Sunday, October 10, 2004

Capillary Formation in Bioengineered Human Skin Constructs (BHSC) Designed to Study Burn Injury

Sanjay Kumar, MD, Jean-Luc Van Tran, MS, Josef Hadeed, MD, Emily Bellavance, MD, Louise Strande, MS, Riva Eydelman, BS, Martha Matthews, MD, Steven W Marra, MD, and Charles Hewitt, PhD.

Introduction: Bioengineered human skin constructs (BHSC), comprised of human fibroblasts and keratinocytes on a scaffold of collagen (Alloderm®), have been used successfully as an animal alternative in burn research. In order to make the model more representative of human skin, we hypothesized that endothelial cells introduced into the BHSC would form capillary beds.

Methods: Human microvascular endothelial cells (HMVEC) were plated onto the BHSC and induced to grow with the addition of growth factors hEGF, hFGF-b, VEGF, IGF and Vitamin C. Samples were fixed at weeks 2 and 3. These samples were stained with H&E and immunostained for the adhesion molecules ICAM and VCAM as well as for the endothelial cell receptor Tie-1. Digital image analysis (DIA) was used to quantify the amount of immunostain within the samples by calculating the average intensity stain index (AISI).

Results: Progressive infiltration of HMVEC was noted from weeks one to three. In areas with significant endothelial cell infiltration, capillary formation was dramatic. For the VCAM immunostain, at week 2, the AISI was 26.4 and 49.0 at week 3. For Tie-1, the AISI was 48.2 at week 2 and 66.8 at week 3. For ICAM, the AISI was 48.2 at week 2 and 36.3 at week 3.

Conclusion: HMVEC do infiltrate BHSCs and appear to form capillaries and neo-vascularization. DIA confirmed the marked presence of endothelial cells. The addition of endothelial cells, the vascular component of tissues, to the burn model greatly enhances the utility and relevance of the BHSC.

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