William R. Bailey

Genome-wide association study of smoking behaviours in patients with COPD

Background Cigarette smoking is a major risk factor for chronic obstructive pulmonary disease (COPD) and COPD severity. Previous genome-wide association studies (GWAS) have identified numerous single nucleotide polymorphisms (SNPs) associated with the number of cigarettes smoked per day (CPD) and a dopamine beta-hydroxylase (DBH) locus associated with smoking cessation in multiple populations. Objective To identify SNPs associated with lifetime average and current CPD, age at smoking initiation, and smoking cessation in patients with COPD. Methods GWAS were conducted in four independent cohorts encompassing 3441 ever-smoking patients with COPD (Global Initiative for Obstructive Lung Disease stage II or higher). Untyped SNPs were imputed using the HapMap (phase II) panel. Results from all cohorts were meta-analysed. Results Several SNPs near the HLA region on chromosome 6p21 and in an intergenic region on chromosome 2q21 showed associations with age at smoking initiation, both with the lowest p=2×10−7. No SNPs were associated with lifetime average CPD, current CPD or smoking cessation with p<10−6. Nominally significant associations with candidate SNPs within cholinergic receptors, nicotinic, alpha 3/5 (CHRNA3/CHRNA5; eg, p=0.00011 for SNP rs1051730) and cytochrome P450, family 2, subfamily A, polypeptide 6 (CYP2A6; eg, p=2.78×10−5 for a non-synonymous SNP rs1801272) regions were observed for lifetime average CPD, however only CYP2A6 showed evidence of significant association with current CPD. A candidate SNP (rs3025343) in DBH was significantly (p=0.015) associated with smoking cessation. Conclusion The authors identified two candidate regions associated with age at smoking initiation in patients with COPD. Associations of CHRNA3/CHRNA5 and CYP2A6 loci with CPD and DBH with smoking cessation are also likely of importance in the smoking behaviours of patients with COPD.

Effect of dietary copper deficiency on the distribution of dopamine and norepinephrine in mice and rats.

Dietary copper deficiency was produced in Swiss albino mice and Sprague Dawley rats to determine the organ specificity of alterations in norepinephrine (NE) and dopamine (DA) concentrations and the relationship with organ copper levels. A 5-week dietary treatment was used, which started 1 week after birth for mice, initially via dams, and 3 weeks after birth for rats. Mice offspring (6 weeks of age) and rats (8 weeks of age) maintained on a copper-deficient (-Cu) treatment were compared with copper-adequate (+Cu) controls. Compared with +Cu animals, -Cu mice and rats were anemic and had low (<1% of +Cu) ceruloplasmin activities but normal body weights. The -Cu mice had organ copper concentrations ranging between 30% and 65% of +Cu values for eight organs studied, with the thymus being the least depleted. For -Cu rats, the range was 15% to 65%. Significant reductions in NE concentration were observed in the heart, pancreas, and spleen of -Cu mice. Elevated DA levels were observed in all organs except the brain. For -Cu rats, the NE level was lower in the heart and the DA level was higher in both the heart and spleen compared with +Cu rats. Dopamine elevation in the heart and spleen for both -Cu mice and rats was four- and fivefold higher, respectively. Adrenal catecholamine levels were only slightly changed by copper deficiency in mice or rats. Urinary levels of both NE and DA were higher in -Cu rats and mice. Plasma and heart tyrosine levels were not altered in -Cu mice. Elevated DA in -Cu rodents may be due to limiting dopamine-beta-monooxygenase. Higher urinary NE and lower organ NE may be due to a combination of decreased synthesis and enhanced turnover. The magnitude of decreased organ copper was not predictive of altered catecholamine pool size.

Regional Specificity in Alterations of Rat Brain Copper and Catecholamines Following Perinatal Copper Deficiency

Abstract: Perinatal copper deficiency was studied in 1‐month‐old female and male Sprague‐Dawley rat offspring to investigate regional changes in brain copper and catecholamine levels. Offspring of dams given the low copper treatment beginning at day 7 of gestation exhibited signs characteristic of deficiency such as impaired growth and 10‐fold lower liver copper levels compared with copper‐adequate controls. Regional analysis of brain copper by graphite furnace atomic absorption spectroscopy revealed uniform and severe reduction of copper to levels 20 ± 3% of controls in all regions, except the hypothalamus, where reductions to 56 and 28% of those in copper‐adequate females and males, respectively, were measured. HPLC analysis revealed significant reductions in norepinephrine levels in cerebrum, midbrain, corpus striatum, cerebellum, and medulla‐pons of copper‐deficient offspring ranging between 39 and 67% of control values. There were no significant differences in norepinephrine concentration in the hypothalamus. There was a significant, one‐third reduction of dopamine in the corpus striatum of copper‐deficient male rats. Consistent with altered in vivo dopamine β‐monooxygenase activity, there were five‐, three‐, and twofold elevations of dopamine in cerebellum, medulla‐pons, and hypothalamus of copper‐deficient rats. Spectrophotometric measurement of in vitro dopamine β‐monooxygenase activity of brain and adrenal homogenates was higher in copper‐deficient rats, confirming prior work. An explanation for the in vitro data is unclear. Changes in copper and catecholamine levels were influenced by diet and were regionally selective, especially in the hypothalamus.

Alterations of Rat Brain Peptidylglycine α-Amidating Monooxygenase and Other Cuproenzyme Activities Following Perinatal Copper Deficiency

Abstract Perinatal copper (Cu) deficiency was studied in month-old female and male Sprague-Dawley rat offspring to investigate regional changes in brain cuproenzymes. Offspring of dams given the low Cu treatment beginning at Day 7 of gestation exhibited signs characteristic of Cu deficiency including a 70% reduction in liver Cu levels compared with Cu-adequate controls. Compared with Cu-adequate rats, Cu-deficient rats had lower activities of the cuproenzymes peptidylglycine α-amidating monooxygenase (PAM), cytochrome c oxydase (CCO), and Cu, Zn-superoxide dismutase (SOD) in all six brain regions studied. Apparent activity of dopamine-β-monooxygenase (DBM) was higher in all regions from Cu-deficient compared with Cu-adequate rats. Activity of the selenoenzyme glutathione peroxidase (GPX) was not greatly altered in brain by Cu deficiency. Following 1 month of Cu repletion, liver but not brain Cu levels were equivalent to control. Brain CCO activity was still lower in Cu-repleted female and male rats. PAM activity was still lower in cerebrum of Cu-repleted rats. DBM activity was no longer significantly elevated in the former Cu-deficient groups except for midbrain. SOD and GPX activity were equivalent between groups. PAM activity, in vitro, is lower in the brain following perinatal Cu deficiency and activity is slow to recover following nutritional supplementation with Cu. Perhaps neuropeptide maturation is compromised by Cu deficiency.

Persistent neurochemical changes following perinatal copper deficiency in rats

Abstract Copper (Cu) levels in the central nervous system are known to be influenced by Cu nutriture during perinatal development. A rat model of dietary Cu deficiency (−Cu), initiated during gestation, was employed to examine changes in regional levels of brain Cu, norepinephrine (NE), dopamine (DA) and selected enzymes of month-old female Sprague Dawley offspring. Nutritional repletion of Cu to −Cu rats was studied following 1, 2, or 4 months. Levels of Cu in six different regions of brain from −Cu rats were reduced 80% or greater compared with levels in Cu-adequate (+Cu) controls. Following 4 months of Cu repletion, brain regional Cu levels were still below those measured in +Cu rats. The norepinephrine/dopamine was higher in +Cu compared with −Cu rats for all brain regions except corpus striatum suggesting impairment of dopamine-β-monooxygenase (DBM). Cu repletion normalized these ratios after 1 month. In vitro DBM activity was higher in samples from −Cu compared with +Cu rats and these differences were reversed by Cu supplementation. Cytochrome c oxidase (CCO) activity of 1-month-old −Cu rats was 18 to 50% of that measured in +Cu rats. Small differences in CCO activity remained in all regions, except hypothalamus, even after 4 months of repletion. Cu,Zn-superoxide dismutase activity (Cu,Zn-SOD) in −Cu rats was 75 to 91% of that measured in +Cu rats and was equivalent following repletion for 1 month. Four months of Cu repletion were not sufficient to restore brain Cu and CCO levels of 1-month-old −Cu rats.

Copper deficiency during neonatal development alters mouse brain catecholamine levels

Abstract Copper (Cu) deficiency was investigated in Swiss Webster albino mice to develop a model that alters brain catecholamine metabolism without serious growth impairment. Cu deficiency was induced by feeding a diet low in Cu (modified AIN-76A) to dams beginning either 7 days prior, 4 days prior, 4 days after, or on the day of parturition. Control dams were given Cu in the drinking water (20 μg Cu/mL); their pups were considered Cu-adequate (+Cu). Mice were weaned when three weeks old and were maintained on the treatment of their dams for an additional week. All four-week-old male Cu-deficient (−Cu) offspring exhibited biochemical characteristics of Cu deficiency when compared to their respective +Cu control mice. However, the best model, which resulted in altered catecholamine metabolism characterized by elevation of dopamine (DA) and depression in norepinephrine (NE) in brain and heart, was when treatment began 4 days prior to birth. Body and brain weight were not altered. However, levels of Cu in brain were markedly reduced to 21% of those measured in +Cu controls. Furthermore, brain NE and DA concentrations of −Cu mice were 72% and 132% of those quantified in +Cu offspring, respectively. Confirmatory studies demonstrated equivalent results in female offspring. One-month dietary Cu repletion studies reversed most biochemical changes in the “−Cu” mice. Brain Cu levels remained lower in repleted mice (55% of those in +Cu controls). These data and previous results support the hypothesis that neonatal Cu deficiency results in permanent changes to the brain.

Ascorbic acid synthesis and concentrations in organs of copper-deficient and brindled mice

Copper deficiency was studied in mice to investigate an interaction between copper and ascorbic acid. Twelve-day-old mutant brindled mice that exhibited signs of copper deficiency were compared to their normal brothers as well as to age-matched suckling mice that were copper deficient (-Cu) because their dams were consuming a copper-deficient diet throughout gestation and lactation, and a fourth group of copper-supplemented ( + Cu) suckling mice that served as dietary controls. Dietary copper deficiency was also produced in older mice by beginning the treatment at birth and continuing for 7 wk. Organ ascorbate levels were determined by high performance liquid chromatography with electrochemical detection. Differences caused by diet and genetics were evident but age-dependent. Compared to controls, liver and kidney ascorbate levels did not change remarkably in young or old copper-deficient mice. Cardiac ascorbate levels were higher in 7-wk-old - Cu mice and lower in 12-d-old - Cu mice, despite hypertrophy in both cases. Spleen ascorbate levels were lower in older -Cu mice and higher in 12-d-old mice, but total spleen ascorbate reflected the hypertrophic and atrophic size in the older and younger -Cu mice, respectively. Brindled mutants had an extremely low level of ascorbate in spleen. Plasma ascorbate was lower in 7-wk-old - Cu mice. Reasons for the alterations in ascorbate levels are not known. Synthesis in liver from D-glucuronate was not altered by dietary copper deficiency in 7-wk-old mice. Synthesis was lower in livers from 12-d-old - Cu and brindled mice compared to control values. However, the difference correlated better with body weight of the mice rather than with degree of copper deficiency. Consequences of the altered organ levels of ascorbate in copper-deficient mice are not completely known.