Tiziana Giuliano

Association between thyroid autoimmunity and fibromyalgic disease severity

Our objectives were to investigate thyroid abnormalities and autoimmunity in 120 patients affected by fibromyalgia (FM) and to study their relationships with clinical data and symptoms. Thyroid assessment by means of antithyroglobulin antibodies, antithyroid peroxidase antibodies, free triiodo–thyronine, free thyroxine, and thyroid stimulating hormone analyses was carried out. The clinical parameters “Fibromyalgia Impact Questionnaire”, pain, tender points, fatigue, and other symptoms, and the presence of depression or anxiety disorders were evaluated. The basal thyroid hormone levels of FM patients were in the normal range, while 41% of the patients had at least one thyroid antibody. Patients with thyroid autoimmunity showed a higher percentage of dry eyes, burning, or pain with urination, allodynia, blurred vision, and sore throat. Correlations found between thyroid autoimmunity and age or with the presence of depression or anxiety disorders were not significant. However, in the cohort of post-menopausal patients, the frequency of thyroid autoimmunity was higher with respect to pre-menopausal patients. In conclusion, autoimmune thyroiditis is present in an elevated percentage of FM patients, and it has been associated with the presence of typical symptoms of the disease.

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.

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.

Cell-free DNA in the plasma of patients with systemic sclerosis

The aim of the present study was to evaluate the concentration of cell-free DNA (cf-DNA) in the plasma of patients with systemic sclerosis (SSc) and to examine the correlation of cf-DNA with clinical variables of the disease. The study population consisted of 122 SSc patients and 16 healthy controls. Epidemiological and clinical data were collected by direct assessment. The β-globin gene was used to determine the total amount of DNA in the plasma by real-time quantitative PCR analysis. cf-DNA was found in all patients (mean concentration 1,420.7 copies/ml) and controls (mean concentration 1,462.5), with no significant difference. In SSc patients, no correlation was found between cf-DNA and the type of organ involvement, but patients with active disease presented significantly higher cf-DNA concentrations than those with inactive disease (p < 0.05). Our data suggest that cf-DNA could provide a useful biomarker for the assessment of disease activity in SSc patients.

Antipolymer antibody in Italian fibromyalgic patients

The objectives of the present study were to evaluate the presence of antipolymer antibody (APA) seropositivity in 285 Italian patients affected by primary fibromyalgia (FM) and to verify whether APA levels correlate with disease severity and with cytokine levels.APA levels were determined on serum samples by an indirect ELISA kit that detects IgG APA. Cytokines (IL-1, IL-6, IL-8, IL-10 and TNFα) were measured by ELISA in plasma. The impact of FM on the quality of life was estimated using the Fibromyalgia Impact Questionnaire, while pain severity was evaluated using a visual analogic scale. Patients were also characterized by the presence of tiredness, stiffness, nonrestorative sleep, anxiety, depression, tension headache, irritable bowel syndrome, temporomandibular dysfunction and Raynauds phenomena.Using a cut-off value of 30 U, APA-positive values were detected in 60 FM patients (21.05%) and in 15 healthy control individuals (15.00%) without significant differences among their levels or the percentage of seropositivity. FM patients with moderate and severe symptoms had slightly higher APA levels with respect to patients with mild symptoms. APA-seropositive patients exhibited significant correlations between APA levels and the Fibromyalgia Impact Questionnaire estimate (P = 0.042), tiredness (P = 0.003) and IL-1 levels (P = 0.0072).In conclusion, APA cannot be considered a marker of disease in Italian FM patients. The presence of APA, however, might permit the identification of a subset of FM patients with more severe symptoms and of patients who may respond differently to different therapeutic strategies.

Comparison of real-time PCR and nested PCR for the detection of Y chromosome sequences in the peripheral blood mononuclear cells of patients with systemic sclerosis

Real-time PCR (RT-PCR) is a sensitive technique that has been recently applied to the determination and quantification of fetomaternal microchimerism.1–3 In a previous manuscript, we detected the presence of Y chromosome sequences in female patients with systemic lupus erythaematosus (SLE) using nested PCR.4 In the present study we evaluated the presence of Y chromosome sequences in the peripheral blood mononuclear cells (PBMCs) of female patients with systemic sclerosis (SSc) using nested and RT-PCR, correlating the results with the clinical and serological manifestations of the patients. Genomic DNA was isolated from …

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