1. Association of CCL2 with systemic inflammation in Schnitzler’s syndrome
Karoline Krause, Robert Sabat, Ellen Witte-Händel, Anne Schulze, Viktoria Puhl, Marcus Maurer,
and Kerstin Wolk
Charité – Universitätsmedizin Berlin, Germany
Dept. of Dermatology and Allergy
Institute of Medical Immunology
Berlin-Brandenburg Centre for Regenerative Therapies
British Journal of Dermatology. DOI: /bjd.17334

2. Karoline Krause, M.D.
Kerstin Wolk, Ph.D.

3. Introduction What’s already known?
Schnitzler’s syndrome (SchS) is a rare, acquired autoinflammatory disease, clinically characterized by urticarial exanthema, arthralgia and osterosclerosis, and episodes of fever
The pathogenesis involves an over-activation of the IL-1 system
The exact pathogenetic pathways are substantially unknown and biomarkers for assessing the inflammatory activity are missing in SchS

4. Objective
This study aimed at the identification and characterization of pathogenetic players in SchS

5. Methods
Collection of clinical, laboratory and demographic data of patients with SchS (n=17)
Quantification of inflammatory parameters including cytokines, chemokines and antimicrobial proteins in the blood of these SchS patients and, for comparison, healthy subjects and patients with psoriasis and hidradenitis suppurativa using ELISA
Analysis of CCL2 (a chemoattractant for monocytic and further mononuclear immune cells) expression in cytokine-stimulated primary cells [immune cells (peripheral blood mononuclear cells), keratino-cytes, fibroblasts, endothelial cells] by RT-qPCR and ELISA

6. Results
CCL2 serum levels are markedly elevated and linked to disease activity in SchS
IL-1b
IL-6
IL-17
IL-19
1.5 30
1.0 12
P=0.003
P=0.024
P=0.098
P=0.461
0.8
1.0 20 8
0.6
serum level [pg/ml]
0.4
0.5 10 4
0.2
0.0
1.00
1.00
IL-22
TNF-a
CCL2
b-def. 2 20 3
600
3000
0.75
0.75
P=0.683
P=0.003
P=0.003
P=0.954
serum level [ng/ml]
CCL2 15
serum level [ng/ml]
CCL2 2
0.50
400
2000
0.50
serum level [pg/ml] 10
0.25 1
200
1000
0.25 5
rs = 0.61, P=0.040
rs=0.53, P=0.064
0.00
0.00 5 10 15 1 2 5 4 3
healthy
Schnitzler’s syndrome
disease activity
[PGA]
bone / muscle pain
[activity grade]
CCL2
800
P=0.000
P=0.366
P=0.785
Serum levels quantified by ELISA;
rs: Spearman’s correlation coefficient.
healthy
600
Schnitzler’s syndrome
serum level [pg/ml]
400
psoriasis vulgaris
hidradenitis suppurativa
200

7. Results
Mononuclear immune cells and fibroblasts produce CCL2 induced by IL-1ß and TNF-α
CCL2
0.8
0.8
0.8
0.8
IL-1b:
CCL2 quantification in cells and culture supernatants of 24h cytokine-stimulated primary immune cells (PBMC), endothelial cells (EC), fibroblasts (Fb) and keratinocytes (KC) using RT-qPCR and ELISA.
0.6
0.6
0.6
0.6
0 ng/ml
1 ng/ml
secreted protein [ng/ml]
0.4
0.4
0.4
0.4
10 ng/ml
0.2
0.2
0.2
0.2
100 ng/ml
0.0
0.0
0.0
0.0
PBMC EC Fb KC
CCL2, PBMC
CCL2, Fb
CCL2, Fb
CCL2, Fb
6000
6000 10
400
300
4000
4000 1
IL-1b only
secreted protein [% control]
mRNA [relative to HPRT]
secreted protein [% of IL-1b]
200
IL-1b
+ indicated cytokine(s)
2000
2000
0.1
100
0.0
0.0
0.01
TNF-a
IL-1b
IL-17
TNF-a
+ IL-17 -
IL-12
IL-23
IL-36b
TNF-a
IFN-g
GM-CSF -
IL-1b
IL-17A
IL-19
IL-22
IL-24
IL-36b
TNF-a
IFN-g -
IL-1b
IL-17A
TNF-a

8. Results CCL2 levels are associated with IL-1ß and TNF-α
levels and anti-IL-1ß therapy response in SchS
CCL2
Disease activity
1.00
1.00
0.6
0.0
0.2
1.0
0.8
0.4
0.6
0.0
0.2
1.0
0.8
0.4 15 5 25 20 10 25
P=0.004
P=0.004 20
0,75
0.75
serum level [ng/ml]
CCL2
serum level [ng/ml]
CCL2 15
0,50
0.50
serum level [ng/ml]
PGA 10
0.25
0.25 5
rs=0.48, P=0.025
rs=0.64, P=0.010
0.00
0.00
0.001
0.01
0.1 1 10
100
0.1 1 10
TNF-a
[pg/ml]
0 wks
1 wk
0 wks
1 wk
0 wks
1 wk
0 wks
1 wk
IL-1b
[pg/ml]
≥ 100 wks
≥ 100 wks
time of patient treatment with canakinumab
Serum levels quantified by ELISA;
rs: Spearman’s correlation coefficient.

9. Discussion
Quantifying CCL2 blood levels may allow the objective estimation of inflammatory disease activity and may help with the therapy decision in SchS patients
CCL2 may be a key element of the pathogenetic cascades in SchS, especially those important for osterosclerosis

10. Discussion
Proposed role of CCL2 in SchS

11. Conclusions What does this study add?
The chemokine CCL2 is significantly upregulated and is linked to global disease activity and treatment response in SchS patients
CCL2 is especially correlated with bone pain in these patients
The cellular sources of CCL2 include mononuclear immune cells and fibroblasts
IL-1β and TNF-α, whose blood levels correlate with CCL2 levels in SchS patients, are important CCL2 inducers

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