Characterisation of the main drivers of intra- and inter- breed variability in the plasma metabolome of dogs

Introduction

Dog breeds are a consequence of artificial selection for specific attributes. These closed genetic populations have metabolic and physiological characteristics that may be revealed by metabolomic analysis.

Objectives

To identify and characterise the drivers of metabolic differences in the fasted plasma metabolome and then determine metabolites differentiating breeds.

Methods

Fasted plasma samples were collected from dogs maintained under two environmental conditions (controlled and client-owned at home). The former (n = 33) consisted of three breeds (Labrador Retriever, Cocker Spaniel and Miniature Schnauzer) fed a single diet batch, the latter (n = 96), client-owned dogs consisted of 9 breeds (Beagle, Chihuahua, Cocker Spaniel, Dachshund, Golden Retriever, Greyhound, German Shepherd, Labrador Retriever and Maltese) consuming various diets under differing feeding regimens. Triplicate samples were taken from Beagle (n = 10) and Labrador Retriever (n = 9) over 3 months. Non-targeted metabolite fingerprinting was performed using flow infusion electrospray-ionization mass spectrometry which was coupled with multivariate data analysis. Metadata factors including age, gender, sexual status, weight, diet and breed were investigated.

Results

Breed differences were identified in the plasma metabolome of dogs housed in a controlled environment. Triplicate samples from two breeds identified intra-individual variability, yet breed separation was still observed. The main drivers of variance in dogs maintained in the home environment were associated with breed and gender. Furthermore, metabolite signals were identified that discriminated between Labrador Retriever and Cocker Spaniels in both environments.

Conclusion

Metabolite fingerprinting of plasma samples can be used to investigate breed differences in client-owned dogs, despite added variance of diet, sexual status and environment.

     

Mechanistic study of the anti‐cancer effect of Gynostemma pentaphyllum saponins in the ApcMin/+ mouse model

Gynostemma pentaphyllum saponins (GpS) have been shown to have anti‐cancer activity. However, the underlying mechanisms remain unclear. In this study, we used the Apc^Min/+ colorectal cancer (CRC) mouse model to investigate the anti‐cancer effect of GpS and we demonstrated that GpS treatment could significantly reduce the number and size of intestinal polyps in Apc^Min/+ mice. In order to identify the potential targets and mechanisms involved, a comparative proteomics analysis was performed and 40 differentially expressed proteins after GpS treatment were identified. Bioinformatics analyses suggested a majority of these proteins were involved in processes related to cellular redox homeostasis, and predicted Raf‐1 as a potential target of GpS. The upregulation of two proteins known to be involved in redox homeostasis, peroxiredoxin‐1 (Prdx1) and peroxiredoxin‐2 (Prdx2), and the downregulation of Raf‐1 were validated using Western blot analysis. After further investigation of the associated signaling networks, we postulated that the anti‐cancer effect of GpS was mediated through the upregulation of Prdx1 and Prdx2, suppression of Ras, RAF/MEK/ERK/STAT, PI3K/AKT/mTOR signaling and modulation of JNK/p38 MAPK signaling. We also examined the potential combinatorial effect of GpS with the chemotherapeutic 5‐fluorouracil (5‐FU) and found that GpS could enhance the anti‐cancer efficacy of 5‐FU, further suppressing the number of polyps in Apc^Min/+ mice. Our findings highlight the potential of GpS as an anti‐cancer agent, the potential mechanisms of its anti‐cancer activities, and its effect as an adjuvant of 5‐FU in the chemotherapy of CRC.     

Relationship between maternal environment and DNA methylation patterns of estrogen receptor alpha in wild Eastern Bluebird (Sialia sialis) nestlings: a pilot study

There is mounting evidence that, across taxa, females breeding in competitive environments tend to allocate more testosterone to their offspring prenatally and these offspring typically have more aggressive and faster‐growing phenotypes. To date, no study has determined the mechanisms mediating this maternal effect's influence on offspring phenotype. However, levels of estrogen receptor alpha (ERα) gene expression are linked to differences in early growth and aggression; thus, maternal hormones may alter gene regulation, perhaps via DNA methylation, of ERα in offspring during prenatal development. We performed a pilot study to examine natural variation in testosterone allocation to offspring through egg yolks in wild Eastern Bluebirds (Sialia sialis) in varying breeding densities and percent DNA methylation of CG dinucleotides in the ERα promoter in offspring brain regions associated with growth and behavior. We hypothesized that breeding density would be positively correlated with yolk testosterone, and prenatal exposure to maternal‐derived yolk testosterone would be associated with greater offspring growth and decreased ERα promoter methylation. Yolk testosterone concentration was positively correlated with breeding density, nestling growth rate, and percent DNA methylation of one out of five investigated CpG sites (site 3) in the diencephalon ERα promoter, but none in the telencephalon (n = 10). Percent DNA methylation of diencephalon CpG site 3 was positively correlated with growth rate. These data suggest a possible role for epigenetics in mediating the effects of the maternal environment on offspring phenotype. Experimentally examining this mechanism with a larger sample size in future studies may help elucidate a prominent way in which animals respond to their environment. Further, by determining the mechanisms that mediate maternal effects, we can begin to understand the potential for the heritability of these mechanisms and the impact that maternal effects are capable of producing at an evolutionary scale.     

Immunogenicity of Outer Membrane Proteins VirB9-1 and VirB9-2, a Novel Nanovaccine against Anaplasma marginale

Anaplasma marginale is the most prevalent tick-borne livestock pathogen and poses a significant threat to cattle industry. In contrast to currently available live blood-derived vaccines against A. marginale, alternative safer and better-defined subunit vaccines will be of great significance. Two proteins (VirB9-1 and VirB9-2) from the Type IV secretion system of A. marginale have been shown to induce humoral and cellular immunity. In this study, Escherichia coli were used to express VirB9-1 and VirB9-2 proteins. Silica vesicles having a thin wall of 6 nm and pore size of 5.8 nm were used as the carrier and adjuvant to deliver these two antigens both as individual or mixed nano-formulations. High loading capacity was achieved for both proteins, and the mouse immunisation trial with individual as well as mixed nano-formulations showed high levels of antibody titres over 10⁷ and strong T-cell responses. The mixed nano-formulation also stimulated high-level recall responses in bovine T-cell proliferation assays. These results open a promising path towards the development of efficient A. marginale vaccines and provide better understanding on the role of silica vesicles to deliver multivalent vaccines as mixed nano-formulations able to activate both B-cell and T-cell immunity, for improved animal health.     

Sex and Gender Medical Education Summit: a roadmap for curricular innovation

The Sex and Gender Medical Education Summit: a roadmap for curricular innovation was a collaborative initiative of the American Medical Women's Association, Laura W. Bush Institute for Women’s Health, Mayo Clinic, and Society for Women's Health Research (www.sgbmeducationsummit.com). It was held on October 18–19, 2015 to provide a unique venue for collaboration among nationally and internationally renowned experts in developing a roadmap for the incorporation of sex and gender based concepts into medical education curricula. The Summit engaged 148 in-person attendees for the 1?1/2-day program. Pre- and post-Summit surveys assessed the impact of the Summit, and workshop discussions provided a framework for informal consensus building. Sixty-one percent of attendees indicated that the Summit had increased their awareness of the importance of sex and gender specific medicine. Other comments indicate that the Summit had a significant impact for motivating a call to action among attendees and provided resources to initiate change in curricula within their home institutions. These educational efforts will help to ensure a sex and gender basis for delivery of health care in the future.     

Using Africa's protected area network to estimate the global population of a threatened and declining species: a case study of the Critically Endangered White‐headed Vulture Trigonoceps occipitalis

The White‐headed Vulture Trigonoceps occipitalis (WhV) is uncommon and largely restricted to protected areas across its range in sub‐Saharan Africa. We used the World Database on Protected Areas to identify protected areas (PAs) likely to contain White‐headed Vultures. Vulture occurrence on road transects in Southern, East, and West Africa was adjusted to nests per km² using data from areas with known numbers of nests and corresponding road transect data. Nest density was used to calculate the number of WhV nests within identified PAs and from there extrapolated to estimate the global population. Across a fragmented range, 400 PAs are estimated to contain 1893 WhV nests. Eastern Africa is estimated to contain 721 nests, Central Africa 548 nests, Southern Africa 468 nests, and West Africa 156 nests. Including immature and nonbreeding birds, and accounting for data deficient PAs, the estimated global population is 5475 ‐ 5493 birds. The identified distribution highlights are alarming: over 78% (n = 313) of identified PAs contain fewer than five nests. A further 17% (n = 68) of PAs contain 5 ‐ 20 nests and 4% (n = 14) of identified PAs are estimated to contain >20 nests. Just 1% (n = 5) of PAs are estimated to contain >40 nests; none is located in West Africa. Whilst ranging behavior of WhVs is currently unknown, 35% of PAs large enough to hold >20 nests are isolated by more than 100 km from other PAs. Spatially discrete and unpredictable mortality events such as poisoning pose major threats to small localized vulture populations and will accelerate ongoing local extinctions. Apart from reducing the threat of poisoning events, conservation actions promoting linkages between protected areas should be pursued. Identifying potential areas for assisted re‐establishment via translocation offers the potential to expand the range of this species and alleviate risk.     

Differential Localization of Glutathione-S-Transferase Yp and Yb Subunits in Oligodendrocytes and Astrocytes of Rat Brain

Glutathione-S-transferase Yb subunits were recently identified in rat brain and localized to astrocytes, ependymal cells lining the ventricles, subventricular zone cells, and tanycytes. Another isoform, Yp (pi family), was detected in rat brain by immunoblotting, and its mRNA was detected by Northern hybridizations. Double immunofluorescence localized Yb and Yp in different glial cells. The strongly Yp-positive cells were identified as oligodendrocytes by virtue of their arrangement in rows in white-matter tracts, colocalization in strongly carbonic anhydrase-positive cells, and association with myelinated tracts in the corpus striatum. Ependymal cells in the choroid plexus and ventricular lining were also strongly Yp positive, whereas Yb was not detected in the choroid plexus. The occurrence of Yp at low levels in astrocytes was indicated after immunostaining by a sensitive peroxidase-antiperoxidase method, which revealed weak staining of those cells in the molecular layer of the cortex. The data suggest that Yb and Yp subunits are primarily localized to astrocytes and oligodendrocytes, respectively, and that both are absent from neurons. The glutathione-S-transferase in oligodendrocytes may participate in the removal of toxins from the vicinity of the myelin sheath. The finding of glutathione-S-transferases in ependymal cells and astrocytes in the brain also suggests that this enzyme could be a first line of defense against toxic substances.