Mikola et al.

Clin Transl Allergy (2018)

8:22
https://doi.org/10.1186/s13601-018-0205-z

RESEARCH Open Access

Tonsillar Tonsillar cytokine cytokine expression

expression
between
betweenpatients patients with with tonsillar
tonsillar
hypertrophyand recurrent tonsillitis
hypertrophy and recurrent
Emilia Mikola1, Varpu Elenius2†, Maria Saarinen2†, Oscar Palomares3,4,5, Matti
tonsillitis
Waris6,7, Riitta Turunen2,
Tuomo Puhakka1,8,
Vuorinen6,7,
Tobias
Lotta Ivaska1, Beate Rückert3,4, Alar Aab3,4, Tero Vahlberg9, Tytti
Allander10, Carlos A. Camargo Jr11,12, Mübeccel Akdis3,4, Cezmi A. Akdis3,4 and
The Harvard community has made this
Tuomas Jartti2*
article openly available. Please share how
this access benefits you. Your story matters
Abstract
Background: Tonsils provide an innovative in vivo model for investigating immune response to
infections
gens. and data
However, aller-are scarce on the diferences in tonsillar virus infections and immune
responses
with between
tonsillar patients or recurrent tonsillitis. We investigated the diferences in virus
hypertrophy
CitationMikola,
detection and
interferon E.,andV. Elenius,
geneT expression
cell M.undergoing
in patients Saarinen,tonsillectomy
O. Palomares, M. tonsillar
due to Waris, R.hypertrophy
or recurrent
Methods: Turunen,
tonsillitis.
Tonsils of T. Puhakka,
89 surgical et al.
patients with 2018. hypertrophy
tonsillar “Tonsillar(n cytokine expression
= 47) or recurrent
tonsillitis
lysed. (n =were
Patients between
42) carefully
were patients with
ana- characterized tonsillar
clinically. hypertrophy
Standard and recurrent
questionnaire was used to
asses preceding
symptoms. and allergy
Respiratory viruses were
tonsillitis.” analysed
Clinical andin Translational
tonsils and nasopharynx
Allergy by8 PCR.
(1): Quantitative
22.
real-time
used PCR was
to analyse intratonsillar
doi:10.1186/ gene expressions of IFN-α, IFN-β, IFN-γ, IL-10, IL-13, IL-
s13601-018-0205-z. http://dx.doi.org/10.1186/s13601-018-0205-z.
17, IL-28, IL-29, IL-37, TGF-β,
GATA3, FOXP3,
RORC2 and Tbet.
Results: Median age of the subjects was 15 years (range 2–60). Patients with tonsillar
hypertrophy
smoked were younger,
less often, Published
had less Versiondoi:10.1186/s13601-018-0205-z
pollen allergy and had more adenovirus, bocavirus-1, coronavirus
and rhinovirus in <nasopharynx P
0.05). Only(all
bocavirus-1 was more often detected
< 0.05). in P
In hypertrophic
age-adjusted
analysis,tonsils ( hypertrophy was associated with higher mRNA expressions
tonsillar P
< 0.05).
Citable linkhttp://nrs.harvard.edu/urn-3:HUL.InstRepos:37298440
Conclusions: of IL-37 ( T cell and interferon gene expressions appeared to be relatively
Intratonsillar
stable
lar for both and
hypertrophy tonsil-
recurrent tonsillitis. Of the studied cytokines, only newly discovered
anti-infammatory
IL-37, cytokineassociated with tonsillar hypertrophy showing slightly stronger anti-
was independently
Terms
infammatory responsearticlein was
of UseThis downloaded
these patients. from Harvard University’s DASH
repository,
Keywords: Allergy,and is made
Asthma, Child, available
Cytokine,under the terms
Interferon, and conditions
Interleukin, T helper
Tonsillar
the
sillar
applicablefeld disease
of
disease
otorhinolaryngology.
centrally
tocdoma include
Other is onerecurrent
located
Posted of
ment the is
at
most
Diferent
tonsillitis
usually
Material,the common
beginning
antibiotics
astypes
disorders
and
set of
of
forththeor respiratory
at http://
cell, Tonsil,
(http://creat
© and
provided
The Author(s)
indicate
you Virus
give
iveco
if2018.
appropriate
changes
mmons
This
publi
.org/licen
were
article
credit
made.
is
in/zero/1.0/)
ses/by/4.0/),
todistributed
The the
Creative
original
applies
under
Commons
which
author(s)
the
permits
to
Public
terms
theand
unrestricted
data
Domain
ofthethe
made
source,
Dedication
Creative
available
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provide
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link
Attribution
(http://creat
article,
to and
the Creative
unless
4.0iveco
nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-
reproduction
International
Commons
mmons .org/ Background
in
ton-
hypertrophy,
tonsillar
license,
inLicense
any medium,otherwisegastrointestinal
gens
play
immune
that
with
breathing,
and
comes
aTonsils
conventional
[1].
between
stated.tonsils
both
into
system.
are use#LAA
Surgically
leading
snoring,
contact
tonsillectomy.
foreign
are
secondary
Our
accessible
tracts
organs
to
previous
with
dyspnea,
removed
pathogens,
symptoms
lymphoid
where
where
infections
source
studies
palatine
apnea
the
immune
ofallergens
organs,
mouth
to
immune
or
agents
demonstrated
study
regulation
tonsils
dysphagia.
which
system
and
and the
the provide
are
aller-
inter-
frst
takes
Treat-
host
 Clinical and
Translational Allergy
 et al. Clin Transl Allergy (2018) Page 2 of 8Mikola
8:22

place in which allergen-specifc regulatory T cells Defnitions can

be generated by Tonsillar
mechanisms hypertrophy
dependinggroup on plasmacy-
was defned as patients who
toid dendriticunderwent cells [2,tonsillectomy
3]. Te expression because of of T cell-
obstructive symp-
and
interferon specifc toms genes
such as in snoring,
tonsils has breathing
been showndifculties
to or swallow-
be closely related ing problems.
to existing Tereviral
were infections,
no tonsillarage, infection problems
and
allergic illnesses in this[4, group.
5]. However,
Recurrentdata tonsillitis
are scarce group
on was defned
the
diferences inas patients
tonsillarwho virusunderwent
infections tonsillectomy
and immunebecause of recur-
responses between rentlypatients
infectedwith tonsils
tonsillar
(viralhypertrophy
or bacterial) during the
or recurrent past tonsillitis
6–12 months. [6–8].
Tose operated because of acute infection
Tonsils appear to provide a or goodperitonsillar
in vivo modelabscess for were excluded.
investigating the mechanisms of infammatory processes
and infections in lymphoid organs [2–5]. Analysis Terefore,
of viruses we and cytokines
studied whether In-house
virus real-time
detection PCR and assays
T cell wereand inter-
used to detect
feron gene expressions human bocavirus-1,
difered betweenrhinovirus,the two enterovirus,
main and
indications oftory respira-
surgery,
syncytial tonsillar
virus hypertrophy
as describedorpreviously [4].
recurrent
tonsillitis. Seeplex RV12 ACE Detection (Seegene, Seoul, Korea) multiplex
PCR assay was used for detection of adenovirus, corona-
Methods viruses (229E/NL63 and OC43/HKU1), infuenza A and
Patients B viruses, metapneumovirus, parainfuenza virus types
Human tonsil samples 1-3, respiratory
used in this syncytial
study were virusacquired
group A and B, and
from 200 consecutive rhi-
novirus tonsillectomy
[4, 5]. Virus patients diagnostics who were
under-carried out in
went tonsillectomy the
Department
in Satakunta
of Virology,
CentralUniversity
Hospital,ofPori, Turku, Turku,
Finland between Fin-
land, April
and 2008
in the andDepartment
March 2009. of TeClinical
inclu- Microbiology,
sion criteria were Karolinska elective tonsillectomy
University Hospital, accordingStockholm,
to Sweden.
clinical indication To isolate
and writtentotal informed
RNA from consentpalatinefrom tonsils, tissues
the study patient (pre-
and/or his/her guardian. viously
) were
Te studystabilized later
homogenized in
in RNA
pro-
tocol was approved grinding by the
tubes ethics
containing
committee CK28ofceramic
Satakuntabeads by using
Central Hospital, a Precellys
Pori, Finland.
24 homogenizer
Study was(Bertin initiated Technologies, Mon-
only
after obtaining tigny
written
le Bretonneux,
consent from France)
the participant
two times at or 6000 rpm
his/her guardian. for 50 s [4]. Total RNA from cell samples was isolated
using the RNeasy mini kit (Qiagen, Hilden, Germany).
Study protocol Reverse and sample transcription
collection was performed with the Revert Aid
A standard questionnaireM-MuLV Reverse was Transcriptase
used to obtain(Fermentas, informa- St. Leon-
tion on allergic Rot,diseases
Germany)and using
respiratory
random hexamer symptoms primers according
within
30 days before to thetheoperation
manufacturers(Additional
protocol. fle 1: Gene expressions of
Table 1).
Tonsillectomy α, IFN-
was IFN-β,
performed IFN-γ,according
IL-10,toIL-13, routine IL-17,
clini-IL-28, IL-29,
cal
viral procedure.
aspirate analyses,
were IL-37,
TGF-β,
Internal
Uvalues
gation
Continuous
stored
systems,
test
a partdueFOXP3,
tonsillar
multiplied
factor
in of
Foster
to
dry GATA3,
variables
the
skewed
1α
tubes tissue
tonsils
City,
by
(EF1α) −and
RORC2
distribution.
at
are
104,
CA, was
and
described
was
80
USA).
where
a°C
usedTbet aswere
Housekeeping
ΔCT
[4]. analysed
Categorical
for corresponds
medians
normalization.
gene
and to
Hospital
Full listand
of author
Turku University,
informationP.O.
is available
Box 52, 20520
at theTurku,
end of the
−
immediately
cut
zation
at
nasopharyngeal
pharyngeal
Naso-
thesia
25(OH)D
(Abbott
els
measurements
published
80
+of
in4°C
3–4
reagent
°C
using
25(OH)D
measurement
Architect,
ofafter
*Correspondence:
Finland
article
Varpu
Department
Elenius formulae.
until
aspirate
andmm
are
diference
variables
athe
(D-binding
by
Supermix
Data
quartile
inter-
7900HT
elon-
cubes,
(Qiagen,
standardized
were
removal
quantitative
Paediatricsshown
next
Chicago,
samples
was
estimated
Fast
stored
working
with
ranges,
as
between
tuomas.jartti@utu.f
Maria Saarinen
and of
Hilden,
done
protein
relative
Real-Time
†the
ROX
Adolescentwere
contributed thelate
procedure
USA)
in
using
real-time
day
using
and
non-absorbed
(Bio-Rad,
RNA
Germany),
and
obtained
and
and
were
expressions,
CT
Medicine,an
equallyrPCR
additional
albumin)
bioavailable
RNA
value
fnally
[4].
immunoassay
Turku
as PCR
analysed
instrument
Hercules,
stabili-
incubated
during
Serum
secondusing
reagent.
for
stored
and
Universityserum
which
the
using
author2 anaes-
total
iTaq
lev-
(Applied
and
CA,
gene
atFor
show
SYBR
Statistical −(ΔCT)
Mann–Whitney
EF1α.
USA)
ofanalysis
Green
2Bio-
interest
on a
 et al. Clin Transl Allergy (2018) Page 3 of 8Mikola
8:22

are expressed as frequencies and percentages, and were Results

analysed using Chi square test or Fisher exact test. Study population
Clini-
cal, viral and immunological diferences between study
Originally, 200 patients participated in the study. Of
groups were analysed
them, 46using unadjusted
subjects did notand multivariable
have remaining tonsil and/
linear regression
or nasopharyngeal samples for the immu-
analysis. Te adjustments for current analysis, and
nologic analyses were chosen using backward stepwise
11 had no intratonsillar virology done in their samples.
multivariableAnother
models 54
that initially
subjects wereincluded
excludedclinical
for having mixed
fac-
tors and virusindications
infections which signifcantly difered
of operation other than hypertrophy or
ton-
between the groups
sillitis (Fig. 1). Tus, 89pollen
(age, self-reported allergy,
patients comprised the
self-smoking, analytic
both adenotomy and tonsillectomy per-
cohort. Forty-seven (53%) of them had tonsillar
hypertro-
formed, respiratory symptomsphyone
andmonth priorhad
42 (47%) to recurrent
the
operation and bioavailable tonsillitis.
25(OH)D level). Te fnal
model was adjusted only for age. Before regression Patient characteristics
anal-
yses, cytokine and transcription factor values
All operations were performed during wereafebrile period
log-
transformed because of positively skewed distributions.
of chronic tonsil condition. Respiratory symptoms on
Te mean diference was computed
the operation day for
werelog-transformed
present equally in the hyper-
values: a recurrently infected group
trophy group and the recurrentminus hypertrophic
tonsillitis group (15
group. Statistical analysis was completed using
= 0.72).vs.
Te 18%,JMPrespectively;
median age ofP the
version 12.0.1 software (SAS Institute Inc. Cary, NC,
patients was 8 years (range 2–46) and 20 years (range
USA). A two-sided P < 0.05 was considered
< 0.001) statistically
(Table
7–60), P
1).respectively
In addition (
to
signifcant. being younger, patients in the hypertrophy group had
more often adenotomy and tonsillectomy done, had
 et al. Clin Transl Allergy (2018) Page 4 of 8Mikola
8:22

Table 1 Patient characteristics

CharacteristicsTonsillar hypertrophy P tonsillitis
Recurrent value
n = 47 n = 42

Age, years (range)8 (2, 46)20 (7, 60)< 0.0001

Male27 (57%)23 (55%)0.80
Tonsillectomy and adenotomy28 (60%)5 (12%)< 0.0001
Self-reported allergy20/43 (47%)23/40 (58%)0.32
Food6/43 (14%)3/39 (8%)0.38
Drug4/43 (9%)4/39 (10%)0.87
Seasonal, i.e. pollen0/43 (0%)7/39 (18%) 0.004
Perennial, i.e. animal or house dust mite3/43 (7%)2/39 (5%)0.74
Other2/43 (5%)2/39 (5%)0.90
Multiple5/43 (12%)4/39 (10%)0.86
Physician-diagnosed atopic dermatitis11/44 (25%)4/38 (11%)0.09
Self-reported allergic rhinitis11/44 (25%)11/39 (28%)0.74
Physician-diagnosed asthma6/42 (14%)6/39 (15%)0.89
Self-smoking2/43 (5%)14/40 (35%) 0.0005
Maternal smoking15/44 (34%)13/41 (32%)0.82
Paternal smoking16/41 (39%)19/36 (53%)0.23
Season of the surgery0.85
Winter (months 12–2)7 (15%)4 (10%)
Spring (months 3–5)14 (30%)13 (31%)
Summer (months 6–8)8 (17%)5 (12%)
Fall (months 9–11)18 (38%)20 (48%)
Respiratory symptoms
The operation day6/39 (15%)7/38 (18%)0.72
Within 2 weeks16/42 (38%)12/37 (32%)0.60
Within 4 weeks23/42 (55%)18/37 (48%)0.59
One month prior the operation
Throat pain8/39 (21%)14/35 (40%) 0.0005
Rhinitis23/39 (59%)6/35 (17%) 0.0002
Cough15/39 (38%)5/36 (14%) 0.02
Acute otitis media2/39 (5%)0/35 (0%)0.17
Wheezing2/39 (5%)0/35 (0%)0.17
25(OH)D level
Total (nmol/l)56.2 (41.7, 66.9)45.7 (34.4, level
25(OH)D
compared 72.3)0.08
toanother
Fig.
8fever
smelling
had1other
Values nare
hypertrophy,
Study
= shown
1).
breath,
indication
fow
29ashad
chart.
and
tonsillitis,
recurrent
medianfeeling
of18 operation
had of
hypertrophy
tonsillitis
and
beat
(interquartile another
than nthe
in hypertrophy
throat
range,and coronavirus
in
15%
indication
except =than
<orrecurrent
1;
age)
0.05)
of patients
patients
indication
(recurrent
recurrent
or
<0.001).
(Table
tonsillitis ortonsillitis
number offever
rhinovirus
inwith
n2).
nIn
(recurrent
fever, the
(chronic
subjects
addition,
However,
hypertrophy
= (%). recurrent
=1;white
otitis
fever
5; group
teethgroup,
nin
recurrent
Data
nasopharyngeal
patches
media
braces
= 8;
were
intratonsillar
n(Table
patients
nhad
=and
recurrent
otitis
in amedia,
only
=tonsils
analysed4;2; 2). in
recurrent
by throat Patients
otitis2%
the
aspirate
virus
of
n =recurrent
abscess
2;
otitis
media
Mann–Whitney ==in
accumulation
U nnmedia
fever
test,1).
1;Chi
nno
Free
and
=321
ofclear
1;
food
square (pg/ml)6.4
fever
had
recurrent
cause = (4.9,
hypertrophy
remnants,
n or
test, = n3;
otitis 8.1)5.0
recurrent
1)
bad
Bioavailable
Fisher’sandmedia 1less
group
tonsillitis
diferences
Signifcantly
virus
n =(3.3,
tonsillitis;
Exact 2). month
48.8)0.14
Viruses
(ng/ml)2.2
Signifcant
test P were
self-reported
throat
(all
prior<tion
detected
(1.7,
values pain,
phy
(all
hypertrophy
the
more
patients
except
0.01)
in
the
detec-
found
group
in
2.9)1.6
are their
didn’t
shownbut
operation
patients
pollen
(Table
for
hypertro-
had
between
nasopharyngeal
bocavirus-1
group
show
allergy,
more
nasopharyngeal
(1.1,
in 2.7)
italic 1).
inandstatistically
the
Otherwise
were
often
higher
two
hypertrophy
smoked
more
which
rhinitis
adenovirus,
aspiratesaspirates
groups.
bioavailable
no
often
less,
was
and signifcant
signifcant
and
group,
detectedPtively;
positive
(79
and
tonsilsbocavirus-1,
cough
vs. P tonsils
haddiferences,
in
38%,
for one
respec-
0.01 in
 et al. Clin Transl Allergy (2018) Page 5 of 8Mikola
8:22

Table 2 Nasopharyngeal and intratonsillar virus detection

VirusNasopharynx P P
valueTonsil
value

Tonsillar hypertrophy Recurrent tonsillitis

Tonsillar
Recurrenthypertrophy
tonsillitis
n = 47 n = 42 n = 42
47

Adenovirus9 (19%)2 (5%) 7 (15%)2 (5%)0.10 0.03

Bocavirus-112 (26%)2 (5%) 7 (15%)1 (2%) 0.005 0.03
Coronavirus3 (6%)0 (0%) 0 (0%)0 (0%)– 0.048
Enteroviruses4 (9%)4 (10%)0.876 (13%)4 (10%)0.63
Infuenza A or B virus1 (2%)0 (0%)0.260 (0%)0 (0%)–
Metapneumovirus0 (0%)1 (2%)0.291 (2%)0 (0%)0.26
Parainfuenza virus types 1-41 (2%)1 (2%)0.944 (9%)1 (2%)0.19
Respiratory syncytial virus1 (2%)0 (0%)0.262 (4%)0 (0%)0.11
Rhinovirus species A, B or C27 (57%)14
2 (4%)2
(33%)(5%)0.91 0.02
Number of positive viruses37 (79%)1617(38%)<
(36%)8 (19%)0.07 0.0001
Positive for 1 virus23 (49%)10 (24%)9 (19%)7 (17%)0.76 0.01
Positive for 2 viruses8 (17%)4 (10%)0.305 (11%)0 (0%) 0.01
Positive for 3 viruses5 (11%)2 (5%)0.302 (4%)1 (2%)0.62
Positive for 4 viruses1 (2%)0 (0%)0.261 (2%)0 (0%)0.26
Positive for ≥ 1 viruses37 (79%)16 (38%)17 (36%)8 (19%)
Positive for ≥ 2 viruses14 (30%)6 (14%)8 (17%)1 (2%)
Positive for ≥ 3 viruses6 (13%)2 (5%)3 (7%)1 (2%)
Positive for ≥ 4 viruses1 (2%)01 (2%)0
Values are shown as number of subjects (%). Data were analysed by Chi square test, or Fisher’s
Exact test
Signifcant values are shown in italic

virus in theirTable
nasopharyngeal aspirates
3 Intratonsillar cytokine (49 vs. 24%) or factor
and transcription
two viruses in expression
their tonsils (11 vs. 0%,
in hypertrophic respectively)
tonsils and recurrent tonsillitis
(both
P < 0.05) (Table 2). Tonsillar
Recurrent
Cytokine
hypertrophy
tonsillitis
or transcription
n =factor
47
42
Cytokine and transcription factor expression
Medianprofles
(IQR)Median (IQR)
in tonsils
T-helper1
In unadjusted analysis, patients in the hypertrophy
IFN-γ58 (37, 90)76 (32, 117)
group
had stronger tonsillar expression of PTbet = 0.03)
( and
Tbet54 (31, 83)34 (22, 60)
IL-37 P( = 0.001) than patients in the recurrent
T-helper2
tonsillitis
group (Tables 3, 4). In the multivariable regression
analy- IL-131.2 (0.03, 3.7)0.45 (0.02, 2.2)
sis, onlythey
recurrent
compared
tively, <toage remained
tonsillitis.
0.05,
patients
also Fig.
had as
with
2).
less arecurrent
Patients
No signifcant
self-reported
otherwith co-factor
diferences
tonsillitis.
tonsillar
pollen in allergy,
(Table 4). interleukin; GATA327
IQR, interquartile
GATA3, (16, 36)18
GATA-binding
range; (12,
Values
IFN,
factorare39)
interferon;
3;arbitrary
RORC, Tbet,
RAR-related
units
T-box
× 104
transcription
orphan
relative
receptor
to
After
or
the
Tis
going
were adjustment
(P cytokine
wasdiferences
group
Discussion
cell
hypertrophy
only
Respec-
no
but transcription
independently
groups.
study
and
typically
bocavirus-1
tonsillectomy
interferon
showswere
C; for
younger, age,
diferences
factor
was
associated
found
gene
due
more
IL-370.26
TGF-β163
IFN-α15
IFN-β24
factor; the
IL-1710
IL-2911
IL-2831
RORC215
IL-1049
FOXP346
IL, expressions
expression
to
and
expressions
in often
infood
tonsillar
had
with
virus
more
found
(0.15,
(105,
(5.6,
(0.59,
(2.3,
(2.7,
(3.1,
(7.2,
(24,
(20, allergies
FOXP,tonsillar
were
detections
inin
0.37)0.14
232)171
74)35
87)48
19)7.3
34)3.7
101)22
88)12
31)20
62)9.7(21,
(24,
(1.4,
(10,
forkhead
EF1α of
hypertrophy
viralfound
patients
tonsils
(4.1,
(1.3,
(2.1,
(120,
(0.27,
(0.10,
64)
80)
box 28)
75)only
fndings,
between
13)
26)
103)
225) IL-37
hypertrophy
between
andwhen
56) under-
or
0.24)
protein; Tbut
the
Type
T-helper17
T-regulatory
TGF, I/III growth
tumour interferons
factor
 et al. Clin Transl Allergy (2018) Page 6 of 8Mikola
8:22

Table 4 Diferences in cytokine and transcription factor expression between hypertrophic tonsils and recurrent
tonsillitis
Cytokine or transcription factorMean diferences recurrently infected minus hypertrophic group

UnivariateMultivariate

nDiference of means (95% CI)Adjusted diference of means (95% CI)Adjustments

T-helper1
IFN-γ880.061 (− 0.32, 0.44) ––
P = 0.75
Tbet89− 0.40 −( 0.76,−0.034) − 0.21 (− 0.61,
Age 0.18)
P = 0.03 P = 0.29
T-helper2
IL-1389− 0.74 (− 1.8, 0.36) ––
P = 0.18
GATA389− 0.12 (− 0.42, 0.18) ––
P = 0.43
T-helper17
IL-1789− 0.35 (− 0.80, 0.096) − 0.087 (− 0.57,
Age 0.40)
P = 0.12 P = 0.72
RORC2890.014 (− 0.35, 0.38) ––
P = 0.94
T-regulatory
IL-1089− 0.26 (− 0.61, 0.0871) − 0.0070 (− 0.38,
Age 0.36)
P = 0.14 P = 0.97
IL-3787− 0.48 −( 0.77,−0.19) −0.31 −
−(
0.0021)
0.63,
Age
P = 0.001 P =0.049
FOXP3890.091 (− 0.31, 0.49) ––
P = 0.65
TGF-β89− 0.0077 (− 0.32, 0.30) ––
P = 0.96
Type I/III interferons
IFN-α87− 0.41 (− 1.5, 0.7) ––
P = 0.47
IFN-β88− 0.21 (− 1.0, 0.61) ––
P = 0.62
IL-2889− 0.64 (− 1.5, 0.2) ––
P = 0.13
IL-2987− 0.57 (− 1.5, 0.31) ––
P = 0.15

Data are expressed as mean diferences as a recurrently infected group minus hypertrophic group. The data were analysed using backward
stepwise linear regression groups.
of
tal
epithelial
IL-7,
analysis
CI, confdence interval; IFN, interferon;
IL-37.
inhibitor
afterIFN-γ,
After
Other
cells
tion/regulatory”
Tbet, logarithmic
cytokines
previously
tory
of
analysis
age-adjusted
of
andthan
IFN-λ
innate
[11].
TNF-α)
cytokine
age,adds
with
After
transformation.
T-box transcription
family
immunity
shown
no
and
analysis,
IL-37
factor; Only
that
antiviral
other
ligand
augments
that
[12,
cytokines
tonsillar
[9,
issignifcant
the
13].
activation,
also
signifcant
IL, interleukin;
tonsillar
10].
activity
the
expression
Tey
(IL-10,
independently
It
expression
level
co-factors are
has
co-
and
were IL-37
produced
IL-17,
been
of
of
their
used
GATA3, GATA-binding asIL-37
ofand
IL-37,
anti-
expression
byis TGF-
adjustments
factor 3; RORC,inRAR-
the
fnal model
was
hypertrophy
were
factors
to
shown
inhibits
infammatory
anti-
IL-37
beassociated
found.
expressed
(formerly
cytokines
IL-10
β,
is
FOXP3,
closely
positively
induced
dendritic
and
infamma-
Interferons
with
inpositively
IL-1
and
macrophages,
GATA3,
higher
byT(especially
family
regulatory
viral
(IFN-α,
RORC2,
tonsillar
associated
member
associated
infection.
monocytes,
Tbet)
IL-1β,
cells
IFN-β,
7)
mRNA is
with
inwith
[11].
IL-28
expressions
but
atonsils
plasma
IFN-γ,
fundamen-
other
also
tonsillar
We
andhave
and
“immune
[2].
IL-6,
IL-29
IL-28, Te
hypertrophy.
are
IL-29)
current
activa-
membersare
related orphan receptor C; FOXP, forkhead
Signifcantboxvalues
protein;
are TGF,
shown tumour
in italic
growth factor
 et al. Clin Transl Allergy (2018) Page 7 of 8Mikola
8:22

adenovirus, bocavirus-1, coronavirus and rhinovirus.

However, intratonsillar virus detection was low and did
not show any statistically signifcant diferences except
for bocavirus-1. Te results of nasopharyngeal and
intra-
tonsillar virus detection vs tonsillar cytokine
responses are discussed in detail in our previous report
[2].
Small diferences in cytokine expression may partly
be explained by diferences concerning tonsillar germi-
nal centers. Te mean follicular area has been found to
be larger, and the number of germinal centers higher,
in
the hypertrophy group compared to the recurrent tonsil-
litis [17–19]. In our study, the samples were taken
from of the tonsils to minimize the margin of error
inside
and possibility to misinterpreted diferences between
the
the
groups. Seasonal changes, e.g. pollen and infuenza sea-
sons, may afect the expression of peripheral T cells
[20],
but we found no diferences in tonsillar expression of
cytokines between the seasons of the surgery. Also,
res-
piratory viruses are continuously detected in children
with chronic tonsillitis throughout the year [21–23].
Cir-
culating serum 25(OH)D level has been shown to been
positively associated with IL-37 level [3], but here it
did not confound the results.
A limitation of the current study is that we did not
investigate bacterial colonization of the tonsils in
these
patients following
cells and macrophages due to fact thatinfection
viral the operation or was done during
acti- an afebrile
vation with bacterial period of
components their chronic
[12–14]. tonsil condition.
We expected
Te
to see diferencesdownstream
in IFN signaling
expressionof(lower
IL-37 responses
is a complex in process and
to show functionality
recurrent tonsillitis than in tonsillar of IL-37 by downstream mediators
hypertophy
group),
since they have wasantiviral
not in the scope of and
properties thisthey
study. In addition to
up-regu-
forming
cell-surface
late the expression of MHC receptor
Class II complexes,
molecules on IL-37cells translocates to
which increases the
nucleus where system’s
the immune it binds to nuclear
ability to DNA and participate
recognize
viruses [14, 15].in
transcription.
However, we[24, did 25]
not IL-37
observe is these
regarded as a “dual
dif- func-
ferences. We speculate thattion” cytokine,
tonsillar similar
hypertophy maytobeIL-1α and IL-
a consequence of chronic infammation33. in tonsils and the
same interferon pathways are equally activated in both Conclusions
conditions.
between
patients,
2–11
rent years)
the
theWe
Age tonsillitis
was In
butage
cell
may summary,
have
feron
stronger
lymphoid
main
two
the
distribution
explain
group.
research
main
narrow
clinical
genein this
previously
organs.
Most study
indications found
expressions
age
the
patients
in provides
strong
characteristic
of
range
lack
often
lymphoid
Our
hypertrophy
patients
of
with
data
of
detected
appear
diference new
tonsillectomy,
their
tissue
tonsillar
suggest
with
to
and
subjects
viruses
from insights
intragroup
relatively
that
recurrent
hypertrophy
theT-cell about
tonsillar
clinical
stable
and
than T
over
correlations
IFN-γ,
previous
the
where
diference
ences
Interestingly,
tonsillitis
(ages
[17].
with
were
diferentiating
Virus
tonsillectomy
hypertrophy
tonsillar
aswas
in IL-28)
fndings
adults
smoking
between
found
aspect
tonsils
of
tory
the
mechanisms
with
inter-
phy
hypertro-
[2].
Reis
hypertrophy
tonsillar
is
have
the
[16],
and
in
and
indication
the
immune
more
and
surgical
the
as
more
in
recurrent
groups
we
acolleagues
common
additional
of
nasopharynx
responses,
good
IFN
found
recurrent
group
infammatory
groups.
also
indication
expression(IFN-α,
in
with
tonsillitis.
that
and
tonsillitis.
vivo
explains
adenotomy
found
younger
tonsillitis.
namely
of
Inmodel
obstruction
38%processes
79%
agreement
for
nothe
in IL-37,
of
children
diference
recur-
tonsillectomy.
However,
for
performed.
Additional patients
difer-
fleAdditional
investigating
IFN-β,
due
Te
and
with
might
1. age
to
infections
anti-infamma-
Health be We
slightly
flefound
thein
questionnaire.
 et al. Clin Transl Allergy (2018) Page 8 of 8Mikola
8:22

Authors’ contributions 5. Elenius V, Palomares O, Waris M, Turunen R, Puhakka T, Rückert B, et

al. The relationship
The study protocol and manuscript were writtenofbyserum the investigators.
vitamins A, D, Tonsil E and LL-37 levels with
samples and clinical data were allergic
collected
status, by tonsillar
TP, MS andvirus LI.detection
Viral analysesand immune response. PLoS ONE.
were
performed by MW, TVu and TA Cytokine analyses were conducted by OP, RT, 2017;12(2):e0172350.
BR, AA and supervised by MA and 6. Andersson
CAA. CAC organized
J, Abramsthe J, Björk
vitaminL,DFuna analyses.
K, Litton M, Agren K, et al.
Data were analyzed by EM, VE,Con- comitant
MS and TJ in andvivobyproduction
statistician of 19 TVa.diferent
All cytokines in human tonsils.
authors
read and approved the fnal manuscript. Immunology. 1994;83(1):16–24.
7. Kim J, Bhattacharjee R, Dayyat E, Snow AB, Kheirandish-Gozal L, Goldman
Author details1 JL, et al. Increased cellular proliferation and infammatory cytokines
Department of Otorhinolaryngology, in
tonsils derived
Turku University
from children Hospital with andobstructive
Turku sleep apnea. Pediatr
Res.
University, Turku, Finland. 2 Department of Paediatrics and Adolescent 2009;66(4):423–8.
Medi-
cine, Turku University Hospital 8. Woonand HG,TurkuBraunUniversity,
A, Li J, Smith P.O. C, BoxEdwards
52, 20520 J, Sierro F, et al.
Turku,
Finland. 3 Swiss Institute Compartmen-
of
talization
Allergy and of total
Asthmaand Research,
virus-specifc University tissue-resident
of memory CD8+ T
Zürich,
Davos, Switzerland. 4 Christine cellsKühne-Center
in human lymphoid
for Allergy organs.Research PLoS andPathog. 2016;12(8):e1005799.
Educa-
tion, Davos, Switzerland. 5 Department 9. Nold MF,ofNold-Petry
Biochemistry CA, and ZeppMolecular
JA, Palmer Biology,
BE, Bufer P, Dinarello CA.
School of Chemistry, Complutense IL-37 Universityis a fundamental
of Madrid, Madrid,inhibitor Spain.
of innate immunity. Nat Immunol.
6 Department of Clinical Virology, Turku University Hospital, Turku, 2010;11(11):1014–22.
Finland. 7 of Virology, University
Department 10. Banchereau
of Turku,J,Turku, Pascual Finland.
V, O’Garra 8 A. From IL-2 to IL-37: the
Department
of Otorhinolaryngology, Satakunta expand-Centraling spectrum
Hospital,ofPori, anti-infammatory
Finland. 9 cytokines. Nat Immunol.
Depart-
ment of Biostatistics, University of Turku and Turku University Hospital, 2012;13(10):925–31.
Turku,
Finland. 10 Department of 11.Clinical
Conti P,Microbiology,
Carinci F, Lessiani Karolinska G, Spinas
UniversityE, Kritas SK, Ronconi G, et al.
Hospital, Potential of
Stockholm, Sweden. 11 Department therapeutic
Emergency use Medicine,
of IL-37: Massachusetts
a key suppressor of innate immunity and
General Hospital, Harvard Medicalallergic School, immuneBoston,responses
USA. 12 Division mediated of by mast cells. Immunol Res.
Rheu-
matology, Allergy and Immunology, Department of Medicine, Massachusetts 2017;65(5):982–6.
General Hospital, Harvard Medical 12. KelmSchool,NE, ZhuBoston,Z, DingUSA. VA, Xiao H, Wakefeld MR, Bai Q, et al. The role
of IL-29 in immunity and cancer. Crit Rev Oncol Hematol. 2016;106:91–8.
Competing interests 13. Witte K, Witte E, Sabat R, Wolk K. IL-28A, IL-28B, and IL-29:
Fig. 2 Relative tonsillar expression of IL-37. Forty-two
The authors declare that they promising
cytokines
have no with competing
type Iinterests.
interferon-like properties. Cytokine Growth
recurrent
tonsillitis samples compared Factorwith 47 hypertrophic tonsil Rev. 2010;21(4):237–51.
samples.
Values are arbitrary units x 104 relative to EF1α. Data are
Ethics approval and consent 14.toAkdis
participate
M, Burgler S, Crameri R, Eiwegger T, Fujita H, Gomez E, et al.
represented
asstudy
median with interquartile
The protocol Inter-
was approved byrange.
leukins, thefrom IL-37,
ethics Interleukin
1 tocommittee
37, and interferon-γ:
of Satakunta receptors, functions, and
37 roles in
Central Hospital, Pori, Finland. Study was initiated diseases.only J Allergy
after obtaining
Clin Immunol. 2011;127(3):701–21.
written
consent from the participant 15. Commins
or his/her S, Steinke
guardian.. JW, Borish L. The extended IL-10 superfamily: IL-
10, IL-19, IL-20, IL-22, IL-24, IL-26, IL-28, and IL-29. J Allergy Clin
Funding Immunol. 2008;121(5):1108–11.
TJ and his laboratory are supported 16. Rosenmannby the E, Academy
Rabinowitz
of FinlandR, Schlesinger
(Grants 114034M. Lymphocyte subsets in
and 132595), the Finnish Medical human Foundation,
tonsils: thethe efect
Sigrid
of age Juselius
and infection.
Foundation, Pediatr Allergy Immunol.
and the Foundation for Pediatric Research. EM and MS are supported by the 1998;9(3):161–7.
Foundation for Pediatric Research.
17. Reis LG, VE is Almeida
supported
EC, da by Silva
Tampere JC,Tuberculosis
GeA Pereira, VeF Barbosa,
Foundation. OP is a RamonEtchebehere
y RM.
Cajal Tonsillar
Scholar hyperplasia
funded by MINECO and recurrent
and the Euro- tonsillitis: clinical-
pean Social Fund. MA’s laboratory histological cor- relation.
is sponsored by EU 7thBraz Framework
J Otorhinolaryngol.
Program 2013;79(5):603–
PREDICTA: Post-Infectious18. Immune PC, Pang YT, 8.
ZhangReprogramming and
Loh Its
KS, Association
Wang DY. Comparison with of histology between
Persistence and Chronicity recurrent
of Respiratory tonsillitis
Allergic andDiseases
tonsillar (No. hypertrophy.
260895). Clin Otolaryngol Allied
Labo-
ratory of CAA is supported Sci. by the Swiss National Science Foundation Grant 2003;28(3):235–9.
32-132899 and Christine Kühne-Center
19. Agren K,for Andersson
AllergyU,Research
Litton M, andFunaEducation.
K, Nordlander B, Andersson J. The
The granting agencies coveredproduction all costs of andimmunoregulatory
played no role in cytokines
study design,
is localized to the extrafol-
data analysis, or manuscript preparation. licular area of human tonsils. Acta Otolaryngol. 1996;116(3):477–
20. Jartti85. T, Burmeister KA, Seroogy CM, Jennens-Clough ML, Tisler CJ,
SalazarLP, et al. Association between CD4+CD25(high) T cells and atopy in
Publisher’s
Springer
2. Jartti
3.
4. Palomares
Kücüksezer
humanNature
tinct tonsils
regulation Note
T, Palomares
O,UC,
remains
Rückert
as
Palomares
potential
of neutral
O,
22.
tonsillar
B,
25.
Waris
24.
Jartti
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MC,
O,
caspase-1
Bulau
chronic
chil-frst-line
Sharma
with
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recurrent
et
M,C,
immune
T,
al.
AM, ıS,
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regard
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rdar
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nuclear
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