Inflammatory regulation of extracellular matrix remodeling in calcific aortic valve stenosis.

BACKGROUND: Calcific aortic stenosis (AS), the most frequent heart valve disorder in developed countries, leads to the calcification and fibrous thickening of the valve. While several studies have addressed the process of valvular calcification, the molecular pathomechanisms of the extensive matrix remodeling remain unclear. Because inflammation is present in stenotic valves, we hypothesized that the proinflammatory cytokine tumor necrosis factor alpha (TNFalpha) might influence cell proliferation and regulate the expression and activation of matrix metalloproteinases (MMPs)--enzymes that are thought to be involved in calcific AS. METHODS: Immunohistochemistry for leukocytes, TNFalpha, MMP-1, and the endogenous MMP inhibitor tissue inhibitor of metalloproteinase (TIMP)-1 was performed on human stenotic (n = 19) and control (n = 8) valves. Primary cultures of human aortic valve myofibroblasts were incubated with and without TNFalpha, and cell proliferation was assessed. The expression and activation of MMP-1 were detected by Western blotting and a specific MMP-1 activity assay. RESULTS: Control valves showed scattered macrophages and low expression of TNFalpha, MMP-1, and TIMP-1. In stenotic valves, leukocyte infiltration and a strong, colocalized expression of TNFalpha and MMP-1 were present, while TIMP-1 remained unchanged. Double-label immunofluorescence localized TNFalpha mainly to macrophages. In cultured human aortic valve myofibroblasts, TNFalpha stimulated proliferation and induced a time-dependent increase in MMP-1 expression and activation, while TIMP-1 remained unchanged. CONCLUSION: The results indicate that matrix remodeling in calcific AS involves the expression and activation of MMPs. Activated leukocytes, by the secretion of TNFalpha, may stimulate valvular myofibroblasts to proliferate and express MMPs, thus regulating actively the matrix remodeling in calcific AS.     

Portal hypertension in Williams syndrome: report of two patients

Williams or Williams-Beuren syndrome (WBS) is a developmental disorder with multisystemic manifestations characterized by distinctive facial features, mental disability with unique cognitive and personality profiles, vascular stenoses, growth retardation, and occasional infantile hypercalcemia, caused by haploinsufficiency for genes deleted in chromosome band 7q11.23. However, with the exception of arterial stenoses caused by haploinsufficiency for the elastin gene (ELN), no specific implication of any other gene in the phenotype has been established. We present two patients with portal hypertension leading to splenomegaly and pancytopenia carrying the common 1.5 Mb WBS deletion. We propose this is an additional severe vascular complication of ELN deficiency and discuss the specific characteristics of the portal venous tract that could explain the impact of ELN deficiency in that venous territory. This complication is potentially lethal and should thus be considered in any patient with WBS and splenomegaly.     

Inflammatory responses in blood samples of human immunodeficiency virus-infected patients with pulmonary infections

We analyzed the characteristics of the inflammatory response occurring in blood during pulmonary infections in human immunodeficiency virus (HIV)-infected patients. A prospective study of consecutive hospital admissions of HIV-infected patients with new-onset radiologic pulmonary infiltrates was carried out in a tertiary university hospital from April 1998 to May 2001. Plasma cyclic AMP receptor protein (CRP), interleukin 1beta (IL-1beta), IL-6, IL-8, IL-10, and tumor necrosis factor alpha (TNF-alpha) levels were determined at the time of admission and 4, 5, and 6 days later. Patients were included in a protocol addressed to study etiology and outcome of disease. A total of 249 episodes of infection were included, with the main diagnoses being bacterial pneumonia (BP) (118 episodes), Pneumocystis carinii pneumonia (PCP) (41 episodes), and mycobacteriosis (36 episodes). For these three patient groups, at the time of admission the median CRP and cytokine levels were as follows: CRP, 10.2, 3.8 and 5 mg/dl, respectively (P = 0.0001); IL-8, 19, 3, and 2.9 pg/ml (P = 0.045); and TNF-alpha, 46.4, 44, and 75 pg/ml, respectively (P = 0.029). There were no significant differences in levels of IL-1beta, IL-6, or IL-10 among the patient groups. A total of 23 patients died. At the time of admission, HIV-infected patients with BP had higher plasma CRP and IL-8 levels than did PCP and mycobacteriosis patients. TNF-alpha levels were higher in patients with mycobacteriosis. An elevated IL-8 level (>61 pg/ml) at the time of admission was an independent factor associated with higher mortality (odds ratio, 12; 95% confidence interval, 1.2 to 235.5).     

Supernumerary humeral heads of the biceps brachii muscle revisited

Supernumerary humeral heads of the biceps brachii muscle were found in 27 (15.4%) of 175 cadavers. They were bilateral in five cadavers and unilateral in 22 (8 left, 14 right), giving a total of 32 examples in 350 arms (9.1%). Depending on their origin and location, the supernumerary heads were classified as superior, infero-medial, and infero-lateral humeral heads. Previous studies were reviewed using this classification. The infero-medial humeral head was observed in 31 of 350 (9%) arms and was therefore the most common variation. The superior humeral head was observed in five (1.5%). The infero-lateral humeral head was the least common variation, observed only in one (0.3%) of 350 arms. A biceps brachii with three heads was observed in 27 of 350 (7.7%) arms and with four heads in five (1.4%) arms.     

The role of Serratia marcescens porins in antibiotic resistance

The outer membrane permeability of Serratia marcescens was studied by comparing porin-deficient mutants with their parental strains. Omp1-deficient strains were selected by moxalactam resistance, whereas mutants lacking the Omp2 porin were obtained by experimental infection with the SMP2 phage, whose primary receptor is the Omp2 porin. The role of porins was demonstrated in quinolone accumulation assays, where semiquantitative differences in accumulation were observed. Permeability coefficients to cephaloridine of Omp1 mutants were determined and compared with those of the parental strain. The clinical isolates S. marcescens HCPR1 and 866 showed 30- to 200-fold reduced permeability coefficients when Omp1 porin was absent.     

Ionotropic glutamate receptor subunits are differentially regulated in the motoneuronal pools of the rat hypoglossal nucleus in response to axotomy

Unilateral hypoglossal nerve axotomy was used as a model to analyse immunohistochemically the expression of the GluR1, GluR2, GluR3, and GluR4 glutamate receptor subunits of the -amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) subtype and the NR1 subunit of the N-methyl-D-aspartate (NMDA) subtype in the different morphofunctional hypoglossal pools from 1 to 45 days postaxotomy. Following hypoglossal nerve axotomy, the percentage of motoneurons that were GluR1-immunopositive and the labeling intensity for this subunit was increased in some hypoglossal pools. Immunolabeling for the GluR2 subunit was undetectable. These results contrast with the unchanged pattern for these two subunits after sciatic nerve axotomy previously described. Image analysis showed a significant decrease in the intensity of immunohistochemical labeling for the GluR2/3 and GluR4 subunits in motoneurons, although most motoneurons were still immunopositive for these 2 subunits after axotomy. The intensity of immunolabeling for the NR1 subunit was slightly decreased postlesion, whereas the percentage of NR1-immunopositive motoneurons increased. Immunoreactivity returned to basal levels 45 days postlesion. These findings show that in axotomized hypoglossal motoneurons, i) AMPA and NMDA receptor subunits are still expressed, ii) the composition of the ionotropic glutamate receptor subunit pool is subjected to continuous changes during the regeneration process, iii) AMPA receptors, if functional, would have physiological properties different to those in intact motoneurons, and iv) the various AMPA receptor subunits are differentially regulated. The present results also suggest a faster recovery of basal levels of immunoreactivity for caudally localised groups of motoneurons which could reflect a caudo-rostral sequential functional revovery in the hypoglossal nucleus.     

Localization of tyrosine hydroxylase-immunoreactivity in the brain of the Senegalese sole, Solea senegalensis

The localization of catecholamines in the brain of the Senegalese sole was determined by immunohistochemical techniques using antibodies against tyrosine hydroxylase. Although the general pattern of distribution of catecholamines is consistent with that reported in other teleosts, some remarkable differences are observed. The most rostral tyrosine hydroxylase immunoreactive (TH-ir) cells were identified in the olfactory bulbs, in which a clear asymmetry in the number and location of TH-ir perikarya and fibers was observed. The number of TH-ir cells is manifestly higher in the right olfactory bulb, especially in the internal cell layer. TH-ir fibers are also much more abundant in the right bulb, principally in the glomerular and internal cell layers. Other TH-ir cell masses were identified in the ventral telencephalon, preoptic area, caudoventral hypothalamus, posterior tuberculum, synencephalon, isthmic region and rhombencephalon. Surprisingly, no ir cell bodies were identified in the ventromedial thalamic nucleus, which exhibits a large number of TH-ir cells in other teleosts. The presence of TH-ir fibers in the brain of sole is particularly evident within and around the nuclei in which immunoreactive cells are found. However, other zones such as the dorsal telencephalon, posterior commissure, optic tectum, torus semicircularis, reticular formation or inferior olive also displayed TH-ir fibers. TH-ir axons also enter the infundibulum, reaching the proximal pars distalis of the adenohypophysis. The distribution of TH-ir cells and fibers is compared with that observed in other teleosts and is discussed in a comparative context.     

Ionotropic glutamate receptor subunit distribution on hypoglossal motoneuronal pools in the rat

The expression of ionotropic glutamate receptor subunits in the motoneuronal pools of the hypoglossal nucleus was studied using specific antibodies against subunits of the -amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), kainate and N-methyl-D-aspartate (NMDA) subtypes. The highest numbers of intensely immunolabelled motoneurons were found in the dorsal tier and caudoventromedial part of the hypoglossal nucleus with all antibodies except that against the GluR1 AMPA subunit. Labelling for the GluR1 subunit was weak except for caudally located groups of motoneurons which innervate tongue muscles related to respiratory activity. By contrast, most motoneurons were intensely immunostained with antibodies against GluR2/3 and GluR4 subunits of the AMPA subtype. The low staining observed using an antibody specific for the GluR2 subunit (which prevents Ca²⁺-entry through AMPA channels) strongly suggests that AMPA receptors in hypoglossal motoneurons are Ca²⁺-permeable. Immunolabelling for the GluR5/6/7 kainate receptor subunits was found in many motoneuronal somata as well as in thin axon-like profiles and puncta that resembled synaptic boutons. Most motoneurons were intensely immunostained for the NMDA receptor subunit NR1. These results show that the hypoglossal nucleus contains five heterogeneous pools of motoneurons which innervate functionally defined groups of tongue muscles. The uneven expression of the different receptor subunits analysed here could reflect diverse phenotypic properties of hypoglossal motoneurons which might be expected to generate different patterns of motor responses under different physiological or pathological conditions.     

Circulating inflammatory mediators during start of fever in differential diagnosis of gram-negative and gram-positive infections in leukopenic rats

Gram-negative and gram-positive infections have been considered the most important causes of morbidity and mortality in patients with leukopenia following chemotherapy. However, discrimination between bacterial infections and harmless fever episodes is difficult. Because classical inflammatory signs of infection are often absent and fever is frequently the only sign of infection, the aim of this study was to assess the significance of serum interleukin-6 (IL-6), IL-10, macrophage inflammatory protein-2 (MIP-2), procalcitonin (PCT), and C-reactive protein (CRP) patterns in identifying bacterial infections during start of fever in normal and cyclophosphamide-treated (leukopenic) rats following an injection of lipopolysaccharide (LPS) or muramyl dipeptide (MDP) as a model for gram-negative and gram-positive bacterial infections. We found that, compared to normal rats, immunosuppressed animals exhibited significantly higher fevers and lesser production of all mediators, except IL-6, after toxin challenge. Moreover, compared to rats that received MDP, both groups of animals that received an equivalent dose of LPS showed significantly higher fevers and greater increase in serum cytokine levels. Furthermore, in contrast to those in immunocompetent rats, serum levels of IL-6 and MIP-2 were not significantly changed in leukopenic animals after MDP injection. Other serum markers such as PCT and CRP failed to discriminate between bacterial stimuli in both groups of animals. These results suggest that the use of the analyzed serum markers at an early stage of fever could give useful information for the clinician for excluding gram-negative from gram-positive infections.     

Immunity in HIV-1-Infected Adults with a Previous State of Moderate-Severe Immune-Suppression and More Than 500 CD4+ T Cell After Highly Active Antiretroviral Therapy

We evaluated phenotypic and functional parameters of immune restoration of 27 HIV-infected patients on highly active antiretroviral therapy (HAART) (HIV-cases) with HIV-RNA levels below detectable limits at least during 18 months, and CD4+ cell per microliter higher than 500 at the moment of the study and lower than 300 anytime before. These patients were compared with 11 HIV-controls that never had less than 500 CD4+ cell per microliter and 20 healthy-controls (HIV seronegative subjects) in a cross-sectional study. HIV-cases had lower counts of naïve CD4+ than HIV-controls and healthy-controls. HIV-patients (both HIV-cases and HIV-controls) showed higher values of naïve and memory CD8+ counts than healthy-controls. TREC-bearing cell levels were significantly lower in HIV-cases than in healthy-controls. Peripheral blood mononuclear cells (PBMC) cultures, HIV-cases had lower values in proliferation to streptokinase (SK) and tetanus toxin (TT) than in healthy-controls. HIV-cases had lower IFN-γ and higher IL-5 production with pokeweed than healthy-controls (P < 0.01). However, IL-5 production of HIV-cases after TT stimulation was lower than in HIV-controls and healthy-controls. Total IgG and IgG1 levels were significantly higher in HIV-cases than in HIV-controls and healthy-controls. Also, IgM levels were significantly higher in HIV-cases than in healthy-controls. Nevertheless, IgG2 levels were significantly lower in HIV-cases and HIV-controls than in healthy-controls. The levels of specific Igs antipneumococcal capsular polysaccharide and TT were significantly lower in HIV-cases than in healthy-controls. HIV-patients with a previous state of severe-moderate immunosuppression normalizing their CD4+ counts have a incomplete immune reconstitution after HAART. Long-term consequences of this subclinical immune deficiency remain to be determined.