Neuroarthropathy in diabetes: pathogenesis of Charcot arthropathy.

Charcot neuroarthropathy is a rare but serious complication of diabetes, causing progressive destruction of the bones and joints of the foot leading to deformity, altered biomechanics and an increased risk of ulceration. Management is complicated by a lack of consensus on diagnostic criteria and an incomplete understanding of the pathogenesis. In this review, we consider recent insights into the development of Charcot neuroarthropathy. It is likely to be dependent on several interrelated factors which may include a genetic pre-disposition in combination with diabetic neuropathy. This leads to decreased neuropeptides (nitric oxide and calcitonin gene-related peptide), which may affect the normal coupling of bone formation and resorption, and increased levels of Receptor activator of nuclear factor kappa-B ligand, potentiating osteoclastogenesis. Repetitive unrecognized trauma due to neuropathy increases levels of pro-inflammatory cytokines (interleukin-1β, interleukin-6, tumour necrosis factor α) which could also contribute to increased bone resorption, in combination with a pre-inflammatory state, with increased autoimmune reactivity and a profile of monocytes primed to transform into osteoclasts - cluster of differentiation 14 (CD14). Increased blood glucose and loss of circulating Receptor for Advanced Glycation End-Products (AGLEPs), leading to increased non-enzymatic glycation of collagen and accumulation of AGLEPs in the tissues of the foot, may also contribute to the pathological process. An understanding of the relative contributions of each of these mechanisms and a final common pathway for the development of Charcot neuroarthropathy are still lacking. Cite this article: S. E. Johnson-Lynn, A. W. McCaskie, A. P. Coll, A. H. N. Robinson. Neuroarthropathy in diabetes: pathogenesis of Charcot arthropathy. Bone Joint Res 2018;7:373-378. DOI: 10.1302/2046-3758.75.BJR-2017-0334.R1

Tumor-Induced IL-6 Reprograms Host Metabolism to Suppress Anti-tumor Immunity

In patients with cancer, the wasting syndrome, cachexia, is associated with caloric deficiency. Here, we describe tumor-induced alterations of the host metabolic response to caloric deficiency that cause intratumoral immune suppression. In pre-cachectic mice with transplanted colorectal cancer or autochthonous pancreatic ductal adenocarcinoma (PDA), we find that IL-6 reduces the hepatic ketogenic potential through suppression of PPARalpha, the transcriptional master regulator of ketogenesis. When these mice are challenged with caloric deficiency, the resulting relative hypoketonemia triggers a marked rise in glucocorticoid levels. Multiple intratumoral immune pathways are suppressed by this hormonal stress response. Moreover, administering corticosterone to elevate plasma corticosterone to a level that is lower than that occurring in cachectic mice abolishes the response of mouse PDA to an immunotherapy that has advanced to clinical trials. Therefore, tumor-induced IL-6 impairs the ketogenic response to reduced caloric intake, resulting in a systemic metabolic stress response that blocks anti-cancer immunotherapy.We also thank the University of Cambridge, Cancer Research UK, the CRUK Cambridge Institute Core Facilities, and Hutchison Whampoa Limited. This work was also supported by the Lustgarten Foundation for Pancreatic Cancer Research, the Ludwig Institute for Cancer Research, the NIHR Biomedical Research Centre, and the Cambridge ECMC. T.R.F. was supported by the Rosetrees Trust and the Cambridge School of Clinical Medicine’s MB/PhD Programme, T.J. was supported by the Wellcome Trust Translational Medicine and Therapeutics Programme and the University of Cambridge Department of Oncology (RJAG/076), C.M.C. was supported by the Cambridge University Hospitals NHS Foundation Trust, E.W.R. was supported by the CRI Irvington Postdoctoral Fellowship Program, and A.P.C. was supported by the Medical Research Council (MRC) Metabolic Diseases Unit (MRC_MC_UU_12012/1). D.T.F. is a Distinguished Scholar of the Lustgarten Foundation

Imaging Mitochondrial Calcium Fluxes with Fluorescent Probes and Single- or Two-Photon Confocal Microscopy

The concentration of calcium ions in the mitochondria has been shown to affect its function, modulating respiratory activity at low levels and causing lethal damage at high concentrations. The rhodamine series of dyes can be used to measure mitochondrial calcium concentration, but the reliability of measurements depends upon correct partitioning of the dye within to the mitochondria. Methods are described to aid verification and quantification of the mitochondrial calcium concentration using single- or two-photon confocal microscopy. The method of linear unmixing to separate fluorescent signals based on either differing excitation or emission spectra is outlined and for the purposes of illustration is applied to the separation of rhod-2 signals originating from the dye within the mitochondria and nucleoli

Development of superlattice CrNNbN coatings for joint replacements deposited by High Power Impulse Magnetron Sputtering

The demand for reliable coating on medical implants is ever growing. In this research, enhanced performance of medical implants was achieved by a CrN/NbN coating utilising nanoscale multilayer/superlattice structure. The advantages of the novel High Power Impulse Magnetron Sputtering technology, namely its unique highly ionised plasma were exploited to deposit dense and strongly adherent coatings on Co-Cr implants. TEM analyses revealed coating superlattice structure with bi-layer thickness of 3.5 nm. CrN/NbN deposited on Co-Cr samples showed exceptionally high adhesion, critical load values of LC2= 50 N in scratch adhesion tests. Nanoindentation tests showed high hardness of 34 GPa and Young's modulus of 447 GPa. Low coefficient of friction (µ) 0.49 and coating wear coefficient (KC) = 4.94 x 10-16 m3N-1m-1 were recorded in dry sliding tests. Metal ion release studies showed a reduction in Co, Cr and Mo release at physiological and elevated temperatures, (70 oC) to almost undetectable levels (<1 ppb). Rotating beam fatigue testing showed a significant increase in fatigue strength from 349±59 MPa (uncoated) to 539±59 MPa (coated). In vitro biological testing has been performed in order to assess the safety of the coating in biological environment, cytotoxicity, genotoxicity and sensitisation testing have been performed, all showing no adverse effects. Keywords: Orthopaedic implant, High Power Impulse Magnetron Sputtering, Superlattice coating, Corrosion, Biocompatibility

Early onset obesity and adrenal insufficiency associated with a homozygous POMC mutation

Isolated hypocortisolism due to ACTH deficiency is a rare condition that can be caused by homozygous or compound heterozygous mutations in the gene encoding proopiomelanocortin (POMC). Loss of function mutations of POMC gene typically results in adrenal insufficiency, obesity and red hair. We describe an 18 month old Hispanic female with congenital adrenal insufficiency, a novel POMC mutation and atypical clinical features. The patient presented at the age of 9 months with hypoglycemia and the endocrine evaluation resulted in a diagnosis of ACTH deficiency. She developed extreme weight gain prompting sequence analysis of POMC, which revealed a homozygous c.231C > A change which is predicted to result in a premature termination codon. The case we report had obesity, hypocortisolism but lacked red hair which is typical for subjects with POMC mutations. Mutations of POMC should be considered in individuals with severe early onset obesity and adrenal insufficiency even when they lack the typical pigmentary phenotype

T wave abnormalities, high body mass index, current smoking and high lipoprotein (a) levels predict the development of major abnormal Q/QS patterns 20 years later. A population-based study

BACKGROUND: Most studies on risk factors for development of coronary heart disease (CHD) have been based on the clinical outcome of CHD. Our aim was to identify factors that could predict the development of ECG markers of CHD, such as abnormal Q/QS patterns, ST segment depression and T wave abnormalities, in 70-year-old men, irrespective of clinical outcome. METHODS: Predictors for development of different ECG abnormalities were identified in a population-based study using stepwise logistic regression. Anthropometrical and metabolic factors, ECG abnormalities and vital signs from a health survey of men at age 50 were related to ECG abnormalities identified in the same cohort 20 years later. RESULTS: At the age of 70, 9% had developed a major abnormal Q/QS pattern, but 63% of these subjects had not been previously hospitalized due to MI, while 57% with symptomatic MI between age 50 and 70 had no major Q/QS pattern at age 70. T wave abnormalities (Odds ratio 3.11, 95% CI 1.18–8.17), high lipoprotein (a) levels, high body mass index (BMI) and smoking were identified as significant independent predictors for the development of abnormal major Q/QS patterns. T wave abnormalities and high fasting glucose levels were significant independent predictors for the development of ST segment depression without abnormal Q/QS pattern. CONCLUSION: T wave abnormalities on resting ECG should be given special attention and correlated with clinical information. Risk factors for major Q/QS patterns need not be the same as traditional risk factors for clinically recognized CHD. High lipoprotein (a) levels may be a stronger risk factor for silent myocardial infarction (MI) compared to clinically recognized MI

Loss of Mrap2 is associated with Sim1 deficiency and increased circulating cholesterol.

Melanocortin receptor accessory protein 2 (MRAP2) is a transmembrane accessory protein predominantly expressed in the brain. Both global and brain-specific deletion of Mrap2 in mice results in severe obesity. Loss-of-function MRAP2 mutations have also been associated with obesity in humans. Although MRAP2 has been shown to interact with MC4R, a G protein-coupled receptor with an established role in energy homeostasis, appetite regulation and lipid metabolism, the mechanisms through which loss of MRAP2 causes obesity remains uncertain. In this study, we used two independently derived lines of Mrap2 deficient mice (Mrap2(tm1a/tm1a)) to further study the role of Mrap2 in the regulation of energy balance and peripheral lipid metabolism. Mrap2(tm1a/tm1a) mice have a significant increase in body weight, with increased fat and lean mass, but without detectable changes in food intake or energy expenditure. Transcriptomic analysis showed significantly decreased expression of Sim1, Trh, Oxt and Crh within the hypothalamic paraventricular nucleus of Mrap2(tm1a/tm1a) mice. Circulating levels of both high-density lipoprotein and low-density lipoprotein were significantly increased in Mrap2 deficient mice. Taken together, these data corroborate the role of MRAP2 in metabolic regulation and indicate that, at least in part, this may be due to defective central melanocortin signalling

Heterogeneity of hypothalamic pro-opiomelanocortin-expressing neurons revealed by single-cell RNA sequencing

$\textbf{Objective}$ Arcuate proopiomelanocortin (POMC) neurons are critical nodes in the control of body weight. Often characterized simply as direct targets for leptin, recent data suggest a more complex architecture. $\textbf{Methods}$ Using single cell RNA sequencing, we have generated an atlas of gene expression in murine POMC neurons. $\textbf{Results}$ Of 163 neurons, 118 expressed high levels of $\textit{Pomc}$ with little/no Agrp expression and were considered “canonical” POMC neurons (P$^{+}$). The other 45/163 expressed low levels of $\textit{Pomc}$ and high levels of $\textit{Agrp}$ (A$^{+}$P$_{+}$). Unbiased clustering analysis of P$^{+}$ neurons revealed four different classes, each with distinct cell surface receptor gene expression profiles. Further, only 12% (14/118) of P$^{+}$ neurons expressed the leptin receptor ($\textit{Lepr}$) compared with 58% (26/45) of A$^{+}$P$_{+}$ neurons. In contrast, the insulin receptor ($\textit{Insr}$) was expressed at similar frequency on P$^{+}$ and A$^{+}$P$_{+}$ neurons (64% and 55%, respectively). $\textbf{Conclusion}$ These data reveal arcuate POMC neurons to be a highly heterogeneous population. Accession Numbers: GSE92707.This work was supported by the UK Medical Research Council (MRC) Metabolic Disease Unit (MRC_MC_UU_12012/1 & MRC_MC_UU_12012/5), a Wellcome Trust Strategic Award (100574/Z/12/Z), and the Helmholtz Alliance ICEMED

The obesity-associated gene TMEM18 has a role in the central control of appetite and body weight regulation

An intergenic region of human chromosome 2 (2p25.3) harbors genetic variants which are among those most strongly and reproducibly associated with obesity. The gene closest to these variants is TMEM18, although the molecular mechanisms mediating these effects remain entirely unknown. Tmem18 expression in the murine hypothalamic paraventricular nucleus (PVN) was altered by changes in nutritional state. Germline loss of Tmem18 in mice resulted in increased body weight, which was exacerbated by high fat diet and driven by increased food intake. Selective overexpression of Tmem18 in the PVN of wild-type mice reduced food intake and also increased energy expenditure. We provide evidence that TMEM18 has four, not three, transmembrane domains and that it physically interacts with key components of the nuclear pore complex. Our data support the hypothesis that TMEM18 itself, acting within the central nervous system, is a plausible mediator of the impact of adjacent genetic variation on human adiposity.RL, YCLT, DR, GSHY, SOR and APC are funded by the Medical Research Council (MRC) Metabolic Disease Unit (MRC_MC_UU_12012/1) and animal work was carried out with the assistance of MRC Disease Model Core of the Wellcome Trust MRC Institute of Metabolic Sciences (MRC_MC_UU_12012/5 and Wellcome Trust Strategic Award (100574/Z/12/Z). F. Bosch is the recipient of an award from the ICREA Academia, Generalitat de Catalunya, Spain. Vector generation and production were funded by Ministerio de Economía y Competitividad (SAF 2014-54866-R), Spain. CD and DWL were supported by the Wellcome Trust (WT098051) and CD was supported by the Wellcome Trust PhD Programme for Clinicians (100679/Z/12/Z)