Abstract
Canine progenitor epidermal keratinocytes (CPEK) are used as canine keratinocyte cell line. Their suitability for skin barrier studies is unknown. Measurement of transepithelial electric resistance (TEER) evaluates epithelial permeability. We compared TEER and tight junction (TJ) expression in CPEKs and normal keratinocytes (NK) harvested from biopsies of normal dogs. CPEKs and NK were grown until confluence (D0) and for 13 additional days. Slides were fixed on D0 and stained with ZO-1 and claudin-1 antibodies. Five images/antibody were taken, randomized and evaluated blindly by three investigators for intensity, staining location, granularity, and continuousness. Cell size and variability were evaluated. TEER increased overtime to 2000 Ohms/cm in NK, while remained around 100–150 Ohms/cm in CPEK. ANOVA showed significant effect of time (p < 0.0001), group (p < 0.0001) and group x time interaction (p < 0.0001) for TEER. Size of CPEKs was significantly (p < 0.0001) smaller and less variable (p = 0.0078) than NK. Intensity of claudin-1 staining was greater in CPEKs (p < 0.0001) while granularity was less in CPEKs (p = 0.0012). For ZO-1, cytoplasmic staining was greater in CPEK (p < 0.0001) while membrane continuousness of staining was greater in NK (p = 0.0002). We conclude that CPEKs grown in monolayer are not representative of NK for permeability studies.
Highlights
Keratinocytes are increasingly recognized as playing an important role in orchestrating the immune response and much attention has been devoted in recent years on the importance of skin barrier function in dermatological diseases like atopic dermatitis [1]
canine progenitor epidermal keratinocyte (CPEK) grown in monolayer did not perform to the cultures of keratinocytes harvested from normal dogs in terms of epithelial permeability measurements like TransEpithelial Electrical Resistance (TEER)
At this point in time, we can say that the location and intensity of tight junction (TJ) staining as well as TEER performance was not equivalent between CPEKs and normal canine keratinocytes grown in culture
Summary
Keratinocytes are increasingly recognized as playing an important role in orchestrating the immune response and much attention has been devoted in recent years on the importance of skin barrier function in dermatological diseases like atopic dermatitis [1]. In veterinary medicine it has been customary to use canine progenitor epidermal keratinocyte (CPEKs) for in vitro studies and consider them as a substitute for canine keratinocytes [2,3]. This is done for convenience as CPEKs can be purchased and no biopsies from dogs are needed. The alternative approach would be to take biopsies from dogs, harvest keratinocytes and establish primary cultures [4]. This approach is more labor intensive and the need for skin biopsies creates some practical limitations
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