Genotype-dependent Effects Research Articles

Developmental stage and genotype dependent behavioral effects of embryonic alcohol exposure in zebrafish larvae

Fetal Alcohol Spectrum Disorders (FASD) represent a worldwide problem. The severity and types of symptoms of FASD vary, which may be due to the genotype of the fetus and the developmental stage at which the fetus is exposed to alcohol. The most prevalent forms of FASD present less severe symptoms, including behavioral and cognitive abnormalities, and arise from exposure to low amounts of alcohol consumed infrequently. Treating or diagnosing FASD patients has been difficult because we do not understand the mechanisms underlying FASD. Animal models, including the zebrafish, have been suggested to answer this question. Here, we present a proof of concept analysis studying the behavioral effects of embryonic alcohol exposure in one-week old juvenile zebrafish. We exposed zebrafish embryos at one of five developmental stages (8, 16, 24, 32, or 40 hour post-fertilization) to 0% (control) or 1% (vol/vol) ethanol for 2 h, and tested the behavior of these fish at their age of 7–9 days post-fertilization. We employed two genetically distinct zebrafish populations, a quasi-inbred AB derivative strain, and a genetically variable WT population. We report significant developmental time and genotype dependent effects of alcohol on certain measures of motor function and/or anxiety-like responses. For example, we found embryonic alcohol exposed AB fish to swim faster, vary their speed more, stop moving more often and turn less compared to control fish, alcohol induced changes that were absent or less robust in WT fish. We conclude that our results open new avenues to the identification of genetic mechanisms that mediate or influence alcohol induced developmental alteration of brain function and behavior, which, on the long run, may allow us to identify diagnostic biomarkers and treatment options for human FASD.

Maternal immune activation modifies the course of Niemann-pick disease, type C1 in a gender specific manner

Niemann-Pick disease, type C1 (NPC1) is a rare neurodegenerative lysosomal storage disease with a wide spectrum of clinical manifestation. Multiple genetic factors influence the NPC1 mouse phenotype, but very little attention has been given to prenatal environmental factors that might have long-term effects on the neuroinflammatory component of NPC1 pathology. Studies in other mouse models of cerebellar ataxia have shown that developmental exposures lead to Purkinje neuron degeneration later in life, suggesting that environmental exposures during development can impact cerebellar biology. Thus, we evaluated the potential effect of maternal immune activation (MIA) on disease progression in an Npc1 mouse model. The MIA paradigm used mimics viral infection using the toll like receptor 3 agonist polyinosinic-polycytidilic acid during gestation. Through phenotypic and pathologic tests, we measured motor and behavioral changes as well as cerebellar neuroinflammation and neurodegeneration. We observed a gender and genotype dependent effect of MIA on the cerebellum. While the effects of MIA have been previously shown to primarily affect male progeny, we observed increased sensitivity of female mutant progeny to prenatal exposure to treatment with polyinosinic-polycytidilic acid. Specifically, prenatal MIA resulted in female NPC1 mutant progeny with greater motor deficits and a corresponding decrease in cerebellar Purkinje neurons. Our data suggest that prenatal environmental exposures may be one factor contributing to the phenotypic variability observed in individuals with NPC1.

P719Endothelial expression of adenylate cyclase type 9 (Adcy9) regulates endothelial cell signalling and vasodilation

Abstract Background/Introduction The clinical benefits of the cholesterol ester transfer protein (CETP) inhibitor dalcetrapib are dependent on the ADCY9 gene, which encodes adenylate cyclase (AC) type 9 (AC9). AC9 is one of nine membrane-bound isoforms of AC producing cyclic adenosine monophosphate (cAMP). We demonstrated that Adcy9 inactivation in the mouse protects from atherosclerosis, but only in absence of CETP. Adcy9 inactivation is also associated with improved endothelial function, including greater endothelial-dependent vasodilation (EDV) in response to acetylcholine (ACh). This suggests that Adcy9 may control endothelial Ca2+, an essential mediator of endothelial cell (EC) signalling. We hypothesized that Adcy9 is expressed in the endothelium and controls EC signalling. Purpose Our aim was to study AC9 expression in the vascular endothelium and the role of AC9 expression on endothelial cAMP signalling and Ca2+ dynamics. Methods The effects of Adcy9 inactivation were studied using wild-type (WT) and Adcy9-inactivated (Adcy9Gt/Gt) mice and siRNA-mediated inactivation of ADCY9 in human coronary artery endothelial cells (HCAEC). In the mouse, AC9 expression was quantified in whole aorta (± endothelium) and primary cultures of lung EC (LEC) by Western blot. Endothelial Ca2+ pulsars were monitored in mouse femoral arteries in response to acetylcholine (ACh, 10 μM). cAMP accumulation to AC activators VIP and forskolin were measured in LEC and HCAEC. EDV to VIP was studied ex vivo in mouse femoral arteries using pressurized arteriography. Results AC9 is expressed in LEC and the aorta, and its detection in the latter decreased by 33% after mechanical removal of the endothelium (P<0.05). AC activation with forskolin (10–4 M) in LEC led to increased cAMP accumulation in response to Adcy9 inactivation (WT: 1129±138 pmol cAMP/mg of protein, Adcy9Gt/Gt: 1965±169, n=3; P<0.01). ADCY9 inactivation in HCAEC also increased cAMP accumulation to forskolin 10–4M (control siRNA: 85±17, ADCY9 siRNA: 150±6, n=3; P<0.01). In LEC, receptor-dependent accumulation of cAMP to VIP (10–5 M) was higher in Adcy9Gt/Gt (1509±258)) compared to WT (915±170, n=3; P<0.05) mice. In mouse femoral arteries, VIP-induced maximal vasodilation was increased by Adcy9 inactivation in the presence of the endothelium (Adcy9Gt/Gt: 85±4%, n=6; WT: 52±9%, n=5, P<0.05), but not in its absence (Adcy9Gt/Gt: 67±12%, n=6 and WT: 59±15%, n=5). In the femoral artery endothelium, ACh increased Ca2+ pulsar frequency more in Adcy9Gt/Gt (243±32%, n=4) than in WT (178±19%, n=5; P<0.05) mice. Conclusion ADCY9 is expressed in the endothelium, and its inactivation potentiates endothelial cell Ca2+ dynamics, cAMP accumulation and VIP-induced vasodilation. We therefore identify ADCY9 as a new molecular pathway regulating endothelial-dependent vasodilation. This suggests that ADCY9's endothelial biology could be involved in the ADCY9 genotype-dependent clinical effects of dalcetrapib. Acknowledgement/Funding DalCor

Influence of histo blood group antigen expression on susceptibility to enteric viruses and vaccines.

Gastroenteritis results in substantial morbidity and mortality worldwide, especially in young children in low-and-middle-income settings. Rotavirus and norovirus are the leading causes of viral gastroenteritis. Although introduction of rotavirus vaccines into childhood immunization programmes has reduced disease burden, vaccine effectiveness remains low in developing countries. Norovirus is replacing rotavirus as the most common cause of diarrhea hospitalization in settings where rotavirus vaccines are highly effective. Genetically determined host factors, such as expression of histo blood group antigens (HBGAs) are hypothesized to play key roles in susceptibility to infections and gastroenteritis caused by these virus, as well as influence vaccine take. Epidemiology studies provide strong support for virus genotype-dependent effects of host HBGA expression, specifically secretor status on susceptibility to rotavirus and norovirus. Secretor-positive persons are significantly more susceptible to gastroenteritis caused by rotavirus P[8] genotype, and to infection with the GII.4 genotype of human norovirus. There is increasing data on the role of secretor status on rotavirus vaccine take but results are currently conflicting. For analyses involving young infants, maternal HBGA status is an important factor to be considered in future studies. Genetically determined HBGA expression influences susceptibility to enteric viruses of public health importance.

Deciphering rhizosphere microbiome assembly of wild and modern common bean (Phaseolus vulgaris) in native and agricultural soils from Colombia

BackgroundModern crop varieties are typically cultivated in agriculturally well-managed soils far from the centers of origin of their wild relatives. How this habitat expansion impacted plant microbiome assembly is not well understood.ResultsHere, we investigated if the transition from a native to an agricultural soil affected rhizobacterial community assembly of wild and modern common bean (Phaseolus vulgaris) and if this led to a depletion of rhizobacterial diversity. The impact of the bean genotype on rhizobacterial assembly was more prominent in the agricultural soil than in the native soil. Although only 113 operational taxonomic units (OTUs) out of a total of 15,925 were shared by all eight bean accessions grown in native and agricultural soils, this core microbiome represented a large fraction (25.9%) of all sequence reads. More OTUs were exclusively found in the rhizosphere of common bean in the agricultural soil as compared to the native soil and in the rhizosphere of modern bean accessions as compared to wild accessions. Co-occurrence analyses further showed a reduction in complexity of the interactions in the bean rhizosphere microbiome in the agricultural soil as compared to the native soil.ConclusionsCollectively, these results suggest that habitat expansion of common bean from its native soil environment to an agricultural context had an unexpected overall positive effect on rhizobacterial diversity and led to a stronger bean genotype-dependent effect on rhizosphere microbiome assembly.

The serotonin transporter gene variants modulate acute stress-induced hippocampus and dorsomedial prefrontal cortex activity during memory retrieval.

The short (s) allele of a polymorphism in the promoter region of the serotonin transporter gene (5-HTTLPR) is related to reduced serotonin transporter efficiency and an increased vulnerability to stress and mental disorders. In the present study, we investigated how 5-HTTLPR impacts on memory retrieval under stress and related neural activity by reanalyzing a small genetic neuroimaging data set. Twenty-seven healthy male volunteers participated in both the Trier Social Stress Test (TSST) and a respective control procedure and then their brain activity was measured with functional MRI (fMRI) while they performed an emotional-face-recognition task. Sixteen participants were carriers of the short allele (ss/sl carriers) and 11 were homozygous for the long allele (ll carriers). Genotype groups were compared with respect to stress-related physiological changes, memory performance, and brain activity. No significant genotype-dependent effects on memory performance or cortisol levels were found. The ss/sl carriers showed significantly higher systolic and diastolic blood pressure than the ll carriers, independent of stress. The ss/sl carriers reported stronger stress-induced nervous mood than the ll carriers. Our fMRI data revealed that the ss/sl carriers showed significantly weaker left hippocampus activation and stronger dorsomedial prefrontal cortex (dmPFC) deactivation when retrieving memories under stress as compared with the ll carriers. Subsequent analyses revealed that the distinct hippocampal activation pattern in both genotypes was associated with stress-induced cortisol elevation, while the distinct dmPFC activation pattern in both genotypes was associated with stress-induced changes in reaction times. Our results thus add new evidence that serotonin signaling modulates neural activity in the hippocampus and dmPFC during memory retrieval under acute psychosocial stress.

Interactions between the host genome and the gut microbiome determine susceptibility to CNS autoimmune disease

Abstract Multiple sclerosis (MS) is a chronic autoimmune central nervous system (CNS) disease and the leading cause of non-traumatic neurological disability in young adults. The etiology of MS is multifactorial and polygenic, with 30% of risk genetic and 70% environmental. Recent studies identified an imbalance in the human gut microbiome within MS patients, including depletion of the Lactobacillus genus. Animal models support a causal role of the gut microbiome in MS, though the mechanisms remain unclear. Here, we leverage natural genetic variation in wild-derived PWD/PhJ (PWD) mice as compared to classic C57BL/6J (B6) mice, modeling human genetic diversity to identify host genome and gut microbiome interactions governing disease susceptibility. In a model of MS, experimental autoimmune encephalomyelitis (EAE), 27 consomic strains carrying PWD chromosomes on the B6 background demonstrated that PWD-derived loci exhibit bidirectional and sex-specific effects on disease severity. 16S rRNA gene sequencing of the gut microbiome in consomic and parental strains identified strain-specific profiles and metabolic functions associated with differential susceptibility to EAE. Functionally, manipulation of the gut microbiome by co-housing or transplantation, demonstrated direct host genotype-dependent effects of the microbiome on EAE susceptibility, with transfer of signature Lactobacillus species associated with changes in circulating indole metabolites and altered EAE severity. These studies reveal the presence of complex bidirectional interactions between the host genome and gut microbiome that modulate susceptibility to CNS autoimmunity, and identify specific microbiome constituents as potential therapeutic candidates.

Genetic variation in the glucocorticoid receptor and psychopathology after dexamethasone administration in cardiac surgery patients

The glucocorticoid receptor (GR) agonist dexamethasone is frequently used for its anti-inflammatory properties. We recently showed that a single high-dose of dexamethasone had long-lasting protective effects on the development of psychopathology after cardiac surgery and postoperative intensive care unit stay. In this study, we investigated whether common genetic variation in the hypothalamic-pituitary-adrenal (HPA)-axis would influence the susceptibility for PTSD and depression after dexamethasone administration.Participants (n = 996) of the Dexamethasone for Cardiac Surgery (DECS) randomized clinical trial were followed after receiving a single high intraoperative dose of dexamethasone (1 mg/kg), a GR agonist, or placebo. PTSD and depressive symptoms were assessed up to four years after cardiac surgery. We focused primarily on five common single nucleotide polymorphisms (SNPs) in the glucocorticoid receptor (GR). Secondarily, we comprehensively assessed common genetic variation in the FK506 binding protein (FKBP5) and the mineralocorticoid receptor (MR).The protective effects of dexamethasone on postoperative PTSD symptoms were dependent on the GR polymorphisms rs41423247 (p = .009), rs10052957 (p = .003), and rs6189 (p = .002), but not on rs6195 (p = .025) or rs6198, (p = .026) after Bonferroni correction. No genotype-dependent effects were found for postoperative depressive symptoms. Also, no associations of FKBP5 and MR polymorphisms were found on PTSD and depression outcomes.Protective effects of dexamethasone on PTSD symptoms after cardiac surgery and ICU stay seem to depend on common genetic variation in its target receptor, the GR. These effects indicate that pre-operative genetic screening could potentially help in stratifying patients for their vulnerability for developing PTSD symptoms after surgery.

A role for the gibberellin pathway in biochar-mediated growth promotion

Biochar is a carbon negative soil amendment that can promote crop growth. However, the effects of biochar on different plant species and cultivars within a species are not well understood, nor is the underlying basis of biochar-mediated plant growth promotion. This knowledge is critical for optimal use of biochar and for breeding biochar-responsive plants. Here, we investigated the genotype-specific effects of biochar on two cultivars of Solanum lycopersicum (tomato), and two wild relatives of tomato, Solanum pimpinellifolium, and Solanum pennelli, in two types of biochar. Biochar promoted shoot growth in all genotypes independent of biochar type but had genotype-dependent effects on other plant traits. Germination tests, exogenous GA4 application and mutant analysis indicated a role for GA in biochar-mediated plant growth promotion. Together, our results suggest that biochar promotes growth partially through stimulation of the GA pathway.

Shaping of Intestinal Microbiota in Nlrp6- and Rag2-Deficient Mice Depends on Community Structure

Contradicting observations have been made regarding the relative contributions of immune sensors to shaping the microbiome, yet the reasons for these discrepancies are not fully understood. Here, we investigated the contribution of environmental factors in shaping the microbiome in mice deficient in adaptive immunity (Rag2-/-) and Nlrp6, an immune sensor proposed to be involved in regulation of microbiota composition. In conventionally housed Nlrp6-/- mice, familial transmission has a significant effect on microbiota composition, complicating theanalysis of genotype-dependent effects. Notably, after rederivation into standardized specific pathogen-free (SPF) conditions devoid of pathobionts, microbiota composition was indistinguishable between WT, Rag2-/-, and Nlrp6-/- mice. However, upon reintroduction of a pathobiont-containing community host, genotype-dependent differences reappear, specifically affecting the relative abundance of pathobionts such as Helicobacter spp. Our results show that the impact of Nlrp6 and also of adaptive immunity on microbiota composition depends on community structure and primarily influences pathobionts but not commensals.

Psoriasis-Associated Late Cornified Envelope (LCE) Proteins Have Antibacterial Activity

Terminally differentiating epidermal keratinocytes express a large number of structural and antimicrobial proteins that are involved in the physical barrier function of the stratum corneum and provide innate cutaneous host defense. Late cornified envelope (LCE) genes, located in the epidermal differentiation complex on chromosome 1, encode a family of 18 proteins of unknown function, whose expression is largely restricted to epidermis. Deletion of two members, LCE3B and LCE3C (LCE3B/C-del), is a widely-replicated psoriasis risk factor that interacts with the major psoriasis-psoriasis risk gene HLA-C*06. Here we performed quantitative trait locus analysis, utilizing RNA-seq data from human skin and found that LCE3B/C-del was associated with a markedly increased expression of LCE3A, a gene directly adjacent to LCE3B/C-del. We confirmed these findings in a 3-dimensional skin model using primary keratinocytes from LCE3B/C-del genotyped donors. Functional analysis revealed that LCE3 proteins, and LCE3A in particular, have defensin-like antimicrobial activity against a variety of bacterial taxa at low micromolar concentrations. No genotype-dependent effect was observed for the inside-out or outside-in physical skin barrier function. Our findings identify an unknown biological function for LCE3 proteins and suggest a role in epidermal host defense and LCE3B/C-del-mediated psoriasis risk.

Abstract 184: Adenylate Cyclase Type 9 Promotes Atherosclerosis in Mice

Polymorphisms in the ADCY9 gene, coding for adenylate cyclase type 9, determine atherosclerotic cardiovascular responses to the CETP inhibitor dalcetrapib in patients. ADCY9 is broadly expressed and involved in various immune cell functions and inflammatory responses. Our objective is to determine the role of ADCY9 in the development of atherosclerosis in the absence of CETP. We used 8-12 week-old wild-type (WT, n=25) or Adcy9 -inactivated ( Adcy9 Gt , n=21) male mice. To induce hypercholesterolemia and atherosclerosis, mice were infected with an adeno-associated virus coding for a gain-of-function mutant of Pcsk9 ( Pcsk9 D377Y ) and were fed an atherogenic high-cholesterol diet for 16 weeks in absence or presence of dalcetrapib treatment (200 mg/kg/day). Percent atherosclerotic lesion area (PALA) in the whole aorta was quantified using en face Oil Red O plaque staining. We used VCAM-1 immunofluorescence staining (n=6 per group) in atherosclerotic lesions of the aortic root to quantify inflammation. We also measured aortic root accumulation of macrophages and vascular smooth muscle cells by immunofluorescence imaging of CD68 and smooth muscle actin (SMA), respectively, in plaque lesions. We also quantified blood leukocytes in normocholesterolemic WT and Adcy9 Gt mice by flow cytometry analysis of CD45 + cells. In hypercholesterolemic animals, Adcy9 Gt mice showed a 60% decrease in atherosclerotic lesions (PALA: 2.7±0.5%) compared to WT mice (6.5±0.7%, P<0.01). Dalcetrapib treatment, in these mice without CETP, did not modify significantly the reduction (63%) of PALA in Adcy9 Gt mice (2.9±0.4%) compared to WT (7.6±1.6%, P<0.01). Macrophage content was reduced by 55% from 17.8±2.8% in WT to 7.2%±1.9% in Adcy9 Gt mice (P<0.05), while SMA staining was similar. VCAM-1 expression also tended to be decreased, from 5.7±3.3% in WT to 2.0±0.4% in Adcy9 Gt mice (P=0.0598). CD45 + blood leukocytes were reduced by 35.4% in Adcy9 Gt compared to WT normocholesterolemic mice (P<0.01). Adcy9 inactivation in mice (without CETP) results in large reductions of atherosclerosis and plaque macrophage and in decreased VCAM-1 expression. These results support the ADCY9 genotype-dependent clinical effects of dalcetrapib.

APOE ε4/ε4 diminishes neurotrophic function of human iPSC-derived astrocytes.

The ε4 allele of the APOE gene encoding apolipoprotein E (apoE) is a strong genetic risk factor for aging-related cognitive decline as well as late-onset Alzheimer's disease (AD) compared to the common ε3 allele. In the central nervous system, apoE is produced primarily by astrocytes and functions in transporting lipids including cholesterol to support neuronal homeostasis and synaptic integrity. Although mouse models and corresponding primary cells have provided valuable tools for studying apoE isoform-dependent functions, recent studies have shown that human astrocytes have a distinct gene expression profile compare with rodent astrocytes. Human induced pluripotent stem cells (iPSCs) derived from individuals carrying specific gene variants or mutations provide an alternative cellular model more relevant to humans upon differentiation into specific cell types. Thus, we reprogramed human skin fibroblasts from cognitively normal individuals carrying APOE ε3/ε3 or ε4/ε4 genotype to iPSC clones and further differentiated them into neural progenitor cells and then astrocytes. We found that human iPSC-derived astrocytes secreted abundant apoE with apoE4 lipoprotein particles less lipidated compared to apoE3 particles. More importantly, human iPSC-derived astrocytes were capable of promoting neuronal survival and synaptogenesis when co-cultured with iPSC-derived neurons with APOE ε4/ε4 astrocytes less effective in supporting these neurotrophic functions than those with APOE ε3/ε3 genotype. Taken together, our findings demonstrate APOE genotype-dependent effects using human iPSC-derived astrocytes and provide novel evidence that the human iPSC-based model system is a strong tool to explore how apoE isoforms contribute to neurodegenerative diseases.

Genotype-dependent alleviation effects of exogenous GSH on salinity stress in cotton is related to improvement in chlorophyll content, photosynthetic performance, and leaf/root ultrastructure.

Soil salinity is a major abiotic stress that is constraining crop growth and productivity. Greenhouse hydroponic experiments were performed using salt-sensitive (cv. Zhongmian 41) and tolerant (Zhong 9806) cotton seedlings to evaluate how different genotypes responded to salinity stress in the presence of exogenous GSH (reduced glutathione). Cotton plants grown in 150mM NaCl showed severe reduction in plant height, root length, and shoot and root fresh/dry weight. Salinity also caused reduction in photosynthesis and chlorophyll content, but increase in malondialdehyde (MDA) content. However, the reduction was more in Zhongmian 41 compared to Zhong 9806. Importantly, Sodium concentration was increased in the two genotypes and the induction was more in Zhongmian 41. Calcium and magnesium concentration was decreased in Zhongmian 41; however, in Zhong 9806 there were no significant differences relative to control. Addition of 50mg L-1 GSH in150mM NaCl solution (Na + GSH) significantly alleviated salinity stress. Compared with salinity treatment alone (NaCl), Na + GSH increased fresh and dry weight of the root, stem, and leaf, photosynthesis, and chlorophyll content. Obvious ultrastructural alterations were observed in the saline-treated leaf- and root-tip cells. Exogenous GSH greatly ameliorated the salinity-induced damage on the leaf/root ultrastructure, especially in Zhongmian 41.These results advocate a positive role for GSH in alleviation of salinity, which is related to significant improvement in chlorophyll content, photosynthetic performance, and leaf/root ultrastructure.

Genotype-dependent effects of phosphorus supply on physiological and biochemical responses to Al-stress in cultivated and Tibetan wild barley

Aluminium (Al) toxicity and phosphorus (P) deficiency often co-exist in acidic soils and limit plant growth and crop production. To investigate the alleviating effects of different levels of phosphorus on Al stress, greenhouse hydroponic experiments were conducted using two contrasting Tibetan wild barley genotypes XZ16 and XZ61 of Al tolerant and sensitive, respectively, and Al tolerant cv. Dayton. The results showed that Al stress induced reduction in P accumulation in plants; and stem and leaf P concentrations of the three genotypes, except of XZ16 under HP + Al (100 µM Al with high level of 360 µM P) which was close to the control level. XZ16 recorded significantly higher P accumulation in plants, compared with XZ61 and Dayton, and P concentrations in leaves under Al stress, and in stems under NP + Al (100 µM Al with normal level of 180 µM P) and HP + Al. Meanwhile, H+-, Ca2+Mg2+-, and Total- ATPase activities in XZ16 and Dayton under Al stress were markedly higher than in XZ61. Normal or high level of P under Al stress could relieve Al stress as enhanced plant biomass, with increased photosystem II photochemistry (Fv/Fm) and P content, relative to the low level of 90 µM P. Compared with XZ61, addition of high P concentration for XZ16 significantly increased the values of Gs and Tr, with higher root GPX and H+-ATPase activities, and such nutrient elements as P, Mg and Ca in stems and leaves, and induced more malate secretion, but less MDA accumulation.

CETP: Pharmacogenomics-Based Response to the CETP Inhibitor Dalcetrapib.

High-density lipoproteins are involved in reverse cholesterol transport and possess anti-inflammatory and antioxidative properties. Paradoxically, CETP (cholesteryl ester transfer protein) inhibitors have been shown to increase inflammation as revealed by a raised plasma level of high-sensitivity C-reactive protein. CETP inhibitors did not improve clinical outcomes in large-scale clinical trials of unselected patients with coronary disease. Dalcetrapib is a CETP modulator for which effects on cardiovascular outcomes were demonstrated in the dal-OUTCOMES trial to be influenced by correlated polymorphisms in the ADCY9 (adenylate cyclase type 9) gene (P=2.4×10-8 for rs1967309). Patients with the AA genotype at rs1967309 had a relative reduction of 39% in the risk of presenting a cardiovascular event when treated with dalcetrapib compared with placebo (95% confidence interval, 0.41-0.92). In contrast, patients with the GG genotype had a 27% increase in risk, whereas heterozygotes (AG) presented a neutral result. Supporting evidence from the dal-PLAQUE-2 study using carotid ultrasonography revealed that the polymorphisms tested in the ADCY9 linkage disequilibrium block were associated with disease regression for patients with the protective genotype, progression for the harmful genotype, and no effect in heterozygotes (P≤0.05 and ≤0.01 for 10 and 3 polymorphisms, respectively) when comparing dalcetrapib to placebo. Strikingly concordant and significant genotype-dependent effects of dalcetrapib were also obtained for changes in high-sensitivity C-reactive protein and cholesterol efflux capacity. The Dal-GenE randomized trial is currently being conducted in patients with a recent acute coronary syndrome bearing the AA genotype at rs1967309 in the ADCY9 gene to confirm the effects of dalcetrapib on hard cardiovascular outcomes.

E-PodoFavalin-15999 (Atremorine®)-Induced Neurotransmitter and Hormonal Response in Parkinson’s Disease

E-PodoFavalin-15999 (Atremorine®) is a novel bioproduct obtained by means of non-denaturing biotechnological procedures from structural components of the Vicia faba L. plant, for the prevention and treatment of Parkinsonian disorders. Preclinical studies have revealed that Atremorine is a powerful neuroprotectant with specific activity on dopaminergic neurons, reversing neurodegeneration and improving motor function in animal models of Parkinson’s disease (PD). Clinical studies indicate that Atremorine is a powerful catecholaminergic enhancer with time- and genotype-dependent effects in PD. In the present study, we investigated the effects of Atremorine on the levels of neurotransmitters (dopamine, adrenaline, noradrenaline, serotonin) and hormones (adrenocorticotropic hormone (ACTH), growth hormone (GH), prolactin (PRL), follicle-stimulating hormone (FSH), luteinizing hormone (LH), cortisol, estrogens, testosterone) in Parkinsonian patients (n=119) 1 hour after Atremorine administration (5 g/day). These effects were also studied after stratification of PD patients according to their cardiovascular function. Atremorine induced a significant increase in the levels of dopamine (p<0.001), noradrenaline (p=0.004) and adrenaline (p=0.01), and a significant decrease in the levels of PRL (p<0.001), cortisol (p<0.001), and GH (p=0.002), with no apparent changes in the levels of serotonin, ACTH, FSH, LH, testosterone or estrogen. The levels of dopamine were significantly higher in patients with normal EKG than in patients with abnormal electrocardiogram (EKG); however, the levels of adrenaline, noradrenaline and serotonin tended to be lower in patients with normal EKG as compared to patients with abnormal EKG. There were also some differences in hormonal levels in patients with normal EKG, compared to abnormal EKG. These results clearly show that Atremorine is an effective enhancer of catecholaminergic neurotransmission, which contributes to optimization of hormonal regulation in PD.

Grafting and 2.4-dichlorophenoxyacetic acid induced flowering in non-flowering sweetpotato in sub-tropics

In sub-tropics, most sweetpotato genotypes do not flower at all. The aim of this study was to develop methods of inducing flowering based on the application of 2.4-dichlorophenoxyacetic acid (2.4D) and grafting of non-flowering genotypes on rootstocks of genotypes that flower. The first experiment was a 3*4*2 factorial experiment repeated in summer and winter seasons. Four levels of 2.4D (0, 40, 80 and 120ppm) were applied at two levels (once and twice) on three sweetpotato genotypes. The second experiment involved grafting scions of two genotypes on rootstocks of flowering genotype. During summer season there were significant differences (P<0.05) in bud and flower counts among genotypes, concentration and number of split applications but non-significant differences on all the interactions. Genotype 1 produced more buds and flowers followed by genotype 3 and 2. The highest numbers of buds and flowers were observed at 40ppm and 80ppm. At higher concentrations (80 and 120ppm), the physiological disorders including leaf epinasty, shoot die back and stem cracking were extensive. The optimum concentration for 2.4D proved to be 40ppm and could induce flowering with less physiological disorders. Split application resulted in bud death and delayed flowering. Results showed that grafting has a genotype dependent effect on flowering. Genotype 3 managed to flower but genotype 2 did not flower at all when their scions were grafted on the flowering rootstocks of genotype 1.

Genotype-Dependent Effects of COMT Inhibition on Cognitive Function in a Highly Specific, Novel Mouse Model of Altered COMT Activity.

Catechol-O-methyltransferase (COMT) modulates dopamine levels in the prefrontal cortex. The human gene contains a polymorphism (Val158Met) that alters enzyme activity and influences PFC function. It has also been linked with cognition and anxiety, but the findings are mixed. We therefore developed a novel mouse model of altered COMT activity. The human Met allele was introduced into the native mouse COMT gene to produce COMT-Met mice, which were compared with their wild-type littermates. The model proved highly specific: COMT-Met mice had reductions in COMT abundance and activity, compared with wild-type mice, explicitly in the absence of off-target changes in the expression of other genes. Despite robust alterations in dopamine metabolism, we found only subtle changes on certain cognitive tasks under baseline conditions (eg, increased spatial novelty preference in COMT-Met mice vs wild-type mice). However, genotype differences emerged after administration of the COMT inhibitor tolcapone: performance of wild-type mice, but not COMT-Met mice, was improved on the 5-choice serial reaction time task after tolcapone administration. There were no changes in anxiety-related behaviors in the tests that we used. Our findings are convergent with human studies of the Val158Met polymorphism, and suggest that COMT's effects are most prominent when the dopamine system is challenged. Finally, they demonstrate the importance of considering COMT genotype when examining the therapeutic potential of COMT inhibitors.

Genotype-dependent effect of exogenous 24-epibrassinolide on chromium-induced changes in ultrastructure and physicochemical traits in tobacco seedlings.

Greenhouse hydroponic experiments were carried out using three different heavy metal accumulation tobacco genotypes to evaluate how different genotypes responded to chromium (Cr) toxicity in the presence of 24-epibrassinolide (EBR; a biologically active brassinosteroid). The results showed that Cr stress caused a marked reduction in plant biomass, chlorophyll content, chlorophyll fluorescence, and photosynthesis parameters but induced malondialdehyde accumulation and ultrastructure damage, with 2010-38 (L) less affected. Foliar application of 24-epibrassinolide (0.1μM) on Cr-stressed plants greatly alleviated Cr-induced inhibition of growth and photosynthesis, oxidative stress and ultrastructure damage, decreased Cr accumulation in different parts of leaves and roots, with the exception of the upper and lower of leaves of genotype L, and maintained ion homeostasis. Regarding genotypes, L was more tolerant than M and H, as it absorbed less Cr and also performed better in all of the studied parameters. These findings suggest a potential role for 24-epibrassinolide in Cr stress alleviation and the utilization of elite genetic resources in future breeding programs to develop low Cr accumulation genotypes. These results advocate a positive role for 24-epibrassinolide in reducing pollutant residues from health point of view.