Inflammation plays a key role in coronary artery disease (CAD), but whether it is involved in the pathogenesis of syndrome X (SX) is not known. Thus, we assessed the presence of systemic inflammation in patients with SX and its possible relation to infections from Helicobacter pylori, Chlamydia pneumoniae, cytomegalovirus, and Epstein-Barr virus. We studied 55 patients with SX (57 +/- 8 years old; <em>27</em> women), 49 with stable angina and obstructive CAD (56 +/- 8 years old; 24 women), and 60 healthy controls (57 +/- 11 years old; 24 women). Plasma levels of high-sensitivity C-reactive protein and <em>interleukin</em>-1 receptor antagonist were measured in all patients. Infection from Helicobacter pylori, Chlamydia pneumoniae, cytomegalovirus, and Epstein-Barr virus was assessed in 43 patients with SX, 40 patients with CAD, and in 39 controls. Patients with SX had lower serum levels of C-reactive protein than did patients with CAD (4.06 +/- 6.8 vs 5.99 +/- 7.8 mg/L, p = 0.013) but higher levels of C-reactive protein than did controls (1.75 +/- 1.98 mg/L; p = 0.008). Plasma levels of <em>interleukin</em>-1 receptor antagonist were higher in patients with CAD (570 +/- 738 pg/ml) and patients with SX (494 +/- 677 pg/ml) than in controls (254 +/- 174, pg/ml; p = 0.0003 vs CAD and p = 0.013 vs SX) but did not differ significantly between patients with CAD or SX (p = 0.20). There were no differences across groups in the prevalence of infection from Helicobacter pylori, Chlamydia pneumoniae, cytomegalovirus, and Epstein-Barr virus and in the prevalence of 1, 2, 3, and 4 infections (p = 0.99). Among patients with SX, no correlation was found between markers of inflammation and indexes of disease activity (angina episodes, exercise test results). Our data show evidence of increased low-grade systemic inflammation in patients with cardiac SX, which was unrelated to an increased infectious pathogen burden.

OBJECTIVE

Levels of some cytokines are significantly higher in the vitreous fluid of patients with acute uveitis than in normal vitreous fluid. The authors sought to determine which proinflammatory cytokines were upregulated in the vitreous fluid of patients with ocular sarcoidosis.

METHODS

Samples of vitreous fluid were collected from patients with sarcoid uveitis and from nonsarcoid control patients with idiopathic epiretinal membrane. The levels of <em>27</em> proinflammatory cytokines were measured with a multiplex beads array system. Postvitrectomy macular thickness was also measured by using spectral domain optical coherence tomography (SD-OCT). To assess the relationship between cytokine levels and disease stage, the authors divided patients into three groups based on macular thickness 1 month after operation.

RESULTS

The vitreous levels of 17 cytokines were significantly higher in patients with ocular sarcoidosis than in nonsarcoid controls. Serum levels of interferon γ-induced protein 10 (IP-10) were also higher in ocular sarcoidosis patients than in nonsarcoid controls. Conversely, serum levels of interleukin (IL) 15 in ocular sarcoidosis patients were lower than in the control group. Analysis of cytokine levels and macular thickness revealed that IL-1ra, IL-4, IL-8, IFN-γ, IP-10, monocyte chemotactic protein (MCP)-1, macrophage inflammatory protein (MIP)-1β, and regulated on activation, normal T-cell expressed and secreted (RANTES) were significantly upregulated in patients with thin cystoid macular edema group.

CONCLUSIONS

Patients with ocular sarcoidosis had elevated levels of proinflammatory cytokines in vitreous fluids. Different cytokines might contribute to different stages of macular edema.

A murine T cell hybridoma, constructed by fusion of alloantigen-activated T cells with the BW5147 T cell lymphoma, which produces a lymphokine capable of inducing tumoricidal activity in macrophages, has been identified. Lymphokine release could be detected only after mitogen stimulation of the T cell hybridoma culture. Upon cloning of the parental hybridoma, 24 out of <em>27</em> clones produced tumoricidal-inducing activity. Seven clones produced more cytocidal-inducing activity than did conventional supernatants, generated by concanavalin A stimulation of normal murine spleen cell cultures, which contained macrophage-activating factor (MAF). The supernatant of hybridoma clone 24/G1 was 25 times more active than conventional MAF preparations. Using supernatants from a variety of clones, the levels of macrophage-activating activity and <em>interleukin</em> 2 were found to vary independently of one another. The lymphokine produced by hybridoma clone 24/G1 appeared to be identical to conventional MAF by a variety of criteria including: (a) a requirement for a second signal for induction of tumoricidal activity in macrophages, (b) inactivation after incubation for 1 h at 65 degrees C, and (c) loss of activity after treatment at pH 4.0 but not at pH 5.0. Like conventional MAF, the hybridoma MAF eluted as a single peak after molecular sieve chromatography on Sephadex G100 and exhibited an apparent molecular weight of 55,000. Although somewhat heterogeneous, the majority of hybridoma 24/G1 MAF displayed an isoelectric point of 5.4 as determined using the chromatofocusing technique. These results thus illustrate the usefulness of T cell hybridomas in distinguishing between various lymphokine activities and indicate that the T cell hybridoma clone 24/G1 will be of particular usefulness in achieving the biochemical purification of substantial quantities of murine MAF.

OBJECTIVE

To study the role of Toll-like receptor (TLR)2 and 4 in the onset of joint inflammation and cartilage destruction during immune complex-mediated arthritis (ICA), and its relationship with FcgammaR expression.

METHODS

ICA was induced in knee joints of TLR2-/- and TLR4-/- mice and their wild-type controls. Joint inflammation and cartilage destruction were measured in the knee joint using histology. mRNA levels were determined in synovial specimens and macrophages using quantitative polymerase chain reaction and cytokine protein levels in synovial washouts using Bioplex.

RESULTS

Joint inflammation and cartilage destruction were not different in arthritic TLR2-/- and wild-type mice. By contrast, at day 1 after ICA induction, joint swelling and proteoglycan depletion in knee joints of TLR4-/- mice were considerably lower (inflammation 68-79% and proteoglycan depletion <em>27</em>-76%) when compared with wild-type controls. Cytokine production at this time point was markedly reduced in TLR4-/- mice (<em>interleukin</em> (IL)1, IL6, macrophage inflammatory chemokine (MIP)-1alpha and keratinocyte-derived chemokine 49%, 72%, 68% and 84%, respectively). In arthritic synovia of TLR4-/- mice, and also after injection of the antigen poly-l-lysine (PLL) lysozyme alone, mRNA levels of FcgammaR, and the FcgammaR regulating cytokine IL10 were considerably lower. Stimulation of peritoneal macrophages with PLL lysozyme up regulated mRNA levels of FcgammaR and IL10, whereas neutralisation by anti-IL10 antibodies largely blocked FcgammaR up regulation. At day 4, joint inflammation and cartilage destruction were comparable in TLR4-/- mice and wild-type controls.

CONCLUSIONS

TLR4 regulates early onset of joint inflammation and cartilage destruction during ICA arthritis by up regulation of FcgammaR expression and enhanced cytokine production. TLR4-mediated up regulation of FcgammaR is largely mediated by IL10.

BACKGROUND

The purpose of this study was to assess proinflammatory markers in blunt trauma patients regarding the relationship of these and blood loss and duration of surgery in different fracture locations.

METHODS

Prospective, multicenter, nonrandomized cohort study.

METHODS

Three level I trauma centers.

METHODS

Sixty-eight blunt trauma patients, who did not require emergency operations and had sustained truncal or extremity fractures, were included. In two index patient groups, patients with spinal fractures (group SF, n = 24) and pelvic and acetabular fractures (group PAF, n = 21) underwent fixation of their fractures and were compared with a group of patients with isolated fractures (group FF, n = 28). Ten healthy volunteers served as controls.

METHODS

Internal fixation of pelvic, acetabular and spinal fractures, intramedullary nailing of femoral fractures, measurement of proinflammatory cytokines.

METHODS

From serially sampled central venous blood, the perioperative concentrations of interleukin-6 (IL-6) and IL-8 were evaluated during a 24-hour period and set into relation with the duration of surgery and the degree of blood loss.

RESULTS

Intramedullary instrumentation for isolated PAF caused a significant perioperative increase in the concentrations of IL-6 (preoperative: 16 pg/mL +/- 12 pg/mL, 7 hours: 89 pg/mL +/- 15 pg/mL, and 24 hours: 107 pg/mL +/- 27 pg/mL, p < 0.05). This increase was comparable with the isolated femoral fracture (group FF: IL-6 preoperative, 52 pg/mL +/- 12 pg/mL; 7 hours, 78 pg/mL +/- 14 pg/mL; and 24 hours, 120 pg/mL +/- 23 pg/mL, p = 0.02). The changes observed after spinal fracture fixations (group SF) were considerably lower (IL-6 preoperative: 11 pg/mL +/- 6 pg/mL, 7 hours: 16 pg/mL +/- 11 pg/mL, and 24 hours: 56 pg/mL +/- 19 pg/mL). The percent change of baseline IL-6 and IL-8 concentrations, and the blood loss in group PAF at 24 hours were positively correlated (IL-6 r = 0.72, p < 0.03, IL-8 0.67, p = 004) after insertion. No correlation with the duration of surgery was found.

CONCLUSIONS

The release of proinflammatory cytokines was higher in patients when their pelvic fractures were operated than in patients with spine fracture fixations, and was associated with the degree of blood loss. A higher increase in cytokine levels occurred when they were performed early (day 1-2) across all patient groups. The level of the released markers seems to be related to the magnitude of surgery, rather than to the duration of the procedure. This study supports the value of immunologic markers in determining subclinical changes during and after orthopedic surgical procedures.

OBJECTIVE

We studied the immunologic molecules in cerebrospinal fluid (CSF) and discussed their evolutional changes in pediatric patients with Rasmussen syndrome (RS).

METHODS

CSF samples collected from <em>27</em> patients with RS (average onset age, 7.5 +/- 5.6 years) were studied. Cell count, protein, glucose, albumin, chloride, and immunoglobulin G (IgG) levels were measured by conventional methods. Surface markers of lymphocytes in CSF were examined by a cell sorter. Granzyme B, interferon gamma (IFNgamma), <em>interleukin</em> 4 (IL-4), tumor necrosis factor alpha (TNFalpha), and IL-12 in CSF were quantitated by enzyme-linked immunosorbent assay (ELISA). Autoantibodies against GluR epsilon2 (NR2B) were examined by immunoblot.

RESULTS

The data of the first CSF examination showed that IgG levels (Mann-Whitney U test, p < 0.01), CD4(+) T cells (p = 0.02), TNFalpha levels (p < 0.01), and Granzyme B levels (p < 0.01) were elevated compared with disease controls. White blood cell count, IFNgamma level, IL-12 level, and Granzyme B level were elevated, especially in the early stage of disease. CD4(+) T cells, CD8(+) cells, CD3(+) T cells, IgG levels, and TNFalpha levels were elevated at all stages of disease evolution. Protein levels and albumin levels were elevated in the progressed stage. Autoantibodies against GluR epsilon2 (NR2B) (IgG) were found in 50% of patients in the early stage, and the positive rate was low at the progressed stage.

CONCLUSIONS

The present findings suggest that complex pathophysiologic mechanisms involving CD4(+) T cells and CD8(+) T cells change evolutionally during the progression of RS. A crucial cytotoxic process occurs in the early stage, and declines in the progressed stage.

Septicemia is the most severe form of melioidosis caused by the Gram-negative bacterium, Burkholderia pseudomallei. Here, we show that levels of IL-<em>27</em>p28 transcript and protein were both significantly elevated in patients with sepsis, particularly melioidosis and in patients with unfavorable disease outcome. Moreover, human monocytes/macrophages and neutrophils were the major source of IL-<em>27</em> during infection. The addition of exogenous IL-<em>27</em> in vitro resulted in significantly increased bacterial survival, reduced B. pseudomallei-induced oxidative burst, and enhanced IL-1β and TNF-α production by purified neutrophils from healthy subjects. Finally, blockade of endogenous IL-<em>27</em> in neutrophils using soluble IL-<em>27</em> receptor antagonist prior to infection led to significantly reduced survival of bacteria and decreased IL-1β, but not TNF-α production. These results indicate a potential role for IL-<em>27</em> in the suppression of anti-bacterial defense mechanisms that might contribute to disease severity in sepsis. The targeting of this cytokine may be beneficial in the management of human sepsis.

<em>Interleukin</em> 6 (IL-6) is a potent immunomodulatory cytokine that has pathogenic and prognostic significance in a number of disorders. Previous studies in Hodgkin's disease (HD) have demonstrated the association between elevated serum levels of IL-6 and unfavourable prognosis, including advanced stage and the presence of 'B' symptoms and with reduced survival. Although IL-6 expression has been demonstrated in both the malignant Hodgkin/Reed-Sternberg (HRS) cells and in the various non-malignant cells present in HD biopsies, a relationship between expression of IL-6 by the tumour and outcome measures has not been established. The study group comprised of 97 patients with advanced HD who were recruited to two related clinical trials. IL-6 expression was determined on paraffin-wax sections of biopsy material by means of an immunohistochemical assay. Of the 97 patients, <em>27</em> (28%) showed staining for IL-6 in HRS cells. IL-6 expression by HRS cells was significantly correlated with a decreased likelihood of achieving a complete response to chemotherapy (P = 0.02) and with an increased prevalence of 'B' symptoms (P = 0.04). IL-6 expression by HRS cells was not associated with Epstein-Barr virus status (P = 0.57). In summary, the results suggest that IL-6 expression by HRS cells may contribute to the presence of 'B' symptoms and to a decreased likelihood to achieve a complete remission in HD patients.

B cells bifurcating along 'type 1' or 'type 2' pathways under the influence of polarizing cytokines can, in turn, influence the direction of an immune response. Here, we compare the capacity of human B cells residing within naïve and memory compartments to participate in type 1 polarizing responses. B-cell receptor (BCR) engagement provided the main signal for <em>interleukin</em> (IL)-12Rbeta1 expression in the two subsets: this was potentiated by CD154 together with interferon-gamma (IFN-gamma) but inhibited by IL-12. IL-12Rbeta2 could be induced on a minority of B cells by the same signals, and also by IFN-gamma alone. WSX-1, a receptor for IL-<em>27</em>, was expressed in both subsets with no evidence for its regulation by the signals studied. While neither subset was capable of secreting much IL-12 p70, memory B cells could produce a small amount of IL-12 p40 on CD40 ligation. Memory B cells also, exclusively, expressed IL-23 p19 mRNA on BCR triggering. Importantly, products of appropriately stimulated memory--but not naive--B cells were shown to promote the synthesis of IFN-gamma in uncommitted T-helper cells. The data indicate an equal capacity for naïve and memory B cells to respond within a type 1 polarizing environment. Although poorly equipped for initiating type 1 responses, B cells--by virtue of the memory subset--reveal a capacity for their maintenance and amplification following T-dependent signalling.

BACKGROUND

The secretion of heat shock protein (HSP) <em>27</em>, HSP60, HSP70, HSP90alpha, 20S proteasome, and their correlations to proinflammatory cytokine <em>interleukin</em>-6 is unknown in patients undergoing on-pump versus off-pump coronary artery bypass graft (CABG) operation.

METHODS

Forty patients were included in this explorative study (on- versus off-pump CABG, each n = 20). Serum samples were obtained before and 30 minutes, 60 minutes, and 24 hours after CABG operation. Enzyme-linked immunosorbent assay technique was utilized to determine soluble HSP<em>27</em>, 60, 70, and 90alpha, 20S proteasome, and levels of <em>interleukin</em>-6.

RESULTS

Serum levels of HSP are increased in patients undergoing on-pump CABG operation as compared with off-pump CABG technique. These differences were highly significant for HSP<em>27</em>, 70, and 90alpha at 60 minutes after initiation of cardiopulmonary bypass (all, p < 0.001). Concentrations of soluble 20S proteasome were increased 24 hours after operation in on- and off-pump CABG patients (p < 0.001) and correlated significantly with the serum content of HSP <em>27</em>, 70, and 90alpha at 60 minutes after initiation of cardiopulmonary bypass (p < 0.001). No correlation was found when comparing <em>interleukin</em>-6 levels with intravascular leakage of HSP and 20S proteasome after CABG operation.

CONCLUSIONS

We conclude from our data that the innate immune system is activated owing to spillage of known immune modulatory and apoptosis-associated proteins after CABG operation.

Atherosclerosis, and the resulting coronary heart disease and stroke, is the most common cause of death in developed countries. Atherosclerosis is an inflammatory process that results in the development of complex lesions or plaques that protrude into the arterial lumen. Plaque rupture and thrombosis result in the acute clinical complications of myocardial infarction (MI) and stroke. Although certain risk factors (dyslipidemias, diabetes, hypertension) and humoral markers of plaque vulnerability (C-reactive protein, <em>interleukin</em>-6, 10 and 18, CD40L) have been identified, a highly sensitive and specific biomarker or protein profile, which could provide information on the stability/vulnerability of atherosclerotic lesions, remains to be identified. In this review, we report several proteomic approaches which have been applied to circulating or resident cells, atherosclerotic plaques or plasma, in the search for new proteins that could be used as cardiovascular biomarkers. First, an example using a differential proteomic approach (2-DE and MS) comparing the secretome from control mammary arteries and atherosclerotic plaques is displayed. Among the different proteins identified, we showed that low levels of HSP-<em>27</em> could be a potential marker of atherosclerosis. Second, we have revised several studies performed in cells involved in the pathogenesis of atherosclerosis (foam cells and smooth muscle cells). Another approach consists of performing proteomic analysis on circulating cells or plasma, which will provide a global view of the whole body response to atherosclerotic aggression. Circulating cells can bear information reflecting directly an inflammatory or pro-coagulant state related to the pathology. As an illustration, we report that circulating monocytes and plasma in patients with acute coronary syndromes has disclosed that mature Cathepsin D is increased both in the plasma and monocytes of these patients. Finally, the problems of applying proteomic approach directly to plasma will be discussed. The purpose of this review is to provide the reader with an overview of different proteomic approaches that can be used to identify new biomarkers in vascular diseases.

BACKGROUND

Repeated exposure to HIV is not always associated with infection and multiple cohorts of HIV-exposed but seronegative individuals (ESN) have been described. HIV-specific CD4 and CD8 T lymphocytes are detected both in HIV patients and in ESN; we verified whether different patterns of HIV-specific memory T lymphocytes would be detected in individuals in whom exposure to HIV results or does not result in infection.

METHODS

Gag-specific T cells were analysed in 15 ESN, 14 HIV patients, and 15 healthy controls using extensive flow cytometry analysis.

RESULTS

Data confirmed that gag-specific T lymphocytes are present in ESN. Gag-specific T cells mainly secrete <em>interleukin</em>-2 in ESN and interferon-gamma in HIV patients. In addition the CD4/CD8 and the memory/naive ratios are altered, central memory (45RA-/CCR7+) CD4 and CD8 T lymphocytes are more abundant, and terminally differentiated (45RA+/CCR7- and <em>27</em>-/28-) CD8 T lymphocytes are augmented in ESN individuals.

CONCLUSIONS

Exposure to HIV occurs in high risk seronegative individuals; the observation that naive cells and CM are skewed in ESN indicate that this exposure is robust enough to modulate the CM/EM ratio. The increase in late effectors and in natural killer cells seen in ESN suggests a role for these cells in preventing actual infection.

BACKGROUND

The role of leukocyte migration and chimerism in organ allograft acceptance has been obscured by the lack of information about the late localization of the donor cells.

METHODS

Male Lewis rat->>female Brown Norway abdominal heart transplantation was performed under tacrolimus immunosuppression (days 0-13, 20, and <em>27</em>) with or without donor bone marrow and (in bone marrow subgroups) a 1-week postoperative course of a possibly chimerism-enhancing drug. Using rat sex-determining region-Y-specific oligonucleotide primers, we determined the donor DNA concentration by polymerase chain reaction in serial venous blood samples for 100 days and in tissue specimens when animals were killed.

RESULTS

Chimerism was detected out to 56 days in 89% of the blood samples but in none of the samples at 100 days. However, donor DNA was detected when animals were killed in 95% of the native hearts, 80% of the skin biopsy specimens, and 23% of the spleens. The presence and quantity of early and late chimerism were strongly correlated the administration of adjunct bone marrow and with a reduction in the vasculopathy and inflammation index in the cardiac allografts. Marginally significant further increases in chimerism and/or reductions in chronic heart rejection beyond those achieved with adjunct bone marrow alone were associated with additional treatment with the growth factors Flt-3 ligand, granulocyte colony-stimulating factor, and a recombinant molecular variant of interleukin-6 (interleukin-6 mutein) but not with hepatocyte growth factor or lisofylline.

CONCLUSIONS

The previously suspected shift of early chimerism in the blood and lymphoid organs to dominance in host nonlymphoid tissues is consistent with the dual mechanisms of clonal exhaustion and immune indifference, governed by antigen migration and localization, that have been postulated elsewhere to account for organ allograft acceptance.

Recently, new complexities in cell fate decision for helper T cells have emerged. One new lineage, which has come to be called Th17 cells, selectively produces proinflammatory cytokines including <em>interleukin</em>-17 (IL-17, A and F), IL-21, and IL-22. In conjunction with transforming growth factor beta-1 (TGFbeta-1), IL-6, IL-21, and IL-23, which activate the transcription factor, signal transducer, and activator of transcription 3 (Stat3), the expression of another transcription factor, retinoic acid-related orphan receptor-gammat (RORgammat) leads to the differentiation of Th17 cells in mice. Other cytokines including IL-2, IL-4, interferon-gamma (IFN-gamma), and IL-<em>27</em> inhibit Th17 differentiation. However, IL-2 acting with TGFbeta-1 induces differentiation of naïve CD4+ T cells to become regulatory T cells (Tregs). Th17 cells are now known to play an important role not only in the pathogenesis of inflammation and autoimmune diseases, but also host defense against extracellular bacteria. Conversely, extensive data substantiate the role of Tregs as essential in maintenance of peripheral tolerance. Selectively targeting Tregs and Th17 cells are likely to be important strategies in the treatment of inflammatory and autoimmune diseases in humans.

<em>Interleukin</em> 1 (IL1) increased phosphorylation of the small heat-shock protein (hsp <em>27</em>) in MRC5 fibroblasts. The increase was maintained for at least 30 min, but levels had returned to pre-stimulation values by 2 h. When hsp <em>27</em> was metabolically labelled with [3H]leucine, about 15% was phosphorylated in resting confluent cells; this rose to 90% upon stimulation by IL1. Peptide maps of the three differently charged phosphorylated forms were consistent with their arising by phosphorylation of increasing numbers of serine residues. IL1 had the same effect on hsp <em>27</em> in pig articular chondrocytes, endothelial cells from human umbilical vein and an epidermoid carcinoma cell line (KB). Certain other agents were found selectively to increase phosphorylation of hsp <em>27</em> in MRC5 cells besides IL1 [and tumour necrosis factor (TNF)]. Platelet-derived growth factor had a similar effect to that of IL1; bradykinin, acid fibroblast growth factor and ATP caused an intermediate effect; phorbol myristate acetate (PMA) and 1-oleoyl-2-acetylglycerol had smaller effects. Dibutyryl cyclic AMP and forskolin had no effects on hsp <em>27</em> phosphorylation. When cells had been depleted of protein kinase C (PKC) by prolonged treatment with PMA, stimulation by IL1, TNF or bradykinin still increased hsp <em>27</em> phosphorylation. The stimulation by all three agents was also unaffected by the PKC inhibitor staurosporine. IL1, TNF and bradykinin each caused hsp <em>27</em> phosphorylation by a pathway independent of PKC. The results are consistent with IL1 activating a serine kinase which remains to be identified.

The effects of ribavirin and interferon (IFN) alpha have been investigated on cultured peripheral blood mononuclear cells, obtained from 15 patients with chronic hepatitis C virus (HCV) infection. At clinically relevant serum concentrations achieved during therapeutic administration, ribavirin did inhibit moderately the mitogen-stimulated mononuclear cell proliferation and growth of the CD4+ and CD8+ T cell subsets without apparent cytolysis. The ribavirin-IFN-alpha combination showed activity against HCV with disappearance of HCV RNA in <em>27</em>% of cases, and a synergy in the inducibility of the intracellular enzyme 2',5'-oligoadenylate synthetase. Such ribavirin concentrations induced modest increases in the T helper 1-like cytokine production by mononuclear cells. Higher ribavirin concentrations markedly inhibited IFN-gamma production, but augmented <em>interleukins</em> (IL) 2, 4, and 12 secretion. Conversely, IFN-alpha tended to suppress IL 2, 4 and 12, but enhanced IFN-gamma and IL-10 secretion. Thus, ribavirin and IFN-alpha appear to cause diverse effects on immunoregulatory cytokine secretion, and when combined, counteracted for production of IL-2 and IL-12, while upregulated mononuclear cell secretion of IFN-gamma and that of the anti-inflammatory cytokine IL-10. These findings suggest a non-cytolytic modulation of inflammatory responses induced by the drug combination, that may be relevant in the pathophysiology of chronic HCV infection.

BACKGROUND

All-trans retinoic acid (ATRA) has antiproliferative and anti-inflammatory effects and is currently used in the treatment of leukemia and dermatologic diseases. We tested the therapeutic potential of ATRA on anti-glomerular basement membrane (GBM) glomerulonephritis rats.

METHODS

Glomerulonephritis was induced in male Wistar-Kyoto rats on day 0 by an intravenous injection of antirat GBM antibody. On day 14 after the induction of anti-GBM glomerulonephritis, some rats were sacrificed (N = 5). Another 10 rats were divided into two groups: the vehicle group (N = 5) and the ATRA treated group (N = 5). ATRA was orally administrated from day 14 to day <em>27</em> after disease induction. Blood pressure, body weight, urinary protein excretion, and blood chemistry was determined on days 1, 14, 21, and <em>27</em>. Kidney samples were obtained on day 28. The kidneys were examined with periodic acid-Schiff staining (PAS) and immunohistochemistry using antibodies against the proliferative cell nuclear antigen (PCNA), rat monocyte and macrophage (ED-1), and alpha-smooth muscle actin (alpha-SMA). Glomerular RNA was extracted from isolated glomeruli, and reverse transcription (RT) followed by polymerase chain reaction (PCR) was performed.

RESULTS

ATRA administration produced a 55% reduction of proteinuria in glomerulonephritis rats. Light microscopic analysis revealed severe necrosis/crescent formation (>50% of the glomerulus) affecting 34% of glomeruli in vehicle rats, whereas ATRA treatment reduced the glomeruli showing severe change to 14%. ATRA also significantly reduced PCNA-positive cells, ED-1-positive cells and alpha-SMA-positive area in the glomeruli. RT-PCR analyses revealed that a wide variety of genes including inflammation related [tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and CCAAT enhancer-binding protein delta (C/EBPdelta)], cell proliferation-related [platelet-derived growth factor (PDGF)] and fibrosis-related [transforming growth factor-beta1 (TGF-beta1), type I collagen, and alpha-SMA) genes were suppressed in the glomeruli of ATRA-treated rats.

CONCLUSIONS

ATRA administration significantly reduced severe necrosis/crescent formation and urinary protein excretion in glomerulonephritis rats. Suppression of a wide variety of gene expression may partly explain the mechanism of ATRA's antiproliferative and anti-inflammatory effects. These data suggest a novel therapeutic application of ATRA toward glomerulonephritis.

OBJECTIVE

This study sought to examine plasma levels of soluble Fas/APO-1 receptor (sFas), an inhibitor of apoptosis, and soluble Fas ligand (sFas-L), an inducer of apoptosis, and their relation to each other and to other clinical variables, such as New York Heart Association functional class, tumor necrosis factor (TNF) and interleukin-6 (IL-6) in congestive heart failure (CHF).

BACKGROUND

It has been recently reported that apoptotic cell death occurs in myocytes of dogs with CHF. Hypoxia is frequently seen in advanced CHF and can stimulate Fas/APO-1 receptors (Fas) to induce apoptosis in cultured myocytes. Fas and Fas ligand (Fas-L) are cell-surface proteins and representative apoptosis-signaling molecules. Fas on the cell membrane induces apoptosis when it binds Fas-L or sFas-L. However, plasma sFas, a molecule lacking the transmembrane domain of Fas, blocks apoptosis by inhibiting binding between Fas and Fas-L or sFas-L on the cell membrane. At present, it is unknown whether plasma sFas-L and plasma sFas increase in the presence of cardiac disease.

METHODS

The study included 70 patients (mean [+/-SEM] age 65 +/- 2 years, range 21 to 93) with chronic CHF (coronary artery disease in 28, dilated cardiomyopathy in 27, valvular heart disease in 15) and 62 age- and gender-matched normal control subjects. Plasma levels of sFas, sFas-L, TNF-alpha and IL-6 were measured by enzyme-linked immunosorbent assays using monoclonal anti-human antibodies.

RESULTS

There was no significant difference in sFas-L levels between normal subjects and patients in functional classes I to IV; however, sFas increased with severity of functional classification, independent of the underlying disease. sFas levels were significantly higher even in patients in functional class II than in normal subjects and those in functional class I, and were highest in patients in functional class IV (normal subjects; 2.2 +/- 0.1 ng/ml; functional class I: 2.2 +/- 0.2 ng/ml; functional class II: 3.1 +/- 0.2 ng/ml; functional class III: 3.9 +/- 0.3 ng/ml; functional class IV: 5.1 +/- 0.6 ng/ml). Plasma sFas levels were significantly higher in patients with elevated pulmonary artery wedge pressure and a decresed cardiac index than in those with values in the normal range. In patients in functional class IV, there was no significant difference in plasma sFas levels between the survivors and non-survivors during 6-month follow-up. However, plasma levels of sFas tended to decrease in nine patients with clinical improvement (baseline sFas: 5.2 +/- 0.8 ng/ml; 6-month sFas: 4.3 +/- 0.5 ng/ml, p = 0.07) but were similar in patients with no change in functional class. TNF-alpha and IL-6 were increased significantly only in patients in functional class IV, as previously reported, but were not related to sFas.

CONCLUSIONS

We found elevated levels of plasma sFas and no increase in plasma sFas-L in human CHF. The increase in sFas may play an important role in the pathophysiologic mechanisms of CHF.

Cytokines are upregulated in prediabetes, but their relationship with Enterovirus (EV) infection and development of islet autoimmunity is unknown. Cytokines (n = 65) were measured using Luminex xMAP technology in a nested case-control study of 67 children with a first-degree relative with type 1 diabetes: <em>27</em> with islet autoantibodies (Ab(+)) and 40 age-matched persistently autoantibody negative (Ab(-)) control subjects. Of 74 samples, 37 (50%) were EV-PCR(+) in plasma and/or stool (EV(+)) and the remainder were negative for EV and other viruses (EV(-)). Fifteen cytokines, chemokines, and growth factors were elevated (P ≤ 0.01) in Ab(+) versus Ab(-) children (<em>interleukin</em> [IL]-1β, IL-5, IL-7, IL-12(p70), IL-16, IL-17, IL-20, IL-21, IL-28A, tumor necrosis factor-α, chemokine C-C motif ligand [CCL]13, CCL26, chemokine C-X-C motif ligand 5, granulocyte-macrophage colony-stimulating factor, and thrombopoietin); most have proinflammatory effects. In EV(+) versus EV(-) children, IL-10 was higher (P = 0.005), while IL-21 was lower (P = 0.008). Cytokine levels did not differ between Ab(+)EV(+) and Ab(+)EV(-) children. Heat maps demonstrated clustering of some proinflammatory cytokines in Ab(+) children, suggesting they are coordinately regulated. In conclusion, children with islet autoimmunity demonstrate higher levels of multiple cytokines, consistent with an active inflammatory process in the prediabetic state, which is unrelated to coincident EV infection. Apart from differences in IL-10 and IL-21, EV infection was not associated with a specific cytokine profile.

The biologic behavior of invasive pituitary adenomas is poorly understood and appears to be intermediate between intrasellar adenomas and pituitary carcinoma. Utilizing immunohistochemistry on tissue sections, we examined 10 invasive pituitary adenomas involving the sphenoid sinus with a panel of cellular and molecular markers and compared the results with 10 noninvasive pituitary adenomas and five normal pituitary glands. The panel included Ki-67 and PCNA (proliferation cell nuclear antigen), cell proliferation indicators; <em>interleukin</em> 6 (IL-6), a multifunctional cytokine; heat shock protein <em>27</em> (HSP <em>27</em>), a stress response protein; and p53, a tumor suppressor gene product. We found strong expression of IL-6 in normal pituitary glands, 20% of noninvasive and 50% of invasive adenomas. HSP <em>27</em> was absent in normal glands and present in 10% of noninvasive and 50% of invasive adenomas. The Ki-67 and PCNA proliferation index was above normal but was essentially the same in the two groups. p53 was not detected in any specimens. We propose that both IL-6 and HSP <em>27</em> may be markers of biologic aggression in pituitary adenomas.

BACKGROUND

Immune mechanisms of extracorporeal photochemotherapy (ECP) in refractory/resistant graft-versus-host disease (GvHD) are complex. We have previously analyzed the role of CD4CD25Foxp3 regulatory T cells (T-regs).

METHODS

In the current study, we have enlarged the size of the population (n=<em>27</em>; chronic GvHD=18, acute GvHD=9) for a median follow-up of 24 months. T-regs were monitored for CD4, CD25, glucocorticoid-induced tumor necrosis factor receptor (GITR), CD62L, CCR7, Foxp3, and STAT-5. Immune analysis by <em>interleukin</em> (IL)-17 Elispot was carried out on circulating T-helper CD4 cells secreting IL-17, a subset of T cells considered relevant in the pathogenesis of GvHD.

RESULTS

We confirm that ECP is accompanied by a significant increase of CD4CD25Foxp3GITRCD62LCCR7 T-regs. Sorted T-regs show augmented phosphorylation of STAT-5. Only ECP-responding patients demonstrate a raise of circulating T-regs, being mostly affected by chronic GvHD. Moreover, this phenomenon corresponds to a diminished secretion of IL-17.

CONCLUSIONS

In conclusion, our study shows that T-regs represent important immune mediators of the clinical benefits of ECP in patients affected by GvHD.

Plasma <em>interleukin</em>-1 beta (Il-1 beta) and tumor necrosis factor-alpha (TNF-alpha) concentrations were measured twice, at a 48-hour interval, in <em>27</em> drug-free obsessive-compulsive patients (12 women and 15 men) and in <em>27</em> sex-age-matched healthy controls. Il-1 beta and TNF-alpha concentrations were significantly lower in patients than in controls, whereas there were no differences in either group between men and women, between the samples of the two days, or, in the patients, between those who had and those who had not been previously treated with psychopharmacologic drugs.

The aim of this study was 2-fold: (1) to examine insulin resistance, blood lipid levels, and inflammatory markers in 9- to 11.5-year-old obese and lean children and (2) to identify factors that influence insulin resistance in this cohort of youths. Body mass index, skinfold thickness, waist circumference, physical activity (4-day triaxial accelerometer), cardiorespiratory fitness (submaximal bicycle ergometer test), and dietary intake (3-day food records) were evaluated in <em>27</em> obese and <em>27</em> lean boys and girls. Fasting blood samples were analyzed for insulin, glucose, lipids and lipoproteins, C-reactive protein (CRP), <em>interleukin</em> 6, soluble intercellular adhesion molecule, and soluble vascular cell adhesion molecule. Homeostasis model assessment (HOMA) was used to evaluate insulin resistance (HOMA-IR). Obese children presented higher HOMA-IR, CRP, and blood lipid levels (all P < .01) compared with lean children. Total body fat and waist circumference were positively associated with fasting insulin (r>> or = 0.51), HOMA-IR (r>> or = 0.56), CRP (r>> or = 0.51), and blood triacylglycerol (r>> or = 0.38), and were inversely correlated with high-density lipoprotein cholesterol (r>> or = -0.39; all P < .01). Cardiorespiratory fitness was inversely associated with HOMA-IR (r = -0.24; P < .05), but this association disappeared when adjusted for age, sex, and fat mass. Waist circumference and total daily physical activity explained 49% of the variance in HOMA-IR in these children. In conclusion, these findings suggest that total and central adiposity are positively associated and physical activity is negatively associated with insulin resistance in children. Interventions to improve glucose metabolism in youth should target at reducing total body and abdominal fat and increasing physical activity. The lack of association between inflammatory markers and HOMA-IR suggests that obesity may precede the elevation of these markers in the evolution of insulin resistance in youth.

Hypertriglyceridemia is often associated with small dense low density lipoprotein (LDL), elevated remnants, and decreased high density lipoprotein (HDL)-cholesterol (C), which comprise the dyslipidemic triad. The objective of this study was to investigate the effect of fenofibrate on the lipoprotein subfraction profile and inflammation markers in hypertriglyceridemic men. Twenty hypertriglyceridemic men were administered fenofibrate, 200 mg daily, for 8 weeks. Lipoprotein subclasses were measured by nuclear magnetic resonance (NMR) spectroscopy. Inflammation markers including C-reactive protein (CRP), <em>interleukin</em>-6 (IL-6), and monocyte chemotactic protein-1 (MCP-1) were also determined. Fenofibrate lowered triglyceride (TG) by 58% and increased HDL-C by 18%. NMR analysis revealed that very low density lipoprotein (VLDL), particularly large VLDL, intermediate density lipoprotein (IDL), and small LDL, were significantly decreased, and LDL distribution shifted towards the larger particles. HDL distribution was altered; there was an increase in small HDL and a decrease in large HDL, resulting in a significant decrease in HDL particle size, from 9.1 to 8.9 nm, as well as a <em>27</em>% increase in HDL particle number. Among inflammation markers, CRP was significantly decreased by 42%. In conclusion, fenofibrate effectively improves atherogenic dyslipidemia by reducing remnants and small LDL, as well as by increasing HDL particles. These effects, together with the favorable effect on inflammation, might provide a clinical benefit in hypertriglyceridemic subjects.