Transcriptional regulation of S100B and identification of novel melanoma biomarkers

Summary Introduction Cutaneous melanoma is an aggressive disease, which is recognised as the most common fatal skin cancer worldwide. Approximately 450 cases are diagnosed in Ireland per year. Between 1994 and 2004, melanoma demonstrated a higher rate of increase in mortality than any other cancer in this country. Although the prognosis for early melanoma is favorable, less than 20 percent of patients with metastatic melanoma survive for five years. S100B is a calcium sensor protein that modulates biological activity via calcium binding, which is routinely used in histological diagnosis of malignant melanoma and which is also a well-recognized serum marker of the disease. HOX proteins are members of the homeodomain family of transcription factors, which are involved in a host of cellular functions including organogenesis, cellular differentiation, cell cycle and apoptosis. As transcription factors, HOX proteins require co-activator proteins to achieve their full function. SRC-1 is one such coactivator and our group has extensively explored its function. In particular, functional interactions between SRC-1 and HOXCI 1 in breast cancer cell lines and tissue have been described. HOXCI 1 is known to enhance expression of the secreted serum marker S100B. Given the strong association between S100B and malignant melanoma, we believe that this pathway may also have a role in melanoma tumour genesis. S100B is a well-described biomarker in melanoma and serum levels have been shown to correlate with disease stage and response to treatment. In spite of this and the availability other prognostic indicators, many patients go on to develop an unpredictable disease course. For this reason, identification of novel biomarkers is an active area of melanoma research. In the second part of this work, an autoantibody microarray screen was undertaken to identify differentially expressed biomarkers in sera from patients with melanoma. Hypothesis Production of S100B in malignant melanoma is regulated by the transcription factor HOXCI 1, in cooperation with coactivator SRC-1. Protein microarray technology may provide a useful means of identification of autoantibody biomarkers in serum from patients with melanoma. Aims To define the molecular role of HOXC11 and SRC-1 in the transcriptional control of S100B in malignant melanoma. To characterise the effect of manipulation of HOXC11 and SRC-1 on S100B expression. To identify new biomarkers in sera from patients with malignant melanoma. Results Expression of SIOOB, HOXC11 and SRC-1 protein in primary (SKMe128) and metastatic (MeWo) melanoma cell lines was confirmed by Western blotting and quantitative Real Time PCR (qRT-PCR) analysis. Colocalisation of HOXCI 1 and SRC-1 in melanoma cells was confirmed by immunofluorescence. Co-immunoprecipitation was carried out and demonstrated interaction of HOXC11 and SRC-1 in cell lysates. Paraffinembedded melanoma and nevi samples were examined by immunofluorescence and a significantly higher nuclear expression of HOXCI 1 and SRC-1 was observed in the melanoma cohort. Colocalisation of the two proteins was also demonstrated in a series of melanoma primary culture specimens. Chromatin-immunoprecipitation was employed to confirm recruitment of HOXCI 1 to the promoter region of the S100B gene. To determine the ability of HOXCI 1 to regulate expression of S1 OOB, HOXCI 1 was transfected into the SKMe128 cell line and it was found to significantly increase the expression of the target gene S100B. When concomitant HOXCI 1 and SRC- 1 knockdowns were performed, a significant reduction in the presence of S100B was noted. Treatment of cell lines with the phospho-Src inhibitor, dasatinib, resulted in decreased coassociation between HOXCI 1 and SRC-1 in both primary and metastatic cell lines as well as decreased expression of S100B in SKMe128 cells. Protein microarray analysis of sera from patients with melanoma and control patients was carried out. A series of differentially expressed autoantibodies was identified and the non-receptor tyrosine kinase, BMX was chosen for further study. Elevated expression of anti-BMX autoantibody in sera from a larger cohort of melanoma patients was confirmed. Furthermore, expression of BMX protein in melanoma cell lines and frozen tissue samples was confirmed by Western blotting. Conclusion In the absence of effective treatment for advanced melanoma, elucidation of novel signalling pathways and therapeutic targets remains at the forefront of molecular research. In this work, translational techniques have provided an insight in to the transcriptional regulation of S100B in melanoma. Furthermore, protein microarray analysis has been utilised to identify potentially useful autoantibody biomarkers. These findings constitute a small fragment of all the potential genetic aberrations that may be implicated in melanoma turnourgenesis. Advanced melanoma is likely to present a significant therapeutic challenge to clinicians and academics for many years to come. What is certain is that translational research methods, as have been employed here, are essential in pushing forward the boundaries of our molecular understanding of this fascinating disease

NovoSorb biodegradable temporizing matrix for reconstruction of multiplanar degloving injury of the upper limb

Originally described as "wringer injuries" by MacCollum in 1938, traumatic multiplanar degloving injuries that occur as the result of the hand, forearm or arm being drawn between the rollers of a machine are functionally devastating and present a significant reconstructive challenge. Revascularization and comprehensive excision of devitalized bone and soft tissue, followed by appropriate skeletal fixation and vascularized soft tissue cover are the mainstays of management. To date, published case series have described local flaps and free tissue transfer for coverage of wounds that involve exposed vital structures such as nerves, vessels, and tendons.NovoSorb biodegradable temporizing matrix (BTM; PolyNovo Biomaterials Pty Ltd, Melbourne, Australia) is a bilayer bioabsorbable synthetic polymer dermal substitute, which has the ability to integrate into large wound beds and is resistant to infection.3BTM comprises a bioabsorbable, polyurethane matrix that allows for cellular infiltration and a temporary nonbiodegradable, nonporous polyurethane layer, which limits moisture loss and provides a barrier to bacteria. Here we describe the successful use of BTM in the staged reconstruction of a high-energy industrial roller injury in an adolescent patient.</p

NovoSorb Biodegradable Temporizing Matrix for Reconstruction of Multiplanar Degloving Injury of the Upper Limb

Originally described as “wringer injuries” by MacCollum in 1938,1 traumatic multiplanar degloving injuries that occur as the result of the hand, forearm or arm being drawn between the rollers of a machine are functionally devastating and present a significant reconstructive challenge. Revascularization and comprehensive excision of devitalized bone and soft tissue, followed by appropriate skeletal fixation and vascularized soft tissue cover are the mainstays of management. To date, published case series have described local flaps and free tissue transfer for coverage of wounds that involve exposed vital structures such as nerves, vessels, and tendons.2 NovoSorb biodegradable temporizing matrix (BTM; PolyNovo Biomaterials Pty Ltd, Melbourne, Australia) is a bilayer bioabsorbable synthetic polymer dermal substitute, which has the ability to integrate into large wound beds and is resistant to infection.3 BTM comprises a bioabsorbable, polyurethane matrix that allows for cellular infiltration and a temporary nonbiodegradable, nonporous polyurethane layer, which limits moisture loss and provides a barrier to bacteria. Here we describe the successful use of BTM in the staged reconstruction of a high-energy industrial roller injury in an adolescent patient

Frostbite injuries from recreational nitrous oxide use

Introduction Recreational use of inhaled nitrous oxide is rapidly increasing. As liquid nitrous oxide (N2O) is cooled to its gaseous form for inhalation, container mishandling can cause frostbite injuries to the face and extremities. Cases (1) 19-year-old male with 2% total body surface area (TBSA) frostbite burn to bilateral inner thighs; (2) 18-year-old female with 1.5% TBSA burn to palm and fingers; (3) 14-year-old female with 4% TBSA burn to bilateral inner thighs; (4) 23-year-old female with 0.5% TBSA burn to fingers; (5) 22-year-old female with 0.2% TBSA frostbite burn to fingers; (6) 21-year-old female with 1.5% TBSA burn to volar forearm; (7) 17-year-old female with 0.5% burn to lips and chin. Outcome Two patients required debridement and skin grafting, which resulted in significant scarring and contour deformity. One patient presented with toxic shock syndrome and required management in the paediatric intensive care unit. All patients required specialised dressings and occupational therapy for scar management and/or hand rehabilitation. Discussion We have observed a cluster of frostbite burns sustained from recreational use of N2O. Healthcare workers should be aware of the practice and be able to recognise and manage the associated injuries.</div

Feasibility of cleft lip and palate repair in personal protective equipment (PPE).

Elective surgery during the evolving COVID-19 pandemic presents unprecedented logistical challenges to surgical teams. Cleft surgery may be considered an aerosol generating procedure (AGP), which may lead to small-droplet transmission of virions. Strict adherence to personal protective equipment (PPE) policy is used with the hope of preventing transmission of the virus between patients and operating theatre staff. </p

Non-interventional factors influencing velopharyngeal function for speech in initial cleft palate repair: a systematic review protocol

Background: This systematic review aims to inform the development of a screening tool which pre-operatively predicts which children are likely to develop velopharyngeal insufficiency, one of the causes of poor speech outcomes, following cleft palate repair. This would be highly beneficial as it would inform pre-operative counselling of parents, allow targeted speech and language therapy, and enable meaningful comparison of outcomes between surgeons, techniques, and institutions. Currently, it is unclear which factors influence speech outcomes. A systematic review investigating the non-interventional factors which potentially influence speech outcomes following cleft palate repair is warranted. This may be illuminating in itself or provide foundations for future studies. Methods: A systematic review will be carried out according to Cochrane methodology and reported according to PRISMA guidelines (PLoS Med 6: e1000097, 2009). Systematic review software will be used to facilitate three-stage screening by two independent reviewers experienced in cleft lip and palate. Thereafter, data extraction and GRADE assessment will be performed in duplicate by five independent reviewers experienced in cleft lip and palate. Studies reporting the proportion of patients who were recommended or underwent secondary speech surgery for velopharyngeal insufficiency following primary surgery for cleft palate will be included. The study findings will be tabulated and summarised. The primary outcome measure will be further speech surgery (either recommended or performed). The secondary outcome measure will be perceptual speech assessment for the presence of velopharyngeal insufficiency. A meta-analysis is planned. However, if this is not possible, due to the anticipated marked heterogeneity of study characteristics, pre-operative assessment, and the recorded outcome measures, a narrative synthesis will be undertaken. Discussion: This systematic review may provide sufficient data to inform the development of a screening tool to predict the risk of velopharyngeal insufficiency prior to cleft palate repair. However, it is anticipated that these findings will provide the foundation for future studies in this area. Systematic review registration: Registered on 19 December 2016 with PROSPERO CRD42017051624.</p

Minimally-invasive airway management and early cleft palate repair in infants born with Robin sequence

The objective of this study was to report outcomes of early cleft palate repair in infants born with Robin sequence (RS). A retrospective case series in a tertiary referral paediatric hospital was carried out, examining a consecutive series of 69 infants born with RS and cleft palate. A minimally invasive approach was taken to upper airway obstruction, with liberal nasopharyngeal airway (NPA) and non-invasive ventilation (NIV) use, guided by sleep studies. The palate was repaired between 6 and 9 months with a modified Malek technique. The most frequently used airway adjunct (59.4% of patients) was an NPA and the median duration of use was 5.6 months. All patients underwent a modified Malek cleft palate repair at a median of 7 months of age. Overnight oximetry demonstrated higher mean oxygen saturation (SpO2) across the group from initial neonatal admission to discharge (median 96.5% (interquartile range [IQR] 95-98%) vs 97.45% (IQR 96.5-98%) (P = 0.2, N = 34). Of those with a cardiorespiratory polysomnogram, the obstructive apnoea-hypopnea index (OAHI) was significantly lower postoperatively (5.9 vs 2.8, P = 0.028). This study supports the use of non-surgical airway strategies and early cleft palate repair in infants born with RS and cleft palate. </p