Control of intestinal stem cell function and proliferation by mitochondrial pyruvate metabolism.

Most differentiated cells convert glucose to pyruvate in the cytosol through glycolysis, followed by pyruvate oxidation in the mitochondria. These processes are linked by the mitochondrial pyruvate carrier (MPC), which is required for efficient mitochondrial pyruvate uptake. In contrast, proliferative cells, including many cancer and stem cells, perform glycolysis robustly but limit fractional mitochondrial pyruvate oxidation. We sought to understand the role this transition from glycolysis to pyruvate oxidation plays in stem cell maintenance and differentiation. Loss of the MPC in Lgr5-EGFP-positive stem cells, or treatment of intestinal organoids with an MPC inhibitor, increases proliferation and expands the stem cell compartment. Similarly, genetic deletion of the MPC in Drosophila intestinal stem cells also increases proliferation, whereas MPC overexpression suppresses stem cell proliferation. These data demonstrate that limiting mitochondrial pyruvate metabolism is necessary and sufficient to maintain the proliferation of intestinal stem cells

A monoclonal antibody against kininogen reduces inflammation in the HLA-B27 transgenic rat

The human leukocyte antigen B27 (HLA-B27) transgenic rat is a model of human inflammatory bowel disease, rheumatoid arthritis and psoriasis. Studies of chronic inflammation in other rat models have demonstrated activation of the kallikrein–kinin system as well as modulation by a plasma kallikrein inhibitor initiated before the onset of clinicopathologic changes or a deficiency in high-molecular-mass kininogen. Here we study the effects of monoclonal antibody C11C1, an antibody against high-molecular-mass kininogen that inhibits the binding of high-molecular-mass kininogen to leukocytes and endothelial cells in the HLA-B27 rat, which was administered after the onset of the inflammatory changes. Thrice-weekly intraperitoneal injections of monoclonal antibody C11C1 or isotype IgG(1 )were given to male 23-week-old rats for 16 days. Stool character as a measure of intestinal inflammation, and the rear limbs for clinical signs of arthritis (tarsal joint swelling and erythema) were scored daily. The animals were killed and the histology sections were assigned a numerical score for colonic inflammation, synovitis, and cartilage damage. Administration of monoclonal C11C1 rapidly decreased the clinical scores of pre-existing inflammatory bowel disease (P < 0.005) and arthritis (P < 0.001). Histological analyses confirmed significant reductions in colonic lesions (P = 0.004) and synovitis (P = 0.009). Decreased concentrations of plasma prekallikrein and high-molecular-mass kininogen were found, providing evidence of activation of the kallikrein–kinin system. The levels of these biomarkers were reversed by monoclonal antibody C11C1, which may have therapeutic potential in human inflammatory bowel disease and arthritis

A Bayesian Partition Method for Detecting Pleiotropic and Epistatic eQTL Modules

Studies of the relationship between DNA variation and gene expression variation, often referred to as “expression quantitative trait loci (eQTL) mapping”, have been conducted in many species and resulted in many significant findings. Because of the large number of genes and genetic markers in such analyses, it is extremely challenging to discover how a small number of eQTLs interact with each other to affect mRNA expression levels for a set of co-regulated genes. We present a Bayesian method to facilitate the task, in which co-expressed genes mapped to a common set of markers are treated as a module characterized by latent indicator variables. A Markov chain Monte Carlo algorithm is designed to search simultaneously for the module genes and their linked markers. We show by simulations that this method is more powerful for detecting true eQTLs and their target genes than traditional QTL mapping methods. We applied the procedure to a data set consisting of gene expression and genotypes for 112 segregants of S. cerevisiae. Our method identified modules containing genes mapped to previously reported eQTL hot spots, and dissected these large eQTL hot spots into several modules corresponding to possibly different biological functions or primary and secondary responses to regulatory perturbations. In addition, we identified nine modules associated with pairs of eQTLs, of which two have been previously reported. We demonstrated that one of the novel modules containing many daughter-cell expressed genes is regulated by AMN1 and BPH1. In conclusion, the Bayesian partition method which simultaneously considers all traits and all markers is more powerful for detecting both pleiotropic and epistatic effects based on both simulated and empirical data

Gene expression profiling in brain of mice exposed to the marine neurotoxin ciguatoxin reveals an acute anti-inflammatory, neuroprotective response

<p>Abstract</p> <p>Background</p> <p>Ciguatoxins (CTXs) are polyether marine neurotoxins and potent activators of voltage-gated sodium channels. This toxin is carried by multiple reef-fish species and human consumption of ciguatoxins can result in an explosive gastrointestinal/neurologic illness. This study characterizes the global transcriptional response in mouse brain to a symptomatic dose of the highly toxic Pacific ciguatoxin P-CTX-1 and additionally compares this data to transcriptional profiles from liver and whole blood examined previously. Adult male C57/BL6 mice were injected with 0.26 ng/g P-CTX-1 while controls received only vehicle. Animals were sacrificed at 1, 4 and 24 hrs and transcriptional profiling was performed on brain RNA with Agilent whole genome microarrays. RT-PCR was used to independently validate gene expression and the web tool DAVID was used to analyze gene ontology (GO) and molecular pathway enrichment of the gene expression data.</p> <p>Results</p> <p>A pronounced 4°C hypothermic response was recorded in these mice, reaching a minimum at 1 hr and lasting for 8 hrs post toxin exposure. Ratio expression data were filtered by intensity, fold change and p-value, with the resulting data used for time course analysis, K-means clustering, ontology classification and KEGG pathway enrichment. Top GO hits for this gene set included acute phase response and mono-oxygenase activity. Molecular pathway analysis showed enrichment for complement/coagulation cascades and metabolism of xenobiotics. Many immediate early genes such as Fos, Jun and Early Growth Response isoforms were down-regulated although others associated with stress such as glucocorticoid responsive genes were up-regulated. Real time PCR confirmation was performed on 22 differentially expressed genes with a correlation of 0.9 (Spearman's Rho, p < 0.0001) with microarray results.</p> <p>Conclusions</p> <p>Many of the genes differentially expressed in this study, in parallel with the hypothermia, figure prominently in protection against neuroinflammation. Pathologic activity of the complement/coagulation cascade has been shown in patients suffering from a chronic form of ciguatera poisoning and is of particular interest in this model. Anti-inflammatory processes were at work not only in the brain but were also seen in whole blood and liver of these animals, creating a systemic anti-inflammatory environment to protect against the initial cellular damage caused by the toxin.</p

Llama-Derived Single Domain Antibodies to Build Multivalent, Superpotent and Broadened Neutralizing Anti-Viral Molecules

For efficient prevention of viral infections and cross protection, simultaneous targeting of multiple viral epitopes is a powerful strategy. Llama heavy chain antibody fragments (VHH) against the trimeric envelope proteins of Respiratory Syncytial Virus (Fusion protein), Rabies virus (Glycoprotein) and H5N1 Influenza (Hemagglutinin 5) were selected from llama derived immune libraries by phage display. Neutralizing VHH recognizing different epitopes in the receptor binding sites on the spikes with affinities in the low nanomolar range were identified for all the three viruses by viral neutralization assays. By fusion of VHH with variable linker lengths, multimeric constructs were made that improved neutralization potencies up to 4,000-fold for RSV, 1,500-fold for Rabies virus and 75-fold for Influenza H5N1. The potencies of the VHH constructs were similar or better than best performing monoclonal antibodies. The cross protection capacity against different viral strains was also improved for all three viruses, both by multivalent (two or three identical VHH) and biparatopic (two different VHH) constructs. By combining a VHH neutralizing RSV subtype A, but not subtype B with a poorly neutralizing VHH with high affinity for subtype B, a biparatopic construct was made with low nanomolar neutralizing potency against both subtypes. Trivalent anti-H5N1 VHH neutralized both Influenza H5N1 clade1 and 2 in a pseudotype assay and was very potent in neutralizing the NIBRG-14 Influenza H5N1 strain with IC50 of 9 picomolar. Bivalent and biparatopic constructs against Rabies virus cross neutralized both 10 different Genotype 1 strains and Genotype 5. The results show that multimerization of VHH fragments targeting multiple epitopes on a viral trimeric spike protein is a powerful tool for anti-viral therapy to achieve "best-in-class" and broader neutralization capacity

Rapid Environmental Change over the Past Decade Revealed by Isotopic Analysis of the California Mussel in the Northeast Pacific

The anthropogenic input of fossil fuel carbon into the atmosphere results in increased carbon dioxide (CO2) into the oceans, a process that lowers seawater pH, decreases alkalinity and can inhibit the production of shell material. Corrosive water has recently been documented in the northeast Pacific, along with a rapid decline in seawater pH over the past decade. A lack of instrumentation prior to the 1990s means that we have no indication whether these carbon cycle changes have precedence or are a response to recent anthropogenic CO2 inputs. We analyzed stable carbon and oxygen isotopes (δ13C, δ18O) of decade-old California mussel shells (Mytilus californianus) in the context of an instrumental seawater record of the same length. We further compared modern shells to shells from 1000 to 1340 years BP and from the 1960s to the present and show declines in the δ13C of modern shells that have no historical precedent. Our finding of decline in another shelled mollusk (limpet) and our extensive environmental data show that these δ13C declines are unexplained by changes to the coastal food web, upwelling regime, or local circulation. Our observed decline in shell δ13C parallels other signs of rapid changes to the nearshore carbon cycle in the Pacific, including a decline in pH that is an order of magnitude greater than predicted by an equilibrium response to rising atmospheric CO2, the presence of low pH water throughout the region, and a record of a similarly steep decline in δ13C in algae in the Gulf of Alaska. These unprecedented changes and the lack of a clear causal variable underscores the need for better quantifying carbon dynamics in nearshore environments

Homeodomain Interacting Protein Kinase 2 Activation Compromises Endothelial Cell Response to Laminar Flow: Protective Role of p21waf1,cip1,sdi1

BACKGROUND: In the cardiovascular system, laminar shear stress (SS) is one of the most important source of endothelial protecting signals. Physical and chemical agents, however, including ionising radiations and anticancer drugs, may injure endothelial cells determining an increase in oxidative stress and genotoxic damage. Whether the SS protective function remains intact in the presence of strong oxidants or DNA damage is currently unclear. METHODS AND RESULTS: To investigate this aspect a series of experiments were performed in which HUVEC were exposed to sub-lethal doses of the radio-mimetic compound Bleomycin (Bleo; 10 microg/ml) which generated free radicals (ROS) without significantly compromising cell survival. Remarkably, the application of a SS of 12 dyne/cm(2) did not protect endothelial cells but markedly accelerated apoptosis compared to controls kept in static culture and in the presence of Bleo. Experiments with the inducible nitric oxide synthase (iNOS) inhibitor GW274150 significantly reduced the SS-dependent apoptosis indicating that the production of NO was relevant for this effect. At molecular level, the ataxia-telangectasia-mutated (ATM) kinase, the homeodomain-interacting protein kinase-2 (HIPK2) and p53 were found activated along a pro-apoptotic signalling pathway while p21(waf1,cip1,sdi1) was prevented from its protective action. RNA interference experiments revealed that HIPK2 and p53 were both important for this process, however, only the forced expression p21(waf1,cip1,sdi1) fully restored the SS-dependent pro-survival function. CONCLUSIONS: This study provides the first evidence that, in the presence of genotoxic damage, laminar flow contributes to endothelial toxicity and death and identifies molecular targets potentially relevant in endothelial dysfunction and cardiovascular disease pathogenesis

A Review on the Oral Health Impacts of Acculturation

The impact of acculturation on systemic health has been extensively investigated and is regarded as an important explanatory factor for health disparity. However, information is limited and fragmented on the oral health implications of acculturation. This study aimed to review the current evidence on the oral health impact of acculturation. Papers were retrieved from five electronic databases. Twenty-seven studies were included in this review. Their scientific quality was rated and key findings were summarized. Seventeen studies investigated the impacts of acculturation on the utilization of dental services; among them, 16 reported positive associations between at least one acculturation indicator and use of dental services. All 15 studies relating acculturation to oral diseases (dental caries and periodontal disease) suggested better oral health among acculturated individuals. Evidence is lacking to support that better oral health of acculturated immigrants is attributable to their improved dental attendance. Further researches involving other oral health behaviors and diseases and incorporating refined acculturation scales are needed. Prospective studies will facilitate the understanding on the trajectory of immigrants’ oral health along the acculturation continuum

Survivors of intensive care with type 2 diabetes and the effect of shared care follow-up clinics: study protocol for the SWEET-AS randomised controlled feasibility study

Published online: 13 October 2016Background: Many patients who survive the intensive care unit (ICU) experience long-term complications such as peripheral neuropathy and nephropathy which represent a major source of morbidity and affect quality of life adversely. Similar pathophysiological processes occur frequently in ambulant patients with diabetes mellitus who have never been critically ill. Some 25 % of all adult ICU patients have diabetes, and it is plausible that ICU survivors with co-existing diabetes are at heightened risk of sequelae from their critical illness. ICU follow-up clinics are being progressively implemented based on the concept that interventions provided in these clinics will alleviate the burdens of survivorship. However, there is only limited information about their outcomes. The few existing studies have utilised the expertise of healthcare professionals primarily trained in intensive care and evaluated heterogenous cohorts. A shared care model with an intensivist- and diabetologist-led clinic for ICU survivors with type 2 diabetes represents a novel targeted approach that has not been evaluated previously. Prior to undertaking any definitive study, it is essential to establish the feasibility of this intervention. Methods: This will be a prospective, randomised, parallel, open-label feasibility study. Eligible patients will be approached before ICU discharge and randomised to the intervention (attending a shared care follow-up clinic 1 month after hospital discharge) or standard care. At each clinic visit, patients will be assessed independently by both an intensivist and a diabetologist who will provide screening and targeted interventions. Six months after discharge, all patients will be assessed by blinded assessors for glycated haemoglobin, peripheral neuropathy, cardiovascular autonomic neuropathy, nephropathy, quality of life, frailty, employment and healthcare utilisation. The primary outcome of this study will be the recruitment and retention at 6 months of all eligible patients. Discussion: This study will provide preliminary data about the potential effects of critical illness on chronic glucose metabolism, the prevalence of microvascular complications, and the impact on healthcare utilisation and quality of life in intensive care survivors with type 2 diabetes. If feasibility is established and point estimates are indicative of benefit, funding will be sought for a larger, multi-centre study. Trial registration: ANZCTR ACTRN12616000206426Yasmine Ali Abdelhamid, Liza Phillips, Michael Horowitz and Adam Dean

Socio-demographic factors associated with smoking and smoking cessation among 426,344 pregnant women in New South Wales, Australia

BACKGROUND: This study explores the socio-demographic characteristics of pregnant women who continue to smoke during the pregnancy, and identifies the characteristics of the smokers who were likely to quit smoking during the pregnancy period. METHODS: This was secondary analysis of the New South Wales (NSW) Midwives Data Collection (MDC) 1999–2003, a surveillance system covering all births in NSW public and private hospitals, as well as home births. Bivariate and multiple logistic regression analyses were performed to explore the associations between socio-demographic characteristics and smoking behaviour during pregnancy. RESULTS: Data from 426,344 pregnant women in NSW showed that 17.0% continued to smoke during pregnancy. The smoking rate was higher among teenage mothers, those with an Aboriginal (indigenous) background, and lower among more affluent and overseas-born mothers. This study also found that unbooked confinements, and lack of antenatal care in the first trimester were strongly associated with increased risk of smoking during pregnancy. About 4.0% of the smoking women reported they may quit smoking during their pregnancy. Findings showed that mothers born overseas, of higher socio-economic status, first time mothers and those who attended antenatal care early showed an increased likelihood of smoking cessation during pregnancy. Those who were heavy smokers were less likely to quit during pregnancy. CONCLUSION: Although the prevalence of smoking during pregnancy has been declining, it remains a significant public health concern. Smoking cessation programs should target the population subgroups of women at highest risk of smoking and who are least likely to quit. Effective antismoking interventions could reduce the obstetric and perinatal complications of smoking in pregnancy