Dispase-Mediated Basal Detachment of Cultured Keratinocytes Induces Urokinase-type Plasminogen Activator (uPA) and Its Receptor (uPA-R, CD87)

Schaefer, Birgit M.; Reinartz, Jeannette; Bechtel, Michael; Inndorf, Stefan; Lang, Ekkehard; Kramer, Michael D.

doi:10.1006/excr.1996.0323

Cited by 12 publications

(7 citation statements)

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“…Inspired by this concept we have previously analyzed alterations of the gene expression pro®le induced in differentiated keratinocyte cultures upon dispase-mediated detachment from the growth substratum. By using immunohistochemistry and northern blotting we found upregulation of a complex array of proliferation-and migrationrelated molecules (Schaefer et al, 1996(Schaefer et al, , 2000Grondahl-Hansen et al, 1988;McNeill and Jensen, 1990;Romer et al, 1991Romer et al, , 1994. Collectively, our ®ndings indicated that dispase-induced detachment induces a reaction, which at ®rst approximation resembles a process termed keratinocyte ``activation'' (Coulombe et al, 1997;Schaefer et al, 2000).…”

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confidence: 60%

The Membrane-Associated Protein pKe#192/MAP17 in Human Keratinocytes

Jaeger

Schaefer²,

Wallich³

et al. 2000

Journal of Investigative Dermatology

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confidence: 60%

The Membrane-Associated Protein pKe#192/MAP17 in Human Keratinocytes

Jaeger

Schaefer²,

Wallich³

et al. 2000

Journal of Investigative Dermatology

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“…Conventionally, cell populations are expanded in vitro through repeated cycles of cell harvesting via protease treatment, which have undesirable effects on cells 12, 13. In our technique, cells were only subjected to one enzymatic treatment, that is, during cell isolation.…”

Section: Discussionmentioning

confidence: 99%

Assimilating cell sheets and hybrid scaffolds for dermal tissue engineering

Tham

Lim

et al. 2005

J Biomedical Materials Res

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Cell sheets can be used to produce neo-tissue with mature extracellular matrix. However, extensive contraction of cell sheets remains a problem. We devised a technique to overcome this problem and applied it to tissue engineer a dermal construct. Human dermal fibroblasts were cultured with poly(lactic-co-glycolic acid)-collagen meshes and collagen-hyaluronic acid foams. Resulting cell sheets were folded over the scaffolds to form dermal constructs. Human keratinocytes were cultured on these dermal constructs to assess their ability to support bilayered skin regeneration. Dermal constructs produced with collagen-hyaluronic acid foams showed minimal contraction, while those with poly(lactic-co-glycolic acid)-collagen meshes curled up. Cell proliferation and metabolic activity profiles were characterized with PicoGreen and AlamarBlue assays, respectively. Fluorescent labeling showed high cell viability and F-actin expression within the constructs. Collagen deposition was detected by immunocytochemistry and electron microscopy. Transforming Growth Factor-alpha and beta1, Keratinocyte Growth Factor and Vascular Endothelial Growth Factor were produced at various stages of culture, measured by RT-PCR and ELISA. These results indicated that assimilating cell sheets with mechanically stable scaffolds could produce viable dermal-like constructs that do not contract. Repeated enzymatic treatment cycles for cell expansion is unnecessary, while the issue of poor cell seeding efficiency in scaffolds is eliminated.

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“…Keratinocytes grown under differentiating conditions in vitro develop into a multilayered ''sheet'' with epidermis-like tissue architecture. In such cultures dispase-mediated detachment induces the expression of uPA and uPA-R (12). Given this finding we hypothesized that dispasemediated detachment may induce a similar activation reaction as observed in vivo.…”

Section: Introductionmentioning

confidence: 78%

“…65136, Progen; Heidelberg, FRG) and a v -integrin (No. A049, Biomol; Hamburg; FRG) as described previously (12). The sections were incubated with the primary antibody, which then was detected by using biotin-labeled goat anti-mouse IgG antibodies and Cy3-labeled streptavidin.…”

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Immunohistochemical and molecular characterization of cultured keratinocytes after dispase‐mediated detachment from the growth substratum

Schaefer

Wallich

Schmolke

et al. 2000

Experimental Dermatology

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Keratinocyte activation comprises changes in protein and gene expression pattern resulting in phenotypic and functional changes necessary for re-epithelialization such as the expression of urokinase-type plasminogen activator (uPA) and its cell surface receptor (uPA-R; CD87). As uPA and uPA-R are rapidly induced after dispase-mediated detachment of cultured normal human epidermal keratinocytes (NHEK) we hypothesized that dispase-mediated detachment may cause a similar "activation" of keratinocytes with uPA and uPA-R being only one aspect of a complex "activation reaction". To test this hypothesis we have comparatively analysed adherent versus detached keratinocyte sheets for selected indicators of keratinocyte activation by immunohistochemistry. Furthermore we have identified genes via subtraction cloning which are up-regulated upon dispase-induced detachment. The analyses provided evidence for an increased transcriptional and translational activity in detached keratinocytes, as indicated by over-expression of several ribosomal components (L3 and S10 ribosomal protein) and transcription factors (initiation factor 4A, elongation factor 1alpha). Increased proliferative activity was indicated by increased expression of the proliferation markers Ki67, keratin 6 and keratin 17. Finally, several markers of keratinocyte activation such as the integrin chain alpha(v), psoriasin, glutathion-S-transferase and heparin-binding epidermal growth factor-like growth factor were up-regulated. Furthermore mevalonate kinase, a molecule as yet unknown to be expressed in keratinocytes, was identified. The findings provide evidence that dispase-mediated detachment in cultured keratinocytes induces a reaction, which comprises the up-regulation of a complex array of proliferation- and migration-related molecules. The pattern of which resembles the activation reaction observed in the re-epithelializing keratinocytes in vivo.

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Dispase-Mediated Basal Detachment of Cultured Keratinocytes Induces Urokinase-type Plasminogen Activator (uPA) and Its Receptor (uPA-R, CD87)

Cited by 12 publications

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The Membrane-Associated Protein pKe#192/MAP17 in Human Keratinocytes

The Membrane-Associated Protein pKe#192/MAP17 in Human Keratinocytes

Assimilating cell sheets and hybrid scaffolds for dermal tissue engineering

Immunohistochemical and molecular characterization of cultured keratinocytes after dispase‐mediated detachment from the growth substratum

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