Effects of interleukin (IL)-6 gene polymorphisms on recurrent aphthous stomatitis
Transcript of Effects of interleukin (IL)-6 gene polymorphisms on recurrent aphthous stomatitis
ORIGINAL PAPER
Effects of interleukin (IL)-6 gene polymorphisms on recurrentaphthous stomatitis
Nevin Karakus • Serbulent Yigit • Aydin Rustemoglu •
Goknur Kalkan • Nihan Bozkurt
Received: 31 May 2013 / Revised: 6 August 2013 / Accepted: 9 August 2013 / Published online: 28 August 2013
� Springer-Verlag Berlin Heidelberg 2013
Abstract Recurrent aphthous stomatitis (RAS) is a
common disease with oral ulceration in which cytokines
are thought to play an important role. High levels of
interleukin (IL)-6, a pro-inflammatory cytokine have been
detected in the circulation of ulcer tissue. The purpose of
the present study was to investigate if the IL-6 gene
polymorphisms are associated with RAS or clinical char-
acteristics of RAS in a cohort of Turkish population. 184
RAS patients and 150 healthy controls were included in the
study. The genotypes of IL-6 gene -572G[C and
-174G[C polymorphisms were determined using poly-
merase chain reaction based restriction fragment length
polymorphism analysis. The genotype frequencies of
-572G[C polymorphism showed statistically significant
differences between RAS patients and controls (p = 0.01).
Frequencies of GG ? GC genotypes and G allele of
-572G[C polymorphism were found higher in RAS
patients (p = 0.0001, OR 10.8, 95 % CI 2.79–70.5;
p = 0.0008, OR 2.06, 95 % CI 1.35–3.17, respectively).
The genotype frequencies of -174G[C polymorphism
also showed statistically significant differences between
RAS patients and controls (p \ 0.0001). Frequencies of
GG genotype and G allele of -174G[C polymorphism
were found higher in RAS patients (p \ 0.0001, OR 4.87,
95 % CI 3.06–7.85; p \ 0.0001, OR 3.82, 95 % CI
2.64–5.59, respectively). GG–GG combined genotype and
G–G haplotype of -174G[C to -572G[C loci were also
significantly higher in RAS patients (p \ 0.0001 and
p = 1.5 9 10-8, respectively). After stratifying clinical
and demographical characteristics of RAS patients
according to IL-6 gene polymorphisms, an association was
observed between family history of RAS and -174G[C
polymorphism (p = 0.011). Susceptibility effects of both
IL-6 gene -572G[C and -174G[C polymorphisms for
RAS were observed. Further studies are necessary to prove
the association of IL-6 gene polymorphisms with RAS.
Keywords Recurrent aphthous stomatitis (RAS) �Interleukin (IL)-6 � Genetic susceptibility � Cytokine �Oral ulceration
Introduction
Recurrent aphthous stomatitis (RAS) is a common disease
with recurrent episodes of oral ulceration [17]. It has been
estimated that approximately 25 % of the general popula-
tion, regardless of age, race and geographic region, might
suffer from RAS at some time in their lives [20]. Family
history of RAS was identified in more than 40 % of RAS
patients [15]. There may be possible factors associated with
RAS such as local trauma, stress, trait anger, anxiety,
drugs, hormonal changes, vitamin and trace element defi-
ciencies, socioeconomics, allergies, mechanical injury and
preservatives [15, 30]. Local and systemic conditions and
genetic, immunological and microbial factors may also
play a role in the pathogenesis of RAS [15].
Immune dysfunction occurs during the ulcerative stage
of RAS [17]. In the earliest stages, focal epithelial cell
degeneration occur [23]. A lympho-monocytic infiltrate
develops adjacent to the damaged cells. It is supposed that
N. Karakus (&) � S. Yigit � A. Rustemoglu � N. Bozkurt
Department of Medical Biology, Faculty of Medicine,
Gaziosmanpasa University, 60100 Tokat, Turkey
e-mail: [email protected]
G. Kalkan
Department of Dermatology, Faculty of Medicine,
Gaziosmanpasa University, Tokat, Turkey
123
Arch Dermatol Res (2014) 306:173–180
DOI 10.1007/s00403-013-1406-x
RAS is formed by a local damage in an individual genet-
ically predisposed to an abnormal cytokine cascade.
Although extreme serum levels of interleukin (IL)-6, a pro-
inflammatory cytokine that regulates a wide range of
activities, was not observed in RAS patients [16], high
levels of IL-6 have been detected in the circulation of ulcer
tissue [24, 29]. IL-6 is produced by a variety of cells
including macrophages, B cells, T cells, and fibroblasts and
plays important roles in regulating hematopoiesis, immune
cell activation, and inflammation [11].
IL-6 gene is located on chromosome 7p21 [5]. IL-6
expression is almost regulated at the transcriptional level
by regulatory elements in the promoter region of the gene
[14]. A number of polymorphisms have been detected in
this region of IL-6. The -174G[C and -572G[C poly-
morphisms in the promoter region are important regulators
of transcription [12]. The IL-6 gene -174G[C polymor-
phism is located immediately upstream of a multiresponse
element at positions -173 to -151 [8]. The IL-6
-572G[C polymorphism is located near a potential glu-
cocorticoid receptor element at position -557 to -552 [6].
IL-6 production is modulated by these polymorphisms and
has been associated with susceptibility to many several
pathological conditions [2, 4, 7, 8, 22, 25]. There have been
only a few studies dealing with the relationship between
IL-6 promoter polymorphisms and RAS, and their results
have been conflicting. While Bazrafshani et al. [3] reported
a positive association between carriage of the IL-6 -174
GG genotype and RAS, others could not confirm this
observation [10].
The purpose of the present study was to investigate if the
IL-6 gene -174G[C and -572G[C polymorphisms and
combined genotypes and haplotypes are associated with
RAS or clinical characteristics of RAS in a cohort of
Turkish population.
Materials and methods
Subjects
The study group consisted of 184 unrelated patients with
RAS [66 males and 118 females; mean age 35.96 ± 11.645
standard deviation (SD) years], and 150 (62 male and 88
female; mean age 37.28 ± 13.091 SD years) unrelated
healthy controls. RAS patients were recruited consecu-
tively and prospectively from those whom were treated and
followed up in the Dermatology Department of Gazios-
manpasa University Research Hospital, Tokat, Turkey
between January 2012 and December 2012. The diagnosis
of RAS was performed by a dermatologist according to the
history and physical examination based on accepted clini-
cal criteria [21]. RAS was divided into three clinical
features: major (larger than 1 cm and deeper than minor
RAS and healing within 10–30 days), minor (less than
1 cm in diameter and healing within 4–14 days) and her-
petiform aphthae (grouped aphthae, 1–2 mm in size and
painful). Only patients with major and minor RAS were
included in this study. All RAS patients recruited had at
least three recurrences of oral ulcers per year. None of the
patients suffered from Behcet’s Disease, inflammatory
bowel disease, or any systemic disease, and none received
any systemic treatment that could influence the results of
tests performed in this study. The study groups were also
non-smokers and non-alcohol users. Patients were other-
wise healthy and had active aphthous lesions during the
study. The following routine laboratory tests (complete
blood count, liver enzymes, fasting glucose, ferritine,
vitamin B12 and folic acid) were performed. The patients
with abnormal laboratory findings were excluded. The
control group was comprised of patients with no history of
RAS or systemic diseases, who were admitted to derma-
tology clinic with other reasons; for nevus examination,
treatment of warts, tinea pedis or for contact dermatitis.
Exclusion criteria for controls included the presence of any
other significant local or systemic diseases; Behcet’s dis-
ease, coeliac disease and other gastrointestinal symptoms
and/or diseases. All participants, patients and healthy
controls, were of Turkish origin, from the inner Central
Black Sea region of Turkey. The healthy controls were
matched for age and gender with RAS patients (Table 1).
The study protocol was approved by the Local Ethics
Committee of Gaziosmanpasa University, Faculty of
Medicine and written informed consent was obtained from
the study participants or from the parents when subjects
were \18 years.
Genotyping
Genomic DNA was extracted from whole venous blood
samples using a commercial DNA isolation kit (Sigma-
Aldrich, Taufkirchen, Germany). The IL-6 gene -572G[C
(rs1800796) and -174G[C (rs1800795) polymorphisms
Table 1 The demographical characteristics of patients with RAS and
healthy controls
RAS patients
n = 184
Healthy controls
n = 150
p value
Gender 0.311
Female 118 (64.1) 88 (58.7)
Male 66 (35.9) 62 (41.3)
Age (years) 35.98 ± 11.871 37.28 ± 13.091 0.344
Data were analyzed by analysis of variance and v2 test. Mean plus
standard deviation values are presented for age
174 Arch Dermatol Res (2014) 306:173–180
123
were analyzed by polymerase chain reaction (PCR) based
restriction fragment length polymorphism (RFLP) analysis.
The PCR amplifications were carried out in a total volume
of 25 ll reaction containing 100 ng of genomic DNA,
2.5 ll of 109 PCR buffer, 200 lM dNTP, 10 pM each
primers, and one unit of Taq DNA polymerase. The IL-6
-174G[C polymorphism was analyzed as previously
described by Tseng et al. [27] using forward (f) 50-TTG
TCA AGA CAT GCC AAA GTG CGG AG-30 and reverse
(r) 50-GTG CAA TGT GAC GTC CCT TAG CAT-30
primers. The amplification conditions consisted of an initial
melting step of 5 min at 94 �C; followed by 40 cycles of
30 s at 94 �C, 30 s at 56 �C and 1 min at 72 �C. After
amplification, the 156 bp PCR product was digested with
FastDigest BseL-I restriction endonuclease (Fermentas) at
37 �C for 30 min and analyzed on a 3 % agarose gel
stained with ethidium bromide. Two fragments (139 and
17 bp) for G allele and three fragments (117, 22 and 17 bp)
for C allele were observed. For IL-6 -572G[C polymor-
phism, the sequences of PCR primers were (f) 50-CAG
CAG CCA ACC TCC TCT AA-30 and (r) 50-CCA AGC
CTG GGA TTA TGA AG-30. The cycling conditions for
-572G[C polymorphism were 40 cycles of 30 s at 95 �C,
30 s at 62 �C and 1 min at 72 �C. 224 bp PCR product was
digested with FastDigest BsrB-I restriction endonuclease
(Fermentas) at 37 �C for 30 min and analyzed on a 3 %
agarose gel stained with ethidium bromide. While two
fragments (150 and 74 bp) were observed for G allele, only
one fragment (224 bp) was observed for C allele. Second
PCR was performed to confirm samples whose results were
not clear.
Statistical analysis
Statistical analysis was performed using the Statistical
Package for the Social Sciences (SPSS 20), Arlequin
software version 3.1.1 and OpenEpi Info software
package version 3.01 (www.openepi.com). Results were
given as mean ± standard deviation. Allelic frequencies,
haplotypes, haplotype frequencies, Hardy–Weinberg
equilibrium (HWE), LD between SNP pairs were per-
formed by Arlequin software. The relationships between
-174G[C and -572G[C polymorphisms and the clin-
ical and demographical characteristics of patients and
clinical characteristics of oral ulcers of patients were
analyzed by using v2 test or analysis of variance
(ANOVA) statistics. v2 test and Fisher’s exact test were
used to compare categorical variables appropriately,
and odds ratio (OR) and 95 % confidence interval (CI)
were used for the assessment of risk factors. All p values
were two-tailed and p values \0.05 were considered as
significant.
Results
The baseline clinical and demographical characteristics
of RAS patients are presented in Table 2. Gender, age,
age at disease onset, family history, papulopustule,
systemic involvement and colchicine use of RAS
patients were analyzed. No patient was found with
erythema nodosum, genital ulcers and pathergy positiv-
ity. Although no association was not found between
clinical and demographical characteristics of RAS
patients and IL-6 gene -572G[C polymorphism, an
association was observed between family history of RAS
and IL-6 gene -174G[C polymorphism (p = 0.011)
(Table 2). The frequency of CC genotype was higher in
RAS patients with no family history of RAS (1.4 %)
than the patients with family history of RAS (5.3 %).
Table 3 shows the clinical characteristics (size, number,
frequency and period of recovery) of oral ulcers of RAS
patients. There was no statistically significant associa-
tion between characteristics of oral ulcers of RAS
patients and IL-6 gene -572G[C and -174G[C
polymorphisms.
Allelic and genotypic distributions of IL-6 gene
-572G[C and -174G[C polymorphisms are shown in
Table 4. The frequencies of GG, GC and CC genotypes of
-572G[C polymorphism in the patients were 78.3, 20.7,
and 1.1 % and in the controls were 68.7, 20.7, and 10.7 %.
Genotype frequencies of IL-6 gene -572G[C polymor-
phism were significantly different between patients and
controls (p = 0.001) due to an increased frequency of G
allele in patient group (p = 0.0008, OR 2.06, 95 % CI
1.35–3.17). This was reflected in the significantly increased
number of GG homozygotes and GC heterozygotes
(p = 0.0001, OR 10.8, 95 % CI 2.79–70.5) among
patients.
The frequencies of GG, GC and CC genotypes of IL-6
gene -174G[C polymorphism in the patients were 75.5,
20.7, and 3.8 % and in the controls were 38.7, 45.3, and
16.0 %. Genotype frequencies of IL-6 gene -174G[C
polymorphism were significantly different between patients
and controls (p \ 0.0001) due to an increased frequency of
G allele in patient group (p \ 0.0001, OR 3.82, 95 % CI
2.64–5.59) (Table 4). This was reflected in the significantly
increased number of GG homozygotes (p \ 0.0001, OR
4.87, 95 % CI 3.06–7.85) among patients.
As IL-6 gene -572G[C and -174G[C polymorphisms
were both associated with RAS, the risk associated
with inheriting the combined genotypes was examined
(Table 5). Homozygosity for GG at -174G[C and
-572G[C loci encoded a p value of\0.0001. Thus, indi-
viduals who are G homozygous at both loci have a higher risk
of developing RAS. However GG–CC, GC–GG, GC–GC
Arch Dermatol Res (2014) 306:173–180 175
123
Ta
ble
2C
lin
ical
and
dem
og
rap
hic
alch
arac
teri
stic
so
fR
AS
pat
ien
tsst
rati
fied
acco
rdin
gto
IL-6
gen
e-
57
2G[
Can
d-
17
4G[
Cp
oly
mo
rph
ism
s
Ch
arac
teri
stic
sT
ota
ln
=1
84
-5
72
G[
Cp
oly
mo
rph
ism
-1
74
G[
Cp
oly
mo
rph
ism
GG
n=
14
4G
Cn
=3
8C
Cn
=2
pv
alu
eG
Gn
=1
39
GC
n=
38
CC
n=
7p
val
ue
Gen
der
,m
ale/
fem
ale,
n(%
)6
6/1
18
(35
.9/6
4.1
)4
9/9
5(7
4.2
/80
.5)
16
/22
(24
.2/1
8.6
)1
/1(1
.5/0
.8)
0.5
98
54
/85
(81
.8/7
2.0
)1
0/2
8(1
5.2
/23
.7)
2/5
(3.0
/4.2
)0
.33
2
Ag
e(y
ears
)3
5.9
8±
11
.87
13
6.4
2±
12
.02
33
4.6
1±
10
.97
53
0.5
0±
13
.43
50
.56
93
6.5
3±
11
.70
73
5.2
9±
12
.45
52
9.0
0±
11
.04
50
.24
3
Ag
eat
dis
ease
on
set
(yea
rs)
28
.55
±1
1.5
94
29
.05
±1
2.0
23
26
.89
±9
.57
12
4.5
0±
19
.09
20
.52
92
8.7
1±
11
.29
32
8.4
7±
13
.05
02
6.0
0±
10
.39
20
.83
5
Fam
ily
his
tory
of
RA
S,
n(%
)0
.17
40
.01
1
Yes
70
(38
.0)
55
(78
.6)
13
(18
.6)
2(2
.9)
47
(67
.1)
22
(31
.4)
1(1
.4)
No
11
4(6
2.0
)8
9(7
8.1
)2
5(2
1.9
)0
92
(80
.7)
16
(14
.0)
6(5
.3)
Pap
ulo
pu
stu
le,
n(%
)0
.89
00
.93
7
Yes
19
(10
.3)
15
(78
.9)
4(2
1.1
)0
14
(73
.7)
4(2
1.1
)1
(5.3
)
No
16
5(8
9.7
)1
29
(78
.2)
34
(20
.6)
2(1
.2)
12
5(7
5.8
)3
4(2
0.6
)6
(3.6
)
Sy
stem
icin
vo
lvem
ent,
n(%
)0
.87
00
.14
5
Yes
1(0
.5)
1(1
00
)0
00
1(1
00
)0
No
18
3(9
9.5
)1
43
(78
.1)
38
(20
.8)
2(1
.1)
13
9(7
6.0
)3
7(2
0.2
)7
(3.8
)
Co
lch
icin
eu
se,
n(%
)0
.81
00
.58
5
Yes
24
(14
.6)
18
(75
.0)
6(2
5.0
)0
19
(79
.2)
5(2
0.8
)0
No
14
0(8
5.4
)1
07
(76
.4)
31
(22
.1)
2(1
.4)
10
7(7
6.4
)2
7(1
9.3
)6
(4.3
)
Ery
them
an
od
osu
m–
––
––
––
––
Gen
ital
ulc
ers
––
––
––
––
–
Pat
her
gy
po
siti
vit
y–
––
––
––
––
Dat
aw
ere
anal
yze
db
yan
aly
sis
of
var
ian
cean
dv2
test
.M
ean
plu
sst
and
ard
dev
iati
on
val
ues
are
pre
sen
ted
for
age
and
age
atd
isea
seo
nse
t
Th
ere
sult
sth
atar
est
atis
tica
lly
sig
nifi
can
tar
esh
ow
nin
bo
ldfa
ce
IL-6
inte
rleu
kin
6,
RA
Sre
curr
ent
aph
tho
us
sto
mat
itis
176 Arch Dermatol Res (2014) 306:173–180
123
and CC–GG combined genotypes of -174G[C to
-572G[C loci were significantly higher in control group
than patient group that these combined genotypes seem to
have protective effect against RAS (p = 0.029, p = 0.004,
p = 0.005, p = 0.0008, respectively). The haplotype
characteristics of RAS patients and controls are given in
Table 6. The G–G haplotype was observed to be signifi-
cantly higher in the patient group (74.46 vs 45.40 %,
p = 1.5 9 10-8), and the two others, C–G (14.13 vs
33.60 %, p = 7.5 9 10-9) and C–C (0 vs 5.07 %,
p = 5.0 9 10-6) were found to be significantly higher in
control subjects (Table 6). The linkage disequilibrium
(LD) between pairs of IL-6 gene polymorphisms in the
RAS patients and the control group are tabulated in
Table 6. No linkage in FMF patients and controls was
detected between -174G[C and -572G[C loci. The
observed and expected frequencies of IL-6 gene -572G[C
polymorphism were in Hardy–Weinberg equilibrium in
patient group but not in control group. However, the
observed and expected frequencies of IL-6 gene -174G[C
polymorphism were in Hardy–Weinberg equilibrium in
both groups.
Discussion
In the present study, a highly significant differences were
detected, between patients with RAS and control subjects,
in the genotype and allele frequencies of -572G[C and
-174G[C polymorphisms located in the promoter region
IL-6 gene. Patients with RAS had a lower frequency of the
minor alleles (C allele, low producer allele) for both the
polymorphisms. These results suggested that having a high
producer phenotype for carriage of IL-6 gene promoter
polymorphisms may cause oral ulcer formation, in accor-
dance to what has been demonstrated by Bazrafshani et al.
[3] who analyzed the -174G[C and -572G[C poly-
morphisms in an American Caucasian population. In this
previous study, a significant difference in the distribution
of IL-6 gene -174G[C polymorphism was observed in 91
Caucasoid RAS patients and 91 controls (p = 0.0002) [3].
In this study, the G allele observed is significantly higher in
RAS patients (p = 0.0001) and greatest risk of RAS was
associated with GG homozygosity (OR = 3.4; 95 % CI
1.9–6.2; p = 0.0001). However, in the same study, differ-
ently from our results, no significant association was
Table 3 Clinical characteristics of oral ulcers of RAS patients stratified according to IL-6 gene -572G[C and -174G[C polymorphisms
Characteristics Total
n = 184
-572G[C polymorphism -174G[C polymorphism
GG
n = 144
GC
n = 38
CC
n = 2
p value GG
n = 139
GC
n = 38
CC
n = 7
p value
Size, n (%) 0.659 0.211
Minor (MiRAS) 105 (57.1) 79 (75.2) 24 (22.9) 2 (1.9) 82 (78.1) 19 (18.1) 4 (3.8)
Major (MaRAS) 26 (14.1) 21 (80.8) 5 (19.2) 0 21 (80.8) 3 (11.5) 2 (7.7)
Minor ? major 53 (28.8) 44 (83.0) 9 (17.0) 0 36 (67.9) 16 (30.2) 1 (1.9)
Number, n (%) 0.327 0.106
One 56 (30.4) 43 (76.8) 11 (19.6) 2 (3.6) 47 (83.9) 9 (16.1) 0
Two 83 (45.1) 64 (77.1) 19 (22.9) 0 57 (68.7) 23 (27.7) 3 (3.6)
Three 27 (14.7) 24 (88.9) 3 (11.1) 0 21 (77.8) 4 (14.8) 2 (7.4)
CFour 18 (9.8) 13 (72.2) 5 (27.8) 0 14 (77.8) 2 (11.1) 2 (11.1)
Frequency, n (%) 0.208 0.519
Once a month 50 (27.2) 36 (72.0) 12 (24.0) 2 (4.0) 39 (78.0) 8 (16.0) 3 (6.0)
Twice a month 31 (16.8) 22 (71.0) 9 (29.0) 0 26 (83.9) 5 (16.1) 0
Three times a month 18 (9.8) 16 (88.9) 2 (11.1) 0 13 (72.2) 5 (27.8) 0
Four times a month 49 (26.6) 37 (75.5) 12 (24.5) 0 35 (71.4) 11 (22.4) 3 (6.1)
Once in 2 months 20 (10.9) 17 (85.0) 3 (15.0) 0 12 (60.0) 7 (35.0) 1 (5.0)
Once in 3 months 16 (8.7) 16 (100) 0 0 14 (87.5) 2 (12.5) 0
Period of recovery, n (%) 0.453 0.970
1 week 95 (51.6) 76 (80.0) 17 (17.9) 2 (2.1) 71 (74.7) 19 (20.0) 5 (5.3)
2 weeks 81 (44.0) 62 (76.5) 19 (23.5) 0 62 (76.5) 17 (21.0) 2 (2.5)
3 weeks 4 (2.2) 4 (100) 0 0 3 (75.0) 1 (25.0) 0
[3 weeks 4 (2.2) 2 (50.0) 2 (50.0) 0 3 (75.0) 1 (25.0) 0
Data were analyzed by v2 test
RAS recurrent aphthous stomatitis, IL-6 interleukin 6
Arch Dermatol Res (2014) 306:173–180 177
123
observed with IL-6 gene -572G[C polymorphism and
RAS. Another previous study composed of 64 RAS
patients and 64 controls, Guimaraes et al. [10] did not
found any difference in allele and genotype frequencies of
the -174G[C polymorphism in a Brazilian cohort with
RAS in comparison to controls.
According to combined genotypes and haplotype anal-
ysis, GG–GG combined genotype and G–G haplotype of
-174G[C to -572G[C loci were found significantly
higher in RAS patients (p \ 0.0001 and p = 1.5 9 10-8,
respectively) in our study and these combined genotype
and haplotype represented by the G allele for the -597 and
-174 loci appear to favor a progressive disease. IL-6 gene
-572G[C polymorphism was not found to be in linkage
disequilibrium with the -174G[C polymorphism [7] as we
confirmed in our study. However, the IL-6 gene -174G[C
and -572G[C polymorphisms are very close that an
association with one might be expected to result in asso-
ciation with the other [26, 28].
In vivo and in vitro studies showed that IL-6 gene
-174G[C and -572G[C polymorphisms were associated
with IL-6 production or protein expression [8, 26]. It was
demonstrated that the -174 GG and -174 GC genotypes
characterized by higher circulating IL-6 levels than the
-174 CC genotype accounted for the low producer phe-
notype [8]. A complex interactive effect on IL-6 expression
might exist for the two polymorphisms. High production of
IL-6 could be an important etiological factor for RAS and
could explain the elevated IL-6 plasma levels reported in
RAS [29]. It is important that corticosteroids and thalido-
mide, two of the most effective therapeutic agents in RAS,
are inhibitors of IL-6 production [1, 19].
Genetic factors play an important role in the develop-
ment of RAS. 24–46 % of RAS patients have a positive
family history of RAS [18]. There was also a significantly
Table 4 Genotype and allele frequencies of IL-6 gene -572G[C and -174G[C polymorphisms in patient and control groups
Polymorphism RAS patients n = 184 (%) Healthy controls n = 150 (%) p OR (95 % CI)
IL-6 (-572G[C)
Genotypes
GG 144 (78.3) 103 (68.7) 0.001
GC 38 (20.7) 31 (20.7)
CC 2 (1.1) 16 (10.7)
GG : GC ? CC 144 (78.3) : 40 (21.7) 103 (68.7) : 47 (31.3) 0.049 1.64 (1.00–2.69)
GG ? GC : CC 182 (98.9) : 2 (1.1) 134 (89.3) : 16 (10.7) 0.0001 10.8 (2.79–70.5)
Alleles
G 326 (88.6) 237 (79.0) 0.0008 2.06 (1.35–3.17)
C 42 (11.4) 63 (21.0)
IL-6 (-174G[C)
Genotypes
GG 139 (75.5) 58 (38.7) <0.0001
GC 38 (20.7) 68 (45.3)
CC 7 (3.8) 24 (16.0)
GG : GC ? CC 139 (75.5) : 45 (24.5) 58 (38.7) : 92 (61.3) <0.0001 4.87 (3.06–7.85)
GG ? GC : CC 177 (96.2) : 7 (3.8) 126 (84.0) : 24 (16.0) 0.0001 4.79 (2.06–12.3)
Alleles
G 316 (85.9) 184 (61.3) <0.0001 3.82 (2.64–5.59)
C 52 (14.1) 116 (38.7)
The results that are statistically significant are shown in boldface
Table 5 Comparative analysis of combined genotypes of RAS
patients and controls
Genotypes Patient (n = 184) Control (n = 150) p
n % n %
-174G[C to -572G[C
GG–GG 103 56.0 34 22.7 <0.0001
GG–GC 34 18.5 16 10.7 0.046
GG–CC 2 1.1 8 5.3 0.029
GC–GG 34 18.5 48 32.0 0.004
GC–GC 4 2.2 14 9.3 0.005
GC–CC 0 0 6 4.0 –
CC–GG 7 3.8 21 14.0 0.0008
CC–GC 0 0 1 0.7 –
CC–CC 0 0 2 1.3 –
The results that are statistically significant are typed in bold
RAS recurrent aphthous stomatitis
178 Arch Dermatol Res (2014) 306:173–180
123
higher disease concordance in monozygotic than dizygotic
twins [13]. This can explain the association between family
history of RAS and -174G[C polymorphism that we
detected in our study.
According to our results, the carriage of the low
producer phenotype of the IL-6 gene -174G[C and
-572G[C polymorphisms appears to protect from RAS.
Cytokines play a fundamental role in the immune response
and may to lead to a protective immunity or the tissue
destruction by the production of different profiles of
cytokines [9]. This may explain why lower producer
genotypes appear to protect from RAS.
In conclusion, our findings demonstrated that IL-6 gene
-174G[C and -572G[C promoter polymorphisms influ-
ence formation and course of RAS. If these results can be
substantiated with further studies in different populations,
evaluation for IL-6 gene -174G[C and -572G[C pro-
moter polymorphisms may become necessary in selected
patients.
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Table 6 Comparative analysis of haplotypes of RAS patients and controls
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2174G[C to 2572G[C
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G-C 0.1141 r2 = 0.0212 0.1593 r2 = 0.0247 NS NS
C-G 0.1413 v2 = 7.80 0.3360 v2 = 7.42 0.32 (0.22–0.47) 7.5 3 1029
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Arch Dermatol Res (2014) 306:173–180 179
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