Autoimmune mechanisms in chronic idiopathic urticaria

Article (PDF Available)inThe Journal of allergy and clinical immunology 130(3):814-816.e4 · June 2012with29 Reads
DOI: 10.1016/j.jaci.2012.04.037 · Source: PubMed
Fig 1, A shows the daily percentage of patients with nasal, oc-
ular, and respiratory symptoms as well as the evolution of daily
pollen levels, weighted according to the number of participants
per town and per day. Although the start of the pollen season is
staggered during the month of April in different cities, significant
peaks are detected mostly between mid-May and mid-July. Fig 1,
B-D displays the shape (with a 95% CIs) of the pollen-symptoms
relationship from the generalized additive mixed models, ad-
justed for correlated response, time trend, and meteorological var-
iables. The effect of a pollen-level increase on the x-axis leads to a
symptoms increase on the y-axis, expressed as ‘ln(OR)’ (natural
logarithm of the odds ratio). The plot suggests that the effect of
grass pollen on nasal and ocular symptoms is more or less linear,
up to a saturation point of 80 and 90 grains/m
3
, respectively.
Above these cutoffs, the effects of pollens on these symptoms re-
main relatively constant. The plot for respiratory symptoms
shows lower slopes and wide CIs, and this relationship has been
considered as linear.
With the generalized estimating equation model, the respira-
tory symptoms’ OR for a 10 grain/m
3
increase in Poaceae is
significant (1.03 [1.01-1.05]). For nasal and ocular symptoms,
piecewise generalized estimating equation models quantified
significant nasal and ocular pollen-symptom relationships up
to the saturation point of 80 and 90 grains/m
3
(OR 5 1.06
[1.03-1.08] and 1.08 [1.05-1.12], respectively). Beyond these cut-
offs, ORs are close to 1 and not significant.
At the start of the season (ie, before peaks occurrence, days on
which pollen levels were below 10 grains/m
3
), the relationships
between pollens and nasal and ocular symptoms are globally
linear and ORs are high (OR 5 2.68 [1.23-5.82] and 1.86
[0.55-6.30], respectively), consistent with no or very low thresh-
old. For nasal symptoms, the relationship is higher in people with
perennial sensitization (OR 5 4.22 [1.39-12.81]) than in people
without perennial sensitization (OR 5 2.37 [0.85-6.55]).
In the literature, most authors assume linear relationships
between the number of subjects with symptoms in grass-
sensitized people and Poaceae levels. However, our results
show a sharp positive linear trend followed by a plateau. This is
consistent with the findings of a recent study (although the
outcome variable is different) from Erbas et al.
2
This study shows
in a time-series analysis that grass pollen has an increasing effect
on asthma hospital admissions up to a saturation point of 30
grains/m
3
, with this effect remaining stable thereafter. Tobias
et al
3
also reported nonlinear short-term effects of airborne grass
levels on asthma emergency room admissions in Madrid. In a
panel study performed in 1973, but that did not use adequate sta-
tistical analyses, Davies and Smith
4
suggested that the number of
grass pollens necessary to induce upper airway symptoms in most
patients was about 50 grains/m
3
(ie, plateau effect). In Spain,
Brito et al
5
suggested that threshold levels for the induction of
symptoms in all monosensitized patients to olive pollens was
162 grains/m
3
.
The higher level of nasal and ocular symptoms, without any
trigger threshold, observed in the whole population at the
beginning of pollination may be explained by a priming effect,
as described by Connell.
6
This means that during natural
exposure, decreasing amounts of pollen are required to elicit
symptoms as the season progresses, and the severity of allergic re-
sponse is therefore increased.
During this same pollination period, the stronger relationship
observed for nasal symptoms in people with animal or dust
sensitization, but without perennial symptoms, may be related to a
nonspecific prepriming phenomenon , as described by Ellis et al,
7
using an Environmental Exposure Unit. However, with provoca-
tion nasal tests, the response to repeated allergen challenge can
show large variations from a strongly enhanced response
8
to a
complete diminution of the response.
9
In conclusion, the clinical response in sensitized patients to
Poaceae varies during the grass season. Our data suggest
prepriming and priming effects at the beginning of the season
without any trigger threshold, a linear response up to a saturation
point, then a plateau.
Denis M. Caillaud, MD, PhD
a
Sylvie Martin, PharmD, MPH
b
Claire Segala, MD, MPH
b
Jean-Pierre Besancenot, PhD
c
Bernard Clot, PhD
d
Michel Thibaudon, PhD
c
on behalf of the French Aerobiology Network
From
a
the Pulmonary and Allergology Department, Gabriel Montpied Hospital, Cler-
mont-Ferrand, France;
b
Sepia-Sant
e, 56150, Baud, France;
c
the French Network of
Aerobiological Monitoring, RNSA, Brussieu, France; and
d
the Federal Office of Me-
teorology and Climatology MeteoSwiss, Payerne, Switzerland. E-mail: dcaillaud@
chu-clermontferrand.fr.
Supported by Agence Nationale de Securit
e Sanitaire: research convention no. EST
2008/1/21.
Disclosure of potential conflict of interest: The authors declare that they have no relevant
conflicts of interest.
REFERENCES
1. Bell ML, Samet JM, Dominici F. Time-series studies of particulate matter. Annu
Rev Public Health 2004;25:247-80.
2. Erbas B, Chang JH, Dharmage S, Ong EK, Hyndman R, Newbigin E, et al. Do levels
of airborne grass pollen influence asthma hospital admissions? Clin Exp Allergy
2007;37:1641-7.
3. Tobias A, Galan I, Banegas JR. Non-linear short-term effects of airborne pollen
levels with allergenic capacity on asthma emergency room admissions in Madrid,
Spain. Clin Exp Allergy 2004;34:871-8.
4. Davies RR, Smith LP. Forecasting the start and severity of the hay fever season. Clin
Allergy 1973;3:263-7.
5. Brito FF, Gimeno PM, Carnes J, Martin R, Fernandez-Caldas E, Lara P, et al. Olea
europaea pollen counts and aeroallergen levels predict clinical symptoms in patients
allergic to olive pollen. Ann Allergy Asthma Immunol 2011;106:146-52.
6. Connell JT. Quantitative intranasal pollen challenge, II: effect of daily pollen chal-
lenge, environmental pollen exposure, and placebo challenge on the nasal mem-
brane. J Allergy 1968;41:123-39.
7. Ellis AK, Ratz JD, Day AG, Day JH. Factors that affect the allergic rhinitis re-
sponse to ragweed allergen exposure. Ann Allergy Asthma Immunol 2010;104:
293-8.
8. de Bruin-Weller MS, Weller FR, De Monchy JG. Repeated allergen challenge as a
new research model for studying allergic reactions. Clin Exp Allergy 1999;29:
159-65.
9. Reinartz SM, van Ree R, Versteeg SA, Zuidmeer L, van Drunen CM, Fokkens WJ.
Diminished response to grass pollen allergen challenge in subjects with concurrent
house dust mite allergy. Rhinology 2009;47:192-8.
Available online June 15, 2012.
http://dx.doi.org/10.1016/j.jaci.2012.04.034
Autoimmune mechanisms in chronic idio-
pathic urticaria
To the Editor:
Chronic idiopathic urticaria (CIU) is defined as repeated
episodes of hives occurring over a period of 6 or more weeks
without an identifiable cause.
1
CIU is a common disorder with a
prevalence of 0.5% in the general population.
2
In the 1980s, the
presence of a serum factor that induced a wheal and flare response
following intradermal injection of autologous serum was
J ALLERGY CLIN IMMUNOL
SEPTEMBER 2012
814 LETTERS TO THE EDITOR
reported
3
and has since been regarded as a helpful surrogate for
autoimmunity in CIU.
1
IgG autoantibodies directed against the
a subunit of the high-affinity IgE receptor (FcεRI) are thought
to be the primary autoimmune mechanism of CIU, and these
are reportedly present in 30% to 60% of the patients with CIU.
4-7
We have developed an immortalized human mast cell line
(LUVA) displaying FcεRI that both degranulates and releases
arachidonic acid-metabolites in an FcεRI-dependent fashion.
8
These cells express high levels of chymase and tryptase
(CT)-positive and therefore display characteristics of connective
tissue mast cells. We investigated mast cell prostaglandin D
2
(PgD
2
) production and Ca
21
influxes by LUVA cells in
response to incubation with sera derived from patients with CIU.
Patients with CIU and matched control subjects were recruited
under a protocol approved by our human investigations commit-
tee.
9
Autologous serum skin tests were performed by using stan-
dard methodology.
1
Serum (diluted 1:1) was incubated with
LUVA cells (5 3 10
4
)at378C for 30 minutes, and activation
was determined via enzyme immunoassay as PgD
2
secretion
(sensitivity >55 pg/mL; Cayman Chemical Co, Ann Arbor,
Mich). To properly assess LUVA activation, we also measured
PgD
2
concentrations in comparably diluted serum samples and
subtracted this from the LUVA supernatant values. Additional
PgD
2
assays were performed after incubating LUVA cells over-
night with monoclonal IgE (100 ng/mL; Millipore, Temecula,
Calif) to upregulate FcεRI expression.
8
As an alternative measure
of mast cell activation by CIU serum, we investigated the induc-
tion of Ca
21
fluxes by using the Fluo-4NW calcium assay kit
(Molecular Devices, Sunnyvale, Calif). Briefly, loading buffer
with 5 mM probenecid (Sigma, St Louis, Mo) was added to
each well and incubated at 378C for 30 minutes. Cells were stim-
ulated with CIU serum and cytoplasmic [Ca
21
]i was determined
at an extinction wavelength of 494 nm and an emission
wavelength of 516 nm by using a Flexstation spectrophotometer
(Molecular Devices). ATP (300 mM) and monoclonal anti-FcεRI
antibodies (Upstate, Lake Placid, NY) were used as positive con-
trols, and data are expressed as arbitrary relative fluorescence
units. To evaluate the role of FcεR1-dependent and independent
pathways in cellular activation,
10-14
LUVA cells were incubated
with the syk inhibitor ER27319 (10-60 mM, Tocris, Ellisville,
Mo) or pertussis toxin (100 ng, Tocris) for 2 hours at 378C. Unless
otherwise specified, data are expressed as the mean 6 SD and the
differences analyzed by paired t test.
Fourteen patients with CIU were enrolled in the study.
Characteristics of patients enrolled are presented in this article’s
Online Repository at www.jacionline.org (Tables E1 and E2). Our
CIU population was predominantly composed of women with an
average age of 40 6 14 years, similar to what other investigators
have found.
15,16
In addition, the majority of our patients were var-
iously noted to have well-recognized markers of autoimmunity in
CIU, including anti-FcεRI antibodies (9 of 14), positive autolo-
gous serum skin test results (9 of 14), and/or antithyroid
antibodies (6 of 14).
As the most prominent activation pathway triggered by LUVA
mast cells,
8
we initially focused on PgD
2
secretion. We observed
increased de novo PgD
2
secretion after the incubation of CIU-
derived serum with LUVA cells in comparison to control serum
(363.3 6 125.9 pg/mL vs 243.5 6 94.18 pg/mL; P < .01; Fig 1).
PgD
2
secretion was modestly (18%) but consistently (14 of 14
subjects) increased after LUVA cells were incubated overnight
with IgE (768.0 6 177.3 pg/mL to 909.0 6 203.1 pg/mL;
P < .001) (see Fig E1 in this article’s Online Repository at
www.jacionline.org). To dissect the role of FcεRI-dependent
and -independent pathways in CIU, PgD
2
secretion using serum
samples obtained from the 6 subjects displaying the highest induc-
tion was measured after exposing LUVA cells to either a syk inhib-
itor or pertussis toxin, as well as after the overnight incubation of
serum with recombinant human FcεRI as described.
9
Preincuba-
tion with the syk inhibitor decreased PgD
2
production in all 6 sub-
jects (mean percent decrease was 27.3% 6 4.7%; P < .05) (Fig 2).
Similarly, partial inhibition of PgD
2
secretion was observed with
pertussis toxin (18.5% 6 3.9% decrease; P < .05). No synergy was
observed when the syk inhibitor and pertussis toxin were com-
bined (30% 6 8.17% decrease). Better inhibition of activation
was observed after overnight ‘capture’ of anti-FcεRI antibodies
FIG 1. PgD
2
Production after incubation with control- and CIU-derived se-
rum. Serum was incubated with LUVA cells at 378C for 30 minutes and se-
creted PgD
2
assayed by enzyme immunoassay. Data represent net
production of PgD
2
after subtracting out PgD
2
present in the applied serum
sample (P < .01).
FIG 2. Inhibition of PgD
2
secretion by Syk inhibitor and pertussis toxin.
LUVA mast cells were incubated with the syk inhibitor ER27319 (60
mM) or pertussis toxin (100 ng) for 2 hours at 378C. LUVA cells were sub-
sequently challenged with CIU patient sera and PgD
2
secretion
determined.
J ALLERGY CLIN IMMUNOL
VOLUME 130, NUMBER 3
LETTERS TO THE EDITOR 815
with excess recombinant human FcεRI (53% 6 28% [n 5 7]; data
not shown). We also established the ability of CIU serum to induce
Ca
21
fluxes (see Fig E2 in this article’s Online Repository www.
jacionline.org). Induction of Ca
21
influxes was consistently ob-
served across the entire cohort of subjects (P < .0001). As with
the PgD
2
data, pretreatment of LUVA cells with the syk inhibitor
produced consistent but incomplete inhibition of Ca
21
influx,
comparable to the extent of inhibition observed by using anti-
FcεRI antibodies (Fig E2).
Previously, basophil activation tests have been considered the
benchmark diagnostic test to identify autoimmune causes of
CIU.
17-19
However, this approach is limited by the inherent vari-
ability of basophils, which requires rigorous screening of baso-
phils from numerous donors to identify ‘releaser’ basophils
capable of FcεRI-dependent activation, and even among donors
displaying this phenotype, there is day-to-day variability in this
capacity.
20
Stringent screening is also required to ensure against
high spontaneous secretion of histamine. While basophils provide
useful diagnostic information, CIU is a disease targeting cutane-
ous, largely connective tissue, mast cells. As basophils are not
considered the primary cellular target of CIU, a mast cell model
would be the more ideal framework to study CIU pathophysiol-
ogy. The concept that CIU involves autoimmune mechanisms
that include anti-FcεRI antibodies is strongly supported by our
current studies. A putative contributing role for anti-FcεRI
antibodies is supported by the increase in PgD
2
secretion after
overnight incubation with IgE, an intervention that increases
cell-surface expression of these receptors and also as the inhibi-
tion of secretion after overnight ‘capture’ of putative anti-FcεRI
antibodies with excess recombinant FcεRI protein.
8,21
It should
be noted, however, that the upregulation of PgD
2
secretion in-
duced by IgE incubation cannot categorically distinguish
FcεRI-dependent effects from nonspecific priming of the LUVA
cells. In further support of a role for FcεRI-mediated cell activa-
tion, application of a syk inhibitor significantly reduced PgD
2
pro-
duction and Ca
21
fluxes. The extent of this inhibition of Ca
21
fluxes was comparable to that produced when cells were stimu-
lated by anti-FcεRI antibodies. Of note, however, pertussis toxin
also reduced PgD
2
production, presumably reflecting additional
complementary or collateral activation pathways.
11
This mast
cell line may also prove useful in dissecting additional mast
cell–targeting autoantibodies or other complementary pathways
such as those mediated by complement, bradykinin, or the clot-
ting cascade that might contribute to the development of CIU.
In summary, we utilized a unique connectiv e tissue mast cell line
to study the effects of CIU patient sera on activ ation, demonstrating
PgD
2
secretion and Ca
21
influxes. Upregulation after overnight incu-
bation with IgE and inhibition of PgD
2
secretion and Ca
21
fluxes af-
ter preincubation with a syk inhibitor or recombinant human FcεRI
points to a role for FcεRI-dependent autoimmun e mechanisms, al-
though additional complementary pathways appear to be in volv ed.
Future studies will be required to determine the relative validity of
diagnostic testing for autoimmune mechanisms in CIU using baso-
phil activation as opposed to assays based on what is ar guably the
more rele v ant cellular target, the connective tissue mast cell.
Jonathon Posthumus, MD
Adrienne Ti~nana, MD, MPH
Jonathan D. Mozena, MD
John W. Steinke, PhD
Larry Borish, MD
From the Asthma and Allergic Disease Center, Carter Center for Immunology Research,
University of Virginia, Charlottesville, Va. E-mail: lb4m@virginia.edu.
Disclosure of potential conflict of interest: J. W. Steinke has received research support
from the National Institutes of Health (NIH) and Medtronic. L. Borish has received
honoraria and research support from Merck, has consulted for Pfizer, and has received
research support from the NIH. The rest of the authors declare that they have no rel-
evant conflicts of interest.
REFERENCES
1. Sabroe RA, Grattan CE, Francis DM, Barr RM, Black AK, Greaves MW. The au-
tologous serum skin test: a screening test for auto-antibodies in chronic idiopathic
urticaria. Br J Dermatol 1999;140:446-52.
2. Charlesworth EN. Urticaria and angioedema. Allergy Asthma Proc 2002;23:
341-5.
3. Grattan CE, Wallington TB, Warin RP, Kennedy CTC, Bradfield JW. A serological
mediator in chronic idiopathic urticaria: a clinical immunological and histological
evaluation. Br J Dermatol 1986;114:583-90.
4. Hide M, Francis DM, Grattan CE, Hakimi J, Kochan JP, Greaves MW. Auto-anti-
bodies against the high affinity IgE receptor as a cause of histamine release in
chronic urticaria. N Engl J Med 1993;328:1599-604.
5. Fiebiger E, Maurer D, Holub H, Reininger B, Hartmann G, Weisetschlager M,
et al. Serum IgG auto-antibodies directed against the a chain of FcεRI: a selective
marker and pathogenetic factor for a distinct subset of chronic urticaria patients?
J Clin Investig 1995;96:2606-12.
6. Ferrer M, Kinet JP, Kaplan AP. Comparative studies of functional and binding as-
says for IgG anti-FcεRIa (a-subunit) in chronic urticaria. J Allergy Clin Immunol
1998;101:672-6.
7. Kaplan AP, Joseph K. Basophil secretion in chronic urticaria: autoantibody-
dependent or not? J Allergy Clin Immunol 2007;120:729-30.
8. Laidlaw TM, Steinke JW, Tinana AM, Feng C, Xing W, Lam BK, et al. Char-
acterization of a novel human mast cell line that responds to stem cell factor
and expresses functional FcepsilonRI. J Allergy Clin Immunol 2011;127:
815-22.e1-5.
9. Mozena JD, Tinana A, Negri J, Steinke JW, Borish L. Lack of a role for cross-
reacting anti-thyroid antibodies in chronic idiopathic urticaria. J Investig Dermatol
2010;130:1860-5.
10. Saito H, Okajima F, Molski TF, Sha’afi RI, Ui M, Ishizaka T. Effects of ADP-
ribosylation of GTP-binding protein by pertussis toxin on immunoglobulin E-de-
pendent and -independent histamine release from mast cells and basophils.
J Immunol 1987;138:3927-34.
11. Oliver JM, Kepley CL, Ortega E, Wilson BS. Immunologically mediated signaling
in basophils and mast cells: finding therapeutic targets for allergic diseases in the
human FcvarepsilonR1 signaling pathway. Immunopharmacology 2000;48:269-81.
12. Kepley CL, Youssef L, Andrews RP, Wilson BS, Oliver JM. Multiple defects in
Fc epsilon RI signaling in Syk-deficient nonreleaser basophils and IL-3-induced
recovery of Syk expression and secretion. J Immunol 2000;165:5913-20.
13. MacGlashan D Jr. Two regions of down-regulation in the IgE-mediated signaling
pathway in human basophils. J Immunol 2003;170:4914-25.
14. Macglashan D, Miura K. Loss of syk kinase during IgE-mediated stimulation of
human basophils. J Allergy Clin Immunol 2004;114:1317-24.
15. O’Donnell BF, Francis DM, Swana GT, Seed PT, Kobza Black A, Greaves MW.
Thyroid autoimmunity in chronic urticaria. Br J Dermatol 2005;153:331-5.
16. Kulthanan K, Jiamton S, Gorvanich T, Pinkaew S. Autologous serum skin test in
chronic idiopathic urticaria: prevalence, correlation and clinical implications.
Asian Pac J Allergy Immunol 2006;24:201-6.
17. Greaves MW, Tan KT. Chronic urticaria: recent advances. Clin Rev Allergy Immu-
nol 2007;33:134-43.
18. Yasnowsky KM, Dreskin SC, Efaw B, Schoen D, Vedanthan PK, Alam R, et al.
Chronic urticaria sera increase basophil CD203c expression. J Allergy Clin Immu-
nol 2006;117:1430-4.
19. Altrich ML, Halsey JF, Altman LC. Comparison of the in vivo autologous skin test
with in vitro diagnostic tests for diagnosis of chronic autoimmune urtic aria. Allergy
Asthma Proc 2009;30:28-34.
20. Lichtenstein LM, MacGlashan DW Jr. The concept of basophil releasability.
J Allergy Clin Immu nol 1986;77:291-4.
21. Yamaguchi M, Lantz CS, Oettgen HC, Katona IM, Fleming T, Miyajima I, et al.
IgE enhances mouse mast cell Fc(epsilon)RI expression in vitro and in vivo: evi-
dence for a novel amplification mechanism in IgE-dependent reactions. J Exp
Med 1997;185:663-72.Appendix
Available online June 12, 2012.
http://dx.doi.org/10.1016/j.jaci.2012.04.037
J ALLERGY CLIN IMMUNOL
SEPTEMBER 2012
816 LETTERS TO THE EDITOR
FIG E1. Upregulation of PgD
2
secretion after incubation with IgE. PgD
2
as-
says were repeated after incubating LUVA cells with monoclonal IgE (100
ng/mL) overnight to upregulate FcεRI expression (*P < .0001).
J ALLERGY CLIN IMMUNOL
SEPTEMBER 2012
816.e1 LETTERS TO THE EDITOR
FIG E2. Induction of Ca
21
fluxes after incubation with CIU serum. Mast cell
activation by CIU-derived serum was determined by using the Fluo-4NW
calcium assay as described in the text. ATP (300 mM) and monoclonal
anti-FcεRI antibodies were used as positive controls. Varying concentra-
tions of syk inhibitor were utilized to determine optimal concentration.
Each condition was performed in triplicate, and the average of the 3 mea-
surements is presented. Data are expressed as arbitrary relative fluores-
cence units (RFUs). inh, Inhibitor.
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VOLUME 130, NUMBER 3
LETTERS TO THE EDITOR 816.e2
TABLE E1. Characteristics of CIU population
Patient Age (y) Sex ASST* Anti-FcεRI antibodiesy Thyroid antibodiesz
1 61 F Positive Positive Negative
2 56 F Negative Positive Positive
3 28 F Positive Positive Positive
4 65 M Positive Positive Positive
5 35 F Positive Positive Negative
6 47 M Negative Positive Negative
7 61 F Negative Negative Positive
8 46 F Positive Negative Negative
9 21 F Positive Negative Positive
10 29 M Negative Negative Negative
11 40 F Positive Positive Positive
12 27 F Positive Positive Negative
13 27 F Positive Positive Negative
14 37 F Negative Negative Negative
ASST, Autologous serum skin test; F, female; M, male.
*Positive ASST result defined as wheal more than 5 mm in diameter more than saline controls.
Positive anti-FcεRI titer defined as more than 3.9 mg/mL.
9
àPositive thyroid antibodies defined as a positive thyroglobulin and/or thyroid peroxidase antibody titer.
J ALLERGY CLIN IMMUNOL
SEPTEMBER 2012
816.e3 LETTERS TO THE EDITOR
TABLE E2. Summary of characteristics of the CIU population
Characteristic n 5 14
Age (y), mean 6 SD 41 6 14
Sex (number of females) 11/14
ASST (number positive)* 9/14
Anti-FcεRI antibodies (number positive) 9/14
Thyroid antibodies (number positive)à 6/14
ASST, Autologous serum skin test; EIA, enzyme immunoassay.
*Positive ASST result defined as wheal more than 5 mm in diameter more than saline
controls.
Positive anti-FcεRI titer defined as more than 3.9 mg/mL via EIA as described.
9
àPositive thyroid antibodies defined as a positive thyroglobulin and/or thyroid
peroxidase antibody titer.
J ALLERGY CLIN IMMUNOL
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LETTERS TO THE EDITOR 816.e4
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