A Spectrum of Skin Disease: How S. aureus Colonization, Barrier Dysfunction, and Cytokines Shape the Skin

Abstract

Cytokines are key mediators of skin homeostasis and disease through their effects on keratinocytes (KCs), skin barrier integrity, immune activation and microbial ecology. Sirobhushanam et al (2020) suggest that the interferon (IFN) signature in lupus erythematosus (LE) alters expression of epithelial barrier and adhesin genes which, in turn, promotes Staphylococcus aureus colonization. This work highlights the need to better understand both barrier function and S. aureus colonization in LE, potential new therapeutic targets for the treatment of LE.

Psoriasis (PS), atopic dermatitis (AD), and cutaneous lupus erythematosus (LE) represent a unique spectrum of skin diseases based on the characteristics of barrier dysfunction, S. aureus colonization, and cutaneous cytokine profiles. These features are thought to contribute to disease onset and exacerbations in a feed forward loop, but without real clarity on which is primary or their relative importance. In this issue, Sirobhushanam et al. (2020) investigate the effects of type 1 IFNs on S. aureus colonization and epithelial barrier gene expression in LE. This study, together with our current understanding of epidermal barrier function and S. aureus colonization in PS and AD, places LE in the center of this barrier disruption and S. aureus colonization continuum (Table 1).

Table 1:

Unifying and Distinguishing Features of Three Common Inflammatory Skin Disorders.

Disease	Skin S. aureus (culture)	Skin S. aureus Adhesin Expression	Reduced Skin Barrier Gene Expression*	NL Skin Barrier Dysfunction (TEWL)	Skin Cytokines	Skin Lesion Location
Atopic Dermatitis	L > NL1 (>90%)	Yes3,4 (FGG & FN1)	FLG, FLG2, CLDN1, CLDN4, CLDN8, CLDN231,5	Yes1,5,6	IL-4, IL-13, IL-22, IFNγ	Flexors
Lupus	L (50%)2 NL (0%)	Yes2 (ITGA5)	DSG1, CLDN1, CLDN11, FLG & FLG22, 10-12	ND	type 1 IFNs, IL-12, IL-17, IL-23	UV exposed
Psoriasis	L (0%)2	ND	Not well studied8	Normal7	TNFα, IFNα, IL-12, IL-17, IL-22, IL-239	Extensors palms & soles

Abbreviations: L=lesional, NL=nonlesional, ND=not done.

^*We have included only epithelial genes whose reduced expression has been shown to affect barrier function.

References:

^1.(Weidinger, Beck et al. 2018)

^2.(Sirobhushanam, Parsa et al, 2020)

^3.(Cho, Strickland et al. 2001)

^4.(Cho, Strickland et al. 2001)

^5.(Brewer, Yoshida et al. 2019)

^6.(De Benedetto, Rafaels et al. 2011)

^7.(Darlenski, Hristakieva et al. 2018)

^8.(Visconti, Paolino et al. 2015)

^9.(Guttman-Yassky, Nograles et al. 2011)

^10.(Polivka, Hadj-Rabia et al. 2018)

^11.(Table 1)

^12.(Fig.4)

S. aureus Skin Colonization

S. aureus skin colonization can play a significant role in the onset, progression and severity of skin disease. Numerous studies have shown that > 90% of AD patients are colonized with S. aureus, which contrasts with ≈20% of healthy controls and rare individuals with PS (Otto 2010). We recently demonstrated that AD subjects colonized with S. aureus have more severe disease, as measured by the Eczema Area and Severity Index (EASI), greater skin barrier disruption as measured by increased transepidermal water loss (TEWL), and more type 2 immune deviation as measured by elevations in serum total IgE, CCL17 levels and absolute eosinophil counts (Simpson, Villarreal et al. 2018). In this issue, Sirobhushanam et al. (2020) evaluated the frequency of S. aureus skin colonization in a small cohort of PS and much larger sample of LE patients using routine culture techniques and PCR validation. They observed a modest increase in the percentage of LE patients who were colonized with S. aureus, identified enhanced epithelial expression of S. aureus adhesins (integrin alpha 5 [IGA5] and fibronectin-1 [FN1]) and greater S. aureus adhesion to LE keratinocytes.

Psoriasis

Psoriasis, in contrast to AD and LE, is a largely IL-17 driven disease. While this IL-17 skewing is relevant for the pathogenesis of a number of autoimmune and inflammatory disorders, it is also recognized as an important component of host defense and repair following infections with S. aureus (Otto 2010). IL-17 is thought to have a number of protective roles in the skin including enhancing production of antimicrobial peptides, such as lipocalin 2 and β-defensin, as well as neutrophil recruitment (Guttman-Yassky and Krueger 2017). More recently, IL-17A has also been shown to enhance tight junction (TJ) barrier function in primary human keratinocytes (Brewer, Yoshida et al. 2019). Collectively, these actions likely contribute to the observation made by Sirobhushanam et al. (2020) that none of their six PS subjects were colonized with S. aureus.

Atopic Dermatitis

AD is characterized by type 2 immunity, epidermal barrier disruption and S. aureus colonization (Weidinger, Beck et al. 2018). Barrier dysfunction is thought to be the consequence of reduced expression of stratum corneum (SC) and TJ structural proteins, an imbalance of proteases and protease-inhibitors, and altered lipid composition and organization. Epidermal barrier disruption promotes the release of alarmins such as TSLP, IL-25 and IL-33, that activate innate lymphoid cell type 2 (ILC2) cells and promote the recruitment of Th2 cells by inducing the release of the chemokines CCL17 and CCL20. The type 2 cytokines IL-4 and IL-13 have been shown to increase keratinocyte sensitivity to S. aureus toxins, suppress antimicrobial peptides, and promote S. aureus attachment by enhancing expression of fibrinogen and fibronectin (Weidinger, Beck et al. 2018), (Cho, Strickland et al. 2001). Collectively, these effects are thought to explain the high rates of S. aureus colonization in this disease, which arguably fuels a vicious cycle of barrier disruption and inflammation, ultimately leading to greater disease severity.

Lupus Erythematosus

While PS is characterized by increased skin barrier function with decreased bacterial colonization, and AD by the opposite, little was known about skin barrier proteins and bacterial colonization in the skin of lupus patients. LE is characterized by an elevated type I IFN signature, both in the skin and systemically. Sirobhushanam et al. suggest that, similar to AD, LE patients may have a feedback loop whereby type 1 IFNs alter the expression of epidermal barrier genes and S. aureus adhesins, thereby promoting S. aureus skin colonization, which drives further cytokine expression. This hypothesis would suggest that LE patients with S. aureus colonization would have more severe systemic disease which should be addressed in future studies.

The effect of the cytokine milieu on barrier function in LE appears to be complex. As in AD, FN1 expression is increased and β-defensins are decreased in LE skin lesions as compared to controls (Fig. 4), which would increase the likelihood of chronic S. aureus colonization. These expression differences are further enhanced in AD by the actions of Th2 cytokines (IL-4 & IL-13). However, the opposite effect was observed in IFNα-stimulated keratinocytes (Fig. 2 & 4). This suggests that other cytokines that are present in LE skin are likely to be responsible for S. aureus colonization. IL-13, which has been shown to be elevated in LE, may contribute to the observed gene expression changes and ultimately S. aureus skin colonization observed in half of LE patients (Morimoto, Tokano et al. 2001).

The epidermal barrier has been studied in AD at the genetic, mRNA, protein and functional levels. Null mutations in SC barrier genes such as filaggrin (FLG) and filaggrin 2 (FLG2) predispose individuals to development of AD, but their precise effects on barrier function and S. aureus colonization are still disputed (Weidinger, Beck et al. 2018), (Hansmann, Ahrens et al. 2012, Kawasaki, Nagao et al. 2012) The allergen sensitization that is characteristic of AD patients is thought to be due to disruption of both SC and TJ barriers, which is supported by the reduced expression of a number of TJ proteins (Claudin [CLDN]s-1, −4, −8 and −23). This construct has also been implicated in canine AD (De Benedetto, Rafaels et al. 2011, Kim, Cronin et al. 2016, Altunbulakli, Reiger et al. 2018). Sirobhushanam et al. showed that mRNA expression of several SC and TJ barrier proteins is decreased in skin biopsies or LE keratinocytes, but only a few of these proteins have been shown to affect barrier function (Table 1 & Fig. 4; FLG, CLDN-1, CLDN-11, desmoglein 1 (DSG1)) (Tamura and Tsukita 2014). Unfortunately, this study did not determine if these alterations in barrier gene expression were reflected in physiological measures of epidermal barrier function in LE patients (in vivo) or in their keratinocytes (in vitro). Future studies will need to evaluate whether reductions in these barrier genes increase TEWL measurements in LE patients or diminish transepithelial electrical resistance in LE keratinocytes propagated ex vivo. Such studies may highlight the potential importance of addressing skin barrier disturbance as a new therapeutic strategy for LE patients.

Future therapeutic approaches

The authors previously demonstrated the importance of IFNκ in cutaneous LE pathogenesis (Sarkar, Hile et al. 2018). The three IFN types; type I IFN (e.g. IFNα and IFNκ), type II (IFNγ) and type III (e.g. IFNλ) should be explored to define their relative importance in epithelial barrier integrity, epithelial innate immune responses and expression of S. aureus adhesins and ultimately in S. aureus colonization. Baricitinib, which is an oral selective Janus kinase (JAK)1 and JAK2 inhibitor that improves the signs and symptoms of LE, was shown to reverse the increased S. aureus adhesion observed in IFNα-stimulated KCs (Fig. 3) (Wallace, Furie et al. 2018). A more targeted therapy to directly address the effect of type 1 IFNs on barrier function and S. aureus colonization in LE patients would be anifrolumab, which is a fully human monoclonal antibody that binds to subunit 1 of the type I IFN receptor. Anifrolumab has recently been shown to be effective in LE (Morand 2019).

Ustekinumab, a monoclonal antibody targeting IL-12 and IL-23 that is FDA-approved to treat PS, has also shown some benefit for the treatment of LE (van Vollenhoven, Hahn et al. 2018). This might seem surprising based on its inhibition of the IL-17 pathway, which one might predict would further aggravate skin barrier dysfunction and promote S. aureus colonization in LE. While IL-17 has been shown to be modestly elevated in LE, there is not much evidence that this is relevant for LE pathogenesis (Martin, Baeten et al. 2014). Therefore, one might assume that ustekinumab is improving LE primarily via its actions on IL-12 blockade, and the inhibitory effect this has on the Th1 response.

New therapeutic approaches to LE, both cutaneous and systemic, are needed as only one medication has been FDA-approved for LE in the past 60 years. By comparison, great strides have been made in the development of targeted and safe treatments for PS and recently for AD as well. This paper highlights a new hypothesis, namely that cutaneous colonization with S. aureus may play a role in LE pathogenesis. The importance of S. aureus colonization in SLE as either a driver of type 1 IFNs or as a consequence of skin barrier disruption (or both) needs to be established. This study also suggests that there is much more to be done to understand the complex relationship between inflammatory skin diseases and cutaneous microbial ecology.

Clinical Implications:

• Patients with lupus are more commonly colonized with S. aureus than patients with psoriasis.

• The epithelium of lupus patients has alterations in gene expression, which may affect S. aureus adhesion, barrier function and innate immune responsiveness.

• Some of these epithelial abnormalities may be explained by the actions of type 1 or type 2 interferons.

Abbreviations:

AD

Atopic dermatitis

CLDN

claudin

EASI

Eczema Area and Severity Index

FGG

fibrinogen

FN1

fibronectin

FLG

filaggrin

FLG2

filaggrin 2

ILC2

innate lymphoid cell type 2

ITGA5

integrin subunit alpha 5

IFN

interferon

JAK

Janus kinase

KCs

keratinocytes

LE

lupus erythematosus

PS

psoriasis

S. aureus

Staphylococcus aureus

SC

stratum corneum

TSLP

thymic stromal lymphopoietin

TJ

tight junction

TEWL

transepidermal water loss

TGM5

transglutaminase 5