Endotoxaemia in Haemodialysis: A Novel Factor in Erythropoetin Resistance?

Background/Objectives Translocated endotoxin derived from intestinal bacteria is a driver of systemic inflammation and oxidative stress. Severe endotoxaemia is an underappreciated, but characteristic finding in haemodialysis (HD) patients, and appears to be driven by acute repetitive dialysis induced circulatory stress. Resistance to erythropoietin (EPO) has been identified as a predictor of mortality risk, and associated with inflammation and malnutrition. This study aims to explore the potential link between previously unrecognised endotoxaemia and EPO Resistance Index (ERI) in HD patients. Methodology/Principal Findings 50 established HD patients were studied at a routine dialysis session. Data collection included weight, BMI, ultrafiltration volume, weekly EPO dose, and blood sampling pre and post HD. ERI was calculated as ratio of total weekly EPO dose to body weight (U/kg) to haemoglobin level (g/dL). Mean haemoglobin (Hb) was 11.3±1.3 g/dL with a median EPO dose of 10,000 [IQR 7,500–20,000] u/wk and ERI of 13.7 [IQR 6.9–23.3] ((U/Kg)/(g/dL)). Mean pre-HD serum ET levels were significantly elevated at 0.69±0.30 EU/ml. Natural logarithm (Ln) of ERI correlated to predialysis ET levels (r = 0.324, p = 0.03) with a trend towards association with hsCRP (r = 0.280, p = 0.07). Ln ERI correlated with ultrafiltration volume, a driver of circulatory stress (r = 0.295, p = 0.046), previously identified to be associated with increased intradialytic endotoxin translocation. Both serum ET and ultrafiltration volume corrected for body weight were independently associated with Ln ERI in multivariable analysis. Conclusions This study suggests that endotoxaemia is a significant factor in setting levels of EPO requirement. It raises the possibility that elevated EPO doses may in part merely be identifying patients subjected to significant circulatory stress and suffering the myriad of negative biological consequences arising from sustained systemic exposure to endotoxin

Transcriptome-Wide Identification of Novel Imprinted Genes in Neonatal Mouse Brain

Imprinted genes display differential allelic expression in a manner that depends on the sex of the transmitting parent. The degree of imprinting is often tissue-specific and/or developmental stage-specific, and may be altered in some diseases including cancer. Here we applied Illumina/Solexa sequencing of the transcriptomes of reciprocal F1 mouse neonatal brains and identified 26 genes with parent-of-origin dependent differential allelic expression. Allele-specific Pyrosequencing verified 17 of them, including three novel imprinted genes. The known and novel imprinted genes all are found in proximity to previously reported differentially methylated regions (DMRs). Ten genes known to be imprinted in placenta had sufficient expression levels to attain a read depth that provided statistical power to detect imprinting, and yet all were consistent with non-imprinting in our transcript count data for neonatal brain. Three closely linked and reciprocally imprinted gene pairs were also discovered, and their pattern of expression suggests transcriptional interference. Despite the coverage of more than 5000 genes, this scan only identified three novel imprinted refseq genes in neonatal brain, suggesting that this tissue is nearly exhaustively characterized. This approach has the potential to yield an complete catalog of imprinted genes after application to multiple tissues and developmental stages, shedding light on the mechanism, bioinformatic prediction, and evolution of imprinted genes and diseases associated with genomic imprinting

The importance of imprinting in the human placenta.

As a field of study, genomic imprinting has grown rapidly in the last 20 years, with a growing figure of around 100 imprinted genes known in the mouse and approximately 50 in the human. The imprinted expression of genes may be transient and highly tissue-specific, and there are potentially hundreds of other, as yet undiscovered, imprinted transcripts. The placenta is notable amongst mammalian organs for its high and prolific expression of imprinted genes. This review discusses the development of the human placenta and focuses on the function of imprinting in this organ. Imprinting is potentially a mechanism to balance parental resource allocation and it plays an important role in growth. The placenta, as the interface between mother and fetus, is central to prenatal growth control. The expression of genes subject to parental allelic expression bias has, over the years, been shown to be essential for the normal development and physiology of the placenta. In this review we also discuss the significance of genes that lack conservation of imprinting between mice and humans, genes whose imprinted expression is often placental-specific. Finally, we illustrate the importance of imprinting in the postnatal human in terms of several human imprinting disorders, with consideration of the brain as a key organ for imprinted gene expression after birth

Postnatal Changes in the Expression Pattern of the Imprinted Signalling Protein XLαs Underlie the Changing Phenotype of Deficient Mice

The alternatively spliced trimeric G-protein subunit XLαs, which is involved in cAMP signalling, is encoded by the Gnasxl transcript of the imprinted Gnas locus. XLαs deficient mice show neonatal feeding problems, leanness, inertia and a high mortality rate. Mutants that survive to weaning age develop into healthy and fertile adults, which remain lean despite elevated food intake. The adult metabolic phenotype can be attributed to increased energy expenditure, which appears to be caused by elevated sympathetic nervous system activity. To better understand the changing phenotype of Gnasxl deficient mice, we compared XLαs expression in neonatal versus adult tissues, analysed its co-localisation with neural markers and characterised changes in the nutrient-sensing mTOR1-S6K pathway in the hypothalamus. Using a newly generated conditional Gnasxl lacZ gene trap line and immunohistochemistry we identified various types of muscle, including smooth muscle cells of blood vessels, as the major peripheral sites of expression in neonates. Expression in all muscle tissues was silenced in adults. While Gnasxl expression in the central nervous system was also developmentally silenced in some midbrain nuclei, it was upregulated in the preoptic area, the medial amygdala, several hypothalamic nuclei (e.g. arcuate, dorsomedial, lateral and paraventricular nuclei) and the nucleus of the solitary tract. Furthermore, expression was detected in the ventral medulla as well as in motoneurons and a subset of sympathetic preganglionic neurons of the spinal cord. In the arcuate nucleus of Gnasxl-deficient mice we found reduced activity of the nutrient sensing mTOR1-S6K signalling pathway, which concurs with their metabolic status. The expression in these brain regions and the hypermetabolic phenotype of adult Gnasxl-deficient mice imply an inhibitory function of XLαs in energy expenditure and sympathetic outflow. By contrast, the neonatal phenotype of mutant mice appears to be due to a transient role of XLαs in muscle tissues

Antagonistic roles in fetal development and adult physiology for the oppositely imprinted Grb10 and Dlk1 genes

BACKGROUND: Despite being a fundamental biological problem the control of body size and proportions during development remains poorly understood, although it is accepted that the insulin-like growth factor (IGF) pathway has a central role in growth regulation, probably in all animals. The involvement of imprinted genes has also attracted much attention, not least because two of the earliest discovered were shown to be oppositely imprinted and antagonistic in their regulation of growth. The Igf2 gene encodes a paternally expressed ligand that promotes growth, while maternally expressed Igf2r encodes a cell surface receptor that restricts growth by sequestering Igf2 and targeting it for lysosomal degradation. There are now over 150 imprinted genes known in mammals, but no other clear examples of antagonistic gene pairs have been identified. The delta-like 1 gene (Dlk1) encodes a putative ligand that promotes fetal growth and in adults restricts adipose deposition. Conversely, Grb10 encodes an intracellular signalling adaptor protein that, when expressed from the maternal allele, acts to restrict fetal growth and is permissive for adipose deposition in adulthood. RESULTS: Here, using knockout mice, we present genetic and physiological evidence that these two factors exert their opposite effects on growth and physiology through a common signalling pathway. The major effects are on body size (particularly growth during early life), lean:adipose proportions, glucose regulated metabolism and lipid storage in the liver. A biochemical pathway linking the two cell signalling factors remains to be defined. CONCLUSIONS: We propose that Dlk1 and Grb10 define a mammalian growth axis that is separate from the IGF pathway, yet also features an antagonistic imprinted gene pair. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12915-014-0099-8) contains supplementary material, which is available to authorized users

Postnatal loss of Dlk1 imprinting in stem cells and niche astrocytes regulates neurogenesis.

The gene for the atypical NOTCH ligand delta-like homologue 1 (Dlk1) encodes membrane-bound and secreted isoforms that function in several developmental processes in vitro and in vivo. Dlk1, a member of a cluster of imprinted genes, is expressed from the paternally inherited chromosome. Here we show that mice that are deficient in Dlk1 have defects in postnatal neurogenesis in the subventricular zone: a developmental continuum that results in depletion of mature neurons in the olfactory bulb. We show that DLK1 is secreted by niche astrocytes, whereas its membrane-bound isoform is present in neural stem cells (NSCs) and is required for the inductive effect of secreted DLK1 on self-renewal. Notably, we find that there is a requirement for Dlk1 to be expressed from both maternally and paternally inherited chromosomes. Selective absence of Dlk1 imprinting in both NSCs and niche astrocytes is associated with postnatal acquisition of DNA methylation at the germ-line-derived imprinting control region. The results emphasize molecular relationships between NSCs and the niche astrocyte cells of the microenvironment, identifying a signalling system encoded by a single gene that functions coordinately in both cell types. The modulation of genomic imprinting in a stem-cell environment adds a new level of epigenetic regulation to the establishment and maintenance of the niche, raising wider questions about the adaptability, function and evolution of imprinting in specific developmental contexts

Evaluation of appendicitis risk prediction models in adults with suspected appendicitis

Background Appendicitis is the most common general surgical emergency worldwide, but its diagnosis remains challenging. The aim of this study was to determine whether existing risk prediction models can reliably identify patients presenting to hospital in the UK with acute right iliac fossa (RIF) pain who are at low risk of appendicitis. Methods A systematic search was completed to identify all existing appendicitis risk prediction models. Models were validated using UK data from an international prospective cohort study that captured consecutive patients aged 16–45 years presenting to hospital with acute RIF in March to June 2017. The main outcome was best achievable model specificity (proportion of patients who did not have appendicitis correctly classified as low risk) whilst maintaining a failure rate below 5 per cent (proportion of patients identified as low risk who actually had appendicitis). Results Some 5345 patients across 154 UK hospitals were identified, of which two‐thirds (3613 of 5345, 67·6 per cent) were women. Women were more than twice as likely to undergo surgery with removal of a histologically normal appendix (272 of 964, 28·2 per cent) than men (120 of 993, 12·1 per cent) (relative risk 2·33, 95 per cent c.i. 1·92 to 2·84; P < 0·001). Of 15 validated risk prediction models, the Adult Appendicitis Score performed best (cut‐off score 8 or less, specificity 63·1 per cent, failure rate 3·7 per cent). The Appendicitis Inflammatory Response Score performed best for men (cut‐off score 2 or less, specificity 24·7 per cent, failure rate 2·4 per cent). Conclusion Women in the UK had a disproportionate risk of admission without surgical intervention and had high rates of normal appendicectomy. Risk prediction models to support shared decision‐making by identifying adults in the UK at low risk of appendicitis were identified