Neuroinflammation, immune system and Alzheimer disease: searching for the missing link.

Due to an increasingly aging population, Alzheimer disease (AD) represents a crucial issue for the healthcare system because of its widespread prevalence and the burden of its care needs. Several hypotheses on AD pathogenesis have been proposed and current therapeutical strategies have shown limited effectiveness. In the last decade, more evidence has supported a role for neuroinflammation and immune system dysregulation in AD. It remains unclear whether astrocytes, microglia and immune cells influence disease onset, progression or both. Amyloid-ß peptides that aggregate extracellularly in the typical neuritic plaques generate a constant inflammatory environment. This causes a prolonged activation of microglial and astroglial cells that potentiate neuronal damage and provoke the alteration of the blood brain barrier (BBB), damaging the permeability of blood vessels. Recent data support the role of the BBB as a link between neuroinflammation, the immune system and AD. Hence, a thorough investigation of the neuroinflammatory and immune system pathways that impact neurodegeneration and novel exciting findings such as microglia-derived microvesicles, inflammasomes and signalosomes will ultimately enhance our understanding of the pathological process. Eventually, we should proceed with caution in defining a causal or consequential role of neuroinflammation in AD, but rather focus on identifying its exact pathological contribution.

Genetic evaluation of AMPD1, CPT2, and PGYM metabolic enzymes in patients with chronic fatigue syndrome.

Chronic fatigue syndrome (CFS) is a disease that can seriously impair one's quality of life; patients complain of excessive fatigue and myalgia following physical exertion. This disease may be associated with abnormalities in genes affecting exercise tolerance and physical performance. Adenosine monophosphate deaminase (AMPD1), carnitine palmitoyltransferase II (CPT2), and the muscle isoform of glycogen phosphorylase (PYGM) genes provide instructions for producing enzymes that play major roles in energy production during work. The aim of this study was to look for evidence of genotype-associated excessive muscle fatigue. Three metabolic genes (AMPD1, CPT2, and PYGM) were therefore fully sequenced in 17 Italian patients with CFS. We examined polymorphisms known to alter the function of these metabolic genes, and compared their genotypic distributions in CFS patients and 50 healthy controls using chi-square tests and odds ratios. One-way analysis of variance with F-ratio was carried out to determine the associations between genotypes and disease severity using CF scores. No major genetic variations between patients and controls were found in the three genes studied, and we did not find any association between these genes and CFS. In conclusion, variations in AMPD1, CPT2, and PGYM genes are not associated with the onset, susceptibility, or severity of CFS.

Affordable, automatic quantitative fall risk assessment based on clinical balance scales and Kinect data.

The problem of a correct fall risk assessment is becoming more and more critical with the ageing of the population. In spite of the available approaches allowing a quantitative analysis of the human movement control system's performance, the clinical assessment and diagnostic approach to fall risk assessment still relies mostly on non-quantitative exams, such as clinical scales. This work documents our current effort to develop a novel method to assess balance control abilities through a system implementing an automatic evaluation of exercises drawn from balance assessment scales. Our aim is to overcome the classical limits characterizing these scales i.e. limited granularity and inter-/intra-examiner reliability, to obtain objective scores and more detailed information allowing to predict fall risk. We used Microsoft Kinect to record subjects' movements while performing challenging exercises drawn from clinical balance scales. We then computed a set of parameters quantifying the execution of the exercises and fed them to a supervised classifier to perform a classification based on the clinical score. We obtained a good accuracy (~82%) and especially a high sensitivity (~83%).

Mitochondrial alterations, oxidative stress and neuroinflammation in Alzheimer's disease.

Alzheimer's disease (AD) is a multifactorial disorder characterized by the progressive deterioration of neuronal networks. The primary cause and sequence of its progression are only partially understood but abnormalities in folding and accumulation of insoluble proteins such as beta-amyloid and Tau-protein are both associated with the pathogenesis of AD. Mitochondria play a crucial role in cell survival and death, and changes in mitochondrial structure and/or function are related to many human diseases. Increasing evidence suggests that compromised mitochondrial function contributes to the aging process and thus may increase the risk of AD. Dysfunctional mitochondria contribute to reactive oxygen species which can lead to extensive macromolecule oxidative damage and the progression of amyloid pathology. Oxidative stress and amyloid toxicity leave neurons chemically vulnerable. The mitochondrial toxicity induced by beta-amyloid is still not clear but may include numerous mechanisms, such as the increased permeability of mitochondrial membranes, the disruption of calcium homeostasis, the alteration of oxidative phosphorylation with a consequent overproduction of reactive oxygen species. Other mechanisms have been associated with the pathophysiology of AD. Inflammatory changes are observed in AD brain overall, particularly at the amyloid deposits, which are rich in activated microglia. Once stimulated, the microglia release a wide variety of pro-inflammatory mediators including cytokines, complement components and free radicals, all of which potentially contribute to further neuronal dysfunction and eventually death. Clinically, novel approaches to visualize early neuroinflammation in the human brain are needed to improve the monitoring and control of therapeutic strategies that target inflammatory and other pathological mechanisms. Similarly, there is growing interest in developing agents that modulate mitochondrial function.

Alzheimer's disease, autoimmunity and inflammation. The good, the bad and the ugly.

Alzheimer's disease (AD) has been recognized as the most common cause of sporadic dementia. It represents both a medical and social problem, as it affects 10% of over-65 population. Even if the elderly are the most involved population, aging alone cannot be considered as the only cause of this disease. In this review we wanted to focus on the last hypotheses on the possible causes of this neuronal affection. We focused in particular on the role of inflammation and alteration of the inflammatory status that is typical of the elderly and may lead to chronic inflammation. The inflammation seems to be a cause of neuronal impairment and loss. Some studies have proposed a protective role of antiinflammatory drugs. Then we analyzed the role of genetic polymorphisms of some pro-inflammatory substances that seem to be linked to some cases of dementia. The complement system seems to have a role too, as some factors have been found in senile plaques, representing a possible involvement of classical complement pathway. One of the latest hypotheses is about the role of blood-brain barrier (BBB), as its loss of integrity may lead to a passage of proteins in cerebro spinal fluid (CSF), causing a compromised role of BBB in preserving the brain as an "immune sanctuary".

HIV-related acute inflammatory leukoencephalopathy of undetermined origin: review of the literature.

HIV-related acute inflammatory leukoencephalopathy of undetermined origin (AIL) has been anecdotally described in literature as being responsible for cognitive and motor deficits. We carried out a review of all the cases of AIL published in literature. Articles were selected according to 2 criteria: acute onset of symptoms; undetermined aetiology and non-fulfilment of multiple sclerosis diagnostic criteria. They were then analyzed in terms of clinical, biological and instrumental features, therapy, diagnostic classification and prognosis. Although rare (21 patients out of about 4,000 publications), AIL is of particular interest, as the comprehension of its mechanisms could give some insight into the direct and immune-mediated actions of HIV within the brain. All the reported patients share several clinical, histopathological, radiological and CSF features, leading to hypothesize a similar aetiopathogenetic mechanism. Conversely, we observed a high heterogeneity of treatment and diagnostic classification, which could have conditioned the broad prognostic variability. The absence of a defined aetiology leads to consider these forms as a particular subgroup of not determined leucoencephalopathies (NDLE), with both MRI and histological pattern dominated by inflammation as distinctive feature.

Chronic fatigue syndrome/myalgic encephalomyelitis: an update.

Chronic Fatigue Syndrome (CFS), also referred to as Myalgic Encephalomyelitis (ME), is a disease of unknown origin. It is classified as Post Viral Fatigue Syndrome (PVFS) in the WHO International Classification of Diseases (ICD) and listed as sub-category at G93.3 under chapter G93, other disorders of the brain. ME/CFS is primarily an endemic disorder but occurs in both epidemic and sporadic forms. It affects all racial-ethnic groups and is seen in all socioeconomic strata. A diagnosis of CFS is a diagnosis of exclusion, meaning other medical conditions, including psychiatric disorders, must be first ruled out. CFS is diagnosed if there is no other explanation for the fatigue and if the other symptoms did not develop before the fatigue. The estimated worldwide prevalence of CFS is 0.4?1 percent. The disease predominantly affects young adults, with a peak age of onset of between 20 and 40 years, and women, with a female to male ratio of 6:1. Mean illness duration ranges from 3 to 9 years. The patho-physiological mechanism of CFS is unclear but the immunological pattern of CFS patients gleaned from various studies indicates that the immune system is chronically activated. Besides the role of environmental insults (xenobiotics, infectious agents, stress) the genetic features of patients are studied to evaluate their role in triggering the pathology. At present there are no specific pharmacological therapies to treat the disease but a variety of therapeutic approaches have been described as benefiting patients. Treatment programs are directed at relief of symptoms, with the goal of the patient regaining some level of preexisting function and well-being.

Antioxidant vitamins reduce oxidative stress and ventricular remodeling in patients with acute myocardial infarction.

We analyzed soluble vascular adhesion molecules (sVCAM-1), reactive oxygen metabolites (ROMs) level, total antioxidant status (TAS) and telediastolic left ventricular volume (TLVV) in patients with myocardial infarction undergoing reperfusion therapy and treated with antioxidant vitamins (AT) or placebo (P) before and for 1 month after reperfusion. After reperfusion, sVCAM-1 serum concentration, reactive oxygen metabolites level, and TLVV were significantly higher in patients treated with placebo than in those treated with antioxidant vitamins, while TAS was significantly higher in patients treated with antioxidant supplementation. We observed that 48 hours after reperfusion sVCAM-1 (P) vs sVCAM-1 (AT) was 2.03+/-0.5 vs 1.63+/-0.7 microg/ml with p < 0.01; ROMs (P) vs ROMs (AT) were 335.60+/-35.80 vs 307.50+/-47.10 U.CARR with p < 0.05; TAS (P) vs TAS (AT) was 526.47+/-44.24 vs 737.65+/-51.15 micromol/l with p < 0.01; 1 week after reperfusion TLVV (P) vs TLVV (AT) was 125.12+/-29.80 vs 119.40+/-29.40 ml with p < 0.05; 1 month after reperfusion TLVV (P) vs TLVV (AV) was 132.00+/-33.50 vs 123.40+/-21.60 ml with p < 0.05. In the first period after infarction, vitamin treatment improves the antioxidant system and reduces oxidative stress, inflammatory process and left ventricular remodeling.

Transient transfection of porcine granulosa cells after 3D culture in barium alginate capsules.

Three-dimensional culture systems in barium alginate capsules can be employed to maintain primary granulosa cells in an undifferentiated state for almost 6 days. This is due to a self-organization of cells in a pseudofollicular structure. The transfection of primary granulosa cells is a necessary condition when employing these culture systems for several purposes, for example as an in vitro toxicity test or the development of oocytes or zygotes. In this work, the feasibility of two transient transfection techniques (liposome-mediated and electroporation) was assessed in primary porcine granulosa cells after a 6-day culture in an artificial extracellular matrix (barium alginate membrane). Human recombinant green fluorescent protein was chosen as a molecular readout, and protein expression was assessed after 48 hours from transfection. Liposome-mediated transfection gave low transfection levels, with increasing yields from 2 to 12 microgDNA/ml of medium; the maximum percentage (85.7%) was reached at 12 microgDNA/ml of medium. Electroporation-mediated transfection yields were higher: the best results (81.7% of transfected cells) were achieved with two 50V pulses and 12 microg/ml DNA. The application of a single or double pulse (50V) at 4 mgDNA/ml gave negligible results. These results indicate that primary granulosa cell cultured in barium alginate capsules can be transfected by electroporation with high transfection yields.

HLA and hypocomplementemia: the disadvantage of carrying the HLA-B35 and the silent alleles of the C4 complement component.

Hypocomplementemia is an extremely complex phenomenon: we devoted our attention to its immunogenetic basis, particularly to the HLA haplotypes involved and to the study of C4 polymorphic genes. With this in mind we analyzed a group of unrelated patients with hypocomplementemia and 15 families suffering from specific C4 deficiency. Firstly, we performed a population analysis in order to identify a statistically significant association: HLA-B35 and C4BQ0 alleles, in the total group of hypocomplementemic individuals, seem to be associated with the primary disease. Secondly, we defined HLA haplotypes clear-cut segregation in the hypocomplementemic families and we identified the most common HLA haplotypes carrying B35 and C4 null allele associated with this condition. With the aid of correspondence analysis and the Transmission Disequilibrium Test (TDT), we measured the strength of this association. In this work, mainly through family analysis, we envisaged a potentially interesting genomic trait, within HLA, close to B locus, that seems to be involved in hypocomplementemia itself and perhaps in hypocomplementemia-related disorders.

Immunological features of neurological paraneoplastic syndromes.

Neurological paraneoplastic syndromes are a rare group of disorders that occur in 1-2% of people with malignancy. They are usually caused by an immune response, triggered by and directed against a tumour, that cross-reacts with protein expressed by the peripheral or central nervous system. Any part of the nervous system can be affected and patients often develop severe and permanent disability. Diagnosis can be difficult as in two-thirds of patients the neurological problems appear up to 5 years before the tumour manifests. However, certain of these syndromes are often associated with specific serum autoantibodies that can be useful both in diagnosis of the neurological syndrome and in focusing the search for a particular tumour. Thus, these antibodies can allow earlier identification and treatment of cancer and, potentially, a reduction in morbidity and mortality. It was only in the 1980s that the first anti-neuronal autoantibodies were characterized and their associations with clinical syndromes and tumours defined. Further antibodies have been isolated over the past 20 years and novel pathogenic mechanisms for several syndromes have been recognized. For example, voltage-gate ion channels seem to be a common target for autoantibodies involved in peripheral nerve diseases such as the Lambert-Eaton myasthenic syndrome and neuromyotonia (Isaacs' syndrome). However, the place of most paraneoplastic antibodies in the pathogenesis of central syndromes is yet to be fully elucidated.

CD40 ligand-dependent maturation of human monocyte-derived dendritic cells by activated platelets.

Atherosclerosis is defined as an inflammatory immunological disease that is triggered by platelet activation, endothelial injury and consequent innate and adaptive immune processes. Dendritic cells are critical for the cell-mediated arm of the immune response as they activate naïve T cells after maturation. Platelets play a crucial role in thrombus formation in the injured vessel walls. We investigated the role of resting and thrombin-activated platelets in dendritic cell maturation in vitro using platelets and monocyte-derived dendritic cells from healthy donors. Resting platelet supernatants did not affect maturation, whereas supernatants from thrombin-activated platelets induced dendritic cell maturation as demonstrated by FACS analysis of HLA-DR expression. This effect was inhibited by anti CD40 ligand antibody, but not by aspirin pretreatment of platelets. Supernatants of platelet-dendritic cell co-cultures induced augmented monocyte migration when platelets were activated by thrombin, again reversible by blocking CD40 ligand. These data show that activated platelets trigger dendritic cell maturation independent of cyclooxygenase-derived arachidonic acid metabolites by mechanisms involving CD40 ligand, which is also involved in monocyte chemotactic mediator release from platelets and dendritic cells. The results of this study suggest a role of CD40 ligand from activated platelets in connecting innate and adaptive immunity.

Inhibition of leukocyte and platelet aggregation in vitro by antithrombin.

Glycosaminoglycan-mediated aggregation of cells occurs through adhesion mechanisms in which heparan sulfate chains bind to counter receptors on these cells. As antithrombin interacts with heparan sulfate proteoglycans through its heparin-binding domain and inhibits leukocyte adhesion in ischaemia/reperfusion, it may affect leukocyte aggregation. Leukocyte aggregation was therefore monitored in vitro as the increase in transmission of light through stirred suspensions in a platelet aggregometer. Aggregation curves were quantified as the area under the curve in the first 6 min following stimulation. Leukocytes in platelet-rich plasma were obtained from heparinized whole blood of healthy donors by centrifugation; the ratio of leukocytes to platelets was about 1:45, and the final concentration of autologous plasma was 80%. Neutrophils were purified by dextran sedimentation, density centrifugation, and hypotonic lysis of erythrocytes. Aggregation was induced by phytohaemaglutinin (0.24 mg/mL) or formyl-Met-Leu-Phe (0.2 x 10(-6) M), with or without various concentrations of antithrombin. During the observation period (6 min) no aggregation of leukocytes in platelet-rich plasma or isolated neutrophils could be induced either with medium or with antithrombin (0.2 x 10(0) to 0.2 x 10(-6) IU/mL). Addition of phytohaemagglutinin or formyl-Met-Leu-Phe stimulated aggregation of leukocytes in platelet-rich plasma and neutrophils to different extents. Additional presence of anti-thrombin significantly inhibited phytohaemagglutinin-induced aggregation of leukocytes in platelet-rich plasma, whereas formyl-Met-Leu-Phe-induced aggregation was not affected by antithrombin. Data show that in the presence of plasma and platelets, aggregation of normal white blood cells after stimulation with lectin but not with chemotaxin is inhibited by antithrombin, suggesting specific interactions of antithrombin with lectin-activated processes of neutrophil aggregation that occur in the presence of platelets and/or plasma.

The inhibition of oxygen radical release from human neutrophils by resting platelets is reversed by administration of acetylsalicylic acid or clopidogrel.

Resting platelets inhibit oxygen radical release from neutrophils. Antiplatelet therapy may support this function by preventing platelet activation. Whether antiplatelet agents affect the antioxidative action of resting platelets in the absence of platelet activation is unknown. The effect of acetylsalicylic acid or clopidogrel administration on the antioxidative action of resting platelets was therefore studied in ten healthy volunteers. Preparations of resting platelets were obtained from 5 subjects each - before, during and after an eight-day course of daily treatment with 100 mg of acetylsalicylic acid or 75 mg of the thienopyridine clopidogrel. Human peripheral blood neutrophils were pretreated with the platelets at a ratio of (1/5)0 for 45 min; then formyl-Met-Leu-Phe-triggered oxygen radical release was measured fluorometrically. The inhibitory effect of platelets on oxygen radical release from neutrophils which was seen before treatment was abolished by antiplatelet therapy with either of the drugs, and inhibition was restored gradually after discontinuing acetlsalicylic acid/ clopidogrel intake. Results suggest that the protective role of resting platelets in controlling oxygen radical release from neutrophils in the absence of platelet activation may be impaired by antiplatelet therapy.

Plasma levels of procalcitonin and interleukin-6 in acute myocardial infarction.

Estimation of cardiac morbidity in patients after major surgery is a difficult problem. In addition, infectious complications seriously decrease potential beneficial outcome after cardiovascular surgery. The present study assessed the use of a newer marker of the inflammatory response, procalcitonin, in the field of myocardial infarction, in conjunction with measurements of interleukin-6. Forty-four consecutive cases with acute myocardial infarction were included in the study 4+/-1.3 h after the onset of symptoms. Plasma levels of procalcitonin and interleukin-6 were obtained at admission, and after 3, 6, 12, 18, 24 and 48 h, using commercially available test kits. The range of levels of interleukin-6 and procalcitonin was about normal at admission. Interleukin-6 levels increased significantly following myocardial infarction, whereas procalcitonin were essentially unchanged, i.e. remained close to the normal level threshold of 0.5 ng/ml; only minor variability occurred with a mean peak level of procalcitonin of 1+/-0.4 ng/ml. Data demonstrate that, in contrast to the acute phase reactant interleukin-6, plasma levels procalcitonin are not significantly elevated during uncomplicated acute myocardial infarction. This observation may support the role of procalcitonin measurements in the differential diagnosis of infectious and cardiovascular complications after major surgery.

Inhibition of human peripheral blood neutrophil respiratory burst by alcohol-based venipuncture site disinfection.

Ethanol inhibits the respiratory burst of neutrophils. Therefore, the effects of alcohol-based skin disinfection on oxygen metabolism in neutrophils were tested using 70% ethanol or an ethanol-isopropanol-n-propanol mixture. Neutrophil respiratory burst activity as assessed fluorometrically by oxidation of 2', 7'-dichlorofluorescein diacetate increased at 10 min after disinfection with 70% ethanol compared to the activity at 30 s. The increase was significant for triggering oxidative burst with formyl-peptide but not with phorbol myristate acetate.

Neutrophil infiltration into human gliomas.

Human gliomas were analysed for the infiltration of neutrophils using immunohistochemistry by staining sections for CD15-positive and myeloperoxidase-positive cells. Over 70% of all glioma samples analysed (n = 105) had significant neutrophil infiltration, but there was a marked and significant correlation between tumour grade and the extent of the neutrophil infiltration. In the low grade tumours only 40-50% had significant infiltration, while in glioblastoma multiforme over 85% of the samples analysed had significant infiltration. Numbers of neutrophils infiltrating glioblastoma multiforme tumours were also greater than in the other tumour groups. Circulating white blood cell counts were elevated above the normal range in all glioma patients, but this elevation was entirely due to increased numbers of circulating neutrophils. Again, the highest numbers of circulating neutrophils were seen in the glioblastoma multiforme patients. These experiments indicate that glioma-derived factors may directly or indirectly affect the number of circulating neutrophils and influence their infiltration into the tumours.