Cutaneous gene therapy – an update

In this paper we review various strategies for gene transfer into the skin, their applications and show some of our own examples. The skin is the most accessible somatic tissue. Inherited skin diseases, such as epidermolysis bullosa and ichthyosis, and various systemic metabolic disorders are under investigation as potential candidate diseases for cutaneous gene transfer. Research is directed against genetic therapy of wounds and malignancies as well. So far, cutaneous gene transfer only has been used experimentally, but several clinical trials are under preparation.     

Comparative Analyses of QTLs Influencing Obesity and Metabolic Phenotypes in Pigs and Humans

The pig is a well-known animal model used to investigate genetic and mechanistic aspects of human disease biology. They are particularly useful in the context of obesity and metabolic diseases because other widely used models (e.g. mice) do not completely recapitulate key pathophysiological features associated with these diseases in humans. Therefore, we established a F2 pig resource population (n = 564) designed to elucidate the genetics underlying obesity and metabolic phenotypes. Segregation of obesity traits was ensured by using breeds highly divergent with respect to obesity traits in the parental generation. Several obesity and metabolic phenotypes were recorded (n = 35) from birth to slaughter (242 ± 48 days), including body composition determined at about two months of age (63 ± 10 days) via dual-energy x-ray absorptiometry (DXA) scanning. All pigs were genotyped using Illumina Porcine 60k SNP Beadchip and a combined linkage disequilibrium-linkage analysis was used to identify genome-wide significant associations for collected phenotypes. We identified 229 QTLs which associated with adiposity- and metabolic phenotypes at genome-wide significant levels. Subsequently comparative analyses were performed to identify the extent of overlap between previously identified QTLs in both humans and pigs. The combined analysis of a large number of obesity phenotypes has provided insight in the genetic architecture of the molecular mechanisms underlying these traits indicating that QTLs underlying similar phenotypes are clustered in the genome. Our analyses have further confirmed that genetic heterogeneity is an inherent characteristic of obesity traits most likely caused by segregation or fixation of different variants of the individual components belonging to cellular pathways in different populations. Several important genes previously associated to obesity in human studies, along with novel genes were identified. Altogether, this study provides novel insight that may further the current understanding of the molecular mechanisms underlying human obesity.     

Elite athletes’ genetic predisposition for altered risk of complex metabolic traits

Background

Genetic variants may predispose humans to elevated risk of common metabolic morbidities such as obesity and Type 2 Diabetes (T2D). Some of these variants have also been shown to influence elite athletic performance and the response to exercise training. We compared the genotype distribution of five genetic Single Nucleotide Polymorphisms (SNPs) known to be associated with obesity and obesity co-morbidities (IGF2BP2 rs4402960, LPL rs320, LPL rs328, KCJN rs5219, and MTHFR rs1801133) between athletes (all male, n = 461; endurance athletes n = 254, sprint/power athletes n = 207), and controls (all male, n = 544) in Polish and Russian samples. We also examined the association between these SNPs and the athletes’ competition level (‘elite’ and ‘national’ level). Genotypes were analysed by Single-Base Extension and Real-Time PCR. Multinomial logistic regression analyses were conducted to assess the association between genotypes and athletic status/competition level.

Results

IGF2BP2 rs4402960 and LPL rs320 were significantly associated with athletic status; sprint/power athletes were twice more likely to have the IGF2BP2 rs4402960 risk (T) allele compared to endurance athletes (OR = 2.11, 95% CI = 1.03-4.30, P <0.041), and non-athletic controls were significantly less likely to have the T allele compared to sprint/power athletes (OR = 0.62, 95% CI =0.43-0.89, P <0.0009). The control group was significantly more likely to have the LPL rs320 risk (G) allele compared to endurance athletes (OR = 1.26, 95% CI = 1.05-1.52, P <0.013). Hence, endurance athletes were the “protected” group being significantly (p < 0.05) less likely to have the risk allele compared to sprint/power athletes (IGF2BP2 rs4402960) and significantly (p < 0.05) less likely to have the risk allele compared to controls (LPL rs320). The other 3 SNPs did not show significant differences between the study groups.

Conclusions

Male endurance athletes are less likely to have the metabolic risk alleles of IGF2BP2 rs4402960 and LPL rs320, compared to sprint/power athletes and controls, respectively. These results suggest that some SNPs across the human genome have a dual effect and may predispose endurance athletes to reduced risk of developing metabolic morbidities, whereas sprint/power athletes might be predisposed to elevated risk.     

Bacteria and Fungi Respond Differently to Multifactorial Climate Change in a Temperate Heathland,Traced with 13C-Glycine and FACE CO2

It is vital to understand responses of soil microorganisms to predicted climate changes, as these directly control soil carbon (C) dynamics. The rate of turnover of soil organic carbon is mediated by soil microorganisms whose activity may be affected by climate change. After one year of multifactorial climate change treatments, at an undisturbed temperate heathland, soil microbial community dynamics were investigated by injection of a very small concentration (5.12 µg C g⁻¹ soil) of ¹³C-labeled glycine (¹³C₂, 99 atom %) to soils in situ. Plots were treated with elevated temperature (+1°C, T), summer drought (D) and elevated atmospheric carbon dioxide (510 ppm [CO2]), as well as combined treatments (TD, TCO2, DCO2 and TDCO2). The ¹³C enrichment of respired CO₂ and of phospholipid fatty acids (PLFAs) was determined after 24 h. ¹³C-glycine incorporation into the biomarker PLFAs for specific microbial groups (Gram positive bacteria, Gram negative bacteria, actinobacteria and fungi) was quantified using gas chromatography-combustion-stable isotope ratio mass spectrometry (GC-C-IRMS).Gram positive bacteria opportunistically utilized the freshly added glycine substrate, i.e. incorporated ¹³C in all treatments, whereas fungi had minor or no glycine derived ¹³C-enrichment, hence slowly reacting to a new substrate. The effects of elevated CO₂ did suggest increased direct incorporation of glycine in microbial biomass, in particular in G⁺ bacteria, in an ecosystem subjected to elevated CO₂. Warming decreased the concentration of PLFAs in general. The FACE CO₂ was ¹³C-depleted (δ¹³C = 12.2‰) compared to ambient (δ¹³C = ∼−8‰), and this enabled observation of the integrated longer term responses of soil microorganisms to the FACE over one year. All together, the bacterial (and not fungal) utilization of glycine indicates substrate preference and resource partitioning in the microbial community, and therefore suggests a diversified response pattern to future changes in substrate availability and climatic factors.     

Promoter polymorphisms in the chitinase 3-like 1 gene influence the serum concentration of YKL-40 in Danish patients with rheumatoid arthritis and in healthy subjects

Introduction  

The present study investigates the association between single nucleotide polymorphisms (SNPs) in the chitinase 3-like 1 (CHI3L1) gene and serum concentrations of YKL-40 in Danish patients with rheumatoid arthritis (RA) and healthy controls as well as the association with RA in the Danish population. The CHI3L1 gene is located on chromosome 1q32.1 and encodes the YKL-40 glycoprotein. YKL-40 concentrations are elevated in the serum of patients with RA compared to healthy subjects, and YKL-40 has been suggested to be an auto-antigen and may play a role in development of RA and in inflammation.     

Identification of genomic regions associated with phenotypic variation between dog breeds using selection mapping

The extraordinary phenotypic diversity of dog breeds has been sculpted by a unique population history accompanied by selection for novel and desirable traits. Here we perform a comprehensive analysis using multiple test statistics to identify regions under selection in 509 dogs from 46 diverse breeds using a newly developed high-density genotyping array consisting of >170,000 evenly spaced SNPs. We first identify 44 genomic regions exhibiting extreme differentiation across multiple breeds. Genetic variation in these regions correlates with variation in several phenotypic traits that vary between breeds, and we identify novel associations with both morphological and behavioral traits. We next scan the genome for signatures of selective sweeps in single breeds, characterized by long regions of reduced heterozygosity and fixation of extended haplotypes. These scans identify hundreds of regions, including 22 blocks of homozygosity longer than one megabase in certain breeds. Candidate selection loci are strongly enriched for developmental genes. We chose one highly differentiated region, associated with body size and ear morphology, and characterized it using high-throughput sequencing to provide a list of variants that may directly affect these traits. This study provides a catalogue of genomic regions showing extreme reduction in genetic variation or population differentiation in dogs, including many linked to phenotypic variation. The many blocks of reduced haplotype diversity observed across the genome in dog breeds are the result of both selection and genetic drift, but extended blocks of homozygosity on a megabase scale appear to be best explained by selection. Further elucidation of the variants under selection will help to uncover the genetic basis of complex traits and disease.     

pH landscapes in a novel five-species model of early dental biofilm

Background

Despite continued preventive efforts, dental caries remains the most common disease of man. Organic acids produced by microorganisms in dental plaque play a crucial role for the development of carious lesions. During early stages of the pathogenetic process, repeated pH drops induce changes in microbial composition and favour the establishment of an increasingly acidogenic and aciduric microflora. The complex structure of dental biofilms, allowing for a multitude of different ecological environments in close proximity, remains largely unexplored. In this study, we designed a laboratory biofilm model that mimics the bacterial community present during early acidogenic stages of the caries process. We then performed a time-resolved microscopic analysis of the extracellular pH landscape at the interface between bacterial biofilm and underlying substrate.

Methodology/Principal Findings

Strains of Streptococcus oralis, Streptococcus sanguinis, Streptococcus mitis, Streptococcus downei and Actinomyces naeslundii were employed in the model. Biofilms were grown in flow channels that allowed for direct microscopic analysis of the biofilms in situ. The architecture and composition of the biofilms were analysed using fluorescence in situ hybridization and confocal laser scanning microscopy. Both biofilm structure and composition were highly reproducible and showed similarity to in-vivo-grown dental plaque. We employed the pH-sensitive ratiometric probe C-SNARF-4 to perform real-time microscopic analyses of the biofilm pH in response to salivary solutions containing glucose. Anaerobic glycolysis in the model biofilms created a mildly acidic environment. Decrease in pH in different areas of the biofilms varied, and distinct extracellular pH-microenvironments were conserved over several hours.

Conclusions/Significance

The designed biofilm model represents a promising tool to determine the effect of potential therapeutic agents on biofilm growth, composition and extracellular pH. Ratiometric pH analysis using C-SNARF-4 gives detailed insight into the pH landscape of living biofilms and contributes to our general understanding of metabolic processes in in-vivo-grown bacterial biofilms.     

Childhood socioeconomic position and objectively measured physical capability levels in adulthood: a systematic review and meta-analysis

Background

Grip strength, walking speed, chair rising and standing balance time are objective measures of physical capability that characterise current health and predict survival in older populations. Socioeconomic position (SEP) in childhood may influence the peak level of physical capability achieved in early adulthood, thereby affecting levels in later adulthood. We have undertaken a systematic review with meta-analyses to test the hypothesis that adverse childhood SEP is associated with lower levels of objectively measured physical capability in adulthood.

Methods and Findings

Relevant studies published by May 2010 were identified through literature searches using EMBASE and MEDLINE. Unpublished results were obtained from study investigators. Results were provided by all study investigators in a standard format and pooled using random-effects meta-analyses. 19 studies were included in the review. Total sample sizes in meta-analyses ranged from N = 17,215 for chair rise time to N = 1,061,855 for grip strength. Although heterogeneity was detected, there was consistent evidence in age adjusted models that lower childhood SEP was associated with modest reductions in physical capability levels in adulthood: comparing the lowest with the highest childhood SEP there was a reduction in grip strength of 0.13 standard deviations (95% CI: 0.06, 0.21), a reduction in mean walking speed of 0.07 m/s (0.05, 0.10), an increase in mean chair rise time of 6% (4%, 8%) and an odds ratio of an inability to balance for 5s of 1.26 (1.02, 1.55). Adjustment for the potential mediating factors, adult SEP and body size attenuated associations greatly. However, despite this attenuation, for walking speed and chair rise time, there was still evidence of moderate associations.

Conclusions

Policies targeting socioeconomic inequalities in childhood may have additional benefits in promoting the maintenance of independence in later life.