Induction of normal and psoriatic phenotypes in submerged keratinocyte cultures

Abstract

Lesional psoriatic epidermis displays a number of phenotypic changes that are distinct from the differentiation program found in normal interfollicular epidermis. In psoriatic epidermis, keratinocytes are hyperproliferative and several differentiation-associated molecules are expressed that are absent in normal skin (e.g., cytokeratins (CK) 6, 16, and 17, and the epidermal proteinase inhibitor SKALP/ elafin). In addition, several molecules which are normally restricted to the stratum granulosum are strongly upregulated in the stratum spinosum (e.g., psoriasis-associated fatty acid binding protein (PA-FABP), psoriasin, involucrin, and transglutaminase). The aim of this study was to develop in vitro culture systems which (a) would allow to study the induction of normal and psoriatic differentiation pathways, and (b) would be amenable for screening of antipsoriatic drugs. Here we have investigated several models for induction of differentiation with respect to the expression of markers for the normal and psoriatic phenotype. Cell cycle parameters and expression levels of CK1, CK10, CK16, SKALP/elafin, transglutaminase, involucrin, psoriasin, and PA-FABP were assessed in these models using flow cytometry, immunocytochemistry, and Northern blot analysis. We observed that induction of differentiation with fetal calf serum resembled the psoriatic phenotype (sustained hyperproliferation; high levels of CK16, SKALP/elafin, transglutaminase, and involucrin; moderate psoriasin expression), whereas differentiation induced by growth factor depletion in a confluent culture resembled the normal differentiation phenotype (low proliferative rate; high expression levels of CK1 and CK10; moderate expression of involucrin and transglutaminase; low expression levels of SKALP/elafin and CK16; absence of psoriasin). We propose that these models can be used to study expression and pharmacological modulation of selected differentiation genes and the coordinated expression of sets of genes associated with epidermal differentiation programs.

Coal tar induces AHR-dependent skin barrier repair in atopic dermatitis

“…Keratinocytes were isolated from human abdominal skin derived from donors who underwent surgery for abdominal wall correction, as previously described (17). For submerged culture, keratinocytes were cultured in keratinocyte growth medium (Lonza) and differentiated by depletion of growth factors, as described previously (58). Coal tar was supplemented during the differentiation phase at indicated concentrations.…”

Psoriasis genetics: breaking the barrier

“…Granular layer products, such as psoriasis associated fatty acid binding protein (FABP5), filaggrin (FLG), corneodesmosin (CDSN), and proteins involved in the formation of the cornified envelope (CE) such as epidermal transglutaminase (TGM3), involucrin (IVL), and loricrin (LOR), can also be prematurely expressed in the spinous layer [21, 22]. The granular layer is sometimes reduced in psoriatic skin [23].…”

Identification and Sequence Analysis of Two New Members of the SKALP/elafin and SPAI-2 Gene Family

“…described by van Ruissen et al (34). Keratinocyte growth medium was composed of KBM (Clonetics; 0.15 mM calcium) supplemented with ethanolamine (Sigma; 0.1 mM), phosphoethanolamine (Sigma; 0.1 mM), bovine pituitary extract (Clonetics; 0.4% v/v), epidermal growth factor (Sigma; 10 ng/ml), insulin (Sigma; 5 g/ml), hydrocortisone (Collaborative Research Inc.; 0.5 g/ml), penicillin (Life Technologies, Inc.; 100 units/ml), and streptomycin (Life Technologies, Inc.; 100 g/ml).…”