the hsd11b1 gene located at 1q32.2 ( tannin et al . , 1991 ) encodes the microsomal enzyme 11-hydroxysteroid dehydrogenase type 1 ( 11-hsd1 ) that is responsible for the conversion of the hormone cortisol ( also known as stress hormone ) to its metabolically inactive form cortisone ( ricketts et al . , 1998 ) .
an imbalance in the levels of cortisol is associated with visceral fat , insulin resistance and hyperlipidemia , all of which are related to obesity , type 2 diabetes mellitus ( t2 dm ) and metabolic syndrome ( paterson et al . , 2004 ) .
various studies have suggested an important functional role of 11-hsd1 in the metabolic processes underlying these pathologies . in knockout animals ,
the absence of 11-hsd1 had a protective effect against insulin resistance and hyperglycemia because of the lack of glucocorticoid regeneration in the liver and adipose tissue ( kotelevtsev et al .
the reverse situation was seen in transgenic animals with over - expression of 11-hsd1 , with an increase in the concentration of intra - adipocyte glucocorticoid , hyperglycemia and a marked central obesity phenotype ( masuzaki et al . , 2001 ) .
in particular , the g allele of snp rs12086634 was associated with lower 11-hsd1 transcription in vitro ( draper et al . , 2003 ) .
the association between the rs846910 polymorphism in the p2 promoter region and rs12086634 in an enhancer of the hsd11b1 gene has been investigated in several clinical contexts ( gambineri et al . , 2011 ; moon et al . , 2011 ; utriainen et al . ,
, these two snps were associated with t2 dm , but not with obesity ( nair et al . , 2004 ) .
( 2011 ) found that the combination of these snps in caucasian women of northern italy was associated with a higher risk of metabolic syndrome , regardless of the diagnosis of polycystic ovary syndrome .
in other studies , both snps were associated with t2 dm and/or hypertension ( freedman et al .
considering the wide range of biochemical and physiological effects of cortisol , it is possible that temporal or tissue - specific changes in the levels of this hormone could influence a wide range of complex diseases , including obesity and metabolic syndrome .
since the occurrence of genetic polymorphisms in the hs11b1 gene could influence cortisol levels , in this study we investigated the influence of two snps of the hsd11b1 gene ( rs846910 and rs12086634 ) on anthropometric and biochemical variables associated with obesity and metabolic syndrome in an adult population from southern brazil .
the sample consisted of 215 workers of euro - brazilian descent employed by the federal university of paran in southern brazil .
since the aim in selecting the volunteers was to obtain a sample representative of the population heterogeneity , no pathology was used as an inclusion or exclusion criterion .
one hundred and forty - seven women ( 2272 years old , 56% overweight and obese ) and 68 men ( 2360 years old , 23% overweight and obese ) participated in the study .
assessment of the physical activity of the volunteers for seven days using a pedometer ( yamax digi - walker sw-700 ) showed that 23% were sedentary , 37% had low physical activity , 26% were active and 14% had high physical activity [ according to criteria proposed by wyatt et al .
individuals were considered obese when the body mass index ( bmi ) was 30 kg / m and non - obese when the bmi was < 30 kg / m .
weight and height were measured with an accuracy of 0.1 kg and 0.1 cm , respectively .
glucose , triglycerides ( tg ) , total cholesterol ( tc ) and hdl - cholesterol ( hdl - c ) were measured by standard automated methods .
ldl - cholesterol ( ldl - c ) levels were calculated using the friedewald equation ( friedewald et al . , 1972 ) .
dna was extracted from peripheral blood by a salting - out method ( lahiri and nurnberger jr , 1991 ) and then diluted to a final concentration of 20 ng/l .
the variant located in the p2 promoter 5urr ( rs846910 ; snp1 ; g / a ) and the variant in the enhancer region of intron 3 ( rs12086634 ; snp2 ; t / g ) were genotyped with a taqman snp genotyping assay ( applied biosystems ) .
the reactions were done in a mastercycler realplex 2 ( eppendorf ) using the following conditions : 50 c for 2 min , 95 c for 10 min and 50 cycles of 95 c for 15 s and 62 c for 1 min .
three previously sequenced control samples , representative of each of the possible genotypes , were included in each reaction for both snps .
frequency distributions , variances , the shapiro - wilk normality test , students t - test and the mann - whitney test were calculated using statistica for windows v. 5.0 ( statsoft inc .
chi - square tests were done using clump ( sham and curtis , 1995 ) .
multiple regression analyses were done using spss for windows v. 13.0 ( spss inc . ,
the sample consisted of 215 workers of euro - brazilian descent employed by the federal university of paran in southern brazil .
since the aim in selecting the volunteers was to obtain a sample representative of the population heterogeneity , no pathology was used as an inclusion or exclusion criterion .
one hundred and forty - seven women ( 2272 years old , 56% overweight and obese ) and 68 men ( 2360 years old , 23% overweight and obese ) participated in the study .
assessment of the physical activity of the volunteers for seven days using a pedometer ( yamax digi - walker sw-700 ) showed that 23% were sedentary , 37% had low physical activity , 26% were active and 14% had high physical activity [ according to criteria proposed by wyatt et al .
individuals were considered obese when the body mass index ( bmi ) was 30 kg / m and non - obese when the bmi was < 30 kg / m .
weight and height were measured with an accuracy of 0.1 kg and 0.1 cm , respectively .
glucose , triglycerides ( tg ) , total cholesterol ( tc ) and hdl - cholesterol ( hdl - c ) were measured by standard automated methods .
ldl - cholesterol ( ldl - c ) levels were calculated using the friedewald equation ( friedewald et al . , 1972 ) .
dna was extracted from peripheral blood by a salting - out method ( lahiri and nurnberger jr , 1991 ) and then diluted to a final concentration of 20 ng/l .
the variant located in the p2 promoter 5urr ( rs846910 ; snp1 ; g / a ) and the variant in the enhancer region of intron 3 ( rs12086634 ; snp2 ; t / g ) were genotyped with a taqman snp genotyping assay ( applied biosystems ) .
the reactions were done in a mastercycler realplex 2 ( eppendorf ) using the following conditions : 50 c for 2 min , 95 c for 10 min and 50 cycles of 95 c for 15 s and 62 c for 1 min .
three previously sequenced control samples , representative of each of the possible genotypes , were included in each reaction for both snps .
frequency distributions , variances , the shapiro - wilk normality test , students t - test and the mann - whitney test were calculated using statistica for windows v. 5.0 ( statsoft inc .
chi - square tests were done using clump ( sham and curtis , 1995 ) .
multiple regression analyses were done using spss for windows v. 13.0 ( spss inc . ,
table 1 shows the allele frequencies of the two snps in the sample stratified by sex and bmi .
the genotype frequencies in the overall sample and in the groups were in hardy - weinberg equilibrium .
there was no difference in allele frequency between obese and non - obese men (  = 0.62 , p = 0.43 ) , nor between obese and non - obese women (  = 0.17 , p = 0.67 ) for snp1 .
a similar result was found in comparisons between obese and non - obese men and women for snp2 allele frequencies (  = 0.33 , p = 0.56 and  = 0.67 , p = 0.41 , respectively ) .
allele frequencies ( % ) of snps 1 and 2 of the hsd11b1 gene in samples stratified by sex ( men and women ) and bmi ( obese and non - obese ) .
obese individuals : bmi  30 kg / m ; non - obese individuals : bmi < 30 kg / m .
analyses performed with the stratification of the sample only by gender , showed that there was no significant difference in the bmi of men and women ( 26.87 4.00 and 27.20 5.44 , respectively ; p = 0.97 ) .
however , there were significant differences in the hdl - c , tg and glucose levels of men and women , regardless of the hsd11b1 genotype ( p = 0.00001 , p = 0.001 and p = 0.006 , respectively ) ( figure 1 ) .
hdl - c , tc , tg , ldl - c and glucose levels in men and women .
combination analysis with the two snps ( rs846910 and rs12086634 ) revealed no significant associations , in contrast to the findings of gambineri et al .
however , when the effects of each of the two hsd11b1 snps on the biochemical variables and bmi were analyzed separately in men and women , significant differences were found only in women ( table 2 ) .
carriers of the a allele ( rare ) of snp1 had significantly higher hdl levels when compared to individuals homozygous for the g allele ( common ) .
similarly , carriers of the g allele ( rare ) of snp2 had higher glucose levels ( tending to significance , p = 0.06 ) compared to women homozygous for the t allele ( common ) .
multiple regression analysis was used to confirm the effect of these genetic variants on hdl and glucose levels ( table 3 ) . when hdl - c was used as the dependent variable and snp1 , age and bmi as the independent variables
, the analyses showed that bmi and snp1 were independent factors in determining the hdl - c levels in women (  = 0.37 0.11 , p = 0.002 and  = 0.22 0.10 , p = 0.03 , respectively ) .
similar results were obtained when glucose was used as the dependent variable and age , bmi and snp2 were the dependent variables , i.e. , bmi and snp2 were independent factors for increasing glucose levels (  = 0.46 0.10 , p = 0.00002 and  = 0.19 0.09 , p = 0.03 , respectively ) .
comparison of the bmi , hdl - c , tc , tg , ldl - c and glucose levels in men and women stratified as carriers of common ( g or t ) or rare ( a or g ) alleles of snps 1 and 2 .
the data are expressed as the mean sem of the number of individuals indicated in parentheses .
statistical comparisons between men and women ( stratified by genotype ) were done using students t - test ( t ) for parametric data and the mann - whitney test ( z ) for non - parametric data .
cellular responses to stress , even before exposure to sex hormones , are different in men and women ( du et al . , 2004 ) and probably reflect gender - related differences in metabolic pathways ( pollitzer , 2013 ) .
indeed , the higher prevalence of obesity and diabetes in women compared to men , especially after the onset of menopause ( ryan , 2009 ) , indicates that gender - related metabolic differences can influence the mechanisms of these diseases .
the greater amount of visceral fat and higher fat content in the liver correlate with the lack of a protective effect of estrogen in premenopausal women ( geer and shen , 2009 ) .
however , the effect of estrogen on the metabolism of adipose tissue is not fully understood .
premenopausal women have a higher density of antilipolytic 2-adrenergic receptors than men ( richelsen , 1986 ) .
pedersen et al . ( 2004 ) demonstrated that estradiol increases the expression of this receptor in human adipocytes through activation of er- receptors only in subcutaneous adipose tissue , with no effect on visceral adipose tissue .
the activity of lpl ( lipoprotein lipase ) , which controls fat uptake in adipocytes , is also influenced by estradiol since this hormone has transcriptional inhibitory effects ( homma et al .
, 2000 ) and decreases the transcription and enzymatic activity of 11-hsd1 in rodents ( new et al . , 2000 ) .
postmenopausal women with normal weight show enhanced 11-hsd1 activity in adipose tissue and liver ( andersson et al . , 2009 ) , suggesting that low estrogen levels may up - regulate 11hsd1 activity and contribute to the imbalance of energy metabolism influenced by cortisol . in a study of inflammatory bowel disease ,
11-hsd1 expression was higher in male than in female patients , whereas 11-hsd2 showed not gender - specific regulation in its expression ( stegk et al . , 2009 ) .
as shown here , only in women was the presence of the rare snp2 allele associated with higher glucose levels , whereas women homozygous for the common snp1 allele showed lower hdl - c levels compared to rare allele carriers . the combination of higher glucose levels and lower hdl - c levels may have an important role in the development of pathologies associated with obesity .
lower than normal levels of hdl - c have been related to the early development of t2 dm ( von eckardstein et al . , 2000 ) , and conditions such as insulin resistance and obesity may also be related to lower hdl - c levels and to the generation of small particles of hdl - c that can result in several functional changes ( goff et al . , 2005 ) .
men and woman show different responses to the same food intake . compared to women ,
men have higher levels of postprandial insulin and tg ( cohn et al . , 1988 ) , suggesting that estrogen also has a beneficial effect on tg levels in response to food ingestion ( westerveld , 1998 ) .
the sexual dimorphism in tg levels was striking in our study : women had a normal mean tg level , whereas men had a borderline mean tg level , based on age- and gender - related reference values for tg .
since the women in this study were more sensitive to the effects of the hsd11b1 gene polymorphisms investigated here , it is possible that such variations influence the modulation established between estrogen and 11-hsd1 .
the presence of the two snps may alter the down - regulation caused by estrogen and possibly lead to an imbalance in the metabolic pathways involved in glucose and fat metabolism .
the g allele of snp rs12086634 was associated with lower transcriptional activity in vitro ( draper et al . , 2003 ) , whereas the less frequent allele combination ( a and g ) for these two snps ( rs846910 and rs12086634 , respectively ) was associated with higher mrna levels and 11-hsd1 activity in adipose tissue in southern european caucasian women with and without polycystic ovary syndrome ( pcos ) ( gambineri et al . , 2011 ) .
other studies found no relationship between one or both variants and 11-hsd1 levels ( nair et al .
, 2004 ; millan et al . , 2005 ; white , 2005 ; malavasi et al . , 2010 ) .
the genetic variants investigated here may have increased the hsd11b1 transcriptional levels , leading to an imbalance of homeostasis via estradiol down - regulation in women .
a similar effect was not detected in males , suggesting that mechanisms other than estradiol suppress the influence of these genetic polymorphisms on the metabolic variables investigated in this study .
our results showed that the snps and bmi had an independent effect on glucose and hdl - c levels in women , although other factors need to be considered .
other variables such as smoking ( brischetto et al . , 1983 ) , abdominal fat distribution ( ostlund et al . ,
1990 ) and aerobic exercise training ( kokkinos and fernhall , 1999 ) have also been related to hdl - c levels . moreover ,
a decline in hdl - c levels simultaneous to the decline in estrogen levels in post - menopausal woman has been described ( li et al . , 1996 ; senoz et al .
1997 ) ; insulin and glucose levels have also been related to hdl - c , which suggests an influence of carbohydrate metabolism and sex hormone status on the levels of this lipoprotein ( sowers and sigler , 1999 ) . among the limitations of this study
was the sample size , which may not have been large enough to detect any other significant results .
in addition , a functional relationship between the variants studied would be better established in a case - control study , especially with post - menopausal women . in conclusion
, we found that the snps investigated here acted as independent factors in determining glucose and hdl - c levels only in women .
this finding suggested a potentially important sexually dimorphic effect that may be related to gene regulation exerted by estrogen .
 in this study , we investigated the influence of two snps ( rs846910 and rs12086634 ) of the hsd11b1 gene that encodes 11-hydroxysteroid dehydrogenase type 1(11-hsd1 ) , the enzyme that catalyzes the conversion of cortisol to cortisone , on variables associated with obesity and metabolic syndrome in 215 individuals of both sexes from southern brazil . 
 the hsd11b1 gene variants were genotyped using the taqman snp genotyping assay . 
 glucose , triglycerides , total cholesterol , hdl - cholesterol and ldl - cholesterol were measured by standard automated methods . 
 significant results were found in women , with carriers of the g allele of snp rs12086634 having higher glucose levels than non - carriers . 
 carriers of the a allele of snp rs846910 had higher levels of hdl - cholesterol . 
 the involvement of both polymorphisms as independent factors in determining the levels of glucose and hdl - cholesterol was confirmed by multiple regression analysis (  = 0.19 0.09 , p = 0.03 and = 0.22  0.10 , p = 0.03 , respectively ) . 
 our findings suggest that the hsd11b1snps studied may indirectly influence glucose and hdl - cholesterol metabolism in women , possibly through down - regulation of the hsd11b1 gene by estrogen . 
Introduction
Materials and Methods
Subjects
DNA analysis
Statistical analysis
Results
Discussion