Correlation between profile of circulating mononuclear cells and clinical manifestations in patients with pemphigus vulgaris

Phenotypes of 38 samples of mononuclear (PBMC) cells from 11 different patients with pemphigus vulgaris (PV) at different stages of the disease were explored looking for a possible relationship between cell immunity, mucocutaneous or mucosal lesion intensity and capacity of serum autoantibodies to elicit the disease in mice. PBMC from 5 patients with mucocutaneous lesions and sera with IgG capable of inducing the disease in neonatal mice had a high proportion of mature monocytes with CD14low DRhigh, and co-expressing CD16 and CD11b. In addition, a high proportion of CD19+CD5+ activated B cells and a very low proportion of naive CD4+CD45RA+ and CD8+CD11b+ T lymphocytes was observed. Monocytes from these patients expressed inducible nitric oxide synthase (iNOS). In contrast, PBMC from 6 patients, with lesions restricted to mucosal membranes and IgG lacking the capacity to induce the disease in mice, contained a high proportion of CD14high DRlow co-expressing CD16 circulating macrophages, CD8+CD11b+ T cells, and a low proportion of activated B lymphocytes. The results suggest a possible association between proportion of different antigen presenting cells (monocytes with high HLA-DR and low CD14 expression and activated B lymphocytes, or differentiated monocytes/macrophages), type of PV and capacity of serum autoantibodies to elicit the disease in mice.     

Differential Adhesion Molecule Expression on Porcine Mononuclear Cell Populations

Adhesion molecule expression was analysed on porcine blood and lymphoid organ CD4⁺ CD8^? naïve T helper (Th) lymphocytes, CD4⁺ CD8⁺ memory Th lymphocytes (particular to the pig), CD4^? CD8^high cytotoxic T (Tc) lymphocytes, CD4^? CD8^low NK cells (CD3^? in the pig), CD4^? CD8^? T-cell receptor-γδ-positive (TCRγδ⁺) lymphocytes, B lymphocytes and monocytes. While CD44 expression was relatively homogeneous amongst mononuclear cells, differences were noted for the integrins. Blood naïve Th lymphocytes were CD49d^low CD11a^low, as were splenic naïve Th cells; blood memory Th lymphocytes were CD49d^high CD11a^low, splenic memory Th cells were CD49d^high CD11a^high with a CD49d^high CD11a^low subpopulation; blood Tc lymphocytes were mainly CD49d^low CD11a^low, and splenic cells were CD49d^high CD11a^high. Lymph node lymphocytes were more homogeneous in their integrin expression. These were relatively CD49d^low CD11a^low, except the memory Th lymphocytes which had higher integrin expression. B lymphocytes related to the majority of integrin^low T cells, while monocytes and NK cells were CD49d^high CD11a^high; γδ T lymphocytes showed variable CD49d expression but a CD11a^high phenotype. CD49d^high CD11a^high co-expression was found, and this phenotype was typical of, but not exclusive to, CD25⁺ (activated) lymphocytes. These results demonstrated that porcine memory Th lymphocytes and NK cells, as well as activated cells, would have increased integrin-dependent activities compared with naïve Th lymphocytes, and integrin-dependent reactions would probably vary between blood and lymphoid organ cells.     

NR4A1‐dependent Ly6Clow monocytes contribute to reducing joint inflammation in arthritic mice through Treg cells

Monocytes are central to the physiopathology of arthritis, but their roles in progression and resolution of the disease remain to be clarified. Using NR4A1^?/? mice, which lack patrolling lymphocyte antigen 6C (Ly6C^low) monocytes, we found that inflammatory Ly6C^high monocytes contribute to rapid development of arthritis in a serum transfer‐induced arthritis (STIA) model. Our experiments suggest that patrolling monocytes do not promote the initiation and progression of arthritis in mice, as severity of symptoms was amplified in NR4A1^?/? mice. Moreover, we show that treatment of arthritic wild type (WT) mice with cytosporone B (Csn‐B), a NR4A1‐specific agonist, significantly reduces severity of disease. Effects of Csn‐B were absent in monocyte‐depleted mice treated with clodronate until Ly6C^low monocytes were restored. Adoptive transfer of Ly6C^low monocytes in arthritic NR4A1^?/? mice treated with Csn‐B reduces joint inflammation, supporting the regulatory role of Ly6C^low subset on disease development. Our results also reveal that administration of Csn‐B to arthritic mice enhances levels of circulating CD4⁺CD25⁺FoxP3⁺ Treg cells, a process requiring the presence of Ly6C^low monocytes. Together, these data indicate that Ly6C^high monocytes are involved in the initiation and progression of arthritis and Ly6C^low monocytes contribute to reduce joint inflammation through the mobilization of Treg cells.     

Elevated circulating CD14<Superscript>low</Superscript>CD16<Superscript>+</Superscript> monocyte subset in primary biliary cirrhosis correlates with liver injury and promotes Th1 polarization

Primary biliary cirrhosis (PBC) is a progressive autoimmune liver disease in which monocytes/macrophages infiltration and skewed T helper type (Th) 1 and Th17 cell responses participate in the development of the disease. Human peripheral blood monocytes are heterogeneous and can be divided into classical CD14^highCD16^?, intermediate CD14^highCD16⁺, and nonclassical CD14^lowCD16⁺ monocyte subsets. Compared to classical monocytes, CD16⁺ monocytes are generally termed pro-inflammatory monocytes and play an important pathogenic role in autoimmune diseases. However, little is known about the immunophenotype and immunopathogenic role of peripheral blood CD16⁺ monocytes in PBC. Thus, we investigated the phenotype and function of these circulating monocyte subsets from PBC patients. The frequencies of circulating CD14^highCD16⁺ and CD14^lowCD16⁺ subpopulation were increased in disease compared with healthy controls. Among them, CD14^lowCD16⁺ monocyte subset positively correlated with disease progress, liver damage indicators and serum C-reactive protein, respectively. Furthermore, the frequencies of Th1 and Th17 cells were upregulated and CD14^lowCD16⁺ monocyte subset was also positively associated with Th1 cell frequency in PBC. Using a vitro coculture model, we further found that CD14^lowCD16⁺ monocytes promoted Th1 cell polarization compared to classical monocytes. Interleukin-12 (IL-12) and direct contact of patient CD4⁺T cell and CD14^lowCD16⁺ monocytes, were responsible for CD14^lowCD16⁺ monocytes promotion of Th1 cells polarization in PBC. Our study demonstrated that the enhanced CD14^lowCD16⁺ monocyte subset participated in fostering liver damage and inflammatory responses, and promoted Th1 cells skewing in PBC.     

Mouse and human CD8+ CD28low regulatory T lymphocytes differentiate in the thymus

Regulatory T (Treg) lymphocytes play a central role in the control of immune responses and so maintain immune tolerance and homeostasis. In mice, expression of the CD8 co‐receptor and low levels of the co‐stimulatory molecule CD28 characterizes a Treg cell population that exerts potent suppressive function in vitro and efficiently controls experimental immunopathology in vivo. It has remained unclear if CD8⁺ CD28^low Treg cells develop in the thymus or represent a population of chronically activated conventional T cells differentiating into Treg cells in the periphery, as suggested by their CD28^low phenotype. We demonstrate that functional CD8⁺ CD28^low Treg cells are present in the thymus and that these cells develop locally and are not recirculating from the periphery. Differentiation of CD8⁺ CD28^low Treg cells requires MHC class I expression on radioresistant but not on haematopoietic thymic stromal cells. In contrast to other Treg cells, CD8⁺ CD28^low Treg cells develop simultaneously with CD8⁺ CD28^high conventional T cells. We also identified a novel homologous naive CD8⁺ CD28^low T‐cell population with immunosuppressive properties in human blood and thymus. Combined, our data demonstrate that CD8⁺ CD28^low cells can develop in the thymus of mice and suggest that the same is true in humans.     

Syncytin-1/HERV-W envelope is an early activation marker of leukocytes and is upregulated in multiple sclerosis patients

Syncytin-1 is the envelope protein of the human endogenous retrovirus W (HERV-W). It has been related to multiple sclerosis (MS) but its role in cellular immunity and its pathogenic mechanism in the autoimmune context are not fully understood. We analyzed syncytin-1 levels in peripheral blood mononuclear cells (PBMC) subsets from healthy donors, MS patients in relapse or remission, and patients with acute infections by flow cytometry. PBMC cultures were also prepared to analyze protein expression kinetics. MS patients had higher levels of syncytin-1 levels than controls. We found that syncytin-1 is elevated in monocytes during MS relapses and infections. Cells expressing syncytin-1, including monocytes, T and B lymphocytes, and NKs presented mainly an activated phenotype and, upon stimulation with LPS, its levels increased rapidly on antigen-presenting cells. Syncytin-1 ligation promoted the activation of monocytes, as demonstrated by the upregulation of CD80 and the nonclassical subset CD14^low CD16⁺. Our results suggest an important role for syncytin-1 in the activation of leukocytes. Given that the expression of syncytin-1 is upregulated in MS patients, this protein might be contributing to the autoimmune cascade in the disease.     

Blood immune cell biomarkers in lung cancer

Characterization of host immune cell parameters prior to treatment is expected to identify biomarkers predictive of clinical outcome as well as to elucidate why some patients fail to respond to immunotherapy. We monitored blood immune cells from 58 patients with non-small- cell lung cancer (NSCLC) undergoing surgery of the primary tumor and from 50 age-matched healthy volunteers. Complete leukocyte blood count, the number of circulating dendritic cells (DC), HLA-DR^low monocytes and several lymphocytic subpopulations were determined by eight-color flow cytometry. Furthermore, the prognostic value of the immune cell parameters investigated was evaluated by patients’ survival analysis. Compared to the control group, blood of NSCLC patients contained more neutrophils resulting in a higher neutrophil-to-lymphocyte ratio (NLR), but a lower number of blood DC, in particular of plasmacytoid DC (pDC), natural killer (NK) cells and naive CD4⁺ and CD8⁺ T cells. Furthermore, a higher frequency of CD4⁺ regulatory T cells (Treg) and HLA-DR^low monocytes was detected, and smoking had a significant impact on these values. HLA-DR^low monocytes were positively correlated to the number of neutrophils, monocytes and NLR, but negatively associated with the number of pDC and naive CD4⁺ T cells. The frequency of Treg, HLA-DR^low monocytes and naive CD4⁺ and CD8⁺ T cells as well as the ratios of CD4/HLA-DR^low monocytes and HLA-DR^low monocytes/pDC correlated with patient’s overall survival. Next to Treg, HLA-DR^low monocytes and naive T cells represent prognostic markers for NSCLC patients and might be useful for monitoring of patients’ responses to immunotherapies in future studies.     

Surface antigen expression in spleen cells of C57Bl/6 mice during ageing: influence of sex and parity

So far all studies on the murine ageing process have been conducted on virgin mice. Immune ageing may be influenced by sex hormone differences related to sex or pregnancies. The aim of this study was to investigate whether pregnancies and gender influence the cell changes observed during ageing in a peripheral lymphoid compartment of C57Bl/6 mice. Using flow cytometry, changes in (Thy1.2⁺) T cell, (B220⁺) B cell and (CD11b/Mac-1) macrophage spleen populations were monitored in 2, 8 (3 months after last pregnancy) 15 and 23-month-old mice including males, virgin and multiparous females. The development of naive (CDCD44^low), memory (CD44^high), activated/memory (MEL-14, CD62L) cells were investigated in CD4⁺ and CE8⁺ T cell subsets. Both short term (at 8 months) and long term (at 15 and 23 months) effects of multiparity were obvious in the lymphocyte/macrophage population changes associated with the ageing process. Short-term effects included delayed appearance of CD4⁺CD44^high memory lymphocytes and increased numbers of both CD4⁺MEL-14^low activated/memory cells and Mac-1⁺ macrophages when compared with virgin control mice. Later effects of multiparity were increased CD8α^dull populations and increased T/B cell ratios and the ratio of memory to naive CD4⁺ cells (CD44^+high/CD44^+low). A sex effect was noticed: males exhibited lower Mac-1⁺ levels and memory/naive ratio in CD4⁺ subset than virgin females throughout life. These results suggest that gender and/or pregnancies affect the age-related distribution of lymphoid and macrophage cell populations in the spleen of C57Bl/6 mice.     

A population of CD62Llow NK1.1− CD4+ T cells that resembles NK1.1+ CD4+ T cells

In MHC class II−/− C57BL/6 (II−/−) mouse spleen, a small population of CD4⁺ T cells is present of which NK1.1⁺ CD4⁺ (NK) T cells comprise 40 to 45 %. We report here that many of the NK1.1⁻ CD4⁺ T cells derived from II−/− mice are also NK T cells. They produce large amounts of IL-4 in response to anti-CD3 ligation and do so without any requirement for the presence of IL-4 in the priming culture, a property characteristic of NK T cells. Their IFN-γ production is large and is enhanced by IL-12. In addition, II−/− NK1.1⁻ CD4⁺ T cells produce IL-4 as a result of culture with L cells expressing murine CD1 (L-CD1). We report that CD49b, a component of integrin VLA-2, is expressed on the majority of both NK1.1⁺ and NK1.1⁻ NK T cells. NK1.1⁻ NK T cells also exist in wild-type C57BL/6 mice. Evidence supporting this is that Vβ8 usage by CD62L^low NK1.1⁻ CD4⁺ T cells was ∼ 5 % higher than that by CD62L^high CD4⁺ T cells in wild-type mice in keeping with the estimated proportion of NK1.1⁻ NK T cells in the CD62L^low population. CD62L^low CD4⁺ T cells from β2-m−/− mice, which lack NK T cells, showed no increase in Vβ8 usage. When activated by anti-CD3 or L-CD1, CD62L^low NK1.1⁻ CD4⁺ T cells from conventional but not β2-m−/− and CD1−/− mice produce IL-4 in a manner indistinguishable from II−/− NK1.1⁻ CD4⁺ T cells. NK1.1⁻ NK T cells in normal mouse spleens are approximately as numerous as NK1.1⁺ NK T cells.     

Maturation of CD4+ Regulatory T Lymphocytes and of Cytokine Secretions in Infants Born Prematurely

Purpose

To characterize the maturation of CD4⁺ regulatory T lymphocytes (Treg) and of cytokine productions in preterm infants during their first 16 months of life.

Methods

The proportions of CD4⁺ Treg cells, their phenotypic characteristics, and the mitogen-induced cytokine productions by peripheral blood mononuclear cells (PBMC) were analysed in 26 very-preterm infants from 2 to 16 months of age, and compared to results obtained for 17 cord blood mononuclear cells (CBMC) from very-preterm infants, 12 from term infants and to blood samples from 40 adults.

Results

High proportion of CD25^+/highCD4⁺ Treg cells was found at birth in preterm CB with a gradual decreased afterwards. However, their percentage at 16 months of age was still higher than in term CB. In contrast to adults, preterm infants were characterized by excellent linear correlations between the proportions of CD25^+/highCD4⁺ and CD25^+/highFoxP3⁺ CD4⁺ or CD25^+/highCD127^low CD4⁺ or CD25^+/highFoxP3⁺CD127^low CD4⁺T lymphocytes. CD45RO⁺ and HLA-DR⁺ expressions were very low on preterm Treg and progressively increased with age. Functionally, preterm compared to term CBMC secreted in response to phytohaemagglutinin lower IFN-γ, higher IL-5 and similar IL-12p70, IL-10, IL-2 and IL-13 concentrations. IFN-γ, IL-12p70 and IL-10 productions were at 16 months still lower than those obtained for adults

Conclusion

Preterm differed from term CBMC both by their proportion and phenotype of CD4⁺ Treg lymphocytes and by their cytokine secretions. Maturation occurred during infancy with similar IFN-γ secretion but with persistently higher proportion of CD4⁺ Treg cells in 1 year preterm infants compared to term neonates.     

Interleukin‐4‐induced loss of CD8 expression and cytolytic function in effector CD8 T cells persists long term in vivo

Activation of naive CD8⁺ T cells in the presence of interleukin‐4 modulates their CD8 co‐receptor expression and functional differentiation, resulting in the generation of CD8^low cells that produce type 2 cytokines and display poor cytolytic and anti‐tumour activity. Although this CD8^low phenotype becomes stable after about a week and can persist with further stimulation in vitro, it is not known whether it can be maintained long term in vivo. Here we report that CD8^low cells derived from oval‐bumin_257–264‐specific T‐cell receptor‐transgenic CD8⁺ T cells activated in the presence of interleukin‐4 could be detected in the spleen for at least 4 months after adoptive transfer into normal mice. A significant proportion of the long‐term surviving cells retained their CD8^low phenotype in vivo and after clonal re‐activation in vitro. Although long‐term surviving CD8^low cells lacked detectable cytolytic activity or perforin expression, they showed some anti‐tumour function in vivo. The persistence of functional cells with a CD8^low phenotype in vivo raises the possibility that such cells can contribute to effector or regulatory responses to tumours or pathogens.     

C-C chemokines,but not the C-X-C chemokines interleukin-8 and interferon-γ inducible protein-10, stimulate transendothelial chemotaxis of T lymphocytes

Eight chemokines were tested for ability to elicit transendothelial chemotaxis of unstimulated peripheral blood T lymphocytes. The C-C chemokines monocyte chemotactic protein (MCP)-2, MCP-3, RANTES, macrophage inflammatory protein (MCP)-lα, MIP-1β, and, as previously described, MCP-1 induced significant, dose-dependent transendothelial chemotaxis of CD3⁺ T lymphocytes. In contrast, the C-X-C chemokines interleukin-8 (IL-8) and interferon-γ inducible protein-10 (IP-10) failed to induce transendothelial chemotaxis of CD3⁺ T lymphocytes or T lymphocyte subsets. RANTES, MIP-1α, and MIP-1β induced significant transendothelial chemotaxis of CD4⁺, CD8⁺, and CD45RO⁺ T lymphocyte subsets. Phenotyping of mononuclear cells that underwent transendothelial migration to MCP-2, MCP-3, RANTES, or MIP-1α showed both monocytes and activated (CD26 high), memory-type (CD45RO⁺) T cells. Both CD4⁺ and CD8⁺ T lymphocytes were recruited, but not natural killer cells or significant numbers of B cells. MCP-2 was the only C-C chemokine tested that attracted a significant number of naive-type (CD45RA⁺) T lymphocytes. In the absence of endothelium, IL-8 but not IP-10 promoted modest but significant chemotoxis of CD3⁺ T lymphocytes. Our data support the hypothesis that C-C, not the C-X-C chemokines IL-8 or IP-10, promote transendothelial chemotaxis of T lymphocytes.     

Staphylococcus aureus convert neonatal conventional CD4+ T cells into FOXP3+ CD25+ CD127low T cells via the PD‐1/PD‐L1 axis

The gut microbiota provides an important stimulus for the induction of regulatory T (Treg) cells in mice, whether this applies to newborn children is unknown. In Swedish children, Staphylococcus aureus has become a common early colonizer of the gut. Here, we sought to study the effects of bacterial stimulation on neonatal CD4⁺ T cells for the induction of CD25⁺ CD127^low Treg cells in vitro. The proportion of circulating CD25⁺ CD127^low Treg cells and their expression of FOXP3, Helios and CTLA‐4 was examined in newborns and adults. To evaluate if commensal gut bacteria could induce Treg cells, CellTrace violet‐stained non‐Treg cells from cord or peripheral blood from adults were co‐cultured with autologous CD25⁺ CD127^low Treg cells and remaining mononuclear cells and stimulated with S. aureus. Newborns had a significantly lower proportion of CD25⁺ CD127^low Treg cells than adults, but these cells were Helios⁺ and CTLA‐4⁺ to a higher extent than in adults. FOXP3⁺ CD25⁺ CD127^low T cells were induced mainly in neonatal CellTrace‐stained non‐Treg cells after stimulation with S. aureus. In cell cultures from adults, S. aureus induced CD25⁺ CD127^low T cells only if sorted naive CD45RA⁺ non‐Treg cells were used, but these cells expressed less FOXP3 than those induced from newborns. Sorted neonatal CD25⁺ CD127^low T cells from S. aureus‐stimulated cultures were still suppressive. Finally, blocking PD‐L1 during stimulation reduced the induction of FOXP3⁺ CD25⁺ CD127^low T cells. These results suggest that newborns have a higher proportion of circulating thymically derived Helios⁺ Treg cells than adults and that S. aureus possess an ability to convert neonatal conventional CD4⁺ T cells into FOXP3⁺ CD25⁺ CD127^low Treg cells via the PD‐1/PD‐L1 axis.     

Depletion of macrophages in mice results in higher dengue virus titers and highlights the role of macrophages for virus control

Monocytes and macrophages are target cells for dengue infection. Besides their potential role for virus replication, activated monocytes/macrophages produce cytokines that may be critical for dengue pathology. To study the in vivo role of monocytes and macrophages for virus replication, we depleted monocytes and macrophages in IFN‐αβγR knockout mice with clodronate liposomes before dengue infection. Although less virus was first recovered in the draining LN in the absence of macrophages, monocyte/macrophage depletion eventually resulted in a ten‐fold higher systemic viral titer. A massive infiltration of CD11b⁺CD11c^lowLy6C^low monocytes into infected organs was observed in parallel with increasing virus titers before viremia was controlled. Depletion of monocytes in the blood before or after local infection had no impact on virus titers, suggesting that monocytes are not required as “virus‐shuttles”. Our data provide evidence that systemic viremia is established independently of tissue macrophages present at the site of infection and blood monocytes. Instead, we demonstrate the importance of monocytes/macrophages for the control of dengue virus.     

Generalized autoimmune disease in interleukin-2-deficient mice is triggered by an uncontrolled activation and proliferation of CD4+ T cells

Interleukin-2-deficient mice (IL-2^?/?) crossed to a BALB/c genetic background develop a lymphoproliferative syndrome with severe hemolytic anemia and die within 5 weeks of age. The presence of autoantibodies of various specificities and inflammatory lesions in several organs are indicative of a generalized autoimmune disease. No alterations of the immune system were observed in 6-day-old animals, but 10-day-old mice already showed an increased proliferation and polyclonal activation of lymphocytes. The treatment of IL-2^?/? mice with anti-gp39(CD40L) antibody prevented the disease and indicated that the appearance of activated CD4⁺ T cells (CD44^high, CD69⁺) represents the first alteration of the immune system in IL-2^?/? mice. Collectively, our results suggest that an essential role of IL-2 in vivo, which is not compensated by other cytokines, is the maintenance of self tolerance.     

CD147 modulates the differentiation of T‐helper 17 cells in patients with rheumatoid arthritis

The role of CD147 in regulation of rheumatoid arthritis (RA) is not fully elucidated. The aim of this study was to investigate the effect of cell‐to‐cell contact of activated CD14⁺ monocytes with CD4⁺ T cells, and the modulatory role of CD147 on T‐helper 17 (Th17) cells differentiation in patients with RA. Twenty confirmed active RA patients and twenty normal controls were enrolled. CD4⁺ T cells and CD14⁺ monocytes were purified by magnetic beads cell sorting. Cells were cultured under different conditions in CD4⁺ T cells alone, direct cell‐to‐cell contact co‐culture of CD4⁺ and CD14⁺ cells, or indirect transwell co‐culture of CD4⁺/CD14⁺ cells in response to LPS and anti‐CD3 stimulation with or without anti‐CD147 antibody pretreatments. The proportion of IL‐17‐producing CD4⁺ T cells (defined as Th17 cells) was determined by flow cytometry. The levels of interleukin (IL)‐17, IL‐6, and IL‐1β in the supernatants of cultured cells were measured by ELISA. The optimal condition for in vitro induction of Th17 cells differentiation was co‐stimulation with 0.1 μg/mL of LPS and 100 ng/mL of anti‐CD3 for 3 days under direct cell‐to‐cell contact co‐culture of CD4⁺ and CD14⁺ cells. Anti‐CD147 antibody reduced the proportion of Th17 cells, and also inhibited the productions of IL‐17, IL‐6, and IL‐1β in PBMC culture from RA patients. The current results revealed that Th17 differentiation required cell‐to‐cell contact with activated monocytes. CD147 promoted the differentiation of Th17 cells by regulation of cytokine production, which provided the evidence for pathogenesis and potential therapeutic targets for RA.     

Proliferative response of human CD4+ T lymphocytes stimulated by the lectin jacalin

The Galβ(1–3)GalNAc-binding lectin jacalin is known to specifically induce the proliferation of human CD4⁺ T lymphocytes in the presence of autologous monocytes and to interact with the CD4 molecule and block HIV-1 infection of CD4⁺ cells. We further show that jacalin-induced proliferation is characterized by an unusual pattern of T cell activation and cytokine production by human peripheral blood mononuclear cells (PBMC). A cognate interaction between T cells and monocytes was critical for jacalin-induced proliferation, and human recombinant interleukin (IL)-1 and IL-6 did not replace the co-stimulatory activity of monocytes. Blocking studies using monoclonal antibodies (mAb) point out the possible importance of two molecular pathways of interaction, the CD2/LFA-3 and LFA-1/ICAM-1 pathways. One out of two anti-CD4 mAb abolished jacalin responsiveness. Jacalin induced interferon-γ and high IL-6 secretion, mostly by monocytes, and no detectable IL-2 synthesis or secretion by PBMC. In contrast, jacalin-stimulated Jurkat T cells secreted IL-2. CD3^? Jurkat cell variants failed to secrete IL-2, suggesting the involvement of the T cell receptor/CD3 complex pathway in jacalin signaling. IL-2 secretion by CD4^? Jurkat variant cells was delayed and lowered. In addition to CD4, jacalin interacts with the CD5 molecule. Jacalin-CD4 interaction and the proliferation of PBMC, as well as IL-2 secretion by Jurkat cells were inhibited by specific jacalin-competitive sugars.     

Ro52/TRIM21‐deficient expression and function in different subsets of peripheral blood mononuclear cells is associated with a proinflammatory cytokine response in patients with idiopathic inflammatory myopathies

The presence of anti‐Ro52/tripartite motif 21 (Trim21) autoantibodies has been associated with a distinctive clinical profile and has gained value as a prognostic marker in idiopathic inflammatory myopathies (IIM). The aim of the present work was to analyse Ro52/Trim21 expression in different subsets of peripheral blood mononuclear cells (PBMCs) of patients with IIM, as well as the ubiquitination profile and its association with proinflammatory cytokine production. We included 18 patients with recent‐onset IIM and 18 age‐ and gender‐matched healthy donors. PBMCs were isolated and different subsets (CD4⁺, CD8⁺, CD14⁺) were purified by magnetic selection. The expression of Ro52/Trim21 in different PBMC subsets of patients with IIM and healthy donors was analysed by Western blot. We assessed the presence of myositis‐specific and associated autoantibodies by enzyme‐linked immunosorbent assay (ELISA). Cytokine levels were measured by cytometric bead array. Patients with IIM showed decreased protein expression of Ro52/Trim21 in comparison to healthy controls in PBMC (0·97 ± 0·60 versus 1·84 ± 0·92, P = 0·016), CD4⁺ lymphocytes (0·79 ± 0·54 versus 2·41 ± 0·78, P = 0·017), and monocytes (0·87 ± 0·35 versus 1·89 ± 0·20, P < 0·001). There were no significant differences among IIM groups. Also, a lower K48‐mediated ubiquitination profile was found, predominantly in CD4⁺ lymphocytes. Furthermore, after mitogenic stimulation, there was a higher synthesis of proinflammatory cytokines by T cells [interleukin (IL)‐17A and tumour necrosis factor (TNF)‐α] and monocytes [IL‐6 and interferon (IFN)‐α] from IIM patients compared with healthy controls. Our data suggest that patients with IIM, mainly DM, are characterized by a deficient expression of Ro52/TRIM21 in different PBMC subsets (CD4⁺ lymphocytes and monocytes), along with lower K48‐mediated ubiquitination, which is associated with a proinflammatory cytokine response.     

Cognate Th2–B Cell Interaction is Essential for the Autoantibody Production in Pemphigus Vulgaris

Pemphigus vulgaris (PV) is a Th2-dominant autoimmune skin disease. We showed that indeed active PV patients had a biased Th2 response and specific IgG4 autoantibodies were dominant. To further investigate the role of antigen-specific Th2 cells in the regulation of pathogenic Dsg3-IgG antibody production, we used recombined Dsg3 protein to immunize wild-type C57BL/6 mice with aluminum hydroxide or complete Freund’s adjuvant as adjuvant. CD4⁺ T cells from Dsg3-immunized mice were adoptively transferred into TCR-β chain deficient mice. The transferred CD4⁺ T cells were readily seen in the peripheral blood and spleen, and interacted with B cells, resulting in B-cell activation. Furthermore, transferred CD4⁺ T cells from mice immunized with Dsg3 plus Alum with Th2 phenotype were able to render unprimed B cells to secrete Dsg3-specific IgG1 antibody in vivo. Taken together, these results provide the first demonstration of direct role of Dsg3-reactive CD4⁺ T (Th2) cells in the regulation of pathologic anti-Dsg3 antibody production.