Characterization of the macromolecular interactions of the key NHEJ components

Non-homologous end-joining (NHEJ) is a major mechanism for repairing DNA double strand breaks in mammalian cells. Six ‘core’ NHEJ components have been identified to date: Ku70, Ku80, the DNA dependent protein kinase catalytic subunit DNA-PKcs, DNA ligase IV, XRCC4, and XLF-Cemunnos. In this thesis, the DNA binding properties of the Ku70/80 heterodimer are characterised by means of fluorescence anisotropy and electrophoretic mobility shift assays. Studies with duplexes holding 1, 2 or 3 heterodimers allowed to develop a binding model that can be applied to analyse the interactions of Ku with duplexes of any length. In addition, salt dependent studies indicate that electrostatic interactions play a major role in the binding of Ku to nucleic acids. The interaction of the Ku heterodimer with the DNA ligase IV/XRCC4 (LX) complex and the role of DNA-PKcs in regulating the NHEJ complex assembly are investigated. Protein-protein interaction experiments show that Ku interacts with DNA ligase IV via its tandem BRCT domain and this interaction is enhanced by the presence of XRCC4 and DNA. In particular, mutagenesis studies indicate that residues 643-748 encompassing the first BRCT domain of DNA ligase IV are necessary for binding. Moreover, Ku needs to be in its heterodimeric form to bind LX and the C-terminal of Ku80 is dispensable for this interaction. The presence of DNA-PKcs favours the interaction between Ku and LX, although its kinase activity induces the disassembly of the complex. Finally, the interaction of Ku with other factors potentially involved in NHEJ such as the replication protein A, PARP-1, and the WRN helicase is investigated. Collectively, these findings provide novel information on the macromolecular interactions that regulate the NHEJ pathway

Assessment of trabecular bone score (TBS) in overweight/obese men: effect of metabolic and anthropometric factors

The "trabecular bone score" (TBS) indirectly explores bone quality, independently of bone mineral density (BMD). We investigated the effects of anthropometric and metabolic parameters on TBS in 87 overweight/obese men. We assessed BMD and TBS by DXA, and some parameters of glucose metabolism, sex-and calciotropic hormone levels. Regression models were adjusted for either age and BMI, or age and waist circumference, or age and waist/hip ratio, also considering BMI >35 (y/n) and metabolic syndrome (MS) (y/n). Correlations between TBS and parameters studied were higher when correcting for waist circumference, although not significant in subjects with BMI >35. The analysis of covariance showed that the same model always had a higher adjusted r-square index. BMD at lumbar spine and total hip, fasting glucose, bioavailable testosterone, and sex hormone-binding globulin are the only covariates having a significant effect (p 35 on TBS values or significant interaction terms between each covariate and either BMI >35 or the presence of MS. Obesity negatively affected TBS, despite unchanged BMD. Alterations of glucose homeostasis and sex hormone levels seem to influence this relationship, while calciotropic hormones have no role. The effect of waist circumference on TBS is more pronounced than that of BMI

High potency of melaleuca alternifolia essential oil against multi-drug resistant gram-negative bacteria and methicillin-resistant staphylococcus aureus

Purpose: Herein, an extended investigation of Tea tree oil (TTO) against a number of multi-drug resistant (MDR) microorganisms in liquid and vapor phases is reported. Methods: The activity of TTO was tested against methicillin-sensitive Staphylococcus aureus (MSSA), Escherichia coli, and clinical strains of methicillin-resistant S. aureus (MRSA), extended-spectrum beta lactamases producer carbapenem-sensitive Klebsiella pneumoniae (ESBL-CS-Kp), carbapenem-resistant K. pneumoniae (CR-Kp), Acinetobacter baumannii (CR-Ab), and Pseudomonas aeruginosa (CR-Pa). Minimal inhibitory/bactericidal concentrations (MIC/MBCs) and synergistic activity between TTO and different antimicrobials were determined. In the vapor assay (VP), TTO-impregnated discs were placed on the lid of a petri dish and incubated for 24 h at 37°C. Results: TTO showed a potent bactericidal activity against all the tested microorganisms. TTO in combination with each reference antimicrobial showed a high level of synergism at sub-inhibitory concentrations, particularly with oxacillin (OXA) against MRSA. The VP assay showed high activity of TTO against CR-Ab. Conclusion: Evaluation of in-vitro activity clearly indicated TTO as a potential effective antimicrobial treatment either alone or in association with known drugs against MDR. Therefore, TTO could represent the basis for a possible role in non-conventional regimens against S. aureus and Gram-negative MDR. TTO in VP might represent a promising option for local therapy of pneumonia caused by CR-Ab

Discriminating among multiple components affecting bulk atmospheric deposition chemistry: a multivariate approach using data from a forest plot in Calabria (Southern Italy)

This study examines the relationships between meteorology and atmospheric deposition chemistry on the basis of 4 years of monitoring in an area of Calabria (Piano Limina) under the National Integrated Programme for the Control of Forest Ecosystems. The location of the area and its low anthropogenic impact meant that phenomena of locally originating alkaline dust deposition could be distinguished from those originating long distances away. The analysis performed on the whole dataset revealed the interaction between temperature, solar radiation and ionic concentrations; the effects of the atmospheric transport of compounds, with lower concentrations during calm conditions; and a marked increase of calcium, alkalinity and pH with winds from W-SW, owing to the transport of alkaline dust from North Africa, in agreement with thematic maps on the synoptic scale. The possible influence of two volcanic events deriving from Stromboli and Etna is discussed. After elimination of the Saharan dust and volcanic events, a multivariate analysis showed the effects of compounds deriving from anthropogenic activities. Sulphate, nitrate and ammonium were closely correlated with NW winds; air masses from this direction come from the continental land mass and the sea, crossing the Calabrian plain before being deposited as precipitation on the Apennine chain. The component from NW also includes a high marine contribution, with maximum values of chloride and sodium

Palliative care training addressed to hospital healthcare professionals by palliative care specialists: a mixed-method evaluation

Despite the great advances in the concept of palliative care (PC) and its benefits, its application seems to be delayed, leaving unfulfilled the many needs of patients and family members. One way to overcome this difficulty could be to develop a new training programme by palliative care specialists to improve PC primary skills in healthcare professionals. The aim of this study was to evaluate the training's impact on trainees within a hospital setting using Kirkpatrick's and Moore's models

Analysis of colorectal cancers for human cytomegalovirus presence

<p>Abstract</p> <p>Background</p> <p>A possible association between human cytomegalovirus (HCMV) infection and colorectal cancer progression has been inferred by the identification in tumour tissues of HCMV antigens and specific viral DNA or RNA sequences. To further investigate the relationship between HCMV and colorectal cancers we developed qualitative and quantitative PCR assay to detect HCMV DNA in 56 formalin-fixed paraffin-embedded (FFPE) tissue samples from patients belonging to 4 different histological phenotypes: adenoma; poorly, moderately and well differentiated adenocarcinomas.</p> <p>Results</p> <p>Of the 56 FFPE tested tissue samples, 6 (11%) were positive for HCMV nested PCR amplification, and more precisely 1 (5%) of 20 cases of adenoma and 5 (21%) of 24 cases of moderately differentiated adenocarcinoma. No PCR positivity was obtained in samples from well and poorly differentiated adenocarcinomas.</p> <p>Conclusion</p> <p>Our observations suggest that there is no evidence of a direct association between HCMV and colorectal cancer. Moreover, the results obtained are not supportive of a causal role of HCMV in the processes of carcinogenesis and/or progression of colorectal cancer. However, the fact that the virus may present a "hit and run" like-mechanism and HCMV can thus only be detectable at a particular stage of a processing adenocarcinoma, suggests that a significant number of colorectal cancers might have been the subject of HCMV infection that could contribute to trigger the oncogenic differentiation. Our analysis does not exclude the possibility of HCMV infection subsequent viral clearance.</p

Is there a difference in phenotype between males and females with non-transfusion-dependent thalassemia? A cross-sectional evaluation.

Non-transfusion-dependent thalassemia includes a variety of phenotypes and genotypes that rarely require regular transfusions. However, these patients can experience a wide range of complications. The objective of this retrospective study was to verify whether there is a significant difference in non-transfusion-dependent thalassemia-related complications and treatment among males and females.We performed a re-analysis of samples evaluated in a previously published cross-sectional study, regarding 96 non-transfusion-dependent thalassemia patients followed at the 'UOSD Malattie Rare del Globulo Rosso' Centre of the Cardarelli Hospital in Naples, Italy.We found that females were more anemic than males, but there was no significant difference in prevalence of common complications among genders, except for hypogonadism. Furthermore, the transitory regular transfusions regimen in women who had been pregnant does not seem to have a significant impact on overall prognosis.In non-transfusion-dependent thalassemia patients, the lower levels of hemoglobin found in females do not seem to indicate a higher prevalence of complications.This data should be considered in studies with experimental treatments aiming to correct anemia in patients with non-transfusion-dependent thalassemia. It should probably also be taken into account in order to set up different transfusion regimens among genders in transfusion-dependent patients

Role of phytotherapy associated with antibiotic prophylaxis in female patients with recurrent urinary tract infections

Objective: Aim of this study is to evaluate the efficacy of a phytotherapic which includes Solidago, Orthosiphon and Birch extract (Cistimev®) in association with antibiotic prophylaxis in female patients affected by recurrent urinary tract infections (UTIr). Materials and methods: Patients affected by UTIr older than 18 years started a 3-months antibiotic prophylaxis (Prulifloxacin 600 mg, 1 cps/week or Phosphomicyn 1 cachet/week) according to antibiogram after urine culture. The patients were divided in 2 groups: Group A: antibiotic prophylaxis plus phytotherapy (1 cps/die for 3 months) and Group B: antibiotic prophylaxis alone. Results: 164 consecutive patients were studied: 107 were included in group A (mean age 59 ± 17.3 years) and 57 (mean age 61 ± 15.7) in group B. During the treatment period the relapse frequencies between the two groups were not significantly different (p = 0.854): 12/107 (11.21%) patients interrupted the treatment for UTIr in group A and 6/57 (10.52%) in group B. In the long term follow-up the relapse UTI risk was significant different in the two groups with a relapse risk 2.5 greater in group B than in group A (p &lt; 0.0001). Conclusion: Our study demonstrated that in female patients affected by recurrent UTI, the association between antibiotic prophylaxis and of a phytotherapic which includes Solidago, Orthosiphon and Birch extract reduced the number of UTI in the 12 months following the end of prophylaxis and obtained a longer relapsing time, greatly improving the quality of life of the patients