De Novo Lipogenesis and Cholesterol Synthesis in Humans with Long-Standing Type 1 Diabetes Are Comparable to Non-Diabetic Individuals

Background

Synthesis of lipid species, including fatty acids (FA) and cholesterol, can contribute to pathological disease. The purpose of this study was to investigate FA and cholesterol synthesis in individuals with type 1 diabetes, a group at elevated risk for vascular disease, using stable isotope analysis.

Methods

Individuals with type 1 diabetes (n = 9) and age-, sex-, and BMI-matched non-diabetic subjects (n = 9) were recruited. On testing day, meals were provided to standardize food intake and elicit typical feeding responses. Blood samples were analyzed at fasting (0 and 24 h) and postprandial (2, 4, 6, and 8 hours after breakfast) time points. FA was isolated from VLDL to estimate hepatic FA synthesis, whereas free cholesterol (FC) and cholesteryl ester (CE) was isolated from plasma and VLDL to estimate whole-body and hepatic cholesterol synthesis, respectively. Lipid synthesis was measured using deuterium incorporation and isotope ratio mass spectrometry.

Results

Fasting total hepatic lipogenesis (3.91±0.90% vs. 5.30±1.22%; P = 0.41) was not significantly different between diabetic and control groups, respectively, nor was synthesis of myristic (28.60±4.90% vs. 26.66±4.57%; P = 0.76), palmitic (12.52±2.75% vs. 13.71±2.64%; P = 0.65), palmitoleic (3.86±0.91% vs. 4.80±1.22%; P = 0.65), stearic (5.55±1.04% vs. 6.96±0.97%; P = 0.29), and oleic acid (1.45±0.28% vs. 2.10±0.51%; P = 0.21). Postprandial lipogenesis was also not different between groups (P = 0.38). Similarly, fasting synthesis of whole-body FC (8.2±1.3% vs. 7.3±0.8%/day; P = 0.88) and CE (1.9±0.4% vs. 2.0±0.3%/day; P = 0.96) and hepatic FC (8.2±2.0% vs. 8.1±0.8%/day; P = 0.72) was not significantly different between diabetic and control subjects.

Conclusions

Despite long-standing disease, lipogenesis and cholesterol synthesis was not different in individuals with type 1 diabetes compared to healthy non-diabetic humans.     

Invasiveness risk of the tropical biofuel crop Jatropha curcas L. into adjacent land use systems: from the rumors to the experimental facts

Jatropha curcas L. produces seeds rich in non‐edible oil suitable for biodiesel but it has been categorized as invasive. Although not scientifically verified, this allegation has resulted in a cultivation ban in several countries. In this article we report an integrated series of observations and experimental findings from invasiveness research in Zambia. We studied the impacts of J. curcas plantations on adjacent land use systems focusing on spontaneous occurrence of seedlings, seed dispersal mechanisms, seed predation by animals, and germination success of dispersed seeds. No spontaneous regeneration was observed in land use systems adjacent to J. curcas plantations. Primary seed dispersal was limited, predominantly under the canopy of the mother plant. Rodents and shrews dispersed and predated J. curcas seeds and fruits. They transported the seeds up to 23 m from the sources and repositioned them in their burrows up to 0.7 m deep, but none of these seeds could establish. Germination experiments in adjacent land use systems revealed 4% germination success at the soil surface, and 65% if buried artificially at 1–2 cm depth, yet the latter is unlikely to occur under natural conditions. These findings show that J. curcas seeds may be dispersed by animals to adjacent land use systems, but no natural recruitment was observed given low germination on the surface and none in burrows. Altogether these results suggest that the plant currently does not show an elevated risk of invasion to adjacent land use systems, at least in the investigated case study. But more long‐term studies, also in other growing areas are needed to corroborate these results.     

Influenza Aerosols in UK Hospitals during the H1N1 (2009) Pandemic – The Risk of Aerosol Generation during Medical Procedures

Background

Nosocomial infection of health-care workers (HCWs) during outbreaks of respiratory infections (e.g. Influenza A H1N1 (2009)) is a significant concern for public health policy makers. World Health Organization (WHO)-defined ‘aerosol generating procedures’ (AGPs) are thought to increase the risk of aerosol transmission to HCWs, but there are presently insufficient data to quantify risk accurately or establish a hierarchy of risk-prone procedures.

Methodology/Principal Findings

This study measured the amount of H1N1 (2009) RNA in aerosols in the vicinity of H1N1 positive patients undergoing AGPs to help quantify the potential risk of transmission to HCWs. There were 99 sampling occasions (windows) producing a total of 198 May stages for analysis in the size ranges 0.86–7.3 µm. Considering stages 2 (4–7.3 µm) and 3 (0.86–4 µm) as comprising one sample, viral RNA was detected in 14 (14.1%) air samples from 10 (25.6%) patients. Twenty three air samples were collected while potential AGPs were being performed of which 6 (26.1%) contained viral RNA; in contrast, 76 May samples were collected when no WHO 2009 defined AGP was being performed of which 8 (10.5%) contained viral RNA (unadjusted OR = 2.84 (95% CI 1.11–7.24) adjusted OR = 4.31 (0.83–22.5)).

Conclusions/Significance

With our small sample size we found that AGPs do not significantly increase the probability of sampling an H1N1 (2009) positive aerosol (OR (95% CI) = 4.31 (0.83–22.5). Although the probability of detecting positive H1N1 (2009) positive aerosols when performing various AGPs on intensive care patients above the baseline rate (i.e. in the absence of AGPs) did not reach significance, there was a trend towards hierarchy of AGPs, placing bronchoscopy and respiratory and airway suctioning above baseline (background) values. Further, larger studies are required but these preliminary findings may be of benefit to infection control teams.     

Development of an Efficient Targeted Cell-SELEX Procedure for DNA Aptamer Reagents

Background

DNA aptamers generated by cell-SELEX offer an attractive alternative to antibodies, but generating aptamers to specific, known membrane protein targets has proven challenging, and has severely limited the use of aptamers as affinity reagents for cell identification and purification.

Methodology

We modified the BJAB lymphoblastoma cell line to over-express the murine c-kit cell surface receptor. After six rounds of cell-SELEX, high-throughput sequencing and bioinformatics analysis, we identified aptamers that bound BJAB cells expressing c-kit but not wild-type BJAB controls. One of these aptamers also recognizes c-kit endogenously expressed by a mast cell line or hematopoietic progenitor cells, and specifically blocks binding of the c-kit ligand stem cell factor (SCF). This aptamer enables better separation by fluorescence-activated cell sorting (FACS) of c-kit⁺ hematopoietic progenitor cells from mixed bone marrow populations than a commercially available antibody, suggesting that this approach may be broadly useful for rapid isolation of affinity reagents suitable for purification of other specific cell types.

Conclusions/Significance

Here we describe a novel procedure for the efficient generation of DNA aptamers that bind to specific cell membrane proteins and can be used as high affinity reagents. We have named the procedure STACS (Specific TArget Cell-SELEX).     

Assessing Early Access to Care and Child Survival during a Health System Strengthening Intervention in Mali: A Repeated Cross Sectional Survey

Background

In 2012, 6.6 million children under age five died worldwide, most from diseases with known means of prevention and treatment. A delivery gap persists between well-validated methods for child survival and equitable, timely access to those methods. We measured early child health care access, morbidity, and mortality over the course of a health system strengthening model intervention in Yirimadjo, Mali. The intervention included Community Health Worker active case finding, user fee removal, infrastructure development, community mobilization, and prevention programming.

Methods and Findings

We conducted four household surveys using a cluster-based, population-weighted sampling methodology at baseline and at 12, 24, and 36 months. We defined our outcomes as the percentage of children initiating an effective antimalarial within 24 hours of symptom onset, the percentage of children reported to be febrile within the previous two weeks, and the under-five child mortality rate. We compared prevalence of febrile illness and treatment using chi-square statistics, and estimated and compared under-five mortality rates using Cox proportional hazard regression. There was a statistically significant difference in under-five mortality between the 2008 and 2011 surveys; in 2011, the hazard of under-five mortality in the intervention area was one tenth that of baseline (HR 0.10, p<0.0001). After three years of the intervention, the prevalence of febrile illness among children under five was significantly lower, from 38.2% at baseline to 23.3% in 2011 (PR = 0.61, p = 0.0009). The percentage of children starting an effective antimalarial within 24 hours of symptom onset was nearly twice that reported at baseline (PR = 1.89, p = 0.0195).

Conclusions

Community-based health systems strengthening may facilitate early access to prevention and care and may provide a means for improving child survival.     

Little evidence that condition,stress indicators,sex ratio,or homozygosity are related to landscape or habitat attributes in declining woodland birds

Habitat loss, fragmentation and degradation are drivers of major declines in biodiversity and species extinctions. The actual causes of species population declines following habitat change are more difficult to discern and there is typically high covariation among the measures used to infer the causes of decline. The causes of decline may act directly on individual fitness and survival, or through disruption of population processes. We examined the relationships among configuration, extent and status of native vegetation and three commonly used indicators of individual body condition and chronic stress (haemoglobin level, haematocrit, residual body mass condition index) in 13 species of woodland‐dependent birds in south‐eastern Australia. We also examined two measures of changes to population processes (sex ratio and individual homozygosity) in ten species and alleic richness in five species. We found little support for relationships between site or landscape characteristics and individual or population response variables, notwithstanding that our simulations showed we had sufficient power to detect relatively small effects. We discuss possible causes of the absence of detectable habitat effects in this system and the implications for the usefulness of individual body condition and easily measured haematological indices as indicators of the response of avian populations to habitat change.     

Estimation of population allele frequencies from next‐generation sequencing data: pool‐versus individual‐based genotyping

Molecular markers produced by next‐generation sequencing (NGS) technologies are revolutionizing genetic research. However, the costs of analysing large numbers of individual genomes remain prohibitive for most population genetics studies. Here, we present results based on mathematical derivations showing that, under many realistic experimental designs, NGS of DNA pools from diploid individuals allows to estimate the allele frequencies at single nucleotide polymorphisms (SNPs) with at least the same accuracy as individual‐based analyses, for considerably lower library construction and sequencing efforts. These findings remain true when taking into account the possibility of substantially unequal contributions of each individual to the final pool of sequence reads. We propose the intuitive notion of effective pool size to account for unequal pooling and derive a Bayesian hierarchical model to estimate this parameter directly from the data. We provide a user‐friendly application assessing the accuracy of allele frequency estimation from both pool‐ and individual‐based NGS population data under various sampling, sequencing depth and experimental error designs. We illustrate our findings with theoretical examples and real data sets corresponding to SNP loci obtained using restriction site–associated DNA (RAD) sequencing in pool‐ and individual‐based experiments carried out on the same population of the pine processionary moth (Thaumetopoea pityocampa). NGS of DNA pools might not be optimal for all types of studies but provides a cost‐effective approach for estimating allele frequencies for very large numbers of SNPs. It thus allows comparison of genome‐wide patterns of genetic variation for large numbers of individuals in multiple populations.     

Function-regulating pharmacophores in a sulfonamide class of glucocorticoid receptor agonists

A class of α-methyltryptamine sulfonamide glucocorticoid receptor (GR) modulators was optimized for agonist activity. The design of ligands was aided by molecular modeling, and key function-regulating pharmacophoric points were identified that are critical in achieving the desired agonist effect in cell based assays. Compound 27 was profiled in vitro and in vivo in models of inflammation. Analogs could be rapidly prepared in a parallel approach from aziridine building blocks.     

Characterization of the liver kinase B1-mouse protein-25 -Ste-20-related adaptor protein complex in adult mouse skeletal muscle

In liver, the AMP-activated protein kinase kinase (AMPKK) complex was identified as the association of liver kinase B1 (LKB1), mouse protein 25 (MO25α/β), and Ste-20-related adaptor protein (STRADα/β); however, this complex has yet to be characterized in skeletal muscle. We demonstrate the expression of the LKB1-MO25-STRAD complex in skeletal muscle, confirm the absence of mRNA splice variants, and report the relative mRNA expression levels of these proteins in control and muscle-specific LKB1 knockout (LKB1(-/-)) mouse muscle. LKB1 detection in untreated control and LKB1(-/-) muscle lysates revealed two protein bands (50 and 60 kDa), although only the heavier band was diminished in LKB1(-/-) samples [55 ± 2.5 and 13 ± 1.5 arbitrary units (AU) in control and LKB1(-/-), respectively, P < 0.01], suggesting that LKB1 is not represented at 50 kDa, as previously cited. The 60-kDa LKB1 band was further confirmed following purification using polyethylene glycol (43 ± 5 and 8.4 ± 4 AU in control and LKB1(-/-), respectively, P < 0.01) and ion-exchange fast protein liquid chromatography. Mass spectrometry confirmed LKB1 protein detection in the 60-kDa protein band, while none was detected in the 50-kDa band. Coimmunoprecipitation assays demonstrated LKB1-MO25-STRAD complex formation. Quantitative PCR revealed significantly reduced LKB1, MO25α, and STRADβ mRNA in LKB1(-/-) muscle. These findings demonstrate that the LKB1-MO25-STRAD complex is the principal AMPKK in skeletal muscle.     

Polymorphisms of the SLC11A1 gene and resistance to bovine tuberculosis in African Zebu cattle

Bovine tuberculosis (BTB) is a considerable health threat to livestock keepers and general communities in many developing countries. Information on genetic resistance or susceptibility because of polymorphisms of candidate genes could be used in making selection decisions for breeding disease tolerant/resistant animals. Here, we investigated associations between polymorphisms at the solute carrier family 11 (proton‐coupled divalent metal ion transporters), member 1 gene (SLC11A1, previously known as natural resistant associated macrophage protein 1, NRAMP1), with BTB phenotypes in Chadian cattle. Phenotypes were (i) single intradermal comparative cervical tuberculin test (SICCT) outcome, (ii) presence of gross visible lung lesions, (iii) a bacteriological culture test outcome and (iv) a predicted true BTB infection status using a Bayesian model. All traits were recorded as binary (presence or absence) traits. A total of 211 cattle were genotyped for a microsatellite within the SLC11A1 candidate gene. Standard linear and threshold‐liability models regressing BTB traits on copy number of SLC11A1 alleles revealed statistically significant effects of SLC11A1 alleles (P < 0.001) on most BTB traits. Polymorphisms (alleles 211, 215 and 217) are significantly related to lower incidence of BTB traits in Chadian cattle. This is the first study to report the association of SLC11A1 gene polymorphisms with BTB traits in Chadian or any other African cattle breeds.