Paolo Bongioanni

A genome-wide association meta-analysis identifies a novel locus at 17q11.2 associated with sporadic amyotrophic lateral sclerosis

Identification of mutations at familial loci for amyotrophic lateral sclerosis (ALS) has provided novel insights into the aetiology of this rapidly progressing fatal neurodegenerative disease. However, genome-wide association studies (GWAS) of the more common (∼90%) sporadic form have been less successful with the exception of the replicated locus at 9p21.2. To identify new loci associated with disease susceptibility, we have established the largest association study in ALS to date and undertaken a GWAS meta-analytical study combining 3959 newly genotyped Italian individuals (1982 cases and 1977 controls) collected by SLAGEN (Italian Consortium for the Genetics of ALS) together with samples from Netherlands, USA, UK, Sweden, Belgium, France, Ireland and Italy collected by ALSGEN (the International Consortium on Amyotrophic Lateral Sclerosis Genetics). We analysed a total of 13 225 individuals, 6100 cases and 7125 controls for almost 7 million single-nucleotide polymorphisms (SNPs). We identified a novel locus with genome-wide significance at 17q11.2 (rs34517613 with P = 1.11 × 10(-8); OR 0.82) that was validated when combined with genotype data from a replication cohort (P = 8.62 × 10(-9); OR 0.833) of 4656 individuals. Furthermore, we confirmed the previously reported association at 9p21.2 (rs3849943 with P = 7.69 × 10(-9); OR 1.16). Finally, we estimated the contribution of common variation to heritability of sporadic ALS as ∼12% using a linear mixed model accounting for all SNPs. Our results provide an insight into the genetic structure of sporadic ALS, confirming that common variation contributes to risk and that sufficiently powered studies can identify novel susceptibility loci.

TNF and sTNFR1/2 plasma levels in ALS patients

The involvement of the immune system has been hypothesized in the pathogenesis of amyotrophic lateral sclerosis (ALS). In this study a significantly higher level of TNF-alpha and its soluble receptors, TNF-R1 and TNF-R2, has been found in plasma of patients affected by the sporadic form of ALS compared to normal subjects. The genetic analysis of the polymorphisms of TNF-alpha, TNF-R1 and TNF-R2 showed no statistically significant differences in alleles and genotype frequencies between patients and controls. These data suggest a participation of the immune system in response to as far unknown intracellular signals.

Assessment of Swallowing by Oropharyngoesophageal Scintigraphy in Patients with Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is the most common degenerative motor neuron disease in adults, and dysphagia is one of its most frequent and disabling symptoms. Oropharyngoesophageal scintigraphy (OPES) permits a functional and semiquantitative study of the various stages of swallowing. We studied 28 ALS patients (12 females and 16 males; mean age = 63.57 ± 10.39 yr SD), who were clinically rated against the ALSFRS scale (Amyotrophic Lateral Sclerosis Functioning Rating Scale) and underwent OPES with 99mTc-nanocolloid using either a liquid or a semisolid bolus. The semiquantitative parameters we analyzed were Oral Transit Time (OTT), Pharyngeal Transit Time (PTT), Esophageal Transit Time (ETT), Retention Index (RI), and Esophageal Emptying Rate (EER10s). Hence, the OPES performed with a semisolid bolus produced a higher proportion of pathologic values for the swallowing variables than when liquid bolus was used. Analyzed by grouping the patients into classes according to their bulbar ALSFRS scores, we found a significant increase in the OTT (p < 0.005), PTT (p < 0.02), and Oropharyngeal Retention Index (OPRI) (p < 0.0004) variables in ALS patients with more severe bulbar involvement. OPES has turned out to be a very important examination for detecting tracheal-bronchial inhalation and it also offers the possibility of acquiring a semiquantitative evaluation of the amount of food inhaled. In our experience, OPES in patients with ALS has been easy to use, economic, well tolerated, and capable of supplying precise indications with regard to the extent of the swallowing disorder, which permits a better clinical definition of the ALS patient.

Quality of life of patients with amyotrophic lateral sclerosis

The aim of this study is to evaluate the correlation between different measures of quality of life (QoL), functional status and mood status in patients with amyotrophic lateral sclerosis (ALS). A sample of 40 patients with ALS was recruited and scales for the evaluation of health-related QoL (SF36), patient-centered QoL (SEIQoL), functional status (ALSFRS) and depression (ZDS) have been administered to them. All the correlations (Pearsons r) between the scores have been considered and the t-test was performed in order to compare male with female patients. No correlation emerged between the different measures of QoL and functional status, apart from the scores of the Physical Functioning subscale of the SF36 which resulted positively correlated (p < 0.01) with those of the ALSFRS. Mood status resulted negatively correlated with many aspects of health-related QoL but not with patient-centred QoL. Data suggest that the relationship between functional status and different domains of QoL in this population of patients with a severely compromising disease is not linear and that the satisfaction with life of the patients themselves is not strongly related to the level of physical impairment.

Lymphocyte subsets in multiple sclerosis A study with two-colour fluorescence analysis

Multiple sclerosis (MS) is postulated to be an immunopathologically mediated disease. This concept is supported by the finding of abnormally distributed peripheral blood T-cell subsets and a decreased T-suppressor function. Thirty-seven MS patients have been selected according to the criteria for definite MS. Fluorescein- or phycoerythrin-conjugated monoclonal antibodies have been used to define different lymphocyte subsets: CD4+, CD5+, CD8+, CD19+, CD38+, CD45RA+, CD4+CD45RA+, CD19+CD5+, CD8+CD38+. In relapsing-remitting (RR)-MS patients a significantly decreased percentage of CD19+ cells and in progressive MS patients a significantly increased percentage of CD19+CD5+ cells have been found. During a relapse in RR-MS, a significantly decreased percentage of CD4+CD45RA+ cells and a significantly increased percentage of CD8+CD38+ cells have been observed. Moreover, in RR-MS patients a significantly increased percentage of CD38+ cells and significantly high IgM amounts have been found. The increased percentage of CD19+CD5+ and CD38+ cells (together with high IgM levels) and the reduced percentage of CD4+CD45RA+ lymphocytes could be related to an activation of both cellular and humoral immune response in acute MS.

Time course of oxidant markers and antioxidant defenses in subgroups of amyotrophic lateral sclerosis patients

Oxidative stress markers have been found in nervous and peripheral tissues of familial and sporadic amyotrophic lateral sclerosis patients. Here, we evaluated the activity of some antioxidant enzymes glutathione peroxidase, glutathione reductase and Cu-Zn superoxide dismutase in erythrocyte, the marker of non-enzymatic antioxidant response (total antioxidant status), as well as plasma reactive oxygen species, at the enrolment and during disease progression in 88 patients affected by the sporadic form of amyotrophic lateral sclerosis. Our study has been performed along 72 months by grouping the patients according to the ALS functional rating score or rate of disease progression. Our results showed a significant impairment of erythrocytes glutathione peroxidase activity in all groups of patients that remained low during disease time course. SOD1 activity significantly decreased along disease course in subjects with a more impaired functional status. A decreasing activity of all assayed enzymes was found in patients who have a faster disease progression rate. By this work we have the evidence that different ALS phenotypes present with different profile of enzymatic and non-enzymatic antioxidant response.

Distribution of VGF peptides in the human cortex and their selective changes in Parkinson's and Alzheimer's diseases

VGF mRNA and its precursor‐derived products are selectively expressed in certain neurons and promptly respond to neurotrophins and to neural/electrical activity. Proteomic studies have previously revealed a reduction in some VGF peptides in the cerebrospinal fluid of patients affected by Alzheimer’s disease and other conditions, suggesting their potential diagnostic and clinical significance. As the presence of VGF peptides within the human cortex has been somewhat elucidated, they were studied postmortem in the frontal, parietal, and temporal cortex areas of control subjects and patients affected by Parkinson’s disease, and in parietal cortex samples from patients with Alzheimer’s disease. We raised antibodies to the C‐/N‐terminal portions of the proVGF precursor protein, to the TPGH and TLQP sequences and to the neuroendocrine regulatory peptide (NERP)‐1, all used for enzyme‐linked immunosorbent assay coupled with gel chromatography and for immunohistochemistry. In the control brain samples, the levels of TPGH and C‐terminus peptides were about 130–200 and 700–2000 pmol g−1, respectively, the N‐terminus and NERP‐1 peptides were less represented (about 10–30 and 4–20 pmol g−1, respectively), and the TLQP peptides were below detection limits. Upon gel chromatography, the VGF antisera mainly revealed small molecular weight forms (i.e. about 0.8–1.3 kDa), whereas VGF immunolocalisation was found within different types of neuron in rat and bovine brain cortices. In the Parkinson’s disease samples, a clear‐cut decrease was revealed in the parietal cortex only, exclusively for TPGH and NERP‐1 peptides, whereas in the Alzheimer’s disease samples, a reduction in all of the VGF peptides was shown. The results suggest the involvement of VGF in the physiological or pathophysiological mechanisms occurring in the parietal cortex of patients with Parkinson’s and Alzheimer’s diseases.

Growth hormone secretion is impaired in amyotrophic lateral sclerosis

Objective   ALS is the most common motor neurone disorder in human adults. Scanty data on endocrine abnormalities have been reported. The aim of the present study was to investigate the GH‐IGF‐I axis in ALS patients.

Increased T-lymphocyte interleukin-6 binding in patients with multiple sclerosis.

Multiple sclerosis (MS) represents a T‐cell‐mediated autoimmune demyelinating disease of the central nervous system associated with altered immunoregulation. Interleukin (IL)‐6 is a cytokine that has several effects on the neuroimmune system. Specific IL‐6 receptors have been found in human lymphocytes and neuroglial cells. The aim of the present study was to assay IL‐6 binding on peripheral blood T lymphocytes in MS patients. We found that T cells from MS patients had significantly more IL‐6 receptors [Bmax: 279 ± 7 vs. 246 ± 8 (mean ± SEM) receptors/cell, in patients and controls, respectively], whereas Kd values were similar to those of healthy subjects [26.8 ± 0.7 vs. 25.4 ± 0.6 (mean ± SEM) pM, in patients and controls, respectively]. Significant (P < 0.05) differences in IL‐6 binding values were observed between stable patients and those relapsing (272 ± 9 vs. 300 ± 12 (mean ± SEM) receptors/cell, respectively). We found significantly (P < 0.001) higher amounts of IL‐6 receptors on CD4+ T cells from MS patients than on CD4+ lymphocytes from controls (434 ± 11 vs. 363 ± 9 (mean ± SEM) receptors/cell, respectively); CD8+ T cells showed very few IL‐6 receptors in both patients and controls. These data are discussed in terms of MS immune pathogenesis and pathophysiology, because T‐cell activation seems to be linked to increased IL‐6 binding. The upregulated IL‐6 system might be involved in antibody‐mediated demyelinating pathways, because IL‐6 is well known to enhance humoral immune response.

T-cell tumour necrosis factor-α receptor binding in demented patients

Abstract Dementia of Alzheimer type (DAT) is a neurodegenerative disease of the central nervous system (CNS), in which an unbalanced cytokine network may lead to an altered immunoregulation. Tumour necrosis factor (TNF)-α is a cytokine with manifold effects on the neuroimmune system. Specific TNF-α receptors have been found on human peripheral blood lymphocytes. The aim of the present study has been to assay TNF-α binding on T cells from DAT patients and healthy sex- and age-matched controls. We found that T lymphocytes from demented patients bear significantly more p60 and p80 TNF-α receptors than those from controls (Bmax: 705, 29 vs 131, 6 (mean, SEM) receptors/cell). Such TNF-α binding sites, of the same type in DAT patients and healthy subjects (Kd: 67.6, 5.0 vs 70.7, 5.6 (mean, SEM) pM), are functional, since they are able to mediate in vitro NF-κB activation. These results are discussed in terms of DAT pathogenesis. Since it has been reported that activated T cells have more TNF-α receptors than resting cells, an increased number of lymphocyte TNF-α receptors might indicate a systemic immune activation in DAT patients as compared with healthy controls.