Barbara E. Brown

The epidermal hyperplasia associated with repeated barrier disruption by acetone treatment or tape stripping cannot be attributed to increased water loss

Acute disruption of the permeability barrier produces marked changes in epidermal metabolism, including increased lipid synthesis, increased DNA synthesis, and the enhanced production of cytokines. Because abnormalities in the barrier are present in a wide variety of skin disorders, we hypothesized that barrier disruption may be an important event that initiates pathological changes in the skin. In the present study, we found that repeated barrier disruption by topical acetone treatment or tape stripping induced epidermal hyperplasia in the flank skin of hairless mice and the ear of ICR mice, as well as inflammation in ear skin. The degree of epidermal hyperplasia correlated with the level and duration of barrier disruption. Likewise, the epidermal mitotic index, which was localized to the basal layer, increased with repeated disruption, indicating that the hyperplasia could be ascribed to increased cell proliferation. However, occlusion with a water-impermeable membrane, which prevents water loss, did not prevent the epidermal hyperplasia. Moreover, immunohistochemical staining for TNFα and IL 1α increased following repeated acetone treatment or tape stripping, and this increase also was not blocked by occlusion. These studies indicate that manipulations of the stratum corneum which disrupt the permeability barrier, such as, repeated acetone treatment or tape stripping, induce a variety of biologic responses in the underlying epidermis. Since neither the increase in epidermal cytokine production nor the described changes in cutaneous pathology were prevented by occlusion, in these two models the changes should not be attributed to increased water loss, but rather to epidermal injury resulting in the production and release of epidermal cytokines.

Topical treatment with thiazolidinediones, activators of peroxisome proliferator-activated receptor-γ, normalizes epidermal homeostasis in a murine hyperproliferative disease model

Abstract:  In a murine model of epidermal hyperplasia reproducing some of the abnormalities of several common skin disorders, we previously demonstrated the antiproliferative and pro‐differentiating effects of peroxisome proliferator‐activated receptor (PPAR)α, PPARβ/δ, and liver X receptor activators. Unlike other subgroups of PPAR activators, thiazolidinediones (TZDs), a family of PPARγ ligands, did not inhibit keratinocyte proliferation in normal murine skin. Here, we studied the effects of two TZDs, namely ciglitazone (10 mM) and troglitazone (1 mM), in the same murine model where epidermal hyperproliferation was reproduced by repeated barrier abrogation with tape stripping. Topical treatment with ciglitazone and troglitazone resulted in a marked and significant decrease in epidermal thickness. Furthermore, in all TZD‐treated groups, we observed a significant decrease in keratinocyte proliferation using proliferating cell nuclear antigen, 5‐bromo‐2′‐deoxyuridine, and tritiated thymidine incorporation. However, using the terminal deoxynucleotidyl transferase‐mediated dUTP nick end‐labeling assay, we found no difference in apoptosis between different treatments, emphasizing that it is the antiproliferative role of these activators that accounts for the decrease of epidermal thickness. Finally, using immunohistochemical methods, we determined the effects of ciglitazone on keratinocyte differentiation in this hyperproliferative model. We observed an increased expression of involucrin and filaggrin following ciglitazone treatment, suggesting a pro‐differentiating action of TZDs in this model. In summary, topical TZDs significantly reduce epidermal keratinocyte proliferation while promoting differentiation in a murine model of hyperproliferative epidermis. Together, these results suggest that in addition to their metabolic effects currently in use in the treatment of type 2 diabetes, topical TZDs could be considered as potential alternative therapeutic agents in hyperproliferative skin diseases such as psoriasis.

Epidermal Vascular Endothelial Growth Factor Production Is Required for Permeability Barrier Homeostasis, Dermal Angiogenesis, and the Development of Epidermal Hyperplasia : Implications for the Pathogenesis of Psoriasis

Primary abnormalities in permeability barrier function appear to underlie atopic dermatitis and epidermal trauma; a concomitant barrier dysfunction could also drive other inflammatory dermatoses, including psoriasis. Central to this outside-inside view of disease pathogenesis is the epidermal generation of cytokines/growth factors, which in turn signal downstream epidermal repair mechanisms. Yet, this cascade, if sustained, signals downstream epidermal hyperplasia and inflammation. We found here that acute barrier disruption rapidly stimulates mRNA and protein expression of epidermal vascular endothelial growth factor-A (VEGF-A) in normal hairless mice, a specific response to permeability barrier requirements because up-regulation is blocked by application of a vapor-impermeable membrane. Moreover, epidermal vegf(-/-) mice display abnormal permeability barrier homeostasis, attributable to decreased VEGF signaling of epidermal lamellar body production; a paucity of dermal capillaries with reduced vascular permeability; and neither angiogenesis nor epidermal hyperplasia in response to repeated tape stripping (a model of psoriasiform hyperplasia). These results support a central role for epidermal VEGF in the maintenance of epidermal permeability barrier homeostasis and a link between epidermal VEGF production and both dermal angiogenesis and the development of epidermal hyperplasia. Because psoriasis is commonly induced by external trauma [isomorphic (Koebner) phenomenon] and is associated with a prominent permeability barrier abnormality, excess VEGF production, prominent angiogenesis, and epidermal hyperplasia, these results could provide a potential outside-inside mechanistic basis for the development of psoriasis.

Activity of the hSPCA1 Golgi Ca2+ pump is essential for Ca2+-mediated Ca2+ response and cell viability in Darier disease

Keratinocyte differentiation, adhesion and motility are directed by extracellular Ca2+ concentration increases, which in turn increase intracellular Ca2+ levels. Normal keratinocytes, in contrast to most non-excitable cells, require Ca2+ release from both Golgi and endoplasmic reticulum Ca2+ stores for efficient Ca2+ signaling. Dysfunction of the Golgi human secretory pathway Ca2+-ATPase hSPCA1, encoded by ATP2C1, abrogates Ca2+ signaling and causes the acantholytic genodermatosis, Hailey-Hailey disease. We have examined the role of the endoplasmic reticulum Ca2+ store, established and maintained by the sarcoplasmic and endoplasmic reticulum Ca2+-ATPase SERCA2 encoded by ATP2A2, in Ca2+ signaling. Although previous studies have shown acute SERCA2 inactivation to abrogate Ca2+ signaling, we find that chronic inactivation of ATP2A2 in keratinocytes from patients with the similar acantholytic genodermatosis, Darier disease, does not impair the response to raised extracellular Ca2+ levels. This normal response is due to a compensatory upregulation of hSPCA1, as inactivating ATP2C1 expression with siRNA blocks the response to raised extracellular Ca2+ concentrations in both normal and Darier keratinocytes. ATP2C1 inactivation also diminishes Darier disease keratinocyte viability, suggesting that compensatory ATP2C1 upregulation maintains viability and partially compensates for defective endoplasmic reticulum Ca2+-ATPase in Darier disease keratinocytes. Keratinocytes thus are unique among mammalian cells in their ability to use the Golgi Ca2+ store to mediate Ca2+ signaling.

Avian epidermal differentiation. II. Adaptive response of permeability barrier to water deprivation and replenishment.

Zebra Finches are the epitome of desert-adapted avian species; i.e. they are able to survive without drinking water for over a year. Whereas transepidermal water loss (TEWL) in naked Zebra Finch nestlings is lower than in adults, and desert adaptation is accompanied by intercellular deposition of epidermal multigranular body (MGB) contents, MGB secretion is reduced as nestlings mature into feathered adults, indicative of less stringent barrier requirements. Here, removal of drinking water resulted in increased intercellular deposition of MGB contents, and TEWL progressively decreased. In contrast, MGB intercellular deposition decreased when birds were rehydrated, with TEWL returning towards normal within 5 days of rehydration. Finally, water-deprivation caused significant changes in epidermal lipid composition that returned toward control levels with rehydration. These studies show that adult Zebra Finches adapt to xeric stress by increased secretion of multigranular bodies resulting in reduced TEWL.

Lipokeratinocytes of the epidermis of a cetacean (Phocena phocena)

Biochemical and ultrastructural analysis of epidermis from the porpoise, Phocena phocena, revealed certain similarities and differences between cetaceans and terrestrial mammals. The predominant cell of cetacean epidermis, not found in normal terrestrial mammals, is a lipokeratinocyte, which elaborates not only keratin filaments, but also two types of lipid organelles: first, lamellar bodies, morphologically identical to those of terrestrial mammals, are elaborated in great abundance in all suprabasal epidermal layers, forming intercellular lipid bilayers in the stratum corneum interstices: and second, non-membrane-bounded droplets appear and persist in all epidermal layers. Although the porpoise lipokeratinocyte morpologically resembles the sebokeratocyte of avians in certain respects, nonmembrane-bounded lipid droplets are not released into the intercorneocyte space as they are in avian stratum corneum. Whereas phospholipid/neutral lipid gradients are similar in porpoise and terrestrial mammals, PAS-positive glycoconjugates, specifically glycosphingolipids, are retained in porpoise stratum corneum, but lost from these layers in terrestrials. The novel, non-polar acylglucosyl-ceramides, which also are lost during cornification in terrestrial mammals, are retained in porpoise stratum corneum. The lipid components of porpoise lipokeratinocytes appear to subserve not only barrier function in a hypertonic milieu, but also underlie the unique buoyancy, streamlining, insulatory, and caloric properties exhibited as adaptations to the cetacean habitat.

Epidermal injury stimulates prenylation in the epidermis of hairless mice

Isoprenylation is the covalent attachment of isoprenyl groups, intermediates of the cholesterol biosynthesis pathway, to carboxy terminal cysteine residues of proteins. Numerous proteins are isoprenylated including small GTP binding proteins, trimeric G proteins, and nuclear lamins, and these prenylated proteins regulate a variety of cell functions, including cell growth, cytokinesis, and differentiation. Here, we quantitated protein prenylation and determined which proteins are prenylated in the epidermis of hairless mice by radiolabeling with 3H-mevalonolactone following acute or chronic epidermal injury. In normal epidermis, four major radiolabeled bands, with molecular weights of 17–26, 48, 54, and 68 kDa, were observed. The levels of each of these bands increased by 24–63% 16 h following acute epidermal injury induced by topical acetone treatment or tape stripping, returning to normal by 24 h. On 2D gel electrophoresis, there were no major differences between the patterns of labeling following barrier disruption. Subacute epidermal injury induced by either acetone or tape stripping twice a day for 7 days and chronic injury induced by feeding an essential fatty acid-deficient (EFAD) diet, also resulted in a significant increase in protein prenylation. As with acute injury, SDS-PAGE and 2D gel electrophoresis did not reveal marked differences in the pattern of protein prenylation. These results demonstrate that the prenylation of proteins in the epidermis is stimulated by injury, suggesting that one or more of these prenylated species may be important in epidermal proliferation or differentiation.

Histochemical and morphological studies on mammalian epidermal peridermal granules

We performed morphological studies on the epidermal peridermal granules (PG) of newborn mice using histological, fluorescent and electron microscopic methods. PG appeared during the 17th to 18th day of foetal life, and remained in the outermost epidermal layers until 5 days after birth. PG were evenly distributed over the entire cutaneous surface of the mouse, except the tail. PG were found to contain protein and corresponded to sites of transglutaminase activity, but they were devoid of lipids and nucleic acids. They could be differentiated clearly from keratohyalin granules. Together, these studies suggest that PG represent a unique epidermal protein product that serves as a marker of late foetal development.