R. Voegeli

Increased stratum corneum serine protease activity in acute eczematous atopic skin

Background  Atopic dermatitis (AD) is a chronic inflammatory disease associated with changes in stratum corneum (SC) structure and function. The breakdown of epidermal barrier function in AD is associated with changes in corneocyte size and maturation, desquamation, lipid profiles, and some protease activities.

Profiling of serine protease activities in human stratum corneum and detection of a stratum corneum tryptase-like enzyme

Seasonal variation in stratum corneum (SC) biophysical and biological characteristics has been described previously. In particular, the winter season has been shown to affect more severely the properties of facial skin compared with forearm skin. Moreover, when compromised, such as in dry skin conditions, facial SC has been shown to contain increased inflammatory cytokines and proteases. Nevertheless, there have been no comparative studies of the activities and depth activity of several proteases in the SC on different body sites and at different times of the year. In this study, we examined the distribution of key serine protease activities (kallikrein 5, kallikrein 7, urokinase, plasmin and a tryptase‐like enzyme) in different layers of the SC on the cheek and the forearm by analysis of consecutive tape strippings of healthy Caucasian subjects during winter and summer. The protein content of the tape strippings was quantified by absorption measurements with a recently developed and novel infrared densitometer SquameScanTM 850A while the SC enzyme activities were determined using fluorogenic peptide substrates. Transepidermal water loss (TEWL), skin pH and skin hydration were higher on the cheek than on the forearm. In the same way, the activity of the inflammatory‐related proteases plasmin, urokinase and tryptase was approximately five to eight times and the activity of the desquamatory‐related proteases kallikrein 5 and kallikrein 7 approximately two to four times higher on the cheek than on the forearm. There were no gender‐related differences in these enzyme activities except for the increased kallikrein 7 in the forearm skin of the female subjects in winter. Reduced kallikrein 5 was associated with increased SC cohesion, as judged by increased protein removal, in forearm skin in the winter months of the year although the skin was clinically normal. It can be concluded that (i) protected skin areas show lower TEWL, skin pH and skin hydration and less protease activities than skin areas that are exposed to the environment, possibly indicating subclinical inflammation on these body sites, (ii) in normal healthy forearm skin, the outer SC exhibits greater serine protease activity than its deeper layers, (iii) compared with the forearm, urokinase‐ and plasmin‐like activities are elevated on SC strippings from the cheek, confirming activation of the plasminogen cascade, and (iv) tryptase‐like activity in the SC is also elevated in samples from the cheek, possibly indicating involvement of mast cells in these barrier‐compromised body sites or the synthesis of a novel tryptase‐like enzyme by keratinocytes. Although elevation of the activities of urokinase, plasmin, kallikrein 5, kallikrein 7 and now a tryptase‐like enzyme was observed on SC derived from skin of clinically normal cheeks, we anticipate even higher activities in skin conditions where the epidermal barrier is further impaired.

Stratum corneum proteases and dry skin conditions

This paper reviews the role of stratum corneum (SC) proteases and their inhibitors in normal and xerotic skin conditions. The importance of the corneodesmosome for SC integrity is also discussed, and the effect of proteases on its disassembly. The relevance of each enzyme class is outlined, as well as their potential inhibitors. It is becoming much clearer, however, that the LEKTI family of inhibitors are critical for SC enzyme control. Delayed desquamation is the accumulation of corneocytes on the surface of the SC that leads ultimately to the cosmetic condition commonly termed as “dry skin”. The reductions of serine protease activity are a consistent theme in dry skin, and non-eczematous atopic dermatitis otherwise known as atopic xerosis leading to retention hyperkeratosis. Flaky skin is normally seen on the body whereas a rough skin is observed on the face. Increased protease activity occurs in most, if not all, inflammatory dermatoses, ranging from the genetic disorders, psoriasis and eczematous atopic dermatitis to sub-clinical barrier abnormalities induced by surfactants or by environmental influences as a result of premature desquamation. In some of these conditions a thinner SC is apparent, e.g., eczematous atopic skin or on photodamaged facial skin. A better understanding of the proteolytic events and of the regulatory mechanisms involved in desquamation should enable the design of new treatments for skin disorders associated with faulty desquamation. This new knowledge will be an important basis for new developments in ‘corneotherapy’ and ‘corneocare’.

Increased basal transepidermal water loss leads to elevation of some but not all stratum corneum serine proteases.

There are indications of elevation of some inflammatory serine proteases in barrier damaged skin (e.g. plasmin and urokinase). Moreover, many other serine protease activities are present such as desquamatory enzymes as well as a newly detected tryptase‐like serine protease. However, the activities of these proteases have never been correlated with stratum corneum (SC) barrier function. The activity of extractable key serine proteases (SC trypsin‐like kallikreins, SC chymotrypsin‐like kallikreins, SC tryptase‐like serine protease, urokinase and plasmin) was measured from the outermost layers of SC obtained from facial tape strippings in clinically normal subjects. The protein content of the tape strippings was quantified by absorption measurements with the novel infrared densitometer SquameScanTM 850A and the protease activities by the use of fluorogenic peptide substrates. SC barrier function, SC hydration and skin surface pH were measured using AquaFluxTM, NOVA dermal phase meter and Skin‐pH‐Meter®, respectively. As expected, SC hydration was reduced with increased transepidermal water loss (TEWL) values indicative of barrier impairment. Surprisingly, SC chymotrypsin‐like activity showed no correlation with hydration or TEWL, whereas all other enzymes positively correlated with impaired barrier function and some were statistically significant: SC trypsin‐like kallikreins (R2 = 0.66, P < 0.01), SC tryptase‐like enzyme (R2 = 0.95, P < 0.001), plasmin (R2 = 0.86, P < 0.001) and urokinase (R2 = 0.50, P < 0.05). All enzymes except urokinase also negatively correlated with SC hydration. Elevated levels of SC serine proteases have been associated with some dermatological disorders, such as atopic dermatitis, psoriasis and rosacea but these results indicate that these enzymes are also elevated with milder forms of barrier disruption, which is not clinically evident as irritated skin. As these proteases are elevated in the SC, they will also be elevated in the epidermis where they can be involved in neurogenic inflammation and epidermal barrier impairment via activation of the protease‐activated receptors. These results highlight the need for using serine protease inhibitors especially for urokinase and plasmin, SC tryptase‐like serine protease and possibly SC trypsin‐like kallikreins even in milder forms of barrier damage.

Increased mass levels of certain serine proteases in the stratum corneum in acute eczematous atopic skin.

Acute eczematous atopic dermatitis (AD) is associated with increases in stratum corneum (SC) serine protease activity. The purpose of this study was to examine whether the increased SC protease activities in acute eczematous atopic dermatitis were associated with increased mass levels of SC proteases. Six subjects with healthy skin and six patients with AD each with non‐lesional skin or lesional acute eczematous skin had the mass levels of their extractable SC kallikreins (KLK), plasmin and urokinase quantified using Luminex multiplex bead‐based assays from SC tape strippings. The mass levels of KLK5 and KLK14 together with urokinase were not elevated in the SC in atopic skin. However, the mass levels of KLK7 and KLK11 together with plasmin were greatly elevated compared with the extracts from the non‐lesional and the healthy skin and correlated with the corresponding enzymatic activities.

A fundamental investigation into aspects of the physiology and biochemistry of the stratum corneum in subjects with sensitive skin.

Sensitive skin is a poorly understood skin condition. Defects in stratum corneum (SC) barrier function and/or extrasensory neuronal networks in the epidermis are believed to be involved in the problem.

Variation in the activities of late stage filaggrin processing enzymes, calpain-1 and bleomycin hydrolase, together with pyrrolidone carboxylic acid levels, corneocyte phenotypes and plasmin activities in non-sun exposed and sun-exposed facial stratum corneum of different ethnicities.

Knowledge of the ethnic differences and effects of photodamage on the relative amounts of natural moisturizing factor (NMF) together with filaggrin processing enzymes in facial stratum corneum is limited. Our aim was to characterize the activities of calpain‐1 (C‐1), bleomycin hydrolase (BH) and the levels of pyrrolidone carboxylic acid (PCA) as a marker for total NMF levels and to relate them to plasmin activities and corneocyte maturation.

A novel continuous colour mapping approach for visualization of facial skin hydration and transepidermal water loss for four ethnic groups

The aim of this exploratory study was to develop a novel colour mapping approach to visualize and interpret the complexity of facial skin hydration and barrier properties of four ethnic groups (Caucasians, Indians, Chinese and Black Africans) living in Pretoria, South Africa.

Facial skin pigmentation is not related to stratum corneum cohesion, basal transepidermal water loss, barrier integrity and barrier repair

Hypotheses have been developed for the evolutionary selection of skin pigmentation one of which relates to improved skin barrier function. The aim of this study was to compare facial skin condition on photoexposed (cheek) and photoprotected (post‐auricular) sites of naturally pigmented subjects of different ethnicities (Fitzpatrick skin phototypes II/III and V/VI) and Albino African subjects to understand better the relationship between facial stratum corneum (SC) barrier function, skin surface pH and skin pigmentation.

Variation in stratum corneum protein content as a function of anatomical site and ethnic group.

Quantification of stratum corneum (SC) protein levels from tape strippings is frequently used to investigate skin conditions, to correct for amounts of SC protein removed in SC biomarker studies and to determine distribution of topically applied ingredients. In recent years, a rapid and convenient method for SC protein quantification from tape strippings has become available using infrared densitometry (IRD). However, standard curves have only been generated for Caucasian forearm and shoulder SC and have been assumed to be correct not only for facial SC but also for SC samples of other ethnic groups. The aim of this study was to investigate whether the use of IRD for SC protein measurement is valid for other body sites such as the cheek and for measuring SC protein content of darkly pigmented skin types.