Evidence for the in vivo safety of insulated foamy viral vectors.

Retroviral vector-mediated stem cell gene therapy is a promising approach for the treatment of hematopoietic disorders. However, genotoxic side effects from integrated vector proviruses are a significant concern for the use of retroviral vectors in the clinic. Insulated foamy viral (FV) vectors are potentially safer retroviral vectors for hematopoietic stem cell gene therapy. We evaluated two newly identified human insulators, A1 and A2, for use in FV vectors. These insulators had moderate insulating capacity and higher titers than previously developed insulated FV vectors. The A1-insulated FV vector was chosen for comparison with the previously described 650cHS4-insulated FV vector in human cord blood CD34+ repopulating cells in an immunodeficient mouse model. To maximize the effects of the insulators on the safety of FV vectors, FV vectors containing a highly genotoxic spleen focus forming virus promoter were used to elicit differences in genotoxicity. In vivo, the A1-insulated FV vector showed an approximate 50% reduction in clonal dominance compared with either the 650cHS4-insulated or control FV vectors, although the transduction efficiency of the A1-insulated vector was higher. This data suggests that the A1-insulated FV vector is promising for future preclinical and clinical studies.

An essential role of Ffar2 (Gpr43) in dietary fibre-mediated promotion of healthy composition of gut microbiota and suppression of intestinal carcinogenesis.

Composition of the gut microbiota has profound effects on intestinal carcinogenesis. Diet and host genetics play critical roles in shaping the composition of gut microbiota. Whether diet and host genes interact with each other to bring specific changes in gut microbiota that affect intestinal carcinogenesis is unknown. Ability of dietary fibre to specifically increase beneficial gut microbiota at the expense of pathogenic bacteria in vivo via unknown mechanism is an important process that suppresses intestinal inflammation and carcinogenesis. Free fatty acid receptor 2 (FFAR2 or GPR43) is a receptor for short-chain fatty acids (acetate, propionate and butyrate), metabolites of dietary fibre fermentation by gut microbiota. Here, we show FFAR2 is down modulated in human colon cancers than matched adjacent healthy tissue. Consistent with this, Ffar2(-/-) mice are hypersusceptible to development of intestinal carcinogenesis. Dietary fibre suppressed colon carcinogenesis in an Ffar2-dependent manner. Ffar2 played an essential role in dietary fibre-mediated promotion of beneficial gut microbiota, Bifidobacterium species (spp) and suppression of Helicobacter hepaticus and Prevotellaceae. Moreover, numbers of Bifidobacterium is reduced, whereas those of Prevotellaceae are increased in human colon cancers than matched adjacent normal tissue. Administration of Bifidobacterium mitigated intestinal inflammation and carcinogenesis in Ffar2(-/-) mice. Taken together, these findings suggest that interplay between dietary fibre and Ffar2 play a key role in promoting healthy composition of gut microbiota that stimulates intestinal health.

Systematic review and critical appraisal of the impact of acellular dermal matrix use on the outcomes of implant-based breast reconstruction.

BACKGROUND: Acellular dermal matrix (ADM) may improve outcomes in implant-based breast reconstruction (IBBR). The aim of this study was critically to appraise and evaluate the current evidence for ADM-assisted IBBR. METHODS: Comprehensive electronic searches identified complete papers published in English between January 2000 and August 2013, reporting any outcome of ADM-assisted IBBR. All systematic reviews, randomized clinical trials (RCTs) and non-randomized studies (NRSs) with more than 20 ADM recipients were included. Studies were critically appraised using AMSTAR for systematic reviews, the Cochrane risk-of-bias tool for RCTs and its adaptation for NRSs. Characteristics and results of identified studies were summarized. RESULTS: A total of 69 papers (8 systematic reviews, 1 RCT, 40 comparative studies and 20 case series) were identified, all of which were considered at high risk of bias, mostly due to patient selection and selective outcome reporting. The median ADM group sample size was 51.0 (i.q.r. 33.0-127.0). Most studies were single-centre (54), and they were often single-surgeon (16). ADM was most commonly used for immediate (40) two-stage IBBR (36) using human ADM (47), with few studies evaluating ADM-assisted single-stage procedures (10). All reported clinical outcomes (for example implant loss) and more than half of the papers (33) assessed process outcomes, but few evaluated cosmesis (16) or patient-reported outcomes (10). Heterogeneity between study design and, especially, outcome measurement precluded meaningful data synthesis. CONCLUSION: Current evidence for the value of ADMs in IBBR is limited. Use in practice should therefore be considered experimental, and evaluation within registries or well designed and conducted studies, ideally RCTs, is recommended to prevent widespread adoption of a potentially inferior intervention.

Invasive Propionibacterium acnes infections in a non-selective patient cohort: clinical manifestations, management and outcome.

An increasing number of reports suggest that Propionibacterium acnes can cause serious invasive infections. Currently, only limited data exist regarding the spectrum of invasive P. acnes infections. We conducted a non-selective cohort study at a tertiary hospital in the UK over a 9-year-period (2003-2012) investigating clinical manifestations, risk factors, management, and outcome of invasive P. acnes infections. Forty-nine cases were identified; the majority were neurosurgical infections and orthopaedic infections (n = 28 and n = 15 respectively). Only 2 cases had no predisposing factors; all neurosurgical and 93.3 % of orthopaedic cases had a history of previous surgery and/or trauma. Foreign material was in situ at the infection site in 59.3 % and 80.0 % of neurosurgical and orthopaedic cases respectively. All neurosurgical and orthopaedic cases required one or more surgical interventions to treat P. acnes infection, with or without concomitant antibiotic therapy; the duration of antibiotic therapy was significantly longer in the group of orthopaedic cases (median 53 vs 19 days; p = 0.0025). All tested P. acnes isolates were susceptible to penicillin, ampicillin and chloramphenicol; only 1 was clindamycin-resistant. Neurosurgical and orthopaedic infections account for the majority of invasive P. acnes infections. Most cases have predisposing factors, including previous surgery and/or trauma; spontaneous infections are rare. Foreign material is commonly present at the site of infection, indicating that the pathogenesis of invasive P. acnes infections likely involves biofilm formation. Since invasive P. acnes infections are associated with considerable morbidity, further studies are needed to establish effective prevention and optimal treatment strategies.

Type 2 cGMP-dependent protein kinase regulates homeostasis by blocking c-Jun N-terminal kinase in the colon epithelium.

Analysis of knockout animals indicates that 3',5'cyclic guanosine monophosphate (cGMP) has an important role in gut homeostasis but the signaling mechanism is not known. The goals of this study were to test whether increasing cGMP could affect colon homeostasis and determine the mechanism. We increased cGMP in the gut of Prkg2(+/+) and Prkg2(-/-) mice by treating with the PDE5 inhibitor Vardenafil (IP). Proliferation, differentiation and apoptosis in the colon mucosa were then quantitated. Vardenafil (Vard) treatment increased cGMP in colon mucosa of all mice, but reduced proliferation and apoptosis, and increased differentiation only in Prkg2(+/+) mice. Vard and cGMP treatment also increased dual specificity protein phosphatase 10 (DUSP10) expression and reduced phospho-c-Jun N-terminal kinase (JNK) levels in the colon mucosa of Prkg2(+/+) but not Prkg2(-/-) mice. Treatment of Prkg2(-/-) mice with the JNK inhibitor SP600125 reversed the defective homeostasis observed in these animals. Activation of protein kinase G2 (PKG2) in goblet-like LS174T cells increased DUSP10 expression and reduced JNK activity. PKG2 also increased goblet cell-specific MUC2 expression in LS174T cells, and this process was blocked by DUSP10-specific siRNA. The ability of cGMP signaling to inhibit JNK-induced apoptosis in vivo was demonstrated using dextran sodium sulfate (DSS) to stress the colon epithelium. Vard was a potent inhibitor of DSS-induced epithelial apoptosis, and significantly blocked pathological endpoints in this model of experimental colitis. In conclusion, Vard treatment activates cGMP signaling in the colon epithelium. Increased PKG2 activity alters homeostasis by suppressing proliferation and apoptosis while promoting differentiation. The PKG2-dependent mechanism was shown to involve increased DUSP10 and subsequent inhibition of JNK activity.

X-ray streak camera cathode development and timing accuracy of the 4ω ultraviolet fiducial system at the National Ignition Facility.

The convergent ablator experiments at the National Ignition Facility (NIF) are designed to measure the peak velocity and remaining ablator mass of an indirectly driven imploding capsule. Such a measurement can be performed using an x-ray source to backlight the capsule and an x-ray streak camera to record the capsule as it implodes. The ultimate goal of this experiment is to achieve an accuracy of 2% in the velocity measurement, which translates to a ±2 ps temporal accuracy over any 300 ps interval for the streak camera. In order to achieve this, a 4ω (263 nm) temporal fiducial system has been implemented for the x-ray streak camera at NIF. Aluminum, titanium, gold, and silver photocathode materials have been tested. Aluminum showed the highest relative quantum efficiency, with five times more peak signal counts per fiducial pulse when compared to Gold. The fiducial pulse data were analyzed to determine the centroiding statistical accuracy for incident laser pulse energies of 1 and 10 nJ, showing an accuracy of ±1.6 ps and ±0.7 ps, respectively.

High performance imaging streak camera for the National Ignition Facility.

An x-ray streak camera platform has been characterized and implemented for use at the National Ignition Facility. The camera has been modified to meet the experiment requirements of the National Ignition Campaign and to perform reliably in conditions that produce high electromagnetic interference. A train of temporal ultra-violet timing markers has been added to the diagnostic in order to calibrate the temporal axis of the instrument and the detector efficiency of the streak camera was improved by using a CsI photocathode. The performance of the streak camera has been characterized and is summarized in this paper. The detector efficiency and cathode measurements are also presented.

Demonstration of ignition radiation temperatures in indirect-drive inertial confinement fusion hohlraums.

We demonstrate the hohlraum radiation temperature and symmetry required for ignition-scale inertial confinement fusion capsule implosions. Cryogenic gas-filled hohlraums with 2.2 mm-diameter capsules are heated with unprecedented laser energies of 1.2 MJ delivered by 192 ultraviolet laser beams on the National Ignition Facility. Laser backscatter measurements show that these hohlraums absorb 87% to 91% of the incident laser power resulting in peak radiation temperatures of T(RAD)=300 eV and a symmetric implosion to a 100 µm diameter hot core.

An interdisciplinary, family-focused approach to relational learning in neonatal intensive care.

OBJECTIVE: The aim of this study is to show the efficacy of the Program to Enhance Relational and Communication Skills-Neonatal Intensive Care Unit (PERCS-NICU). STUDY DESIGN: In this study, 74 practitioners attended workshops and completed baseline, post-training and follow-up questionnaires. RESULT: On yes/no questions, 93 to 100% reported improved preparation, communication skills and confidence post-training and follow-up. A total of 94 and 83% improved their ability to establish relationships, and 76 and 83% reported reduced anxiety post-training and follow-up, respectively. On Likert items, 59 and 64% improved preparation, 45 and 60% improved communication skills and confidence, 25 and 53% decreased anxiety and 16 and 32% improved relationships post-training and follow-up, respectively. Qualitative themes included integrating new communication and relational abilities, honoring the family perspective, appreciating interdisciplinary collaboration, personal/human connection and valuing the learning. In total, 93% applied skills learned, three-quarters transformed practice and 100% recommended PERCS-NICU. CONCLUSION: After PERCS-NICU, clinicians improved preparation, communication and relational abilities, confidence and reduced anxiety when holding difficult neonatal conversations.

PKG inhibits TCF signaling in colon cancer cells by blocking beta-catenin expression and activating FOXO4.

Activation of cGMP-dependent protein kinase (PKG) has anti-tumor effects in colon cancer cells but the mechanisms are not fully understood. This study has examined the regulation of beta-catenin/TCF signaling, as this pathway has been highlighted as central to the anti-tumor effects of PKG. We show that PKG activation in SW620 cells results in reduced beta-catenin expression and a dramatic inhibition of TCF-dependent transcription. PKG did not affect protein stability, nor did it increase phosphorylation of the amino-terminal Ser33/37/Thr41 residues that are known to target beta-catenin for degradation. However, we found that PKG potently inhibited transcription from a luciferase reporter driven by the human CTNNB1 promoter, and this corresponded to reduced beta-catenin mRNA levels. Although PKG was able to inhibit transcription from both the CTNNB1 and TCF reporters, the effect on protein levels was less consistent. Ectopic PKG had a marginal effect on beta-catenin protein levels in SW480 and HCT116 but was able to inhibit TCF-reporter activity by over 80%. Investigation of alternative mechanisms revealed that cJun-N-terminal kinase (JNK) activation was required for the PKG-dependent regulation of TCF activity. PKG activation caused beta-catenin to bind to FOXO4 in colon cancer cells, and this required JNK. Activation of PKG was also found to increase the nuclear content of FOXO4 and increase the expression of the FOXO target genes MnSOD and catalase. FOXO4 activation was required for the inhibition of TCF activity as FOXO4-specific short-interfering RNA completely blocked the inhibitory effect of PKG. These data illustrate a dual-inhibitory effect of PKG on TCF activity in colon cancer cells that involves reduced expression of beta-catenin at the transcriptional level, and also beta-catenin sequestration by FOXO4 activation.

Analysis of enacted difficult conversations in neonatal intensive care.

OBJECTIVE: To analyze the communicative contributions of interdisciplinary professionals and family members in enacted difficult conversations in neonatal intensive care. STUDY DESIGN: Physicians, nurses, social workers, and chaplains (n=50) who attended the Program to Enhance Relational and Communication Skills, participated in a scenario of a preterm infant with severe complications enacted by actors portraying family members. Twenty-four family meetings were videotaped and analyzed with the Roter Interaction Analysis System (RIAS). RESULT: Practitioners talked more than actor-family members (70 vs 30%). Physicians provided more biomedical information than psychosocial professionals (P<0.001), and less psychosocial information than nurses, and social workers and chaplains (P<0.05; P<0.001). Social workers and chaplains asked more psychosocial questions than physicians and nurses (MD=P<0.005; RN=P<0.05), focused more on family's opinion and understanding (MD=P<0.01; RN=P<0.001), and more frequently expressed agreement and approval than physicians (P<0.05). No differences were found across disciplines in providing emotional support. CONCLUSION: Findings suggest the importance of an interdisciplinary approach and highlight areas for improvement such as using silence, asking psychosocial questions and eliciting family perspectives that are associated with family satisfaction.

An exploratory study of the safety, tolerability and bioactivity of a single intravitreal injection of vascular endothelial growth factor Trap-Eye in patients with diabetic macular oedema.

AIM: The aim of the study was to assess the safety and bioactivity of a single intravitreal injection of vascular endothelial growth factor (VEGF) Trap-Eye in subjects with diabetic macular oedema (DMO). METHODS: Five subjects with DMO, foveal thickness > or =250 microm measured by optical coherence tomography (OCT), and best-corrected visual acuity (BCVA) between 20/40 and 20/320, were enrolled. Each participant received a single intravitreal injection of 4.0 mg of VEGF Trap-Eye followed by a 6-week observation period. Outcome measures included safety and biological activity, including changes in BCVA and excess retinal thickness assessed by OCT. RESULTS: Injections of VEGF Trap-Eye were well tolerated with no ocular toxicity. One patient had an unrelated serious adverse event: hospitalisation for cellulitis of the left foot 27 days after injection of VEGF Trap-Eye. Median baseline BCVA was 36 ETDRS letters read at 4 m (not ETDRS visual acuity score; Snellen equivalent: 20/50) and median baseline excess central 1 mm foveal thickness (FTH) was 108 microm. At 4 weeks after injection, the median excess FTH was 59 microm and the median improvement in BCVA was nine letters. At 6 weeks after injection, four of the five patients showed improvement in excess FTH (median 74 microm; 31% reduction from baseline, p = 0.0625) and four of the five showed improvement in BCVA (median improvement of three letters). CONCLUSIONS: A single intravitreal injection of 4.0 mg of VEGF Trap-Eye was well tolerated and preliminary evidence of bioactivity was detected. These findings support additional studies investigating multiple injections of VEGF Trap-Eye in patients with DMO.

Robotic pyeloplasty using internet protocol and satellite network-based telesurgery.

BACKGROUND: In North America, the urological community has embraced surgical robotic technology in the performance of complex laparoscopic surgery. The performance of complex long-distance telesurgery requires further investigation prior to clinical application. METHODS: The feasibility of laparoscopic robot-assisted pyeloplasty in a porcine model was assessed using the Zeus robot and the internet protocol virtual private network (IP-VPNe) and satellite links. Eighteen pyeloplasty procedures were performed, using real-time, IP-VPNe and satellite network connection (six of each). Network and objective operative data were collected. RESULTS: Despite network delays and jitter, it was feasible to perform the pyeloplasty procedure without significant detriment in operative time or surgical results compared with real-time surgery. CONCLUSION: The completion of complex tasks such as robotic pyeloplasty is feasible using both land-line and satellite telesurgery. However, the clinical relevance of telesurgery requires further assessment.

Randomized trial of peribulbar triamcinolone acetonide with and without focal photocoagulation for mild diabetic macular edema: a pilot study.

OBJECTIVE: To provide pilot data on the safety and efficacy of anterior and posterior sub-Tenon injections of triamcinolone either alone or in combination with focal photocoagulation in the treatment of mild diabetic macular edema (DME). DESIGN: Prospective, phase II, multicenter, randomized clinical trial. PARTICIPANTS: One hundred nine patients (129 eyes) with mild DME and visual acuity 20/40 or better. METHODS: The participants were assigned randomly to receive either focal photocoagulation (n = 38), a 20-mg anterior sub-Tenon injection of triamcinolone (n = 23), a 20-mg anterior sub-Tenon injection followed by focal photocoagulation after 4 weeks (n = 25), a 40-mg posterior sub-Tenon injection of triamcinolone (n = 21), or a 40-mg posterior sub-Tenon injection followed by focal photocoagulation after 4 weeks (n = 22). Follow-up visits were performed at 4, 8, 17, and 34 weeks. MAIN OUTCOME MEASURES: Change in visual acuity and retinal thickness measured with optical coherence tomography (OCT). RESULTS: At baseline, mean visual acuity in the study eyes was 20/25 and mean OCT central subfield thickness was 328 mum. Changes in retinal thickening and in visual acuity were not significantly different among the 5 groups at 34 weeks (P = 0.46 and P = 0.94, respectively). There was a suggestion of a greater proportion of eyes having a central subfield thickness less than 250 mum at 17 weeks when the peribulbar triamcinolone was combined with focal photocoagulation. Elevated intraocular pressure and ptosis were adverse effects attributable to the injections. CONCLUSIONS: In cases of DME with good visual acuity, peribulbar triamcinolone, with or without focal photocoagulation, is unlikely to be of substantial benefit. Based on these results, a phase III trial to evaluate the benefit of these treatments for mild DME is not warranted.

Transcriptional regulation of a hybrid cluster (prismane) protein.

HCP (hybrid-cluster protein) contains two Fe/S clusters, one of which is a hybrid [4Fe-2S-2O] cluster. Despite intensive study, its physiological function has not been reported. The Escherichia coli hcp gene is located in a two-gene operon with hcr, which encodes an NADH-dependent HCP reductase. E. coli HCP is detected after anaerobic growth with nitrate or nitrite: possible roles for it in hydroxylamine or nitric oxide reduction have been proposed. To study the regulation and role of HCP, an hcp::lacZ fusion was constructed and transformed into fnr, arcA and norR mutant strains of E. coli. Transcription from the hcp promoter was induced during anaerobic growth. Only the fnr mutant was defective in hcp expression. Nitrate- and nitrite-induced transcription from the hcp promoter was activated by the response regulator proteins NarL and NarP. Gel retardation assays were used to show that FNR (fumarate-nitrate regulation) and NarL form a complex with the hcp promoter. Transcription of the hcp-hcr operon initiates at a thymine nucleotide located 31 bp upstream of the translation-initiation codon. HCP has been overexpressed from a recombinant plasmid for physiological studies.

Switching patients with migraine from sumatriptan to other triptans increases primary care costs.

Treatment of migraine with triptans is highly effective, although cost considerations may prompt a change in therapy. This retrospective database analysis of 3196 patients with migraine, established on sumatriptan therapy, found that 54% of the 292 experiencing a triptan switch returned to sumatriptan within 15 months, suggesting that the alternative was less acceptable. Excluding patients with unusually high use of triptans (> or = 208 tablets/year), switching therapy resulted in a significant increase of pounds sterling 53/patient in total costs, compared with patients continuing on sumatriptan (p = 0.014). Cost savings (pounds sterling 17/patient over 15 months) were observed only among the 41% of patients in whom the initial switch was successful and did not result in a further switch or return to sumatriptan. Among patients who were relatively low cost initially, switching resulted in increased costs, irrespective of the outcome. This study suggests that there is no economic justification for switching from sumatriptan to another triptan.

Bupropion as an aid to smoking cessation: a review of real-life effectiveness.

Clinical trials rigorously demonstrate the efficacy of new products and justify their marketing. However, it is only after use in real-life settings that the clinical value (effectiveness) of a new treatment is fully known. The purpose of this review was to summarise the effectiveness data for bupropion SR as an aid to smoking cessation. Available reports of effectiveness data for bupropion SR were obtained from the literature, presentations at smoking cessation meetings and from the manufacturer. Twelve sources of effectiveness data were found and included clinical practice trials, observational studies/surveys, motivational support programme results and employer-based cessation programme results. The 6-month point prevalence smoking cessation rates ranged from 25 to 49%. There is a growing body of evidence supporting the effectiveness of bupropion SR as an aid to smoking cessation. Real-life quit rates for bupropion SR are similar to those seen in the original clinical trial programme.