Oral Health Status in Subjects with Amnestic Mild Cognitive Impairment and Alzheimer's Disease: Data from the Zabút Aging Project

Background: The relationship between Alzheimer's disease (AD) and periodontitis has been recently investigated with heterogenous results. Objective: This study aims to evaluate the oral health status and its relationship with cognitive impairment of participants, enrolled in the Zabút Aging Project, a community-based cohort study performed in rural community in Sicily, Italy. Methods: A case-control study (20 subjects with AD, 20 with amnestic mild cognitive impairment (aMCI), and 20 controls) was conducted. The protocol included a comprehensive medical and cognitive-behavioral examination. Full-mouth evaluation, microbial analysis of subgingival plaque samples (by RT-PCR analysis), and oral health-related quality of life (OHR-QoL) were evaluated. Results: The decayed, missing, and filled teeth (DMFT) total score of AD subjects was significantly higher than for aMCI (p = 0.009) and controls (p = 0.001). Furthermore, the "M" component of DMFT (i.e., the number of missing teeth) was significantly higher in AD than in aMCI (p < 0.001) and controls (p < 0.001). A Poisson regression model revealed that age (p < 0.001), male gender (p = 0.001), and AD (p = 0.001) were positively correlated with DMFT. Concerning oral microbial load, the presence of Fusobacterium nucleatum was significantly higher in AD than in controls (p = 0.02), and a higher load of Treponema denticola was found in aMCI than with AD (p = 0.004). OHR-QoL scores did not differ among the groups. Conclusion: The current research suggests that AD is associated with chronic periodontitis, which is capable of determining tooth loss due to the pathogenicity of Fusobacterium nucleatum. These data remain to be confirmed in larger population-based cohorts

Generation of two human iPSC lines, FINCBi002-A and FINCBi003-A, carrying heteroplasmic macrodeletion of mitochondrial DNA causing Pearson's syndrome

Pearson marrow pancreas syndrome (PMPS) is a sporadic mitochondrial disease, resulting from the clonal expansion of a mutated mitochondrial DNA (mtDNA) molecule bearing a macro-deletion, and therefore missing essential genetic information. PMPS is characterized by the presence of deleted (Δ) mtDNA that co-exist with the presence of a variable amount of wild-type mtDNA, a condition termed heteroplasmy. All tissues of the affected individual, including the haemopoietic system and the post-mitotic, highly specialized tissues (brain, skeletal muscle, and heart) contain the large-scale mtDNA deletion in variable amount. We generated human induced pluripotent stem cells (hiPSCs) from two PMPS patients, carrying different type of large-scale deletion

Generation of a human iPSC line, FINCBi001-A, carrying a homoplasmic m.G3460A mutation in MT-ND1 associated with Leber's Hereditary optic Neuropathy (LHON)

Leber's Hereditary Optic Neuropathy (LHON) is a maternally inherited disorder caused by homoplasmic mutations of mitochondrial DNA (mtDNA). LHON is characterized by the selective degeneration of the retinal ganglion cells (RGC). Almost all LHON maternal lineages are homoplasmic mutant (100% mtDNA copies are mutant) for one of three frequent mtDNA mutations now found in over 90% of patients worldwide (m.11778G > A/MT-ND4, m.3460G > A/MT-ND1, m.14484 T > C/MT-ND6). Human induced pluripotent stem cells (hiPSCs) were generated from a patient carrying the homoplasmic m.3460G > A/MT-ND1 mutation using the Sendai virus non-integrating virus

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Batteri produttori di β-lattamasi ad ampio spettro in latte ovino massale

L'utilizzo di antibiotici β-lattamici nell'allevamento da latte ha portato alla comparsa di batteri resistenti, soprattutto Gram-negativi, grazie alla produzione di enzimi specifici come β-lattamasi a spettro esteso (ESBL). Ciò rappresenta un problema di salute pubblica a causa del potenziale rischio di trasferimento orizzontale dei geni codificanti questi enzimi, attraverso elementi genetici mobili, nei prodotti di origine animale come il latte. Sebbene siano molti gli studi sulla presenza di germi produttori di ESBL nel latte bovino, sono minori le informazioni riguardanti il latte ovino. Per tale motivo, l'obiettivo di questo studio è stato quello di verificare la presenza di batteri produttori di ESBL in campioni di latte ovino provenienti da due regioni del centro Italia e valutarne alcune caratteristiche quali antibiotico resistenza e patogenicità. Il 66% (64/ 97) dei campioni di latte analizzati è risultato positivo per la presenza di microrganismi in grado di sviluppare su terreno ESBL medium. Di questi, solo il 3% (2/64) era presumibilmente riconducibile a E. coli per il tipico colore rosso magenta delle colonie, mentre il 97% (62/64) era ascrivibile a Pseudomonas spp, Acinetobacter spp. o altri batteri Gram-negativi non fermentanti. Successivamente, il sequenziamento del gene 16S rRNA, effettuato su un gruppo di isolati selezionati (n=25) e affiancato a prove biochimiche, ha permesso di rilevare la presenza di Strenotrophomonas malthophilia (20%) e Stenotrophomonas pavanii (16%), seguiti da Pseudomonas aeruginosa (20%) e altre specie di Pseudomonas di minor interesse (16%), E. coli e Proteus terrae (8% entrambi), Achromobacter mucicolens e Achromobacter xylosoxidans (4% entrambi), e infine Hafnia paralvei (4%). I risultati dell'antibiogramma hanno evidenziato una spiccata resistenza nei confronti dei β-lattamici da parte di alcune specie di interesse clinico quali E. coli e S. malthophilia. Soltanto la specie S. malthophilia ha mostrato resistenza nei confronti dei carbapenemi, mentre nel caso di Pseudomonas spp. l'unica resistenza riscontrata è stata quella nei confronti di sulfametossazolo/trimetoprim. In seguito, le PCR condotte sugli isolati di E. coli, risultati essere commensali, non hanno evidenziato la presenza di geni codificanti enzimi ESBL. Solo nel caso di S. malthophilia è stato possibile rilevare geni codificanti β-lattamasi a spettro esteso come L1 e L2. The use of β-lactam antibiotics in dairy farming has led to the emergence of resistant bacteria, especially Gram-negative ones; Thanks to the production of specific enzymes such as extended spectrum β-lactamases (ESBL). This represents a public health concern due to the potential risk of horizontal transfer of the genes encoding these enzymes, via mobile genetic elements, in products of animal origin such as milk. Although there are many studies on the presence of ESBL-producing germs in bovine milk, there is less information on sheep's milk. For this reason, the aim of this study was to verify the presence of ESBL-producing bacteria in sheep milk samples from two regions of central Italy and to evaluate some characteristics such as antibiotic resistance and pathogenicity. Sixty-six percent (64/97) of milk samples tested were positive for the presence of microrganisms able to growth on ESBL medium. Of these, only 3% (2/64) was presumably attributable to E. coli due to the typical magenta red colour of the colonies, while 97% (62/64) was attributable to Pseudomonas spp, Acinetobacter spp. and other non-fermenting Gram-negative bacteria. Subsequently, the sequencing of the 16S rRNA gene, carried out on a group of selected isolates, (n=25) with others biochemical tests, allowed us to detect the presence of Stenotrophomonas malthophilia (20%) and Stenotrophomonas pavanii (16%), followed by Pseudomonas aeruginosa (20%) and others Pseudomonas species of minor interest (16%), E. coli and Proteus terrae (8% both), Achromobacter mucicolens and Achromobacter xylosoxidans (4% both), and finally Hafnia paralvei (4%). The results of antibiogram test showed a marked resistance against β-lactams in some species of clinical interest such as E. coli and S. malthophilia. Only S. malthophilia showed resistance against carbapenems, while in the case of Pseudomonas spp. the only resistance found was against sulfamethoxazole/trimethoprim. Subsequently, PCR carried out on E. coli isolates, which were commensal, did not show the presence of genes encoding ESBL enzymes. Only in the case of S. malthophilia it was possible to detect genes encoding extended-spectrum β-lactamases such as L1 and L2

Dendritic pathology, spine loss and synaptic reorganization in human cortex from epilepsy patients

Neuronal dendritic arborizations and dendritic spines are crucial for a normal synaptic transmission and may be critically involved in the pathophysiology of epilepsy. Alterations in dendritic morphology and spine loss mainly in hippocampal neurons have been reported both in epilepsy animal models and in human brain tissues from patients with epilepsy. However, it is still unclear whether these dendritic abnormalities relate to the cause of epilepsy or are generated by seizure recurrence. We investigated fine neuronal structures at the level of dendritic and spine organization using Golgi impregnation, and analysed synaptic networks with immunohistochemical markers of glutamatergic (vGLUT1) and GABAergic (vGAT) axon terminals in human cerebral cortices derived from epilepsy surgery. Specimens were obtained from 28 patients with different neuropathologically defined aetiologies: type Ia and type II focal cortical dysplasia, cryptogenic (no lesion) and temporal lobe epilepsy with hippocampal sclerosis. Autoptic tissues were used for comparison. Three-dimensional reconstructions of Golgi-impregnated neurons revealed severe dendritic reshaping and spine alteration in the core of the type II focal cortical dysplasia. Dysmorphic neurons showed increased dendritic complexity, reduction of dendritic spines and occasional filopodia-like protrusions emerging from the soma. Surprisingly, the intermingled normal-looking pyramidal neurons also showed severe spine loss and simplified dendritic arborization. No changes were observed outside the dysplasia (perilesional tissue) or in neocortical postsurgical tissue obtained in the other patient groups. Immunoreactivities of vGLUT1 and vGAT showed synaptic reorganization in the core of type II dysplasia characterized by the presence of abnormal perisomatic baskets around dysmorphic neurons, in particular those with filopodia-like protrusions, and changes in vGLUT1/vGAT expression. Ultrastructural data in type II dysplasia highlighted the presence of altered neuropil engulfed by glial processes. Our data indicate that the fine morphological aspect of neurons and dendritic spines are normal in epileptogenic neocortex, with the exception of type II dysplastic lesions. The findings suggest that the mechanisms leading to this severe form of cortical malformation interfere with the normal dendritic arborization and synaptic network organization. The data argue against the concept that long-lasting epilepsy and seizure recurrence per se unavoidably produce a dendritic pathology

Valutazione dell’esposizione passiva a pesticidi a lungo termine mediante flussi informativi di popolazione: aspetti metodologici

INTRODUZIONE: I fattori ambientali di rischio delle malattie cronico-degenerative quali i contaminanti chimici necessitano con ogni probabilità di un periodo d’azione assai prolungato per esercitare tale effetto, specie per livelli espositivi ridotti. Negli studi caso-controllo di popolazione basati su flussi informativi, tuttavia, la ricostruzione storica dei tempi e della tipologia dell’esposizione è di difficile effettuazione e talora addirittura impossibile. METODI: Abbiamo utilizzato informazioni raccolte nell’ambito di una indagine sulle cause ambientali della sclerosi laterale amiotrofica (SLA) per valutare eventuali differenze nelle stime di rischio per esposizione passiva a pesticidi, utilizzando dati recenti (residenza alla diagnosi nel periodo 1998-2011 in prossimità di coltivazioni specifiche quali vigneti, seminativi, culture orticole e frutteti) e storici (cioè dati residenziali e di uso del suolo riferiti agli anni ‘70). Si è trattato di uno studio caso- controllo di popolazione condotto in tre province emiliane (Parma, Reggio E. e Modena) e in quella di Catania, ricostruendo e georeferenziando la storia residenziale dei soggetti e confrontando tali dati con la cartografia regionale dell’uso del territorio. RISULTATI: I soggetti inclusi nello studio al momento della diagnosi nel periodo 1998-2011 sono stati complessivamente 3440 (703 casi): per 2235 di essi abbiamo potuto ricostruire i dati espositivi relativi agli anni ’70. I valori di rischio relativo di SLA riferiti ai due periodi sono risultati pressochè identici, in entrambi i contesti geografici, pur con una minore stabilità statistica nel periodo meno recente per la ridotta numerosità della casistica. CONCLUSIONI: Nel complesso, questi risultati suggeriscono come nel caso dei pesticidi l’utilizzazione di dati espositivi recenti non pregiudichi in modo significativo la valutazione dell’esposizione a lungo termine

Valutazione dell’esposizione passiva a pesticidi a lungo termine mediante flussi informativi di popolazione: aspetti metodologici

INTRODUZIONE: I fattori ambientali di rischio delle malattie cronico-degenerative quali i contaminanti chimici necessitano con ogni probabilità di un periodo d’azione assai prolungato per esercitare tale effetto, specie per livelli espositivi ridotti. Negli studi caso-controllo di popolazione basati su flussi informativi, tuttavia, la ricostruzione storica dei tempi e della tipologia dell’esposizione è di difficile effettuazione e talora addirittura impossibile. METODI: Abbiamo utilizzato informazioni raccolte nell’ambito di una indagine sulle cause ambientali della sclerosi laterale amiotrofica (SLA) per valutare eventuali differenze nelle stime di rischio per esposizione passiva a pesticidi, utilizzando dati recenti (residenza alla diagnosi nel periodo 1998-2011 in prossimità di coltivazioni specifiche quali vigneti, seminativi, culture orticole e frutteti) e storici (cioè dati residenziali e di uso del suolo riferiti agli anni ‘70). Si è trattato di uno studio caso- controllo di popolazione condotto in tre province emiliane (Parma, Reggio E. e Modena) e in quella di Catania, ricostruendo e georeferenziando la storia residenziale dei soggetti e confrontando tali dati con la cartografia regionale dell’uso del territorio. RISULTATI: I soggetti inclusi nello studio al momento della diagnosi nel periodo 1998-2011 sono stati complessivamente 3440 (703 casi): per 2235 di essi abbiamo potuto ricostruire i dati espositivi relativi agli anni ’70. I valori di rischio relativo di SLA riferiti ai due periodi sono risultati pressochè identici, in entrambi i contesti geografici, pur con una minore stabilità statistica nel periodo meno recente per la ridotta numerosità della casistica. CONCLUSIONI: Nel complesso, questi risultati suggeriscono come nel caso dei pesticidi l’utilizzazione di dati espositivi recenti non pregiudichi in modo significativo la valutazione dell’esposizione a lungo termine