ARTHRITIS & RHEUMATISM

Vol. 52, No. 9, September 2005, pp 29262935

DOI 10.1002/art.21250
2005, American College of Rheumatology

Prevalence and Clinical Significance of

Antineutrophil Cytoplasmic Antibodies in
Churg-Strauss Syndrome
Renato A. Sinico,1 Lucafrancesco Di Toma,1 Umberto Maggiore,2 Paolo Bottero,3
Antonella Radice,1 Cinzia Tosoni,4 Chiara Grasselli,2 Laura Pavone,2 Gina Gregorini,4
Stefano Monti,4 Micol Frassi,4 Filomena Vecchio,3 Caterina Corace,1
Emanuela Venegoni,3 and Carlo Buzio2
specificity for proteinase 3, was found in 3 of 35 patients
(8.6%). Atypical patterns were found in 6 of 30 patients
with anti-MPO antibodies (20.0%). ANCA positivity was
associated with higher prevalences of renal disease
(51.4% versus 12.1%; P < 0.001) and pulmonary hemorrhage (20.0% versus 0.0%; P 0.001) and, to a lesser
extent, with other organ system manifestations (purpura and mononeuritis multiplex), but with lower frequencies of lung disease (34.3% versus 60.3%; P
0.019) and heart disease (5.7% versus 22.4%; P 0.042).
Conclusion. ANCAs are present in 40% of patients with CSS. A pANCA pattern with specificity for
MPO is found in most ANCA-positive patients. ANCA
positivity is mainly associated with glomerular and
alveolar capillaritis.

Objective. Churg-Strauss syndrome (CSS) is classified among the so-called antineutrophil cytoplasmic
antibodyassociated systemic vasculitides (AASVs) because of its clinicopathologic features that overlap with
the other AASVs. However, while antineutrophil cytoplasmic antibodies (ANCAs) are consistently found in
7595% of patients with Wegeners granulomatosis or
microscopic polyangiitis, their prevalence in CSS varies
widely and their clinical significance remains uncertain.
We undertook this study to examine the prevalence and
antigen specificity of ANCAs in a large cohort of patients with CSS. Moreover, we evaluated the relationship
between ANCA positivity and clinicopathologic features.
Methods. Immunofluorescence and enzymelinked immunosorbent assay were used to determine the
presence or absence of ANCAs in 93 consecutive patients at the time of diagnosis. The main clinical and
pathologic data, obtained by retrospective analysis, were
correlated with ANCA status.
Results. ANCAs were present by immunofluorescence in 35 of 93 patients (37.6%). A perinuclear ANCA
(pANCA) pattern was found in 26 of 35 patients
(74.3%), with specificity for myeloperoxidase (MPO) in
24 patients, while a cytoplasmic ANCA pattern, with

Churg-Strauss syndrome (CSS) is defined as an

eosinophil-rich and granulomatous inflammation involving the respiratory tract, coupled with necrotizing vasculitis affecting small to medium-size vessels, and associated with asthma and eosinophilia (1,2). CSS is a rare
disorder with an incidence of 1.36.8 cases per 1,000,000
patients per year and an overall prevalence of 10.713
per 1,000,000 adults (35).
Three different phases can usually be recognized
in CSS. Asthma and atopic allergies such as rhinitis may
precede by months, and sometimes by several years, the
development of an eosinophilic infiltrative disease with
eosinophilic pneumonia or gastroenteritis, followed by
the vasculitic phase (69). Nasal polyposis, sinusitis, and
nonfixed pulmonary infiltrates are seen in the majority
of patients. Vasculitis commonly affects the skin, peripheral nerves, gastrointestinal tract, and heart. Coronary
arteritis and myocarditis are the main causes of morbidity
and mortality (1,69). Pathologic confirmation is based on

1
Renato A. Sinico, MD, Lucafrancesco Di Toma, MD, Antonella Radice, PhD, Caterina Corace, PhD: Azienda Ospedaliera
Ospedale San Carlo Borromeo, Milan, Italy; 2Umberto Maggiore,
MD, Chiara Grasselli, MD, Laura Pavone, MD, Carlo Buzio, MD:
Ospedale Maggiore, Parma, Italy; 3Paolo Bottero, MD, Filomena
Vecchio, MD, Emanuela Venegoni, MD: Ospedale di Magenta,
Magenta, Italy; 4Cinzia Tosoni, MD, Gina Gregorini, MD, Stefano
Monti, MD, Micol Frassi, MD: Spedali Civili, Brescia, Italy.
Address correspondence and reprint requests to Renato A.
Sinico, MD, Dipartimento di Nefrologia e Immunologia, Azienda
Ospedaliera Ospedale San Carlo Borromeo, Via Pio Secondo, 3-20153
Milan, Italy. E-mail: renato.sinico@oscb.sined.net.
Submitted for publication December 29, 2004; accepted in
revised form May 24, 2005.

2926

ANCAs IN CHURG-STRAUSS SYNDROME

the presence of extravascular granulomas in association

with necrosis and predominant extravascular eosinophils,
as well as necrotizing vasculitis (1,2), but all of these lesions
are rarely found together in biopsy specimens (6). The vast
majority of patients have blood eosinophilia (1,500 cells/
mm3 or 10%), and the exceptions are those previously
treated with glucocorticoids for asthma (79).
CSS is considered to be one of the so-called
antineutrophil cytoplasmic antibodyassociated systemic
vasculitides (AASVs) because of its clinical and pathologic features that overlap with those of the other
AASVs, Wegeners granulomatosis (WG) and microscopic polyangiitis (MPA) (10,11). However, these 2
diseases differ from CSS clinically, by the absence of
asthma, and pathologically, by the absence of eosinophilia and eosinophil-rich tissue infiltrates (912). Moreover, while antineutrophil cytoplasmic antibodies
(ANCAs) are consistently found in 7595% of patients
with active WG or MPA (10,11), the prevalence of
ANCAs in CSS has been reported to be variable, ranging
from 40% to 70% (7,8,1215). This is probably due
to the small number of patients studied in most series, to
selection bias, to the different classification criteria used
for CSS, and to the various methods used to test for
ANCA positivity (for review, see ref. 8). The immunofluorescence pattern is usually perinuclear (pANCA)
with specificity for myeloperoxidase (MPO) by enzymelinked immunosorbent assay (ELISA). Only a minority
of patients have cytoplasmic ANCAs (cANCA) with
antibodies to proteinase 3 (PR3) (7,14,15). Data are
scarce concerning the clinical significance, if any, of
ANCA positivity.
The aim of the present study was to examine the
prevalence, fluorescence patterns, and antigen specificity of ANCAs in a large cohort of consecutive unselected
patients with CSS. Moreover, we examined the relationship between ANCA positivity and clinicopathologic
features and long-term outcome.
PATIENTS AND METHODS
Patients. We identified 106 unselected consecutive
patients in whom CSS was diagnosed clinically at internal
medicine departments (nephrology, clinical immunology and
rheumatology, pulmonology, neurology, and others) in 4 general hospitals in northern Italy between 1985 and 2004. Ninetythree patients, in whom CSS was diagnosed between 1989 and
2004, were tested for the presence of ANCAs at the time of
diagnosis and were therefore eligible for the present study. The
clinical characteristics of the 13 excluded patients were similar
to those of the included patients (data not shown). CSS was
diagnosed, as described by Guillevin et al (7), when asthma,
hypereosinophilia (10% or 1,500 cells/mm3), and clinical
manifestations consistent with systemic vasculitis, with or

2927

without histologic evidence, were present. The following exceptions were allowed: 1) hypereosinophilia was not considered mandatory in the case of a few patients previously treated
with oral steroids for asthma in the presence of histologic
confirmation (vasculitis plus extravascular eosinophils); 2) the
absence of asthma was not considered an exclusion criterion in
the case of 4 patients with hypereosinophilia (1,500 cells/
mm3) and histologic confirmation. We examined the medical
records of all these patients. CSS was defined according to the
Chapel Hill Consensus Conference nomenclature (2). The
classification criteria for CSS of the American College of
Rheumatology (ACR) (16) as well as Lanhams (Hammersmith) criteria (6) were retrospectively applied to the study
population. All patients had direct (histologic) or indirect
(using surrogate markers) evidence of vasculitis (14).
Organ system involvement was assessed using the
Birmingham Vasculitis Activity Score (BVAS) item list (17).
Clinical manifestations were usually confirmed by instrumental
examinations as appropriate and/or tissue biopsy. Unusual
manifestations not included in BVAS items (e.g., cholecystitis
and genitourinary involvement) were also considered.
Definitions and pathologic studies. A patients disease
was considered to be in full remission when there was a
complete absence of clinical disease activity for at least 6
months according to the BVAS item list, with the exception of
asthma or neurologic and renal sequelae due to scars (7). A
relapse was defined as the occurrence or recurrence of a clinical
manifestation attributable to CSS. Persistent asthma or an isolated increase of eosinophilia was not considered a relapse (7).
One or more biopsy specimens from affected tissues
were obtained when considered necessary by the clinicians. A
biopsy specimen was considered consistent with a diagnosis of
CSS when eosinophilic tissue infiltration and/or vasculitis were
found (9). The Disease Extent Index and the prognostic
Five-Factors Score (FFS) were also calculated as described by
their respective authors (18,19).
Biochemical and serologic markers. Routine laboratory tests were performed in all cases at the time of diagnosis
and at followup. The presence of ANCAs was determined in all
patients at the time of diagnosis, before starting immunosuppressive treatment, using indirect immunofluorescence on
ethanol-fixed granulocytes and antigen-specific PR3 and MPO
ELISAs, as previously described (2022). Antigen-specific
ELISAs were performed on all serum samples, including those
that were ANCA negative by immunofluorescence. Because
PR3 and MPO ELISAs were not available in 2 laboratories
until 1995, these assays were performed on stored serum
samples in 6 cases. ANCA positivity was also tested at the end
of the followup period in 37 patients and at the time of a
relapse of disease in 16 patients. ANCA serology was tested
first in each hospital and rechecked centrally, on frozen serum
samples, in a laboratory that participated in the EC/BCR study
for ANCA assay standardization (2325). The staining patterns
were defined according to the nomenclature of the International Consensus Statement, as follows: cANCA cytoplasmic
fluorescence with interlobular accentuation; cANCA atypical other types of cytoplasmic fluorescence (e.g., homogeneous); pANCA perinuclear or granulocyte-specific nuclear
fluorescence; atypical ANCAs other, less common patterns,
such as mixed cytoplasmic and perinuclear fluorescence (26).
Perinuclear ANCA and atypical ANCApositive serum samples were also tested in the central laboratory on

2928

SINICO ET AL

Table 1. Main clinical features of the 93 patients with Churg-Strauss

syndrome*
No. (%) of patients
Asthma
Constitutional symptoms
Sinusitis
Skin involvement
Lung involvement
Heart involvement
Gastrointestinal involvement
Peripheral neuropathy
CNS involvement
Renal involvement

89 (95.7)
63 (67.7)
72 (77.4)
49 (52.7)
47 (50.5)
15 (16.1)
20 (21.5)
60 (64.5)
13 (14.0)
25 (26.9)

* CNS central nervous system.

formalin-fixed neutrophils, as previously described, to confirm

that they truly contained ANCAs and not, for example,
antinuclear antibodies (22). Antigen specificity was also confirmed by inhibition study in selected cases as previously
described (20).
Treatment. All patients were treated with corticosteroids (1 mg/kg/day for 34 weeks with subsequent tapering),
preceded by methylprednisolone pulses in the case of 22
patients. Cyclophosphamide (daily oral or pulses), as induction
treatment, was added for 36 months in 42 patients (45.2%),
usually those with the most severe disease (27). Other induction treatment consisted of methotrexate (5 patients) and
azathioprine (1 patient). Plasma exchange and intravenous
immunoglobulin were added in 4 patients and in 1 patient,
respectively. Maintenance treatment consisted of low doses of
glucocorticoids (512.5 mg/day) in all patients plus immunosuppressive drugs in 33 patients (35.5%) (azathioprine in 14,
methotrexate in 12, and cyclosporin A in 7). Long-term
followup and outcome were established after the patients last
visit or death.
Statistical analysis. All analyses were performed using
Stata statistical software, release 8.2 (Stata Corporation, Col-

lege Station, TX). The differences between ANCA-positive

and ANCA-negative patients in continuous variables were
tested by the Mann-Whitney U test and in categorical variables
by Fishers exact test. We also computed the prevalence ratio
and the prevalence difference of organ involvement, together
with their associated 95% confidence intervals, using the Stata
program epitab (28,29). The cumulative risk of relapse and
death was estimated by the life-table (actuarial) method since
survival curves were calculated at yearly intervals, and therefore individual times at which the events (or censoring)
occurred were not precisely known. The difference between
the risk curves was tested by the likelihood ratio test. These
latter computations were performed using the Stata program
ltable (29). All reported P values are 2-sided. P values less than
0.05 were considered significant.

RESULTS
Clinical and histologic characteristics. There were
39 male patients and 54 female patients with a mean age of
51.6 years (median 51 years, range 1886 years). All but 4
patients had bronchial asthma, which usually preceded
the diagnosis of CSS. Seven patients had been receiving
low doses of oral corticosteroids (15 mg/day of prednisone or equivalent) for asthma for a few weeks at the
time of diagnosis. Eosinophilia (10%) was present in
88 patients (94.6%), and the 5 patients without eosinophilia had received previous steroid treatment for
asthma. Patients had a median of 4,400 eosinophils/mm3
(range 60028,815). Of the 93 patients with CSS, 85
(91.4%) met the ACR classification criteria for CSS;
Lanhams criteria were met by 77 patients (82.8%), and
90 patients (96.8%) met at least 1 of the 2 criteria. The
main clinical features of these patients are summarized
in Table 1. One or more tissue biopsies were performed

Table 2. ANCA assay results in Churg-Strauss syndrome*

Indirect immunofluorescence
Author, year (ref.)

No. of
patients

Total
ANCAs

cANCA

Present study
Keogh and Specks, 2003 (14)
Della Rossa et al, 2002 (13)
Solans et al, 2001 (15)
Guillevin et al, 1999 (7)
Reid et al, 1998 (12)
Schnabel et al, 1996 (33)

93
30
18
18
42
17
17

35 (37.6)
22 (73.3)
NR
14 (77.8)
20 (47.6)
10 (58.8)
7 (41.2)

3/35 (8.6)
1/22 (4.5)
NR
1/14 (7.1)
1/20 (5.0)
0/10 (0.0)
5/7 (71.4)

ELISA

pANCA

Not classifiable
ANCAs

Anti-PR3 ANCAs

Anti-MPO ANCAs

26/35 (74.3)
21/22 (95.5)
NR
13/14 (92.9)
15/20 (75.0)
0/10 (0.0)
2/7 (28.6)

6/35 (17.1)
0/22 (0.0)
NR
0/14 (0.0)
4/20 (20.0)
10/10 (100.0)
0/7 (0.0)

3/93 (3.2)
NR
0/18 (0.0)
1/14 (7.1)
0/11 (0.0)
NR
5/17 (29.4)

30/93 (32.3)
28/37 (75.7)
7/18 (38.9)
13/14 (92.9)
10/11 (90.9)
5/?
2/17 (11.8)

* Values are the number (%) of antineutrophil cytoplasmic antibody (ANCA)positive patients, the number of patients with a given type of
ANCA/total number of ANCA-positive patients (%) within a given series in the case of immunofluorescence, or the number of ANCA-positive
patients by enzyme-linked immunosorbent assay (ELISA)/total number of patients tested (%) in the case of ELISA. In the case of ELISA,
denominators may differ from numbers of patients due to differences between studies in methods of reporting antibody positivity. Only series
describing at least 15 patients are shown. cANCA cytoplasmic ANCA; pANCA perinuclear ANCA; PR3 proteinase 3; MPO
myeloperoxidase; NR not reported.
Atypical patterns.
Pooled data from patients tested at the time of diagnosis and at the time of relapse.
The number of patients tested for anti-MPO antibodies was not stated.

ANCAs IN CHURG-STRAUSS SYNDROME

2929

Figure 1. Patterns of antineutrophil cytoplasmic antibodies (ANCAs). A, Cytoplasmic ANCA pattern on ethanol-fixed neutrophils obtained from
a patient with Wegeners granulomatosis and antiproteinase 3 antibodies. B, Perinuclear ANCA pattern on ethanol-fixed neutrophils obtained from
a patient with microscopic polyangiitis and antimyeloperoxidase (anti-MPO) antibodies. C, Atypical ANCA pattern (perinuclear cytoplasmic) on
ethanol-fixed neutrophils obtained from a patient with Churg-Strauss syndrome (CSS) and anti-MPO antibodies. D, Cytoplasmic ANCA atypical
pattern on ethanol-fixed neutrophils obtained from a patient with CSS and anti-MPO antibodies. See Patients and Methods for descriptions of
staining patterns. (Original magnification 400.)

in 63 patients (67.7%). Biopsy samples were suggestive

of CSS in 57 patients. Six biopsy samples were negative
or aspecific.
Prevalence of ANCAs and ANCA antigen specificity. ANCAs were present by immunofluorescence in
35 of 93 patients (37.6%) tested at the time of diagnosis
(Table 2). Of the 7 patients treated with oral corticosteroids at the time of ANCA testing, 3 (42.9%) were
ANCA positive. The prevalence of ANCA positivity
varied largely according to the specialty unit of origin,
ranging from 0% to 12.5% in patients from pulmonology
units to 90100% in patients from nephrology units. A

pANCA pattern was found in 26 of 35 patients (74.3%),

with specificity for MPO in 24 patients, while a cANCA
pattern with specificity for PR3 was found in 3 of 35
patients (8.6%). Two pANCA-positive samples were
negative by ELISA. Interestingly, a cANCA pattern and
an atypical ANCA pattern (Figure 1) were found in 3
patients (8.6%) and in 3 patients (8.6%), respectively,
with anti-MPO antibodies by ELISA (Table 3). When
the immunofluorescence assay was repeated in the central laboratory, the 3 cANCA-positive sera (with antiMPO specificity) were defined as cANCA atypical due
to the absence of interlobular accentuation, usually seen

2930

SINICO ET AL

Table 3. ANCA assay results in 6 patients showing a cANCA or atypical ANCA pattern and anti-MPO antibodies*
Indirect immunofluorescence pattern
Ethanol fixed
Patient
1
2
3
4
5
6

ELISA, arbitrary units

Original laboratory

Central laboratory
(home-made)

Central laboratory
(commercial kit)

Formalin
fixed

PR3
ANCAs

MPO
ANCAs

cANCA
Atypical (C P)
Atypical (C P)
cANCA
Atypical (C P)
cANCA

cANCA atypical
cANCA atypical
Atypical (C P)
Atypical (C P)
Atypical (C P)
cANCA atypical

cANCA atypical
cANCA atypical
Atypical (C P)
Atypical (C P)
Atypical (C P)
cANCA atypical

cANCA
cANCA
cANCA
cANCA
cANCA
cANCA

6
6
6
8
10
11

100
62
163
107
129
200

* cANCA pattern cytoplasmic fluorescence with interlobular accentuation; cANCA atypical pattern other types of cytoplasmic fluorescence;
atypical (C P) pattern other, less common patterns, such as mixed cytoplasmic and perinuclear fluorescence (see Table 2 for other definitions).
PR3 ANCAs anti-PR3 antibodies (normal values 20, standard curve range 6400); MPO ANCAs anti-MPO antibodies (normal values 20,
standard curve range 5320).

with PR3 ANCAs (Figure 1). The same patterns were

also demonstrated using commercially available kits
(Inova; Menarini Diagnostics, Florence, Italy) as well as
home-made granulocyte preparations (Table 3).
Inhibition studies confirmed that the cANCA and
atypical ANCA patterns were caused by antibodies to
MPO. Preincubation with MPO, but not with an irrelevant protein, could inhibit the binding both in ELISA
and in immunofluorescence. All pANCA and atypical

Table 4.

ANCA serum samples showed a cANCA pattern when

tested on formalin-fixed neutrophils (Table 3).
Twenty-five consecutive unselected serum samples, with various levels of anti-MPO antibodies, from
patients with MPA and WG were also retested by
immunofluorescence to exclude a casual finding (due to
a particular batch of slides). None of these samples
showed a cANCA or an atypical ANCA fluorescence
pattern.

Clinical features in ANCA-positive and ANCA-negative patients*

Asthma
Constitutional symptoms
Sinusitis
Skin involvement
Purpura
Lung involvement, all kinds
Pulmonary hemorrhage
Heart involvement
Gastrointestinal involvement
Peripheral neuropathy, all kinds
Mononeuritis multiplex
CNS involvement
Renal involvement
RPGN
ACR criteria
Lanhams criteria
Eosinophilia 10%
Eosinophils/mm3, median (range)
BVAS, 063, median (range)
DEI, 021, median (range)
VDI, 011, median (range)
FFS 2

ANCA positive
(n 35)

ANCA negative
(n 58)

34 (97.1)
30 (85.7)
27 (77.1)
21 (60.0)
9 (25.7)
12 (34.3)
7 (20.0)
2 (5.7)
7 (20.0)
25 (71.4)
18 (51.4)
6 (17.1)
18 (51.4)
10 (28.6)
30 (85.7)
30 (85.7)
32 (91.4)
4,881 (1,07428,815)
22 (740)
6 (310)
0 (02)
9 (25.7)

55 (94.8)
33 (56.9)
45 (77.6)
28 (48.3)
4 (6.9)
35 (60.3)
0 (0.0)
13 (22.4)
13 (22.4)
35 (60.3)
14 (24.1)
7 (12.1)
7 (12.1)
3 (5.2)
55 (94.8)
47 (81.0)
56 (96.6)
3,544 (60025,637)
17 (640)
6 (310)
0 (05)
7 (12.1)

1.00
0.006
1.00
0.29
0.015
0.019
0.001
0.042
1.00
0.37
0.013
0.54
0.001
0.004
0.15
0.78
0.36
0.51
0.15
0.85
0.30
0.15

* Except where indicated otherwise, values are the number (%) of patients. ANCA antineutrophil
cytoplasmic antibody; CNS central nervous system; RPGN rapidly progressive glomerulonephritis;
ACR American College of Rheumatology; BVAS Birmingham Vasculitis Activity Score; DEI
Disease Extent Index; VDI Vasculitis Damage Index; FFS Five-Factors Score.
Fishers exact test for categorical variables and Mann-Whitney U test for continuous variables.

ANCAs IN CHURG-STRAUSS SYNDROME

2931

Figure 2. Ratios of and differences in prevalence of organ involvement in antineutrophil cytoplasmic antibody (ANCA)positive and ANCAnegative patients. Left, Ratio of the prevalence of a given type of organ involvement in ANCA-positive patients to the corresponding prevalence in
ANCA-negative patients. A prevalence ratio of 2 means that ANCA-positive patients are twice as likely to be affected as ANCA-negative patients,
while a prevalence ratio of 0.5 means that ANCA-positive patients are half as likely to be affected as ANCA-negative patients. Right, Difference
in the prevalence of a given type of organ involvement in ANCA-positive patients from the corresponding prevalence in ANCA-negative patients.
A prevalence difference of 20% means that the prevalence of a given type of organ involvement in ANCA-positive patients is 20 percentage points
higher than the corresponding prevalence in ANCA-negative patients. Solid circles represent prevalence ratios or prevalence differences. Horizontal
bars represent 95% confidence intervals. Arrows indicate confidence interval upper bounds extending beyond the limits of the plot. The prevalence
ratio for pulmonary hemorrhage, which cannot be estimated since this did not occur in any ANCA-negative patient, is artificially plotted as an open
circle. Vertical dotted lines represent the null value, i.e., the number corresponding to identical prevalence of organ involvement in ANCA-positive
and ANCA-negative patients, which is 1 for the prevalence ratio and 0 for the prevalence difference. CNS central nervous system; RPGN
rapidly progressive glomerulonephritis.

Clinical significance of ANCA status. ANCA

positivity was associated with a significantly higher prevalence of renal involvement (51.4% versus 12.1% in
ANCA-negative patients; P 0.001) and, in particular,
with a clinical picture of rapidly progressive glomerulonephritis (28.6% versus 5.2%; P 0.004) (Table 4 and
Figure 2). Worth noting is the fact that all 11 patients
with a histologic picture of pauciimmune necrotizing
crescentic glomerulonephritis were ANCA positive
(data not shown). Moreover, ANCA positivity correlated with constitutional symptoms (85.7% versus
56.9%; P 0.006) and with certain organ system man-

ifestations, such as mononeuritis multiplex (51.4% versus 24.1%; P 0.013), purpura (25.7% versus 6.9%; P
0.015), and pulmonary hemorrhage (20.0% versus 0.0%;
P 0.001). In contrast, ANCA positivity was associated
with lower prevalences of lung involvement (with the
exception of alveolar hemorrhage) (34.3% versus 60.3%;
P 0.019) and heart involvement (5.7% versus 22.4%; P
0.042). ANCA-positive patients tended to have a
higher BVAS, but the difference was not statistically
significant. Other clinical and serologic parameters did
not differ between ANCA-positive and ANCA-negative
patients (Table 4 and Figure 2). The interval between

2932

SINICO ET AL

onset of asthma and diagnosis was shorter in ANCApositive patients, but the difference was not statistically
significant (median 3.5 years, range 047 years versus
median 7.5 years, range 053 years; P 0.077).
Among the 63 patients in whom at least 1 tissue
biopsy was performed, 57 had pathologic alterations
compatible with a diagnosis of CSS. The most frequent
histologic finding in ANCA-negative patients was eosinophilic tissue infiltration (58.8% versus 13.8%; P
0.001), usually with a perivascular pattern, whereas a
(necrotizing) small-vessel vasculitis and/or capillaritis
(including necrotizing crescentic glomerulonephritis)
was more commonly found in ANCA-positive patients
(75.9% versus 32.4%; P 0.001).
An FFS 2 was found more frequently in
ANCA-positive patients, even though the difference was
not statistically significant (Table 4). Moreover, we did
not find statistical evidence that ANCA positivity carried
a worse prognosis. In fact, the 5-year survival rate was
91.8% for ANCA-positive patients compared with
97.1% for ANCA-negative patients (P 0.74), whereas
the risks of relapse at 5 years were 46.3% and 35.4%,
respectively (P 0.20). It should be noted, however, that
ANCA-positive patients were more likely to be treated
with cyclophosphamide (65.7% versus 32.7%; P
0.003). Patients with anti-MPO antibodies and atypical
patterns by immunofluorescence did not differ from
other ANCA-positive patients.
ANCAs were present in only 4 of 37 patients
(10.8%) tested at the end of the followup period;
however, 1 of these patients still had active disease and
died of a complication of an infection. ANCAs were
present in 3 of 16 patients (18.8%) tested at the time of
a relapse.
DISCUSSION
The present study shows that 40% of patients
with CSS are ANCA positive. Although a classic
pANCA pattern with specificity for MPO is found in
most patients, atypical patterns are frequently encountered, including a cANCA pattern with anti-MPO antibodies. ANCA-positive patients are more likely than
ANCA-negative patients to present with the typical
clinicopathologic picture of the other small-vessel vasculitides and are less likely to have heart and nonhemorrhagic lung involvement.
ANCAs were first detected in sera from patients
with necrotizing glomerulonephritis and systemic vasculitis (30). Later, these autoantibodies were found in a
high proportion of patients with WG or MPA, including
their renal-limited form (idiopathic pauciimmune cres-

centic necrotizing glomerulonephritis) (31,32). CSS is

usually classified among the so-called AASVs (10,11);
however, data on the prevalence and clinical significance
of ANCAs in CSS are scarce and conflicting (8).
Considering the studies with at least 15 patients,
ANCAs were found by immunofluorescence in 41.2
77.7% of patients and in 73 of 124 patients (58.9%)
reported overall in these studies (7,12,14,15,33). We
found ANCAs in 40% of our population, which is
approximately in the range previously reported by some
investigators (38.947.6%) (7,13,33), but considerably
below the range found by others (58.877.7%)
(12,14,15). However, in the latter 3 studies (12,14,15),
only 17, 30, and 18 patients, respectively, were tested for
ANCAs. In our experience, ANCA positivity in CSS
depends very much on the specialty department of
origin, which in turn reflects the pattern of organ system
involvement of patients. This might explain, at least in
part, some discrepancies. Worth noting is the fact that a
recent, large, multicenter study of 100 patients with
CSS, reported only in abstract form, has found ANCA
positivity in 38% of patients (34), which is exactly the
prevalence we found in our population. We confirm that
pANCA with specificity for MPO are found in approximately three-fourths (or more) of ANCA-positive patients. However, in addition to the classical MPO
pANCA and PR3 cANCA patterns, we also found
cANCA and atypical ANCA patterns due to anti-MPO
antibodies (35,36).
In order to exclude a casual finding and/or an
artifact due to the fixation procedure that we used,
ANCA testing was repeated in a central laboratory with
home-made and commercial kits. Moreover, we tested
a similar number of consecutive serum samples with
anti-MPO antibodies from patients with MPA or WG.
The MPO cANCA pattern is reported very rarely in
MPA and WG, and it has been postulated that this
particular staining pattern might be due to the different
epitopes recognized by these antibodies (35). However,
it has also been reported that atypical staining patterns
may be found more frequently with some commercially
available ethanol-fixed neutrophil substrates, including
the one used by ourselves in the reference laboratory
(37,38). It has been speculated that this phenomenon
might be caused by factors in the ethanol fixation
conditions of these slides resulting in the differential
redistribution of different MPO epitopes (38).
In our series of patients, we found that ANCA
positivity was correlated with renal involvement, especially with the histologic picture of necrotizing crescentic
glomerulonephritis, and, to a lesser extent, with constitutional symptoms. Moreover, ANCA-positive patients

ANCAs IN CHURG-STRAUSS SYNDROME

had a significantly higher frequency of certain organ

system clinical manifestations, such as pulmonary hemorrhage, purpura, and mononeuritis multiplex. In contrast, ANCA-negative patients had a higher frequency of
heart and (less severe) lung disease.
Limited data have been reported on the correlation between ANCA positivity and the clinical features
in CSS, even though it should be noted that most (if not
all) reported cases of necrotizing crescentic glomerulonephritis in CSS (3943) involved ANCA-positive (usually MPO pANCA) patients, as in our cohort. Moreover,
the results of studies of small series of patients have
suggested that MPO ANCAs may be associated with the
onset of glomerular disorder in CSS (44,45).
In the largest series of CSS patients reported so
far, Guillevin et al looked for ANCAs in the sera of 42
patients at the time of diagnosis. ANCAs were detected
in 20 of 42 patients (47.6%), a percentage close to ours.
No details were given about the clinical associations with
ANCA positivity; however, a histologically proven, rapidly progressive necrotizing glomerulonephritis was documented in only 3 patients (7). More recently, the same
group has reported, in abstract form, that ANCAs were
detected in 43 of 112 patients with CSS (38%), and that
ANCA positivity at the time of diagnosis was associated
with a significantly higher frequency of renal involvement (P 0.001) with a relative risk of 14.3, whereas
ANCA-negative patients had higher rates of cardiac
involvement (P 0.001) (34). In addition, Booth et al
found, in a retrospective multicenter study on ANCAassociated renal vasculitis, that ANCAs were present in
92% of 256 patients, including most of the 11 patients
with CSS (46). Keogh and Specks found that central
nervous system involvement was the only clinical manifestation that correlated with ANCA status (14). In that
study, only 30 of 91 patients were tested at the time of
diagnosis; moreover, the prevalence of ANCA positivity
was much higher (73.3%) than that in our cohort, and
details about renal disease definition and renal histology
were not given (14).
In CSS, the histopathology and possibly the
pathogenesis of tissue lesions can be different in the
various organs and within the same target organ (69).
Although the pulmonary infiltrates in the prodromal
and vasculitic phases of the disease may be similar
radiographically, the histologic picture in the former
phase is usually that of extensive eosinophilic infiltration
of alveoli and interstitium, whereas necrotizing vasculitis
and granulomas are seen in the latter phase (6). Another
example is provided by gastrointestinal involvement,
which includes features of both eosinophilic gastroenteritis and necrotizing vasculitis (6,9). Peripheral nerve

2933

involvement might also be due to either vasculitis of vasa

nervorum or perineural eosinophilic infiltrate (6). The
eosinophil may also be directly responsible for some of
the classic disease features of CSS, by virtue of the
release of its stored cationic proteins, such as eosinophilic cationic protein, which is implicated in the cardiotoxicity that is seen both in CSS and in the hypereosinophilic syndrome, and eosinophil-derived neurotoxin,
which may contribute to the development of peripheral
neuropathy (8).
The clinical manifestations resulting from these
different pathogenic mechanisms cannot be easily distinguished from each other without histologic examination. It is tempting to speculate that the clinical manifestations might be due mainly to the eosinophilic
infiltrative disease in ANCA-negative patients, whereas
the necrotizing vasculitis component might prevail in
ANCA-positive patients. Indeed, in our experience, the
most frequent histologic finding in ANCA-negative patients was perivascular and/or tissue eosinophil infiltration, whereas an eosinophil-rich, necrotizing smallvessel vasculitis was found mainly in ANCA-positive
patients.
Emerging clinical and in vivo (animal model)
observations provide compelling evidence that ANCAs
are primarily and directly involved in the pathogenesis of
AASVs (47). In particular, 2 different groups of investigators have demonstrated that anti-MPO antibodies
alone can cause necrotizing crescentic glomerulonephritis and pulmonary hemorrhage in experimental models
(47,48). Moreover, Fienberg et al have reported that of
6 patients with negative test results for ANCAs and
histologically diagnosed WG, none had evidence of renal
involvement (49). In contrast, Cohen Tervaert et al
found that all patients with WG and anti-MPO antibodies had renal involvement (50). In MPA, which is usually
MPO ANCA positive, renal disease is almost invariably
present (10,11,32). The presence of ANCAs in CSS has
suggested that these autoantibodies could be an integral
part of the inflammatory diathesis of this disease (9).
ANCAs are capable of activating neutrophils in a wide
variety of ways, resulting in the release of reactive
oxygen species, granule proteins, cytokines, chemokines,
and adhesion molecules. Leukocytes that have been
activated by ANCAs adhere to endothelium and cause
endothelial damage (47). Although we do not think that
they are the main cause of the disease, they might act by
amplifying inflammation in a subset of patients, and they
might contribute directly to the development of necrotizing glomerulonephritis, as our results would suggest.
In conclusion, ANCAs are found less frequently
in CSS than in other ANCA-associated vasculitides, such

2934

SINICO ET AL

as WG and MPA. The main fluorescence pattern is

perinuclear with antibodies to MPO; however, PR3
cANCA can be found in a minority of patients. In
addition to the classical patterns, cANCA and atypical
ANCAs with MPO specificity are demonstrable in a
significant proportion of patients (apparently a much
higher proportion than in patients with MPA or WG).
ANCA positivity correlates with renal involvement (in
particular, with necrotizing crescentic glomerulonephritis), with constitutional symptoms, and with other particular organ system manifestations, such as mononeuritis multiplex, purpura, and pulmonary hemorrhage. In
contrast, ANCA negativity correlates with higher frequencies of lung involvement (with the exception of
alveolar hemorrhage) and heart involvement.

16.

17.
18.
19.

20.

21.

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