The respiratory DC/macrophage network at steady-state and upon influenza infection in the swine biomedical model.

Human and mouse respiratory tracts show anatomical and physiological differences, which will benefit from alternative experimental models for studying many respiratory diseases. Pig has been recognized as a valuable biomedical model, in particular for lung transplantation or pathologies such as cystic fibrosis and influenza infection. However, there is a lack of knowledge about the porcine respiratory immune system. Here we segregated and studied six populations of pig lung dendritic cells (DCs)/macrophages (Mθs) as follows: conventional DCs (cDC) 1 and cDC2, inflammatory monocyte-derived DCs (moDCs), monocyte-derived Mθs, and interstitial and alveolar Mθs. The three DC subsets present migratory and naive T-cell stimulation capacities. As observed in human and mice, porcine cDC1 and cDC2 were able to induce T-helper (Th)1 and Th2 responses, respectively. Interestingly, porcine moDCs increased in the lung upon influenza infection, as observed in the mouse model. Pig cDC2 shared some characteristics observed in human but not in mice, such as the expression of FCɛRIα and Langerin, and an intra-epithelial localization. This work, by unraveling the extended similarities of the porcine and human lung DC/Mθ networks, highlights the relevance of pig, both as an exploratory model of DC/Mθ functions and as a model for human inflammatory lung pathologies.

Mycobacterium bovis BCG killed by extended freeze-drying induces an immunoregulatory profile and protects against atherosclerosis.

OBJECTIVES: Atherosclerosis is an inflammatory disease of the arterial wall that leads to myocardial infarction and stroke. Regulatory T cells (Tregs) and IL-10 exert significant anti-atherogenic effects in experimental models of atherosclerosis by modulating vascular inflammation. We have previously shown that Mycobacterium bovis BCG killed by extended freeze-drying (EFD BCG) decreases lung and colon inflammation by recruiting IL-10-producing Tregs. Therefore, the aim of this study was to investigate the effect of EFD BCG on the development of atherosclerosis. DESIGN: We used two strains of atherosclerosis-prone mice: Ldlr(-/-) (four or six EFD BCG injections) and Apoe(-/-) (six injections). RESULTS: In both models, EFD BCG significantly reduced the size of atherosclerotic lesions, increased IL-10 production and reduced the serum levels of pro-inflammatory cytokines (IL-6, IL-13, KC and tumour necrosis factor-α). Shortly after treatment with EFD BCG, the number of plasmacytoid dendritic cells (pDCs) and Foxp3(+) Tregs in the draining lymph nodes increased. EFD BCG also led to accumulation of Tregs, but not of pDCs in the spleen, and reduced activity of NF-κB and increased activity of PPAR-γ in both the spleen and vascular tissue of treated mice. CONCLUSION: EFD BCG has atheroprotective effects through IL-10 production and Treg expansion. These findings support a novel approach to the prevention and treatment of atherosclerosis.

Efficacy of particle-based DNA delivery for vaccination of sheep against FMDV.

As an alternative strategy to classical inactivated viral vaccine against FMDV, naked DNA vaccine is attractive because of safety, flexibility and low cost. However DNA vaccination is usually poorly efficient in target species. Indeed we found that naked DNA plasmids encoding for P1-2A3C3D and GM-CSF proteins did not induce any detectable immunity against FMDV in sheep. Interestingly, we demonstrate herein that formulations of DNA on poly(D,L-lactide-co-glycolide) (PLG) or in lipofectin triggered divergent types of immune responses: PLG stimulated a T cell response and could elicit significant neutralising antibody titers, whereas lipofectin generated even higher antibody titers but no significant T cell response. The DNA/PLG regimen used in five sheep protected against clinical symptoms and viraemia and prevented the carrier state in four of them. Thus formulated DNA can be remarkably efficient against FMDV in a ruminant species that is usually refractory to DNA vaccination.

B-1-like cells exist in sheep. Characterization of their phenotype and behaviour.

Two populations of B lymphocytes, B-1 (CD5+ and/or CD11b+) and B-2 (CD5- and CD11b-) cells have been described. In mice, which is the species of reference for B-1 and B-2 cell studies, these two subsets present different developmental schemes, phenotypes, antibody repertoires, localization and behaviours. Interestingly, in sheep, B cells rearrange their immunoglobulin (Ig) loci around the neonatal period, similarly to murine B-1 cells. However, the phenotype of the sheep B cells has not been characterized with regards to their developmental pathway. In this report, we show that two sheep B-cell subsets can be distinguished on the basis of CD11b expression. Relative to CD11b- B cells, the CD11b+ B cells frequently co-express CD5, CD11c, higher levels of surface IgM (sIgM), show larger cell size and higher cell-cycling activity, and thus present a B-1-like phenotype. However, unlike murine B-1 cells, sheep B-1 like cells mainly localize in blood, display a higher propensity to spontaneous apoptosis relative to B-2-like cells, and proliferate after sIgM stimulation. Our data show that despite neonatal immunoglobulin loci rearrangements, sheep B cells do not all express a B-1-like phenotype. However, B-1-and B-2-like cells co-exist and present phenotypic and behavioural specificities. Nevertheless, sheep B-1-and B-2-like cells differ from the murine B-1 and B-2 cells in their cell behaviour. These subsets can thus not be considered as true homologues among species.

Bovine leukemia virus-induced lymphocytosis and increased cell survival mainly involve the CD11b+ B-lymphocyte subset in sheep.

In this study, we show that bovine leukemia virus (BLV)-induced persistent lymphocytosis (PL) results from the in vivo expansion of the CD11b+ B-lymphocyte population. This subset shares phenotypic characteristics with murine and human B-1 cells. BLV interactions with the sheep B-1-like subset were explored. We found that B-1- and B-2-like cells are initially infected to similar extents. However, in long-term-infected sheep, the viral load is higher in B-1-like cells and only B-1- and not B-2-like cells show increased ex vivo survival compared to that in uninfected sheep. Ex vivo viral expression was found in both B-1- and B-2-like cells, indicating that both cell types support viral replication. Finally, cycloheximide and a protein kinase C inhibitor (H7) that blocks the ex vivo activation of viral expression did not affect the increased survival in B-1-like cells, suggesting that resistance to apoptosis is acquired in vivo. Collectively, these results indicate a peculiar susceptibility of sheep B-1-like cells to BLV transforming effects and further support the involvement of increased survival in BLV pathogenesis.

Two- and three-dimensional cell structures govern epidermal growth factor survival function in human bladder carcinoma cell lines.

Human bladder carcinomas often express high levels of the epidermal growth factor (EGF) receptor. In three human bladder carcinoma cell lines (OBR, T24, and 647V), we show that two EGF receptor ligands, namely EGF and transforming growth factor alpha, enhanced the apoptosis due to serum starvation on cells cultured as monolayers. Conversely, EGF and transforming growth factor alpha prevented apoptosis when the same serum-starved cells were cultured as three-dimensional spheroids. Both stimulation and inhibition of apoptosis by EGF were associated with p21 WAF1/CIP1 overexpression. In 647V spheroids, EGF protection against radiation-induced apoptosis was negated by genistein and tyrphostin AG1478, suggesting that blockade of the EGF signal transduction in patients with bladder cancer may improve the radiotherapy efficacy.

Bovine leukaemia virus-induced lymphocytosis in sheep is associated with reduction of spontaneous B cell apoptosis.

Experimental inoculation of sheep with bovine leukaemia virus (BLV), a retrovirus homologous to the human T-lymphotropic virus type 1 (HTLV-1), induces a chronic expansion of the B lymphocyte population (persistent lymphocytosis) and the development of a B cell leukaemia/lymphosarcoma syndrome. To gain insight into the mechanisms of BLV-induced lymphocytosis, we tested B cell survival capacity and cycling activity in peripheral blood mononuclear cells (PBMCs) from lymphocytotic, asymptomatic and control sheep. Interestingly, B cells from lymphocytotic sheep presented a lower level of spontaneous apoptosis (29%) in ex vivo cultures compared to that obtained with infected asymptomatic (42%) and control (57%/o) sheep PBMCs. Virus capsid (CA) synthesis was mainly found among surviving B cells and the percentage of CA-producing B cells correlated with the extent of B cell survival, indicating that BLV replication in B lymphocytes may promote protection from cell death. B cell survival was not linked with increases in expression of Bcl-2 mRNA or membrane leukosialin (CD43), although both are documented to be involved in some aspects of the B cell life-span. Finally, cell cycle analyses in freshly isolated PBMCs from lymphocytotic sheep revealed a slightly increased proportion of B cells in S phase compared to controls. Altogether, these data suggest that both BLV-induced B cell proliferation and extended survival are involved in the lymphocytotic stage encountered in BLV infection in sheep.

Bovine immunodeficiency virus: facts and questions.

Bovine immunodeficiency virus (BIV) is a lentivirus whose serologic prevalence is worldwide. Little is known about its impact on animal health status, pathogenesis and mode of transmission. Understanding BIV biology implies isolation of new viral strains and long-term studies on experimentally-infected cows and surrogate hosts such as rabbits.