scholarly journals Expression of beta 1, beta 3, beta 4, and beta 5 integrins by human epidermal keratinocytes and non-differentiating keratinocytes.

1991 ◽  
Vol 115 (3) ◽  
pp. 829-841 ◽  
Author(s):  
J C Adams ◽  
F M Watt
Keyword(s):  
Expression Profiles ◽  
Basal Layer ◽  
Cell Types ◽  
Epidermal Keratinocytes ◽  
Integrin Expression ◽  
Adhesive Properties ◽  
Human Epidermal Keratinocytes ◽  
Normal Keratinocytes ◽  
Beta 1 Integrin ◽  
Alpha V Beta 3

We have compared the adhesive properties and integrin expression profiles of cultured human epidermal keratinocytes and a strain of nondifferentiating keratinocytes (ndk). Both cell types adhered to fibronectin, laminin, and collagen types I and IV, but ndk adhered more rapidly and at lower coating concentrations of the proteins. Antibody blocking experiments showed that adhesion of both cell types to fibronectin was mediated by the alpha 5 beta 1 integrin and to laminin by alpha 3 beta 1 in synergy with alpha 2 beta 1. Keratinocytes adhered to collagen with alpha 2 beta 1, but an antibody to alpha 2 did not inhibit adhesion of ndk to collagen. Both cell types adhered to vitronectin by alpha v-containing integrins. Immunoprecipitation of surface-iodinated and metabolically labeled cells showed that in addition to alpha 2 beta 1, alpha 3 beta 1, and alpha 5 beta 1, both keratinocytes and ndk expressed alpha 6 beta 4 and alpha v beta 5. ndk expressed all these integrins at higher levels than normal keratinocytes. ndk, but not normal keratinocytes, expressed alpha v beta 1 and alpha v beta 3; they also expressed alpha 1 beta 1, an integrin that was not consistently detected on normal keratinocytes. Immunofluorescence experiments showed that in stratified cultures of normal keratinocytes integrin expression was confined to cells in the basal layer; terminally differentiating cells were unstained. In contrast, all cells in the ndk population were integrin positive. Our observations showed that the adhesive properties of ndk differ from normal keratinocytes and reflect differences in the type of integrins expressed, the level of expression and the distribution of integrins on the cell surface. ndk thus have a number of characteristics that distinguish them from normal basal keratinocytes.

Decreased expression of fibronectin and the alpha 5 beta 1 integrin during terminal differentiation of human keratinocytes

1991 ◽  
Vol 98 (2) ◽  
pp. 225-232 ◽  
Author(s):  
L.J. Nicholson ◽  
F.M. Watt
Keyword(s):  
Messenger Rna ◽  
Basal Layer ◽  
Terminal Differentiation ◽  
Human Keratinocytes ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Fibronectin Receptor ◽  
Beta 1 Integrin ◽  
Differentiation Marker ◽  
Unit Gravity Sedimentation

We have examined the expression of fibronectin and the alpha 5 beta 1 fibronectin receptor during terminal differentiation of human epidermal keratinocytes, using involucrin as a terminal differentiation marker. The levels of mRNAs encoding fibronectin and the alpha 5 and beta 1 integrin subunits were measured in keratinocyte populations that had been enriched for involucrin-negative or -positive cells by unit gravity sedimentation or suspension-induced terminal differentiation. All three mRNAs decreased in abundance during terminal differentiation, and the corresponding proteins were localised by immunofluorescence to the basal layer in stratified colonies. We also examined expression in ndk, a strain of epidermal cells with a complete block in terminal differentiation, which, as a result, do not express involucrin. Messenger RNA levels for fibronectin and the alpha 5 and beta 1 subunits were higher in ndk, than in unfractionated keratinocytes and the corresponding proteins were expressed by all ndk, consistent with a basal keratinocyte phenotype. We conclude that expression of fibronectin and the alpha 5 beta 1 fibronectin receptor decreases during terminal differentiation and that such changes are likely to play a role in the selective migration of terminally differentiating cells from the basal epidermal layer.

scholarly journals Differential and Overlapping Effects of 20,23(OH)2D3 and 1,25(OH)2D3 on Gene Expression in Human Epidermal Keratinocytes: Identification of AhR as an Alternative Receptor for 20,23(OH)2D3

2018 ◽  
Vol 19 (10) ◽  
pp. 3072 ◽  
Author(s):  
Andrzej Slominski ◽  
Tae-Kang Kim ◽  
Zorica Janjetovic ◽  
Anna Brożyna ◽  
Michal Żmijewski ◽  
...  
Keyword(s):  
Gene Expression ◽  
Molecular Modeling ◽  
Vitamin D3 ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Canonical Pathway ◽  
Epidermal Keratinocytes ◽  
Hydroxyl Groups ◽  
Dependent Manner ◽  
Human Epidermal Keratinocytes

A novel pathway of vitamin D activation by CYP11A has previously been elucidated. To define the mechanism of action of its major dihydroxy-products, we tested the divergence and overlap between the gene expression profiles of human epidermal keratinocytes treated with either CYP11A1-derived 20,23(OH)2D3 or classical 1,25(OH)2D3. Both secosteroids have significant chemical similarity with the only differences being the positions of the hydroxyl groups. mRNA was isolated and examined by microarray analysis using Illumina’s HumanWG-6 chip/arrays and subsequent bioinformatics analyses. Marked differences in the up- and downregulated genes were observed between 1,25(OH)2D3- and 20,23(OH)2D3-treated cells. Hierarchical clustering identified both distinct, opposite and common (overlapping) gene expression patterns. CYP24A1 was a common gene strongly activated by both compounds, a finding confirmed by qPCR. Ingenuity pathway analysis identified VDR/RXR signaling as the top canonical pathway induced by 1,25(OH)2D3. In contrast, the top canonical pathway induced by 20,23(OH)2D3 was AhR, with VDR/RXR being the second nuclear receptor signaling pathway identified. QPCR analyses validated the former finding by revealing that 20,23(OH)2D3 stimulated CYP1A1 and CYP1B1 gene expression, effects located downstream of AhR. Similar stimulation was observed with 20(OH)D3, the precursor to 20,23(OH)2D3, as well as with its downstream metabolite, 17,20,23(OH)3D3. Using a Human AhR Reporter Assay System we showed marked activation of AhR activity by 20,23(OH)2D3, with weaker stimulation by 20(OH)D3. Finally, molecular modeling using an AhR LBD model predicted vitamin D3 hydroxyderivatives to be good ligands for this receptor. Thus, our microarray, qPCR, functional studies and molecular modeling indicate that AhR is the major receptor target for 20,23(OH)2D3, opening an exciting area of investigation on the interaction of different vitamin D3-hydroxyderivatives with AhR and the subsequent downstream activation of signal transduction pathways in a cell-type-dependent manner.

Effects of Sulfur Mustard on Cytokines Released from Cultured Human Epidermal Keratinocytes

1998 ◽  
Vol 17 (3) ◽  
pp. 223-229 ◽  
Author(s):  
Elien M. Kurt ◽  
Robert J. Schafer ◽  
Carmen M. Arroyo
Keyword(s):  
Sulfur Mustard ◽  
Cell Types ◽  
Epidermal Keratinocytes ◽  
Cytokine Release ◽  
Experimental Conditions ◽  
Human Epidermal Keratinocytes ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
Cytokine Levels ◽  
Factor Alpha

The release of the cytokines interleukin (IL)-1α, IL-1β, IL-6, IL-8, and tumor necrosis factor alpha (TNF-α was measured from epiderm alkeratinocytes in an attempt to characterize the immunologic response in keratinocytes following exposure to bis (2-chloroethyl)sulfide (sulfur mustard, HD). Enzyme-linked immunosorbentassay (ELISA) was used to measure cytokine levels in adult and neonatal culture human epidermal keratinocytes (HEK) 3 h after exposure to 0.50 and 1.0 m M HD. A two-way analysis of variance was carried out for cell type and HD concentration. That analysis showed significant differences between cell types for IL-1α and IL-1β(p =.001 and p =.015, respectively). In both of these cytokines, release in neonatal HEK decreased less than in adult HEK. A significant effectof HD concentration was shown only for IL-1β (P <.001), with cytokine release decreasing with increasing HD dose. In addition, a significant cell donor type by HD concentration interaction effect was found for IL-1β under the experimental conditions described in materials and methods. With increasing HD concentration, the relative decrease in cytokine release was much greater for adult than for neonatal HEK.

scholarly journals Evidence that cadherins play a role in the downregulation of integrin expression that occurs during keratinocyte terminal differentiation

1994 ◽  
Vol 124 (4) ◽  
pp. 589-600 ◽  
Author(s):  
KJ Hodivala ◽  
FM Watt
Keyword(s):  
De Novo ◽  
Lectin Binding ◽  
Basal Layer ◽  
Terminal Differentiation ◽  
Ion Concentration ◽  
Epidermal Keratinocytes ◽  
Integrin Expression ◽  
Standard Medium ◽  
Proliferating Cells ◽  
Low Calcium

In epidermis the onset of terminal differentiation normally coincides with inhibition of integrin function and expression, thereby ensuring that differentiating cells are selectively expelled from the basal layer. However, when stratification of cultured human epidermal keratinocytes is prevented by reducing the calcium concentration of the medium to 0.1 mM, keratinocytes initiate terminal differentiation while still attached to the culture substrate. We have examined the mechanism by which differentiating keratinocytes adhere to extracellular matrix proteins in low calcium medium and the consequences of inducing stratification by raising the calcium ion concentration to 1.8 mM (Standard Medium). In low calcium medium keratinocytes co-expressed integrins and terminal differentiation markers such as involucrin and peanut lectin-binding glycoproteins: differentiating cells contained integrin mRNA, synthesized integrin proteins de novo and expressed functional mature integrins. There were no differences in integrin synthesis, maturation or break down in low calcium or standard medium, although the level of beta 1 integrins on the surface of proliferating cells was higher in standard medium. Within 6 h of transfer from low calcium to standard medium integrin mRNA was no longer detectable in terminally differentiating cells, integrins were being lost from the cell surface, and selective migration out of the basal layer had begun. Antibodies to P- and E-cadherin, which block calcium-induced stratification, prevented the selective loss of integrin mRNA and protein from terminally differentiating cells. This suggests that cadherins may play a role in the down-regulation of integrin expression that is associated with terminal differentiation.

scholarly journals Immunochemical analysis of the integrin expression in retinal pigment epithelial cells

2020 ◽  
Author(s):  
Janosch P Heller ◽  
Tristan G Heintz ◽  
Jessica CF Kwok ◽  
Keith R Martin ◽  
James W Fawcett
Keyword(s):  
Retinal Pigment Epithelial ◽  
Expression Profiles ◽  
Retinal Pigment ◽  
Cell Types ◽  
Human Cell Line ◽  
Retinal Pigment Epithelial Cells ◽  
Proper Function ◽  
Integrin Expression ◽  
Rpe Cells ◽  
Pigment Epithelial

AbstractRetinal pigment epithelial (RPE) cells have been used in disease modelling and transplantation studies in retinal diseases. Several types of RPE cells have been trialed, ranging from primary cells and immortalized cell lines to stem cell-derived RPE cells. During aging and in disease, the extracellular environment of the RPE cells changes, interfering with RPE cell adhesion. We hypothesize that this could be a key problem in transplantation studies that have reported lack of adhesion and survival of the transplanted RPE cells. Integrins are essential for the proper function of the RPE, mediating adhesion to Bruch’s membrane, and the binding and subsequent phagocytosis of photoreceptor outer segments. Variability that has been found in clinical trials might be due to the variability of cell types used and their expression profiles of surface molecules. Here, we set out to analyze integrin expression in primary rat RPE cells and in the human cell line ARPE-19 using immunochemistry. We found that both cell types express integrins to varying degrees. After long-term culturing, ARPE-19 cells resemble mature RPE cells, and increase integrin expression. We hence argue that it is important to test the properties of these cells prior to transplantation to avoid failure of adhesion and to facilitate correct function.

scholarly journals Ultraviolet B, melanin and mitochondrial DNA: Photo-damage in human epidermal keratinocytes and melanocytes modulated by alpha-melanocyte-stimulating hormone

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 881 ◽  
Author(s):  
Markus Böhm ◽  
Helene Z. Hill
Keyword(s):  
Mitochondrial Dna ◽  
Copy Number ◽  
Cell Types ◽  
Ultraviolet B ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Mtdna Damage ◽  
Melanocyte Stimulating Hormone ◽  
Copy Numbers ◽  
Stimulating Hormone

Alpha-melanocyte-stimulating hormone (alpha-MSH) increases melanogenesis and protects from UV-induced DNA damage. However, its effect on mitochondrial DNA (mtDNA) damage is unknown. We have addressed this issue in a pilot study using human epidermal keratinocytes and melanocytes incubated with alpha-MSH and irradiated with UVB. Real-time touchdown PCR was used to quantify total and deleted mtDNA. The deletion detected encompassed the common deletion but was more sensitive to detection. There were 4.4 times more mtDNA copies in keratinocytes than in melanocytes. Irradiation alone did not affect copy numbers. Alpha-MSH slightly increased copy numbers in both cell types in the absence of UVB and caused a similar small decrease in copy number with dose in both cell types. Deleted copies were nearly twice as frequent in keratinocytes as in melanocytes. Alpha-MSH reduced the frequency of deleted copies by half in keratinocytes but not in melanocytes. UVB dose dependently led to an increase in the deleted copy number in alpha-MSH-treated melanocytes. UVB irradiation had little effect on deleted copy number in alpha-MSH-treated keratinocytes. In summary, alpha-MSH enhances mtDNA damage in melanocytes presumably by increased melanogenesis, while α-MSH is protective in keratinocytes, the more so in the absence of irradiation.

scholarly journals Peptide‐Functionalized Hydrogels: Peptide‐Functionalized Hydrogels Modulate Integrin Expression and Stemness in Adult Human Epidermal Keratinocytes (Adv. Biosys. 10/2019)

2019 ◽  
Vol 3 (10) ◽  
pp. 1970101
Author(s):  
Duncan Davis‐Hall ◽  
Vy Nguyen ◽  
Tyler J. D'Ovidio ◽  
Ethan Tsai ◽  
Ganna Bilousova ◽  
...  
Keyword(s):  
Epidermal Keratinocytes ◽  
Integrin Expression ◽  
Human Epidermal Keratinocytes ◽  
Adult Human

scholarly journals Analysis and modeling of time-course gene-expression profiles from nanomaterial-exposed primary human epidermal keratinocytes

2009 ◽  
Vol 10 (Suppl 11) ◽  
pp. S10 ◽  
Author(s):  
Amin Zollanvari ◽  
Mary Jane Cunningham ◽  
Ulisses Braga-Neto ◽  
Edward R Dougherty
Keyword(s):  
Gene Expression ◽  
Time Course ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes

scholarly journals Regulation of cell surface beta 1 integrin levels during keratinocyte terminal differentiation.

1995 ◽  
Vol 128 (6) ◽  
pp. 1209-1219 ◽  
Author(s):  
N A Hotchin ◽  
A Gandarillas ◽  
F M Watt
Keyword(s):  
Cell Surface ◽  
Intracellular Transport ◽  
De Novo ◽  
Terminal Differentiation ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Confocal Immunofluorescence Microscopy ◽  
Affinity Modulation ◽  
Confocal Immunofluorescence ◽  
Beta 1 Integrin

Integrins of the beta 1 family play a central role in controlling adhesion and terminal differentiation within the epidermis. When human epidermal keratinocytes undergo terminal differentiation, intracellular transport of newly synthesized integrins is inhibited, and mature receptors are lost from the cell surface. We have examined the mechanisms underlying these processes, using an experimental model in which keratinocytes are placed in suspension to induce terminal differentiation. The block in intracellular transport was keratinocyte- and integrin-specific since it was not observed when fibroblasts were placed in suspension and did not affect E-cadherin synthesis in suspended keratinocytes. Newly synthesized beta 1 integrins associated with an endoplasmic reticulum resident protein, calnexin; the association was prolonged when keratinocytes were placed in suspension, suggesting a role for calnexin in the inhibition of transport. After 24 h, the level of beta 1 integrin mRNA declines in suspended keratinocytes, reflecting inhibition of gene transcription, but in fibroblasts, the level remained constant. Transport of integrins could be blocked in both adherent keratinocytes and fibroblasts by inhibiting total protein synthesis, raising the possibility that transport is coupled to de novo integrin synthesis. The fate of receptors on the surface of keratinocytes was followed by confocal immunofluorescence microscopy, immunoelectron microscopy, and biochemical analysis: with the onset of terminal differentiation, endocytosed receptors were transported to the lysosomes. These experiments reveal novel mechanisms by which integrin levels can be controlled. Together with our earlier evidence for transcriptional regulation and affinity modulation of integrins, they highlight the complexity of the mechanisms which ensure that the onset of terminal differentiation is linked to detachment of keratinocytes from the underlying basement membrane.

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