE2) production was higher in CD14++CD16- compared with CD14+CD16+ monocytes. In line with the increased production of IL-1β and PGE2, central mediators for inducing Th17 responses, CD14++CD16- monocytes induced greater Th17 responses upon stimulation with heat-killed C. albicans yeast. The percentage of cells that expressed mannose receptor (MR) was higher in the CD14++CD16- monocyte subset, and MR-specific stimulation induced higher Th17 responses only in co-cultures of CD14++CD16- monocytes and CD4 lymphocytes. In conclusion, both monocyte subsets have potent innate antifungal properties, but only CD14++CD16- monocytes are capable of inducing a potent Th17 response to C. albicans, an important component of antifungal host defense.

We identified the C-type-lectin-like receptor, Dectin-1, as the major receptor for fungal β-glucans on murine macrophages and have demonstrated that it plays a significant role in the cellular response to these carbohydrates. Using two novel, isoform-specific mAb, we show here that human Dectin-1, the β-glucan receptor (βGR), is widely expressed and present on all monocyte populations as well as macrophages, DC, neutrophils and eosinophils. This receptor is also expressed on B cells and a subpopulation of T cells, demonstrating that human Dectin-1 is not myeloid restricted. Both major functional βGR isoforms - βGR-A and βGR-B - were expressed by these cell populations in peripheral blood; however, only βGR-B was significantly expressed on mature monocyte-derived macrophages and immature DC, suggesting cell-specific control of isoform expression. Inflammatory cells, recruited in vivo using a new skin-window technique, demonstrated that Dectin-1 expression was not significantly modulated on macrophages during inflammation, but is decreased on recruited granulocytes. Despite previous reports detailing the involvement of other β-glucan receptors on mature human macrophages, we have demonstrated that Dectin-1 acted as the major β-glucan receptor on these cells and contributed to the inflammatory response to these carbohydrates.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第20808号 / 医博第4308号 / 新制||医||1025(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 椛島 健治, 教授 玉木 敬二, 教授 生田 宏一 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Monocytes are professional antigen presenting cells (APC). They serve as precursors of macrophages and dendritic cells (DC). We have used cytopathic (cp) and non-cytopathic (ncp) Bovine Viral Diarrhea Viruses (BVDV) to determine the genes and proteins expression levels in bovine monocytes. Four specific aims were accomplished in this study. The first aim was to assess the baseline expression of the proteins involved in professional antigen presentation in bovine monocytes. The results showed that the differential detergent fractionation (DDF) approach can provide interpretable and meaningful functional information in bovine monocytes. The second aim was to evaluate the role of in vitro cp and ncp BVDV infection in the expression of the selected bovine genes involved in professional antigen presentation. The results showed that both BVDV could escape innate immune responses by modulating toll-like receptor (TLR) gene expression, followed by pro-inflammatory, type I interferon (IFN), Th1/Th2 type cytokine genes expression, and decreasing the expression levels of CD80/CD86 in professional APC. The third objective was to determine how the two biotypes affect selective antigen uptake, receptor-mediated endocytosis and non-selective uptake, macropinocytosis in bovine monocytes. The results indicated that bovine monocytes use macropinocytosis for a bulklow uptake of soluble antigens. The final aim was to characterize protein profiles in peripheral blood monocytes infected with cp BVDV isolate in vitro. Comparative profiling of the membrane and cytosolic proteins related to professional antigen presentation were assessed. The results showed that 47 bovine proteins, involved in immune function of professional APC have been significantly altered after cp BVDV infection. Overall, we hypothesize that by modulating expression levels of multiple proteins and genes related to immune responses BVDV could significantly compromise immune defense mechanisms resulting in uncontrolled immune activation or suppression.

Psoriasis is an immune mediated inflammatory disease which affects 2-3% of the world’s population. Over the last decade, psoriasis has been acknowledged as an independent risk factor for atherosclerosis. The precise mechanism or mechanisms of the heightened risk is widely speculated. Endothelial cells and macrophages are central players in the immunopathological development of both diseases. Interleukin-36 cytokines (IL-36) have been heavily implicated in psoriasis immunopathology. Significant upregulation of epidermal IL-36 is a recognised characteristic of psoriatic skin inflammation. IL-36 induces inflammatory responses in dendritic cells, fibroblasts and epithelial cells. While vascular alterations are a hallmark of psoriatic lesions and dermal endothelial cells are well known to play a critical role in dermal inflammation, the effects of IL-36 on endothelial cells have not been defined. We report that endothelial cells including dermal microvascular cells express a functionally active IL-36 receptor. Adhesion molecules VCAM-1 and ICAM-1 are upregulated following IL-36γ stimulation, and this is reversed in the presence of the endogenous IL-36 receptor antagonist. IL-36γ-stimulated endothelial cells secrete the proinflammatory chemokines IL-8, CCL2 and CCL20. Chemotaxis assays showed increased migration of T-cells following IL-36γ stimulation of endothelial cells. Both resident and infiltrating inflammatory myeloid cells contribute to the immunopathology of psoriasis by promoting the IL-23/IL-17 axis. We show that IL-36γ induces the production of psoriasis-associated cytokines from macrophages (IL-23, TNFα) and that this response is enhanced in macrophages from psoriasis patients. This effect is specific for IL-36γ and could not be mimicked by other IL-1 family cytokines such as IL-1α. Furthermore, IL-36γ stimulated macrophages potently activated endothelial cells as illustrated by ICAM-1(CD54) upregulation, and led to increased adherence of monocytes, effects that were markedly more pronounced for psoriatic macrophages. Interestingly, regardless of stimulus, monocytes isolated from psoriasis patients showed increased adherence to both the stimulated and unstimulated endothelium when compared to monocytes from healthy individuals. Collectively, these findings add to the growing evidence for IL-36γ having roles in psoriatic responses, by enhancing endothelium directed leukocyte infiltration into the skin and strengthening the IL-23/IL-17 pathway. Our findings also point to a cellular response which could potentially support cardiovascular comorbidities in psoriasis.

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