Slit and Receptor Tyrosine Phosphatase 69D Confer Spatial Specificity to Axon Branching via Dscam1.

Axonal branching contributes substantially to neuronal circuit complexity. Studies in Drosophila have shown that loss of Dscam1 receptor diversity can fully block axon branching in mechanosensory neurons. Here we report that cell-autonomous loss of the receptor tyrosine phosphatase 69D (RPTP69D) and loss of midline-localized Slit inhibit formation of specific axon collaterals through modulation of Dscam1 activity. Genetic and biochemical data support a model in which direct binding of Slit to Dscam1 enhances the interaction of Dscam1 with RPTP69D, stimulating Dscam1 dephosphorylation. Single-growth-cone imaging reveals that Slit/RPTP69D are not required for general branch initiation but instead promote the extension of specific axon collaterals. Hence, although regulation of intrinsic Dscam1-Dscam1 isoform interactions is essential for formation of all mechanosensory-axon branches, the local ligand-induced alterations of Dscam1 phosphorylation in distinct growth-cone compartments enable the spatial specificity of axon collateral formation.

Dscam1 overexpression impairs the function of the gut nervous system in Drosophila.

BACKGROUND: Down syndrome (DS) patients have a 100-fold increase in the risk of Hirschsprung syndrome of the colon and rectum (HSCR), a lack of enteric neurons in the colon. The leading DS candidate gene is trisomy of the Down syndrome cell adhesion molecule (DSCAM). RESULTS: We find that Dscam1 protein is expressed in the Drosophila enteric/stomatogastric nervous system (SNS). Axonal Dscam1 phenotypes can be rescued equally by diverse isoforms. Overexpression of Dscam1 resulted in frontal and hindgut nerve overgrowth. Expression of dominant negative Dscam1-ΔC led to a truncated frontal nerve and increased branching of the hindgut nerve. Larval locomotion is influenced by feeding state, and we found that the average speed of larvae with Dscam1 SNS expression was reduced, whereas overexpression of Dscam1-ΔC significantly increased the speed. Dscam1 overexpression reduced the efficiency of food clearance from the larval gut. CONCLUSION: Our work demonstrates that overexpression of Dscam1 can perturb gut function in a model system.

Proteolytic cleavage of Slit by the Tolkin protease converts an axon repulsion cue to an axon growth cue in vivo.

Slit is a secreted protein that has a canonical function of repelling growing axons from the CNS midline. The full-length Slit (Slit-FL) is cleaved into Slit-N and Slit-C fragments, which have potentially distinct functions via different receptors. Here, we report that the BMP-1/Tolloid family metalloprotease Tolkin (Tok) is responsible for Slit proteolysis in vivo and in vitro. In Drosophilatok mutants lacking Slit cleavage, midline repulsion of axons occurs normally, confirming that Slit-FL is sufficient to repel axons. However, longitudinal axon guidance is highly disrupted in tok mutants and can be rescued by midline expression of Slit-N, suggesting that Slit is the primary substrate for Tok in the embryonic CNS. Transgenic restoration of Slit-N or Slit-C does not repel axons in Slit-null flies. Slit-FL and Slit-N are both biologically active cues with distinct axon guidance functions in vivo Slit signaling is used in diverse biological processes; therefore, differentiating between Slit-FL and Slit fragments will be essential for evaluating Slit function in broader contexts.

The use of NovoSorb biodegradable temporising matrix in wound management: a literature review and case series.

OBJECTIVE: NovoSorb (Poly-Novo Ltd, Australia) biodegradable temporising matrix (BTM) is a novel artificial dermal matrix. Previous literature is weighted towards its use in burns reconstruction; however, this paper describes its use within a range of wound aetiologies. The authors present one of the largest and most diverse case series to date, and aim to provide an independent benchmark of clinical practice. METHOD: A retrospective observational study was performed. Patient demographics and clinical data (wound aetiology, site, total body surface area (TBSA), wound bed, number of debridements, time to BTM integration, time to skin grafting) were collected and subgroup analysis preformed. RESULTS: The cohort consisted of 37 patients (acute trauma wounds, n=19; hard-to-heal wounds, n=9; acute infections, n=6; cancer, n=3). Successful BTM integration, allowing subsequent split-thickness skin grafting (STSG), occurred in 70% of cases, despite an overall complication rate of 51%. Mean time from BTM application to STSG was 53 days. There was no difference in STSG outcomes when grafting was performed either before or after the six-week BTM application period. There was no difference when various wound beds (fascia, tendon, periosteum) were compared. Patients >65 years of age were more likely to experience complications; however, this did not affect the speed of integration. The relation of diabetes and smoking to overall integration had no statistical significance. CONCLUSION: In comorbid patients in particular, the time between BTM application and STSG may be longer than the manufacturer's recommendation. Furthermore, data suggest greater wound bed optimisation and closer interval monitoring in hard-to-heal/malignant wounds, and in older patients and patients with comorbidities. However, BTM appears robust (even in infection) and is showing promise as a useful reconstructive tool.

Solve study: a study to capture global variations in practices concerning laparoscopic cholecystectomy.

BACKGROUND: There is a lack of published data on variations in practices concerning laparoscopic cholecystectomy. The purpose of this study was to capture variations in practices on a range of preoperative, perioperative, and postoperative aspects of this procedure. METHODS: A 45-item electronic survey was designed to capture global variations in practices concerning laparoscopic cholecystectomy, and disseminated through professional surgical and training organisations and social media. RESULTS: 638 surgeons from 70 countries completed the survey. Pre-operatively only 5.6% routinely perform an endoscopy to rule out peptic ulcer disease. In the presence of preoperatively diagnosed common bile duct (CBD) stones, 85.4% (n = 545) of the surgeons would recommend an Endoscopic Retrograde Cholangio-Pancreatography (ERCP) before surgery, while only 10.8% (n = 69) of the surgeons would perform a CBD exploration with cholecystectomy. In patients presenting with gallstone pancreatitis, 61.2% (n = 389) of the surgeons perform cholecystectomy during the same admission once pancreatitis has settled down. Approximately, 57% (n = 363) would always administer prophylactic antibiotics and 70% (n = 444) do not routinely use pharmacological DVT prophylaxis preoperatively. Open juxta umbilical is the preferred method of pneumoperitoneum for most patients used by 64.6% of surgeons (n = 410) but in patients with advanced obesity (BMI > 35 kg/m2, only 42% (n = 268) would use this technique and only 32% (n = 203) would use this technique if the patient has had a previous laparotomy. Most surgeons (57.7%; n = 369) prefer blunt ports. Liga clips and Hem-o-loks® were used by 66% (n = 419) and 30% (n = 186) surgeons respectively for controlling cystic duct and (n = 477) 75% and (n = 125) 20% respectively for controlling cystic artery. Almost all (97.4%) surgeons felt it was important or very important to remove stones from Hartmann's pouch if the surgeon is unable to perform a total cholecystectomy. CONCLUSIONS: This study highlights significant variations in practices concerning various aspects of laparoscopic cholecystectomy.

The Drosophila Ret gene functions in the stomatogastric nervous system with the Maverick TGFß ligand and the Gfrl co-receptor.

The RET receptor tyrosine kinase is crucial for the development of the enteric nervous system (ENS), acting as a receptor for Glial cell line-derived neurotrophic factor (GDNF) via GFR co-receptors. Drosophila has a well-conserved RET homolog (Ret) that has been proposed to function independently of the Gfr-like co-receptor (Gfrl). We find that Ret is required for development of the stomatogastric (enteric) nervous system in both embryos and larvae, and its loss results in feeding defects. Live imaging analysis suggests that peristaltic waves are initiated but not propagated in mutant midguts. Examination of axons innervating the midgut reveals increased branching but the area covered by the branches is decreased. This phenotype can be rescued by Ret expression. Additionally, Gfrl shares the same ENS and feeding defects, suggesting that Ret and Gfrl might function together via a common ligand. We identified the TGFß family member Maverick (Mav) as a ligand for Gfrl and a Mav chromosomal deficiency displayed similar embryonic ENS defects. Our results suggest that the Ret and Gfrl families co-evolved before the separation of invertebrate and vertebrate lineages.

The WAGR syndrome gene PRRG4 is a functional homologue of the commissureless axon guidance gene.

WAGR syndrome is characterized by Wilm's tumor, aniridia, genitourinary abnormalities and intellectual disabilities. WAGR is caused by a chromosomal deletion that includes the PAX6, WT1 and PRRG4 genes. PRRG4 is proposed to contribute to the autistic symptoms of WAGR syndrome, but the molecular function of PRRG4 genes remains unknown. The Drosophila commissureless (comm) gene encodes a short transmembrane protein characterized by PY motifs, features that are shared by the PRRG4 protein. Comm intercepts the Robo axon guidance receptor in the ER/Golgi and targets Robo for degradation, allowing commissural axons to cross the CNS midline. Expression of human Robo1 in the fly CNS increases midline crossing and this was enhanced by co-expression of PRRG4, but not CYYR, Shisa or the yeast Rcr genes. In cell culture experiments, PRRG4 could re-localize hRobo1 from the cell surface, suggesting that PRRG4 is a functional homologue of Comm. Comm is required for axon guidance and synapse formation in the fly, so PRRG4 could contribute to the autistic symptoms of WAGR by disturbing either of these processes in the developing human brain.

Correction: The WAGR syndrome gene PRRG4 is a functional homologue of the commissureless axon guidance gene.

[This corrects the article DOI: 10.1371/journal.pgen.1006865.].

Dscam1 Forms a Complex with Robo1 and the N-Terminal Fragment of Slit to Promote the Growth of Longitudinal Axons.

The Slit protein is a major midline repellent for central nervous system (CNS) axons. In vivo, Slit is proteolytically cleaved into N- and C-terminal fragments, but the biological significance of this is unknown. Analysis in the Drosophila ventral nerve cord of a slit allele (slit-UC) that cannot be cleaved revealed that midline repulsion is still present but longitudinal axon guidance is disrupted, particularly across segment boundaries. Double mutants for the Slit receptors Dscam1 and robo1 strongly resemble the slit-UC phenotype, suggesting they cooperate in longitudinal axon guidance, and through biochemical approaches, we found that Dscam1 and Robo1 form a complex dependent on Slit-N. In contrast, Robo1 binding alone shows a preference for full-length Slit, whereas Dscam1 only binds Slit-N. Using a variety of transgenes, we demonstrated that Dscam1 appears to modify the output of Robo/Slit complexes so that signaling is no longer repulsive. Our data suggest that the complex is promoting longitudinal axon growth across the segment boundary. The ability of Dscam1 to modify the output of other receptors in a ligand-dependent fashion may be a general principle for Dscam proteins.

Microsatellite alteration and immunohistochemical expression profile of chromosome 9p21 in patients with sporadic renal cell carcinoma following surgical resection.

BACKGROUND: Long-term prognostic significance of loss of heterozygosity on chromosome 9p21 for localized renal cell carcinoma following surgery remains unreported. The study assessed the frequency of deletions of different loci of chromosome 9p along with immunohistochemical profile of proteins in surgically resected renal cancer tissue and correlated this with long-term outcomes. METHODS: DNA was extracted from renal tumours and corresponding normal kidney tissues in prospectively collected samples of 108 patients who underwent surgical resection for clinically localized disease between January 2001 and December 2005, providing a minimum of 9 years follow-up for each participant. After checking quality of DNA, amplified by PCR, loss of heterozygosity (LOH) on chromosome 9p was assessed using 6 microsatellite markers in 77 clear cell carcinoma. Only 5 of the markers showed LOH (D9S1814, D9S916, D9S974, D9S942, and D9S171). Protein expression of p15(INK4b), p16(INK4a), p14(ARF), CAIX, and adipose related protein (ADFP) were demonstrated by immunostaining in normal and cancer tissues. Loss of heterozygosity for microsatellite analysis was correlated with tumour characteristics, recurrence free, cancer specific, and overall survival, including significance of immunohistochemical profile of protein expressions. RESULTS: The main deletion was found at loci telomeric to CDKN2A region at D9S916. There was a significant correlation between frequency of LOH stage (p = 0.005) and metastases (p = 0.006) suggesting a higher LOH for advanced and aggressive renal cell carcinoma. Most commonly observed LOH in the 3 markers: D9S916, D9S974, and D9S942 were associated with poor survival, and were statistically significant on multivariate analysis. Immunohistochemical expression of p14, p15, and p16 proteins were either low or absent in cancer tissue compared to normal. CONCLUSIONS: Loss of heterozygosity of p921 chromosome is associated with aggressive tumours, and predicts cancer specific or recurrence free survival on long-term follow-up.

Implant reconstruction after mastectomy-A review and summary of current literature.

INTRODUCTION: The landscape of breast reconstruction has changed significantly with a shift in focus to include the restoration of a patient's quality of life after cancer. Reconstructive options can be divided into alloplastic (implant based) and autologous (tissue based). This paper aims to provide a current educational summary regarding implant-based reconstruction after breast cancer surgery and review the current literature. METHOD: A review of the literature was conducted utilising standard PRISMA flowchart. Databases searched included Pubmed, EMBASE, and MEDLINE. RESULTS: Current practice is explored within the text, including types of implants, indications, and surgical approaches. Heterogenous cohorts, surgical technique variation, and selection bias can make comparison of the literature challenging. The major evidence reviews of implant-based reconstruction topics are discussed including, ADM use, radiotherapy, and complications. Despite the benefits of autologous reconstruction, implant-based techniques still represent a significant proportion of reconstructive breast procedures. However, implant-reconstruction is not without its risks and limitations and, with such variety in practice, there remains a lack of high-quality evidence guiding practice. Most importantly, patients need to be counselled about the pros and cons of each choice, particularly with the increasing utilisation of radiotherapy post-reconstruction. Ultimately, the patient and surgeon should reach a decision in full knowledge of the risks and potential outcomes. CONCLUSIONS: Further research is required into implant-based reconstructive therapy, which will allow a greater consensus for management and a pathway for both surgeons and patients.

Collection, Fixation, and Antibody Staining of Drosophila Embryos.

The Drosophila embryonic central nervous system has been used for decades as a model for understanding the genetic regulation of axon guidance and other aspects of neural development. Foundational studies using antibody staining to examine the embryonic ventral nerve cord in wild-type and mutant animals led to the discovery of evolutionarily conserved genes that regulate fundamental aspects of axon guidance, including midline crossing of axons. The development of the regular, segmentally repeating structure of axon pathways in the ventral nerve cord can illustrate basic principles of axon guidance to beginning students and can also be used by expert researchers to characterize new mutants, detect genetic interactions between known genes, and precisely quantify variations in gene function in engineered mutant lines. Here, we describe a protocol for collecting and fixing Drosophila embryos and visualizing axon pathways in the embryonic ventral nerve cord using immunofluorescence or immunohistochemical staining methods. As embryogenesis in Drosophila takes ∼24 h to complete, a 1-d collection yields embryos representing all stages of development from newly fertilized through ready-to-hatch larvae, allowing investigation of multiple developmental events within a single batch of collected embryos. The methods described in this protocol should be accessible to introductory laboratory courses as well as seasoned investigators in established research laboratories.

Ventral Nerve Cord Dissection and Microscopy of Drosophila Embryos.

The technique of visualizing axon pathways in the embryonic ventral nerve cord using antibody labeling has been fundamental to our understanding of the genetic and developmental mechanisms underlying nervous system wiring in Drosophila. High-resolution microscopic examination of the ventral nerve cord remains an essential component of many experiments in Drosophila developmental neuroscience. Although it is possible to examine the ventral nerve cord in intact whole-mount embryos, to collect the highest-quality images it is often useful to isolate the nervous system away from the other embryonic tissues through embryo dissection. This protocol describes methods for dissecting ventral nerve cords from Drosophila embryos that have been fixed and stained via immunofluorescence or horseradish peroxidase (HRP) immunohistochemistry. The process of making fine dissection needles for this purpose from electrolytically sharpened tungsten wire is also described. Dissected and mounted ventral nerve cords can be examined and imaged using a variety of microscopy techniques including differential interference contrast (DIC) optics, epifluorescence, or confocal microscopy.

Analysis of Axon Guidance in the Drosophila Embryo.

The establishment of neural connectivity is a major part of neural development. The central nervous system (CNS) midline is the most characterized axon guidance choice point, and work in Drosophila has played a pivotal role in understanding the molecular mechanisms responsible. Axons respond to attractive cues such as Netrin via the Frazzled receptor, and repulsive cues such as Slit via Robo receptors. Both signals are expressed at the CNS midline, affect pioneer axons, and have dramatic effects on the axon scaffold as a whole. Here, we focus on previous research analyzing classic mutants in the Slit/Robo pathway, which can readily be detected with a dissecting microscope. We also discuss analyzing these mutants in a teaching lab situation. The combination of sophisticated genetics and reliable axonal markers in Drosophila allows phenotypic analysis to be performed at the single-cell level. The elaborate architecture of neurons is very sensitive to disruption by genetic mutations, allowing the effects of novel mutations to be easily detected and assessed.

Axons will not be silenced! A balancing act of attractive receptors.

Axons crossing the CNS midline regulate their responsiveness to both attractive and repulsive cues. In this issue, Dailey-Krempel et al. find different modes of action for DCC isoforms and uncover evidence against the silencing model of axon guidance.

Short- and Long-Term Complications of Free Anterolateral Thigh Flap Reconstructions: A Single-Centre Experience of 92 Consecutive Cases.

Background: The anterolateral thigh (ALT) flap has been amongst the most versatile components of the reconstructive surgeon's armamentarium. The authors utilise these flaps for a variety of reconstructive procedures including lower limb reconstruction; postsarcoma excision; and open fractures. Few studies have discussed the extent of recipient site morbidity and subsequent revisional procedures. We will report our experience of the ALT flap in 92 consecutive reconstructions with focus on recipient site complications and revisional procedures. Methods: Retrospective data collection was done from 92 patients who underwent ALT flap reconstruction-for various large soft tissue defects-at our unit at the Royal Free Hospital, London. We evaluated primary recipient site complications and the requirements for secondary operations after flap transfer. Results: All flaps survived with the exception of 3 cases (97% survival rate) in which irreversible venous thrombosis was encountered. 16 of 92 patients (17%) required a second recipient site operation for the following: 7 patients experienced major recipient site complications that warranted early return to theatre and 9 patients required a secondary revision thinning procedure(s). 8 of the 16 patients (50%) requiring second operations had construction on their lower leg/ankle/feet (p value = 0.10). Conclusions: Our data demonstrated effective use of the ALT flap in the management of soft tissue reconstructive surgery. Partial flap necrosis was the main complication at the recipient site. In future work, secondary thinning procedures, particularly at the ankle/foot, should be separated from flap-specific complications. Furthermore, we demonstrate tailoring ALT thickness can be performed safely without compromising flap viability.

Closed Rhinoplasty: A Single Surgeon Experience of 238 Cases over 2 Years.

Closed rhinoplasty is currently losing favour as a method of rhinoplasty reshaping procedure. Open rhinoplasties are usually performed by surgeons because of the greater degree of visualisation of the cartilages and bones within the nose that need to be reshaped. Because of the criticisms of closed rhinoplasty the senior author performed an audit of his outcomes to determine whether closed rhinoplasty still has a role to play in the armamentarium of the plastic surgeon. We conducted this study as a retrospective review of all closed rhinoplasty operations performed by a single surgeon over a two-year period (from 1st January 2016 to 31st December 2017). The operative technique is provided. An independent panel of 3 assessors (2 board certified consultant plastic surgeons and one lay member of the public) rated outcomes of the closed technique based on photographic series. Statistical analysis was performed using Cohen's Kappa and Friedman test. Additionally, the length of follow up period, revision rates, and post-operative complications (general and aesthetic) were examined. A total of 242 cases of rhinoplasty were performed (8 open and 234 closed rhinoplasty; 225 primary and 17 revision operations). The first consultant surgeon gave a score in a range from 4.2/5 to 4.9/5 with a mode of 4.8 and a mean score of 4.7/5. The second consultant surgeon gave a range of 4.1/5 to 4.9/5 with a mode of 4.4 and mean score of 4.3/5. The lay panel member scored the cases in a range from 4.4/5 to 5/5 with a mode of 4.8 and a mean score of 4.9/5. Cohen's coefficient was 0.72 showing substantial agreement across the panel. The complication rate for the recorded data was 0.8% with two cases of soft tissue infection treated with oral antibiotics. The results of this paper advocates the benefits of the closed approach in aesthetic rhinoplasty. Surgeons should appreciate that both open and closed approaches are complimentary. The favouring of a single approach is indicated in the scenarios discussed. The remaining cases seem to produce equivocal results and the choice of procedure should be based on patient anatomy, outcome aims, and the ability of a surgeon to perform their preferred technique.

Avoiding the SCAMs.

Dendrites from the same neuron usually avoid contact with one another, a behavior known as self-avoidance. In this issue of Neuron and in the upcoming May 4, 2007 issue of Cell, a pair of studies by Soba et al. and Hughes et al. and a study by Matthews et al., respectively, identify products from the highly alternatively spliced Dscam gene as central to this behavior in Drosophila. Signaling induced by adhesion between identical isoforms triggers repulsion between sister dendrites.

Attitudes, Influences and Perceptions towards Plastic Surgery amongst Medical Students.

INTRODUCTION: Plastic surgery is a dynamic and evolving field but remains poorly understood due to lack of knowledge, media misconceptions and recent changes to medical undergraduate curricula. To address issues around student interest and recruitment into the speciality, it is imperative to understand the factors influencing medical students and future clinicians. AIMS: To examine influences, interest and perceptions of plastic surgery amongst Scottish medical students and explore methods to increase undergraduate engagement. METHOD: Cross-sectional survey distributed online via Scottish undergraduate medical school offices comprising 6 domains: demographics; career interest; perceptions, interests and influences in plastic surgery; curriculum and trainer views; understanding the role of a plastic surgeon; and undergraduate engagement. RESULTS: A total of 193 students responded with no statistically significant relationship between year group, gender, and interest in plastic surgery. Phrases most strongly identified with plastic surgery included private practice, reconstruction and cosmetics. Placements, teaching staff and workshops/courses were found to influence perception of plastic surgery. Fortunately, only 6% of students encountered antagonism towards plastic surgery encompassing themes of negative stereotypes of surgeons and connotations surrounding cosmetic surgery. Importantly, many students were largely unaware of the range of common procedures undertaken by plastic surgeons. To overcome this lack of awareness and generate greater interest, students suggested greater plastics exposure, consultant-led teaching and workshops showcasing the specialty. CONCLUSION: Medical students want varied, stimulating and flexible careers - something which plastic surgery can provide. However it seems the understanding of the scope of plastic surgery is poorly understood amongs future trainees. To increase uptake and interest, negative perceptions need to be addressed and greater engagement is required from medical school upwards.

Head and neck research in plastic surgery-Ahead of the rest? An analysis of abstracts presented at British association of plastic reconstructive and aesthetic surgeons meetings.

INTRODUCTION & AIMS: Historically, the head and neck (H&N) discipline has been integral to the service a plastic surgeon provides. Recently, it has been postulated that its popularity is declining. The output of scientific meetings may indicate the popularity of each sub-speciality interest, also allowing comparison with other H&N conferences. AIM: To analyse the proportion of H&N themed, podium and poster presentations from British Association of Plastic Reconstructive and Aesthetic Surgeons' (BAPRAS) scientific meetings and the resulting contribution to published literature. MATERIAL AND METHODS: H&N-themed abstracts were identified from finalised programmes of the biannual BAPRAS meetings between 2008 and 2015. PubMed and Google Scholar databases were searched using keywords and author names from each abstract to identify subsequent publication in a peer-reviewed journal. RESULTS: Overall, 19.3% (350/1815) of BAPRAS abstracts were H&N themed. The publication rate of H&N abstracts was 40.3% (141/350), comprising 43.0% (114/265) of podium and 31.8% (27/85) of poster presentations. H&N reconstruction and cleft and craniofacial were the most frequent topics, with facial palsy having the lowest conversion rate at 15.4%. The mean time to publication was 17.8 months. Research was published in 39 journals, with a mean impact factor of 2.151 (range = 0.772-11.541). The most popular journal was Journal of Plastic, Reconstructive & Aesthetic Surgery (JPRAS; 31.7%; 45/141). Published abstracts originated from 17 different countries. The senior author was a plastic surgeon in 77.3% of abstracts. CONCLUSIONS: From the multiple potential sub-specialties, H&N-themed abstracts consistently contributed 20% of all research presented at BAPRAS. The 40.3% publication rate exceeds the international average of scientific meetings. H&N remains a prominent field in the armamentarium of a plastic surgeon.