Mechanisms underlying clinical efficacy of Angiotensin II type 2 receptor (AT2R) antagonist EMA401 in neuropathic pain: clinical tissue and in vitro studies.

BACKGROUND: The clinical efficacy of the Angiotensin II (AngII) receptor AT2R antagonist EMA401, a novel peripherally-restricted analgesic, was reported recently in post-herpetic neuralgia. While previous studies have shown that AT2R is expressed by nociceptors in human DRG (hDRG), and that EMA401 inhibits capsaicin responses in cultured hDRG neurons, the expression and levels of its endogenous ligands AngII and AngIII in clinical neuropathic pain tissues, and their signalling pathways, require investigation. We have immunostained AngII, AT2R and the capsaicin receptor TRPV1 in control post-mortem and avulsion injured hDRG, control and injured human nerves, and in cultured hDRG neurons. AngII, AngIII, and Ang-(1-7) levels were quantified by ELISA. The in vitro effects of AngII, AT2R agonist C21, and Nerve growth factor (NGF) were measured on neurite lengths; AngII, NGF and EMA401 effects on expression of p38 and p42/44 MAPK were measured using quantitative immunofluorescence, and on capsaicin responses using calcium imaging. RESULTS: AngII immunostaining was observed in approximately 75% of small/medium diameter neurons in control (n = 5) and avulsion injured (n = 8) hDRG, but not large neurons i.e. similar to TRPV1. AngII was co-localised with AT2R and TRPV1 in hDRG and in vitro. AngII staining by image analysis showed no significant difference between control (n = 12) and injured (n = 13) human nerves. AngII levels by ELISA were also similar in control human nerves (4.09 ± 0.36 pmol/g, n = 31), injured nerves (3.99 ± 0.79 pmol/g, n = 7), and painful neuromas (3.43 ± 0.73 pmol/g, n = 12); AngIII and Ang-(1-7) levels were undetectable (<0.03 and 0.05 pmol/g respectively). Neurite lengths were significantly increased in the presence of NGF, AngII and C21 in cultured DRG neurons. AngII and, as expected, NGF significantly increased signal intensity of p38 and p42/44 MAPK, which was reversed by EMA401. AngII mediated sensitization of capsaicin responses was not observed in the presence of MAP kinase inhibitor PD98059, and the kinase inhibitor staurosporine. CONCLUSION: The major AT2R ligand in human peripheral nerves is AngII, and its levels are maintained in injured nerves. EMA401 may act on paracrine/autocrine mechanisms at peripheral nerve terminals, or intracrine mechanisms, to reduce neuropathic pain signalling in AngII/NGF/TRPV1-convergent pathways.

Summary document on safety and recommendations on liposuction for lipoedema: Joint British association of aesthetic plastic surgeons (BAAPS)/British association of plastic reconstructive and aesthetic surgeons (BAPRAS) expert liposuction group.

Liposuction plays an important role as a surgical treatment option for lipoedema. This article serves to critically review the evidence in the literature, as well as explain the differences between the lipoedema population compared with the aesthetic surgery population undergoing liposuction. It is not a comprehensive text on lipoedema management but serves to guide surgeons. This guidance was produced on behalf of the British Association of Aesthetic Plastic Surgeons (BAAPS) and the British Association of Plastic Reconstructive and Aesthetic Surgeons (BAPRAS) by the expert liposuction group. The guidance is based on the evidence available in the literature, along with a specialist expert opinion on liposuction for lipoedema, to provide plastic surgeons with a consensus recommendation for surgical treatment. The aim is to identify best practice to maximise the safety of patients. This article summarises current practices and safety considerations and outlines recommendations covering various aspects of patient care.

Summary document on safety and recommendations for aesthetic liposuction: Joint British Association of Aesthetic Plastic Surgeons (BAAPS)/British Association of Plastic Reconstructive and Aesthetic Surgeons (BAPRAS) expert liposuction group.

Liposuction is one of the commonest surgical aesthetic procedures performed worldwide. Despite being perceived to be a technically simple procedure, poor patient selection, sub-optimal technical execution or sub-optimal peri­operative management could lead to significant harm. This guidance was produced on behalf of the British Association of Aesthetic Plastic Surgeons (BAAPS) and British Association of Plastic Reconstructive and Aesthetic Surgeons (BAPRAS) by the expert liposuction group. The guidance is based on the evidence available in the literature along with specialist expert opinion in aesthetic liposuction to provide plastic surgeons with consensus recommendation. The aim is to identify best practice to maximise the safety of patients. This article summarises current practices and safety considerations and outlines recommendations covering various aspects of patient care.

Quantitative assessment of tissue sections by Reverse Colour Coding.

Stains and dyes are frequently used to emphasise tissue structures for viewing under microscopy but few simple objective quantification methods exist. We describe the derivation of a mathematical formula enabling calculation of the relative contribution of three different coloured components to an image, which can be applied to rapid batch assessment of tissue sections for quantification of area proportions of differently stained elements. In order to validate this method, termed Reverse Colour Coding (RCC), we compared RCC estimations with known area proportions in artificially created images to calculate absolute accuracy, and compared RCC with panel visual estimation (VE) for the assessment of actual NCAM-stained muscle slides. Our results indicate that RCC has an absolute accuracy of 98-98.5% and superior inter-observer agreement and both inter- and intra-observer variability compared with VE. Results also suggest that cognitive bias occurring with VE may be eliminated by use of RCC. We submit RCC as a more accurate and less labour-intensive method of quantifying area proportions of stained tissues on microscopic images.

Granulocyte-macrophage colony-stimulating factor receptor expression in clinical pain disorder tissues and role in neuronal sensitization.

INTRODUCTION: Granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) is highly expressed in peripheral macrophages and microglia, and is involved in arthritis and cancer pain in animal models. However, there is limited information on GM-CSFR expression in human central nervous system (CNS), peripheral nerves, or dorsal root ganglia (DRG), particularly in chronic pain conditions. OBJECTIVES: Immunohistochemistry was used to quantify GM-CSFR expression levels in human tissues, and functional sensory effects of GM-CSF were studied in cultured DRG neurons. RESULTS: Granulocyte-macrophage colony-stimulating factor receptor was markedly increased in microglia at lesional sites of multiple sclerosis spinal cords (P = 0.01), which co-localised with macrophage marker CD68 (P = 0.009). In human DRG, GM-CSFR was expressed in a subset of small/medium diameter cells (30%) and few large cells (10%), with no significant change in avulsion-injured DRG. In peripheral nerves, there was a marked decrease in axonal GM-CSFR after chronic painful nerve injury (P = 0.004) and in painful neuromas (P = 0.0043); CD-68-positive macrophages were increased (P = 0.017) but did not appear to express GM-CSFR. Although control synovium showed absent GM-CSFR immunostaining, this was markedly increased in macrophages of painful osteoarthritis knee synovium. Granulocyte-macrophage colony-stimulating factor receptor was expressed in 17 ± 1.7% of small-/medium-sized cultured adult rat DRG neurons, and in 27 ± 3.3% of TRPV1-positive neurons. Granulocyte-macrophage colony-stimulating factor treatment sensitized capsaicin responses in vitro, which were diminished by p38 MAPK or TrkA inhibitors. CONCLUSION: Our findings support GM-CSFR as a therapeutic target for pain and hypersensitivity in clinical CNS and peripheral inflammatory conditions. Although GM-CSFR was decreased in chronic painful injured peripheral nerves, it could mediate CNS neuroinflammatory effects, which deserves study.

Glucagon-like peptide 1 receptor (GLP-1R) expression by nerve fibres in inflammatory bowel disease and functional effects in cultured neurons.

INTRODUCTION: Glucagon like-peptide 1 receptor (GLP-1R) agonists diminish appetite and may contribute to the weight loss in inflammatory bowel disease (IBD). OBJECTIVES: The aim of this study was to determine, for the first time, the expression of GLP-1R by colon nerve fibres in patients with IBD, and functional effects of its agonists in cultured rat and human sensory neurons. METHODS: GLP-1R and other nerve markers were studied by immunohistochemistry in colon biopsies from patients with IBD (n = 16) and controls (n = 8), human dorsal root ganglia (DRG) tissue, and in GLP-1R transfected HEK293 cells. The morphological effects of incretin hormones oxyntomodulin, exendin-4 and glucagon were studied on neurite extension in cultured DRG neurons, and their functional effects on capsaicin and ATP signalling, using calcium imaging. RESULTS: Significantly increased numbers of colonic mucosal nerve fibres were observed in IBD biopsies expressing GLP-1R (p = 0.0013), the pan-neuronal marker PGP9.5 (p = 0.0008), and sensory neuropeptide CGRP (p = 0.0014). An increase of GLP-1R positive nerve fibres in IBD colon was confirmed with a different antibody to GLP-1R (p = 0.016). GLP-1R immunostaining was intensely positive in small and medium-sized neurons in human DRG, and in human and rat DRG cultured neurons. Co-localization of GLP-1R expression with neuronal markers in colon and DRG confirmed the neural expression of GLP-1R, and antibody specificity was confirmed in HEK293 cells transfected with the GLP-1R. Treatment with oxyntomodulin, exendin-4 and GLP-1 increased neurite length in cultured neurons compared with controls, but did not stimulate calcium influx directly, or affect capsaicin responses. However, exendin-4 significantly enhanced ATP responses in human DRG neurons. CONCLUSION: Our results show that increased GLP-1R innervation in IBD bowel could mediate enhanced visceral afferent signalling, and provide a peripheral target for therapeutic intervention. The differential effect of GLP-1R agonists on capsaicin and ATP responses in neurons suggest they may not affect pain mechanisms mediated by the capsaicin receptor TRPV1, but may enhance the effects of purinergic agonists.

Free Functional Muscle Transfer Tendon Insertion Secondary Advancement Procedure to Improve Elbow Flexion.

One method of restoring elbow flexion in patients with brachial plexus injuries is through the use of free functional muscle transfer. Most of our patients achieve good outcome, resulting in improved usefulness of the affected limb. However, a minority of our patients have successful reinnervation of the transferred muscle but with a suboptimal strength and range of motion (ROM) of elbow flexion. In this subset of patients, we perform a secondary procedure to improve the moment arm of the muscle on the elbow joint. This involves advancing the transferred muscle distally along the radius. The technique of the advancement procedure is described. We also report on our initial series of 5 patients who underwent this advancement procedure. In one patient, there was improvement in strength alone. In another, there was improvement of the ROM alone. The remaining 3 patients had improvement of both strength and ROM.

Arterial and venous anatomical features of the pectoralis minor muscle flap pedicle.

The pectoralis minor muscle has been used as an innervated, vascularized, free-muscle graft in the field of facial reanimation for 20 years. Throughout this period, several centers have demonstrated consistent success with functional muscle transfer; however, opinions regarding the arterial pedicle of the flap have varied. The lateral thoracic and thoracoacromial arteries have been proposed as the predominant arterial sources. It has been the experience of our unit that a vessel (not described in anatomy textbooks) arising directly from the axillary artery and entering the muscle from its dorsal surface provides the dominant supply to the flap and is capable of sustaining it for free-tissue transfer. The vascular pedicle encountered was recorded and photographed in 97 consecutive cases in which the pectoralis minor muscle flap was raised. The findings demonstrated that the dominant supply to the muscle was from a single artery in 77 percent of cases and took the form of an artery arising directly from the axillary vessel in 72 percent of cases. More than one major arterial source was noted in the remainder of the cases. The venous outflow was usually through single or multiple veins running directly from the muscle into the axillary vein.

Reanimation of the middle and lower face in facial paralysis: review of the literature and personal approach.

Facial paralysis refers to a condition in which all or portions of the facial nerve are paralysed. The facial nerve controls the muscles of facial expression, paralysis which results in a lack of facial expression which is not only an aesthetic issue, but has functional consequences as the patient cannot communicate effectively. The treatment of long-standing facial paralysis has challenged plastic surgeons for centuries, and still the ultimate goal of normality of the paralysed hemi-face with symmetry at rest as well as the generation of a spontaneous symmetrical smile with corneal protection has not yet fully been reached. Until the end of the 19th century, the treatment of this condition involved non-surgical means such as ointments, medicines and electrotherapy. With the advent and refinement of microvascular surgical techniques in the latter half of the 20th century, vascularised free muscle transfers coupled with cross-facial nerve grafts were introduced, allowing the possibility of spontaneous emotion being restored to the paralysed face became reality. The aim of this article is to revisit the surgical evolution and current options available as well as outcomes for patients suffering from facial paralysis concentrating on middle and lower face reanimation.

The incidence of anterior belly of digastric agenesis in patients with hemifacial microsomia.

BACKGROUND: : The study examined the incidence of anterior belly of digastric muscle agenesis in patients with hemifacial microsomia, to determine the need for routine imaging of the floor of the mouth in patients within this group requesting dynamic lower lip reanimation. METHODS: : Patients presenting with microtia were assessed according to the OMENS [orbital deformity, mandibular hypoplasia, ear deformity, nerve (cranialnerve VII) involvement, and soft-tissue deficiency] criteria, and also imaged to establish the presence or absence of the anterior belly of the digastric muscle. Each affected hemiface was treated as an individual case (n = 50), with unaffected hemifaces being assigned as controls (an additional group of unaffected controls were also included). The hemifaces with microtia were then subdivided into three groups: those with isolated microtia (with anterior belly of the digastric muscle present), those with features consistent with hemifacial microsomia (with the anterior belly of the digastric muscle present), and those with absent anterior belly of the digastric muscle. RESULTS: : Anterior belly of the digastric muscle agenesis rate was 40 percent in those patients displaying features consistent with hemifacial microsomia. Median OMENS scores were significantly different in intergroup analysis (p < 0.0001), with a trend noticed between the median value for the hemifacial microsomia group and those with absent anterior belly of the digastric muscle. There were no noted incidences of anterior belly of the digastric muscle agenesis in the control group. CONCLUSIONS: : The incidence of anterior belly of the digastric muscle agenesis in patients with hemifacial microsomia is high. Before any attempt to undertake lower lip reanimation using this muscle, the floor of the mouth should be imaged to check for its presence.

Digitalization of the second finger in type 2 central longitudinal deficiencies (clefting) of the hand.

In central longitudinal deficiency of the hand type 2 (Manske and Halikis), the second finger presents itself anatomically and functionally as a second thumb. It is therefore necessary to undertake digitalization of the index, performed exactly as a reverse pollicization technique, with the same principles: minimum volar scarring and reconstruction of a large first web space without scars at the fold of the commissure. The incision surrounds the second digit at the level of the midproximal phalanx, extends over the dorsal edge of the cleft, and finishes on the radial side of the third finger where the second web space is to be created. Through this approach, the index metacarpal is freed (extraperiosteally), preserving the dorsal venous network, and translocated into the space of the missing third ray. After internal bone fixation, the flap, with its wide and safe volar cutaneous pedicle, is easily transposed to reconstruct the first web space, avoiding the need for skin grafting. This technique is easier and safer and does not impair the normal thumb musculature compared with the classic Snow-Littler procedure.

Functional muscle transfer and the variance of reinnervating axonal load: part I. The facial nerve.

BACKGROUND: Results of functional muscle transfer for the treatment of facial palsy vary. Surgical technique in such cases remains constant, with only the selected ramus of the buccal branch of the facial nerve changing. In this study, different-sized branches of the facial nerve were used to reinnervate a constant-sized muscle flap to see whether this might explain the spectrum of clinical results seen. METHODS: Twenty-four New Zealand White rabbits were divided into two groups of 12 animals. The rectus femoris muscle was raised as a free flap and transplanted to the cheek, after being coapted to a large (buccal) or small (marginal mandibular) branch of the facial nerve. Physiological testing of the flaps and histological examination of the muscle and motor nerve were undertaken. Half the animals from each group were euthanized at 6 months and the remaining at 9 months. In all cases, the contralateral rectus femoris muscle served as the control. RESULTS: Improved physiological results were noted in the groups reinnervated by the larger nerve at both time points. Significant improvement in force production was also noted between time points in the large nerve group. These results were confirmed by histological studies. CONCLUSION: Varying sizes of reinnervating axonal load may be responsible for the varied outcomes observed clinically and may also explain the phenomenon of late-onset tightening.

Functional muscle transfer and the variance of reinnervating axonal load: part II. Peripheral nerves.

BACKGROUND: Peripheral limb reconstruction using functional muscle transfer following injury or tumor resection has been widely reported in the literature. Such procedures often fail to deliver the physiologic contractile strength that might be hoped for in relation to the size of the transplanted muscle. Pure motor nerves of differing sizes were used to reinnervate a constant-sized muscle flap to see whether functional results could be improved in an experimental model analogous to clinical peripheral limb reconstruction. METHODS: Twenty-four New Zealand White rabbits were divided into two groups of 12 animals each. The rectus femoris muscle was raised as a free flap and transplanted back to the donor site orthotopically (coapted back to its original motor nerve) in the rectus femoris group or heterotopically (coapted to the larger motor nerve to the vastus lateralis-double the axonal load of the indigenous nerve) in the vastus lateralis group. Half the animals from each group were euthanized at 6 months, and the remaining animals were at 9 months. In all cases, the contralateral rectus femoris muscle served as a control. RESULTS: Physiologic assessment of the transplants for maximal tetanic contractural force was undertaken, together with immunohistochemical and histologic analysis of muscle and nerve tissue. Result analysis demonstrated that the double-sized nerve failed to produce significant improvements in physiologic outcome between the two groups at either the 6- or 9-month time point. The results were further endorsed by the histologic findings. CONCLUSION: Increasing the axonal load to a functional muscle transplant beyond that of its indigenous motor nerve does not improve outcome.

Neurovascular anatomy of the rectus femoris flap in the rabbit.

The rectus femoris muscle in the rabbit represents a reliable flap for experimental work in many aspects of microsurgery. The operative exposure of the flap is often tricky for those unfamiliar with the anatomy, as is isolation of its neurovascular pedicle. Experience with 59 consecutive rectus femoris flaps demonstrated five separate patterns of arteral and venous pedicles. The nervous anatomy was found to be constant. Once recognized, these vessel patterns can be used to facilitate pedicle dissection, thus minimizing intraoperative mistakes which may endanger the viability of the flap.