G. Mascia

Alteration of serotonin transporter density and activity in fibromyalgia

The aim of the study was to evaluate the kinetic parameters of a specific serotonin transporter (SERT) and serotonin uptake in a mentally healthy subset of patients with fibromyalgia. Platelets were obtained from 40 patients and 38 healthy controls. SERT expression and functionality were evaluated through the measurement of [3H]paroxetine binding and the [3H]serotonin uptake itself. The values of maximal membrane binding capacity (Bmax) were statistically lower in the patients than in the healthy volunteers, whereas the dissociation constant (Kd) did not show any statistically significant variations. Moreover, a decrease in the maximal uptake rate of SERT (Vmax) was demonstrated in the platelets of patients, whereas the Michaelis constant (Km) did not show any statistically significant variations. Symptom severity score (tiredness, tender points index and Fibromyalgia Impact Questionnaire) were negatively correlated with Bmax and with Vmax, and positively correlated with Km. A change in SERT seems to occur in fibromyalgic patients, and it seems to be related to the severity of fibromyalgic symptoms.

The trace element content of top-soil and wild edible mushroom samples collected in Tuscany, Italy

The amount of the trace elements As, Ba, Cd, Cr, Cu, Hg, Li, Mn, Ni, Pb, Rb, Se, Sr, and Zn was measured in top soils and edible mushrooms, Boletus edulis, Macrolepiota procera, collected at five distinct green microhabitats inside the Lucca province, North-Central Italy (years 2008–2009). Results showed a top soil element content within the Italian statutory limits. Concerning the amount of mushroom elements, we observed significant species-differences obtaining higher levels of Ni, Rb, and Se in B. edulis or As, Pb, Cu in M. procera. Bioaccumulation factors (BCFs: element in mushroom/element in soil) resulted species-dependent and element-selective: in particular, B. edulis preferentially accumulated Se (BCFs varying from 14 to 153), while M. procera mainly concentrated Cu (BCFs varying from 5 to 15). As well, both species displayed between-site BCF differences. By a multivariate principal component approach, cluster analysis (CA), we could resolve two main clusters of soil element composition, corresponding to the most ecologically divergent sites. Besides, CA showed no cluster relating to element contents of B. edulis at the different collection sites, while a separation in groups was found for M. procera composition with respect to harvesting locations, suggesting uptake systems, in this saprotrophic species, sensitive to microhabitat. Regarding consumer safety, Cd, Hg, Pb levels resulted sometime relevant in present samples, never reaching values from current literature on mushrooms collected in urban-polluted areas. Our findings encourage a deeper assessment of the molecular mechanisms of metal intake by edible mushrooms, encompassing genetic biochemical and geo-ecological variables, with particular awareness to element bioavailability in soils and fungi.

ATP, calcium and magnesium levels in platelets of patients with primary fibromyalgia

OBJECTIVES To evaluate the intracellular levels of the high energy adenosine triphosphate nucleotide ATP and essential divalent cations, calcium and magnesium, in platelets of patients affected by primary fibromyalgia syndrome (FMs). DESIGN AND METHOD Platelet ATP and cation concentrations were measured in 25 patients affected by FMs and 25 healthy volunteers through a chemiluminescent and a fluorimetric assay, respectively. RESULTS Significant lower ATP levels were observed inside platelets of FM patients (fmol ATP/plt: 0.0169+/-0.0012 vs. healthy controls, fmol ATP/plt: 0.0306+/-0.0023, mean+/-SEM) (*** P<0.0001). A trend towards higher calcium concentrations (P=0.06) together with significant increased magnesium levels were also reported in platelets of patients by comparison with controls (P=0.02). CONCLUSIONS This preliminary study suggests that disturbances in the homeostasis of platelet ATP metabolism-signaling and calcium-magnesium flows might have a relevance in the pathogenesis of FMs.

Neuropeptide Y in the Brain and Retina of the Adult Teleost Gilthead Seabream (Sparus aurata L.)

The presence of neuropeptide Y (NPY) in the brain and retina of gilthead seabream (Sparus aurata L.) was investigated for the first time. For this investigation we employed an immunoperoxidase technique and the western immunoblot analysis using an antiserum raised against porcine NPY. The results showed that NPY‐immunoreactivity was widely distributed in the brain of S. aurata. In particular, we have found NPY‐immunoreactive (ir) neurons in the area ventralis telencephali pars centralis and pars lateralis, in the area dorsali telencephali pars centralis subdivision two and in nucleus intermedius thalami. An intense NPY‐ir was detected in the telencephalon, in the optic tectum, in the thalamus, hypothalamus and in the vagal lobes. Scarce positive fibres were seen in the olfactory bulbs. NPY‐ir amacrine cells were observed in the retina. The western immunoblot analysis revealed a protein band with a mobility corresponding to that of synthetic NPY. Our findings are, in general, in agreement with those obtained in other teleosts. The extensive distribution of NPY indicates for this peptide a key role in basic physiological actions, including visual and gustatory inputs processing.

Autoradiographic localization and binding study of benzodiazepines receptor sites in carp brain (Cyprinus carpio L.)

This study demonstrates, for the first time, by both autoradiography and binding assay that [3H]Ro 15-1788 binds to carp brain with a high degree of anatomical selectivity. Saturation binding of the radioligand was determined in seven anatomically defined regions and suggested the presence of one class of binding sites (Type I-lke). In general, there was a good correlation between the autoradiographic and the binding data. By far, the optic tectum and the vagal, facial, and glossopharyngeal lobes showed the majority of [3H]Ro 15-1788 binding sites. Low to negative concentration of binding sites was detected in the cerebellum. The location of [3H]Ro 15-1788 binding sites in particular brain regions, indicates that benzodiazepine receptors could be associated with pathways involved in the control of basic central functions as spatial learning acquisition and retention, and feeding behaviour.

Autoradiographic distribution of neuropeptide Y binding sites in the brain of the carp Cyprinus carpio L. (Cyprinidae, Teleostei)

The present study reports the distribution of neuropeptide Y (NPY)-binding sites in the brain of the adult carp Cyprinus carpio L. Radioiodinated NPY was used as tracer in the autoradiographic procedure. The NPY-binding sites (NPY-bs) were widely distributed in the carp brain. Generally, a good match was observed between the distribution of NPY-bs and the distribution of NPY-immunoreactive (NPY-ir) elements previously reported in the forebrain of the carp. Low to moderate concentration of NPY-bs were found in the telencephalon, this finding indicates that NPY may play a role in the processing of olfactory inputs and in more complex behaviours like spatial learning acquisition and retention, whose importance could correlated with similar results obtained in mammals. Moreover, in the rhombencephalon, the presence of NPY-bs at level of lobus vagus and the lobus facialis suggests that NPY may be implicated in food-seeking behaviour and swallowing reflex.

Decreased elastase activity in circulating granulocytes of systemic sclerosis patients : a possible pathogenetic role

The central event in the pathogenesis of systemic sclerosis (SSc) is an abnormal accumulation of extracellular matrix (ECM) components (1). Human neutrophil elastase (HNE) is a serine protease that might be implicated directly or indirectly in the excessive deposition of ECM, being involved in marked conversion of latent metalloproteinases (MMPs) to lower molecular weight forms, consistent with active MMPs (2) that contribute to matrix remodelling. The control of its degradative activity is of considerable importance because of its wide substrate specificity leading to activity towards a variety of proteins with relevance to SSc: elastin, types III and IV collagen, proteoglycans, and fibronectin (3). Granulocytes and lymphomonocytes are involved in the inflammatory process and our aim was to characterize the biochemical properties of elastase in circulating granulocytes and lymphomonocytes of SSc patients, by means of enzymatic assays, Western blot, and gel chromatography techniques. The activity and levels of elastase and of the major elastase inhibitor a1-anti-trypsin (a1-PI) was also investigated in serum. A total of 30 patients (26 females, four males, mean age 56.1 years) with SSc, classified as limited (n520) or diffuse cutaneous (n510) (4), were studied. The activity score varied between 0.5 and 3.5 (5) and the severity score varied between 2 and 6 (6). Most of the patients (80%) were treated with a calcium antagonist; those who had been taking corticosteroids, D-penicillamine, or non-steroidal anti-inflammatory drugs (NSAIDs) that inhibited HNE were excluded. A further group of 30 controls with age and sex comparable to the patients was studied. Elastase activity and a1-PI concentration were assayed spectrophotometrically (7). HNE activity was lower in circulating granulocytes and sera of SSc patients compared with control subjects [granulocytes: median (25th and 75th percentiles): 93.0 (16.5–42.3) vs. 230.5 (186.5–291.5) mIU/mg, pv0.0001; sera: 15.5 (13.0–19.0) vs. 19.5 (17.0–21.0) mIU/mg, pv0.05] while the values obtained in lymphomonocytes were comparable. No differences in KMvalues were found (KM5Michaelis Menten constant. A constant that is equal to the substrate concentration at which an entyme reaction proceeds at help the maximum velocity). No differences in enzymatic activity were found between patients with diffuse SSc and those with limited cutaneous SSc. The correlation between HNE activity and several parameters (disease onset, activity, severity score, and age) was not significant. Serum levels of a1-PI of the SSc patients [28.0 (23.5–30.0) mmol/L] were not different from the control subjects [24.5 (23.5–27.0) mmol/L], although appreciable individual variation was present in the SSc subjects. As a significantly lower HNE activity was found only in the granulocytes, sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) followed by immunoblot analysis was performed on granulocyte

Autoradiographic distribution of peripheral benzodiazepine receptors in the retina of the albino rabbit, Lepus cunicula

The distribution of peripheral benzodiazepine receptors (PBRs) in the retina of the albino rabbit, Lepus cunicula, was studied by autoradiography using [3H]-PK11195, a isoquinoline carboxamide, as a tracer. Autoradiograms obtained by directly placing the slides containing the retina sections on tritium-sensitive film provide evidence for the presence of PBRs in rabbit retina. Furthermore, the dark field examination of photomicrographs taken from autoradiograms showed two dense horizontal bands corresponding to the outer and inner photoreceptor segments, and to the inner plexiform layer. The retinal regions where [3H]-PK11195 binding was more dense are rich in mitochondria, suggesting that as in other neuronal tissues, retinal PBRs are involved in the mitochondrial activity.