Dag Kvale

IL-10 in HIV infection: increasing serum IL-10 levels with disease progression--down-regulatory effect of potent anti-retroviral therapy.

To examine the potential pathogenic role of IL‐10 in HIV infection, we measured serum IL‐10 levels in 51 HIV‐infected patients and 23 healthy controls both on cross‐sectional and longitudinal testing. All clinical groups (Centers for Disease Control (CDC) categories) of HIV‐infected patients had significantly higher circulating IL‐10 levels than controls, with the highest levels among the AIDS patients, particularly in patients with ongoing Mycobacterium avium complex (MAC) infection. Among 32 HIV‐infected patients followed with longitudinal testing (median observation time 39 months), patients with disease progression had increasing IL‐10 levels in serum, in contrast to non‐progressing patients where levels were stable. While both IL‐10 and tumour necrosis factor‐alpha (TNF‐α) increased in patients with disease progression, the IL‐10/TNF‐α ratio decreased in these patients, suggesting imbalance between these two cytokines. Finally, we found that highly active anti‐retroviral therapy (HAART) induced a significant, gradual decrease in IL‐10 levels but without normalization. These findings suggest a pathogenic role for IL‐10 in HIV infection, and may suggest a possible role for immunomodulating therapy which down‐regulates IL‐10 activity in addition to concomitant potent anti‐retroviral therapy in HIV‐infected patients.

Tumor Necrosis Factor (TNF) System Levels in Human Immunodeficiency Virus—Infected Patients during Highly Active Antiretroviral Therapy: Persistent TNF Activation Is Associated with Virologic and Immunologic Treatment Failure

Because persistent tumor necrosis factor (TNF)-alpha activation may play a pathogenic role in human immunodeficiency virus infection, TNF component levels were assessed over 78 weeks in plasma and peripheral blood mononuclear cells (PBMC) during highly active antiretroviral therapy (HAART) in 40 HIV-infected patients. HAART induced a significant decline in plasma levels of TNF-alpha and soluble TNF receptors and was associated with a fall in the abnormally increased unstimulated and a rise in the abnormally low Mycobacterium avium complex-purified-protein derivative-stimulated TNF-alpha released from PBMC. However, concentrations of these TNF components were not normalized. Patients with virologic and immunologic treatment failure after 52 weeks had higher levels of several TNF components than other patients early after initiation of therapy, also during periods with adequate virologic response. Although TNF components significantly decreased during HAART, these results support data indicating that full immunologic normalization is not achieved during such therapy. The persistent activation of the TNF system in a subgroup of persons may be involved in treatment failure.

Expression and regulation of adhesion molecules ICAM-1 (CD54) and LFA-3 (CD58) in human intestinal epithelial cell lines

T cells and other leucocytes regularly occur within and subjacent to the gut epithelium. Recent data suggest that intestinal epithelial cells may exert accessory immunological functions. Although interactions between leucocytes and accessory cells usually require expression of adhesion molecules, intestinal epithelium has generally been considered negative for intercellular adhesion molecule‐1 (ICAM‐1) by immunohistochemical techniques. We therefore studied the expression of ICAM‐1 and lymphocyte function‐associated antigen‐3 (LFA‐3) by two colonic epithelial cell lines and found that both adhesion molecules were constitutively present at low levels. ICAM‐1 protein expression could be enhanced within 4 h by cytokines, particularly alter co‐incubation with interferon‐γ(IFN) and tumour necrosis factor‐α(TNF), interleukin‐1β(IL‐1), or IL‐6. IFN also resulted in accumulation of ICAM‐1 mRNA. Conversely, the LFA‐3 expression was virtually unaffected by cytokine stimulation. These data imply that intestinal epithelial cells under certain conditions may bear adhesion molecules required for cooperation with juxtaposed leucocytes in situ, and that the expression of some of these molecules is modulated by cytokines from activated mucosal leucocytes.

Epithelial expression of HLA, secretory component (poly-Ig receptor), and adhesion molecules in the human alimentary tract.

Epithelial HLA class II is differentially expressed (DR >> DP) only after birth in salivary glands and small intestinal mucosa, in contrast to class I determinants and secretory component (SC) which appear early in gestation. However, there is a brisk postnatal increase in SC expression along with the class II induction, suggesting stimulation by cytokines from activated immune cells. T lymphocytes remain quite scanty in postnatal salivary glands, and the striking SC and class II expression might reflect a synergistic effect of IFN-gamma and TFN-alpha on immature epithelial cells. Enhanced epithelial expression of both SC and class II in salivary glands from sudden infant death victims could be the effect of immunostimulation caused by an infectious agent. Strikingly upregulated SC and epithelial class II expression (DR > DP > DQ) is seen in various inflammatory lesions such as obstructive sialadenitis, Sjögrens syndrome, chronic gastritis, and celiac disease. IFN-gamma and TNF-alpha are most likely involved as the expression patterns can be reproduced with these cytokines in vitro on colonic epithelial cell lines. However, these molecules of the Ig supergene family do not show a selective response in epithelia of inflammatory lesions because increased expression is also seen for lysozyme, lactoferrin and some other proteins. ICAM-1 can be upregulated on epithelial cells by various cytokines in vitro although the situation remains uncertain in mucosal inflammation. The expression pattern in IBD is complicated by dysplastic epithelial changes leading to reduced SC levels which may thus, in turn, jeopardize the poly-Ig transport mechanism. Epithelial class II molecules appear to have antigen-presenting properties, but the immunopathologic role of their increased expression in inflammatory disease in terms of induction of autoimmunity and/or abrogation of oral tolerance is a matter of continuing dispute.

Induction of Various HLA Class II Molecules in a Human Colonic Adenocarcinoma Cell Line

The colonic carcinoma cell line HT‐29 had no constitutive expression of HLA class II molecules. Gamma interferon (IFN‐γ) induced expression of HLA class II molecules in a dosedependent manner with 100 U/ml as an optimal dose. The expression of HLA‐DR, HLA‐DP, and HLA‐DQ molecules seemed to follow different kinetics. While DR and DP molecules were maximally induced after 2 days, DQ molecules appeared later with maximum percentage positive cells after 8 days. Treatment with a prostaglandin synthesis inhibitor (indomethacin) neither induced class II expression nor altered the dose‐response curve for IFN‐γ; this indicated that possible endogenous production of prostaglandins iin this cell line did not interfere with its class II expression. The lectins phytohaemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM), and wheat germ agglutinin (WGA) did not induce class II expression.

An Exploratory Trial of Cyclooxygenase Type 2 Inhibitor in HIV-1 Infection: Downregulated Immune Activation and Improved T Cell-Dependent Vaccine Responses

ABSTRACT Chronic HIV infection is characterized by chronic immune activation and dysfunctional T cells with elevated intracellular cyclic AMP (cAMP), which inhibits the T cell activation capability. cAMP may be induced by prostaglandin E2 following lipopolysaccharide (LPS)-induced upregulation of cyclooxygenase type 2 (COX-2) in monocytes due to the elevated LPS levels in patients with chronic HIV infection. This hypothesis was tested using celecoxib, a COX-2 inhibitor, for 12 weeks in HIV-infected patients without antiretroviral treatment in a prospective, open, randomized exploratory trial. Thirty-one patients were randomized in the trial; 27 completed the study, including 13 patients on celecoxib. Celecoxib reduced chronic immune activation in terms of CD38 density on CD8+ T cells (−24%; P = 0.04), IgA levels (P = 0.04), and a combined score for inflammatory markers (P < 0.05). Celecoxib further reduced the inhibitory surface receptor programmed death 1 (PD-1) on CD8+ T cells (P = 0.01), including PD-1 on the HIV Gag-specific subset (P = 0.02), enhanced the number of CD3+ CD4+ CD25+ CD127lo/− Treg or activated cells (P = 0.02), and improved humoral memory recall responses to a T cell-dependent vaccine (P = 0.04). HIV RNA (P = 0.06) and D dimers (P = 0.07) tended to increase in the controls, whereas interleukin-6 (IL-6) possibly decreased in the treatment arm (P = 0.10). In conclusion, celecoxib downmodulated the immune activation related to clinical progression of chronic HIV infection and improved T cell-dependent functions in vivo.

Microbial translocation in HIV infection is associated with dyslipidemia, insulin resistance, and risk of myocardial infarction.

Objective:Microbial translocation has been suggested to be a driver of immune activation and inflammation. It is hypothesized that microbial translocation may be related to dyslipidemia, insulin resistance, and the risk of coronary heart disease in HIV-infected individuals. Design:Cross-sectional study of 60 HIV-infected patients on combination antiretroviral therapy with viral suppression >2 years and 31 healthy age-matched controls. Methods:Lipopolysaccharide (LPS) was analyzed by limulus amebocyte lysate colorimetric assay. Lipids, including cholesterol, low-density lipoprotein (LDL), and triglycerides, were measured. Glucose metabolism was determined using an oral glucose tolerance test. Body composition was determined using whole-body dual-energy x-ray absorptiometry scans and magnetic resonance imaging. The Framingham risk score was used to assess risk of cardiovascular disease and myocardial infarction. Results:HIV-infected patients had higher level of LPS compared with controls (64 pg/mL vs. 50 pg/mL, P = 0.002). Likewise, HIV-infected patients had higher triglycerides, LDL, and fasting insulin as well as evidence of lower insulin sensitivity compared with controls. Among HIV-infected patients, high LPS was associated with a higher level of triglycerides and LDL and with lower insulin sensitivity. Importantly, among HIV-infected patients, high LPS was associated with a higher Framingham risk score. Conclusions:HIV-infected patients with suppressed viral replication had increased level of microbial translocation as measured by LPS. LPS was associated with cardiometabolic risk factors and increased Framingham risk score. Hence, the gastrointestinal mucosal barrier may be a potential therapeutic target to prevent dyslipidemia and future cardiovascular complications in HIV infection.

HLA- and dose-dependent immunogenicity of a peptide-based HIV-1 immunotherapy candidate (Vacc-4x).

Objective: The Vacc-4x immunotherapy candidate is composed of four modified peptides corresponding to conserved domains of the HIV-1 protein p24 that preferentially include HLA-A2 restricted elements. Dose-dependent safety and immunogenicity of Vacc-4x and the significance of a HLA-A2 haplotype were examined. Design: Non-AIDS, HIV-1 infected healthy patients (n = 40) stable on HAART with CD4 counts > 300 × 106 cells/l were randomized to receive either low-dose or high-dose Vacc-4x over 26 weeks in an open, prospective phase II clinical trial. Methods: Patients received a total of 10 intradermal injections, using recombinant granulocyte-macrophage colony stimulating factor as a local adjuvant. Vacc-4x-specific cellular responses were monitored in vivo by delayed-type hypersensitivity (DTH) skin test infiltrates and in vitro by both T-cell proliferation, and induction /secretion of cytokines. Results: Most patients developed Vacc-4x-specific DTHs (90%) and proliferative T-cell responses (80%) that were inter-related in magnitude. High-dose Vacc-4x generally induced stronger specific immune responses than low dose in terms of DTH areas and CD4 and CD8 T-cell proliferation. Only HLA-A2 negative patients had a definite dose advantage, and this subgroup had in fact the best overall DTH and proliferative responses. In contrast, no significant dose difference was observed for HLA-A2 positive patients. No serious adverse events were reported. Conclusions: HIV-associated specific responses were safely induced in most patients by Vacc-4x in a dose-dependent manner and were also influenced by the HLA haplotype.

Different regulatory pathways employed in cytokine‐enhanced expression of secretory component and epithelial HLA class I genes

The transmembrane secretory component (SC, or pIg receptor) plays a crucial role in mucosal immunity by translocating dimeric IgA and pentameric IgM through exocrine epithelia. This receptor is up‐regulated by cytokines in parallel with increased epithelial HLA expression. By use of the human epithelial cell line HT‐29m3, we show that IFN‐γ, TNF‐α and IL‐4 activate transcription of the SC gene. This activation was slow, suggesting mediation via newly synthesized protein factors. IFN‐γ and TNF‐α, but not IL‐4, also up‐regulated expression of HLA class I genes. However, this gene induction was rapid and did not depend on new protein synthesis. Nuclear run‐on experiments showed that the transcription rate of HLA class I genes nearly peaked after only 30 min of IFN‐γ or TNF‐α stimulation, whereas the SC transcription rate did not peak until after 20 – 36 h of IFN‐γ, TNF‐α or IL‐4 stimulation. Gel electrophoresis mobility shift assays demonstrated binding of nuclear proteins from cytokine‐stimulated HT‐29 cells to consensus elements in the promoter of the SC gene, involving the binding site for the nuclear factor‐κB p50 subunit after TNF‐α stimulation, and IFN‐stimulated response element after IFN‐γ stimulation (and weakly after TNF‐α). Our observations in vitro likely parallel events in vivo by which activated mucosal T cells and macrophages enhance pIg receptor‐mediated external transport of secretory IgA and IgM and up‐regulate epithelial HLA expression.

Immune modulation of adhesion molecules ICAM-1 (CD54) and LFA-3 (CD58) in human hepatocytic cell lines.

Interactions between leukocytes such as T cells and accessory or target cells are promoted by adhesion molecules, in particular intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function-associated antigen-3 (LFA-3). Hepatocytes are usually negative for these surface membrane proteins which, however, may be up-regulated in inflammatory processes within the liver. Because the regulatory signals for, and tissue distribution of, these adhesion molecules vary among different tissues, expression of ICAM-1 and LFA-3 was studied in the Hep-G2 and SK-Hep-1 human hepatocytic cell lines in vitro. Low, constitutive membrane expression of the two molecules was detected in both cell lines. ICAM-1, but not LFA-3, was rapidly up-regulated by interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF), and to some extent by interferon-gamma (IFN) and IL-6, whereas IL-4 had variable low effects, if any. Considerable synergism on ICAM-1 protein levels was observed after stimulation with TNF, IL-1, and IFN, whereas co-incubation with actinomycin D abolished these effects. ICAM-1 mRNA levels increased 16-20 times after cytokine incubation. Our data indicated that hepatocytes share the regulatory pathways for ICAM-1 described for several other cell types. Absence of these molecules in vivo may reflect a dominance of negative modulation signals in the normal liver, which might also explain the low levels of HLA class I molecules.