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Archives of Toxicology

, Volume 91, Issue 4, pp 1783–1794 | Cite as

Understanding chemical allergen potency: role of NLRP12 and Blimp-1 in the induction of IL-18 in human keratinocytes

  • Angela Papale
  • Elena Kummer
  • Valentina Galbiati
  • Marina Marinovich
  • Corrado L. Galli
  • Emanuela Corsini
Immunotoxicology

Abstract

Keratinocytes (KCs) play a key role in all phases of skin sensitization. We recently identified interleukin-18 (IL-18) production as useful end point for determination of contact sensitization potential of low molecular weight chemicals. The aim of this study was to identify genes involved in skin sensitizer-induced inflammasome activation and to establish their role in IL-18 production. For gene expression analysis, cells were treated for 6 h with p-phenylenediamine (PPD) as reference contact allergen; total RNA was extracted and examined with a commercially available Inflammasome Polymerase Chain Reaction (PCR) array. Among genes induced, NLRP12 (Nod-like receptor P12) was selected for further investigation. NLRP12 promoter region contains Blimp-1 (B-lymphocyte-induced maturation protein-1)/PRDM1 binding site, and from the literature, it is reported that Blimp-1 reduces NLRP12 activity and expression in monocytes/macrophages. Their expression and role in KCs are currently unknown. To confirm NLRP12 expression and to investigate its relationship with Blimp-1, cells were exposed for different times (3, 6 and 24 h) to the extreme sensitizer 2,4-dinitrochlorobenzene (DNCB) and the strong sensitizer PPD. Allergens were able to induce both genes, however, with different kinetic, with DNCB more rapidly upregulating Blimp-1 and inducing IL-18 production, compared to PPD. NLRP12 and Blimp-1 expression appeared to be inversely correlated: Blimp-1 silencing resulted in increased NLRP12 expression and reduced contact allergen-induced IL-18 production. Overall results indicate that contact allergens of different potency differently modulate Blimp-1/NLRP12 expression, with strong allergen more rapidly downregulating NLRP12, thus more rapidly inducing IL-18 production. Data confirm that also in KCs, NLRP12 has an inhibitory effect on inflammasome activation assessed by IL-18 maturation.

Keywords

Allergen potency Contact allergy In vitro IL-18 Inflammasome 

Abbreviations

ACD

Allergic contact dermatitis

AOO

Acetone olive oil

ASC

Apoptosis-associated speck-like protein

Blimp-1

B-lymphocyte-induced maturation protein-1

CARD

Caspase activation and recruitment domain

DAMPs

Damage-associated molecular patterns

DiSFeB

Dipartimento di Scienze Farmacologiche e Biomolecolari

DMSO

Dimethyl sulfoxide

DNCB

2,4-Dinitrochlorobenzene

ELISA

Enzyme-linked Immunosorbent assay

IFN-γ

Interferon-γ

IL-18

Interleukin-18

IL-1β

Interleukin-1β

KC

Keratinocytes

LLNA

Local lymph node assay

LLR

Leucine-rich repeat protein

NF-κB

Nuclear factor-κB

NLR

NOD-like receptor

NLRP12

Nod-like receptor P12

PAMPs

Pathogen-associated molecular patterns

PBS

Phosphate-buffered saline

PCR

Polymerase chain reaction

PPD

p-Phenylenediamine

PVDF

Polyvinylidene difluoride

PYD

Pyrin domain

REACH

Registration evaluation authorization and restriction of chemicals

RHE

Reconstituted human epidermis

SDS

Sodium dodecyl sulfate

siRNA

Small interference RNA

TLRs

Toll-like receptors

TNF-α

Tumor necrosis factor-α

Notes

Acknowledgments

This research was partially supported by the Alternatives Research & Development Foundation, 2014—Alternatives Research Grant Program.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Angela Papale
    • 1
  • Elena Kummer
    • 1
  • Valentina Galbiati
    • 1
  • Marina Marinovich
    • 1
  • Corrado L. Galli
    • 1
  • Emanuela Corsini
    • 1
  1. 1.University of MilanMilanItaly

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