Patrick Blanco

Dendritic cells and cytokines in human inflammatory and autoimmune diseases

Dendritic cells (DCs) produce cytokines and are susceptible to cytokine-mediated activation. Thus, interaction of resting immature DCs with TLR ligands, for example nucleic acids, or with microbes leads to a cascade of pro-inflammatory cytokines and skewing of T cell responses. Conversely, several cytokines are able to trigger DC activation (maturation) via autocrine, for example TNF and plasmacytoid DCs, and paracrine, for example type I IFN and myeloid DCs, pathways. By controlling DC activation, cytokines regulate immune homeostasis and the balance between tolerance and immunity. The increased production and/or bioavailability of cytokines and associated alterations in DC homeostasis have been implicated in various human inflammatory and autoimmune diseases. Targeting these cytokines with biological agents as already is the case with TNF and IL-1 represents a success of immunology and the coming years will expand the range of cytokines as therapeutic targets in autoinflammatory and autoimmune pathology.

Platelet CD154 Potentiates Interferon-α Secretion by Plasmacytoid Dendritic Cells in Systemic Lupus Erythematosus

In the autoimmune disease lupus, platelets activated by self-antigens contribute to pathology by triggering the secretion of interferon from immune cells. Taming the Big Bad Wolf Systemic lupus erythematosus (SLE)—a name some attribute to this disorder’s wolf-like ability to “devour” the affected organs—is an autoimmune inflammatory disease. It can affect virtually any part of the body, but often targets skin, kidney, and joints. A variety of immunological factors have been proposed to contribute to SLE, in particular the type I interferon (IFN) system, which is normally activated in response to viruses. Here, Duffau et al. point to platelets as the culprits in causing aberrant activation of IFN-α (a member of the type I IFN group) in lupus patients and suggest that a drug that blocks platelet activation could be a promising new treatment. A protein called CD154 (CD40 ligand) is found on T cells, where it helps to defend the body by activating cytotoxic immune cells during viral infections. It is also found on the surface of platelets that are activated for clotting and may contribute to the pathogenesis of inflammatory states such as atherosclerosis and autoimmune disorders, including SLE. Here, the authors collected platelets from patients experiencing SLE flare-ups of varying severity, as well as healthy controls, and demonstrated that CD154 abundance and shedding from platelets correlated with disease severity. Moreover, exposure of platelets from healthy donors to serum from patients with active SLE or to immune complexes similar to those in SLE patients triggered an increase in activation and CD154 production. These activated platelets, in turn, signaled to antigen-presenting cells to produce IFN-α, thus propagating an inflammatory cycle, both in vitro and in a murine model of lupus. To further test these ideas, Duffau et al. depleted the platelets in lupus-prone mice, which decreased inflammation in the animals’ kidneys, a commonly affected organ in lupus. They achieved a similar outcome by treating the mice with clopidogrel, an inhibitor of platelet activation already commonly used in patients with heart disease and stroke. In addition to experiencing less kidney damage, the clopidogrel-treated mice with lupus lived for an extra 3 months. The current mainstay of treatment for SLE is immunosuppressive therapy, achieved with steroids and chemotherapy-like medications. These drugs have numerous toxic effects, not the least of which is the immunosuppression itself, which predisposes patients to infections. Being able to treat lupus with an antiplatelet medication such as clopidogrel, which has few side effects, would markedly improve these patients’ safety and quality of life. A similar approach may prove useful in other autoimmune diseases such as rheumatoid arthritis, where it would also provide a badly needed alternative to immunosuppression. Systemic lupus erythematosus (SLE) is a systemic inflammatory autoimmune disease characterized by the involvement of multiple organs and an immune response against nuclear components. Although its pathogenesis remains poorly understood, type I interferon (IFN) and CD40 ligand (CD154) are known to contribute. Because platelets are involved in inflammatory processes and represent a major reservoir of CD154, we hypothesized that they participate in SLE pathogenesis. Here, we have shown that in SLE patients, platelets were activated by circulating immune complexes composed of autoantibodies bound to self-antigens through an Fc-γ receptor IIa (CD32)–dependent mechanism. Further, platelet activation correlated with severity of the disease and activated platelets formed aggregates with antigen-presenting cells, including monocytes and plasmacytoid dendritic cells. In vitro, activated platelets enhanced IFN-α secretion by immune complex–stimulated plasmacytoid dendritic cells through a CD154-CD40 interaction. Finally, in lupus-prone mice, depletion of platelets or administration of the P2Y(12) receptor antagonist (clopidogrel) improved all measures of disease and overall survival; transfusion of activated platelets worsened the disease course. Together, these data identify platelet activation as an important contributor to SLE pathogenesis and suggest that this process and its sequelae may provide a new therapeutic target.

Blood dendritic cells and DC-poietins in systemic lupus erythematosus

Dendritic cells (DCs) control immunity and tolerance. Hence, we surmised that systemic lupus erythematosus (SLE), a systemic autoimmune disease with autoreactive T and B cells, might be due to alterations in DC homeostasis. Taken together, our results demonstrate profound alterations of DCs and DC-poietins homeostasis in SLE. Elevated levels of interferon-alpha (IFN) in serum of SLE patients coexist with decreased numbers of cells producing IFN-alpha, i.e., plasmacytoid dendritic cells (PDCs). Decreased numbers of circulating DCs correlate with increased levels of soluble tumor necrosis factor (TNF) receptors, thus suggesting the potential role of TNF pathway in the observed DC alterations. Finally, increased FMS-like tyrosine kinase 3-ligand (FLT3-L) and its correlation with soluble TNF receptors suggest a physiologic response to compensate low DC numbers. Although IFN-alpha remains at the center of immunologic aberrations in SLE, it remains to be determined whether increased shedding of soluble TNF receptors could also be ascribed to IFN-alpha.

Role for toll-like receptors in autoimmune disease: the example of systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a multisystem disease characterized by an autoimmune response to nuclear antigens. Although the pathophysiology of SLE remains incompletely understood, many recent studies indicate a major role for innate immunity. The toll-like receptors (TLRs), which play a key role in innate responses to infections, are also involved in acute and chronic inflammatory processes induced by endogenous ligands. Numerous in vitro studies have established that TLR7 and TLR9 are involved in immune complex recognition. Activation of these receptors leads to activation of immune cells, most notably B cells and dendritic cells, and to the inappropriate production of many cytokines known to be directly involved in the pathogenesis of SLE. These data prompted studies in several murine models of SLE to assess the impact of inactivation or overexpression of genes encoding TLRs or molecules involved in TLR signaling pathways. The results confirmed the major role for TLR7 and suggested involvement of TLR4 in the induction of an aggressive autoimmune response. However, in vivo data suggest a protective effect of TLR9, thus contradicting the in vitro results. In humans, genetic studies have identified polymorphisms associated with increased susceptibility to SLE.

The interplay of dendritic cell subsets in systemic lupus erythematosus.

Dendritic cells (DC) control immunity and tolerance. Hence, we surmised that systemic lupus erythematosus (SLE), a systemic autoimmune disease with autoreactive T and B cells, might be due to DC alterations. Based on our findings, we are proposing a model of SLE where autoimmune responses are driven by unabated activation of myeloid DC through IFN‐α produced by plasmacytoid DC. Thus, interplay between DC subsets might represent a key component of SLE pathogenesis.

Expression of specific inflammasome gene modules stratifies older individuals into two extreme clinical and immunological states

Low-grade, chronic inflammation has been associated with many diseases of aging, but the mechanisms responsible for producing this inflammation remain unclear. Inflammasomes can drive chronic inflammation in the context of an infectious disease or cellular stress, and they trigger the maturation of interleukin-1β (IL-1β). Here we find that the expression of specific inflammasome gene modules stratifies older individuals into two extremes: those with constitutive expression of IL-1β, nucleotide metabolism dysfunction, elevated oxidative stress, high rates of hypertension and arterial stiffness; and those without constitutive expression of IL-1β, who lack these characteristics. Adenine and N4-acetylcytidine, nucleotide-derived metabolites that are detectable in the blood of the former group, prime and activate the NLRC4 inflammasome, induce the production of IL-1β, activate platelets and neutrophils and elevate blood pressure in mice. In individuals over 85 years of age, the elevated expression of inflammasome gene modules was associated with all-cause mortality. Thus, targeting inflammasome components may ameliorate chronic inflammation and various other age-associated conditions.

Omalizumab-induced decrease of FcɛRI expression in patients with severe allergic asthma

BACKGROUND It is documented that omalizumab treatment reduces the cell surface expression of immunoglobulin E high-affinity receptor (FcɛRI) on several cell types. This has not been investigated in patients with uncontrolled severe persistent allergic asthma. METHODS In a double-blind, randomized, placebo-controlled study, patients with severe allergic asthma uncontrolled by high dose inhaled corticosteroids and long-acting β(2)-agonist received either omalizumab (n = 20) or placebo (n = 11) over 16 weeks at appropriate doses and frequencies. Baseline and end of study (week 16) FcɛRI expression on basophils and plasmacytoid dendritic cells was determined by flow cytometry for the primary endpoint. Secondary efficacy endpoints included asthma control and lung function as part of an initial investigation into the use of FcɛRI expression as a marker of response. RESULTS In the omalizumab group, and with respect to placebo, FcɛRI expression was significantly reduced at end of study on basophils (-82.6%, p < 0.01) and plasmacytoid dendritic cells (-44.2%, p = 0.029). FcɛRI expression reduction was not found to be correlated with clinical response. CONCLUSIONS Long-term omalizumab treatment induced reduction of FcɛRI expression on circulating basophils and plasmacytoid dendritic cells. These changes were not associated with those of clinical features related to severe asthma, which does not support further investigation into its use as a predictive marker of response. TRIAL REGISTRATION The study was registered with ClinicalTrials.gov (identifier: NCT00454051) and the European Clinical Trials Database, EudraCT (identifier: 2006-003591-35).

Comparative analysis of NK cell subset distribution in normal and lymphoproliferative disease of granular lymphocyte conditions

We have characterized the heterogeneity of human blood NK cell subsets defined by expression of KIR, lectin like receptors and NK cell differentiation markers within a cohort of 51 healthy Caucasian individuals. High inter‐individual variability in cell surface expression of most NK cell markers is observed. Range values defining NK cell subsets in healthy donors were further used as references to characterize 14 patients with NK‐type lymphoproliferative disease of granular lymphocytes (NK‐LDGL). Alterations of the KIR repertoire were noted in all NK‐LDGL patients. NK cell expansions were classified as oligoclonal KIR+ or as non‐detectable KIR (ndKIR) using anti‐KIR2DL1/2DS1, anti‐KIR2DL2/2DL3/2DS2, anti‐KIR3DL1 and anti‐KIR2DS4 monoclonal antibodies. A major reduction in the size of the CD56bright NK cell subset was a constant feature of NK‐LDGL. Altered distribution of CD94+, CD161+, and CD162R+ NK cell subsets was also observed in NK‐LDGL patients. Considering the potential role of NK cells in eliminating tumors or virus‐infected cells, the reference values defined in this study should be valuable to characterize both quantitative and qualitative alterations of the NK cell repertoire in pathological conditions and to monitor NK cell reconstitution following hematopoietic transplantation.

Altered dendritic cell distribution in patients with common variable immunodeficiency

Recent data suggest a critical role for dendritic cells (DCs) in the generation of immunoglobulin-secreting plasma cells. In the work reported herein, we analyzed the frequency of peripheral blood plasmacytoid DCs (pDCs) and myeloid DCs (mDCs) in a cohort of 44 adults with common variable immunodeficiency (CVID) classified according to their CD27 membrane expression status on B cells. A deep alteration in the distribution of DC subsets, especially of pDCs, in the peripheral blood of CVID patients was found. Patients with a reduced number of class-switched CD27+IgD-IgM- memory B cells and patients with granulomatous disease had a dramatic decrease in pDCs (P = 0.00005 and 0.0003 vs controls, respectively) and, to a lesser extent, of mDCs (P = 0.001 and 0.01 vs controls, respectively). In contrast, patients with normal numbers of switched memory B cells had a DC distribution pattern similar to that in controls. Taken together, our results raise the possibility that innate immunity contributes to pathogenesis in CVID.

Immunological markers after long-term treatment interruption in chronically HIV-1 infected patients with CD4 cell count above 400 x 10(6) cells/l.

Objective:To analyse immunological markers associated with CD4+ lymphocyte T-cell count (CD4+) evolution during 12-month follow-up after treatment discontinuation. Method:Prospective observational study of chronically HIV-1 infected patients with CD4+ above 400 × 106 cells/l. Results:CD4+ changes took place in two phases: an initial rapid decrease in the first month (−142 × 106 cells/l on average), followed by a slow decline (−17 × 106 cells/l on average) The second slope of CD4+ decline was not correlated with the first and only baseline plasma HIV RNA was associated with it. The decline in CD4+ during the first month was steeper in patients with higher CD4+ and weaker plasma HIV RNA baseline levels. Moreover, the decline was less pronounced (P < 10−4) in patients with CD4+ nadir above 350 × 106 cells/l (−65 × 106 cells/l per month) in comparison with those below 350 × 106 cells/l (−200 × 106 cells/l per month). A high number of dendritic cells (DCs) whatever the type was associated with high CD4+ at the time of treatment interruption and its steeper decline over the first month. Moreover, the myeloid DC level was stable whereas the lymphoid DC count, which tended to decrease in association with decrease in CD4+, was negatively correlated with the HIV RNA load slope. Conclusions:The results support the use of the CD4+ nadir to predict the CD4+ dynamic after treatment interruption and consideration of the CD4+ count after 1-month of interruption merely reflects the 12-month level of CD4+. Although DCs seem to be associated with the CD4+ dynamic, the benefit of monitoring them has still to be defined.