Analisi dell'immuno-trascrittoma di cavallo nelle patologie IAD e RAO

The research project has been developed on the equine inflammatory respiratory diseases, which can be divided in Recurrent Airway Obstruction (RAO) and Inflammatory Airway Disease (IAD). The aim of this study was to investigate immune-related genes expression in the respiratory tract of IAD and RAO-affected horses. Clinical examination and endoscopy were performed. On the Broncho-Alveolar Lavage (BAL) fluid, obtained during endoscopy, cytological and microbiological analysis were performed to evaluate correlations between the gene expressions values and the clinical parameters. A first analysis was developed by real time RT-PCR comparing the gene expression profile of 10 immune-related target genes (IL-1ß, IL-6, IL-8, IL-13, IL-17, TNFa, INF?, TGF-ß1, NF?-ß and TRL 4) in the BAL of healthy horses and RAO-affected ones, on which sampling was performed twice within 15 days. The aim was to deepen the effects of the respiratory disease on the equine immune system and to assess a potential temporal evolution of the gene expression values and of the other parameters considered. In addition, biopsies of the bronchial tissue were obtained and subsequent, histological evaluation and gene expression analyses were performed. Six of the target genes showed a significant expression values increase in the RAO group compared to the control one. A positive statistical correlation between the amount of mucus in the airways and the expression of some genes investigated was found. Regarding inflammatory mediators expression in the biopsy tissue, neither of target genes was significantly differentially expressed between the RAO horses and the control group. The second part of the research project included also the study of IAD. On all horses, clinical investigations and assessments of gene expression profiles were carried out twice within 15 days, at the diagnosis moment and at the end of the pharmacological treatment. Considering the results of the first study, no biopsies of the respiratory tissue were performed. The development of a microarray platform specific for equine immune-related genes, provide a global view of the pathways involved in the IAD and RAO inflammatory response. The statistical analyses showed that 379 transcripts (55 up-regulated and 324 down-regulated) were significantly differentially expressed between the IAD group and control horses and 1763 genes (903 up-regulated and 860 down-regulated) between the RAO-affected horses and the healthy animals. Between IAD-affected horses and RAO animals, were showed significant differences of the respiratory rate at rest and of the amount of mucus in the airways. Some transcripts involved in the genesis, length and motility of the respiratory epithelium cilia, were down-regulated both in IAD and in RAO horses. In the IAD population, has been demonstrated the over-expression of genes coding for inflammatory mediators. Some of the transcripts up-regulated in the RAO group, are involved in the inflammatory response, bronchoconstriction, apoptosis and hypoxia pathway. In the same disease, some genes involved in the genesis of the protective muco-protein film of the respiratory epithelium were under-expressed. The analyses carried out by the software Gene Sets Enrichment Analysis (GSEA) showed that the pathway activated during human asthma, is also enriched in equine RAO, albeit marginally significant (False Discovery Rate <25%, p value 0.08 ). The low quality of the RNA extracted from the BAL of some samples, did not allow to reach a significant number of horses, considered before and after the pharmacological treatment, to assess the effect of the therapy on gene expression profiles. In conclusion, the present studies provided information about the immunological mechanisms activated during the most important equine respiratory diseases. In the future, the information obtained could lead to the development of new therapies for IAD and RAO, by the inhibition of molecules involved in the pathogenesis of these diseases, as is already done in human medicine. The involvement of the same pathway in human asthma and equine RAO, could suggest a possible role of horses as animal model for the study of human chronic respiratory diseases

Aging, Cognitive Decline and Hearing Loss: Effects of Auditory Rehabilitation and Training with Hearing Aids and Cochlear Implants on Cognitive Function and Depression among Older Adults

A growing interest in cognitive effects associated with speech and hearing processes is spreading throughout the scientific community essentially guided by evidence that central and peripheral hearing loss is associated with cognitive decline. For the present research, 125 participants older than 65 years of age (105 with hearing impairment and 20 with normal hearing) were enrolled, divided into 6 groups according to their degree of hearing loss and assessed to determine the effects of the treatment applied. Patients in our research program routinely undergo an extensive audiological and cognitive evaluation protocol providing results from the Digit Span test, Stroop color-word test, Montreal Cognitive Assessment and Geriatric Depression Scale, before and after rehabilitation. Data analysis was performed for a cross-sectional and longitudinal study of the outcomes for the different treatment groups. Each group demonstrated improvement after auditory rehabilitation or training on shortand long-term memory tasks, level of depression and cognitive status scores. Auditory rehabilitation by cochlear implants or hearing aids is effective also among older adults (median age of 74 years) with different degrees of hearing loss, and enables positive improvements in terms of social isolation, depression and cognitive performance

A new sampling device for faecal immunochemical testing: haemoglobin stability is still an open issue

Abstract Background: The detection of faecal occult blood is a fundamental step in making an early diagnosis of colorectal cancer. The aim of the present study was to evaluate the stability of haemoglobin in faeces collected with two sampling devices specific for faecal immunochemical testing (FOB Gold Tube Screen and FOB Gold Tube NG) that contain different preservative buffers (buffer H, BH, and buffer N, BN, respectively). Methods: Fifteen true positive faecal samples were collected with both devices. A pool from each sample was made. Each pool was portioned and stored at +4\ub0C, +21\ub0C and +32\ub0C for 10 days. One aliquot of each pool stored at each of the respective temperatures was tested at five time intervals between sampling and analysis. The same procedure was followed for three synthetic haemoglobin solutions in both buffers. Results: The percentage of cumulative faecal haemoglobin decrease (HbCD%) was evaluated. No significant difference was found between BH and BN in HbCD% at +4\ub0C (p=0.106); at +21\ub0C and +32\ub0C, HbCD% was lower in BH than in BN samples (p=0.002 and

Performance criteria and quality indicators for the post-analytical phase

Background: Quality indicators (QIs) used as performance measurements are an effective tool in accurately estimating quality, identifying problems that may need to be addressed, and monitoring the processes over time. In Laboratory Medicine, QIs should cover all steps of the testing process, as error studies have confirmed that most errors occur in the pre- and post-analytical phase of testing. Aim of the present study is to provide preliminary results on QIs and related performance criteria in the post-analytical phase. Methods: This work was conducted according to a previously described study design based on the voluntary -participation of clinical laboratories in the project on QIs of the Working Group "Laboratory Errors and Patient Safety" (WG-LEPS) of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC). Results: Overall, data collected highlighted an improvement or stability in performances over time for all reported indicators thus demonstrating that the use of QIs is-effective in the quality improvement strategy. Moreover, QIs data are an important source for defining the state-of- the-art concerning the error rate in the total testing process. The definition of performance specifications based on the state-of-the-art, as suggested by consensus documents, is a valuable benchmark point in evaluating the performance of each laboratory. Conclusions: Laboratory tests play a relevant role in the monitoring and evaluation of the efficacy of patient outcome thus assisting clinicians in decision-making. Laboratory performance evaluation is therefore crucial to providing patients with safe, effective and efficient care

Macro- And Microvascular Functions In Cystic Fibrosis Adults Without Cardiovascular Risk Factors: A Case-Control Study.

Increasing survival from cystic fibrosis show untypical systems involvement, such as cardiocirculatory. In particular, the presence of CFTR in smooth muscle and endothelial cells, systemic inflammation and oxidative stress could explain vascular alterations in these patients. We aimed at noninvasely evaluating macro- and microvascular dysfunction in cystic fibrosis adults without cardiovascular risk factors. Twenty-twoadults affected by cystic fibrosis and 24 healthy volunteers matched for age and sex were enrolled. None had known cardiovascular risk factors. All people underwent blood pressure measurement, microvascular function assessment by EndoPAT-2000 device (calculating RH-PAT index) and macrovascular evaluation by pulse wave velocity (PWV). RH-PAT index was significantly lower in patients than in controls (1.74±0.59 vs 2.33±0.34; p<0.001). Thirteen patients of 22 had a value inferior to the threshold of 1.67 (59.1%), while no controls had (p<0.001). Carotid-femoral PWV did not differ between the two groups (5.2±1.5 m/s vs 5.4±1.1; p=0.9), while brachial-ankle one did (11.0±2.2 m/s vs 10.1±0.8 m/s; p=0.04).Adults patients affected by cystic fibrosis show peripheral endothelial dysfunction, which is the first alteration in atherosclerotic phenomenon. Moreover, arterial stiffness measured by PWV unclearly seems to differ respect of healthy people, perhaps because PWV alterations are typical of above 50 years old people. It is unclear what prognostic role of future developing of atherosclerotic disease these findings could be, but it seems evident that cystic fibrosis directly affects cardiovascular system itself

A Randomized Trial of Pharmacogenetic Warfarin Dosing in Naive Patients with Non-Valvular Atrial Fibrillation

Genotype-guided warfarin dosing have been proposed to improve patient's management. This study is aimed to determine whether a CYP2C9- VKORC1- CYP4F2-based pharmacogenetic algorithm is superior to a standard, clinically adopted, pharmacodynamic method. Two-hundred naive patients with non-valvular atrial fibrillation were randomized to trial arms and 180 completed the study. No significant differences were found in the number of out-of-range INRs (INR3.0) (p = 0.79) and in the mean percentage of time spent in the therapeutic range (TTR) after 19 days in the pharmacogenetic (51.9%) and in the control arm (53.2%, p = 0.71). The percentage of time spent at INR>4.0 was significantly lower in the pharmacogenetic (0.7%) than in the control arm (1.8%) (p = 0.02). Genotype-guided warfarin dosing is not superior in overall anticoagulation control when compared to accurate clinical standard of care

Analisi dell'immuno-trascrittoma di cavallo nelle patologie IAD e RAO

The research project has been developed on the equine inflammatory respiratory diseases, which can be divided in Recurrent Airway Obstruction (RAO) and Inflammatory Airway Disease (IAD). The aim of this study was to investigate immune-related genes expression in the respiratory tract of IAD and RAO-affected horses. Clinical examination and endoscopy were performed. On the Broncho-Alveolar Lavage (BAL) fluid, obtained during endoscopy, cytological and microbiological analysis were performed to evaluate correlations between the gene expressions values and the clinical parameters. A first analysis was developed by real time RT-PCR comparing the gene expression profile of 10 immune-related target genes (IL-1ß, IL-6, IL-8, IL-13, IL-17, TNFa, INF?, TGF-ß1, NF?-ß and TRL 4) in the BAL of healthy horses and RAO-affected ones, on which sampling was performed twice within 15 days. The aim was to deepen the effects of the respiratory disease on the equine immune system and to assess a potential temporal evolution of the gene expression values and of the other parameters considered. In addition, biopsies of the bronchial tissue were obtained and subsequent, histological evaluation and gene expression analyses were performed. Six of the target genes showed a significant expression values increase in the RAO group compared to the control one. A positive statistical correlation between the amount of mucus in the airways and the expression of some genes investigated was found. Regarding inflammatory mediators expression in the biopsy tissue, neither of target genes was significantly differentially expressed between the RAO horses and the control group. The second part of the research project included also the study of IAD. On all horses, clinical investigations and assessments of gene expression profiles were carried out twice within 15 days, at the diagnosis moment and at the end of the pharmacological treatment. Considering the results of the first study, no biopsies of the respiratory tissue were performed. The development of a microarray platform specific for equine immune-related genes, provide a global view of the pathways involved in the IAD and RAO inflammatory response. The statistical analyses showed that 379 transcripts (55 up-regulated and 324 down-regulated) were significantly differentially expressed between the IAD group and control horses and 1763 genes (903 up-regulated and 860 down-regulated) between the RAO-affected horses and the healthy animals. Between IAD-affected horses and RAO animals, were showed significant differences of the respiratory rate at rest and of the amount of mucus in the airways. Some transcripts involved in the genesis, length and motility of the respiratory epithelium cilia, were down-regulated both in IAD and in RAO horses. In the IAD population, has been demonstrated the over-expression of genes coding for inflammatory mediators. Some of the transcripts up-regulated in the RAO group, are involved in the inflammatory response, bronchoconstriction, apoptosis and hypoxia pathway. In the same disease, some genes involved in the genesis of the protective muco-protein film of the respiratory epithelium were under-expressed. The analyses carried out by the software Gene Sets Enrichment Analysis (GSEA) showed that the pathway activated during human asthma, is also enriched in equine RAO, albeit marginally significant (False Discovery Rate <25%, p value 0.08 ). The low quality of the RNA extracted from the BAL of some samples, did not allow to reach a significant number of horses, considered before and after the pharmacological treatment, to assess the effect of the therapy on gene expression profiles. In conclusion, the present studies provided information about the immunological mechanisms activated during the most important equine respiratory diseases. In the future, the information obtained could lead to the development of new therapies for IAD and RAO, by the inhibition of molecules involved in the pathogenesis of these diseases, as is already done in human medicine. The involvement of the same pathway in human asthma and equine RAO, could suggest a possible role of horses as animal model for the study of human chronic respiratory diseases.Il lavoro di ricerca svolto nell’arco dei tre anni di dottorato, è stato articolato in due progetti sviluppati nell’ambito delle malattie respiratorie su base infiammatoria che colpiscono gli equini. Tali patologie possono essere distinte in due grandi gruppi: Recurrent Airway Obstruction (RAO) ed Inflammatory Airway Disease (IAD). Lo scopo dei progetti di ricerca si è basato sull’indagine dei profili di espressione di geni immuno-correlati nell’albero respiratorio di cavalli affetti da IAD e RAO, in relazione ad un gruppo di controllo. Su tutti i soggetti, sono stati eseguiti gli esami clinici mirati alla valutazione dell’apparato respiratorio, l’esame endoscopico e l’esame citologico e microbiologico da Broncho-Alveolar Lavage (BAL), per valutare le potenziali correlazioni esistenti tra i profili di espressione genica ed i parametri clinici. Il primo progetto è stato sviluppato comparando cavalli sani con soggetti affetti da RAO, su cui i campionamenti sono stati ripetuti due volte nell’arco di 15 giorni, al fine valutare una potenziale evoluzione temporale dell’espressione genica e degli altri parametri considerati nella ricerca. Inoltre, sono state eseguite biopsie del tessuto bronchiale, sottoposto sia a valutazione istologica che ad analisi di espressione genica. Mediante real time RT-PCR, sono stati indagati i profili di espressione di 10 geni target immuno-correlati (IL-1ß, IL-6, IL-8, IL-13, IL-17, TNFa, INF?, TGF-ß1, NF?-ß e TRL 4), sei dei quali hanno dimostrato una aumento statisticamente significativo dei livelli di espressione nel gruppo RAO rispetto al gruppo di controllo. Le analisi statistiche condotte, hanno riscontrato una correlazione positiva tra la quantità di muco nelle vie aeree e l’ espressione di alcuni dei geni indagati. Non sono state evidenziate differenze di espressione, dei geni inclusi nello studio, tra i tessuti bioptici prelevati dai soggetti affetti da RAO e quelli ottenuti dal gruppo di controllo. Il secondo progetto di ricerca, è stato sviluppato ampliando la casistica dei cavalli affetti da RAO ed introducendo lo studio della IAD. Su tutti i soggetti, le indagini cliniche e le valutazioni dei profili di espressione genica sono state condotte sia al momento della diagnosi che al termine del trattamento farmacologico della durata di 15 giorni. Valutati i risultati del primo lavoro, non sono state eseguite biopsie del tessuto respiratorio. Lo sviluppo di una piattaforma microarray specifica per i geni immuno-correlati di cavallo ha permesso di ottenere una visione globale dei pathway coinvolti nella risposta infiammatoria delle due patologie. Le analisi statistiche effettuate hanno evidenziato una differenza di espressione significativa per 379 trascritti (di cui 55 sovra-espressi e 324 sotto-espressi) tra il gruppo IAD ed il gruppo di controllo e per 1763 geni (di cui 903 sovra-espressi e 860 sotto-espressi) tra i pazienti affetti da RAO ed i soggetti sani. Da un punto di vista clinico, sono state riscontrate differenze statisticamente significative sia della frequenza respiratoria a riposo che della quantità di muco presente nelle vie aeree dei cavalli affetti da IAD rispetto ai soggetti RAO. Tra i geni sotto-espressi nei due gruppi di cavalli affetti da malattia respiratoria, hanno acquistato importanza alcuni trascritti coinvolti nella genesi, lunghezza e motilità dell’apparato ciliare dell’epitelio respiratorio. Nella popolazione IAD, è stata dimostrata la sovra-espressione di geni codificanti per mediatori coinvolti nella risposta infiammatoria. I geni sovra-espressi nel gruppo RAO, caratterizzati da maggior rilievo, sono coinvolti nella risposta infiammatoria, nella broncocostrizione, nella via apoptotica e nel pathway dell’ipossia. Nella medesima patologia, si sono mostrati sotto-espressi anche alcuni geni coinvolti nella genesi del film muco-proteico di protezione dell’epitelio respiratorio. Lo studio condotto mediante Gene Set Enrichment Analysis (GSEA), ha evidenziato che il pathway attivato in corso di asma umano, viene arricchito anche nella patologia RAO equina, sebbene la significatività statistica sia marginale (False Discovery Rate < 25%, p value 0,08). Non è stato possibile valutare l’effetto della terapia farmacologica sui profili di espressione genica, poiché la bassa qualità dell’RNA estratto dal BAL di alcuni campioni non ha permesso di raggiungere un numero significativo di soggetti valutati prima e dopo il trattamento terapeutico. Gli studi effettuati hanno quindi permesso di far luce su alcuni dei meccanismi immunologici che stanno alla base delle patologie respiratorie equine di maggiore importanza veterinaria ed economica. In futuro, le informazioni ottenute, potrebbero condurre allo sviluppo di nuovi mezzi terapeutici per l’inibizione delle molecole coinvolte nello sviluppo di IAD e RAO, come già avviene in medicina umana. Infine, il coinvolgimento di un medesimo pathway nell’asma umano e nella RAO equina, potrebbe condurre all’utilizzo di tale specie come modello animale per lo studio delle patologie respiratorie croniche umane

Gene Expression Profiles of the Immuno-Transcriptome in Equine Asthma

Background: Mild equine asthma (MEA) and severe equine asthma (SEA) are two of the most frequent equine airway inflammatory diseases, but knowledge about their pathogenesis is limited. The goal of this study was to investigate gene expression differences in the respiratory tract of MEA- and SEA-affected horses and their relationship with clinical signs. Methods: Clinical examination and endoscopy were performed in 8 SEA- and 10 MEA-affected horses and 7 healthy controls. Cytological and microbiological analyses of bronchoalveolar lavage (BAL) fluid were performed. Gene expression profiling of BAL fluid was performed by means of a custom oligo-DNA microarray. Results: In both MEA and SEA, genes involved in the genesis, length, and motility of respiratory epithelium cilia were downregulated. In MEA, a significant overexpression for genes encoding inflammatory mediators was observed. In SEA, transcripts involved in bronchoconstriction, apoptosis, and hypoxia pathways were significantly upregulated, while genes involved in the formation of the protective muco-protein film were underexpressed. The SEA group also showed enrichment of gene networks activated during human asthma. Conclusions: The present study provides new insight into equine asthma pathogenesis, representing the first step in transcriptomic analysis to improve diagnostic and therapeutic approaches for this respiratory disease