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Summary: Mir483 is a conserved and highly expressed microRNA in placental mammals, embedded within the Igf2 gene. Its expression is dysregulated in a number of human diseases, including metabolic disorders and certain cancers. Here, we investigate the developmental regulation and function of Mir483 in vivo. We find that Mir483 expression is dependent on Igf2 transcription and the regulation of the Igf2/H19 imprinting control region. Transgenic Mir483 overexpression in utero causes fetal, but not placental, growth restriction through insulin-like growth factor 1 (IGF1) and IGF2 and also causes cardiovascular defects leading to fetal death. Overexpression of Mir483 post-natally results in growth stunting through IGF1 repression, increased hepatic lipid production, and excessive adiposity. IGF1 infusion rescues the post-natal growth restriction. Our findings provide insights into the function of Mir483 as a growth suppressor and metabolic regulator and suggest that it evolved within the INS-IGF2-H19 transcriptional region to limit excessive tissue growth through repression of IGF signaling.

Abstract Background The aim of this paper is to describe the utility of various recruitment modalities utilized in the Working to Increase Stability through Exercise (WISE) study. WISE is a pragmatic randomized trial that is testing the impact of a 3-year, multicomponent (strength, balance, aerobic) physical activity program led by trained volunteers or delivered via DVD on the rate of serious fall-related injuries among adults 65 and older with a past history of fragility fractures (e.g., vertebral, fall-related). The modified goal was to recruit 1130 participants over 2 years in three regions of Pennsylvania. Methods The at-risk population was identified primarily using letters mailed to patients of three health systems and those over 65 in each region, as well as using provider alerts in the health record, proactive recruitment phone calls, radio advertisements, and presentations at community meetings. Results Over 24 months of recruitment, 209,301 recruitment letters were mailed, resulting in 6818 telephone interviews. The two most productive recruitment methods were letters (72% of randomized participants) and the research registries at the University of Pittsburgh (11%). An average of 211 letters were required to be mailed for each participant enrolled. Of those interviewed, 2854 were ineligible, 2,825 declined to enroll and 1139 were enrolled and randomized. Most participants were female (84.4%), under age 75 (64.2%), and 50% took an osteoporosis medication. Not having a prior fragility fracture was the most common reason for not being eligible (87.5%). The most common reason provided for declining enrollment was not feeling healthy enough to participate (12.6%). Conclusions The WISE study achieved its overall recruitment goal. Bulk mailing was the most productive method for recruiting community-dwelling older adults at risk of serious fall-related injury into this long-term physical activity intervention trial, and electronic registries are important sources and should be considered.

El desarrollo de software debe estar enfocado en la imperiosa realidad de garantizar calidad tanto en todo su proceso como en sus resultados (producto), para lograr a través de éste, las ventajas competitivas en un entorno globalizado. Estar consciente de esta realidad en las instituciones desarrolladoras de software, se traducirá en dar formalidad a la disciplina de pruebas como apoyo a este proceso. Sin embargo, esta formalidad esperada en la disciplina y el uso de herramientas automatizadas es escasa en muchas instituciones en el campo. El presente trabajo se enfoca en el estudio de las pruebas automatizadas, algunos aspectos para el proceso de selección de las herramientas para dichas pruebas, siendo más específicos en las pruebas no funcionales, dado que el enfoque en esta área se ve positivamente alterado, pues los tiempos de los procesos de control de calidad serán significativamente más cortos y los resultados más precisos, por ende, se podrán invertir mejor sus recursos y fortalecer actividades como análisis de resultados, mejoramiento de procesos, productos y servicios.

Human infection with Mycobacterium bovis is reported infrequently in the United Kingdom. Most cases involve previous consumption of unpasteurized milk. We report a rare occurrence of 2 incidents of cat-to-human transmission of M. bovis during a cluster of infection in cats.

Despite the inverse relationship between physical activity (PA) and physical function, few older adults achieve PA recommendations. In response to observations that “lack of time” underlies reduced PA among older adults, recent work suggests even short bouts of PA can improve health and fitness. In addition, because they are frequently visited by older adults, an important conduit for PA promotion could be the primary care physician (PCP). However, most PCPs receive little training related to PA, rendering it difficult for them to offer meaningful counseling. Therefore, we explored the feasibility and impact of a PCP-prescribed one-minute daily functional exercise program, consisting of 30 s each of bodyweight push-ups and squats, among 24 patients 60 years of age or older. 42% of patients who were contacted started the exercise prescription and, over 24-weeks, completed approximately 114 sessions, while 75% completed at least half of the possible daily exercise sessions. As a group, the patients demonstrated increases in both maximal push-up and squat performance, though these increases plateaued following week-12. These preliminary results suggest that a PCP prescription of one-minute of daily functional exercise among older adult patients was feasible, acceptable, and effective for improving functional physical fitness. Given these findings, formal controlled research with recruitment from multiple clinics, random assignment to treatment conditions, and blinded assessments of objective functional physical performance should be pursued.

Cientos de objetos de metal de la Edad del Bronce excavados en yacimientos de Chipre han sido analizados para conocer su composición elemental y sus isótopos de plomo en el Isotrace Laboratory de la Universidad de Oxford entre los años 1982 y 2002. Especialmente con posterioridad a 1995 cientos de muestras de minerales y escorias recogidas de minas y sitios de reducción de minerales localizadas en el entorno de las montañas de Troodos también fueron analizadas. La mayoría de los resultados fueron publicados en varios artículos a lo largo del tiempo, pero la interpretación de algunos análisis de isótopos de plomo necesitan de una revisión a la vista de las nuevas investigaciones realizadas en España, Cerdeña y Sur de Francia. Es sabido que los datos de isótopos de plomo de objetos metálicos de yacimientos chipriotas de la Edad del Bronce muestran que no todo el cobre es consistente con un origen de minerales de Chipre. Además, los objetos de plomo y plata debieron ser importados debido a que no existen minerales de plomo o plata en la isla. La re-evaluación de los datos muestra que alrededor del 11 % de los objetos de metal analizados son consistentes con un origen en los depósitos del Egeo y Turquía, mientras que un 14 % lo son con minerales del Mediterráneo Occidental. Este artículo discute en detalle la interpretación actual de la investigación sobre el metal importado recuperado en contextos de la Edad del Bronce en Chipre.

The properties of the mature and pro-forms of recombinant apolipoprotein A-I (apoA-I) were compared with those of apoA-I isolated from human plasma. When the synthesis and secretion of pro- and mature forms of apoA-I from a baculovirus/insect cell expression system were compared in parallel experiments, the amount of the pro-form of apoA-I synthesized and secreted was severalfold higher than that of the mature form of apoA-I. A comparison of the properties of the pro- and mature forms of recombinant apoA-I and human plasma apoA-I showed no difference between all three in their secondary structure, their ability to self-associate, lipid-binding capacity, lecithin: cholesterol acyltransferase activation, and binding to the phospholipid transfer protein. The properties of reconstituted high density lipoprotein (HDL) particles formed from the proteins and their ability to promote cholesterol and phospholipid efflux from human skin fibroblasts were also similar. However, their ability to bind to plasma HDL subfractions differed, because twice as much proapoA-I associated with preβ1-HDL and preβ2-HDL subfractions compared with both mature recombinant and plasma apoA-I. Correspondingly, the amount of proapoA-I in α-HDL subfractions, especially in α1-HDL and α2-HDL, was decreased. We conclude that while the propeptide of apoA-I is required for the effective synthesis and secretion of apoA-I, cleavage of this peptide is a requisite for the effective interconversion of HDL subfractions.—Sviridov, D., L. E. Pyle, M. Jauhiainen, C. Ehnholm, and N. H. Fidge. Deletion of the propeptide of apolipoprotein A-I reduces protein expression, but stimulates effective conversion of preβ-high density lipoprotein to α-high density lipoprotein. J. Lipid Res. 2000. 41: 1872–1882.

Several HDL binding proteins, quite disparate in structure, have recently been cloned and their role in HDL metabolism is currently being assessed. High density lipoprotein binding protein, HBP (vigilin), which lacks a transmembrane domain is responsive to cell cholesterol levels, but its physiological significance remains unknown. On the other hand much is known about SR-B1, a member of the class B scavenger receptors. The level of SR-B1 expression correlates with both the selective transfer of cholesteryl ester into cells and cholesterol efflux from cells, the transfers probably mediated after docking of HDL at the cell surface. SR-B1 exhibits broad ligand specificity and, in animal models, appears to be regulated by the action of pituitary hormones that stimulate steroidogenesis, suggesting an important role for steroid hormone production in supplying precursor cholesterol. Another candidate HDL receptor, HB2, one of a pair of liver HDL binding proteins, shows high sequence homology with adhesion molecules, particularly activated leukocyte-cell adhesion molecule (ALCAM). When HB2 is overexpressed in cells, HDL binding increases. After PMA-induced differentiation of monocytes into macrophages, HB2 mRNA is strikingly elevated, which correlates with increased binding of HDL, but is down-regulated by cholesterol loading of macrophages. The ligand specificity of the HDL receptors, confounded by nonspecific lipid interactions, remains controversial. Their affinity for apoA-I versus apoA-I/A-II-rich HDL particles has clinical implications; both specific sequences in apoA-I and amphipathic α-helices may determine binding events. Post-receptor-mediated signalling events may regulate cell functions which, although not primarily related to lipid transport, nevertheless protect against coronary artery disease. Growing evidence for the involvement of lipid-poor apoA-I as a mediator of such pathways is also discussed.—Fidge, N. H. High density lipoprotein receptors, binding proteins, and ligands. J. Lipid Res. 1999. 40: 187–201.

The interaction of plasma phospholipid transfer protein (PLTP) with HDL has not been characterized in detail, although we have reported that the apoA-I/apoA-II molar ratio in the HDL particle influences PLTP-mediated HDL conversion, but not phospholipid transfer. The aim of this study was to examine whether PLTP binds apoA-I or apoA-II, and if this occurs, then determine the PLTP-binding domain of the apoA-I molecule. To study the PLTP/apolipoprotein interaction we used a solid phase ligand binding assay, the ELISA technique, and apoA-I and apoA-II affinity chromatography. PLTP bound to both apoA-I and apoA-II affinity columns, a finding subsequently utilized in the purification of PLTP. PLTP also bound to both apoA-I and apoA-II on ELISA plates in a concentration-dependent manner, and the binding could be displaced by preincubating the PLTP sample with purified apolipoproteins. To determine which portion of apoA-I is recognized by PLTP, we coated ELISA plates with either recombinant full-length apoA-I or three shortened apoA-I forms sequentially truncated from the C-terminus. To characterize the PLTP binding ability of the C-terminal region of apoA-I, we used both C-terminal CNBr-fragment and a synthetic C-terminal peptide of apoA-I. To further confirm the identity of the binding region, we probed the interaction with a polyclonal and several monoclonal anti-apoA-I antibodies. The antibodies that inhibited the interaction between PLTP and apoA-I were directed towards apoA-I epitopes localized between amino acids 27–141. The polyclonal antibody, R33, and the monoclonal antibody A-I-1 (epitope between amino acids 27–48) were most effective and reduced PLTP binding by 70%. These results show that PLTP binds to both apoA-I and apoA-II, and that the PLTP binding domain of apoA-I resides in the amino terminal region.—Pussinen, P. J., M. Jauhiainen, J. Metso, L. E. Pyle, Y. L. Marcel, N. H. Fidge, and C. Ehnholm. Binding of phospholipid transfer protein (PLTP) to apolipoproteins A-I and A-II: location of a PLTP binding domain in the amino terminal region of apoA-I. J. Lipid Res. 1998. 39: 152–161.

Bovine tuberculosis (bTB) caused by Mycobacterium bovis or M. caprae has recently (re-) emerged in livestock and wildlife in all countries bordering Switzerland (CH) and the Principality of Liechtenstein (FL). Comprehensive data for Swiss and Liechtenstein wildlife are not available so far, although two native species, wild boar (Sus scrofa) and red deer (Cervus elaphus elaphus), act as bTB reservoirs elsewhere in continental Europe. Our aims were (1) to assess the occurrence of bTB in these wild ungulates in CH/FL and to reinforce scanning surveillance in all wild mammals; (2) to evaluate the risk of a future bTB reservoir formation in wild boar and red deer in CH/FL. Tissue samples collected from 2009 to 2011 from 434 hunted red deer and wild boar and from eight diseased ungulates with tuberculosis-like lesions were tested by direct real-time PCR and culture to detect mycobacteria of the Mycobacterium tuberculosis complex (MTBC). Identification of suspicious colonies was attempted by real-time PCR, genotyping and spoligotyping. Information on risk factors for bTB maintenance within wildlife populations was retrieved from the literature and the situation regarding identified factors was assessed for our study areas. Mycobacteria of the MTBC were detected in six out of 165 wild boar (3.6%; 95% CI: 1.4-7.8) but none of the 269 red deer (0%; 0-1.4). M. microti was identified in two MTBC-positive wild boar, while species identification remained unsuccessful in four cases. Main risk factors for bTB maintenance worldwide, including different causes of aggregation often resulting from intensive wildlife management, are largely absent in CH and FL. In conclusion, M. bovis and M. caprae were not detected but we report for the first time MTBC mycobacteria in Swiss wild boar. Present conditions seem unfavorable for a reservoir emergence, nevertheless increasing population numbers of wild ungulates and offal consumption may represent a risk.

M. africanum West African 2 constitutes an ancient lineage of the M. tuberculosis complex that commonly causes human tuberculosis in West Africa and has an attenuated phenotype relative to M. tuberculosis.In search of candidate genes underlying these differences, the genome of M. africanum West African 2 was sequenced using classical capillary sequencing techniques. Our findings reveal a unique sequence, RD900, that was independently lost during the evolution of two important lineages within the complex: the "modern" M. tuberculosis group and the lineage leading to M. bovis. Closely related to M. bovis and other animal strains within the M. tuberculosis complex, M. africanum West African 2 shares an abundance of pseudogenes with M. bovis but also with M. africanum West African clade 1. Comparison with other strains of the M. tuberculosis complex revealed pseudogenes events in all the known lineages pointing toward ongoing genome erosion likely due to increased genetic drift and relaxed selection linked to serial transmission-bottlenecks and an intracellular lifestyle.The genomic differences identified between M. africanum West African 2 and the other strains of the Mycobacterium tuberculosis complex may explain its attenuated phenotype, and pave the way for targeted experiments to elucidate the phenotypic characteristic of M. africanum. Moreover, availability of the whole genome data allows for verification of conservation of targets used for the next generation of diagnostics and vaccines, in order to ensure similar efficacy in West Africa.

Bovine tuberculosis (bTB), caused by Mycobacterium bovis, is a debilitating disease of cattle. Ethiopia has one of the largest cattle populations in the world, with an economy highly dependent on its livestock. Furthermore, Ethiopia has one of the highest incidence rates of human extrapulmonary TB in the world, a clinical presentation that is often associated with transmission of M. bovis from cattle to humans.Here we present a comprehensive investigation of the prevalence of bTB in Ethiopia based on cases identified at slaughterhouses. Out of approximately 32,800 inspected cattle, approximately 4.7% showed suspect tuberculous lesions. Culture of suspect lesions yielded acid-fast bacilli in approximately 11% of cases, with M. bovis accounting for 58 of 171 acid-fast cultures, while 53 isolates were non-tuberculous mycobacteria. Strikingly, M. tuberculosis was isolated from eight cattle, an unusual finding that suggests human to animal transmission.Our analysis has revealed that bTB is widely spread throughout Ethiopia, albeit at a low prevalence, and provides underpinning evidence for public health policy formulation.

Rates of lipolysis, esterification, and free fatty acid release were estimated in isolated epididymal fat cells prepared from rats fed either ad lib. or with a restricted caloric intake. Basal and epinephrine- or theophylline-stimulated rates of lipolysis correlated positively with cell size in the ad lib.-fed group only. Rates of esterification, both basal and epinephrine-stimulated, correlated positively with cell size in the ad lib.-fed group but negatively in the caloric-restricted group. These findings indicate that nutritional factors can modify any possible influence of adipose cell size on lipolysis and esterification. On the other hand, in both groups of rats, epinephrine- and theophylline-stimulated rates of lipolysis correlated positively with the basal rates of lipolysis. Also, rates of esterification in the presence of epinephrine correlated positively with the basal rates of esterification, suggesting that stimulated rates of lipolysis and esterification are at least partly determined by the basal rates regardless of nutritional status. The activity of glycerokinase measured in homogenates of isolated fat cells, if applicable to intact fat cells, was sufficient to cause considerable underestimations of the basal rates of lipolysis (using glycerol production as an index). When lipolysis was stimulated, the potential errors of estimating lipolysis by glycerol production alone were negligible.

The chemical and anatomical pathways of absorption of dietary retinal, retinoic acid, and retinol were examined in rats containing lymph, bile, and duodenal cannulae. The experiments were designed to maintain physiological conditions to the greatest possible extent. In each rat an uninterrupted flow of bile into the duodenum was maintained by connecting the duodenal cannula to the bile duct of a second rat.Labeled vitamin A compounds were introduced into the duodenum in very small amounts (7-14 μg) in the form of a bile-lipid mixture resembling normal intestinal contents. Under these conditions, most (70-80%) of the radioactivity recovered after the feeding of labeled retinol or retinal was found in the lymph, predominantly in saturated retinyl esters. In contrast, 92-95%, of the radioactivity recovered after the feeding of labeled retinoic acid was found in the bile, and was contained in a mixture of polar metabolites, most of them more polar than free retinoic acid. Two-thirds of the small amount of radioactivity found in lymph after retinoic acid-14 feeding was in the form of free retinoic acid.The results indicate that under normal conditions the major pathway of retinal absorption involves its reduction to retinol, which is then esterified and transported via the lymphatics in a manner similar to that of dietary retinol. A small proportion of retinal is apparently normally oxidized, and is then transported via the portal vein and excreted in the bile in a manner similar to that of dietary retinoic acid. The relative importance, in quantitative terms, of these two pathways of retinal metabolism can vary, depending on the status of the animal.

The time course of oxidative damage in different brain regions was investigated in the gerbil model of transient cerebral ischemia. Animals were subjected to both common carotid arteries occlusion for 5 min. After the end of ischemia and at different reperfusion times (2, 6, 12, 24, 48, 72, 96 h and 7 days), markers of lipid peroxidation, reduced and oxidized glutathione levels, glutathione peroxidase, glutathione reductase, manganese-dependent superoxide dismutase (MnSOD) and copper/zinc containing SOD (Cu/ZnSOD) activities were measured in hippocampus, cortex and striatum. Oxidative damage in hippocampus was maximal at late stages after ischemia (48-96 h) coincident with a significant impairment in glutathione homeostasis. MnSOD increased in hippocampus at 24, 48 and 72 h after ischemia, coincident with the marked reduction in the activity of glutathione-related enzymes. The late disturbance in oxidant-antioxidant balance corresponds with the time course of delayed neuronal loss in the hippocampal CA1 sector. Cerebral cortex showed early changes in oxidative damage with no significant impairment in antioxidant capacity. Striatal lipid peroxidation significantly increased as early as 2 h after ischemia and persisted until 48 h with respect to the sham-operated group. These results contribute significant information on the timing and factors that influence free radical formation following ischemic brain injury, an essential step in determining effective antioxidant intervention.