Patterning in stratified epithelia depends on cell-cell adhesion.

Epithelia consist of proliferating and differentiating cells that often display patterned arrangements. However, the mechanism regulating these spatial arrangements remains unclear. Here, we show that cell-cell adhesion dictates multicellular patterning in stratified epithelia.

Substrate membrane bearing close-packed array of micron-level pillars incrassates air-exposed three-dimensional epidermal equivalent model.

Background: We showed previously that a thick three-dimensional epidermal equivalent can be constructed with passaged keratinocytes on a patterned surface.

Material And Methods: We first carried out computer simulations of a three-dimensional epidermal equivalent model built on close-packed arrays of 10 µm, 15 µm, 20 µm, 30 µm, and 60 µm diameter pillars. Based on these predictions, we evaluated epidermal equivalents built on a series of porous plastic membranes bearing arrays of pillars 15 µm, 20 µm, 25 µm, 30 µm, and 50 µm in diameter.

Mathematical-model-guided development of full-thickness epidermal equivalent.

Epidermal equivalents prepared with passaged keratinocytes are typically 10-20 μm thick, whereas intact human epidermis is up to 100 μm thick. Our established mathematical model of epidermal homeostasis predicted that the undulatory pattern of the papillary layer beneath the epidermis is a key determinant of epidermal thickness. Here, we tested this prediction by seeding human keratinocytes on polyester textiles with various fiber-structural patterns in culture dishes exposed to air, aiming to develop a more physiologically realistic epidermal model using passaged keratinocytes.

Real-time imaging of human epidermal calcium dynamics in response to point laser stimulation.

Background: Changes of epidermal calcium ion concentration are involved in regulation of barrier homeostasis and keratinocyte differentiation. Moreover, intracellular calcium dynamics might play a role in skin sensation. But, although calcium dynamics of cultured keratinocytes in response to mechanical stresses has been well studied, calcium propagation in stimulated human epidermis is still poorly understood.

Japanese Cedar (Cryptomeria japonica) pollen allergen induces elevation of intracellular calcium in human keratinocytes and impairs epidermal barrier function of human skin ex vivo.

Cry j1 is the major peptide allergen of Japanese cedar (Sugi), Cryptomeria japonica. Since some allergens disrupt epidermal permeability barrier homeostasis, we hypothesized that Cry j1 might have a similar effect. Intracellular calcium level in cultured human keratinocytes was measured with a ratiometric fluorescent probe, Fura-2 AM.

Abnormal Morphology of Blood Vessels in Erythematous Skin From Atopic Dermatitis Patients.

Previous studies suggest that altered peripheral blood circulation might be associated with erythema or inflammation in atopic dermatitis (AD) patients. However, the overall structure of blood vessels and capillaries in AD skin is poorly understood because most studies have involved light-microscopic observation of thin skin sections. In the present study, we compared the 3-dimensional structures of peripheral blood vessels of healthy subjects and AD patients in detail by means of 2-photon microscopy.

Effects of medium flow on axon growth with or without nerve growth factor.

Axon growth is a crucial process in regeneration of damaged nerves. On the other hand, elongation of nerve fibers in the epidermis has been observed in skin of atopic dermatitis patients. Thus, regulation of nerve fiber extension might be an effective strategy to accelerate nerve regeneration and/or to reduce itching in pruritus dermatosis.

Mathematical modeling of calcium waves induced by mechanical stimulation in keratinocytes.

Recent studies have shown that the behavior of calcium in the epidermis is closely related to the conditions of the skin, especially the differentiation of the epidermal keratinocytes and the permeability barrier function, and therefore a correct understanding of the calcium dynamics is important in explaining epidermal homeostasis. Here we report on experimental observations of in vitro calcium waves in keratinocytes induced by mechanical stimulation, and present a mathematical model that can describe the experimentally observed wave behavior that includes finite-range wave propagation and a ring-shaped pattern. A mechanism of the ring formation hypothesized by our model may be related to similar calcium propagation patterns observed during the wound healing process in the epidermis.

Coculture system of keratinocytes and dorsal-root-ganglion-derived cells for screening neurotrophic factors involved in guidance of neuronal axon growth in the skin.

The density of peripheral nerve fibres is increased in atopic dermatitis. Moreover, reduction in the fibres in a mouse model of atopic dermatitis reduces scratching behaviour. Thus, regulation of nerve fibre extension could be an effective strategy to reduce itching in pruritus dermatosis.

Low environmental humidity induces synthesis and release of cortisol in an epidermal organotypic culture system.

Dry environmental conditions induce a variety of skin pathologies and a recent report indicating that cortisol synthesis in epidermis was increased during wound healing led us to hypothesize that environmental dryness might induce increased cortisol secretion in epidermis. Therefore, we incubated a skin equivalent model under dry (relative humidity: less than 10%) and humid (relative humidity: approximately 100%) conditions for 48 hours and evaluated cortisol secretion and mRNA levels of cortisol-synthesizing enzyme (steroid 11β-hydroxylase, CYP11B1) and IL-1β. Cortisol secretion was increased threefold, and CYP11B1 and IL-1β mRNAs were increased 38-fold and sixfold, respectively, in the dry condition versus the humid condition.

External negative electric potential accelerates exocytosis of lamellar bodies in human skin ex vivo.

Exocytosis of lamellar bodies at the uppermost nucleated layer of the epidermis is a crucial process for epidermal permeability barrier homoeostasis. We have previously suggested that skin surface electric potential might be associated with barrier homoeostasis. Thus, we hypothesized that the potential might drive exocytosis of lamellar bodies.

Distinct intracellular calcium responses of individual cultured human keratinocytes to air pressure changes.

Background: We previously showed that application of hydraulic pressure to cultured human keratinocytes induced elevation of intracellular calcium concentration ([Ca(2+) ]i ), but the absolute value of the pressure could not be determined.

Purpose: To evaluate the effect of the absolute value of pressure on keratinocytes and other skin cells.

Methods: In the present work, we examined the effect of changes in absolute pressure level by observing the [Ca(2+) ]i responses of cultured human keratinocytes and other cells cultured at the bottom of a hermetically sealed plastic flask as the air pressure in the flask was increased gradually, held stable, and then decreased abruptly, using the Ca(2+) -indicator fura-2.

In vitro formation of organized structure between keratinocytes and dorsal-root-ganglion cells.

We recently found that the morphology in a co-culture system of keratinocytes and dorsal-root ganglion-derived cells depended on the timing of seeding of the two cell types. In skin, epidermis is formed first, followed by construction of peripheral nerve structure. Therefore, we hypothesized that formation of peripheral nerve structure in the epidermis might be driven by interaction between keratinocytes and nerve cells.

Oxytocin is expressed in epidermal keratinocytes and released upon stimulation with adenosine 5'-[γ-thio]triphosphate in vitro.

Oxytocin is a neuropeptide produced primarily in the hypothalamus and is best known for its roles in parturition and lactation. It also influences behaviour, memory and mental state. Recent studies have suggested a variety of roles for oxytocin in peripheral tissues, including skin.

Ryanodine receptors are expressed in epidermal keratinocytes and associated with keratinocyte differentiation and epidermal permeability barrier homeostasis.

Ryanodine receptors (RyRs) have an important role as calcium channels in the regulation of intracellular calcium levels in the nervous system and muscle. In the present study, we investigated the expression of RyR in human epidermis. Immunohistochemical studies and reverse transcription-PCR indicated the expression of RyR type 1, 2, and 3 proteins in epidermal keratinocytes.

Intracellular calcium response to high temperature is similar in undifferentiated and differentiated cultured human keratinocytes.

A series of thermo-sensitive proteins is expressed in epidermal keratinocytes, where they function as cation channels that are activated at temperatures in the range of 28-42°C. Here, we examined the influence of high temperature on intracellular calcium level in undifferentiated and differentiated cultured human keratinocytes, which are considered to be representative of deeper and upper epidermal layers, respectively. We found that the percentages of responsive keratinocytes among undifferentiated and differentiated cells exposed to temperatures in the warm-hot range (30-50°C) were essentially the same.