- Email: [email protected]
NEW CONCEPTS IN THE PATHOPHYSIOLOGY OF ALZHEIMER’S DISEASE
P804
Poster Presentations: Monday, July 17, 2017
top predictors of mixed pathology. The quality-controlled rectangular data set was comprised of 39 predictors that included demographics such as age at death, sex, and education along with vascular risk factors, medical conditions, IADLs, APOE ε2 and ε4 alleles, and slopes for episodic memory, semantic memory, working memory, perceptual speed, and visuospatial ability estimated from annual follow-up visits prior to death (mean 8.9 (s.d.¼4.7) years). We performed a Poisson regression with robust standard errors on the top five predictors from the RF model to look at the magnitude and direction of the predictors. Results: The optimal RF model classified the two groups with 81% accuracy. The top five predictors of mixed neuropathology were greater decline in episodic and semantic memory, greater IADL impairment at final visit, slower decline in perceptual speed, and no history of thyroid disease. Rate of decline in episodic memory (p-value < 0.001) and older age at death (p-value ¼ 0.01) were statistically significant in our Poisson regression model that included all five predictors, age at death and sex, with a one standard deviation greater decline in episodic memory associated with an 11% increase in the “risk” of mixed neuropathology. Conclusions: Investigation of longitudinal cognitive symptoms could provide a more accurate approach for ante-mortem differentiation of pure AD and other major dementia syndromes.
P2-438
COGNITION AND HIPPOCAMPAL PATHOLOGIC DENSITY OF a-SYNUCLEIN, TAU AND AMYLOID-b IN DEMENTIAWITH LEWY BODIES
Tanis J. Ferman1, Naoya Aoki2, Koji Kasanuki1, Dennis W. Dickson1, 1 Mayo Clinic, Jacksonville, FL, USA; 2Yokohama City University Medical Center, Yokohama, Japan. Contact e-mail: [email protected] Background: In DLB, limbic pathology contributes to the cognitive difficulties of DLB, but little is known about the role of the hippocampus. We sought to determine whether the density of a-synuclein, tau and amyloid-b pathology in hippocampal subregions is associated with baseline cognitive performance in dementia with Lewy bodies (DLB). Methods: Patients with clinically probable DLB were followed prospectively until autopsy (n¼49). Quantitative image analyses of a-synuclein (LB509), tau (PHF-1) and amyloid-b (4G8) density in the CA1, CA2/3, CA4 and subiculum regions of the hippocampus were carried out. A Memory and Naming composite score was composed of the mean age-adjusted scaled scores of Logical Memory delay, Auditory Verbal Learning Test delay, and Boston Naming Test. An Attention and Visual composite score was composed of the mean age-adjusted scaled scores of Trail Making A, Stroop Word, Rey Complex Figure copy, and Block Design. Spearman rank correlations between pathology and cognitive composite and individual tasks were carried out. Results: Mean duration of illness was 7 6 3 years, mean age at death was 75 6 7, and baseline mean MMSE was 23 6 4. Deposition of tau and amyloid-b was greatest in the subiculum and CA1, while deposition of a-synuclein was greatest in the subiculum and CA2/ 3 (p<0.01). The Memory and Naming composite was correlated with tau in the CA1 (r ¼ -0.35, p<0.01) and subiculum (r ¼ -0.41, p<0.01), with a-synuclein in the subiculum (r¼ -0.47, p<0.01), and with amyloid-b from all hippocampal sectors (but strongest in the subiculum, r ¼ -0.45, p<0.01). The Attention and Visual composite score was correlated with a-synuclein in the subiculum (r¼ -0.31, p¼0.03) and in CA2/3 (r ¼ -0.30, p¼0.03). Conclusions: Memory and naming difficulties were associated with a greater burden of
tau and amyloid-b in the CA1 region, and with greater a-synuclein, tau and amyloid-b in the subiculum. Worse attention and visual processing performance was associated with greater a-synuclein density in the subiculum and CA2/3 hippocampal subregions, a pattern that may be related to the dense interconnectivity to neighboring limbic regions known to be affected in DLB.
P2-439
NEURONAL AND GLIAL CHANGES IN HIPPOCAMPUS IN MODEL OF CHRONIC CEREBRAL ISCHEMIA
Alexey Vladimirovich Smirnov1,2, Ivan Nikolaevich Tyurenkov2, Dmitry Mednikov2, Maxim Schmidt1,2, Maria Ekova2, Denis Kurkin2, Elena Volotova2, 1Volgograd Medical Research Center, Volgograd, Russian Federation; 2Volgograd State Medical University, Volgograd, Russian Federation. Contact e-mail: [email protected] Background: The chronic cerebral ischemia and hypoxemic damage
of neurons, which is frequently caused by atherosclerosis and arterial hypertension, stimulate development of vascular dementia. The impairment in learning, memory functions, cognitive skills, emotions and quality of life are usually associated with injury of hippocampal neurons and synaptic plasticity. Methods: Chronic cerebral ischemia was modeled in rats using a gravitational overload with a force of 9 G in a caudo-cranial vector for 5 minutes twice per day with interval of 12 hours for 28 days. The behavioral status, cognitive functions and memory, the nature of damage of neurons and level of an expression of glial fibrillary acidic protein (GFAP) in brain paraffin sections were measured. Results: After 28 days of chronic cerebral ischemia modeling functional and morphological manifestations of vascular dementia in rats were noted. Cognitive violations manifested by decrease in orientingexploratory activity, decrease in memory and ability to memorize new objects, decreasing in safety of main reflexes were also found. Significant increased amount of pyknotic neurons and neurons with the hyperchromatosis of cytoplasm in perikarya in all zones of hippocampus (p<0.05) were revealed. Decrease in relative density of pyramidal neurons (p<0.05) and focal loss of neurons of pyramidal layer, pericellular edema were combined with increased expression of GFAP in astrocytes of CA2 and CA4. Conclusions: The cognitive violations and behavioral disorders found in the functional tests in vascular dementia modeling are caused by both reversible and irreversible progressing injuries of hyppocampal neurons. Decrease of pyramidal neurons relative density and increase in expression of GFAP in CA2 and CA4 demonstrates the emergence of neurodegenerative changes, activation of astrocytes and gliosis in these zones.
P2-440
NEW CONCEPTS IN THE PATHOPHYSIOLOGY OF ALZHEIMER’S DISEASE
Andre Lopez Fernandez, Sr., Fernanda Betinas Gutierre da Costa Ramos, Sr., M^onica Maria Agata Stiepcich, Sr., Centro Universitario S~ao Camilo, S~ao Paulo, Brazil. Contact e-mail: andre.lopez13@ hotmail.com Background: The purpose of this literature review was to review the major pathophysiological hypothesis of AD. It was analyzed the secondary importance but no less important the cholinergic and glutamatergic hypotheses; beside that, it was observed the central role of the amyloid cascade hypothesis, with a greater emphasis
Poster Presentations: Monday, July 17, 2017
P805
Martin N. Rossor1, Nick C. Fox1, Tamas Revesz5, Tammaryn Lashley5, Janice Holton5, Jonathan D. Rohrer1, 1Dementia Research Centre, Institute of Neurology, University College London, London, United Kingdom; 2 Translational Imaging Group, University College London, London, United Kingdom; 3UCL Institute of Neurology, London, United Kingdom; 4NIHR Queen Square Dementia Biomedical Research Unit, Institute of Neurology, University College London, London, United Kingdom; 5Queen Square Brain Bank, Department of Molecular Neuroscience, Institute of Neurology, University College London, London, United Kingdom; 6Centre for Medical Image Computing, University College London, London, United Kingdom. Contact e-mail: [email protected] Background: White matter hyperintensities (WMH) are present on
in the soluble oligomers bA in relation to Neuritic Plaques (NC), the influence of factors such as apolipoprotein E (ApoE), Tau protein and genetic mutations. And this review also emphasized the contribution of oxidative stress, neuroinflammation and metabolic syndrome as the main triggers of the disease, especially in its resemblance to the Diabetes mellitus, changes of intestinal flora and the promising hypothesis of microbiot. Methods: For the accomplishment of this review we used scientific articles found in the following online databases: LILACS, MEDLINE, PUBMED and SciELO, prioritizing publications of the last 15 years (20162001). Descriptors were used: Alzheimer’s, Amyloid b, Ab, Alzheimer’s and Oxidative Stress, Alzheimer’s and Neuroinflammation, Alzheimer’s and microbiot, Amyloid Precursos Protein, Amyloid Hypothesis, Diabetes Type 3, Alzheimer’s and NMDA, Alzheimer’s and Cholinergic, Alzheimer’s and ApoE, Alzheimer’s and Mitochondria, B-oligomers, Alzheimer’s and tau, Alzheimer’s and Neurofibrillary tangle, Alzheimer’s and Aging. As a selection criteria, information contained in the abstracts and discussions of the articles found in the cited databases was analyzed, prioritizing pathophysiological aspects and molecular mechanisms, reaching a total of 117 scientific articles selected and used, 6 of them from the period before 2001, given their relevance and scientific validity. In addition to the articles were used the United Nations report (World Population Aging 2015) and the Tratado de Neurologia da Academia Brasileira de Neurologia. Results: All the hypothesis analysed - Oxidative Stress, Neuroinflammation, Cholinergic, Glutamatergic, Metabolic and Diabetes Type 3, Microbiot and Amyloid Hypothesis are related, often with positive feedback systems perpetuating and worsing the neurodegeneration. Conclusions: It is concluded the increasing importance of environmental and systemic factors in the initiation of disease and the need to identify new triggers that will improve prevention and therapies and, consequently, will avoid in early stages these positive feedback systems that make the current therapy for Alzheimer’s disease ineffective.
P2-441
PATHOLOGICAL CORRELATES OF WHITE MATTER HYPERINTENSITIES ON CADAVERIC MRI IN PROGRANULINASSOCIATED FRONTOTEMPORAL DEMENTIA
Ione OC. Woollacott1, Martina Bocchetta1, Carole H. Sudre2,3, Basil H. Ridha4, Catherine Strand5, Robert Courtney5, Sebastien Ourselin1,6, Jorge M. Cardoso1,6, Jason D. Warren1,
magnetic resonance imaging (MRI) brain scans of patients with frontotemporal dementia (FTD) associated with progranulin (GRN) mutations. However, their histopathological correlates are unknown. Cadaveric MRI scanning provides undistorted, in situ brain images, allowing precise spatial and quantitative correlations of neuroimaging abnormalities with histology in post-mortem brain tissue. We report a patient with behavioural variant FTD and a GRN Q130fs mutation, with extensive WMH, who underwent detailed cadaveric MRI and characterisation of WMH pathological correlates. Methods: The patient underwent cadaveric in situ brain MRI (GE Signa 1.5T scanner) within 24 hours after death, including volumetric T1 and FLAIR sequences. MRI had also been performed in vivo 2.6 years earlier. Five brain regions were selected for histopathological analysis, corresponding to areas of severe or absent WMH on FLAIR MRI: right frontal pole, anterior frontal, posterior frontal, temporal and occipital lobes. Histological examination was performed using sections stained with haematoxylin and eosin, Luxol fast blue and Perl stains and immunohistochemical staining for TDP-43, Ab, phosphorylated neurofilament (SMI31), myelin basic protein and glial fibrillary acidic protein. Vascular pathology was semi-quantitatively assessed in regions of interest using the recommendations of the Vascular Cognitive Impairment Neuropathology Guidelines. Results: In vivo and cadaveric MRI showed progressive, asymmetric (left worse than right) frontotemporal and parietal atrophy, and asymmetric WMH, predominantly affecting frontal white matter. TDP-43 type A pathology was present in the neocortex. Brain regions with most severe WMH on MRI (frontal pole and anterior frontal lobe) displayed the most severe cortical pathology when assessed for neuronal loss, spongiosis, gliosis, myelin loss and TDP-43 burden. In regions found to have WMH on MRI, there was no or minimal vascular pathology. The severity of cortical pathology corresponded with the degree of white matter pathology in all regions analysed. Conclusions: Vascular pathology does not appear to underlie the extensive WMH seen in GRN-mutation associated FTD. WMH could be due to axonal degeneration secondary to cortical neuronal loss, or, given the known role of GRN in inflammatory pathways, excessive inflammation leading to demyelination. Further study of WMH histopathological correlates across a larger cohort of patients is required.
P2-442
NEURODEGENERATION-ASSOCIATED PROTEINS IN HUMAN OLFACTORY NEURONS
Gianluigi Zanusso1, Luca Sacchetto1, Lorenzo Brozzetti1, Dario Olivieri1, Sergio Ferrari1, Luca Faccioli1, Matilde Bongianni1, Matteo Pedrazzoli1, Mario Buffelli1, Salvatore Monaco1, Maurizio Pocchiari2, Byron Caughey3, Bernardino Francesco Ghetti4, 1University of Verona, Verona, Italy;
Poster Presentations: Monday, July 17, 2017
top predictors of mixed pathology. The quality-controlled rectangular data set was comprised of 39 predictors that included demographics such as age at death, sex, and education along with vascular risk factors, medical conditions, IADLs, APOE ε2 and ε4 alleles, and slopes for episodic memory, semantic memory, working memory, perceptual speed, and visuospatial ability estimated from annual follow-up visits prior to death (mean 8.9 (s.d.¼4.7) years). We performed a Poisson regression with robust standard errors on the top five predictors from the RF model to look at the magnitude and direction of the predictors. Results: The optimal RF model classified the two groups with 81% accuracy. The top five predictors of mixed neuropathology were greater decline in episodic and semantic memory, greater IADL impairment at final visit, slower decline in perceptual speed, and no history of thyroid disease. Rate of decline in episodic memory (p-value < 0.001) and older age at death (p-value ¼ 0.01) were statistically significant in our Poisson regression model that included all five predictors, age at death and sex, with a one standard deviation greater decline in episodic memory associated with an 11% increase in the “risk” of mixed neuropathology. Conclusions: Investigation of longitudinal cognitive symptoms could provide a more accurate approach for ante-mortem differentiation of pure AD and other major dementia syndromes.
P2-438
COGNITION AND HIPPOCAMPAL PATHOLOGIC DENSITY OF a-SYNUCLEIN, TAU AND AMYLOID-b IN DEMENTIAWITH LEWY BODIES
Tanis J. Ferman1, Naoya Aoki2, Koji Kasanuki1, Dennis W. Dickson1, 1 Mayo Clinic, Jacksonville, FL, USA; 2Yokohama City University Medical Center, Yokohama, Japan. Contact e-mail: [email protected] Background: In DLB, limbic pathology contributes to the cognitive difficulties of DLB, but little is known about the role of the hippocampus. We sought to determine whether the density of a-synuclein, tau and amyloid-b pathology in hippocampal subregions is associated with baseline cognitive performance in dementia with Lewy bodies (DLB). Methods: Patients with clinically probable DLB were followed prospectively until autopsy (n¼49). Quantitative image analyses of a-synuclein (LB509), tau (PHF-1) and amyloid-b (4G8) density in the CA1, CA2/3, CA4 and subiculum regions of the hippocampus were carried out. A Memory and Naming composite score was composed of the mean age-adjusted scaled scores of Logical Memory delay, Auditory Verbal Learning Test delay, and Boston Naming Test. An Attention and Visual composite score was composed of the mean age-adjusted scaled scores of Trail Making A, Stroop Word, Rey Complex Figure copy, and Block Design. Spearman rank correlations between pathology and cognitive composite and individual tasks were carried out. Results: Mean duration of illness was 7 6 3 years, mean age at death was 75 6 7, and baseline mean MMSE was 23 6 4. Deposition of tau and amyloid-b was greatest in the subiculum and CA1, while deposition of a-synuclein was greatest in the subiculum and CA2/ 3 (p<0.01). The Memory and Naming composite was correlated with tau in the CA1 (r ¼ -0.35, p<0.01) and subiculum (r ¼ -0.41, p<0.01), with a-synuclein in the subiculum (r¼ -0.47, p<0.01), and with amyloid-b from all hippocampal sectors (but strongest in the subiculum, r ¼ -0.45, p<0.01). The Attention and Visual composite score was correlated with a-synuclein in the subiculum (r¼ -0.31, p¼0.03) and in CA2/3 (r ¼ -0.30, p¼0.03). Conclusions: Memory and naming difficulties were associated with a greater burden of
tau and amyloid-b in the CA1 region, and with greater a-synuclein, tau and amyloid-b in the subiculum. Worse attention and visual processing performance was associated with greater a-synuclein density in the subiculum and CA2/3 hippocampal subregions, a pattern that may be related to the dense interconnectivity to neighboring limbic regions known to be affected in DLB.
P2-439
NEURONAL AND GLIAL CHANGES IN HIPPOCAMPUS IN MODEL OF CHRONIC CEREBRAL ISCHEMIA
Alexey Vladimirovich Smirnov1,2, Ivan Nikolaevich Tyurenkov2, Dmitry Mednikov2, Maxim Schmidt1,2, Maria Ekova2, Denis Kurkin2, Elena Volotova2, 1Volgograd Medical Research Center, Volgograd, Russian Federation; 2Volgograd State Medical University, Volgograd, Russian Federation. Contact e-mail: [email protected] Background: The chronic cerebral ischemia and hypoxemic damage
of neurons, which is frequently caused by atherosclerosis and arterial hypertension, stimulate development of vascular dementia. The impairment in learning, memory functions, cognitive skills, emotions and quality of life are usually associated with injury of hippocampal neurons and synaptic plasticity. Methods: Chronic cerebral ischemia was modeled in rats using a gravitational overload with a force of 9 G in a caudo-cranial vector for 5 minutes twice per day with interval of 12 hours for 28 days. The behavioral status, cognitive functions and memory, the nature of damage of neurons and level of an expression of glial fibrillary acidic protein (GFAP) in brain paraffin sections were measured. Results: After 28 days of chronic cerebral ischemia modeling functional and morphological manifestations of vascular dementia in rats were noted. Cognitive violations manifested by decrease in orientingexploratory activity, decrease in memory and ability to memorize new objects, decreasing in safety of main reflexes were also found. Significant increased amount of pyknotic neurons and neurons with the hyperchromatosis of cytoplasm in perikarya in all zones of hippocampus (p<0.05) were revealed. Decrease in relative density of pyramidal neurons (p<0.05) and focal loss of neurons of pyramidal layer, pericellular edema were combined with increased expression of GFAP in astrocytes of CA2 and CA4. Conclusions: The cognitive violations and behavioral disorders found in the functional tests in vascular dementia modeling are caused by both reversible and irreversible progressing injuries of hyppocampal neurons. Decrease of pyramidal neurons relative density and increase in expression of GFAP in CA2 and CA4 demonstrates the emergence of neurodegenerative changes, activation of astrocytes and gliosis in these zones.
P2-440
NEW CONCEPTS IN THE PATHOPHYSIOLOGY OF ALZHEIMER’S DISEASE
Andre Lopez Fernandez, Sr., Fernanda Betinas Gutierre da Costa Ramos, Sr., M^onica Maria Agata Stiepcich, Sr., Centro Universitario S~ao Camilo, S~ao Paulo, Brazil. Contact e-mail: andre.lopez13@ hotmail.com Background: The purpose of this literature review was to review the major pathophysiological hypothesis of AD. It was analyzed the secondary importance but no less important the cholinergic and glutamatergic hypotheses; beside that, it was observed the central role of the amyloid cascade hypothesis, with a greater emphasis
Poster Presentations: Monday, July 17, 2017
P805
Martin N. Rossor1, Nick C. Fox1, Tamas Revesz5, Tammaryn Lashley5, Janice Holton5, Jonathan D. Rohrer1, 1Dementia Research Centre, Institute of Neurology, University College London, London, United Kingdom; 2 Translational Imaging Group, University College London, London, United Kingdom; 3UCL Institute of Neurology, London, United Kingdom; 4NIHR Queen Square Dementia Biomedical Research Unit, Institute of Neurology, University College London, London, United Kingdom; 5Queen Square Brain Bank, Department of Molecular Neuroscience, Institute of Neurology, University College London, London, United Kingdom; 6Centre for Medical Image Computing, University College London, London, United Kingdom. Contact e-mail: [email protected] Background: White matter hyperintensities (WMH) are present on
in the soluble oligomers bA in relation to Neuritic Plaques (NC), the influence of factors such as apolipoprotein E (ApoE), Tau protein and genetic mutations. And this review also emphasized the contribution of oxidative stress, neuroinflammation and metabolic syndrome as the main triggers of the disease, especially in its resemblance to the Diabetes mellitus, changes of intestinal flora and the promising hypothesis of microbiot. Methods: For the accomplishment of this review we used scientific articles found in the following online databases: LILACS, MEDLINE, PUBMED and SciELO, prioritizing publications of the last 15 years (20162001). Descriptors were used: Alzheimer’s, Amyloid b, Ab, Alzheimer’s and Oxidative Stress, Alzheimer’s and Neuroinflammation, Alzheimer’s and microbiot, Amyloid Precursos Protein, Amyloid Hypothesis, Diabetes Type 3, Alzheimer’s and NMDA, Alzheimer’s and Cholinergic, Alzheimer’s and ApoE, Alzheimer’s and Mitochondria, B-oligomers, Alzheimer’s and tau, Alzheimer’s and Neurofibrillary tangle, Alzheimer’s and Aging. As a selection criteria, information contained in the abstracts and discussions of the articles found in the cited databases was analyzed, prioritizing pathophysiological aspects and molecular mechanisms, reaching a total of 117 scientific articles selected and used, 6 of them from the period before 2001, given their relevance and scientific validity. In addition to the articles were used the United Nations report (World Population Aging 2015) and the Tratado de Neurologia da Academia Brasileira de Neurologia. Results: All the hypothesis analysed - Oxidative Stress, Neuroinflammation, Cholinergic, Glutamatergic, Metabolic and Diabetes Type 3, Microbiot and Amyloid Hypothesis are related, often with positive feedback systems perpetuating and worsing the neurodegeneration. Conclusions: It is concluded the increasing importance of environmental and systemic factors in the initiation of disease and the need to identify new triggers that will improve prevention and therapies and, consequently, will avoid in early stages these positive feedback systems that make the current therapy for Alzheimer’s disease ineffective.
P2-441
PATHOLOGICAL CORRELATES OF WHITE MATTER HYPERINTENSITIES ON CADAVERIC MRI IN PROGRANULINASSOCIATED FRONTOTEMPORAL DEMENTIA
Ione OC. Woollacott1, Martina Bocchetta1, Carole H. Sudre2,3, Basil H. Ridha4, Catherine Strand5, Robert Courtney5, Sebastien Ourselin1,6, Jorge M. Cardoso1,6, Jason D. Warren1,
magnetic resonance imaging (MRI) brain scans of patients with frontotemporal dementia (FTD) associated with progranulin (GRN) mutations. However, their histopathological correlates are unknown. Cadaveric MRI scanning provides undistorted, in situ brain images, allowing precise spatial and quantitative correlations of neuroimaging abnormalities with histology in post-mortem brain tissue. We report a patient with behavioural variant FTD and a GRN Q130fs mutation, with extensive WMH, who underwent detailed cadaveric MRI and characterisation of WMH pathological correlates. Methods: The patient underwent cadaveric in situ brain MRI (GE Signa 1.5T scanner) within 24 hours after death, including volumetric T1 and FLAIR sequences. MRI had also been performed in vivo 2.6 years earlier. Five brain regions were selected for histopathological analysis, corresponding to areas of severe or absent WMH on FLAIR MRI: right frontal pole, anterior frontal, posterior frontal, temporal and occipital lobes. Histological examination was performed using sections stained with haematoxylin and eosin, Luxol fast blue and Perl stains and immunohistochemical staining for TDP-43, Ab, phosphorylated neurofilament (SMI31), myelin basic protein and glial fibrillary acidic protein. Vascular pathology was semi-quantitatively assessed in regions of interest using the recommendations of the Vascular Cognitive Impairment Neuropathology Guidelines. Results: In vivo and cadaveric MRI showed progressive, asymmetric (left worse than right) frontotemporal and parietal atrophy, and asymmetric WMH, predominantly affecting frontal white matter. TDP-43 type A pathology was present in the neocortex. Brain regions with most severe WMH on MRI (frontal pole and anterior frontal lobe) displayed the most severe cortical pathology when assessed for neuronal loss, spongiosis, gliosis, myelin loss and TDP-43 burden. In regions found to have WMH on MRI, there was no or minimal vascular pathology. The severity of cortical pathology corresponded with the degree of white matter pathology in all regions analysed. Conclusions: Vascular pathology does not appear to underlie the extensive WMH seen in GRN-mutation associated FTD. WMH could be due to axonal degeneration secondary to cortical neuronal loss, or, given the known role of GRN in inflammatory pathways, excessive inflammation leading to demyelination. Further study of WMH histopathological correlates across a larger cohort of patients is required.
P2-442
NEURODEGENERATION-ASSOCIATED PROTEINS IN HUMAN OLFACTORY NEURONS
Gianluigi Zanusso1, Luca Sacchetto1, Lorenzo Brozzetti1, Dario Olivieri1, Sergio Ferrari1, Luca Faccioli1, Matilde Bongianni1, Matteo Pedrazzoli1, Mario Buffelli1, Salvatore Monaco1, Maurizio Pocchiari2, Byron Caughey3, Bernardino Francesco Ghetti4, 1University of Verona, Verona, Italy;