Cristina Fain

TH1 and TH2 cytokine production by peripheral blood mononuclear cells from HIV-infected patients

ObjectiveTo study the TH1->TH2 cytokine switch, thought to occur during the progression of HIV infection. DesignWe investigated interleukin (IL)-2, interferon (IFN)-γ, IL-4, IL-6 and IL-10 production by phytohaemagglutinin (PHA)-stimulated and unstimulated peripheral blood mononuclear cell (PBMC) cultures from HIV-negative controls and HIV-positive subjects, stratified according to the Centers for Disease Control and Prevention (CDC) criteria. We correlated the above parameters with markers of disease progression. MethodsCytokine production was measured in supernatants using enzyme-linked immunosorbent assay (ELISA) in 40 patients and 17 controls. To evaluate disease progression, we also determined CD4+ cell counts, PHA-induced proliferative response, p24 release and spontaneous immunoglobulin (Ig) G and IgM production. ResultsIn agreement with the TH1->TH2 switch hypothesis, we found that in the course of HIV disease mitogen-stimulated IL-2 production decreased, spontaneous and stimulated IL-6 production and spontaneous IL-10 secretion increased; IL-4 showed an increasing trend, although it was reduced in HIV-positive subjects. Finally, immunoglobulin production increased over time. In contrast, mitogen-stimulated IFN-γ and IL-10 production did not change among the CDC categories, although the former decreased and the latter increased in comparison with HIV-negative controls. ConclusionsOur data partially agree with the TH1->TH2 switch hypothesis. Since IL-6 and IL-10 are produced by different cell types, whose proportions and functional features vary in the course of the disease, further experiments with purified lymphocyte subsets and monocytes are required. Nevertheless, as already suggested, we believe that a switch from a type 1 to a type 2 response occurs in HIV infection.

In vitro type-1 and type-2 cytokine production in systemic lupus erythematosus: lack of relationship with clinical disease activity:

Objective: to investigate the relationship between disease activity and in vitro cytokine, soluble(s)CD23 and polyclonal and anti-DNA antibody production by PBMC from patients with active systemic lupus erythematosus (SLE). Methods: cytokines, sCD23 and immunoglobulins were estimated by ELISA in unstimulated and polyclonal mitogen-stimulated culture supernatants. Results: PHA-induced IL-2 and IFN-y production were decreased, whereas spontaneous and PHA-induced IL-6 and IL-10 production were increased in cultures of SLE lympho cytes. Conversely, spontaneous and PHA-stimulated IL-4 and sCD23 production was com parable between patients and controls. Finally, we found an increase in in vitro spontaneous polyclonal and anti-DNA IgG secretion. Conclusions: We demonstrated an expanded type-2 cytokine profile with no correlation with parameters of disease activity.

In vitro production of type 1 and type 2 cytokines by peripheral blood mononuclear cells from high-risk HIV-negative intravenous drug users

ObjectiveTo study type 1 and type 2 cytokine patterns in HIV-negative high-risk intravenous drug users (IVDU). DesignWe investigated interleukin (IL)-2, interferon (IFN)-γ, IL-4, IL-6 and IL-10 production by phytohaemagglutinin (PHA)-stimulated and unstimulated peripheral blood mononuclear cell (PBMC) cultures from HIV-negative high-risk IVDU, HIV-negative controls and HIV-positive subjects. MethodsCytokine production was measured in supernatants using enzyme-linked immunosorbent assay (ELISA) in 10 HIV-negative high-risk IVDU, 25 HIV-negative controls, and 12 HIV-positive IVDU. We also determined spontaneous in vitro immunoglobulin (Ig) G and IgM production. ResultsHIV-negative high-risk IVDU showed increased IFN-γ and decreased IL-4, IL-10 and IL-2, although the latter was not significant compared with HIV-negative controls. Further, HIV-negative high-risk IVDU had reduced IgG production and impaired IgM-IgG switch. ConclusionsThe reduced IL-2 and IL-4 production suggest an impaired CD4+ T-cell function in HIV-negative high-risk IVDU. The increased IFN-γ production along with the decreased type 2 cytokine profile is consistent with the hypothesis that protective immunity against HIV may reside in type 1 responses and cell-mediated immunity.

Interleukin-10-induced HIV-1 expression is mediated by induction of both membrane-bound tumour necrosis factor (TNF)-alpha and TNF receptor type 1 in a promonocytic cell line

OBJECTIVE To investigate whether the upregulatory effect of interleukin (IL)-10 on HIV expression in a model of latent HIV infection is mediated by induction of endogenous tumour necrosis factor (TNF)-alpha and TNF receptors (TNFR). DESIGN The latently HIV-infected promonocytic cell line U1 was examined, because in this in vitro model IL-10 has been shown to synergize with multiple cytokines, including TNF-alpha, in enhancing HIV production. METHODS Membrane-bound TNF-alpha, TNFR-1 and TNFR-2 surface expression were determined by flow cytometry. TNF-alpha mRNA was estimated by competitive polymerase chain reaction (PCR), and TNF-alpha, soluble TNFR-1 and soluble TNFR-2 supernatant content by enzyme-linked immunosorbent assay. HIV-1 expression was quantitated by reverse transcriptase assay and p24 antigen release. RESULTS We demonstrated that IL-10 induces a time and cell-concentration dependent upregulation of HIV expression in U1 cells. This effect is mediated through the endogenous production of TNF-alpha as demonstrated by blocking experiments with anti-TNF-alpha antibodies and by detection of IL-10-induced increase of TNF-alpha mRNA by competitive PCR. More importantly, IL-10 is able to upregulate membrane-bound TNF-alpha and TNFR-1, along with a consistent increase in the shedding of soluble TNFR-1 without inducing detectable TNF-alpha secretion. CONCLUSIONS IL-10 activates HIV expression through the membrane-bound TNF-alpha/TNFR-1 pathway, suggesting an amplification mechanism of HIV expression that might occur during cell-to-cell interaction. This positive regulatory effect of IL-10 in an in vitro model of chronic HIV infection is consistent with the inexorable progression of disease seen in advanced patients when both IL-10 and TNF-alpha are elevated.

β-Endorphin content in HIV-infected HuT78 cell line and in peripheral lymphocytes from HIV-positive subjects

We investigated beta-endorphin (BE) content in an HIV-infected cell line and in peripheral blood mononuclear cells (PBM) from HIV-positive subjects. HIV infection increased BE content in HuT78 cell line compared to uninfected cells. Accordingly, BE content was greater in HIV-positive subjects than in healthy controls, both in fresh PBM and in mitogen-stimulated or unstimulated cultured cells. Further, in PHA-stimulated cultures, BE increase was correlated with disease progression. Opioids are known to decrease immune responsiveness in vivo, and it may be that the increased BE concentrations contribute to HIV-associated immune deficiency. In HIV-positive subjects, but not in healthy controls, intracellular BE concentration was positively correlated with PHA-induced PBM proliferation. The latter data suggest an alternative explanation: that the increased BE content represents a paradoxical response of the host in an attempt to balance virus-induced immunodepression. Thus, BE may be important in fine-tuning of the immune response with its up- and downregulation dependent upon differences in immune status.

Immunopharmacological Activity of Cefodizime in Young and Elderly Subjects: In Vitro and Ex Vivo Studies

SummaryThe biological response modifying activities of cefodizime (CDZ), a new third-generation cephalosporin, were investigatedin vitro andex vivo. In vitro investigations using cells isolated from the blood of young healthy donors showed no stimulating activity of CDZ on peripheral blood lymphocytes, natural killer cell activity, IL-1 production by adherent mononuclear cells, PMN chemiluminescence or PMN chemotaxis. A slight but statistically insignificant increase in PMN phagocytosis and phagocytic index was observed in the same population. IL-1 production was increased in three subjects with low resting state values. In a controlledex vivo study, 20 health elderly subjects selected on the basis of depressed phagocytic function were treated with CDZ 1 g i.m. b.i.d. or placebo for eight days. PMN function was determined at baseline and on the day after the last dose. In the CDZ group a significant increase in both phagocytosis and phagocytic index was found, while there were no changes in the placebo group. In conclusion, CDZ restored depressed PMN phagocytic function in a population of elderly subjects. Patients with impaired PMN function who require antibiotic treatment may benefit from this activity of CDZ.ZusammenfassungDie immunologischen Wirkungen von Cefodizim (CDZ), einem neuen Cephalosporin, wurden untersucht. In einerin vitro Studie mit Zellen, die aus dem Blut junger, gesunder Spender isoliert wurden, zeigte CDZ keine stimulierenden Wirkungen auf Lymphozyten, NK-Zell-Aktivität, IL-1 Produktion durch haftende mononukleäre Zellen, die Chemilumineszenz und Chemotaxis von polymorphkernigen neutrophilen Granulozyten (PMN). Ein geringer, statistisch nicht signifikanter Anstieg der PMN Phagozytose und des Phagozytose-Index wurde beobachtet. Bei drei Personen mit niedriger IL-1 Produktion in Ruhe konnte ein Anstieg derselben induziert werden. Im Rahmen einer kontrolliertenex vivo Studie erhielten 20 gesunde ältere Versuchspersonen, die aufgrund einer verminderten PMN Phagozytose ausgewählt worden waren, CDZ 2 × 1 g i.m. oder Plazebo über einen Zeitraum von acht Tagen. Die Funktion der PMN wurde vor Behandlungsbeginn und am Tag nach der letzten Dosis bestimmt. In der CDZ-Gruppe kam es zu einem signifikanten Anstieg der Phagozytose und des Phagozytose-Index, während in der Plazebo-Gruppe keine Veränderungen beobachtet wurden. Schlußfolgerung: CDZ konnte bei älteren gesunden Versuchspersonen eine verminderte Phagozytosetätigkeit der PMN wiederherstellen. Patienten mit gestörter PMN-Funktion, die einer antibiotischen Behandlung bedürfen, könnte diese Eigenschaft von CDZ zugute kommen.

Immunosuppressive activity of 15-deoxyspergualin on normal and autoimmune peripheral blood mononuclear cells

Several experimental conditions were used in this study to evaluate the in vitro effects of 15-deoxyspergualin on the function of T lymphocytes, B lymphocytes and monocytes from healthy subjects and patients suffering from systemic lupus erythematosus. Whilst the secretion of polyclonal immunoglobulin (Ig) M and IgG from the B lymphocytes of the healthy subjects was diminished by 15-deoxyspergualin, neither the proliferative response of normal T and B cells to mitogenic stimulation nor the cytokine secretory capacity of these cells (e.g. interleukin-2, -4, -6 and gamma-interferon) and monocytes (e.g. interleukin-1 beta and -6) were affected by the drug. In contrast, on the mononuclear cells obtained from the lupus patients not only did 15-deoxyspergualin inhibit the spontaneous production of polyclonal and anti-DNA IgG antibodies but also suppressed interleukin-1 beta secretion from the monocytes. Other functional responses of T and B cells and monocytes from lupus patients, including mitogenic activation and cytokine secretion, were not altered by the drug. These data suggest that 15-deoxyspergualin possesses a novel mechanism of pharmacological immunosuppression apparently different from that of other immunosuppressants, such as cyclosporin A, FK506 and corticosteroids, that seems to be primarily displayed at the level of autoreactive B cells and monocytes.

In vitro and ex vivo effect of tiaprofenic acid on human peripheral blood mononuclear cells.

The effect of the non-steroidal anti-inflammatory drug (NSAID) tiaprofenic acid on different human immune parameters was investigated in vitro or following in vivo administration in healthy adult volunteers. Results from the in vitro study demonstrated an increased mitogen-induced blastogenesis and interleukin 2 (IL-2) production together with a reduced polyclonal immunoglobulin (Ig) secretion in the presence of the drug. Results from the ex vivo study showed that treatment with tiaprofenic acid had no significant effects on the immune parameters investigated, i.e. unstimulated and mitogen-induced proliferation and IL-2 production, spontaneous and stimulated Ig synthesis, lymphocyte subpopulations, serum Ig and complement levels.