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Visualization of Morphological and Molecular Features Associated with Chronic Ischemia in Bioengineered Human Skin

Published online by Cambridge University Press:  04 March 2010

Erin M. Gill ,
Joely A. Straseski ,
Cathy A. Rasmussen ,
Sara J. Liliensiek ,
Kevin W. Eliceiri ,
Nirmala Ramanujam ,
John G. White  and
B. Lynn Allen-Hoffmann
Erin M. Gill
Affiliation:
Department of Biomedical Engineering, University of Wisconsin, Madison, WI 53706, USA Laboratory for Optical and Computational Instrumentation, University of Wisconsin, Madison, WI 53706, USA
Joely A. Straseski
Affiliation:
Department of Pathology, Johns Hopkins Medical Institutions, Baltimore, MD 21287, USA
Cathy A. Rasmussen
Affiliation:
Department of Pathology and Laboratory Medicine, University of Wisconsin Medical School, Madison, WI 53706, USA Stratatech Corporation, Research and Development, Madison, WI, USA
Sara J. Liliensiek
Affiliation:
School of Veterinary Medicine, University of Wisconsin, Madison, WI 53706, USA
Kevin W. Eliceiri
Affiliation:
Laboratory for Optical and Computational Instrumentation, University of Wisconsin, Madison, WI 53706, USA
Nirmala Ramanujam
Affiliation:
Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA
John G. White
Affiliation:
Laboratory for Optical and Computational Instrumentation, University of Wisconsin, Madison, WI 53706, USA
B. Lynn Allen-Hoffmann*
Affiliation:
Department of Pathology and Laboratory Medicine, University of Wisconsin Medical School, Madison, WI 53706, USA Stratatech Corporation, Research and Development, Madison, WI, USA
*
Corresponding author. E-mail: [email protected]
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Abstract

We present an in vitro model of human skin that, together with nonlinear optical microscopy, provides a useful system for characterizing morphological and structural changes in a living skin tissue microenvironment due to changes in oxygen status and proteolytic balance. We describe for the first time the effects of chronic oxygen deprivation on a bioengineered model of human interfollicular epidermis. Histological analysis and multiphoton imaging revealed a progressively degenerating ballooning phenotype of the keratinocytes that manifested after 48 h of hypoxic exposure. Multiphoton images of the dermal compartment revealed a decrease in collagen structural order. Immunofluorescence analysis showed changes in matrix metalloproteinase (MMP)-2 protein spatial localization in the epidermis with a shift to the basal layer, and loss of Ki67 expression in proliferative basal cells after 192 h of hypoxic exposure. Upon reoxygenation MMP-2 mRNA levels showed a biphasic response, with restoration of MMP-2 levels and localization. These results indicate that chronic oxygen deprivation causes an overall degeneration in tissue architecture, combined with an imbalance in proteolytic expression and a decrease in proliferative capacity. We propose that these tissue changes are representative of the ischemic condition and that our experimental model system is appropriate for addressing mechanisms of susceptibility to chronic wounds.

Keywords

hypoxiaischemiaulcerationbioengineered skinmultiphotonMMP-2
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 16 , Issue 2 , April 2010 , pp. 117 - 131
Copyright
Copyright © Microscopy Society of America 2010

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