Matti K. Karvonen

ASSOCIATION OF A LEUCINE(7)-TO-PROLINE(7) POLYMORPHISM IN THE SIGNAL PEPTIDE OF NEUROPEPTIDE Y WITH HIGH SERUM CHOLESTEROL AND LDL CHOLESTEROL LEVELS

High serum levels of total and LDL cholesterol are important risk factors in the development of atherosclerotic coronary artery disease. Cholesterol metabolism is affected by nutritional, environmental and genetic factors. Neuropeptide Y (NPY), which is widely expressed in both the central and peripheral nervous systems, has an important role in the hypothalamic regulation of energy balance by stimulating food intake and favoring energy storage through increased lipoprotein lipase activity in white adipose tissue. As a part of ongoing study of the genetic basis of obesity, we screened the NPY gene for sequence variants. We report here the identification of a common Leu(7)-to-Pro(7) polymorphism in the signal peptide of NPY. Presence of this Pro(7) in NPY was associated with higher serum levels of total and LDL cholesterol in obese subjects participating in two independent Finnish and Dutch studies. Furthermore, normal-weight Finns with Pro(7) also had higher serum levels of total and LDL cholesterol than did subjects with Leu(7)/Leu(7), as analyzed in three subsequent determinations at 5-year intervals during a 10-year follow-up period. The NPY polymorphism was not associated with higher cholesterol levels in normal-weight Dutch. Our study provides evidence that NPY is linked to cholesterol metabolism and that the polymorphism producing Pro(7) in NPY is one of the strongest genetic factors identified thus far affecting serum cholesterol, particularly in obese subjects.

Association between the functional variant of the catechol-O-methyltransferase (COMT) gene and type 1 alcoholism.

Catechol-O-methyltransferase (COMT) is an enzyme which has a crucial role in the metabolism of dopamine. It has been suggested that a common functional genetic polymorphism in the COMT gene, which results in 3 to 4-fold difference in COMT enzyme activity,1,2 may contribute to the etiology of mental disorders such as bipolar disorder and alcoholism.1 Since ethanol-induced euphoria is associated with the rapid release of dopamine in limbic areas, it is conceivable that subjects who inherit the allele encoding the low activity COMT variant would have a relatively low dopamine inactivation rate, and therefore would be more vulnerable to the development of ethanol dependence. The aim of this study was to test this hypothesis among type 1 (late-onset) alcoholics. The COMT polymorphism was determined in two independent male late onset (type 1) alcoholic populations in Turku (n = 67) and Kuopio (n = 56). The high (H) and low (L) activity COMT genotype and allele frequencies were compared with previously published data from 3140 Finnish blood donors (general population) and 267 race- and gender-matched controls. The frequency of low activity allele (L) was markedly higher among the patients both in Turku (P = 0.023) and in Kuopio (P = 0.005) when compared with the general population. When all patients were compared with the general population (blood donors), the difference was even more significant (P = 0.0004). When genotypes of all alcoholics (n = 123) were compared with genotypes of matched controls, the odds ratio (OR) for alcoholism for those subjects having the LL genotype vs those with HH genotype was 2.51, 95% CI 1.22–5.19, P = 0.006. Also, L allele frequency was significantly higher among alcoholics when compared with controls (P = 0.009). The estimate for population etiological (attributable) fraction for the LL genotype in alcoholism was 13.3% (95% CI 2.3–25.7%). The results indicate that the COMT polymorphism contributes significantly to the development of late-onset alcoholism.

Altered intracellular processing and release of neuropeptide Y due to leucine 7 to proline 7 polymorphism in the signal peptide of preproneuropeptide Y in humans.

SPECIFIC AIMSTo study the functional role of the recently found leucine 7 to proline 7 (Leu7Pro) polymorphism in the signal peptide of preproneuropeptide Y, sympathetic responses, including neurope...

An insertion/deletion polymorphism in the α2b-adrenergic receptor gene is a novel genetic risk factor for acute coronary events

OBJECTIVES Our aim was to study whether an insertion/deletion (I/D) polymorphism in the a2Badrenoceptor gene is associated with the risk for cardiovascular diseases. BACKGROUND a2-adrenoceptors mediate contraction of vascular smooth muscle and induce coronary vasoconstriction in humans. The a2-adrenoceptor subtype B mediates vasoconstriction in mice. A variant of the human a2B-adrenoceptor gene that encode saDo fthree residues in an intracellular acidic motif has been shown to confer decreased receptor desensitization. This receptor variant could, therefore, be involved in diseases associated with enhanced vasoconstriction. METHODS This study was part of a prospective population-based study investigating risk factors for cardiovascular diseases in a cohort of middle-aged men from eastern Finland. Nine hundred twelve men aged 46 to 64 years were followed for an average time of 4.5 years. RESULTS In this study population, 192 men (21%) had the D/D genotype; 256 (28%) had the I/I genotype, and 464 (51%) had a heterozygous genotype. In a Cox model adjusting for other coronary risk factors, men with the D/D genotype had 2.2 times (95% confidence interval: 1.1 to 4.4, p 5 0.02) the risk to experience an acute coronary event (n 5 15 for D/D, 10 for I/I and 12 for I/D) compared with men carrying either of the other two genotypes. The a2B-adrenoceptor genotype was not associated with hypertension in this study population. CONCLUSIONS The D/D genotype of the a2B-adrenoceptor is a novel genetic risk factor for acute coronary events, but not for hypertension. (J Am Coll Cardiol 2001;37:1516 ‐22)

Suicidal behavior in patients with schizophrenia is related to COMT polymorphism

&NA; A common functional polymorphism that results in a three‐ to four‐fold difference in catechol‐O‐methyltransferase (COMT) enzyme activity has been related to psychiatric disorders such as ultra‐ultra rapid cycling bipolar disorder, drug abuse and alcoholism (Lachman et al., 1996a; Karayiorgou et al., 1997; Vandenbergh et al., 1997; Papolos et al., 1998; Tiihonen et al., 1999). Several studies have also reported associations between the allele encoding the low enzyme activity COMT variant (L allele) and other‐directed aggression (Strous et al., 1997; Lachman et al., 1998; Kotler et al., 1999) in schizophrenic and schizoaffective patients. The current study investigated whether the COMT L allele is also associated with suicide attempts in schizophrenic and schizoaffective patients. COMT genotypes were determined and history of suicide attempts was retrospectively investigated in a Finnish sample (n = 94) and a US sample (n = 54). Significant associations were observed between COMT genotype and suicide; specifically, history of violent suicide attempts. The COMT L allele was more frequent in subjects who had attempted suicide by violent means. These associations were significant in males but not females. These findings support a common neurobiological substrate for self‐ and other‐directed aggression, and suggest that catecholaminergic alterations may contribute to these behaviors in schizophrenic and schizoaffective patients.

Leucine7 to proline7 polymorphism in the preproneuropeptide Y is associated with the progression of carotid atherosclerosis, blood pressure and serum lipids in Finnish men

A rather common leucine7-to-proline7 (Leu7Pro) polymorphism in the preproneuropeptide Y (prepro-NPY) gene signal peptide may be important in blood pressure regulation, cholesterol metabolism and the pathogenesis of atherosclerosis in humans. We examined the associations of the Leu7Pro polymorphism with carotid atherosclerotic progression, blood pressure and serum lipids in a population-based sample of 966 men aged 42-60 years in Finland. The Pro7 substitution (carrier frequency 12.2%) was associated with accelerated four-year increase in the mean (P=0.01) and maximal (P=0.007) common carotid intima-media thickness (IMT) and with slightly increased systolic (P=0.03) and diastolic (P=0.02) blood pressures, adjusted for other major risk factors. Men with Pro7 substitution had 30.6% (95% CI 6.9-54.0%) greater increase in the mean IMT and 20.0% (95% CI 5.3-34.4%) greater increase in the maximal IMT than men with Leu7/Leu7 genotype. The Pro7 substitution was also related to increased serum total cholesterol (P=0.01) and LDL cholesterol (P=0.02) in obese (body mass index (BMI)>30 kg/m(2)) men. This study provides important evidence suggesting that the Pro7 substitution in the prepro-NPY is an important risk factor for accelerated atherosclerotic progression, increased blood pressure and increased serum cholesterol in humans.

Neuropeptide Y and Y2‐receptor are involved in development of diabetic retinopathy and retinal neovascularization

BACKGROUND: Neuropeptide Y is a sympathetic neurotransmitter, a potent endothelium‐derived angiogenic factor and a vascular mitogen. We have studied the role of the functional leucine7 to proline7 polymorphism of the signal peptide region of preproneuropeptide Y (prepro‐NPY) as a genetic susceptibility factor for diabetic retinopathy. In addition, we investigated the role of the NPY Y2‐receptor as a putative mediator of angiogenic NPY signaling in the retina. METHODS: Frequencies of proline7 (Pro7) carriers in the prepro‐NPY were determined in type 1 and type 2 diabetes patients having retinopathy, in type 2 diabetes patients without retinopathy and in healthy control subjects. The role of Y2‐receptor in hyperoxemia‐induced retinal neovascularization was investigated in Y2‐receptor knockout mice (Y2 −/− ) and in rats administered Y2‐receptor mRNA antisense oligonucleotide. RESULTS: The carriers having Pro7 in the preproNPY are markedly over‐represented among type 2 diabetes patients with retinopathy compared to type 2 diabetes patients without retinopathy and to the population control. Neonatal exposure to hyperoxia resulted in development of retinal neovascularization that was prevented in Y2 −/− ‐mice, and significantly inhibited in rats treated with the Y2‐receptor antisense oligonucleotide. CONCLUSIONS: NPY and Y2‐receptor play important roles in diabetic retinopathy and retinal neovascularization and are thus potential new targets for drug molecules for treatment of retinopathy.

Analysis of monoamine oxidase A (MAOA) promoter polymorphism in Finnish male alcoholics

Alterations in monoamine oxidase A (MAOA) expression and enzyme activity may be associated with alcoholism and impulsive behavior. Therefore, functional polymorphisms in the MAOA gene would be good candidates to consider in the interindividual differences that exist in the susceptibility to alcoholism. One variant that has been considered as a candidate in alcoholism is a repeat polymorphism in the MAOA gene promoter. We analyzed a cohort of Finnish males with either type 1 or type 2 alcoholism, as well as controls, for differences in the distribution of MAOA promoter alleles. Based on other studies, we postulated that type 2 alcoholism, which is associated with antisocial behavior, but not type 1 alcoholism, would be correlated with the inheritance of the low promoter activity allele. However, we failed to find a difference in allele distribution in type 1 and type 2 alcoholics. In addition, there was no difference in the allele distribution when each group of alcoholics was compared with controls. However, when both groups of alcoholics were pooled and compared with controls, the difference in allele distribution reached a trend towards significance. Our results suggest a minimal association between the MAOA low activity promoter alleles and alcoholism, regardless of the presence or absence of antisocial behavior. Interestingly, approximately 3% of type 2 alcoholics were found to be heterozygous for the MAOA promoter polymorphism. Since MAOA is X-linked, the heterozygotes are probable cases of Klinefelters syndrome (47,XXY) suggesting that X-chromosome aneuploidy may increase the risk for developing type 2 alcoholism.

Lack of association between the functional variant of the catechol-o-methyltransferase (COMT) gene and early-onset alcoholism associated with severe antisocial behavior

Addictive drugs, including ethanol, increase the brains dopaminergic transmission, and catechol-o-methyltransferase (COMT) enzyme has a crucial role in dopamine inactivation. A common functional polymorphism in the COMT gene results in a three- to four-fold variation in enzyme activity. In a previous study, we found an association between type 1 (with late-onset but without prominent antisocial behavior) alcoholism and the low activity allele of the COMT gene. In this work we analyzed whether the COMT polymorphism has any effect on the development of type 2 (with early-onset and habitual impulsive violent behavior) alcoholism. The COMT genotype was determined in 62 impulsive violent recidivist offenders with early-onset (type 2) alcoholism, 123 late-onset nonviolent (type 1) alcoholics, and 267 race and gender-matched controls. The allele and genotype frequencies of these groups were compared with each other and also with previously published data from 3,140 Finnish blood donors. The type 2 alcoholics did not differ from either the blood donors or the controls. The low activity (L) allele frequency was higher among type 1 alcoholics (chi(2) = 4.98, P = 0.026) when compared with type 2 cases. The odds ratio for type 1 alcoholism as compared with type 2 alcoholism for those subjects with the LL genotype versus the HH genotype was 3.0 (95% confidence interval 1.1-8.4, P = 0.017). The results suggest that COMT genotype has no major role in the development of early-onset alcoholism with severe antisocial behavior.

Enhanced endothelium-dependent vasodilation in subjects with Proline7 substitution in the signal peptide of neuropeptide Y

Neuropeptide Y (NPY) is a molecule that may have both vasoconstrictive and vasodilatory actions. A common polymorphism in the human NPY gene that results in the Leucine7 to Proline7 substitution (Leu7Pro) in the signal peptide part of the NPY was recently identified. This substitution has been associated with elevated serum cholesterol levels and with slightly accelerated progression rate of carotid intima-media thickness, thus suggesting increased risk of atherosclerosis in carriers of Pro7 substitution. Recent data also indicate that subjects with Pro7 substitution may have increased endothelial release of NPY. This study was undertaken to elucidate the effects of Leu7Pro polymorphism on arterial endothelial function. We measured flow-mediated endothelial-dependent dilatation (FMD) of the brachial artery in two separate populations: in 152 middle-aged men and in 95 prepubertal children. In both study populations, subjects with Pro7 substitution had 48-52% higher FMD compared with subjects having the wildtype (Leu7/Leu7) signal peptide sequence. We conclude that Pro7 substitution in signal peptide of the NPY is associated with enhanced endothelial-dependent vasodilation. Prospective studies are needed to determine whether Pro7 substitution is associated with increased or decreased risk of cardiovascular morbidity and mortality.