Caroline Thielen

Lung interstitial macrophages alter dendritic cell functions to prevent airway allergy in mice

The respiratory tract is continuously exposed to both innocuous airborne antigens and immunostimulatory molecules of microbial origin, such as LPS. At low concentrations, airborne LPS can induce a lung DC-driven Th2 cell response to harmless inhaled antigens, thereby promoting allergic asthma. However, only a small fraction of people exposed to environmental LPS develop allergic asthma. What prevents most people from mounting a lung DC-driven Th2 response upon exposure to LPS is not understood. Here we have shown that lung interstitial macrophages (IMs), a cell population with no previously described in vivo function, prevent induction of a Th2 response in mice challenged with LPS and an experimental harmless airborne antigen. IMs, but not alveolar macrophages, were found to produce high levels of IL-10 and to inhibit LPS-induced maturation and migration of DCs loaded with the experimental harmless airborne antigen in an IL-10-dependent manner. We further demonstrated that specific in vivo elimination of IMs led to overt asthmatic reactions to innocuous airborne antigens inhaled with low doses of LPS. This study has revealed a crucial role for IMs in maintaining immune homeostasis in the respiratory tract and provides an explanation for the paradox that although airborne LPS has the ability to promote the induction of Th2 responses by lung DCs, it does not provoke airway allergy under normal conditions.

Levels and profiles of PCDDs, PCDFs and cPCBs in Belgian breast milk. Estimation of infant intake.

Congener-specific analyses of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs) and non-ortho (coplanar) polychlorinated biphenyls (cPCBs) were performed on 20 non-pooled breast milk samples collected in or close to an industrial area of Wallonia (Belgium). PCDD/F concentrations ranged between 16.0 and 52.1 pg TEQ/g fat, with a mean value of 29.4 pg TEQ/g fat. If coplanar PCBs (77, 126, 169) are included in TEQ calculations, levels ranged between 22.2 and 100.2 pg TEQ/g fat, with a mean value of 40.8 pg TEQ/g fat. It appears that 2,3,7,8-TCDD, 1,2,3,7,8-PeCDD, 2,3,4,7,8-PeCDF and PCB-126 account for more than 90% of the TEQ. Estimated PCDD/F dietary intake is 76 pg TEQ/kg body weight (bw)/day. This value is almost 20 times higher than the World Health Organization tolerable daily intake. A value of 103 pg TEQ/kg bw/day represents the intake of PCDDs, PCDFs and cPCBs (no mono-ortho PCBs included).

Lymphotoxin-beta receptor-dependent genes in lymph node and follicular dendritic cell transcriptomes.

Affinity maturation and Ab class switches occur in lymphoid germinal centers (GCs), in which differentiation and maintenance depend on lymphotoxin (LT) signaling and include differentiation of follicular dendritic cells (FDCs). The events leading to FDC and GC maturation are poorly defined. Using several approaches of functional genomics, we enumerated transcripts affected in mice by suppressing LT β receptor (LTβR) signaling and/or overrepresented in FDC-enriched GC isolates. Protein expression analysis of 3 of 12 genes both enriched in FDCs and down-regulated by LTβR signaling suppression validated them as FDC markers. Functional analysis of one of these three, clusterin, suggests a role as an FDC-derived trophic factor for GC B cells. Hence, the set of genes presented in this study includes markers emanating from LTβR signaling and transcripts relevant to GC and FDC function.

Molecular classification of T-cell lymphomas

T-cell neoplasms encompass a heterogeneous group of relatively rare disease entities. This review, focused on lymphoblastic tumors (T-ALL/LBL) and nodal-based peripheral T-cell lymphomas (PTCL), summarizes recent advances in the molecular characterization of these diseases. In T-ALL/LBL, molecular subgroups delineated by gene expression profiling correlate with leukemic arrest at specific stages of normal thymocyte development and different oncogenic pathways, and seem to be of interest for prognosis prediction. Angioimmunoblastic T-cell lymphoma (AITL), one of the most common PTCL entities, comprises neoplastic cells with a molecular signature similar to normal follicular helper T cells, and this cellular derivation might account for several of the peculiar aspects of this disease. Except in ALK-positive anaplastic large cell lymphoma, defined by ALK gene fusions, chromosomal translocations are otherwise rare in PTCLs, but some recurrent rearrangements might be associated with distinct lymphoma subtypes. In PTCL, not otherwise specified (PTCL, NOS), novel molecular biomarkers of potential therapeutic interest have been recently identified.

An Improved Protocol for Efficient Engraftment in NOD/LTSZ-SCIDIL-2RγNULL Mice Allows HIV Replication and Development of Anti-HIV Immune Responses

Cord blood hematopoietic progenitor cells (CB-HPCs) transplanted immunodeficient NOD/LtsZ-scidIL2Rγnull (NSG) and NOD/SCID/IL2Rγnull (NOG) mice need efficient human cell engraftment for long-term HIV-1 replication studies. Total body irradiation (TBI) is a classical myeloablation regimen used to improve engraftment levels of human cells in these humanized mice. Some recent reports suggest the use of busulfan as a myeloablation regimen to transplant HPCs in neonatal and adult NSG mice. In the present study, we further ameliorated the busulfan myeloablation regimen with fresh CB-CD34+cell transplantation in 3–4 week old NSG mice. In this CB-CD34+transplanted NSG mice engraftment efficiency of human CD45+cell is over 90% in peripheral blood. Optimal engraftment promoted early and increased CD3+T cell levels, with better lymphoid tissue development and prolonged human cell chimerism over 300 days. These humanized NSG mice have shown long-lasting viremia after HIV-1JRCSF and HIV-1Bal inoculation through intravenous and rectal routes. We also saw a gradual decline of the CD4+T cell count, widespread immune activation, up-regulation of inflammation marker and microbial translocation after HIV-1 infection. Humanized NSG mice reconstituted according to our new protocol produced, moderate cellular and humoral immune responses to HIV-1 postinfection. We believe that NSG mice reconstituted according to our easy to use protocol will provide a better in vivo model for HIV-1 replication and anti-HIV-1 therapy trials.

TARC and IL-5 expression correlates with tissue eosinophilia in peripheral T-cell lymphomas

The current study attempts to characterize the eosinophilia associated with T-cell lymphomas and to investigate its possible relationship with the secretion of eosinophil-stimulating factors by lymphoma cells and/or intra-tumoral surrounding cells. Paraffin-embedded specimens from 50 patients diagnosed with peripheral T-cell lymphomas, either unspecified (PTCL-U, n=30) or angioimmunoblastic (AITL, n=20) were morphologically assessed for intra-tumoral eosinophilia and analyzed by immunohistochemistry using specific antibodies directed against TARC, IL-5, RANTES, and eotaxin. The AITL and PTCL-U cases contained a mean of 147+/-41 and 102+/-37 eosinophils per 10 high power fields, respectively. Thirty-two of 47 cases (68%) showed IL-5-positive lymphoma cells while 15/50 (30%) tumors showed variable staining for TARC in scattered non-lymphoid cells with dendritic morphology. TARC and IL-5-positive cases possessed significantly more eosinophils. Our data indicate that IL-5 and TARC expression highly correlate with eosinophilia in T-cell lymphomas, suggesting that these chemokines are involved in the recruitment of eosinophils into the tumors.

Expression of SV2 isoforms during rodent brain development

BackgroundSV2A, SV2B and SV2C are synaptic vesicle proteins that are structurally related to members of the major facilitator superfamily (MFS). The function and transported substrate of the SV2 proteins is not clearly defined although they are linked to neurotransmitters release in a presynaptic calcium concentration-dependent manner. SV2A and SV2B exhibit broad expression in the central nervous system while SV2C appears to be more restricted in defined areas such as striatum. SV2A knockout mice start to display generalized seizures at a late developmental stage, around post-natal day 7 (P7), and die around P15. More recently, SV2A was demonstrated to be the molecular target of levetiracetam, an approved anti-epileptic drug (AED). The purpose of this work was to precisely analyze and quantify the SV2A, SV2B and SV2C expression during brain development to understand the contribution of these proteins in brain development and their impact on epileptic seizures.ResultsFirst, we systematically analyzed by immunohistofluorescence, the SV2A, SV2B and SV2C expression during mouse brain development, from embryonic day 12 (E12) to P30. This semi-quantitative approach suggests a modulation of SV2A and SV2B expression in hippocampus around P7. This is the reason why we used various quantitative approaches (laser microdissection of whole hippocampus followed by qRT-PCR and western blot analysis) indicating that SV2A and SV2B expression increased between P5 and P7 and remained stable between P7 and P10. Moreover, the increase of SV2A expression in the hippocampus at P7 was mainly observed in the CA1 region while SV2B expression in this region remains stable.ConclusionsThe observed alterations of SV2A expression in hippocampus are consistent with the appearance of seizures in SV2A−/− animals at early postnatal age and the hypothesis that SV2A absence favors epileptic seizures around P7.

Isolation of bovine follicular dendritic cells allows the demonstration of a particular cellular prion protein

Abstract. As interaction of cellular prion protein (PrPc) and the infectious agent (PrPres) appears to be a crucial pathogenic step promoted by homology, variation in PrPc isoforms on bovine immune cells may explain the absence of infectivity in most bovine lymph organs. In this study, we examined PrPc expression in bovine lymph organs (tonsils and lymph nodes) and on isolated follicular dendritic cells (FDCs). We used a panel of different monoclonal antibodies (MoAbs) raised against different epitopes of prion protein. Two MoAbs recognise amino acids 79–92 (SAF 34 and SAF 32 MoAbs); the 6H4 antibody reacts with a specific peptide comprising the 144–152 amino acids, and the 12F10 MoAb recognises the sequence 142–160. After immunolabelling of frozen sections of lymph organs with 6H4 or 12F10 MoAbs, we detected cellular prion protein in germinal centres. However, using the SAF 34 or SAF 32 antibodies, PrPc was revealed outside the lymphoid tissues. No PrPc was observed in the germinal centres. Therefore, we adapted the method of FDC isolation, making it suitable for the study of PrPc expression on their surface. Using electron microscopy, the presence of PrPc on the surface of FDCs was demonstrated only with 6H4 MoAb. These results suggest that bovine follicular dendritic cells express a particular form of prion protein. Either the N-terminal part of PrPc is cleaved or the accessibility of the specific epitope (79–92) of SAF 34 MoAb is abolished by interaction with other molecules. This particular isoform of PrPc on bovine FDCs might be related to the apparent absence of infectivity in lymph organs in cattle affected by bovine spongiform encephalopathy.

Human Fdc Express Prpc in Vivo and in Vitro

Prion diseases are fatal neurodegenerative disorders caused by accumulation of abnormal prion protein (protease-resistant prion, PrPres). PrPres accumulation is also detected in lymphoid organs after peripheral infection. Several studies suggest that follicular dendritic cells (FDC) could be the site of PrPres retention and amplification. Here we show that human follicular dendritic cells can express normal cellular prion protein (PrPc) both in situ and in vitro. When tonsillar cryosections were treated with anti-PrP antibody, the label was found on some very delicate cell extensions inside the lymphoid follicles, especially in the germinal centres. These extensions react with DRC1 antibody, used frequently to label FDC. Other structures labelled with anti-PrP antibody were the keratinocytes. To confirm the ability of FDC to synthesise PrPc, we isolated FDC by a non-enzymatic procedure and cultured them. By cytochemistry and flow cytometry it was clearly shown that FDC do produce PrPc.

c-Maf expression in angioimmunoblastic T-cell lymphoma reflects follicular helper T-cell derivation rather than oncogenesis

Vicky S Sabine Dana Faratian Tove Kirkegaard-Clausen John MS Bartlett Endocrine Cancer Group, Edinburgh Cancer Research Centre, Institute of Genetics and Molecular Medicine, University of Edinburgh, Western General Hospital, Edinburgh Breakthrough Research Unit, Institute of Genetics and Molecular Medicine, University of Edinburgh, Western General Hospital, Edinburgh, Scotland, UK, and Department of Breast Cancer Research, Institute of Cancer Biology, Danish Cancer Society, Copenhagen, Denmark 1. Kerr JF, Wyllie AH, Currie AR. Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br. J. Cancer 1972; 26; 239–257. 2. Ellis PA, Smith IE, Detre S et al. Reduced apoptosis and proliferation and increased Bcl-2 in residual breast cancer following preoperative chemotherapy. Breast Cancer Res. Treat. 1998; 48; 107–116. 3. Porter AG, Janicke RU. Emerging roles of caspase-3 in apoptosis. Cell Death Differ. 1999; 6; 99–104. 4. Kirkegaard T, Naresh A, Sabine VS et al. Expression of tumor necrosis factor alpha converting enzyme in endocrine cancers. Am. J. Clin. Pathol. 2008; 129; 735–743. 5. Faratian D, Kay C, Robson T et al. Automated image analysis for high-throughput quantitative detection of ER and PR expression levels in large-scale clinical studies: the TEAM trial experience. Histopathology 2009; 55; 587–593.