Maria Luisa Villa

Altered maturation of peripheral blood dendritic cells in patients with breast cancer.

Tumours have at least two mechanisms that can alter dendritic cell (DC) maturation and function. The first affects the ability of haematopoietic progenitors to differentiate into functional DCs; the second affects their differentiation from CD14+ monocytes, promoting an early but dysfunctional maturation. The aim of this study was to evaluate the in vivo relevance of these pathways in breast cancer patients. For this purpose, 53 patients with invasive breast cancer were compared to 68 healthy controls. To avoid isolation or culture procedures for enrichment of DCs, analyses were directly performed by flow cytometry on whole-blood samples. The expression of surface antigens and intracellular accumulation of regulatory cytokines upon LPS stimulation were evaluated. The number of DCs, and in particular of the myeloid subpopulation, was markedly reduced in cancer patients (P<0.001). Patient DCs were characterized by a more mature phenotype compared with controls (P=0.016), and had impaired production of IL-12 (P<0.001). These alterations were reverted by surgical resection of the tumour. To investigate the possible role of some tumour-related immunoactive soluble factors, we measured the plasmatic levels of vascular endothelial growth factor, IL-10 and spermine. A significant inverse correlation between spermine concentration and the percentage of DCs expressing IL-12 was found. Evidence was also obtained that in vitro exposure of monocyte-derived DCs to spermine promoted their activation and maturation, and impaired their function. Taken together, our results suggest that both the above-described mechanisms could concomitantly act in breast cancer to affect DC differentiation, and that spermine could be a mediator of dysfunctional maturation of DCs.

Human Immunodeficiency Virus (HIV)-Specific IgA and HIV Neutralizing Activity in the Serum of Exposed Seronegative Partners of HIV-Seropositive Persons

The presence and activity of human immunodeficiency virus (HIV)-specific antibodies were analyzed in the sera of 15 sexually exposed seronegative persons who had systemic HIV-specific cell-mediated immunity and IgA-mediated mucosal immunity and in their HIV-infected partners. The HIV-positive subjects had HIV-specific serum IgG and IgA; the seronegative persons had HIV-specific serum IgA in the absence of IgG. Testing of the seronegative persons 1 year after the interruption of at-risk sex showed that no IgG seroconversion had occurred and that HIV-specific IgA serum concentrations had declined. Serum from the HIV-exposed seronegative persons was analyzed for the ability to neutralize primary HIV-1 isolates. Neutralizing activity was detected in 5 of 15 sera and in 2 cases was retained by serum-purified IgA. Thus, the immunologic picture for resistance to HIV infection should include HIV-specific cell-mediated immunity as well as HIV-specific IgA-mediated mucosal and systemic immunity.

Mucosal and systemic HIV-1-specific immunity in HIV-1-exposed but uninfected heterosexual men.

Background: Despite multiple, repeated exposures to HIV-1, some individuals never seroconvert. Mucosal and systemic immune correlates of this condition have been analysed in HIV-1-exposed women but no data are available concerning mucosal immunity and HIV-1-specific immune responses in exposed but uninfected men. Design: We analysed cellular and humoral immune parameters in peripheral lymphocytes, seminal fluid and urethral swabs of 14 recently HIV-1-exposed seonegative (ESN) heterosexual men, seven HIV-seropositive patients and seven healthy controls. Results: HIV-1-specific IgA were detected in urethral swabs of 11 out of 14 ESN and of six out of seven HIV-seropositive patients; Env- and Gag-specific IFNγ-producing CD4 and CD8 peripheral lymphocytes were present in ESN and HIV-seropositive patients; seminal lymphocytes, but not peripheral blood lymphocytes, of ESN were enriched in activated populations (CD8CD38RO and CD4CD25). p24-specific cytotoxic T lymphocytes were correlated with the percentage of CD4 in the HIV-seropositive partners. High urethral concentrations of HIV-1-specific IgA were seen in those ESN with the most recent unprotected sexual episode. Conclusions: This is the first report of HIV-specific mucosal immunity in ESN men. These data add to the body of knowledge of the immune correlates present in exposed, uninfected individuals and might be important in vaccine design.

A possible role for the cortisol/anticortisols imbalance in the progression of human immunodeficiency virus

The progression of HIV infection is accompanied by complex alterations in the production of adrenal steroids. Cortisol levels are increased in HIV infection whereas those of dehydroepiandrosterone (DHEA), a physiologic antagonist of the immunoregulatory activities of cortisol, decrease. The progression of HIV infection to AIDS is also characterised by a shift from a type 1 to type 2 cytokine production. Thus, defective production of interferon gamma (IFN gamma), interleukin (IL)-2, and IL-12 as well as increased production of IL-4, IL-5, IL-6, and IL-10 are observed in HIV-seropositive individuals and are proposed to be in vitro immunologic marker of progression. Cortisol and pharmacological doses of glucocorticoids (GC) suppress IL-2 and IFN gamma production and favour the production of IL-4. Furthermore, GC and IL-4 stimulate the differentiation of B lymphocytes into IgE producing plasma cells, the concentration of which augments in HIV infection. Finally, GC induce programmed cell death (PCD) in a variety of different cells, including mature T lymphocytes, and type 2 cytokines were recently proposed to augment the susceptibility of T lymphocytes to PCD. It was suggested that the progressive shift from type 1 to type 2 cytokine production characteristic of HIV infection could be at least partially provoked by the increase in the production of cortisol and the reduction of DHEA. This hypothesis is discussed within the scenario of an endrocrinologic imbalance being responsible for HIV progression at least partially via increased susceptibility of HIV + CD4 lymphocyte to PCD.

Immune Profiles of Patients with Chronic Idiopathic Urticaria

Background: The immunologic characterization of chronic idiopathic urticaria (CIU) is still incomplete. In particular, it is not known if positivity to the intradermal autologous serum skin test (ASST) identifies an immunologic subset of CIU patients. Methods: Nineteen CIU patients and 15 healthy controls were enrolled in the study. A diagnostic flowchart was designed to select CIU patients, who were then analyzed by ASST. Cytokine and chemokine production and the expression of adhesion molecules was measured in patients and controls. Results: In CIU patients compared to controls, it was found that (1) TNF-α, IL-10, MIP-1α and RANTES production was augmented and IL-2 and INF-γ reduced, and (2) CD44, CD11a and CD62L expression on CD4 and CD8 cells was augmented. Additionally, TNF-α and chemokine production was significantly increased in CIU patients with a negative ASST (p–; n = 10) compared to patients with a positive response to the test. Conclusions: The presence of an inflammatory process in CIU patients is suggested by the findings that the production of both TNF-α and chemokines as well as the expression of adhesion molecules is increased in these patients. Similarly to what is seen in rheumatoid arthritis, augmented IL-10 production might be secondary to the attempt to hamper the inflammatory milieu. Immune profiles are particularly altered in CIU p– patients, in whom a more aggressive therapeutic strategy might be considered.

Mucosal and Systemic Immune Activation Is Present in Human Immunodeficiency Virus—Exposed Seronegative Women

Immune parameters were analyzed in peripheral blood mononuclear cells (PBMC) and cervical mucosa biopsy specimens of human immunodeficiency virus (HIV)-seronegative women sexually exposed to HIV (exposed seronegative [ESN]), HIV-infected women, and healthy women without HIV exposure. HIV was not detected in PBMC or cervical mucosa biopsy specimens of ESN women. However, interleukin (IL)-6, IL-10, IL-12, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha and -beta mRNA were elevated in PBMC and cervical mucosa biopsy specimens of ESN and HIV-infected women; CCR5 and CXCR4 mRNA were augmented in cervical mucosa biopsy specimens, but not in PBMC, of ESN and HIV-infected women; HIV-specific IFN-gamma-secreting cells were detected in vaginal washes of ESN and HIV-infected women; and phenotypic alterations were present in PBMC of ESN women. These results suggest that active HIV infection is not required for T cell activation; immune alterations occur in women in whom HIV infection cannot be detected virologically or clinically.

Human defensins activate monocyte-derived dendritic cells, promote the production of proinflammatory cytokines, and up-regulate the surface expression of CD91

Defensins are endogenous defense peptides with well‐defined antimicrobial activity against a broad spectrum of pathogens including bacteria, fungi, viruses, and parasites. Several lines of evidence suggest that defensins might also contribute to the regulation of host innate and adaptive immunity, but their immunomodulatory functions are still poorly understood. Herein, we studied the impact of human defensins on multiple functions of DCs, which are a central player in all immune responses, bridging innate and adaptive immunity. We challenged DCs differentiated in vitro from human moDCs with HNP‐1 α‐defensin or HBD‐1. HNP‐1 and HBD‐1 were chemotactic for moDCs. Both defensins promoted the activation and maturation of moDCs, as assessed by up‐regulation of surface expression of the costimulatory molecules CD80, CD86, and CD40, the maturation marker CD83, and HLA‐DR. HNP‐1 and HBD‐1 also enhanced the production of the proinflammatory cytokines TNF‐α, IL‐6, and IL‐12p70 but did not affect the production of the regulatory cytokine IL‐10. According to these stimulatory effects, HNP‐1 and HBD‐1 increased the allostimulatory activity of moDCs significantly. Finally, HNP‐1 and HBD‐1 promoted the up‐regulation of CD91 on the DC surface. CD91 is a scavenger receptor involved in the recognition of multiple ligands including defensins, thus suggesting that defensins may amplify their own effects through the activation of an autocrine loop. Taken together, our observations may provide new insight into the immunomodulatory properties of human defensins and may aid the exploration of new therapeutic strategies to potentiate antimicrobial and antitumor immunity.

Decrease and dysfunction of dendritic cells correlate with impaired hepatitis C virus-specific CD4+ T-cell proliferation in patients with hepatitis C virus infection

Through the production of stimulatory and suppressive cytokines, dendritic cells (DCs) regulate virus‐specific immune responses that are crucial to virus eradication. To explore a possible role of DCs in the persistence of hepatitis C virus (HCV) infection, in this study we analysed peripheral blood DCs (PBDCs) in patients with chronic hepatitis C (CHC) compared with those in both healthy seronegative (HSN) controls and a group of subjects who had spontaneously resolved infection, defined as healthy HCV‐seropositive (HSP), and we evaluated the relationships between PBDCs and HCV‐specific CD4+ T‐cell reactivity. The number of PBDCs, their immunophenotype and expression of regulatory cytokines were evaluated by flow cytometry on whole‐blood samples. HCV‐specific CD4+ T‐cell activation, proliferation and cytokine production were evaluated in cultures of peripheral blood mononuclear cells (PBMCs) stimulated in vitro with HCV peptides. We found that PBDCs from CHC subjects were numerically reduced and showed lower interleukin‐12 (IL‐12) and higher IL‐10 expression than those from HSN controls. PBDCs from HSP subjects were similar to those from HSN controls. HCV‐specific CD4+ T‐cell proliferation was less frequent and vigorous in CHC than in HSP patients and was directly related to the number of PBDCs and their IL‐12 production but inversely related to their IL‐10 production. Taken together, these results seem to suggest that cytokines of DC origin contribute to the regulation of HCV‐specific immunity in CHC patients and indicate that PBDCs may represent a novel non‐invasive tool for immune monitoring of these patients.

Functional repertoire of dendritic cells generated in granulocyte macrophage‐colony stimulating factor and interferon‐α

Monocyte‐derived dendritic cells (DCs) generated in granulocyte macrophage‐colony stimulating factor GM‐CSF) and interleukin‐4 (IL‐4–DCs) are used to enhance antitumor immunity in cancer patients, although recent evidence suggests that their functional repertoire may be incomplete; in particular, IL‐4–DCs appear unable to induce type 2 cytokine‐producing T helper (Th) cells. To assess whether type 1 interferon (IFN) could replace IL‐4 and generate DCs with a more complete repertoire, we characterized in detail DCs generated from human monocytes cultured with GM‐CSF and IFN‐α (IFN–DCs). We found that IFN‐α induces DC differentiation more efficiently than IL‐4, yielding similar numbers of DCs in a shorter time and that this differentiation persists upon removal of cytokines. Although IFN–DCs had a more mature immunophenotype than IL‐4–DCs, showing higher expression of CD80, CD86, and CD83, they still preserved comparable endocytic and phagocytic capacities and responsiveness to maturation stimuli. IFN–DCs had strong antigen‐presenting capacity, inducing intense proliferation of T cells to alloantigens or influenza virus. Moreover, IFN–DCs produced lower levels of IL‐12p70 and higher levels of IFN‐α, IL‐4, and IL‐10 than IL‐4–DCs. As a consequence of this different pattern of cytokine secretion, IFN–DCs induced T cells to produce type 1 (IFN‐γ) and type 2 (IL‐4 and IL‐10) cytokines, and as expected, IL‐4–DCs induced only Th1 differentiation. As immune responses with extreme Th1 bias are considered inadequate for the induction of optimal, systemic antitumor immunity, the ability of IFN–DCs to promote more balanced cytokine responses may suggest the advisability to consider these cells in the development of future, DC‐based immunotherapy trials.

Immunologic characterization of children vertically infected with human immunodeficiency virus, with slow or rapid disease progression

Cytokine production of unstimulated and mitogen-stimulated peripheral blood mononuclear cells of 31 children vertically infected with human immunodeficiency virus type 1 (HIV) and with different patterns of disease progression was evaluated to establish possible correlations between the immunologic and the clinical findings. Production of interferon gamma and interleukin-2 (type 1 cytokines), and of interleukin-4 and interleukin-10 (type 2 cytokines), was analyzed in seven symptom-free patients (Centers for Disease Control and Prevention class P-1B), 10 patients with mild symptoms (class P-2A), and 14 patients with severe symptoms (class P-2B-F). Cytokine production was compared with that of 10 age- and sex-matched control subjects who were seronegative for HIV. The HIV-infected patients produced significantly fewer type 1 cytokines and significantly more type 2 cytokines than the uninfected control subjects. No differences in the production of interferon gamma and interleukin-2 were detected among the different clinical categories of HIV-infected patients. In contrast, interleukin-4 production was augmented in the patients with class P-2A (p < 0.05) and class P-2B-F HIV infection (p < 0.03), in comparison with the children with class P-1B infection. The increase in interleukin-4 production was paralleled by an increase in the number of children with hyperimmunoglobulinemia E in each of the clinical groups (0% in class P-1B; 40% in class P-2A; and 71% in class P-2 B-F infection). Similarly, interleukin-10 production was increased both in patients with class P-2A and in those with class P-2B-F infection, in comparison with the children with class P-1B disease (p < 0.006 and < 0.04, respectively). These data indicate (1) that vertically acquired HIV infection results in decreased production of type 1 cytokines and in increased production of type 2 cytokines, and (2) that an increased production of type 2 cytokines correlates with hyperimmunoglobulinemia E and is present in, and may be characteristic of, the symptomatic phases of childhood HIV infection.