Intravenous infusions of glucose stimulate key lipogenic enzymes in adipose tissue of dairy cows in a dose-dependent manner

The present study was investigated whether increasing amounts of glucose supply have a stimulatory effect on the mRNA abundance and activity of key lipogenic enzymes in adipose tissue of midlactation dairy cows. Twelve Holstein-Friesian dairy cows in midlactation were cannulated in the jugular vein and infused with either a 40% glucose solution (n=6) or saline (n=6). For glucose infusion cows, the infusion dose increased by 1.25%/d relative to the initial net energy for lactation (NEL) requirement until a maximum dose equating to a surplus of 30% NEL was reached on d 24. This maximum dose was maintained until d 28 and stopped thereafter (between d 29-32). Cows in the saline infusion group received an equivalent volume of 0.9% saline solution. Samples of subcutaneous adipose tissue were taken on d 0, 8, 16, 24, and 32 when surplus glucose reached 0, 10, 20, and 30% of the NEL requirement, respectively. The mRNA abundance of fatty acid synthase, cytoplasmic acetyl- coenzyme A synthetase, cytoplasmic glycerol 3-phosphate dehydrogenase-1, and glucose 6-phosphate dehydrogenase showed linear treatment × dose interactions with increasing mRNA abundance with increasing glucose dose. The increased mRNA abundance was paralleled by a linear treatment × dose interaction for fatty acid synthase and acetyl-coenzyme A synthetase enzymatic activities. The mRNA abundance of ATP-citrate lyase showed a tendency for linear treatment × dose interaction with increasing mRNA abundance with increasing glucose dose. The mRNA abundance of all tested enzymes, as well as the activities of fatty acid synthase and acetyl-coenzyme A synthetase, correlated with plasma glucose and serum insulin levels. In a multiple regression model, the predictive value of insulin was dominant over that of glucose. In conclusion, gradual increases in glucose supply upregulate key lipogenic enzymes in adipose tissue of midlactating dairy cows with linear dose dependency. Insulin appears to be critically involved in this regulation. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved

Simple high-cell density fed-batch technique for high-level recombinant protein production with Pichia pastoris: Application to intracellular production of Hepatitis B surface antigen

<p>Abstract</p> <p>Background</p> <p>Hepatitis B is a serious global public health concern. Though a safe and efficacious recombinant vaccine is available, its use in several resource-poor countries is limited by cost. We have investigated the production of Hepatitis B virus surface antigen (HBsAg) using the yeast <it>Pichia pastoris </it>GS115 by inserting the <it>HBsAg </it>gene into the alcohol oxidase 1 locus.</p> <p>Results</p> <p>Large-scale production was optimized by developing a simple fed-batch process leading to enhanced product titers. Cells were first grown rapidly to high-cell density in a batch process using a simple defined medium with low salt and high glycerol concentrations. Induction of recombinant product synthesis was carried out using rather drastic conditions, namely through the addition of methanol to a final concentration of 6 g L^-1. This methanol concentration was kept constant for the remainder of the cultivation through continuous methanol feeding based on the <it>on-line </it>signal of a flame ionization detector employed as methanol analyzer in the off-gas stream. Using this robust feeding protocol, maximum concentrations of ~7 grams HBsAg per liter culture broth were obtained. The amount of soluble HBsAg, competent for assembly into characteristic virus-like particles (VLPs), an attribute critical to its immunogenicity and efficacy as a hepatitis B vaccine, reached 2.3 grams per liter of culture broth.</p> <p>Conclusion</p> <p>In comparison to the highest yields reported so far, our simple cultivation process resulted in an ~7 fold enhancement in total HBsAg production with more than 30% of soluble protein competent for assembly into VLPs. This work opens up the possibility of significantly reducing the cost of vaccine production with implications for expanding hepatitis B vaccination in resource-poor countries.</p

Application of simple fed-batch technique to high-level secretory production of insulin precursor using Pichia pastoris with subsequent purification and conversion to human insulin

<p>Abstract</p> <p>Background</p> <p>The prevalence of diabetes is predicted to rise significantly in the coming decades. A recent analysis projects that by the year 2030 there will be ~366 million diabetics around the world, leading to an increased demand for inexpensive insulin to make this life-saving drug also affordable for resource poor countries.</p> <p>Results</p> <p>A synthetic insulin precursor (IP)-encoding gene, codon-optimized for expression in <it>P. pastoris</it>, was cloned in frame with the <it>Saccharomyces cerevisiae </it>α-factor secretory signal and integrated into the genome of <it>P. pastoris </it>strain X-33. The strain was grown to high-cell density in a batch procedure using a defined medium with low salt and high glycerol concentrations. Following batch growth, production of IP was carried out at methanol concentrations of 2 g L^-1, which were kept constant throughout the remaining production phase. This robust feeding strategy led to the secretion of ~3 gram IP per liter of culture broth (corresponding to almost 4 gram IP per liter of cell-free culture supernatant). Using immobilized metal ion affinity chromatography (IMAC) as a novel approach for IP purification, 95% of the secreted product was recovered with a purity of 96% from the clarified culture supernatant. Finally, the purified IP was trypsin digested, transpeptidated, deprotected and further purified leading to ~1.5 g of 99% pure recombinant human insulin per liter of culture broth.</p> <p>Conclusions</p> <p>A simple two-phase cultivation process composed of a glycerol batch and a constant methanol fed-batch phase recently developed for the intracellular production of the Hepatitis B surface antigen was adapted to secretory IP production. Compared to the highest previously reported value, this approach resulted in an ~2 fold enhancement of IP production using <it>Pichia </it>based expression systems, thus significantly increasing the efficiency of insulin manufacture.</p

MicroRNA expression signature in human abdominal aortic aneurysms

Background: Abdominal aortic aneurysm (AAA) is a dilatation of the aorta affecting most frequently elderly men. Histologically AAAs are characterized by inflammation, vascular smooth muscle cell apoptosis, and extracellular matrix degradation. The mechanisms of AAA formation, progression, and rupture are currently poorly understood. A previous mRNA expression study revealed a large number of differentially expressed genes between AAA and non-aneurysmal control aortas. MicroRNAs (miRNAs), small non-coding RNAs that are post-transcriptional regulators of gene expression, could provide a mechanism for the differential expression of genes in AAA. Methods: To determine differences in miRNA levels between AAA (n = 5) and control (n = 5) infrarenal aortic tissues, a microarray study was carried out. Results were adjusted using Benjamini-Hochberg correction (adjusted p\u3c 0.05). Real-time quantitative RT-PCR (qRT-PCR) assays with an independent set of 36 AAA and seven control tissues were used for validation. Potential gene targets were retrieved from miRNA target prediction databases Pictar, TargetScan, and MiRTarget2. Networks from the target gene set were generated and examined using the network analysis programs, CytoScape® and Ingenuity Pathway Core Analysis®. Results: A microarray study identified eight miRNAs with significantly different expression levels between AAA and controls (adjusted p \u3c 0.05). Real-time qRT-PCR assays validated the findings for five of the eight miRNAs. A total of 222 predicted miRNA target genes known to be differentially expressed in AAA based on a prior mRNA microarray study were identified. Bioinformatic analyses revealed that several target genes are involved in apoptosis and activation of T cells. Conclusions: Our genome-wide approach revealed several differentially expressed miRNAs in human AAA tissue suggesting that miRNAs play a role in AAA pathogenesis. Keywords: Apoptosis, Microarray analysis, Vascular biology, miRNA-mRNA analysis, Network analysi

Goettingen Minipigs (GMP): Comparison of Two Different Models for Inducing Diabetes

Purpose: Preclinical experiments on large animals are indispensable for evaluating the effectiveness of diabetes therapies. Miniature swine are well suited for such studies due to their physiological and pathophysiological responses. Methods: We compare two methods for inducing diabetes in Goettingen minipigs (GMP), in five with the beta cell toxin streptozotocin (STZ) and in five other GMP by total pancreatectomy (PE). Glucose homeostasis was assessed with the intravenous glucose-tolerance test (IVGTT) and continual monitoring of interstitial glucose levels. At conclusion of the observation period, the pancreata were examined histologically. Three non-diabetic GMP served as control group. Results: The IVGTT revealed markedly diabetic profiles in both GMP groups. STZ-GMP were found to harbor residual C-peptides and scattered insulin-positive cells in the pancreas. PE-GMP survived the total pancreatectomy only with intensive postoperative care. Conclusions: Although both methods reliably induced diabetes in GMP, the PE-GMP clearly had more health problems and required a greater expenditure of time and resources. The PE-GMP model, however, was better at eliminating endogenous insulin and C-peptide than the STZ-GMP model

Mental health of refugees following state-sponsored repatriation from Germany

von Lersner U, Elbert T, Neuner F. Mental health of refugees following state-sponsored repatriation from Germany. BMC Psychiatry. 2008;8(1): 88.BACKGROUND: In recent years, Voluntary Assisted Return Programmes (VARPs) have received increasing funding as a potential way of reducing the number of refugees in EU member states. A number of factors may affect the mental well-being of returnees. These include adjustment to the home country following return, difficult living conditions, and long-term effects resulting from the severe traumatic stress that had originally driven the affected out of their homes. Little is known about the extent to which these and other factors may promote or inhibit the willingness of refugees to return to their country of origin. The present pilot study investigated refugees who returned to their country of origin after having lived in exile in Germany for some 13 years. METHODS: Forty-seven VARP participants were interviewed concerning their present living conditions, their views of their native country, and their attitudes towards a potential return prior to actually returning. 33 participants were interviewed nine months after returning to their country of origin. Mental health and well-being were assessed using the questionnaires Posttraumatic Stress Diagnostic Scale (PDS) and EUROHIS and the structured Mini International Neuropsychiatric Interview (M.I.N.I.).Our objectives were to examine the mental health status of refugees returning to their home country following an extended period of exile. We also aimed to assess the circumstances under which people decided to return, the current living conditions in their home country, and retrospective returnee evaluations of their decision to accept assisted return. RESULTS: Prior to returning to their home country, participants showed a prevalence rate of 53% for psychiatric disorders. After returning, this rate increased to a sizeable 88%. Substantial correlations were found between the living situation in Germany, the disposition to return, and mental health. For two thirds of the participants, the decision to return was not voluntary. CONCLUSION: Psychological strain among study participants was of a considerable magnitude. As a result of traumatic stress experienced during war and refuge, victims were vulnerable and not well equipped to cope with either post-migration stressors in exile or with a return to their country of origin. It is noteworthy that the majority returned under pressure from immigration authorities. Living conditions after return (such as housing, work, and health care) were poor and unstable. Participants also had great difficulty readapting to the cultural environment after having lived abroad for an average of 13 years. Current VARPs do not take these factors into account and are therefore not able to assist in a humanitarian reintegration of voluntary returnees

Mental health of returnees: refugees in Germany prior to their state-sponsored repatriation

von Lersner U, Wiens U, Elbert T, Neuner F. Mental health of returnees: refugees in Germany prior to their state-sponsored repatriation. BMC International Health and Human Rights. 2008;8(1): 8

A large-scale genome-wide association study meta-analysis of cannabis use disorder

Summary Background Variation in liability to cannabis use disorder has a strong genetic component (estimated twin and family heritability about 50–70%) and is associated with negative outcomes, including increased risk of psychopathology. The aim of the study was to conduct a large genome-wide association study (GWAS) to identify novel genetic variants associated with cannabis use disorder. Methods To conduct this GWAS meta-analysis of cannabis use disorder and identify associations with genetic loci, we used samples from the Psychiatric Genomics Consortium Substance Use Disorders working group, iPSYCH, and deCODE (20 916 case samples, 363 116 control samples in total), contrasting cannabis use disorder cases with controls. To examine the genetic overlap between cannabis use disorder and 22 traits of interest (chosen because of previously published phenotypic correlations [eg, psychiatric disorders] or hypothesised associations [eg, chronotype] with cannabis use disorder), we used linkage disequilibrium score regression to calculate genetic correlations. Findings We identified two genome-wide significant loci: a novel chromosome 7 locus (FOXP2, lead single-nucleotide polymorphism [SNP] rs7783012; odds ratio [OR] 1·11, 95% CI 1·07–1·15, p=1·84 × 10−9) and the previously identified chromosome 8 locus (near CHRNA2 and EPHX2, lead SNP rs4732724; OR 0·89, 95% CI 0·86–0·93, p=6·46 × 10−9). Cannabis use disorder and cannabis use were genetically correlated (rg 0·50, p=1·50 × 10−21), but they showed significantly different genetic correlations with 12 of the 22 traits we tested, suggesting at least partially different genetic underpinnings of cannabis use and cannabis use disorder. Cannabis use disorder was positively genetically correlated with other psychopathology, including ADHD, major depression, and schizophrenia. Interpretation These findings support the theory that cannabis use disorder has shared genetic liability with other psychopathology, and there is a distinction between genetic liability to cannabis use and cannabis use disorder. Funding National Institute of Mental Health; National Institute on Alcohol Abuse and Alcoholism; National Institute on Drug Abuse; Center for Genomics and Personalized Medicine and the Centre for Integrative Sequencing; The European Commission, Horizon 2020; National Institute of Child Health and Human Development; Health Research Council of New Zealand; National Institute on Aging; Wellcome Trust Case Control Consortium; UK Research and Innovation Medical Research Council (UKRI MRC); The Brain & Behavior Research Foundation; National Institute on Deafness and Other Communication Disorders; Substance Abuse and Mental Health Services Administration (SAMHSA); National Institute of Biomedical Imaging and Bioengineering; National Health and Medical Research Council (NHMRC) Australia; Tobacco-Related Disease Research Program of the University of California; Families for Borderline Personality Disorder Research (Beth and Rob Elliott) 2018 NARSAD Young Investigator Grant; The National Child Health Research Foundation (Cure Kids); The Canterbury Medical Research Foundation; The New Zealand Lottery Grants Board; The University of Otago; The Carney Centre for Pharmacogenomics; The James Hume Bequest Fund; National Institutes of Health: Genes, Environment and Health Initiative; National Institutes of Health; National Cancer Institute; The William T Grant Foundation; Australian Research Council; The Virginia Tobacco Settlement Foundation; The VISN 1 and VISN 4 Mental Illness Research, Education, and Clinical Centers of the US Department of Veterans Affairs; The 5th Framework Programme (FP-5) GenomEUtwin Project; The Lundbeck Foundation; NIH-funded Shared Instrumentation Grant S10RR025141; Clinical Translational Sciences Award grants; National Institute of Neurological Disorders and Stroke; National Heart, Lung, and Blood Institute; National Institute of General Medical Sciences.Peer reviewe

Shared genetic risk between eating disorder- and substance-use-related phenotypes:Evidence from genome-wide association studies

First published: 16 February 202

Transancestral GWAS of alcohol dependence reveals common genetic underpinnings with psychiatric disorders

Liability to alcohol dependence (AD) is heritable, but little is known about its complex polygenic architecture or its genetic relationship with other disorders. To discover loci associated with AD and characterize the relationship between AD and other psychiatric and behavioral outcomes, we carried out the largest genome-wide association study to date of DSM-IV-diagnosed AD. Genome-wide data on 14,904 individuals with AD and 37,944 controls from 28 case-control and family-based studies were meta-analyzed, stratified by genetic ancestry (European, n = 46,568; African, n = 6,280). Independent, genome-wide significant effects of different ADH1B variants were identified in European (rs1229984; P = 9.8 x 10(-13)) and African ancestries (rs2066702; P = 2.2 x 10(-9)). Significant genetic correlations were observed with 17 phenotypes, including schizophrenia, attention deficit-hyperactivity disorder, depression, and use of cigarettes and cannabis. The genetic underpinnings of AD only partially overlap with those for alcohol consumption, underscoring the genetic distinction between pathological and nonpathological drinking behaviors.Peer reviewe