Effects of miRNA-15 and miRNA-16 expression replacement in chronic lymphocytic leukemia: implication for therapy.

Chronic lymphocytic leukemia (CLL) clones are characterized by loss of a critical region in 13q14.3, (del(13)(q14)) involving the microRNA (miRNA) cluster miR-15a and miR-16-1. We have investigated the effects of replacement of miR-15a and miR-16-1. CLL cells transfected with these miRNA mimics exhibited a decrease in cell viability in vitro and impaired capacity for engraftment and growth in NOD/Shi-scid,γcnull (NSG) mice. No synergistic effects were observed when the two miRNA mimics were combined. The phenomena were not restricted to CLL with the del(13)(q14) lesion. Similar effects induced by miRNA mimics were seen in cells with additional chromosomal abnormalities with the exception of certain CLL clones harboring TP53 alterations. Administration of miRNA mimics to NSG mice previously engrafted with CLL clones resulted in substantial tumor regression. CLL cell transfection with miR-15a and miR-16-1-specific inhibitors resulted in increased cell viability in vitro and in an enhanced capacity of the engrafted cells to grow in NSG mice generating larger splenic nodules. These data demonstrate that the strong control by miR-15a and miR-16-1 on CLL clonal expansion is exerted also at the level of full-blown leukemia and provide indications for a miRNA-based therapeutic strategy.

lncRNA profiling in early-stage chronic lymphocytic leukemia identifies transcriptional fingerprints with relevance in clinical outcome.

Long non-coding RNAs (lncRNAs) represent a novel class of functional RNA molecules with an important emerging role in cancer. To elucidate their potential pathogenetic role in chronic lymphocytic leukemia (CLL), a biologically and clinically heterogeneous neoplasia, we investigated lncRNAs expression in a prospective series of 217 early-stage Binet A CLL patients and 26 different subpopulations of normal B-cells, through a custom annotation pipeline of microarray data. Our study identified a 24-lncRNA-signature specifically deregulated in CLL compared with the normal B-cell counterpart. Importantly, this classifier was validated on an independent data set of CLL samples. Belonging to the lncRNA signature characterizing distinct molecular CLL subgroups, we identified lncRNAs recurrently associated with adverse prognostic markers, such as unmutated IGHV status, CD38 expression, 11q and 17p deletions, and NOTCH1 mutations. In addition, correlation analyses predicted a putative lncRNAs interplay with genes and miRNAs expression. Finally, we generated a 2-lncRNA independent risk model, based on lnc-IRF2-3 and lnc-KIAA1755-4 expression, able to distinguish three different prognostic groups in our series of early-stage patients. Overall, our study provides an important resource for future studies on the functions of lncRNAs in CLL, and contributes to the discovery of novel molecular markers with clinical relevance associated with the disease.

Bendamustine in combination with ofatumumab in relapsed or refractory chronic lymphocytic leukemia: a GIMEMA Multicenter Phase II Trial.

We conducted a phase II, noncomparative, open-label, multicenter GIMEMA (Gruppo Italiano Malattie EMatologiche dell'Adulto) study (CLL0809) to assess the efficacy and safety of bendamustine in combination with ofatumumab (BendOfa) in relapsed/refractory chronic lymphocytic leukemia (CLL). Forty-seven patients from 14 centers were evaluated. Therapy consisted of bendamustine (70 mg/m(2)) for 2 consecutive days every 28 days, and ofatumumab 300 mg on day 1 and 1000 mg on day 8 during the first cycle, and 1000 mg on day 1 subsequently. Treatment was administered up to six cycles. The overall response rate (ORR), as per intention-to-treat analysis, was 72.3% (95% confidence of interval (CI), 57-84%), with 17% complete responses. After a median follow-up of 24.2 months, the overall survival was 83.6% (95% CI, 73.0-95.7%) and the progression-free survival (PFS) was 49.6% (95% CI, 35.9-68.6%). The median PFS was 23.6 months. Univariate and multivariate analyses were used to identify clinical and biological characteristics associated with ORR and PFS. Myelosuppression was the most common toxicity; grade ≥3 neutropenia was observed in 61.7% of patients; however, grade ≥3 infections occurred in 6% of patients. BendOfa is feasible and effective in relapsed/refractory CLL patients, including patients with high-risk clinical and biological features.

Rehabilitation needs after bariatric surgery.

BACKGROUND: Bariatric surgery has grown from an obscure experimental procedure to one of the most popular operations in the world. Such accelerated progress left many gaps, notably concerning subsequent rehabilitation needs of this population. AIM: In the present study, a brief description of both the patients and the interventions is provided, along with potentially disabling features especially concerning the locomotor system, which has received comparatively little attention . DESIGN: Based on reported protocols and actual experience, major issues are addressed. SETTING: Bariatric patients are initially managed in the hospital, however long-term and even lifetime needs may be recognized, requiring major lifestyle and physical activity changes. These have to be focused in all settings, inside and outside the healthcare institutions. POPULATION: Initially only adults were considered bariatric candidates, however currently also adolescents and the elderly are admitted in many centers. RESULTS: Bariatric weight loss is certainly successful for remission or prevention of metabolic, cardiovascular and cancer comorbidities. Yet benefits for bones, joints and muscles, along with general physical performance are still incompletely established. This should be no reason for denying continued care to such individuals, within the context of well-designed protocols, as available evidence points toward favorable rehabilitation in the realms of physical, social and workplace activities. CONCLUSION: The importance of a physiatric curriculum in medical schools has been emphasized. Even more crucial is the presence of such a specialists in obesity and bariatric teams, a requirement recognized in a few countries but not in others. CLINICAL REHABILITATION IMPACT: The relevance of obesity as a disabling condition is reviewed, along with the positive changes induced by surgical weight loss. Although obesity alleviation is a legitimate end-point it is not a sufficient one. The shortcomings of such result from the point of view of physical normalization are outlined, and recommendations are suggested.

The expression pattern of small nucleolar and small Cajal body-specific RNAs characterizes distinct molecular subtypes of multiple myeloma.

Small nucleolar RNAs (snoRNAs) and small Cajal body-specific RNAs (scaRNAs) are non-coding RNAs involved in the maturation of other RNA molecules and generally located in the introns of host genes. It is now emerging that altered sno/scaRNAs expression may have a pathological role in cancer. This study elucidates the patterns of sno/scaRNAs expression in multiple myeloma (MM) by profiling purified malignant plasma cells from 55 MMs, 8 secondary plasma cell leukemias (sPCLs) and 4 normal controls. Overall, a global sno/scaRNAs downregulation was found in MMs and, even more, in sPCLs compared with normal plasma cells. Whereas SCARNA22 resulted the only sno/scaRNA characterizing the translocation/cyclin D4 (TC4) MM, TC2 group displayed a distinct sno/scaRNA signature overexpressing members of SNORD115 and SNORD116 families located in a region finely regulated by an imprinting center at 15q11, which, however, resulted overall hypomethylated in MMs independently of the SNORD115 and SNORD116 expression levels. Finally, integrative analyses with available gene expression and genome-wide data revealed the occurrence of significant sno/scaRNAs/host genes co-expression and the putative influence of allelic imbalances on specific snoRNAs expression. Our data extend the current view of sno/scaRNAs deregulation in cancer and add novel information to the bio-molecular complexity of plasma cell dyscrasias.

Importance of semicore states in GW calculations for simulating accurately the photoemission spectra of metal phthalocyanine molecules.

The quasi-particle energy levels of the Zn-Phthalocyanine (ZnPc) molecule calculated with the GW approximation are shown to depend sensitively on the explicit description of the metal-center semicore states. We find that the calculated GW energy levels are in good agreement with the measured experimental photoemission spectra only when explicitly including the Zn 3s and 3p semicore states in the valence. The main origin of this effect is traced back to the exchange term in the self-energy GW approximation. Based on this finding, we propose a simplified approach for correcting GW calculations of metal phthalocyanine molecules that avoids the time-consuming explicit treatment of the metal semicore states. Our method allows for speeding up the calculations without compromising the accuracy of the computed spectra.

Becoming a nurse in Italy: a multi-method study on expenditures by families and students.

Potential nursing students and their families are faced with difficult decisions regarding the amount of time and money required to complete the nursing programme and the availability of funds to cover the costs and this seems to have received little no attention to date. With the aim of describing the costs incurred by Italian nursing students and/or their families per academic year and compare cost trends incurred from 2004-05 to 2010-11, a multi-centre qualitative/quantitative study design was adopted. Italian Nursing students attending the first, second and third academic years in 2004-05 and those attending the first, second and third academic years in 2010-11 were eligible. Five hundred and six students were involved: 215 (out of 300 eligible, 71.6%) attended the bachelor's degree in nursing in 2004-05 and 291 (out of 383 eligible, 75.9%) in 2010-11. On an annual basis, the average annual expenditures increased by 12% for nursing education from 2004-05 to 2010-11. Given that qualification as a nurse requires at least three years, and considering inflation, for a student who matriculated in 2005 an average of 2485.7€ per year (7457.0€ in total) was required. Data suggest that students have modified their spending behaviour (limiting lunches at public bars, buying books and photocopies) in order to handle the rise of non-discretionary costs, such as tuition fees and the costs of attending lectures and hospital/district trainings. Policies supporting nursing education in general and for those students who are motivated but unable to undertake the course for economic reasons are urgently needed.

The cost of the objective structured clinical examination on an Italian nursing bachelor's degree course.

The OSCE (Objective Structured Clinical Examination) is considered the most valid and reliable method for assessing the clinical skills of students training for health professions, but its use is limited by the related high costs. We analyzed the cost retrospectively of using an OSCE designed for second-year students (2009) in our degree course, adopting the Reznick et al. guidelines (1993), which recommend assessing both high-end costs and low-end costs. The high-end costs adopting the OSCE amounted to € 145.23 per student, while the low-end costs were € 31.51 per student. Considering the economic crisis and the cost-containment measures applied also in nursing education, strategies for further reducing costs are discussed.

[Assessing quality in clinical educational setting: Italian validation of the clinical learning environment and supervision (CLES) scale]. / La valutazione dellA qualità degli ambienti di apprendimento clinico: studio di validazione del Clinical Learning Environment and Supervision (CLES) nel contesto Italiano.

The clinical learning in the wards is one of the most important moment in nursing education: Instruments assessing the quality of the clinical settings as perceived by students may inform and support the best didactic and tutorial choices. In the last 10 years, three clinical learning environment assessment tools for Bachelor Nursing students has been validated. The aim of this study was to validate in the Italian context the Clinical Learning Environment and Supervision scale (CLES) developed in Finland and considered the gold standard within the instruments available. CLES's face and content validity has been evaluated with forward and backward translation and involving a selected group of experts in the field. 117 Bachelor Nursing Students from Udine and Verona University have been enrolled in the study. 28 out of 117 students have filed the instrument twice in order to measure test-retest reliability. The internal consistency has been evaluated with Cronbach alpha. Kurtosis and skewness was measured in order to verify the answers variability distribution. The CLES Cronbach' alpha was 0.96 and the test-retest reliability was r = -0.89. The validity and reliability of the scale has been confirmed. Further research is needed to define a cut-off in order to recognize positive clinical learning environments. The CLES practical use could support in the process of the teaching hospital accreditation and in the evaluation of the effectiveness of the tutorial strategies activated in the wards.

Three-dimensional tomography of the beryllium fermi surface: surface charge redistribution.

The discontinuity in the lattice periodic potential at surfaces often leads to the creation of new electronic surface states. We developed a photoemission based Fermi surface tomography whose surface sensitivity allowed us to quantify the charge redistribution on the Be(0001) surface. The volume enclosed by the bulklike Fermi surface is significantly reduced at the surface, consistent with the charge transfer to the two surface states as estimated from the area within their two-dimensional Fermi contours. This result represents the first quantification of the charge redistribution on a natural surface termination.

An update on xenotransplantation.

Xenotransplantation is one of the possible avenues currently being explored to address the shortage problem of human organs. With this in mind, this article will briefly review the current situation with respect to the immunological, physiological and biosafety aspects related to the transplantation of pig organs into primates. Acute humoral xenograft rejection (AHXR) currently remains the central immunological obstacle and the development of strategies for both a better control of the elicited anti-pig humoral immune response or the prevention of the onset of coagulation disorders that accompany AHXR are the two primary focuses of research. To date, porcine xenografts have been shown to sustain the life of nonhuman primates for several months. Such preclinical studies have also demonstrated the absence of insurmountable physiological incompatibilities between pig and primate. In addition, reassuring findings regarding biosafety aspects have been generated and pro-active research aimed at the identification of an organ source with a higher safety profile is also underway. These advancements, in conjunction with ongoing research in pig genetic engineering, immunosuppression and tolerance are expected to further extend the survival of porcine xenografts transplanted into primates. However, until further physiological, efficacy and safety data are generated in relevant primate models, clinical xenotransplantation should not be considered.

Polyketone polymer: a new support for direct enzyme immobilization.

Polyketone polymer -[-CO-CH(2)-CH(2)-](n)-, obtained by copolymerization of ethene and carbon monoxide, is utilized for immobilization of three different enzymes, one peroxidase from horseradish (HRP) and two amine oxidases, from bovine serum (BSAO) and lentil seedlings (LSAO). The easy immobilization procedure is carried out in diluted buffer, at pH 7.0 and 3 degrees C, gently mixing the proteins with the polymer. No bifunctional reagents and spacer arms are required for the immobilization, which occurs exclusively via a large number of hydrogen bonds between the carbonyl groups of the polymer and the -NH groups of the polypeptidic chain. Experiments demonstrate a high linking capacity of polymer for BSAO and an extraordinary strong linkage for LSAO. Moreover, activity measurements demonstrate that immobilized LSAO totally retains the catalytic characteristics of the free enzyme, where only a limited increase of K(M) value is observed. Finally, the HRP-activated polymer is successfully used as active packed bed of an enzymatic reactor for continuous flow conversion and flow injection analysis of hydrogen peroxide containing solutions.

Construction of a GP integration model.

There are frequent calls to improve integration of health services, within and between primary and secondary care sectors. In Australia, general medical practitioners (GPs) are central to these endeavours. This paper aims to better conceptualise GP integration and to develop a model and index based on this. A conceptualisation of integration is proposed based on integration fundamentally as an activity or process not structure. Integration process is the frequency and quality of episodes of information exchange involving the GP and another practitioner or patient and aimed at fulfilling the objectives of the health care system with regard to patient care. These are both direct responses to structural forces and emergent GP capacities and dispositions. The content of this typology was studied using Concept Mapping in 11 groups of GPs, consumers and other practitioners. Clusters of related statements within thematic domains were used as the basis for a provisional model. This was tested using confirmatory factor analysis in a data set derived from a national probability sample of 501 GPs. Some re-specification of the model was necessary, with three integration process factors needing to be subdivided. One factor congeneric model assumptions were used to identify the constituent items for these factors. The result was a model in which 50 items measured nine integration process factors and 20 items measured five enabling factors. Two distinct but correlated higher order factors, relating to individual patient care and public (or community) health--in contrast to a single higher order factor for integration--were identified. The re-specified model was tested with a new sample of 151 GPs and exhibited strong psychometric properties. Reliability and validity were acceptable to this stage of the indices' development. Further testing of the index is necessary to demonstrate factor invariance of the indices in other contexts as well as their utility in cross-structural analysis. That said, the indices have immediate uses.

Analysis of FGFR3 gene mutations in multiple myeloma patients with t(4;14).

The t(4;14)(p16.3;q32) in multiple myeloma (MM) leads to an apparent deregulation of the FGFR3 and WHSC1/MMSET genes. FGFR3 mutations, known to be associated with genetic skeletal disorders, have also been identified in a few cases of MM (mainly cell lines) with t(4;14). We investigated FGFR3 mutations in a series of 53 MM cases; 11 cases with t(4;14) and FGFR3 overexpression were analysed using reverse transcription polymerase chain reaction, while the remaining cases were studied at DNA level. The Arg248Cys mutation, which is associated with some lethal forms of skeletal disorders, was found in one case with t(4;14). Our results indicate that FGFR3 mutations occur in only a small fraction of MM cases with t(4;14).

Interaction of linear mono- and diamines with dimyristoylphosphatidylcholine and dimyristoylphosphatidylglycerol multilamellar liposomes.

The effect of linear monoamines on dimyristoylphosphatidylglycerol and dimyristoylphosphatidylcholine multilamellar liposomes was studied as a function of their length and compared with the behavior of linear carboxylic acids. The role of the hydrophobic interactions was demonstrated and the free energy of the binding for each interacting carbon atom was determined. The thermotropic behavior of the liposomes was characterized by differential scanning calorimetry and it was shown that these molecules affect the temperature and the cooperativity of the gel to fluid state transition of the membrane differently. In particular, it appeared that membrane perturbation was maximum when the chain length of the amphipathic molecules ranged between 7 and 9 carbon atoms, with more pronounced effects in the case of monoamines. Molecules shorter than 3-4 carbon atoms did not produce any observable change in the transition temperature. The study was extended to linear alpha,omega-diamines to investigate the amphipathic character of long diamines and to investigate the role of bridging bonds established with neighboring phospholipids.

Detection of t(4;14)(p16.3;q32) chromosomal translocation in multiple myeloma by reverse transcription-polymerase chain reaction analysis of IGH-MMSET fusion transcripts.

We and others have recently identified a novel recurring t(4;14)(p16.3; q32) translocation in multiple myeloma (MM) that leads to an apparent deregulation of the FGFR3 and WHSC1/MMSET genes. Because the presence of IGH-MMSET hybrid transcripts has been found in MM cell lines with t(4;14), they may represent a specific tumor-associated marker in MM. In this study, we developed a reverse transcription-PCR (RTPCR) assay for detecting chimeric transcripts from all of the 4p16.3 breakpoints identified thus far, and we used it to investigate a representative panel of 53 MM patients and 16 patients with monoclonal gammopathy of uncertain significance; in addition, t(4;14) was investigated in all of the MM patients by means of two-color fluorescence in situ hybridization. IGH-MMSET transcripts were found in 11 of the 53 (20%) MM cases and 1 of 16 (6%) cases of monoclonal gammopathy of uncertain significance. There was complete concordance between the RT-PCR and fluorescence in situ hybridization analyses of the MM cases. The results of this study indicate that RT-PCR is a sensitive and reliable method of detecting t(4;14) and suggest that it may be useful for monitoring the disease in a significant proportion of patients.

Immunohistochemical analysis of cyclin D1 shows deregulated expression in multiple myeloma with the t(11;14).

The t(11;14)(q13;q32) chromosomal translocation, the hallmark of mantle cell lymphoma (MCL), is recurrently found in multiple myelomas (MM) by means of conventional cytogenetics. Unlike MCL, recent molecular studies of MM-derived cell lines with t(11;14) have indicated that the breakpoints are highly dispersed over the 11q13 region; however, the fact that cyclin D1 is generally overexpressed in these cell lines suggests that this gene is the target of the translocation. To evaluate further the involvement of cyclin D1 in MM, we used immunohistochemistry and fluorescence in situ hybridization to investigate cyclin D1 expression and the presence of chromosome 11 abnormalities in a representative panel of 48 MM patients (40 at diagnosis and 8 at relapse). Cyclin D1 overexpression occurred in 12/48 (25%) of cases; combined immunohistochemistry and fluorescence in situ hybridization analyses in 39 patients showed cyclin D1 positivity in all of the cases (7/7) bearing the t(11;14), in two of the 13 cases with trisomy 11, and in one of the 19 cases with no apparent abnormalities of chromosome 11. Our data indicate that the t(11;14) translocation in MM leads to cyclin D1 overexpression and that immunohistochemical analysis may represent a reliable means of identifying this lesion in MM.