INVESTIGATIVE REPORT
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ActaDV ActaDV Advances in dermatology and venereology Acta Dermato-Venereologica
The ” Alarmins ” HMBG1 and IL-33 Downregulate Structural Skin Barrier Proteins and Impair Epidermal Growth
Uffe NYGAARD 1 , 2 , Ellen H . VAN DEN BOGAARD 3 , Hanna NIEHUES 3 , Malene HVID 2 , Mette DELEURAN 1 , Claus JOHANSEN 1 and Christian VESTERGAARD 1
1
Department of Dermatology and Venereology , Aarhus University Hospital , Aarhus University , 2 Department of Clinical Medicine , Aarhus University , Aarhus , Denmark , and 3 Department of Dermatology , Radboud Institute for Molecular Life Sciences , Radboud UMC , Nijmegen , The Netherlands
The epidermal-derived “ alarmins ” high-mobility group box 1 ( HMGB1 ) protein and interleukin-33 ( IL-33 ) are upregulated in patients with atopic dermatitis . However , their capacity as pro-inflammatory cytokines and their derived effects on skin barrier regulation are poorly elucidated . We investigated the impact of HMGB1 and IL-33 on gene transcription , protein expression and epidermal differentiation across 3 distinct keratinocyte in vitro models . Primary keratinocytes from healthy donors were used in submerged monolayer cultures , 3D human epidermis equivalents and 3D human skin equivalents . All keratinocyte models underwent 96-h stimulation with HMGB1 ( 100 μM ) or IL-33 ( 100 ng / ml ) using IL-4 ( 50 ng / ml ) as positive control of regulation and vehicle as negative control . We found that HMGB1 and IL-33 downregulated transcription of several genes from members of the epidermal differentiation complex , including filaggrin . Furthermore , HMGB1 downregulated the expression of the encoded proteins in the upper epidermis . Finally , IL-33 and HMGB1 ultimately led to impaired epidermal growth and maturation . In conclusion , HMGB1 and IL-33 could play a significant role in the atopic dermatitis pathophysiology due to negative regulation of structural proteins , stratum corneum formation and epidermal growth .
Key words : filaggrin ; involucrin ; loricrin ; IL-33 ; HMGB1 ; IL- 4 ; skin ; keratinocytes ; cell culture ; human skin equivalents ; atopic dermatitis .
Accepted Oct 12 , 2016 ; Epub ahead of print Oct 14 , 2016 Acta Derm Venereol 2017 : 97 : 305 – 312 .
Corr : Uffe Harboe Nygaard , Department of Dermatology and Venereology , Aarhus University Hospital , P . P . Oerumsgade 11 , DK-8000 Aarhus , Denmark . E-mail : uffenygaard @ clin . au . dk
The skin is a vital first line of defence between the environment and the human body . Studies indicate that defects in epidermal barrier function contribute to triggering and perpetuating skin inflammation in atopic dermatitis ( AD ) ( 1 – 5 ). Furthermore , disease activity and barrier function seem to be intimately related , and fluctuations in either can drive the disease ( 6 ). The role of skin barrier dysfunction as an important contributing factor in AD is further substantiated by loss-of-function mutations seen in the filaggrin gene which is a major risk factor for development of AD ( 3 , 7 , 8 ). Filaggrin is a key protein in epidermal homeostasis and barrier integrity ; and through its processing to free amino acids , it facilitates retention of water in the stratum corneum ( SC ) ( natural moisturising factor ) ( 3 , 9 ). In conjunction with filaggrin , other proteins encoded by genes in the epidermal differentiation complex , including loricrin and involucrin , are indispensable elements of the epidermal barrier ( 3 , 10 ). Skin alterations in AD are characterised by increased transepidermal water loss ( TEWL ) and a defective terminal keratinocyte differentiation entailing reduced levels of filaggrin , involucrin and loricrin ( 11 – 13 ).
Loss-of-function mutations in the filaggrin gene are found in approximately one third of all AD patients and are considered responsible for the decreased skin barrier function , the increased TEWL and increased disease severity . Yet , all patients with AD display a reduced expression of filaggrin and also involucrin and loricrin . This phenomenon is explained in part by the impact of cytokines , e . g . interleukin ( IL ) -4 , IL-13 and IL-25 that reduce the expression of barrier proteins ( 12 – 14 ).
The cytokine IL-33 is a member of the IL-1-family . IL-33 is constitutively expressed in healthy skin and is upregulated in skin of AD patients and in AD-like lesions in mice ( 15 – 18 ). Furthermore , we recently found elevated IL-33 serum levels in children with AD ( 19 , 20 ). IL-33 is classified as an alarmin due to the passive release upon cellular stress like allergen stimulation , scratching or non-apoptotic cell death , though these mechanism are still to be fully elucidated ( 16 , 21 , 22 ). It displays Th2 immunomodulatory effects by inducing IL-5 and IL-13 . It also displays a Th1 response via upregulation of IFN-γ ( 23 ). Keratinocytes produce IL-33 and also express its receptor complex consisting of ST2 and IL-1RAcP on their surface ( 16 , 24 ). Moreover , IL-33 appears to exert receptor-independent transcriptional effects in the nucleus of epithelial cells ( 25 ), thereby operating as a dualfunction immune mediator similarly to high-mobility group box 1 ( HMGB1 ) ( 26 ). HMGB1 is localised in the nucleus in almost all cells and is rapidly mobilised to other sites within the cell , and into the extracellular space in response to a variety of stimuli ( 26 , 27 ). Through RAGE and TLR4 signalling , HMGB1 mediates immune responses in non-infectious inflammation and shows
This is an open access article under the CC BY-NC license . www . medicaljournals . se / acta Journal Compilation © 2017 Acta Dermato-Venereologica . doi : 10.2340 / 00015555-2552 Acta Derm Venereol 2017 ; 97 : 305 – 312