Angela Del Rosso

Activin, a Grape Seed-derived Proanthocyanidin Extract, Reduces Plasma Levels of Oxidative Stress and Adhesion Molecules (ICAM-1, VCAM-1 and E-selectin) in Systemic Sclerosis

This study evaluated whether a new generation antioxidant Activin derived from the grape seed proanthocyanidins, could reduce the induction of the adhesion molecules as a result of inflammatory response in the plasma of systemic sclerosis (SSc) patients. SSc patients were divided into two groups: one group was treated with Activin, a grape seed‐derived proanthocyanidins, while the other group served as control. Patients were given Activin 100 mg/day orally for one month after which the blood samples were withdrawn from both groups of the patients. Blood was also taken from normal human volunteers. Plasma was obtained in fasting state between 8 to 9 A.M. from two groups of SSc patients and controls. Soluble adhesion molecules including ICAM‐1, VCAM‐1, E‐selectin and P‐selectin as well as malonaldehyde, a marker for oxidative stress, were measured. The results of our study demonstrated up‐regulation of these soluble adhesion molecules except for P‐selectin, in the plasma of the SSc patients compared to those obtained from human volunteers. Activin significantly attenuated the increased expression of these adhesion molecules. In addition, there was a significant increase in the amount of malondialdehyde formation in the plasma of the SSc patients, which was also attenuated by Activin. The results of this study demonstrated that Activin could reduce the inflammatory response and the oxidative stress developed in SSc patients.

Flow‐Mediated Vasodilation and Carotid Intima‐Media Thickness in Systemic Sclerosis

Abstract:  Increased evidence suggests an accelerated macrovascular disease in systemic sclerosis (SSc). Brachial artery flow‐mediated vasodilation (FMD) and carotid intima‐media thickness (IMT) are two indicators of subclinic cardiovascular disease and are frequently used as surrogate measures of subclinic atherosclerosis. The aim of this study was to evaluate macrovascular involvement in SSc. We studied 35 SSc patients (6 males and 29 females; 11 with diffuse and 24 with limited disease) and 20 healthy controls. Brachial artery FMD was assessed by method described by Celermajer in all patients and 13 control subjects. IMT was measured using high‐resolution B‐mode ultrasonography in patients and controls. Traditional risk factors for atherosclerosis (hypertension, dyslipidemia, and smoke) were also assessed. FMD was significantly impaired (3.41%± 4.56% versus 7.66%± 4.24%; P < 0.037) and IMT was significantly elevated compared with healthy controls (0.93 ± 0.29 mm versus 0.77 ± 0.13 mm; P < 0.005). FMD was not significantly different in SSc with increased IMT compared with those with normal IMT). No correlation was found between risk factors for atherosclerosis and the impairment of FMD or IMT in SSc patients. The impairment of endothelial function and structural changes of large vessels are evident in SSc, but do not seem associated with traditional risk factors for atherosclerosis. Prospective studies including also clinical outcomes are needed to assess the features and significance of macrovacular involvement in SSc.

Effect of N-acetyl-l-cysteine on peroxynitrite and superoxide anion production of lung alveolar macrophages in systemic sclerosis

Lung macrophages may play a relevant role in oxidative processes producing both superoxide anion (O(2)(-)) and NO. In this view, an antioxidant therapy can be useful in the treatment of systemic sclerosis (SSc) patients. N-Acetylcysteine (NAC) is able to expand natural antioxidant defenses by increasing intracellular gluthatione concentration and it has been proposed as an antioxidant therapy in respiratory distress syndromes. The aim of our study was to determine whether lung macrophages obtained from SSc patient bronchoalveolar lavage (BAL) express the inducible form of nitric oxide synthase (iNOS) and whether NAC can reduce the peroxynitrite (ONOO(-)) and O(2)(-) production of these cells. Alveolar macrophages were isolated from BAL of 32 patients and used for the immunocytochemical determination of iNOS, and the production of ONOO(-) and O(2)(-) was measured by fluorimetric or spectrophotometric methods, respectively. Lung macrophages obtained from SSc patients expressed a higher level of iNOS compared to healthy subject cells. NAC preincubation (5 x 10(-5)M, 24h) significantly reduced (-21%) the ONOO(-) production in formyl Met-Leu-Phe (fMLP)-activated cells and slightly reduced it under resting conditions, whereas NAC preincubation was unable to modify the release of O(2)(-) both in basal condition and in fMLP-stimulated cells. We conclude that since SSc lung macrophages express high levels of iNOS and produce a significant quantity of ONOO(-), NAC administration reduces ONOO(-) production and can be an useful treatment to alleviate SSc symptoms.

Cell Invasion Is Affected by Differential Expression of the Urokinase Plasminogen Activator/Urokinase Plasminogen Activator Receptor System in Muscle Satellite Cells from Normal and Dystrophic Patients

The aim of this study was to evaluate the differential expression and the function in cell movement and proliferation of the urokinase plasminogen activator (u-PA) system in muscle satellite cells (MSC) of normal individuals and patients with Duchenne muscular dystrophy (DMD). By immunoenzymatic, zymographic, and radioligand binding methods and by quantitative polymerase chain reaction of the specific mRNA we have shown that both normal and DMD MSC produce u-PA and the plasminogen activator inhibitor-1 and express u-PA receptors (u-PAR). During the proliferation phase of their growth-differentiation program, MSC from DMD patients show more u-PAR than their normal counterpart, produce more plasminogen activator inhibitor-1, and release low amounts of u-PA into the culture medium. By Boyden chamber Matrigel invasion assays we have shown that normal MSC are more prone than DMD cells to spontaneous invasion but, when subjected to a chemotactic gradient of u-PA, DMD MSC sense the ligand much better and to a greater extent than normal MSC. u-PA also stimulates proliferation of MSC, but no difference is observable between normal and DMD patients. Antagonization of u-PA/u-PAR interaction with specific anti–u-PA and anti–u-PAR monoclonal antibodies and with antisense oligonucleotides inhibiting u-PAR expression indicates that u-PA/u-PAR interaction is required in spontaneous and u-PA–induced invasion, as well as in u-PA–induced proliferation.

Antisense oligodeoxynucleotides for urokinase-plasminogen activator receptor have anti-invasive and anti-proliferative effects in vitro and inhibit spontaneous metastases of human melanoma in mice

We have targeted the urokinase‐type plasminogen activator receptor (uPAR) with phosphorothioate antisense oligonucleotides (aODN) in vitro to evaluate the anti‐invasive and anti‐proliferative effects of uPAR down‐regulation, as well as in vivo to evaluate anti‐tumor and anti‐metastatic activity. aODN‐dependent uPAR downregulation in vitro was induced in cells of human melanoma, mammary carcinoma, ovarian carcinoma and SV‐40‐transformed embryonic lung fibroblasts. uPAR was determined by an antibody‐based assay and by semiquantitative polymerase chain reaction. Cell invasion was evaluated by Matrigel invasion assay and cell proliferation by direct cell counting. aODN reduced uPAR, invasion and proliferation in all the treated cell lines. Following aODN treatment, human melanoma cells exhibited a strong decrease of uPAR‐dependent ERK1/2 activation and were used in vivo to control metastasis in CD‐1 male nude (nu/nu) mice by uPAR aODN injection. 60 mice were injected in the hind leg muscles with a suspension of 106 melanoma cells. After 4 days, when a tumor mass of about 350 mg was evident in all the mice injected, 20 mice were treated i.v. with aODN and 20 with dODN at 0.5 mg/day for 5 consecutive days. Twenty control mice were not treated. A second and third cycle of treatment was administered at 2‐day intervals. Treatment with aODN resulted into a 78% reduction of lung metastases and 45% reduction of the primary tumor mass with no loss of body weight. Our results suggest to evaluate the utility of uPAR aODN in controlling the metastatic spreading of human melanoma.

The pathogenesis of inflammatory muscle diseases:: On the cutting edge among the environment, the genetic background, the immune response and the dysregulation of apoptosis

Inflammatory muscle diseases (IMD), including dermatomyositis (DM) and polymyositis (PM), affect skeletal muscle, leading to profound tissue modification. The etiology of IMD is unknown, but multiple steps of the disease pathogenesis have been identified. The main alterations involve the immune response. Cellular infiltrates found in the muscle provide strong evidence for the involvement of a preferential immune mechanism of muscle damage. The pathologic differences found between PM and DM indicate a different role played by cell-mediated and humoral immune alterations. It is well accepted that in the pathogenetic pathway both host genes and environmental factors are involved. Apoptosis, or programmed cell death, is a complex process that plays a key role in many physiological events. It regulates the turnover of immune cells and is one of the mechanisms involved in ensuring a competent, non-autoreactive repertoire of lymphocytes. Apoptosis as a mechanism of muscle fibre death has been described in several neuromuscular disorders and muscular dystrophies, and evidence of a lack of apoptosis in IMD suggests a failure of apoptotic clearance of inflammatory cells playing a role in the maintenance of chronic cytotoxic muscle fibre damage. Most likely, the failure of apoptosis seems to be the main hallmark of the pathogenesis of IMD.

Disease activity and antinucleosome antibodies in systemic lupus erythematosus.

Objective: To evaluate the correlation between antinucleosome antibodies and disease activity in patients with systemic lupus erythematosus (SLE). Methods: We evaluated antinucleosome antibodies (by ELISA) in 48 SLE patients. They were divided in 2 groups: positive (Group A, nr=18) and negative (Group B, nr=30). The groups were evaluated for antinucleosome antibodies and for clinical, humoral parameters (hemoglobin, blood cell count, urinanalysis, ESR, ANA, anti‐dsDNA, anticardiolipin antibodies, LAC), and ECLAM. Results: C3,C4, and hemoglobin were lower in Group A than (vs) group B (C3: 0.61±0.16 g/L vs 0.88±0.08 g/L, p<0.001; C4: 0.086±0.03 g/L vs 0.18±0.07 g/L, p<0.05; hemoglobin: 8.7±5.8 g/dL vs 12.7±1.44 g/dL; p<0.02). ECLAM was higher in group A 7.56±2.19 vs group B 4.67±1.35 (p<0.001). Urinary sediment was more altered in group A (88.8%) vs group B (33.3%; p<0.001). Conclusion: We found a correlation between antinucleosome antibodies and SLE disease activity as expressed by the higher ECLAM score in group A.

The autonomic nervous system in systemic sclerosis. A review

The autonomic nervous system is an underestimated target of systemic sclerosis alterations. In this review we analyzed the major manifestations of its involvement, reconsidering the main theories of its pathogenesis.

Exercise Doppler Echocardiography Identifies Preclinic Asymptomatic Pulmonary Hypertension in Systemic Sclerosis

Abstract:  In systemic sclerosis (SSc), the involvement of the interstitium or vascular system of the lung may lead to pulmonary arterial hypertension (PAH). PAH is often asymptomatic or oligosymptomatic in early SSc and, when it becomes symptomatic, pulmonary vascular system is already damaged. Exercise echocardiography (ex‐echo), measuring pulmonary artery pressure (PAP) during exercise and allowing to differentiate physiologic from altered PAP responses, may identify subclinical PAH. Our aims were (a) to evaluate by ex‐echo the change of PAP in patients with SSc without lung involvement; and (b) to correlate PAP during exercise (ex‐PAP) values to clinical and biohumoral parameters of PAH. Twenty‐seven patients with limited SSc (lSSc) without interstitial lung involvement were studied. Patients underwent rest and exercise two‐dimensional and Doppler echocardiography by supine cycloergometer. Systolic PAP was calculated using the maximum systolic velocity of the tricuspid regurgitant jet at rest and during exercise values of systolic PAP exceeding 40 mmHg at ex‐echo were considered as abnormal, and biohumoral markers potentially related to PAH were assessed. Eighteen of 27 SSc patients presented an ex‐PAP >40 mmHg, while in 9 of 27 patients ex‐PAP values remained <40 mmHg (48.8 ± 4.5 mmHg versus 36.2 ± 3.1 mmHg; P < 0.001). Other echocardiographic and ergometric parameters, clinical tests, and biohumoral markers were not different in the two groups. Ex‐PAP significantly correlated with D‐dimer (P= 0.0125; r2= 0.2029). Ex‐echo identifies a cluster of SSc patients with subclinical PAH that may develop PAH. This group should be followed up and may be considered for specific therapies to prevent disease evolution.

Melatonin is a safe and effective treatment for chronic pulmonary and extrapulmonary sarcoidosis

Abstract:   Chronic sarcoidosis (CS) is often unresponsive to usual treatments. Melatonin, an immunoregulatory drug, was employed in CS patients in whom usual treatments were ineffective or induced severe side effects. Melatonin was given for 2 yr (20 mg/day in the first year, 10 mg/day in the second year) to 18 CS patients. Pulmonary function tests, chest X rays, pulmonary computed tomography, Ga67 scintigraphy and angiotensin‐converting enzyme (ACE) were assayed at baseline and in the follow‐up. Normalization of ACE, improvement of pulmonary parameters and resolution of skin involvement were found in the patients given melatonin. After 24 months of melatonin therapy, hylar adenopathy completely resolved in eight patients and parenchymal lesions were markedly improved in all patients; in the five patients with reduced diffusion capacity of the lung for carbon monoxide, the values normalized after 6 months of therapy and remained stable until month 24. After 24 months, Ga67 pulmonary and extra‐pulmonary uptake was totally normalized in seven patients and, at month 12 months, ACE was normalized in six patients in which the values were high at the baseline. Skin lesions, present in three patients, completely disappeared at month 24 months. No side effects were experienced and no disease relapse was observed during melatonin treatment. Melatonin may be an effective and safe therapy for CS when other treatments fail or cause side effects.