Monitoring hyperproliferative disorders in human skin: Flow cytometry of changing cytokeratin expression

Franssen, M.E.J.; Boezeman, J.B.M.; Kerkhof, P.C.M. van de; Erp, P.E.J. van

doi:10.1002/cyto.b.10069

Cited by 14 publications

(10 citation statements)

References 25 publications

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“…K16 expression, which is not expressed in healthy epidermis, was first explained as the specific result of hyperproliferation. However, more recent studies show K16 expression to be a marker of general trauma and stress of the skin ([37], [38]. Elafin and hBD-2 are anti-microbial peptide/proteins secreted by keratinocytes, they constitute part of the innate immune defense and participate in skin protection against invading microorganisms [39].…”

Section: Discussionmentioning

confidence: 99%

Humanized Mouse Model of Skin Inflammation Is Characterized by Disturbed Keratinocyte Differentiation and Influx of IL-17A Producing T Cells

et al. 2012

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Humanized mouse models offer a challenging possibility to study human cell function in vivo. In the huPBL-SCID-huSkin allograft model human skin is transplanted onto immunodeficient mice and allowed to heal. Thereafter allogeneic human peripheral blood mononuclear cells are infused intra peritoneally to induce T cell mediated inflammation and microvessel destruction of the human skin. This model has great potential for in vivo study of human immune cells in (skin) inflammatory processes and for preclinical screening of systemically administered immunomodulating agents. Here we studied the inflammatory skin response of human keratinocytes and human T cells and the concomitant systemic human T cell response.As new findings in the inflamed human skin of the huPBL-SCID-huSkin model we here identified: 1. Parameters of dermal pathology that enable precise quantification of the local skin inflammatory response exemplified by acanthosis, increased expression of human β-defensin-2, Elafin, K16, Ki67 and reduced expression of K10 by microscopy and immunohistochemistry. 2. Induction of human cytokines and chemokines using quantitative real-time PCR. 3. Influx of inflammation associated IL-17A-producing human CD4+ and CD8+ T cells as well as immunoregulatory CD4+Foxp3+ cells using immunohistochemistry and -fluorescence, suggesting that active immune regulation is taking place locally in the inflamed skin. 4. Systemic responses that revealed activated and proliferating human CD4+ and CD8+ T cells that acquired homing marker expression of CD62L and CLA. Finally, we demonstrated the value of the newly identified parameters by showing significant changes upon systemic treatment with the T cell inhibitory agents cyclosporine-A and rapamycin.In summary, here we equipped the huPBL-SCID-huSkin humanized mouse model with relevant tools not only to quantify the inflammatory dermal response, but also to monitor the peripheral immune status. This combined approach will gain our understanding of the dermal immunopathology in humans and benefit the development of novel therapeutics for controlling inflammatory skin diseases.

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Section: Discussionmentioning

confidence: 99%

Humanized Mouse Model of Skin Inflammation Is Characterized by Disturbed Keratinocyte Differentiation and Influx of IL-17A Producing T Cells

et al. 2012

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“…Routine flow cytometry, for example, allows the detection of up to 17 fluorescence colors in one probe (multiplex or polychromatic flow cytometry) [21] . Monitoring of enzymatically disintegrated skin biopsies by multiparameter flow cytometry has been used highly successfully in the study of hyperproliferative skin disorders [22] . Most histopathology and dermatohistology laboratories are currently utilizing immunofluorescence or immunohistochemistry, employing at most double or triple labeling in one tissue section, sometimes using digital image analysis [23] .…”

Section: Discussionmentioning

confidence: 99%

The CD11a Binding Site of Efalizumab in Psoriatic Skin Tissue as Analyzed by Multi-Epitope Ligand Cartography Robot Technology

Bonnekoh

Böckelmann

Pommer

et al. 2006

Skin Pharmacol Physiol

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Efalizumab (Raptiva^TM) is an immunomodulating recombinant humanized IgG1 monoclonal antibody that binds to CD11a, the α-subunit of leukocyte function antigen-1 (LFA-1). By blocking the binding of LFA-1 to ICAM-1, efalizumab inhibits the adhesion of leukocytes to other cell types and interferes with the migration of T lymphocytes to sites of inflammation (including psoriatic skin plaques). Analysis of the response in patients treated with efalizumab to date shows that distinct groups of responders and nonresponders to the drug exist. It would therefore be of great practical value to be able to predict which patients are most likely to respond to treatment, by identifying key parameters in the mechanism of action of efalizumab. Detailed investigation and detection of multiple epitopes in microcompartments of skin tissue has until recently been restricted by the available technology. However, the newly developed technique of Multi-Epitope Ligand Cartography (MELC) robot technology combines proteomics and biomathematical tools to visualize protein networks at the cellular and subcellular levels in situ, and to decipher cell functions. The MELC technique, which is outlined in this paper, was used to help characterize the binding of efalizumab to affected and unaffected psoriatic skin as compared to normal control skin under ex vivomodel conditions. Efalizumab was labeled with fluorescein isothiocyanate and integrated into a MELC library of more than 40 antibodies. These antibodies were selected for their potential to detect epitopes which may be indicative of (a) various cell types, (b) structural components of the extracellular matrix, or (c) the processes of cell proliferation, activation and adhesion. Efalizumab bound to CD11a in affected psoriatic skin by a factor 15× and 32× higher than in unaffected psoriatic skin and normal control skin, respectively. CD11a and the efalizumab binding site were primarily expressed in the extravascular dermis, whereas CD54 (ICAM-1) as its ligand was most prevalent in the dermal vessels. T lymphocytes (for which the markers were CD3, CD8, CD4, and CD45R0) were the major cellular targets of efalizumab. In contrast, NK cells were only a minor target of efalizumab. Our study demonstrated that efalizumab represents a treatment for psoriasis that primarily targets memory CD4+ and CD8+ T cells and has a high specificity for psoriatic disease activity. Moreover, we hereby introduce the novel principle of a biological drug-binding biochip assay being especially useful for the future monitoring of psoriatic skin lesions under efalizumab treatment conditions.

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“…Finally, the cells were fixed and resuspended at 1 · 10 6 cells/L for flow cytometry analysis of expression. 20 …”

Section: Flow Cytometric Analysis Of the Keratin-10 (K10) Expressionmentioning

confidence: 99%

Helium-Neon Laser Irradiation Promotes the Proliferation and Migration of Human Epidermal Stem Cells In Vitro: Proposed Mechanism for Enhanced Wound Re-epithelialization

Liao

Xie

Liu

et al. 2014

Photomedicine and Laser Surgery

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Objective: The present study was conducted to investigate the effects of helium-neon (He-Ne) laser irradiation on the proliferation, migration, and differentiation of cultured human epidermal stem cells (ESCs). Background data: A He-Ne laser with a wavelength of 632.8 nm is known to have photobiological effects, and is widely used for accelerating wound healing; however, the cellular mechanisms involved have not been completely understood. Methods: The ESCs were prepared from human foreskin, and irradiated by using He-Ne laser at 632.8 nm with 2 J/cm 2 . The ESC proliferation, migration, and differentiation were examined by using XTT assay, scratch assay, and flow cytometry technology, respectively. The phosphorylation of extracellular signalregulated kinases (ERK) was analyzed by using Western blotting. Results: He-Ne laser irradiation markedly promoted cell proliferation and migration accompanied by an increase in the phosphorylation of ERK, but did not significantly influence cell differentiation. Conclusion: Our data indicated that photostimulation with a HeNe laser resulted in a significant increase in human ESC proliferation and migration in vitro, which might contribute, at least partially, to accelerated wound re-epithelialization by low-level laser therapy.

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Monitoring hyperproliferative disorders in human skin: Flow cytometry of changing cytokeratin expression

Cited by 14 publications

References 25 publications

Humanized Mouse Model of Skin Inflammation Is Characterized by Disturbed Keratinocyte Differentiation and Influx of IL-17A Producing T Cells

Humanized Mouse Model of Skin Inflammation Is Characterized by Disturbed Keratinocyte Differentiation and Influx of IL-17A Producing T Cells

The CD11a Binding Site of Efalizumab in Psoriatic Skin Tissue as Analyzed by Multi-Epitope Ligand Cartography Robot Technology

Helium-Neon Laser Irradiation Promotes the Proliferation and Migration of Human Epidermal Stem Cells In Vitro: Proposed Mechanism for Enhanced Wound Re-epithelialization

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