A study was carried out to obtain a more detailed picture of the phenotypes of human otosclerotic and normal bone cells and to analyse the response of both populations to treatment with TGF beta 1. ...Total collagen synthesis was found to be decreased, but fibronectin secretion increased in otosclerotic with respect to normal cells. Although overall glycosaminoglycan (GAG) synthesis was lower in otosclerotic cells, the sulphated GAG to hyaluronic acid (HA) ratio was higher, in particular there was greater expression of chondroitin (CS) and dermatan sulphates (DS). TGF beta 1 induced a more marked increase in collagen and fibronectin release and greater production of sulphated GAGs as DS and heparan sulphate (HS) in the otosclerotic cells. The fact that the phenotype of the otosclerotic cells differed from that of the normal cells and could be modified by TGF beta 1 treatment, suggests that TGF beta 1 is implicated in the pathogenesis of otosclerosis.
In the present study, we demonstrate that both interleukin-1 (IL-1) and interleukin-6 (IL-6) induced a significant decrease in glycosaminoglycan (GAG) synthesis and, more strikingly, secretion by 7 ...and 13 day-old chick embryo skin fibroblasts. We demonstrated that interleukin treatment also inhibited the synthesis of collagenase-digestible proteins (type I collagen). In addition, tissue culture supernatants (conditioned media, CM) were tested for reactivity for IL specific ELISAs and for their ability to stimulate proliferative responses in mouse thymocytes and hybridoma cells. Our findings demonstrate that chick embryo skin fibroblasts spontaneously produce IL-1 and, in even greater amounts, IL-6. Highest levels of interleukin secretion were found in the CM of 13 day-old fibroblasts and the IL-1 beta isoform was predominant over IL-1 alpha. Pretreatment of the fibroblasts with either IL-1 or IL-6 increased the secretion of both cytokines. Increased IL-1 levels were correlated with enhanced IL-1 bioactivity in the CM of IL-6 treated fibroblasts. By contrast, the raised concentrations of IL-1 in the CM of IL-1 treated cells and IL-6 in the CM of IL-1 or IL-6 treated fibroblasts failed to translate into augmented bioactivity. These observations, taken together, indicated that IL-1 and IL-6 are able to regulate the synthesis and secretion of ECM macromolecules of developing connective tissues and the cytokine release by chick embryo skin fibroblasts.
Caprine arthritis/encephalitis (CAE) of goats and occasionally sheep are persistent virus infections caused by a lentivirus (CAEV). This viral infection results in arthritis in adult animals and ...encephalitis in kids. Prognosis for the encephalitic form is normally poor, with substantial economic loss for the farm. In this context an early/fast laboratory diagnosis for CAEV infection could be useful for effective prophylactic action. In this work we performed a quantitative real time PCR designed on the CAEV env gene to detect/quantify in goat/sheep samples, viral RNA or proviral DNA forms of CAEV. This procedure was validated in 15 sheep, experimentally infected with CAEV or with a highly correlated lentivirus (visna maedi, MVV); in addition, a total of 37 clinical goat specimens recruited in CAEV positive herds were analyzed and compared using serological analysis (Elisa and AGID). All samples infected with MVV resulted negative. In sheep experimentally infected with CAEV, proviral DNA was detectable 15 days post infection, whereas the serological methods revealed an indicative positivity after 40-60 days.This method showed a sensitivity of 10(2) env fragments/PCR) with a linear dynamic range of quantitation from 10(3) to 10(7)env fragments/PCR; the R2 correlation coefficient was 0.98. All subjects with a clinical diagnosis for Caprine Arthritis-Encephalitis (CAE) resulted CAEV DNA positive.
Abstract Background Right ventricular dysfunction (RVD) is a major predictor of cardiovascular mortality. Inadequate suppression of the renin-angiotensin-aldosterone system (RAAS) after postural ...manoeuvres favours alterations of left ventricular (LV) function. The effects of RAAS dysregulation on RV performance remain elusive. The present study investigated RV function in hypertensive patients with or without altered RAAS activation. Methods Plasma renin activity (PRA) and plasma aldosterone concentration (PAC) were measured in 104 newly diagnosed hypertensive patients after both supine and upright positioning to assess dynamic changes of RAAS induced by antigravitational stress. Twenty-four–hour ambulatory blood pressure monitoring and echocardiographic evaluation of the right ventricle including tissue Doppler imaging (TDI) were performed. Patients were divided as follows: (1) normal PRA and PAC (N group n = 58), (2) suppressible RAAS after supine positioning (SR group n = 24), and (3), nonsuppressible RAAS (NSR group n = 22). RVD was identified by the TDI-derived myocardial performance index (MPI) calculated with a multisegmental approach. Results Patients in the NSR group had reduced indices of RV function compared with patients in the N and SR groups. MPI of the right ventricle as well as prevalence of RVD were also significantly higher in the NSR group. Regression models showed that inadequate RAAS suppression was independently associated with RVD, regardless of blood pressure values and LV dysfunction (LVD). Conclusions Patients without supine normalization of RAAS display a significant impairment of RV function. Our findings suggest that a dynamic RAAS evaluation may help to identify hypertensive patients at higher risk of RVD.
Interleukin-12 (IL-12) production by human monocytes stimulated with mannoproteins (MPs) of Cryptococcus neoformans was investigated. The results reported show that secreted or cell-associated MPs ...induce an early and significant production of IL-12. MPs show different capabilities to quantitatively affect IL-12 production; MP2, an 8. 2-kDa MP purified from the culture supernatant of C. neoformans, appears to be the most potent stimulator. Cytochalasin B inhibits both internalization and IL-12 induction by MP. In addition, a drastic reduction of IL-12 was observed when monocytes were cultured in the absence of normal human serum or treated with soluble mannan. Early production of IL-12 promotes early secretion of gamma interferon by T cells but does not influence the magnitude of the MP-induced lymphoproliferative response. Overall our results identify cryptococcal antigens responsible for rapid and potent induction of IL-12 in monocytes. MPs appear to regulate IL-12 secretion by internalization via the endocytic pathway and by interaction with monocyte receptors or serum factors.
The existence of a close relationship between apoptosis associated with oxidative stress and the increase of viral progeny in chronically HIV-infected cells has been previously reported. The ...possibility of modulating both phenomena by using an antioxidant such as N-acetylcysteine (NAC) has also been demonstrated. The present investigation was designed to study the role of the nuclear enzyme poly-(ADP-ribose)-polymerase (PARP) when HIV- infected cells are treated with tumour necrosis factor alpha (TNFα), a cytokine capable of inducing both apoptosis and intracellular oxygen free radical production. PARP overexpression may result in a rapid drop of intracellular NAD
+
and ATP concentration, thus contributing to cellular redox imbalance. We have used the specific PARP inhibitor 3- aminobenzamide (3-ABA), alone or in a combination with NAC. 3-ABA was only partially capable of inhibiting viral replication and apoptosis induced by TNFα. In contrast, the combination of NAC and 3-ABA led to an inhibition of apoptosis as well as to a marked decrease in viral particle production, with a parallel replenishment of intracellular reduced glutathione content. The results reported here confirm the potential role of antioxidant drug treatment in specific phases of HIV infection.
The present study was carried out to investigate the potential involvement of cholesterol-rich membrane microdomains in the mobilization of calcium induced by NMDA-receptors (NMDA-R). We herein ...provide evidence that agents interfering with plasma membrane cholesterol (namely, filipin and methyl-β-cyclodextrin (Cdex)) inhibit the NMDA-stimulated influx of calcium in hippocampal cells in culture. Filipin-treated cells maintained their morphology and were able to respond with a calcium influx to high K
+ challenge, whereas Cdex altered both cellular parameters. These results suggest that the NMDA-R can be located in cholesterol-rich membrane microdomains or alternatively that the mechanisms coupling their dynamics in the post-synaptic membrane are dependent on the integrity of the microdomains.
This paper investigates the ability of macrophages and of non-typically phagocitic cells such as fibroblasts to internalize 51Cr-labelled C. albicans in presence or in absence of lectin concanavalin ...A (Con A). The results obtained demonstrate that fibroblasts are also able to internalize C. albicans and that this property is potentiated by the presence of Con A. Lectin modifies only the phenotype of the fibroblast, which, poorly attached to the substrate, is globular in shape. Despite reduced cellular spreading, phagocytosis is stimulated by the lectin. In both cell populations, changes in the organization of some cytoskeletal proteins such as tubulin, actin and alpha-actinin are evident during the C. albicans infection; such rearrangements are more evident and longlasting in the fibroblasts treated with Con A.
We recently suggested that, in muscular dystrophies, the excessive accumulation of adenosine as a result of an altered purine metabolism may contribute to progressive functional deterioration and ...muscle cell death. To verify this hypothesis, we have taken advantage of C2C12 myoblastic cells, which can be differentiated in vitro into multinucleated cells (myotubes). Exposure of both proliferating myoblasts and differentiated myotubes to adenosine or its metabolically-stable analog, 2-chloro-adenosine, resulted in apoptotic cell death and myotube disruption. Cytotoxicity by either nucleoside did not depend upon extracellular adenosine receptors, but, at least in part, by entry into cells via the membrane nitro-benzyl-thio-inosine-sensitive transporter. The adenosine kinase inhibitor, 5-iodotubercidin, prevented 2-chloro-adenosine-induced (but not adenosine-induced) effects, suggesting that an intracellular phosphorylation/activation reaction plays a key role in 2-chloro-adenosine-mediated cytotoxicity. Conversely, adenosine cytotoxicity was aggravated by the addition of homocysteine, suggesting that adenosine effects may be due to the accumulation of S-adenosyl-homocysteine, which blocks intracellular methylation-dependent reactions. Both nucleosides markedly disrupted the myotube structure via an effect on the actin cytoskeleton; however, also for myotubes, there were marked differences in the morphological alterations induced by these two nucleosides. These results show that adenosine and 2-chloro-adenosine induce apoptosis of myogenic cells via completely different metabolic pathways, and are consistent with the hypothesis that adenosine accumulation in dystrophic muscles may represent a novel pathogenetic pathway in muscle diseases.
PDF
