Leland Shapiro

Overview of interleukin-18: more than an interferon-gamma inducing factor.

Initially described in 1989 as interferon‐γ (IFN‐γ) inducing factor (IGIF), interleukin‐18 (IL‐18) is a novel proinflammatory cytokine that is clearly more than an inducer of IFN‐γ The cytokine possesses several biological properties such as activation of nuclear factor‐κB (NF‐κB), Fas ligand expression, the induction of both CC and CXC chemokines, and increased production of competent human immunodeficiency virus. Most activities are due to a receptor complex that recruits the IL‐1 receptor‐activating kinase (IRAK), leading to translocation of NF‐κB. This property and others support the concept that IL‐18 is related to the IL‐1 family. Indeed, one of the IL‐18 receptor chains is the IL‐1 receptor‐related protein, a member of the IL‐1R family. In addition, IL‐18 is structurally similar to IL‐1β and like IL‐1β is first synthesized as a leaderless precursor requiring the IL‐1β converting enzyme for cleavage into an active molecule. The biology of IL‐18 is reviewed in the overview and the implication for a role for this cytokine in disease is presented. J. Leukoc. Biol. 63: 658–664; 1998.

Cytokines in acute myocardial infarction: selective increase in circulating tumor necrosis factor, its soluble receptor, and interleukin-1 receptor antagonist.

Summary Cytokines play a pathogenetic role in a variety of infective and inflammatory diseases. In the present study, we had two objectives: (a) to define the kinetics of tumor necrosis factor (TNF) in plasma after acute myocardial infarction (AMI) in patients treated with early thrombolysis, and (b) to measure other cytokines, interleukin-1 (IL-1) and TNF receptor antagonists, in plasma. TNF-α, but not IL-1β or IL-8, was present in plasma of 6 of 7 patients with severe AMI (Killip class 3 or 4). No TNF (<50 pg/ml) was detected in a group of 11 patients with uncomplicated myocardial infarction (Killip class 1) or in control patients without AMI. Soluble TNF receptor type I and IL-1 receptor antagonist (IL-1Ra) were also significantly increased in the group with severe AMI compared with those with uncomplicated AMI. Circulating TNF is increased only in AMI complicated by heart failure at hospital admission. This finding may have diagnostic and therapeutic relevance.

Interleukin-1β-induced Rat Pancreatic Islet Nitric Oxide Synthesis Requires Both the p38 and Extracellular Signal-regulated Kinase 1/2 Mitogen-activated Protein Kinases

Interleukin-1β (IL-1β) is cytotoxic to rat pancreatic β-cells by inhibiting glucose oxidation, causing DNA damage and inducing apoptosis. Nitric oxide (NO) is a necessary but not sufficient mediator of these effects. IL-1β induced kinase activity toward Elk-1, activation transcription factor 2, c-Jun, and heat shock protein 25 in rat islets. By Western blotting with phosphospecific antibodies and by immunocomplex kinase assay, IL-1β was shown to activate extracellular signal-regulated kinase (ERK) 1/2 and p38 mitogen-activated protein kinase (p38) in islets and rat insulinoma cells. Specific ERK1/2 and p38 inhibitors individually reduced but in combination blocked IL-1β-mediated islet NO synthesis, and reverse transcription-polymerase chain reaction of inducible NO synthase mRNA showed that ERK1/2 and p38 controlled IL-1β-induced islet inducible NO synthase expression at the transcriptional level. Hyperosmolarity caused phosphorylation of Elk-1, activation transcription factor 2, and heat shock protein 25 and activation of ERK1/2 and p38 in islets comparable to that induced by IL-1β but did not lead to NO synthesis. Inhibition of p38 but not of ERK1/2 attenuated IL-1β-mediated inhibition of glucose-stimulated insulin release. We conclude that ERK1/2 and p38 activation is necessary but not sufficient for IL-1β-mediated β-cell NO synthesis and that p38 is involved in signaling of NO-independent effects of IL-1β in β-cells.

Exercise improvement and plasma biomarker changes with intravenous treprostinil therapy for pulmonary arterial hypertension: A placebo-controlled trial

BACKGROUNDnPulmonary arterial hypertension (PAH) remains a poorly understood and frequently lethal disease with few treatment options.nnnMETHODSnWe conducted a placebo-controlled trial of intravenous treprostinil, a prostacyclin analog, in treatment-naive PAH patients. During 12 weeks of treatment with treprostinil or placebo, we quantified 6-minute walk distance (6MW), clinical symptoms and 11 cytokines/growth factors.nnnRESULTSnForty-two of 44 study patients had idiopathic/familial PAH in New York Heart Association (NYHA) Class III. Treprostinil increased 6MW by a placebo-corrected median of 83 meters (p = 0.008; mean increase 93 +/- 42 meters), reduced Borg score by a median 2.0 units (p = 0.02), and improved NYHA class by a median of 1.0 (p = 0.02). There was a trend toward improved survival with treprostinil (p = 0.051). Baseline plasma angiopoietin-2 (Ang-2), vascular endothelial growth factor (VEGF), matrix metalloproteinase-9 (MMP-9) and platelet-derived growth factor (PDGF) were elevated compared with reported normal ranges. Treatment with treprostinil was associated with decreased Ang-2 levels. Improvement in 6MW distance after treatment was associated with reductions in Ang-2 and MMP-9 levels. Most of the cytokines and growth factors studied were not abnormal with disease nor did they change with treatment.nnnCONCLUSIONSnWe conclude that treprostinil treatment significantly improved exercise capacity, dyspnea and functional class. Several plasma proteins that might track disease were abnormal at baseline, and changes were associated with improved exercise capacity.

α1-Antitrypsin monotherapy induces immune tolerance during islet allograft transplantation in mice

Human pancreatic islet transplantation offers diabetic patients tight glucose control but has low graft survival rates. The immunosuppressive drugs that are administered to graft recipients lack the antiinflammatory benefits of corticosteroids because of their diabetogenic effects. The serum protease inhibitor α1-antitrypsin (AAT) possesses antiinflammatory properties and reduces cytokine-mediated islet damage. In the present study, diabetic mice were grafted with allogeneic islets and treated with AAT monotherapy (n = 24). After 14 days of treatment, mice remained normoglycemic and islet allografts were functional for up to 120 treatment-free days. After graft removal and retransplantation, mice accepted same-strain islets but rejected third-strain islets, thus confirming that specific immune tolerance had been induced. Explanted grafts exhibited a population of T regulatory cells in transplant sites. According to RT-PCR, grafts contained high levels of mRNA for foxp3, cytotoxic T lymphocyte antigen-4, TGF-β, IL-10, and IL-1 receptor antagonist; expression of proinflammatory mediators was low or absent. After implantation of skin allografts, AAT-treated mice had greater numbers of foxp3-positive cells in draining lymph nodes (DLNs) compared with control treatment mice. Moreover, dendritic cells in DLNs exhibited an immature phenotype with decreased CD86 activation marker. Although the number of CD3 transcripts decreased in the DLNs, AAT did not affect IL-2 activity in vitro. Thus, AAT monotherapy provides allografts with antiinflammatory conditions that favor development of antigen-specific T regulatory cells. Because AAT treatment in humans is safe, its use during human islet transplantation may be considered.

Alpha-1-antitrypsin inhibits human immunodeficiency virus type 1

Several observations suggest the existence of potent endogenous suppressors of human immunodeficiency virus type 1 (HIV‐1) production, and inhibitors of serine proteases may participate in this effect. Alpha‐1‐antitrypsin (AAT) is the most abundant circulating serine protease inhibitor. Physiological AAT concentrations inhibited HIV‐1 production in chronically infected U1 monocytic cells, reduced virus replication in freshly infected peripheral blood mononuclear cells, and blocked infection of permissive HeLa cells. In U1 cells, AAT suppressed activation of the HIV‐1‐inducing transcription factor NF‐κB. Similar results were obtained using CE‐2072, a synthetic inhibitor of host serine proteases. HIV‐1 did not replicate in blood obtained from healthy volunteers, but marked replication was observed in blood from individuals with hereditary AAT deficiency. These results identify AAT as a candidate circulating HIV‐1 inhibitor in vivo. Two different mechanisms of AAT‐induced HIV‐1 inhibition were identified, including reduced HIV‐1 infectivity and blockade of HIV‐1 production. A novel host pathogen interaction is suggested, and an alternative strategy to treat HIV‐1‐related disease may be possible.—Shapiro, L., Pott, G. B., Ralston, A. H. Alpha‐1‐antitrypsin inhibits human immunodeficiency virus type 1. FASEB J. 15, 122–115 (2001)

α-1-Antitrypsin is an endogenous inhibitor of proinflammatory cytokine production in whole blood

Several observations suggest endogenous suppressors of inflammatory mediators are present in human blood. α‐1‐Antitrypsin (AAT) is the most abundant serine protease inhibitor in blood, and AAT possesses anti‐inflammatory activity in vitro and in vivo. Here, we show that in vitro stimulation of whole blood from persons with a genetic AAT deficiency resulted in enhanced cytokine production compared with blood from healthy subjects. Using whole blood from healthy subjects, dilution of blood with RPMI tissue‐culture medium, followed by incubation for 18 h, increased spontaneous production of IL‐8, TNF‐α, IL‐1β, and IL‐1R antagonist (IL‐1Ra) significantly, compared with undiluted blood. Dilution‐induced cytokine production suggested the presence of one or more circulating inhibitors of cytokine synthesis present in blood. Serially diluting blood with tissue‐culture medium in the presence of cytokine stimulation with heat‐killed Staphylococcus epidermidis (S. epi) resulted in 1.2‐ to 55‐fold increases in cytokine production compared with S. epi stimulation alone. Diluting blood with autologous plasma did not increase the production of IL‐8, TNF‐α, IL‐1β, or IL‐1Ra, suggesting that the endogenous, inhibitory activity of blood resided in plasma. In whole blood, diluted and stimulated with S. epi, exogenous AAT inhibited IL‐8, IL‐6, TNF‐α, and IL‐1β significantly but did not suppress induction of the anti‐inflammatory cytokines IL‐1Ra and IL‐10. These ex vivo and in vitro observations suggest that endogenous AAT in blood contributes to the suppression of proinflammatory cytokine synthesis.

Patients with Nontuberculous Mycobacterial Lung Disease Exhibit Unique Body and Immune Phenotypes

RATIONALEnAmong patients with nontuberculous mycobacterial lung disease is a subset of previously healthy women with a slender body morphotype, often with scoliosis and/or pectus excavatum. We hypothesize that unidentified factors predispose these individuals to pulmonary nontuberculous mycobacterial disease.nnnOBJECTIVESnTo compare body morphotype, serum adipokine levels, and whole-blood cytokine responses of patients with pulmonary nontuberculous mycobacteria (pNTM) with contemporary control subjects who are well matched demographically.nnnMETHODSnWe enrolled 103 patients with pNTM and 101 uninfected control subjects of similar demographics. Body mass index and body fat were quantified. All patients with pNTM and a subset of control subjects were evaluated for scoliosis and pectus excavatum. Serum leptin and adiponectin were measured. Specific cytokines important to host-defense against mycobacteria were measured in whole blood before and after stimulation.nnnMEASUREMENTS AND MAIN RESULTSnPatients with pNTM and control subjects were well matched for age, gender, and race. Patients with pNTM had significantly lower body mass index and body fat and were significantly taller than control subjects. Scoliosis and pectus excavatum were significantly more prevalent in patients with pNTM. The normal relationships between the adipokines and body fat were lost in the patients with pNTM, a novel finding. IFN-γ and IL-10 levels were significantly suppressed in stimulated whole blood of patients with pNTM.nnnCONCLUSIONSnThis is the first study to comprehensively compare body morphotype, adipokines, and cytokine responses between patients with NTM lung disease and demographically matched controls. Our findings suggest a novel, predisposing immunophenotype that should be mechanistically defined.

Clinical, hematologic, and immunologic effects of interleukin-10 in humans.

We conducted a double-blind, placebo-controlled study to investigate the safety, pharmacokinetics, and immunological properties of interleukin-10 (IL-10) administration in healthy humans. Volunteers received a single intravenous bolus injection of recombinant human IL-10 (1, 10, or 25μg/kg) or placebo. Cytokine production in whole blood and peripheral blood mononuclear cells (PBMC) was assessed before and 3, 6, 24, and 48 hr after the injection. Peak serum concentrations of IL-10 (15±1.1, 208±20.1, and 505±22.3 ng/ml) occurred after 2–5 min for 1, 10, and 25μg/kg IL-10, respectively. The terminal-phase half-life was 3.18 hr. A transient leukocytosis (24–63% above baseline) was observed 6 hr after injection, which coincided with a dose-dependent decrease (12–24%) in neutrophil superoxide generation. There was a marked inhibition (60–95%) of endotoxin-induced IL-6 production from whole blood in each group receiving IL-10. Production of IL-8 in endotoxin-stimulated blood was reduced in the 10μg/kg group. In PBMC stimulated with phytohemagglutinin and phorbol ester, there was a decrease (72–87%) in interferon-γ (IFNγ) production 6 hr after IL-10 with a return to pre-IL-10 levels after 24 hr. This reduction was only partially associated with a decrease in the number of CD2-bearing cells. We conclude that IL-10 administration into humans is without significant side effects, and a single injection reducesex vivo production of IL-6, IL-8, and IFNγ.

Monokine Antagonism Is Reduced in Patients With IDDM

Interleukin-1 (IL-1) and tumor necrosis factor α (TNF-α) have been implicated as immune effector molecules in the pathogenesis of insulin-dependent diabetes mellitus (IDDM). Recently, an increased frequency of the A1/A1 genotype of an IL-1 receptor antagonist (IL-1Ra) gene polymorphism was observed in patients with IDDM. Therefore, we investigated plasma IL-1Ra and soluble TNF p55 receptor (TNFsRp55) levels in 18 men with recent-onset IDDM, 10 men with long-standing IDDM, and 35 age-matched healthy men. No differences in plasma IL-1Ra were found among the three groups. However, when the plasma IL-1Ra levels in the subjects with IDDM and the control subjects were analyzed according to IL-1Ra genotypes, we found a 30% lower level of plasma IL-1Ra in subjects with IDDM carrying the A1/A1 genotype compared with the levels in those carrying the A1/A2 genotype (372 ± 40 vs. 530 ± 54 ng/l, respectively, P = 0.025). In contrast, no significant association was seen between plasma IL-1Ra and IL-1Ra genotype in the control subjects. The TNFsRp55 level in heparinized plasma was 17% lower in patients with IDDM than in control subjects (3.93 ± 0.22 vs. 4.72 ± 0.24 μg/l, respectively, P = 0.038). These findings could not be explained by metabolic derangement in the IDDM patients. Although based on a limited number of patients, these preliminary findings suggest that μ-cells in IDDM patients may be more sensitive to the cytotoxic effects of TNF-α and IL-1 because of less production of TNFsRp55 and, in a subset of IDDM patients, of IL-IRa during the inflammatory challenge of insulitis.