RESEARCH

Faculty Research Areas

David T. Woodley, M.D.

• Purification of Epidermolysis Bullosa Acquisita Antigen
• Mechanism of Keratinocyte Motility
• Re-epithelialization of Skin Wounds
• Animal Model for Acquired Autoimmune Disease Epidermolysis Bullosa Acquisita

Mei Chen, Ph.D.

• Strategies Toward Gene Therapy for Dystrophic Epidermolysis Bullosa
• Structure and Function of Type VII Collagen
• Animal Model for Acquired Autoimmune Disease Epidermolysis Bullosa Acquisita

Wei Li, Ph.D.

• Signal Transduction in Migrating Human Keratinocytes
• Mechanisms of Human Dermal Fibroblast Migration
• Re-epithelialization of Skin Wounds

David E. Sawcer, M.D., Ph.D.

• Mechanism of keratinocyte motility
• Non-melanoma and melanoma skin cancer treatment
• Laser applications

David Peng, M.D.

• Melanoma
• Epidemiology of Skin Disease

Manjunath S. Vadmal, M.D.

• Cutaneous Mesenchymal Tumors: A systematic approach to the classification and
  histopathological diagnosis
• Biologic determinants of invasive potentially metastatic and metastatic
  cutaneous malignant melanoma; application on new immunohistochemical
  markers such as MART, MAGE1, 3, CD63, CD99 and to quantify the level of
  expression of those novel markers in melanoma that further determines the
  therapeutic outcome
• Cutaneous adnexal neoplasms with follicular differentiation

Research by Dr. Mei Chen

Dr. Chen’s current specific research involves structures in the skin called anchoring fibrils. These structures are critical for the adherence of the outer skin epidermal layer to the inner dermal connective tissue layer. Anchoring fibrils are composed of type VII collagen. When children are born with defects in the gene that encodes for type VII collagen, they develop a disease of the skin called dystrophic epidermal bullosa (DEB). DEB is characterized by skin fragility, chronic blistering of the skin, scarring and aggressive squamous cell carcinomas. It is incurable. In the last few years, Dr. Chen with her colleague, Dr. Woodley, made significant progress in developing various therapeutic strategies for DEB. First, in an effort to develop ex vivo gene therapy for DEB, we applied a highly efficient lentiviral gene delivery approach to restore type VII collagen expression in RDEB keratinocytes and fibroblasts. This corrected the RDEB cell phenotype in vitro. We then used these gene-corrected cells to regenerate a human skin equivalent transplanted onto immunodeficient mice. Human skin regenerated by gene-corrected RDEB cells demonstrated restoration of type VII collagen expression and anchoring fibril formation at the dermal-epidermal junction (DEJ) in vivo. Second, in an effort to develop a cell-based therapy for DEB, we showed that intradermal injection of normal human or gene-corrected RDEB fibroblasts into mouse skin resulted in the stable expression of human type VII collagen at the mouse DEJ. Third, in an effort to develop an in vivo gene therapy, we engineered a self-inactivating lentiviral vector expressing human type VII collagen and injected this vector intradermally into hairless, immunodeficient mice and into human DEB composite skin equivalents grafted onto immunodeficient mice. A single lentiviral vector injection provided stable type VII collagen at the BMZ for at least 3 months and reversed the DEB phenotype. Lastly, in an effort to develop a protein-based therapy for DEB, we intradermally injected human recombinant type VII collagen into mice. The injected human type VII collagen stably incorporated into the mouse’s BMZ and formed anchoring fibrils. Further, intradermal injection of recombinant type VII collagen into transplanted human DEB skin equivalents also stably restored type VII collagen expression at the BMZ in vivo and reversed RDEB disease features. Our studies provide the first evidence for using protein therapy to correct a skin disease due to a gene defect in a structural protein. All these studies resulted in the several publications in the highest caliber biomedical journals including Nature Genetics, Journal for Investigative Dermatology, Molecular Therapy and Nature Medicine. In summary, studies from last few years provide strong in vitro and in vivo evidence for potential therapeutic strategies for DEB in an intact mouse model or in mice transplanted with a human DEB skin equivalent. Prior to testing any of these approaches for DEB in humans, we need to utilize a preclinical animal model to determine the safety and efficacy of these approaches and address potential immune responses. The aims of our future studies are: 1) To verify the feasibility of protein-based therapy for DEB using DEB mouse and dog models, 2) To determine the safety and efficacy of lentiviral vector-based in vivo gene therapy in DEB animal models, 3) To validate the fibroblast-based approach for correction of RDEB defects in these animal models, 4) To characterize immune responses to a neo-antigen and develop strategies to blunt these responses in the DEB mouse model, and 5) To evaluate the feasibility of intravenous injection of gene-corrected fibroblasts that home to skin for DEB treatment. These studies will advance the prospects for therapy for DEB patients and bring therapy for DEB one step closer to reality.

Anchoring fibrils are also involved in an adult acquired disease called epidermolysis bullosa acquisita (EBA). In this disease, the patient makes IgG autoantibodies against his or her own type VII collagen in the anchoring fibrils. The result of this autoimmune disease is the same as genetic DEB, namely skin fragility, blister formation, scarring and chronic skin wounds. We has also developed animal model of EBA by passively transferring the affinity purified EBA patients’ anti-type VII collagen autoantibodies into the mice and inducing the blistering disease in the animals. This murine model, with features similar to the clinical, histological and immunological features of EBA, will be useful for the fine dissection of immunopathogenic mechanisms in EBA and for development of new therapeutic intervention.

Dr. Chen has extramural funding for these projects from her RO1 grant awarded from National Institute of Health and Dr. David Woodley’s RO1 grant from National Institute of Health (She is the Co-Principal Investigator).

Research by Dr. Wei Li

The goal of Dr. Li’s research is to understand the cell motility signaling, which initiates at the cell surface, propagates via cytosolic signaling networks and executes by newly expressed and secreted gene products. The experimental model systems are un-manipulated primary human skin cells, whose migration is essential for both physiological processes, such as skin wound healing, and pathological occurrence, such as skin cancer.

Cell migration is the result of repeated cycles of cytoskeletal-mediated protrusion, polarization, formation of adhesive contacts, cell contraction, and retraction at the trailing edge. Initiation of these sequential processes is mainly triggered by two extracellular microenvironmental cues, extracellular matrices (ECMs) and soluble growth factors (GFs), which act synergistically to stimulate optimal cell migration. In vivo, ECMs are substratum-immobilized molecules. In contrast, GFs are mostly soluble polypeptides. Cell migration towards a substratum-bound ECM gradient in the absence of growth factors is referred to as “haptotaxis”. Cell migration toward a gradient of soluble GFs is known as “chemotaxis”. Our lab shows that chemotaxis cannot occur in the absence of ECMs. In contrast, haptotaxis can occur without GFs. A schematic four-step cell cell migration model is shown below. In this model, ECMs and GFs are cell motility “triggers”, which hit the cell surface and then “vanish”. The unanswered questions are what the intracellular motility networks and the final motility “executers” are for preparation of cell migration.

The ongoing research projects in his laboratory are at three levels:

Four-Step Model for Cell Motility »

1. To identify physiological ECMs and GFs for different types of human skin cells. Knowledge gained from this study could be used to look into chronic skin wounds for possible defects in these factors or uncontrolled production of these factors during skin cancer progression.
   
   
2. To molecularly reconstitute the intracellular cell motility signaling networks, which is not a single linear pathway. They are using lentiviral gene delivery systems to up- and down- regulate any gene of our interest, alone or in combination, to gain insights into how a cell interprets a motogenic signal versus mitogenic signal, both of which often come from the same cell surface stimulus.
   
   
3. To identify the molecules which execute the final steps of motility signaling prior to cell migration in response to the initial cell surface stimulation by ECMs and GFs. We focus on secreted proteins. They purify them from serum-conditioned medium of the cells and functionally characterize them. Identification of these molecules could lead to new therapies of human skin wounds and skin cenacer.

Note: Their recent contributions include 1) human serum, but not human plasma, promotes keratinocyte migration (LANCET, 2003); 2) identification of PDGF-BB as the major factor in human blood that controls dermal fibroblasts’ motility (Mol. Biol. Cell, 2004); 3) identification of TGF-alpha as the major factor in human blood that controls epidermal keratonocytes’ motility (J. Inv. Derm, 2006); 4) protein kinase C-delta clusters at the cell leading edge and mediates PDGF-BB-stimulated fibroblast migration (J. Inv. Derm., 2006); 4) the naturally occurring “plasma-to-serum-to-plasma transition” serves as a “traffic control” for the dermal and epidermal cell motility during wound healing, in which TGF?3 in serum acts as the “traffic controller” and the cell surface levels of type II TGF? receptor operate as the “sensor” to determine the order of the skin cell migration (J. Cell. Biol. 2006). In particular, the work in our recent JCB paper was selected as an “Editor’s Choice” by Science (Science, 2006).