Shinshu Med J, 65⑶：153～162, 2017

Lack of Association of Serotonin 2A Receptor Gene in JapanesePatients with Obstructive Sleep Apnea Syndrome
Kayoko Ikegawa1）, Masao Ota2）, Nobumitsu Kobayashi1）, Yunden Droma1）
Hironobu Yaegashi3）, Michiko Nishizawa1）, Masanori Yasuo1）, Kazuhisa Urushihata1）
Hiroshi Yamamoto1） and Masayuki Hanaoka1）＊
1） The First Department of Internal Medicine, Shinshu University School of Medicine
2） Department of Legal Medicine, Shinshu University School of Medicine
3） Hiro Internal Medicine Clinic

Background : The contraction of the genioglossus muscle is realized by the binding of serotonin with serotonin
2A receptor through modulating the hypoglossal motor output. When the genioglossus muscle relaxes, it causes
glossoptosis and upper airway obstruction. Therefore, the variations of the serotonin 2A receptor gene (HTR2A)
are hypothesized to be associated with obstructive sleep apnea syndrome (OSAS) according to the pathogenesisof OSAS. To investigate the association of the HTR2A gene with OSAS in the Japanese population, we conduct­
ed the current case-control association study.
Methods : The subjects included 145 male patients with OSAS who were diagnosed by overnight polysomnogra­phy (PSG) and 133 male controls who were normal in PSG. All the subjects were of Japanese origin with respect
to ethnicity. Ten tag single nucleotide polymorphisms (SNPs) in the HTR2A gene were genotyped with
TaqMan SNP genotyping. A multivariate logistic regression analysis was applied with adjustments of age andbody mass index (BMI).
Results : There were no significant differences of allelic frequencies of the ten tag SNPs between patient andcontrol groups. In addition, in sub-analyses among the patients with OSAS, we did not detect any associationsof these SNPs with the severity of OSAS (apnea hypopnea index cutoff : 40 events/h) and with the degree ofobesity (BMI cut off : 25 kg/m2).
Conclusions : This study did not prove the hypothesis regarding the association of variations of the serotonin
2A receptor gene (HTR2A) with OSAS. The HTR2A gene variations were less likely to participate in thepathogenesis of OSAS in Japanese. Shinshu Med J 65 : 153―162, 2017
(Received for publication October 27, 2016 ; accepted in revised form December 27, 2016)
Key words : 5-hydroxytryptamine, gene, obstructive sleep apnea syndrome, polymorphism

Ⅰ Introduction

arousal from sleep1）. OSAS is associated with meta­
bolic syndrome, cardiovascular diseases, and neu­

Obstructive sleep apnea syndrome (OSAS) is char­ropsychological sequelae2）. In addition, traffic andacterized by repeated partial or complete collapse ofwork-related accidents are frequently attributed tothe pharynx during sleep, which results in apnea or

OSAS, which leads substantial social and economic

hypopnea, associated with oxygen desaturation andcosts2）. Clarifying the risk factors that confer suscep­tibility to OSAS would contribute not only to the＊Corresponding author : Masayuki Hanaoka
The First Department of Medicine, Shinshu University
School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano
390-8621, Japan
E-mail : masayuki@shinshu-u.ac.jp

No. 3, 2017

identification of diagnostic and prognostic biomark­ers but also to the promotion of therapeutic andpreventive strategies for individuals with a highrisk of OSAS. In addition to the risk factors of age,
153

Ikegawa・Ota・Kobayashi et al.

gender, and body mass index (BMI), recent studies havepresent, it is unclear about the association of SNPs

identified that genetic factors are closely associatedin the HTR2A gene with susceptibility to OSAS in

3）-5）. For example, a family study suggest­

the Japanese population because of insufficient ge­

ed that the risk of OSAS might be higher in rela­

netic data about this issue12）. In order to understandtives of patients with OSAS than in controls3）. In ad­

the associations of the HTR2A gene with OSAS indition, Redline and Tishler reviewed data in relation

the Japanese, we genotyped and analyzed a large

to OSAS and suggested that nearly 40 % of the vari­number of tag SNPs in the HTR2A gene in a case-ance in the apnea hypopnea index (AHI) in patients

control association study with a relatively large sample

with OSAS

4）

with OSAS might be explained by genetic factors .

size of Japanese male patients with OSAS.

Strong evidences suggested that genetic factors wereⅡinteractively associated with craniofacial structure,body fat distribution, and neural control of the upper4）

airway muscles to produce the OSAS phenotype .

A

Patients and Methods

Patients
This study was approved by the Ethics Commit­

The neurotransmitter, 5-hydroxytryptamine (5-tee of Shinshu University (permission number : 298).

HT, or serotonin), works in the central nervous sys­

Written informed consent was obtained from all pa­

tem to regulate various visceral and physiologic

tients and controls prior to their inclusion in thefunctions, including sleep, appetite, pain perception,

study. All procedures performed in the study were

hormone secretion, thermoregulation, and sexual be­in accordance with the ethical standards of the insti­

6）

havior . In addition, several lines of pharmacological,

tutional research committee and with the 1964 Hel­

neurobehavioral, and therapeutic evidences have im­sinki declaration and its later amendments.

plicated serotonin is involved in the pathogenesis of
OSAS

6）
-9）

This study enrolled 145 male Japanese patients. Serotonin controls genioglossus muscle ac­with OSAS. All subjects were unrelated Japanesetivity by binding the serotonin 2A receptor (HTR2A),

individuals with permanent residences in Japan.

which modulates hypoglossal motor output. Contrac­

Of these, 125 patients were consecutive referrals

tion of the genioglossus muscle, which is innervatedto Shinshu University Hospital and Hiro Internalby 5-HT neurons, prevents collapse of the upper

Medicine Clinic from April 2001 to March 2012 ; the7）8）airway. Previous studies in obese rats demonstrat­other 20 patients were long-distance truck drivers

ed that increased expression of HTR2A could effec­

diagnosed with OSAS through an OSAS screening

tively maintain stable upper airways and normal

check-up at Shinshu University from 2006 to 2007.

9）breathing . Experiments in vitro showed that poly­

The diagnosis of OSAS was based on criteria deter­morphisms in the HTR2A gene could influence themined by the American Academy of Sleep Medicine

10）(AASM)21）. These criteria were AHI ≥ 15 events/hlevel of receptor expression .

The human HTR2A gene comprises 3 exons andor AHI ≥ 5 events/h plus a clinical presentation oflocates in the q14-21 region of chromosome 1311）.

OSAS symptoms. The AHI was monitored continu­

Several important single nucleotide polymorphisms

ously during a night of sleep with polysomnography(SNPs) in the HTR2A gene were studied in order to(PSG). The clinical OSAS symptoms were defined as

detect associations with susceptibility to OSAS,a score ≥11 by the degrees of habitual snoring andhowever, diverse results were shown by various

daytime sleepiness on the Epworth Sleepiness Scaleethnic populations regarding the association between(ESS)22）. The patients were excluded when they had

SNPs of the HTR2A gene with susceptibility torenal failure, hypothyroidism, acromegaly, central

12）
-17）

OSAS

. Indeed, racial and ethnic differences in OSAS

sleep apnea, or psychiatric disorders. For sub-analy­

18）
-20）

sis classified by BMI, WHO defines overweight as ahave been evidenced by international studies

in which the emerging data suggested that certain

BMI greater than or equal to 25. The patients were

ethnic groups may be at increased risk for OSAS. At

further classified into obese OSAS (BMI ≥ 25 kg/154

Shinshu Med J

Vol. 65

The SNPs of the serotonin 2A receptor gene and OSAS in Japanesem2 ; n＝51) and non-obese OSAS (BMI <25 kg/m2 ; nwere : rs3803189 (in the 3’-UTR), rs977003, rs9567737,＝94) subgroups. For sub-analyses classified by AHI,

rs9316232, rs2224721, rs2770296, rs731779, rs9567746,

23）

we followed the criteria in previous studies . The

rs2070036, and rs6311 (in the 5’-UTR). The ten SNPs

patients were further classified into severe OSAS

were selected based on the following information(AHI ≥ 40 events/h ; n＝70) and mild or moderatefrom the NCBI dbSNP database : (a) located within

OSAS (AHI <40 events/h ; n＝75) subgroups.

the HTR2A gene ; (b) minor allele frequency over 10 %Control subjects consisted of 133 healthy, unrelated

in Japanese populations ; (c) average heterozygosity

male Japanese through an OSAS screening check-up

of 30 % ; (d) density of at least one SNP per 5 kb ; andat Shinshu University from 2006 to 2007. To ensure(e) availability for validation assays. Furthermore,

the control subjects were free from sleep-related

these ten SNPs could tag another 38 SNPs in thebreathing disorders, they were selected with the

HTR2A genes in a Japanese population by produc­

following criteria : absence of sleep disturbances ; noing a coefficient of determination (r2) >0.8, when evalu­

symptoms related to any disordered breathing during

ated with tagger software from the International

sleep ; AHI <5 events/h ; and oxygen saturation by

HapMap project24） (Table 1).

pulse oximetry (SpO2) >90 % in an overnight PSG.
B

Polysomnography (PSG)

The SNP Genotyping Assay Mix contained for­ward and reverse primers and FAMTM and VICTM

All patients with OSAS and control subjects un­

dye-minor groove binder-labeled probes (Appliedderwent overnight PSG (Alice III ; Chest Ltd ; Tokyo,

Biosystems Inc., Tokyo, Japan). Allelic discrimination

Japan). Polysomnography consisted of a continuousof the ten SNPs was performed according to thepolygraphic recording from multiple surface leads,

manufacturer’s instructions for the TaqMan®SNP

including leads for an electroencephalography (EEG,

Genotyping Assay with an Applied Biosystems 7500

C3-A2, C4-A1, O2-A1, and O3-A2), for a bilateral

Fast Real-time PCR System (Applied Biosystems

electro-oculography, for chin and lower leg electro­

Inc., Foster City, CA, USA). After thermal cycling,

myography, and for electrocardiography (ECG). Re­

genotype data were acquired automatically and ana­

cordings also tracked output from thermistors forlyzed with sequence detection software (SDS v1.3.1,

nasal and oral airflows, thoracic and abdominal im­

Applied Biosystems Inc.).

pedance belts for respiratory effort, a pulse oximeterD

Statistical analysis

for SpO2, a tracheal microphone for snoring, andQuantitative data were expressed as the mean ±sensors for detecting body position during sleep. An

standard deviation (SD). The Mann-Whitney U test

apnea episode was defined as the complete cessation

was used to evaluate significant differences betweenof airflow for at least 10 seconds (s). Hypopnea wascases and controls in age, BMI, and AHI. Frequen­

defined as at least a 50 % reduction in airflow for atcies of genotypes and alleles were expressed in deci­

least 10s, accompanied by a reduction in SpO2 of atmals. The Hardy-Weinberg equilibrium (HWE) for

least 4 %. AHI was the key indicator for OSAS diag­each SNP was confirmed with the Chi-square test.

nosis ; AHI was defined as the number of apnea or

Significant differences in allele frequencies between

hypopnea events per hour during sleep time, based

two groups were evaluated with the Chi-square test

on results from the overnight PSG.

(2×2 contingency table). The effects of ancestral al­Cleles on inheritance of OSAS were evaluated with

Genotyping
DNA was extracted from whole blood with a Quick­multivariate logistic regression analyses, assuming a

Gene 800 (Fuji Film, Tokyo, Japan). Genomic DNA

dominant mode and a recessive mode. The values of

was prepared at 10-15 ng/µL for the TaqMan SNP

pair-wise linkage disequilibrium (LD) of the ten

genotyping assay. We genotyped ten SNPs that spanned

SNPs were measured with Haploview software25）.

the region between the 3ʼ-untranslated region (UTR)

Results are expressed with odds ratios (OR) with 95

and the 5’-UTR of the HTR2A gene. These SNPs

% confidence interval (CI) values, after adjusting for

No. 3, 2017155

Ikegawa・Ota・Kobayashi et al.
Table 1

Tagging efficiency of the ten SNPs of HTR2A for a Japanese population

Test SNPs
rs3803189
rs977003
rs9567737
rs9316232
rs2224721
rs2770296
rs731779
rs9567746
rs2070036
rs6311Alleles captured

Number of SNPs

rs977003, rs1923882, rs7322347, rs3125
rs3803189, rs1923882, rs977003, rs7322347, rs3125
rs6561333, rs6561333
rs1923888, rs1923888, rs2296972, rs9567739, rs655888, rs3742279, rs1745837,
rs622337, rs655854
rs2224721
rs2770297, rs2770298, rs1928040
rs9567746, rs2770293, rs582854, rs9567746, rs9316235, rs9526245
rs731779, rs2770293, rs582854, rs9316235, rs9526245
rs2070036
rs6311, 6313
Total

4
5
2
9
1
3
6
5
1
2
38

Evaluated at coefficient of determination (r2) >0.8 by tagger software through International HapMap project(http : //hapmap.ncbi.nlm.nih.gov/)

Table 2

Characteristics of subjects with OSAS and controls
Patients with OSAS

Controls

Number of subjects
Age (years)

145
56.9±13.6＊43.3±12.7

BMI (kg/m2)

27.2±5.0＊42.2±19.2＊

AHI (events/h)

13323.4±3.2
3.5±3.7

All subjects were male. Data are expressed as mean±SD.
＊ p <0.001 versus controls by Mann-Whitney U test.

age and BMI26）. P values <0.05 indicated statisticaladjusting for age and BMI, the multivariate logistic

significance. Corrected P values (Pc) were calculated

regression analysis did not show any effects of theby multiplying the number of alleles in a given locus.

ancestral SNP alleles on OSAS inheritance, assuming

Ⅲ
A

Results

Characteristics of subjects with OSAS and con-either the dominant mode or the recessive mode
(Table 3). Moreover, there were no significant differ­ences in frequencies of the observed haplotypes be­trols

tween the controls and OSAS patients.

The final analyses were based on genetic data

C

Associations of the ten tag SNPs with obesityfrom 145 male patients with OSAS and 133 male

and with severity of OSAS

controls. The average AHI was significantly higher

In the sub-analysis concerning obese and non-in the OSAS group than in the control group (42.2±

obese OSAS subgroups classified by BMI (cut-off

19.2 vs. 3.5±3.7 events/h, P <0.001, Table 2). The av­

value : 25 kg/m2), significant associations were noterage age and BMI were significantly greater in thedetected regarding the ten tag SNPs of the HTR2A

patients with OSAS than in the controls (Table 2).

with obese-OSAS (Table 4).

Associations of the ten tag SNPs with OSAS

In the sub-analysis concerning severe and mild or

All the ten SNPs were in HWE for both the OSAS

moderate OSAS subgroups classified by AHI (cut-off

and control groups. There were no significant differ­value : 40 events/h), rs2770296 and rs731779 seemedences in the allelic frequencies of the ten tag SNPs

to be associated with severe OSAS (P＝0.040, 0.047,

between the two groups (Table 3). In addition, after

respectively, Table 5) ; however, such significant

B

156

Shinshu Med J

Vol. 65

The SNPs of the serotonin 2A receptor gene and OSAS in Japanese
Table 3 A allele frequencies and genotype distributions of tag SNPs of HTR2A gene in patients with
OSAS (N＝145) and controls (N＝133)
dbSNPsLocation

Alleles(1/2)＊

rs3803189
3ʼ-UTR

Allele 1
Frequency

Genotype distributions
11

†

P

＊

P‡

＊

＊

12

22

11/12＋22

P‡
11＋12/22

OSAS

Controls

OSAS

Controls

OSAS

Controls

OR (95 % CI)

OR (95 % CI)

0.71

0.593

0.564

0.352

0.383

0.055

0.053

0.741

0.83

0.559

0.534

0.379

0.413

0.062

0.053

0.62

0.621

0.99

0.361

0.379

0.519

0.483

0.120

0.138

A/G

0.548

0.485

0.14

0.290

0.241

0.517

0.408

0.193

0.271

rs2224721
Intron

G/T

0.603

0.526

0.07

0.386

0.286

0.435

0.481

0.179

0.233

rs2770296
Intron

T/C

0.700

0.673

0.49

0.490

0.443

0.421

0.459

0.089

0.098

0.28
0.47
(0.12-1.88)
0.51
1.23
(0.67-2.25)
0.76
0.90
(0.45-1.80)
0.66
1.14
(0.63-2.07)
0.57
0.84
(0.46-1.54)
0.40
1.29
(0.71-2.33)

rs731779
Intron

A/C

0.766

0.759

0.87

0.600

0.579

0.331

0.361

0.069

0.06

rs9567746
Intron

A/G

0.741

0.726

0.67

0.565

0.519

0.352

0.413

0.083

0.068

rs2070036
Intron

T/G

0.686

0.68

0.88

0.455

0.466

0.462

0.421

0.083

0.105

rs6311
5ʼ-UTR

C/T

0.5

0.534

0.43

0.241

0.278

0.518

0.511

0.241

0.211

0.45
0.79
(0.44-1.44)
0.91
1.07
(0.31-3.69)
0.44
0.76
(0.38-1.52)
0.71
1.25
(0.40-3.91)
0.84
0.91
(0.36-2.31)
0.99
1.06
(0.37-2.76)
0.43
0.67
(0.25-1.80)
0.76
1.21
(0.36-4.07)
0.19
1.52
(0.82-2.83)
0.82
0.93
(0.47-1.82)

OSAS

Controls

T/G

0.769

0.756

rs977003
IntronA/C

0.748

rs9567737
IntronT/C

rs9316232
Intron0.81
1.14(0.40-3.25)
0.33
0.74
(0.41-1.35)
0.15
1.72
(0.81-3.64)
0.72
1.13
(0.57-2.26)

Allelic frequencies and genotypic distributions are expressed as decimals.
＊1/2 indicates ancestral allele/derived allele according to the NCBI dbSNP database.
† P values obtained by Chi-square test (2×2 contingency table).
‡ P values of the dominant mode (11/12＋22) and the recessive mode (11＋12/22) as well as their corresponding OR (95% CI) values are obtained by multivariate logistic regression analysis after adjustment for age and body mass index(http : //statpages.org/logistic.html).

associ­ations vanished after correcting with the P values

bility to OSAS in a Japanese population. In addition,(Pc＝0.40, 0.47, respectively, Table 5). No other signifi­

no genetic associations were detected between thesecant associations were detected regarding the ten

SNPs and the severity of OSAS (AHI ≥40 events/h)

tag SNPs of the HTR2A with severe OSAS (Table 5).

or overweight in OSAS (BMI ≥25 kg/m2). The reli­

Ⅳ Discussion

ability of these results were convinced by the facts
that all patients and controls were strictly diagnosed

In the present study, we densely genotyped ten

with standard PSG examinations and that multivari­

SNPs of the HTR2A gene (rs3803189 in the 3ʼ-UTR,

ate logistic regression analyses were adjusted for

rs977003, rs9567737, rs9316232, rs2224721, rs2770296,

significant differences in age and BMI.

rs731779, rs9567746, rs2070036, and rs6311 in the 5’-

The biological pathways underlying OSAS are me­

UTR), those that could tag another 38 SNPs along

diated by genes involved in serotonergic receptor

the HTR2A gene (Table 1), in 145 male patients withtransmission ; thus, these genes attract interest as

OSAS and 133 male controls. The results demon­candidate genes that might confer susceptibility

strated that there was no association between theto OSAS. Serotonin plays important roles in sleep-

ten tag SNPs of the HTR2A gene and the suscepti­

wake behavior and appetite regulation ; it is also in­

No. 3, 2017157

Ikegawa・Ota・Kobayashi et al.
Table 4 A allelic frequencies of the ten tag SNPs of the HTR2A gene between sub­groups classified by BMI (cuto-ff : 25 kg/m2) among the patients with OSAS

dbSNPsAlleles(1/2)＊

rs3803189
rs977003
rs9567737
rs9316232
rs2224721
rs2770296
rs731779
rs9567746
rs2070036
rs6311

T/G
A/C
T/C
A/G
G/T
T/C
A/C
A/G
T/G
C/T

Allele 1 Frequency
Obese OSAS
(N＝94)

Non-obese OSAS
(N＝51)

P†

Pc‡

0.777
0.761
0.628
0.580
0.601
0.691
0.724
0.745
0.686
0.521

0.755
0.725
0.608
0.490
0.608
0.716
0.833
0.735
0.686
0.461

0.676
0.510
0.740
0.143
0.910
0.668
0.062
0.862
0.999
0.325

6.756
5.102
7.398
1.432
9.103
6.677
0.616
8.616
9.985
3.252

Allelic frequencies and genotypic distributions are expressed as decimals.
＊1/2 indicates ancestral allele/derived allele according to the NCBI dbSNP database.
† P values obtained by Chi-square test (2×2 contingency table).
‡ Corrected P value calculated by multiplying the number of alleles in a given locus.

Table 5 A allelic frequencies of the tag SNPs of the HTR2A gene between subgroups
classified by AHI (cut-off : 40 events/hour) among the patients with OSAS

dbSNPsAlleles(1/2)＊

rs3803189
rs977003
rs9567737
rs9316232
rs2224721
rs2770296
rs731779
rs9567746
rs2070036
rs6311

T/G
A/C
T/C
A/G
G/T
T/C
A/C
A/G
T/G
C/T

Allele 1 Frequency
Severe OSAS
(N＝70)

Mild & Moderate OSAS
(N＝75)

P†

Pc‡

0.800
0.753
0.647
0.567
0.640
0.753
0.813
0.787
0.713
0.513

0.736
0.743
0.593
0.529
0.564
0.643
0.714
0.693
0.657
0.486

0.194
0.837
0.345
0.515
0.188
0.040
0.047
0.068
0.303
0.638

1.943
8.373
3.453
5.148
1.878
0.402
0.467
0.683
3.028
6.383

Allelic frequencies and genotypic distributions are expressed as decimals.
＊1/2 indicates ancestral allele/derived allele according to the NCBI dbSNP database.
† P values obtained by Chi-square test (2×2 contingency table).
‡ Corrected P calculated by multiplying the number of alleles in a given locus.

volved in upper airway dilator muscle activity through8）
9）gene with OSAS was detected in an African-Ameri­. In par­can population, however, which was greatly attenu­

ticular, the serotonin 2A receptor was found to be theated after adjusting for BMI (the P value was atten­

predominant excitatory serotonin receptor subtype inuated from 0.0000523 to 0.0126 after adjustment)29）.

hypoglossal motor neurons27）; indeed, administration

The rs9526240 SNP is located in the intron of theof a serotonin 2A receptor agonist improved upper

HTR2A gene, and its function is currently unknown.

its modulation of hypoglossal motor output

28）

airway stability in an animal model . A significant

This association attenuation suggested that HTR2A

association of the rs9526240 SNP in the HTR2A

may influence OSAS through pleiotropic pathways

158

Shinshu Med J

Vol. 65

The SNPs of the serotonin 2A receptor gene and OSAS in Japanesethat influence both airway stability and obesity.

susceptibility to OSAS in the present Japanese pa­

Moreover, the positive association of the rs6311

tients.

(-1438G/A) in the HTR2A gene with OSAS were15）

The HTR2A receptor is located primarily in the

16）
17）

neurocortex, caudate nucleus, nucleus accumbens,

populations, yet those significances were uncertain

olfactory tubercle, and the hippocampus, in the cen­because of the absence of adjustment for BMI

tral nervous system, while being marginally distrib­

reported in Chinese

13）
14）

, Turkish , and Brazilian

13）
-17）

. It is well known that obesity

uted in the hypoglossal motor nucleus in the periph­

is the most common characteristic of adults witheral nervous system35）. Thus, the HTR2A receptor

OSAS. There are probably both shared and un­

mainly targets biological molecules in serotonergic-

shared genetic factors that underlie the susceptibili­rich areas of the central nervous system involved

4）ties to OSAS and obesity . Thus, the association be­in neuronal excitation, behavioral effects, learning,

tween the HTR2A polymorphisms and OSAS might

and anxiety, but has a minor function in excitatory

be partially explained by a common causal pathway

transmission at the serotonergic-poor area of theinvolving both AHI and BMI pathogeneses30）. Never­hypoglossal motor nucleus in the peripheral nervous

theless, it is absolutely necessary to adjust BMI insystem35）. We interpreted this to mean that the scantstatistical analyses to minimize the possibility ofdensity and minor function of the HTR2A receptor

false-positive or conflicting results in genetic associ­in the hypoglossal motor nucleus might partly ex­ation studies on OSAS.

plain the negative results found in the present study.

in these studies

The rs6311 and rs6313 SNPs of the HTR2A gene

At present, the relations of the hypoglossal nerve

were the most attractive candidates, based on previ­and serotonin receptor have been demonstrated inous studies on genetic variants associated withanimals by animal experiments ; however, the distri­OSAS

12-17）. One meta-analysis revealed that rs6311

bution of the serotonin receptor in the medulla ob­

was significantly associated with susceptibility

longata (where the nucleus of the hypoglossal nerve

to OSAS, but not rs631331）. The rs6311 is a polymor­

exists) has not yet been evidenced in humans. Addi­

phism in the promoter of HTR2A, with functionaltional mechanisms other than the HTR2A receptor

significance in serotonergic neurotransmission. A

might be involved in OSAS pathophysiology as well.

structure-function equation model suggested that

For example, craniofacial morphologic abnormalities

this promoter polymorphism might affect both tran­

are more severe in Asian populations than in Cauca­

scription factor binding and promoter methylation,

sians with the same range of BMI or the degree ofand thus, it might alter the rate of HTR2A tran­obesity36）37）. Endothelin-receptor-A38） and transform­

scription in a methylation-dependent manner32）. The

ing growth factor-beta 239） are concerned with cra­

rs6311 is in complete linkage disequilibrium (r2＝1.0)

niofacial morphologic abnormalities, and it is sug­with rs6313 in the Japanese population on the genet­gested that these genes be analyzed regarding theic dataset of HapMap. Regarding the rs6313, it is a

genetic background of OSAS pathophysiology.

synonymous variant, with no resulting change in the

The obvious limitation of the present study wasamino acid sequence, though it may affect thethat the age and BMI of the patient group did notmRNA stability, quantity, and/or translation, whichmatch those of the control group, although adjust­33）

could affect protein expression . Additionally, thements were applied to the statistical analyses for

rs6313 SNP may also affect methylation of thetheoretical correlations. We did not restrict age or

34）

HTR2A promoter . Pollesskaya and Sokolov ob­

BMI in the process of selecting subjects because we

served that the T allele of rs6313 was associatedaimed to include a relatively large sample size towith an elevated number of HTR2A receptors inachieve adequate statistical power. In practice, it is33）

the central nervous system . Although true, we diddifficult to recruit large sample sizes of an OSAS

not find any associations of these two SNPs with thegroup and control group matched in the age andNo. 3, 2017159

Ikegawa・Ota・Kobayashi et al.

BMI, these being the two major risk factors for de­

Respiratory Failure Research Group in the Ministryveloping OSAS.

of Health, Labour and Welfare, Japan. The funding
source had no roles in study design ; in the collection,

Ⅴ Conclusion

analysis and interpretation of data ; in the writing ofThis study showed that ten SNPs in the HTR2A
g e n e ( r s 3803189 , r s 977003 , r s 9567737 , r s 9316232 ,the report ; and in the decision to submit the article
for publication.

rs2224721, rs2770296, rs731779, rs9567746, rs2070036,

Ⅶand rs6311) and their tagged 38 SNPs were not asso­
ciated with susceptibility to OSAS in a Japanese

Conflict of Interest Disclosure Statement

No potential conflicts of interest were disclosed.

population. Further studies on different genes that

Ⅷ

might be associated with OSAS, such as genes in­volved with craniofacial morphology38）, transforming
39）

Acknowledgement

We are very grateful to Dr. Yoshimichi Komatsu

growth factor-beta 2 , endothelin-receptor-A, or a

(Suwatoyoda Clinic, Suwa) and Dr. Keisaku Fujimoto

whole genome scan, might elucidate the role of ge­(Shinshu University School of Medicine, Matsumoto)netics in the pathogenesis of OSAS in the Japanese

for their contributions to the collection of samples

population.

for this study.
Ⅵ Funding SourceThis research was financially supported by the
References
1） Douglas NJ, Polo O : Pathogenesis of obstructive sleep apnoea/hypopnoea syndrome. Lancet 344 : 653-655, 1994
2） Redline S, Young T : Epidemiology and natural history of obstructive sleep apnea. Ear Nose Throat J 72 : 20-21, 2426, 1993
3） Redline S, Tishler PV, Tosteson TD, Williamson J, Kump K, Browner I, Ferrette V, Krejci P : The familial aggrega­tion of obstructive sleep apnea. Am J Respir Crit Care Med 151 : 682-687, 1995
4） Redline S, Tishler PV : The genetics of sleep apnea. Sleep Med Rev 4 : 583-602, 2003
5） Casale M, Pappacena M, Rinaldi V, Bressi F, Baptista P, Salvinelli F : Obstructive sleep apnea syndrome : from phe­notype to genetic basis. Curr Genomics 10 : 119-126, 20096） Richter DW, Manzke T, Wilken B, Ponimaskin E : Serotonin receptors : guardians of stable breathing. Trends Mol
Med 9 : 542-548, 2003
7） Sood S : Role of endogenous serotonin in modulating genioglossus muscle activity in awake and sleeping rats. Am J
Respir Crit Care Med 172 : 1338-1347, 2005
8） Kraiczi H : Effect of serotonin uptake inhibition on breathing during sleep and daytime symptoms in obstructive
sleep apnea. Sleep 22 : 61-67, 1999
9） Sood S, Liu X, Liu H, Horner RL : Genioglossus muscle activity and serotonergic modulation of hypoglossal motor
output in obese Zuker rats. J Appl Physiol 102 : 2240-2250, 200710） Myers RL, Airey DC, Manier DH, Shelton RC, Sanders-Bush E : Polymorphisms in the regulatory region of the hu­
man serotonin 5-HT2A receptor gene (HTR2A) influence gene expression. Biol Psychiatry 61 : 167-173, 2007
11） Chen K, Yang W, Grimsby J, Shih JC : The human 5-HT2 receptor is encoded by a multiple intron-exon gene.
Molec Brain Res 14 : 20-26, 1992
12） Sakai K, Takada T, Nakayama H, Kubota Y, Nakamata M, Satoh M, Suzuki E, Akazawa K, Gejyo F : Serotonin-2Aand 2C receptor gene polymorphisms in Japanese patients with obstructive sleep apnea. Intern Med 44 : 928-933,
2005
160

Shinshu Med J

Vol. 65

The SNPs of the serotonin 2A receptor gene and OSAS in Japanese
13） Chen H, Hu K, Zhu J, Xianyu Y, Cao X, Kang J, He J, Zhao P, Mei Y : Polymorphisms of the 5-hydroxytryptamine
2A/2C receptor genes and 5-hydroxytryptamine transporter gene in Chinese patients with OSAHS. Sleep Breath
17 : 1241-1248, 201314） Yin G, Ye J, Han D, Zhang Y, Zeng W, Liang C : Association of the 5-HT2A receptor gene polymorphisms with ob­
structive sleep apnea syndrome in Chinese Han population. Acta Otolaryngol 132 : 203-209, 201215） Bayazit YA, Yilmaz M, Ciftci T, Erdal E, Kokturk O, Gokdogan T, Kemaloglu YK, Inal E : Association of the -1438G/
A polymorphism of the 5-HT2A receptor gene with obstructive sleep apnea syndrome. ORL J Otorhinolaryngol
Relat Spec 68 : 123-128, 200616） de Carvalho TB, Suman M, Molina FD, Piatto VB, Maniglia JV : Relationship of obstructive sleep apnea syndrome
with the 5-HT2A receptor gene in Brazilian patients. Sleep Breath 17 : 57-62, 201217） Piatto VB, Carvalho TB, De Marchi NS, Piatto VB, Maniglia JV : Polymorphisms in the 5-HTR2A gene related toobstructive sleep apnea syndrome. Braz J Otorhinolaryngol 77 : 348-355, 201118） Redline S, Tishler PV, Hans MG, Tosteson TD, Strohl KP, Spry K : Racial differences in sleep-disordered breathing
in African-Americans and Caucasians. Am J Respir Care Med 155 : 186-192, 1977
19） Baldwin M, Kolbe J, Troy K, Belcher J, Gibbs H, Frankel A, Eaton T, Christmas T, Veale A : Racial differences in se­verity of sleep apnea between Maori, Pacific Islands and Europeans. Am J Respir Crit Care Med 153 : A357 (abstr),
199620） Ng TP, Seow A, Tan WC : Prevelence of snoring and sleep breathing-related disorders in Chinese, Malay and Indi­an adults in Singapore. Eur Respir J 12 : 198-203, 199821） The AASM manual for the scoring of sleep and associated events : Rules, terminology, and technical specification.
American Academy of Sleep Medicine, Westchester, 200722） Johns MW : A new method for measuring daytime sleepiness : the Epworth sleepiness scale. Sleep 14 : 540-545, 1991
23） Young T, Palta M, Dempsey J, Skatrud J, Weber S, Badr S : The occurrence of sleep-disordered breathing among
middle-aged adults. N Engl J Med 328 : 1230-1235, 199324） International HapMap project (http : //hapmap.ncbi.nlm.nih.gov/), Accessed 26 June 201625） Barret JC, Fry B Maller J, Daly MJ : Haploview : analysis and visualization of LD and haplotype maps. Bioinfomatics
21 : 263-265, 200526） Pezzullo JC (2015) : Logistic Regression. http : //statpages.org/logistic.html Accessed 26 June 201627） Fenik P, Veasey SC : Pharmacological characterization of serotonergic receptor activity in the hypoglossal nucleus.
Am J Respir Crit Care Med 167 : 563-569, 2003
28） Ogasa T, Ray AD, Michlin CP, Farkas GA, Grant BJ, Magalang UJ : Systemic administration of serotonin 2A/2C ag­
onist improves upper airway stability in Zucker rats. Am J Respir Crit Care Med 170 : 804-810, 2004
29） Larkin E, Patel S, Goodloe R, Li Y, Zhu X, Gray-McGuire C, Adams MD, Redline S : A candidate gene study of ob­
structive sleep apnea in European Americans and African Americans. Am J Respir Crit Care Med 182 : 947-953,
201030） Palmer LJ, Buxbaum SG, Larkin E, Patel SR, Elston RC, Tishler PV, Redline S : A whole-genome scan for obstruc­tive sleep apnea and obesity. Am J Hum Genet 72 : 340-350, 2002
31） Zhao Y, Tao L, Nie P, Lu X, Xu X, Chen J, Zhu M : Association between 5-HT2A receptor polymorphisms and riskof obstructive sleep apnea and hypopnea syndrome : A systematic review and meta-analysis. Gene 530 : 287-294,
201332） Falkenberg VR, Gurbaxani BM, Unger ER, Rajeevan MS : Functional genomics of serotonin receptor 2A (HTR2A) :Interaction of polymorphism, methylation, expression and disease association. Neuromolecular Med 13 : 66-76, 201133） Polesskaya OO, Sokolov BP : Differential expression of the 'C' and 'T' alleles of the 5-HT2A receptor gene in thetemporal cortex of normal individuals and schizophrenics. J Neurosci Res 67 : 812-822, 2002
No. 3, 2017161

Ikegawa・Ota・Kobayashi et al.
34） Polesskaya OO, Aston C, Sokolov BP : Allele C-specific methylation of the 5-HT2A receptor gene : Evidence for cor­
relation with its expression and expression of DNA methylase DNMT1. J Neurosci Res 83 : 362-373, 200635） Hoyer D, Hannon JP, Martin GR : Molecular, pharmacological and functional diversity of 5-HT receptors. Pharmacol
Biochem Behav 71 : 533-554, 200236） Li KK, Kushida C, Powell NB, Riley RW, Guilleminault C : Obstructive sleep apnea syndrome : a comparison between
Far-East Asian and white men. Laryngoscope 110 : 1689-1693, 2000
37） Liu Y, Lowe AA, Zeng X, Fu M, Fleetham JA : Cephalometric comparisons between Chinese and Caucasian patientswith obstructive sleep apnea. Am J Orthod Dentofacial Orthop 117 : 479-485, 200038） Kurihara Y, Kurihara H, Suzuki H, Kodama T, Maemura K, Nagai R, Oda H, Kuwaki T, Cao WH, Kamada N : Ele­vated blood pressure and craniofacial abnormalities in mice deficient in endotheline-1. Nature 368 : 703-710, 1994
39） Sanford LP, Ormsby I, Gittenberger-de Groot AC, Sariola H, Friedman R, Boivin GP, Cardell EL, Doetschman T : TGF
Beta-2 knockout mice have multiple development defects that are non-overlapping with other TGF Beta knockout
phenotypes. Development 124 : 2659-2670, 1997（2016. 10. 27 received；2016. 12. 27 accepted）

162

Shinshu Med J

Vol. 65