Non-battle injury among repatriated UK armed forces since cessation of combat operations: a prospective observational study.

AIM: To describe the mechanisms, burden of injury, inpatient management and rehabilitation requirements of wounded military personnel at the UK Role 4 (R4) facility within the first 12 months following cessation of combat operations in Afghanistan. METHODS: All aeromedical evacuations were recorded prospectively between October 2014 and October 2015. Demographic, logistical and clinical data were derived manually from referring medical unit and patient movement requests in addition to host nation and R4 medical records. RESULTS: Ninety-five patients were repatriated to R4 following traumatic injury: 98.9% (n=94) were male, and median age was 27 years (IQR 25-36 years). The most common mechanisms of injury (MOIs) were sports 26.3% (n=25), falls <2 m 11.6% (n=11) and road traffic collisions 9.8% (n=9). The most common anatomical regions of injury were isolated lower limb 24.1% (n=22), isolated hand 20.0% (n=19) and polytrauma 14.7% (n=14). Median Injury Severity Score was 4 (IQR 4-9), mean 8 (range 1-41). Eleven patients (11.6%) were discharged to rehabilitation units, of whom 7 (63.6%) required neurorehabilitation. CONCLUSION: Although service personnel sustain civilian-type injuries, the specific rehabilitation goals and shift in the acute rehabilitation requirements for military personnel must be considered in the absence of enduring combat operations. It is notable that permanent medical downgrading secondary to trauma still occurs outside of warfare. The colocation of civilian major trauma services and R4 has ensured a mutually beneficial partnership that contributes to institutional memory and improves the coordination of patient pathways. The importance of relevant resource allocation, training, support and logistical considerations remain, even during the current scale of military activity overseas.

Targeting protein kinase C in sarcoma.

Protein kinase C (PKC) is a family of serine/threonine tyrosine kinases that regulate many cellular processes including division, proliferation, survival, anoikis and polarity. PKC is abundant in many human cancers and aberrant PKC signalling has been demonstrated in cancer models. On this basis, PKC has become an attractive target for small molecule inhibition within oncology drug development programmes. Sarcoma is a heterogeneous group of mesenchymal malignancies. Due to their relative insensitivity to conventional chemotherapies and the increasing recognition of the driving molecular events of sarcomagenesis, sarcoma provides an excellent platform to test novel therapeutics. In this review we provide a structure-function overview of the PKC family, the rationale for targeting these kinases in sarcoma and the state of play with regard to PKC inhibition in the clinic.

Abdominal Aortic Junctional Tourniquet - Stabilized (AAJTS) can be applied both successfully and rapidly by Combat Medical Technicians (CMTs).

BACKGROUND: 'Non-compressible' haemorrhage is the leading cause of preventable battlefield death, often requiring surgical or radiological intervention, which is precluded in the pre-hospital environment. One-fifth of such bleeds are junctional and therefore potentially survivable. We examine the use of the Abdominal Aortic Junctional Tourniquet - Stabilized (AAJTS) among UK Combat Medical Technicians (CMTs) as a device to control junctional haemorrhage with external compression of the abdominal aorta-compression of junctional haemorrhage previously considered 'non-compressible.' This follows animal studies showing that the AAJTS achieves control of haemorrhage and improves physiological parameters. METHODS: CMTs were selected and applied the AAJTS to each other following a 1-hour training package. A consultant radiologist-operated hand-held ultrasound monitored flow changes in the subjects' common femoral artery. CMTs were then surveyed for their opinions as to utility and function. RESULTS: 21 CMTs were screened and 17 CMTs participated with 34 total applications (16 day and 18 low-light). 27/34 (79%) achieved a successful application. The median application time was 75 s in daylight and 57 s in low-light conditions. There was no significant difference in Body Mass Index (p=0.23), median systolic blood pressure (p=0.19), nor class of CMT (p=0.10) between successful and unsuccessful applications. Higher systolic blood pressure was associated with longer application times (p=0.03). Users deemed the device easy to use (median score 4.4 on a 5-point Likert scale). CONCLUSION: CMTs can use AAJTS successfully after a 1-hour training session in the majority of applications. Application was successful in both daylight and low-light conditions. Self-reported usability ratings were high.

Initial extremity war wound debridement: a multidisciplinary consensus.

A joint meeting of the Limb Trauma and Wounds Working Groups resulted in the establishment of 29 consensus recommendations for the conduct of initial extremity war wound debridement. Pre-operative, operative and post-operative phases of debridement were considered along with wound irrigation and dressings. Wounds where a different surgical approach is required, such as superficial soft tissue wounds at one end of the spectrum and complex wounds sustained in close proximity to explosions at the other, were also discussed. The recommendations represent the consensus opinion of orthopaedic, vascular and plastic surgeons, as well as nursing officers, from across the Defence Medical Services and are intended to provide useful guidance to the deploying surgeon, regardless of their own personal experience.

Protein kinase Cepsilon is required for macrophage activation and defense against bacterial infection.

To assess directly the role of protein kinase C (PKC)epsilon in the immune system, we generated mice that carried a homozygous disruption of the PKCepsilon locus. PKCepsilon(-/-) animals appeared normal and were generally healthy, although female mice frequently developed a bacterial infection of the uterus. Macrophages from PKCepsilon(-/-) animals demonstrated a severely attenuated response to lipopolysaccharide (LPS) and interferon (IFN)gamma, characterized by a dramatic reduction in the generation of NO, tumor necrosis factor (TNF)-alpha, and interleukin (IL)-1beta. Further analysis revealed that LPS-stimulated macrophages from PKCepsilon(-/-) mice were deficient in the induction of nitric oxide synthase (NOS)-2, demonstrating a decrease in the activation of IkappaB kinase, a reduction in IkappaB degradation, and a decrease in nuclear factor (NF)kappaB nuclear translocation. After intravenous administration of Gram-negative or Gram-positive bacteria, PKCepsilon(-/-) mice demonstrated a significantly decreased period of survival. This study provides direct evidence that PKCepsilon is critically involved at an early stage of LPS-mediated signaling in activated macrophages. Furthermore, we demonstrate that in the absence of PKCepsilon, host defense against bacterial infection is severely compromised, resulting in an increased incidence of mortality.

The complete primary structure of protein kinase C--the major phorbol ester receptor.

Protein kinase C, the major phorbol ester receptor, was purified from bovine brain and through the use of oligonucleotide probes based on partial amino acid sequence, complementary DNA clones were derived from bovine brain complementary DNA libraries. Thus, the complete amino acid sequence of bovine protein kinase C was determined, revealing a domain structure. At the amino terminal is a cysteine-rich domain with an internal duplication; a putative calcium-binding domain follows, and there is at the carboxyl terminal a domain that shows substantial homology, but not identity, to sequences of other protein kinase.

Multiple, distinct forms of bovine and human protein kinase C suggest diversity in cellular signaling pathways.

A new family of protein kinase C-related genes has been identified in bovine, human, and rat genomes. The alpha-, beta-, and gamma-type protein kinase sequences are highly homologous, include a kinase domain, and potential calcium-binding sites, and they contain interspersed variable regions. The corresponding genes are located on distinct human chromosomes; the possibility of even greater genetic complexity of this gene family is suggested by Northern and Southern hybridization analyses.

Protein kinase C isotypes controlled by phosphoinositide 3-kinase through the protein kinase PDK1.

Phosphorylation sites in members of the protein kinase A (PKA), PKG, and PKC kinase subfamily are conserved. Thus, the PKB kinase PDK1 may be responsible for the phosphorylation of PKC isotypes. PDK1 phosphorylated the activation loop sites of PKCzeta and PKCdelta in vitro and in a phosphoinositide 3-kinase (PI 3-kinase)-dependent manner in vivo in human embryonic kidney (293) cells. All members of the PKC family tested formed complexes with PDK1. PDK1-dependent phosphorylation of PKCdelta in vitro was stimulated by combined PKC and PDK1 activators. The activation loop phosphorylation of PKCdelta in response to serum stimulation of cells was PI 3-kinase-dependent and was enhanced by PDK1 coexpression.

Imaging protein kinase Calpha activation in cells.

Spatially resolved fluorescence resonance energy transfer (FRET) measured by fluorescence lifetime imaging microscopy (FLIM), provides a method for tracing the catalytic activity of fluorescently tagged proteins inside live cell cultures and enables determination of the functional state of proteins in fixed cells and tissues. Here, a dynamic marker of protein kinase Calpha (PKCalpha) activation is identified and exploited. Activation of PKCalpha is detected through the binding of fluorescently tagged phosphorylation site-specific antibodies; the consequent FRET is measured through the donor fluorophore on PKCalpha by FLIM. This approach enabled the imaging of PKCalpha activation in live and fixed cultured cells and was also applied to pathological samples.

Training tomorrow's military surgeons: lessons from the past and challenges for the future.

The nature of conflict is evolving, with current warfare being associated with an initial "shock and awe" phase followed by protracted periods ofcounter-insurgency and peace support missions. As conflict has changed, so have the munitions deployed and the resulting patterns of injury. Improvised Explosive Devices have become the preferred weapon of the insurgent and the resultant explosive and fragmentation injuries are the hallmark of modern military wounding. These injuries pose a significant challenge to deployed medical forces, requiring a well-defined, seamless approach from injury to rehabilitation. Traditionally, military medical services demonstrate a poor 'institutional memory' in the maintenance of combat surgical skills. Numerous publications detail the re-learning of key tenets of war surgery by generations of surgeons deploying onto the field of battle. While the maintenance of military surgical capability in trained surgeons may be addressed through combat surgical courses, concern exists as to the generic competency of those currently in training and their ability to deal with the burden of injury associated with modern conflict. The training of junior doctors in the United Kingdom and further afield is in a state of flux. New curriculum development, streamlined and run-through training programmes have combined with the legal requirements of the European Working Time Directive to produce a training landscape almost unrecognisable with that of previous years. This article investigates the development of current military wounding patterns and modern surgical training programmes. It describes processes already in place to address the unique training needs of military surgeons and proposes a framework for enabling appropriate training opportunities in the future.

Optimal clinical timelines--a consensus from the academic department of military surgery and trauma.

There are no published studies directly addressing the issue of what is an acceptable timeline from point of wounding to surgical intervention within the military context. The proximal threshold has previously been determined by personal opinion, tactical, logistic and practical imperatives rather than by clinical demands. The aim of this paper is to review all relevant military and civilian studies where timelines have been quoted and to reach a number of unambiguous consensus statements to state the perceived ideal upper limits from point of wounding to holistic and realistic surgical care in modern war. An injured casualty should be transferred to an appropriate surgeon in an appropriate facility in as short a time from wounding as practical. It is clear that the best trauma surgery is performed in large, well resourced, well-supplied, air-conditioned hospitals. Current advances aimed to stretch timelines from wounding to surgical intervention are exciting and hold potential but remain scientifically unproven and are currently without any firm evidence base. Further critical research is therefore necessary. The effect of pre-hospital haemostatic resuscitation, provided by the enhanced Medical Emergency Response Team (MERTe) on patient outcome and effective timelines is currently unknown and unproven: it does have intuitive medical merit. There is also a very significant moral and morale component. MERTe serves two main functions; reduction in time from point of wounding to advanced / haemostatic resuscitation and provision of in-flight diagnostics. Continuation of in-flight resuscitation then allows physician-led decision making on critically unstable casualties. This allows either an expedited straight move from the HLS direct to the operating theatre or direct transfer to a regional neurosurgical centre. To prevent avoidable death,our unequivocal conclusion is that there must be an upper limit of 2 hours from wounding to surgical intervention (surgical haemorrhage control) for all casualties.

Inositol lipid 5-phosphatases--traffic signals and signal traffic.

Several novel phosphoinositide 5-phosphatases have been identified and characterised, revealing a growing family of regulators of inositol lipid dependent processes. The features of these proteins, their likely partners and their involvement in signal transduction and membrane traffic is discussed.

Protein kinase C--a question of specificity.

Following the initial identification of protein kinase C (PKC) by Nishizuka and co-workers in the late seventies, a wealth of information on this protein kinase has accumulated. Perhaps most striking was the realization that PKC is not just a single polypeptide but in fact consists of a large family of related proteins. These PKC isotypes are unique, not only with respect to primary structure, but also on the basis of expression patterns, subcellular localization, activation in vitro and responsiveness to extra-cellular signals. This review focuses on the heterogeneity within the PKC family and highlights some of the recent evidence that the isotypes might have separate and unique functions in the cell.

Specific involvement of PKC-epsilon in sensitization of the neuronal response to painful heat.

Pain is unique among sensations in that the perceived intensity increases, or sensitizes, during exposure to a strong stimulus. One important mediator of sensitization is bradykinin (BK), a peptide released as a consequence of tissue damage. BK enhances the membrane ionic current activated by heat in nociceptive neurons, using a pathway that involves activation of protein kinase C (PKC). We find that five PKC isoforms are present in sensory neurons but that only PKC-epsilon is translocated to the cell membrane by BK. The heat response is sensitized when constitutively active PKC-epsilon is incorporated into nociceptive neurons. Conversely, BK-induced sensitization is suppressed by a specific peptide inhibitor of PKC-epsilon. We conclude that PKC-epsilon is principally responsible for sensitization of the heat response in nociceptors by bradykinin.

Spatially selective sampling of single cells using optically trapped fusogenic emulsion droplets: a new single-cell proteomic tool.

We present a platform for the spatially selective sampling of the plasma membrane of single cells. Optically trapped lipid-coated oil droplets (smart droplet microtools, SDMs), typically 0.5-5 microm in size, composed of a hexadecane hydrocarbon core and fusogenic lipid outer coating (mixture of 1,2-dioleoyl-phosphatidylethanolamine and 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine) were brought into controlled contact with target colon cancer cells leading to the formation of connecting membrane tethers. Material transfer from the cell to the SDM across the membrane tether was monitored by tracking membrane-localized enhanced green fluorescent protein.

Femoral neck stress fractures in military personnel--a case series.

Femoral neck stress fractures (FNSF) are uncommon, representing 5% of all stress fractures. In military personnel, FNSF represents one of the more severe complications of training, which can result in medical discharge. Clinical examination findings are often non-specific and plain radiography may be inconclusive--leading to missed or late diagnosis of FNSF This paper highlights the significance of FNSFs in military personnel and alerts physicians to the potential diagnosis. We identified all military recruits, aged 17 to 26, who attended the Infantry Training Centre (Catterick, U.K.), over a four-year period from the 1st July 2002 to 30th June 2006, who had suffered a FNSF. The medical records, plain radiographs, bone scans and MRIs of the recruits were retrospectively reviewed. Of 250 stress fractures 20 were of the femoral neck; representing 8% of all stress fractures and an overall FNSF rate of 12 in 10,000 military recruits. FNSFs were most prevalent amongst Parachute Regiment recruits (1 in 250, p < 0.05). Onset of symptoms was most commonly between weeks 13-16 of training. The majority (17/20, 85%) of FNSFs were undisplaced, these were all treated conservatively. Three FNSFs were displaced on presentation and were treated surgically. Overall, the medical discharge rate was 40% (8/20). FNSFs are uncommon and the diagnosis remains a challenge to clinicians and requires a high index of suspicion in these young athletic individuals. In such individuals early referral for MRI is recommended, to aid prompt diagnosis and treatment and to prevent more serious sequelae.

Intracellular signalling. PI 3-kinase puts GTP on the Rac.

Phosphoinositide 3-kinase, an enzyme that is known to transduce signals received by a variety of receptor types, has been found to mediate agonist-dependent membrane ruffling via the small GTP-binding protein Rac.

Regulation of epidermal growth factor receptor traffic by the small GTPase rhoB.

Members of the Rho family of small GTPases control cell adhesion and motility through dynamic regulation of the actin cytoskeleton. Although twelve family members have been identified, only three of these - RhoA, Rac and Cdc42 - have been studied in detail. RhoA regulates the formation of focal adhesions and the bundling of actin filaments into stress fibres. It is also involved in other cell signalling pathways including the regulation of gene expression and the generation of lipid second messengers [1] [2]. RhoA is very closely related to two other small GTPases about which much less is known: RhoB and RhoC (which are approximately 83% identical). Perhaps the most intriguing of these is RhoB. RhoA is largely cytosolic but translocates to the plasma membrane on activation. RhoB, however, is entirely localised to the cytosolic face of endocytic vesicles [3] [4]. This suggests a potential role for RhoB in regulating endocytic traffic; however, no evidence has been presented to support this. RhoA has been shown to act at the plasma membrane to regulate the clathrin-mediated internalisation of transferrin receptor [5] and of the muscarinic acetylcholine receptor [6]. We have recently demonstrated that RhoB binds the RhoA effector, PRK1 and targets it to the endosomal compartment [7]. We show here that RhoB acts through PRK1 to regulate the kinetics of epidermal growth factor receptor traffic.

Phosphorylation of threonine 638 critically controls the dephosphorylation and inactivation of protein kinase Calpha.

BACKGROUND: It has been widely reported that multisite phosphorylation plays an essential role in the regulation of protein kinases. However, our understanding of how these events modify protein function in vitro and in vivo is poorly understood. Protein kinase C (PKC) affords an interesting example of how phosphorylation control is coupled to effector control. PKC is acutely regulated by the second messenger diacylglycerol; however, it is also known to undergo multisite phosphorylation. Previously, we and others have shown that one site in the 'activation loop' of PKCalpha (a threonine residue at position 497; T497) and PKCbeta (T500) is essential for the catalytic competence of these proteins. More recently, a carboxy-terminal site (T638 in PKCalpha) has been implicated. In this report, we investigate the role of this site and its interaction with the catalytic core site. RESULTS: We have analyzed mutant PKCalpha proteins, in which amino-acid substitutions were made at the T638 site, and shown that phosphorylation at this site affects the conformation of the protein, as judged by thermal stability, and sensitivity to oxidation, trypsin and phosphatase treatment. This supersensitivity to dephosphorylation in vitro was also seen in an agonist-dependent context in vivo. We have also shown that phosphorylation of this site is not essential for catalytic activity of the purified protein. The molecular basis of the control operating through the T638 site was provided by the evidence of a functional interaction with the previously described catalytic core site, T497. This inter-relationship was further established by the demonstration that the E497 mutant protein had a thermal instability and phosphatase supersensitivity similar to that of the A638 and E638 mutants. CONCLUSIONS: The T638 phosphorylation site is not required for the catalytic function of PKCalpha per se, but serves to control the duration of activation by regulating the rate of dephosphorylation and inactivation of the protein. This is achieved through the cooperative interaction between the T638 and T497 sites; if either of these residues is not phosphorylated, the protein is supersensitive to phosphatase action. This model of PKCalpha function is likely to be of general significance to the protein kinase superfamily, where similarly juxtaposed sites exist. We conclude that dephosphorylation of PKCalpha, and, by inference, other protein kinases, is regulated by multisite phosphorylation.

The activation of phosphatidylinositol 3-kinase by Ras.

BACKGROUND: Activation of the mammalian phosphatidylinositol 3-kinase complex can play a critical role in transducing growth factor responses. The lipid kinase complex, which is made up of p85 alpha and p110 alpha regulatory and catalytic subunits, becomes associated with a number of activated receptor protein tyrosine kinases, but the mechanism of its activation has not yet been defined. Recent evidence indicates that Ras can bind to the p85 alpha/p110 alpha complex. We describe here the functional regulation of the mammalian phosphatidylinositol 3-kinase complex by Ras. RESULTS: Expression of p110 alpha, the catalytic subunit of phosphatidylinositol 3-kinase, in the fission yeast, Schizosaccharomyces pombe, has been used to demonstrate an inhibitory effect of p85 alpha on p110 alpha activity in intact cells; inhibition did not result from a decrease in p110 alpha expression. In this cellular context, we have investigated the effect of a constitutively active mutant of Ras, v-Ras, either on p85 alpha or p110 alpha-alone, or on the p85 alpha/p110 alpha complex. In the presence of the p85 alpha/p110 alpha complex, v-Ras suppressed cell growth, but an effector-domain mutant of v-Ras did not. The growth-suppressive effect of v-Ras was not seen for any other combination of expressed proteins. The phenotype induced by v-Ras was consistent with activation of the p85 alpha/p110 alpha complex: it was sensitive to the phosphatidylinositol 3-kinase inhibitor, wortmannin, and the cells accumulated 3-phosphorylated polyphosphoinositides. Activation of purified p85 alpha/p110 alpha by purified recombinant Ras in vitro was also demonstrated. CONCLUSIONS: The phosphatidylinositol 3-kinase complex, p85 alpha/p110 alpha, shows a suppressed catalytic function in vivo when compared with free p110 alpha. This complex can, however, be activated by Ras. We suggest that the phosphatidylinositol 3-kinase p85 alpha/p110 alpha complex is a downstream effector of Ras.