Formation of a Unique Cluster of G-Quadruplex Structures in the HIV-1 nef Coding Region: Implications for Antiviral Activity

G-quadruplexes are tetraplex structures of nucleic acids that can form in G-rich sequences. Their presence and functional role have been established in telomeres, oncogene promoters and coding regions of the human chromosome. In particular, they have been proposed to be directly involved in gene regulation at the level of transcription. Because the HIV-1 Nef protein is a fundamental factor for efficient viral replication, infectivity and pathogenesis in vitro and in vivo, we investigated G-quadruplex formation in the HIV-1 nef gene to assess the potential for viral inhibition through G-quadruplex stabilization. A comprehensive computational analysis of the nef coding region of available strains showed the presence of three conserved sequences that were uniquely clustered. Biophysical testing proved that G-quadruplex conformations were efficiently stabilized or induced by G-quadruplex ligands in all three sequences. Upon incubation with a G-quadruplex ligand, Nef expression was reduced in a reporter gene assay and Nef-dependent enhancement of HIV-1 infectivity was significantly repressed in an antiviral assay. These data constitute the first evidence of the possibility to regulate HIV-1 gene expression and infectivity through G-quadruplex targeting and therefore open a new avenue for viral treatment. © 2013 Perrone et al

Surgical management of complex ileocolonic Crohn’s disease: a survey of IBD colorectal surgeons to assess variability in operative strategy

Cirurgia colorectal; Malaltia de Crohn; Resecció ileocecalCirugía colorrectal; Enfermedad de Crohn; Resección ileocecalColorectal surgery; Crohn’s disease; Ileocaecal resectionIntroduction To explore the reported variability in the surgical management of ileocolonic Crohn’ s disease and identify areas of standard practice, we present this study which aims to assess how different colorectal surgeons with a subspecialty interest in inflammatory bowel disease (IBD) surgery may act in different clinical scenarios of ileocolonic Crohn’s disease. Methods Anonymous videos demonstrating the small bowel walkthrough and anonymised patients’ clinical data, imaging and pathological findings were distributed to the surgeons using an electronic tool. Surgeons answered on operative strategy, bowel resections, management of small bowel mesentery, type of anastomosis and use of stomas. Results Eight small bowel walkthrough videos were registered and 12 assessors completed the survey with a questionnaire completion rate of 87.5%. There was 87.7% agreement in the need to perform an ileocolonic resection. However, the agreement for the need to perform associated surgical procedures such as strictureplasties or further bowel resections was only 57.4%. When an anastomosis was fashioned, the side to side configuration was the most commonly used. The preferred management of the mesentery was dissection close to the bowel. Conclusions The decision on the main procedure to be performed had a high agreement amongst the different assessors, but the treatment of multifocal disease was highly controversial, with low agreement on the need for associated procedures to treat internal fistulae and the use of strictureplasties. At the same time, there was significant heterogeneity in the decision on when to anastomose and when to fashion an ileostomy

An immunohistochemically positive E-cadherin status is not always predictive for a good prognosis in human breast cancer

BACKGROUND: in primary breast cancers dichotomic classification of E-cadherin expression, according to an arbitrary cutoff, may be inadequate and lead to loss of prognostic significance or contrasting prognostic indications. We aimed to assess the prognostic value of high and low E-cadherin levels in a consecutive case series (204 cases) of unilateral node-negative non-lobular breast cancer patients with a 8-year median follow-up and that did not receive any adjuvant therapy after surgery. METHODS: expression of E-cadherin was investigated by immunohistochemistry and assessed according to conventional score (0, 1+, 2+, 3+). Multiple correspondence analysis was used to visualise associations of both categorical and continuous variables. The impact of E-cadherin expression on patients outcome was evaluated in terms of event-free survival curves by the Kaplan-Meier method and proportional hazard Cox model. RESULTS: respect to intermediate E-cadherin expression values (2+), high (3+) or low (0 to 1+) E-cadherin expression levels had a negative prognostic impact. In fact, both patients with a low-to-nil (score 0 to 1+) expression level of E-cadherin and patients with a high E-cadherin expression level (score 3+) demonstrated an increased risk of failure (respectively, hazard ratio (HR)=1.71, confidence interval (CI)=0.72-4.06 and HR=4.22, CI=1.406-12.66) and an interesting association with young age. CONCLUSIONS: the findings support the evidence that high expression values of E-cadherin are not predictive for a good prognosis and may help to explain conflicting evidence on the prognostic impact of E-cadherin in breast cancer when assessed on dichotomic basis

Epidemiology and outcome of Clostridium difficile infections in patients hospitalized in Internal Medicine: findings from the nationwide FADOI-PRACTICE study.

BACKGROUND: Clostridium difficile (CD) is a leading cause of diarrhoea among hospitalized patients. The objective of this study was to evaluate the rate, the optimal diagnostic work-up, and outcome of CD infections (CDI) in Internal Medicine (IM) wards in Italy. METHODS: PRACTICE is an observational prospective study, involving 40 IM Units and evaluating all consecutive patients hospitalized during a 4-month period. CDI were defined in case of diarrhoea when both enzyme immunoassay for GDH, and test for A/B toxin were positive. Patients with CDI were followed-up for recurrences for 4 weeks after the end of therapy. RESULTS: Among the 10,780 patients observed, 103 (0.96 %) showed CDI, at admission or during hospitalization. A positive history for CD, antibiotics in the previous 4 weeks, recent hospitalization, female gender and age were significantly associated with CDI (multivariable analysis). In-hospital mortality was 16.5 % in CD group vs 6.7 % in No-CD group (p\u2009<\u20090.001), whereas median length of hospital stay was 16 (IQR\u2009=\u200913) vs 8 (IQR\u2009=\u20098) days (p\u2009<\u20090.001) among patients with or without CDI, respectively. Rate of CD recurrences was 14.6 %. As a post-hoc evaluation, 23 out of 34 GDH+/Tox- samples were toxin positive, when analysed by molecular method (a real-time PCR assay). The overall CD incidence rate was 5.3/10,000 patient-days. CONCLUSIONS: Our results confirm the severity of CDI in medical wards, showing high in-hospital mortality, prolonged hospitalization and frequent short-term recurrences. Further, our survey supports a 2-3 step algorithm for CD diagnosis: EIA for detecting GDH, A and B toxin, followed by a molecular method in case of toxin-negative samples

A video guide of five access methods to the splenic flexure: the concept of the splenic flexure box

Aim: The aim of this study was to describe all the possible approaches for laparoscopic splenic flexure mobilization (SFM), each suitable for specific situations, and create an illustrated system to show SFM approaches in an easy and practical way to make it easy to learn and teach. Methods: Two different phases. First part: Cadaver-based study of the colonic splenic flexure anatomy. In order to demonstrate the different approaches, a balloon was placed through the colonic hepatic flexure in the lesser sac without sectioning any of the fixing ligaments of the splenic flexure. Second part: A real case series of laparoscopic SFM. Results: First part: 11 cadavers were dissected. Five potential approaches to SFM were found: anterior, trans-omentum, lateral, medial infra-mesocolic, and medial trans-mesocolic. The illustrative system developed was named: Splenic Flexure “Box”(SFBox). Second part: One of the types of SFM described in first part was used in five patients with colorectal cancer. Each laparoscopic approach to the splenic flexure was illustrated in a video accompanied by illustration aids delineating the access. Conclusion: With the cadaver dissection and subsequent demonstration in real-life laparoscopic surgery, we have shown five types of laparoscopic splenic flexure mobilization. The Splenic Flexure “Box” is a useful way to learn and teach this surgical maneuver

Apraxia and motor dysfunction in corticobasal syndrome

Background: Corticobasal syndrome (CBS) is characterized by multifaceted motor system dysfunction and cognitive disturbance; distinctive clinical features include limb apraxia and visuospatial dysfunction. Transcranial magnetic stimulation (TMS) has been used to study motor system dysfunction in CBS, but the relationship of TMS parameters to clinical features has not been studied. The present study explored several hypotheses; firstly, that limb apraxia may be partly due to visuospatial impairment in CBS. Secondly, that motor system dysfunction can be demonstrated in CBS, using threshold-tracking TMS, and is linked to limb apraxia. Finally, that atrophy of the primary motor cortex, studied using voxel-based morphometry analysis (VBM), is associated with motor system dysfunction and limb apraxia in CBS.   Methods: Imitation of meaningful and meaningless hand gestures was graded to assess limb apraxia, while cognitive performance was assessed using the Addenbrooke's Cognitive Examination - Revised (ACE-R), with particular emphasis placed on the visuospatial subtask. Patients underwent TMS, to assess cortical function, and VBM.   Results: In total, 17 patients with CBS (7 male, 10 female; mean age 64.4+/2 6.6 years) were studied and compared to 17 matched control subjects. Of the CBS patients, 23.5% had a relatively inexcitable motor cortex, with evidence of cortical dysfunction in the remaining 76.5% patients. Reduced resting motor threshold, and visuospatial performance, correlated with limb apraxia. Patients with a resting motor threshold <50% performed significantly worse on the visuospatial sub-task of the ACE-R than other CBS patients. Cortical function correlated with atrophy of the primary and pre-motor cortices, and the thalamus, while apraxia correlated with atrophy of the pre-motor and parietal cortices.   Conclusions: Cortical dysfunction appears to underlie the core clinical features of CBS, and is associated with atrophy of the primary motor and pre-motor cortices, as well as the thalamus, while apraxia correlates with pre-motor and parietal atrophy