scholarly journals Canine Epidermal Keratinocytes (CPEK) Grown in Monolayer Are Not Representative of Normal Canine Keratinocytes for Permeability Studies: Pilot Studies

2022 ◽  
Vol 9 (1) ◽  
pp. 25
Author(s):  
Rosanna Marsella ◽  
Rachel Wilkes ◽  
Kim Ahrens
Keyword(s):  
Skin Barrier ◽  
Epidermal Keratinocytes ◽  
Epithelial Permeability ◽  
Electric Resistance ◽  
Pilot Studies ◽  
Cytoplasmic Staining ◽  
Time Interaction ◽  
Normal Keratinocytes ◽  
Keratinocyte Cell ◽  
Permeability Studies

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.

scholarly journals Central role of α7 nicotinic receptor in differentiation of the stratified squamous epithelium

2002 ◽  
Vol 159 (2) ◽  
pp. 325-336 ◽  
Author(s):  
Juan Arredondo ◽  
Vu Thuong Nguyen ◽  
Alexander I. Chernyavsky ◽  
Dani Bercovich ◽  
Avi Orr-Urtreger ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Terminal Differentiation ◽  
Skin Barrier ◽  
Epidermal Keratinocytes ◽  
Cycle Progression ◽  
Cell Monolayers ◽  
Α7 Nachr ◽  
Keratinocyte Cell ◽  
Functional Inactivation

Several ganglionic nicotinic acetylcholine receptor (nAChR) types are abundantly expressed in nonneuronal locations, but their functions remain unknown. We found that keratinocyte α7 nAChR controls homeostasis and terminal differentiation of epidermal keratinocytes required for formation of the skin barrier. The effects of functional inactivation of α7 nAChR on keratinocyte cell cycle progression, differentiation, and apoptosis were studied in cell monolayers treated with α-bungarotoxin or antisense oligonucleotides and in the skin of Acra7 homozygous mice lacking α7 nAChR channels. Elimination of the α7 signaling pathway blocked nicotine-induced influx of 45Ca2+ and also inhibited terminal differentiation of these cells at the transcriptional and/or translational level. On the other hand, inhibition of the α7 nAChR pathway favored cell cycle progression. In the epidermis of α7−/− mice, the abnormalities in keratinocyte gene expression were associated with phenotypic changes characteristic of delayed epidermal turnover. The lack of α7 was associated with up-regulated expression of the α3 containing nAChR channels that lack α5 subunit, and both homomeric α9- and heteromeric α9α10-made nAChRs. Thus, this study demonstrates that ACh signaling through α7 nAChR channels controls late stages of keratinocyte development in the epidermis by regulating expression of the cell cycle progression, apoptosis, and terminal differentiation genes and that these effects are mediated, at least in part, by alterations in transmembrane Ca2+ influx.

Anti-proliferative effects of an herbal formulated cream on human keratinocytes and its implication for psoriasis treatment

2016 ◽  
Vol 5 (07) ◽  
pp. 4686 ◽  
Author(s):  
Harsha M. R.* ◽  
Baidyanath Mishra ◽  
Chaithra C. S. ◽  
Vivekananda Ramana
Keyword(s):  
Cell Model ◽  
Test Material ◽  
Epidermal Keratinocytes ◽  
Recurrence Rates ◽  
Inflammatory Infiltration ◽  
Keratinocyte Proliferation ◽  
Psoriasis Treatment ◽  
Study Material ◽  
Keratinocyte Cell ◽  
Promising Source

Psoriasis is a chronic inflammatory skin disorder which affects more than 3% of the population worldwide and is characterized histopathologically by proliferative imbalance and abnormal differentiation of epidermal keratinocytes and inflammatory infiltration. Hence, loss of regulation in keratinocyte proliferation and differentiation makes it a typical pathophysiological phenomenon in psoriasis manifestation. Traditionally, herbal products used in treating psoriasis have shown promising effects in several clinical studies with relatively fewer adverse effects, higher remission and lower recurrence rates. In our previous study, the polyherbal formulation of InnoVision’s test material was found to induce AQP-3 expression in keratinocyte cell line. In the present study, we screened the study material for its anti-proliferative properties using cultured human HACAT keratinocyte cell model. Our experimental results suggest that InnoVision’s Psoriderm Cream is a promising source which can be effectively used as an herb-based topical agent for psoriasis treatment. Evidence is provided that inhibition of keratinocyte proliferation and improving skin hydration via induction of aquaporin-3 stimulation is the possible underlying mechanism for the observed anti-psoriatic action of study material. 

scholarly journals EGFR inhibitors switch keratinocytes from a proliferative to a differentiative phenotype affecting epidermal development and barrier function

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Nicolas Joly-Tonetti ◽  
Thomas Ondet ◽  
Mario Monshouwer ◽  
Georgios N. Stamatas
Keyword(s):  
Growth Factor ◽  
Barrier Function ◽  
Kinase Inhibitors ◽  
Treatment Protocol ◽  
Growth Factor Receptor ◽  
Skin Barrier ◽  
Keratinocyte Differentiation ◽  
Skin Barrier Function ◽  
Epidermal Keratinocytes ◽  
Epidermal Structure

Abstract Background Cutaneous adverse drug reactions (CADR) associated with oncology therapy involve 45–100% of patients receiving kinase inhibitors. Such adverse reactions may include skin inflammation, infection, pruritus and dryness, symptoms that can significantly affect the patient’s quality of life. To prevent severe skin damages dose adjustment or drug discontinuation is often required, interfering with the prescribed oncology treatment protocol. This is particularly the case of Epidermal Growth Factor Receptor inhibitors (EGFRi) targeting carcinomas. Since the EGFR pathway is pivotal for epidermal keratinocytes, it is reasonable to hypothesize that EGFRi also affect these cells and therefore interfere with the epidermal structure formation and skin barrier function. Methods To test this hypothesis, the effects of EGFRi and Vascular Endothelial Growth Factor Receptor inhibitors (VEGFRi) at therapeutically relevant concentrations (3, 10, 30, 100 nM) were assessed on proliferation and differentiation markers of human keratinocytes in a novel 3D micro-epidermis tissue culture model. Results EGFRi directly affect basal keratinocyte growth, leading to tissue size reduction and switching keratinocytes from a proliferative to a differentiative phenotype, as evidenced by decreased Ki67 staining and increased filaggrin, desmoglein-1 and involucrin expression compared to control. These effects lead to skin barrier impairment, which can be observed in a reconstructed human epidermis model showing a decrease in trans-epidermal water loss rates. On the other hand, pan-kinase inhibitors mainly targeting VEGFR barely affect keratinocyte differentiation and rather promote a proliferative phenotype. Conclusions This study contributes to the mechanistic understanding of the clinically observed CADR during therapy with EGFRi. These in vitro results suggest a specific mode of action of EGFRi by directly affecting keratinocyte growth and barrier function.

scholarly journals Apoptosis as a Mechanism for Keratinocyte Death in Canine Toxic Epidermal Necrolysis

2016 ◽  
Vol 54 (2) ◽  
pp. 249-253 ◽  
Author(s):  
F. Banovic ◽  
S. Dunston ◽  
K. E. Linder ◽  
P. Rakich ◽  
T. Olivry
Keyword(s):  
Cell Death ◽  
Toxic Epidermal Necrolysis ◽  
Caspase 3 ◽  
Skin Lesions ◽  
Terminal Deoxynucleotidyl Transferase ◽  
Epidermal Keratinocytes ◽  
Biopsy Specimens ◽  
Significant Difference ◽  
Keratinocyte Cell ◽  
Life Threatening

In humans and dogs, toxic epidermal necrolysis (TEN) is a life-threatening dermatosis characterized by sudden epidermal death resulting in extensive skin detachment. There is little information on the pathogenesis of keratinocyte cell death in canine TEN. We studied the occurrence of apoptosis in skin lesions of dogs with TEN to determine if apoptosis contributes to the pathogenesis of this disease. Immunostaining with antibodies to activated caspase-3 and the terminal deoxynucleotidyl-transferase (TdT)–mediated deoxyuridine triphosphate (dUTP) nick-end labeling technique revealed positive apoptotic keratinocytes in basal and suprabasal epidermal compartments in 17 biopsy specimens collected from 3 dogs with TEN and 16 from 3 dogs with erythema multiforme (EM). There was no significant difference in the number of positively stained epidermal cells between TEN and EM. These results suggest that apoptosis of epidermal keratinocytes and lymphocytic satellitosis represent one of the early steps in the pathogenesis of canine TEN, as in the human disease counterpart.

Epidermal keratinocytes form a functional skin barrier in the absence of Atg7 dependent autophagy

2013 ◽  
Vol 71 (1) ◽  
pp. 67-75 ◽  
Author(s):  
Heidemarie Rossiter ◽  
Ulrich König ◽  
Caterina Barresi ◽  
Maria Buchberger ◽  
Minoo Ghannadan ◽  
...  
Keyword(s):  
Skin Barrier ◽  
Epidermal Keratinocytes

Glycoproteins and glycosaminoglycans of cultured normal human epidermal keratinocytes

1983 ◽  
Vol 61 (1) ◽  
pp. 325-338
Author(s):  
K.W. Brown ◽  
E.K. Parkinson
Keyword(s):  
Chondroitin Sulphate ◽  
Heparan Sulphate ◽  
Sodium Dodecyl ◽  
Immunological Methods ◽  
Epidermal Keratinocytes ◽  
Immunoperoxidase Staining ◽  
Human Epidermal Keratinocytes ◽  
Molecular Weights ◽  
Keratinocyte Cell ◽  
Normal Human

[3H]glucosamine has been used to label metabolically keratinocyte cell-surface glycoconjugates. The major labelled bands identified on sodium dodecyl sulphate/polyacrylamide gels had apparent molecular weights of greater than 250 000, and 150 000-80 000. Most of these components were trypsin-sensitive, indicating that the label was protein-bound. Some of the labelled components were shown to be proteoglycans and the labelled glycosaminoglycans released from them by trypsin were identified as hyaluronic acid (54%), heparan sulphate (33%) and chondroitin sulphate (13%). Specific immunological methods (immunoperoxidase staining and immunoprecipitation) showed that keratinocytes produced fibronectin. Immunoperoxidase staining showed keratinocytes produce only small ‘stitches’ of fibronectin at cell edges; no large fibrils were seen nor any staining over or between cells.

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