Maria Giovanna Quaranta

Leptin promotes differentiation and survival of human dendritic cells and licenses them for Th1 priming

Leptin is an adipocyte-derived hormone/cytokine that links nutrition, metabolism, and immune homeostasis. Leptin is capable of modulating several immune responses. However, the effect of leptin on dendritic cells (DCs) has not yet been recognized. Because DCs are instrumental in the development of immune responses, in this study, we evaluated the impact of leptin on DC activation. We demonstrated the presence of leptin receptor in human immature and mature DCs both at mRNA and protein level and its capacity to transduce leptin signaling leading to STAT-3 phosphorylation. We found no consistent modulation of DC surface molecules known to be critical for their APC function in response to leptin. In contrast, we found that leptin induces rearrangement of actin microfilaments, leading to uropod and ruffle formation. At a functional level, leptin up-regulates the IL-1β, IL-6, IL-12, TNF-α, and MIP-1α production. Coincident with this, leptin-treated DCs stimulate stronger heterologous T cell responses. Furthermore, we found that leptin down-regulates IL-10 production by DCs and drives naive T cell polarization toward Th1 phenotype. Finally, we found that leptin partly protects DCs from spontaneous and UVB-induced apoptosis. Consistent with the antiapoptotic effect of leptin, we observed the activation of NF-κB and a parallel up-regulation of bcl-2 and bcl-xL gene expression. These results provide new insights on the immunoregulatory function of leptin demonstrating its ability to improve DC functions and to promote DC survival. This is of relevance considering a potential application of leptin in immunotherapeutic approaches and its possible use as adjuvant in vaccination protocols.

Intraindividual genome expression analysis reveals a specific molecular signature of psoriasis and eczema

Signatures from patients with both psoriasis and eczema contribute to understanding disease pathogenesis and diagnosis. Discrimination That’s Skin Deep A molecular signature may be able to aid in the differential treatment of psoriasis from eczema. Quaranta et al. examined a group of individuals affected by both psoriasis and eczema and compared molecular signatures from psoriasis and eczema lesions on the same individual. This approach limited noise from interindividual variability, allowing the authors to focus on genes involved in disease pathogenesis. They found that psoriasis-specific genes involved not only immune mediators but also regulators of metabolism. In contrast, eczema-related genes included those related to the epidermal barrier and inflammasome activation. These insights provide not only new targets for disease-specific therapies but also an independently verified classifier that can be used in difficult to diagnose patients. Previous attempts to gain insight into the pathogenesis of psoriasis and eczema by comparing their molecular signatures were hampered by the high interindividual variability of those complex diseases. In patients affected by both psoriasis and nonatopic or atopic eczema simultaneously (n = 24), an intraindividual comparison of the molecular signatures of psoriasis and eczema identified genes and signaling pathways regulated in common and exclusive for each disease across all patients. Psoriasis-specific genes were important regulators of glucose and lipid metabolism, epidermal differentiation, as well as immune mediators of T helper 17 (TH17) responses, interleukin-10 (IL-10) family cytokines, and IL-36. Genes in eczema related to epidermal barrier, reduced innate immunity, increased IL-6, and a TH2 signature. Within eczema subtypes, a mutually exclusive regulation of epidermal differentiation genes was observed. Furthermore, only contact eczema was driven by inflammasome activation, apoptosis, and cellular adhesion. On the basis of this comprehensive picture of the pathogenesis of psoriasis and eczema, a disease classifier consisting of NOS2 and CCL27 was created. In an independent cohort of eczema (n = 28) and psoriasis patients (n = 25), respectively, this classifier diagnosed all patients correctly and also identified initially misdiagnosed or clinically undifferentiated patients.

HIV-1 Nef triggers Vav-mediated signaling pathway leading to functional and morphological differentiation of dendritic cells.

The accessory HIV‐1 Nef protein plays a key role in AIDS pathogenesis. We recently demonstrated that exogenous Nef triggers phenotypic and functional differentiation of immature dendritic cells (DCs). Here we investigated whether the Nef‐induced DC differentiation occurs with morphological remodeling and have focused on the interference of Nef in the signaling pathways that regulates DC maturation. We found that exogenous Nef enters immature DCs, promoting their functional and morphological differentiation. Specifically, Nef promotes interleukin (IL) ‐12 release, which closely fits with nuclear factor (NF) ‐κB activation. Nef induces rearrangement of actin microfilaments, leading to uropod and ruffle formation. Moreover, Nef increases the capacity of DCs to form clusters with allogeneic CD4+ T cells, improving immunological synapse formation. Searching for molecules involved in Nef‐triggered signaling pathways driving the DC maturation, we found that Nef targets Vav and promotes its tyrosine phosphorylation, associated with its nucleus‐to‐cytoplasm redistribution. This has a direct effect on Vav guanine nucleotide exchange factor activity for the small GTPase Rac1. We hypothesize that targeting Vav, Nef modulates both early signaling events (such as cytoskeletal rearrangement) and delayed responses (such as transcriptional regulation), promoting DC differentiation. Our results highlight how Nef may enhance T lymphocyte activation, thus fostering virus dissemination, manipulating the DC arm of the immune response.—Quaranta, M. G., Mattioli, B., Spadaro, F., Straface, E., Giordani, L., Ramoni, C., Malorni, W., Viora, M. HIV‐1 Nef triggers Vav‐mediated signaling pathway leading to functional and morphological differentiation of dendritic cells. FASEB J. 17, 2025–2036 (2003)

Leptin exerts an anti-apoptotic effect on human dendritic cells via the PI3K-Akt signaling pathway

Leptin is an adipocyte‐derived hormone/cytokine that modulates immune responses. It induces functional and morphological changes in human dendritic cells (DCs), licensing them towards Th1 priming and promoting DC survival. Here we found that leptin protects DCs from spontaneous, UVB and H2O2‐induced apoptosis, by triggering the activation of nuclear factor‐kappa B (NF‐kB) and a parallel up‐regulation of bcl‐2 and bcl‐XL gene expression and Akt activation. We found that leptin activates the PI3K‐Akt signaling pathway as demonstrated by the suppression of the effect of leptin on DC survival by wortmannin and API‐2, which suppress the leptin‐induced activation of Akt, NF‐kB, bcl‐2, bcl‐XL and protection from apoptosis. These results provide insights on the immunoregulatory function of leptin, supporting a potential application in immunotherapeutic approaches.

IL-22 suppresses IFN-γ–mediated lung inflammation in asthmatic patients

BACKGROUND IL-22 controls tissue homeostasis by both proinflammatory and anti-inflammatory effects. However, the anti-inflammatory mechanisms of IL-22 remain poorly investigated. OBJECTIVE We sought to investigate the anti-inflammatory role for IL-22 in human asthma. METHODS T-cell lines derived from lung biopsy specimens of asthmatic patients were characterized by means of flow cytometry. Human bronchial epithelial cells from healthy and asthmatic subjects were stimulated with IL-22, IFN-γ, or the combination of both cytokines. Effects of cytokine stimulation were investigated by using whole-genome analysis, ELISA, and flow cytometry. The functional consequence of cytokine stimulation was evaluated in an in vitro wound repair model and T cell-mediated cytotoxicity experiments. In vivo cytokine expression was measured by using immunohistochemistry and Luminex assays in bronchoalveolar lavage fluid of healthy and asthmatic patients. RESULTS The current study identifies a tissue-restricted antagonistic interplay of IL-22 and the proinflammatory cytokine IFN-γ. On the one hand, IFN-γ antagonized IL-22-mediated induction of the antimicrobial peptide S100A7 and epithelial cell migration in bronchial epithelial cells. On the other hand, IL-22 decreased epithelial susceptibility to T cell-mediated cytotoxicity by inhibiting the IFN-γ-induced expression of MHC-I, MHC-II, and CD54/intercellular adhesion molecule 1 molecules. Likewise, IL-22 inhibited IFN-γ-induced secretion of the proinflammatory chemokines CCL5/RANTES and CXCL10/interferon-inducible protein 10 in vitro. Consistently, the IL-22 expression in bronchoalveolar lavage fluid of asthmatic patients inversely correlated with the expression of CCL5/RANTES and CXCL10/interferon-inducible protein 10 in vivo. CONCLUSIONS IL-22 might control the extent of IFN-γ-mediated lung inflammation and therefore play a tissue-restricted regulatory role.

IFN-α amplifies human naïve B cell TLR-9-mediated activation and Ig production

TLRs are a family of molecules that function as sensors for the detection of pathogens. TLR‐9, expressed on B cells and pDCs, recognizes CpG motifs of unmethylated bacterial DNA and plays a role in the development of autoimmunity. The present study was designed to investigate the effects of IFN‐α in combination with CpG ODN on the activation of CD27– naïve B cells and on Ig production. We provide evidence that CpG ODN not only induces a total and T‐dependent, specific IgM response by naïve B cells but also their phenotypic differentiation in plasma cells, as demonstrated by the up‐regulation of CD38 expression. We found that TLR‐9 stimulation with CpG ODN induces IL‐1β, TNF‐α, IL‐10, and IL‐6 production. Interestingly, we also found that CpG ODN induces naïve B cell maturation into memory cells, as demonstrated by the induction of CD27, AID mRNA expression, and IgG production. More importantly, our results demonstrate that IFN‐α amplifies the inductive effect of CpG ODN on naïve B activation and on Ig production through a mechanism involving TLR‐9/MyD88‐dependent signaling. Moreover, we found that IFN‐α enhances the frequency of CpG ODN‐induced memory B cells. Our results may contribute to clarify the events promoting IFN‐α‐induced amplification of naïve B cell activation via TLR‐9 for a better understanding of the pathogenesis of autoimmune disorders and may guide treatments targeting this pathway within B cells.

HIV-1 Nef equips dendritic cells to reduce survival and function of CD8+ T cells: a mechanism of immune evasion

The accessory HIV‐1 Nef protein is a crucial determinant for viral replication and pathogenesis. During HIV infection, loss of immune control in the setting of a strong and broad HIV‐specific T‐lymphocyte response, leads to a lethal outcome through AIDS. Moreover, dysfunction of dendritic cells (DCs) may contribute to the immune suppression associated with AIDS progression. We recently demonstrated that exogenous Nef selectively activates immature DCs manipulating their phenotypical, morphological, and functional developmental program. Here, we tracked whether Nef, targeting DCs, could be involved in the dysregulation of CD8+ T cell responses. We found that Nef inhibits the capacity of DCs to prime alloreactive CD8+ T cell responses down‐regulating their proliferation and functional competence. This coincides with the induction of CD8+ T cell apoptosis. Nef oversees apoptotic killing of CD8+ T cells up‐regulating TNF‐α and FasL production by DCs and interfering with the death receptor pathway in CD8+ T cells and thus activating caspase 8. Our findings suggest that Nef may contribute to the immune evasion associated with HIV‐1 infection, subverting DC biology. This may help explain the pleiotropic function that Nef plays during infection and makes this protein an attractive target for preventive and therapeutic intervention.

Leptin as an immunological adjuvant: enhanced migratory and CD8+ T cell stimulatory capacity of human dendritic cells exposed to leptin

Leptin is an adipocyte‐derived hor‐ mone/cytokine that links nutrition, metabolism, and immune homeostasis and is endowed to modulate several immune responses. We previously demon strated that both immature and mature human den dritic cells (DCs) express a functional leptin receptor, and we found that leptin activates DCs, licenses them for Th1 priming, and promotes DC survival. Moreover, we found that leptin induces rearrangement of actin microfilaments, leading to uropod and ruffle formation. Here we monitor the effects of leptin on DC migratory capacities, focusing on the intracellular sig naling driving cytoskeleton rearrangement. We found that leptin increases immature DC migratory perfor mance both by favoring cytoskeleton dynamics and by up‐regulating CCR7 surface expression, thus favoring chemotactic responsiveness. We found that in immature DCs, leptin activates cofilin, favoring the turnover of actin microfilaments, and, by triggering Vav phos phorylation, promotes Rac1 activation. Finally, we found that in immature DCs, leptin up‐regulates inter‐ leukin‐12p70 production on CD40 stimulation and, more importantly, increases their capacity to stimulate activation of autologous CD8+ T cells. Taken alto gether, the findings herein highlight the potential use of leptin as an adjuvant tool in vaccination protocols employing ex vivo‐generated autologous DCs.— Mat‐ tioli, B., Straface, E., Matarrese, P., Quaranta, M. G., Giordani, L., Malorni, W., Viora, M. Leptin as an immunological adjuvant: enhanced migratory and CD8+T cell stimulatory capacity of human dendritic cells exposed to leptin. FASEB J. 22, 2012–2022 (2008)

Redox imbalance and immune functions: opposite effects of oxidized low‐density lipoproteins and N‐acetylcysteine

This study investigates the in vitro effects of oxidized low‐density lipoproteins (ox‐LDL), ‘physiological’ pro‐oxidants, N‐acetylcysteine (NAC), a free radical scavenger and glutathione precursor, and their combination on human peripheral blood mononuclear cell functions. We found that treatment with ox‐LDL induced a significant down‐regulation of proliferative response to mitogens, antigens and interleukin‐2. Lipid extracts from ox‐LDL were able to reproduce the same effect as the lipoprotein. On the other hand, NAC exposure induced a significant up‐regulation of proliferative responses to all the stimuli used. Moreover, we showed that natural killer (NK) cell‐mediated cytotoxic activity was significantly down‐regulated by ox‐LDL while treatment with NAC induced a significant up‐regulation of NK‐cell activity. Finally, we found that ox‐LDL and NAC exerted opposite effects on the cytokine network, interfering both at the protein secretion level and the messenger RNA synthesis level. More importantly, when NAC was used in combination with ox‐LDL the proliferative responses, NK‐cell‐mediated cytotoxic activity and cytokine production were restored to values comparable to controls. These data indicate that ox‐LDL and NAC modulate immune functions, exerting opposite effects reflecting their pro‐oxidant and antioxidant behaviours. Our results add new insights to the key role played by redox imbalance as a modulator of immune system homeostasis and suggest that an antioxidant drug such as NAC could be useful against pathologies associated with an increase in lipid peroxidation.

The immunoregulatory effects of HIV-1 Nef on dendritic cells and the pathogenesis of AIDS

Dendritic cells (DC) play a crucial role in the generation and regulation of immunity, and their interaction with HIV is relevant in the pathogenesis of AIDS favoring both the initial establishment and spread of the infection and the development of antiviral immunity. HIV‐1 Nef is an essential factor for efficient viral replication and pathogenesis, and several studies have been addressed to assess the possible influence of endogenous or exogenous Nef on DC biology. Our findings and other reported data described in this review demonstrate that Nef subverts DC biology interfering with phenotypical, morphological, and functional DC developmental programs, thus representing a viral tool underlying AIDS pathogenesis. This review provides an overview on the mechanism by which Nef, hijacking DC functional activity, may favor both the replication of HIV‐1 and the escape from immune surveillance. Overall, the findings described here may contribute to the understanding of Nef function, mechanism of action, and cellular partners. Further elucidation of genes induced through Nef signaling in DC could reveal pathways used by DC to drive HIV spread and will be critical to identify therapeutic strategies to bias the DC system toward activation of antiviral immunity instead of facilitating virus dissemination.—Quaranta, M. G., Mattioli, B., Giordani, L., Viora, M. The immunoregulatory effects of HIV‐1 Nef on dendritic cells and the pathogenesis of AIDS. FASEB J. 20, 2198–2208 (2006)