Transgenic human tissue: a powerful new tool for research and therapeutic targeting.

Lead Research Organisation: University of Sheffield
Department Name: Medicine and Biomedical Science

Abstract

Tissue engineering is a rapidly advancing medical technique that uses human cells and specialised materials to create living human tissues or organs in the laboratory. Such tissue can be used to repair diseased or damaged tissues in patients. Current examples of this include bone, skin, cartilage and bladder; however it is hoped that more complex tissues and organs will one day also be created, decreasing the demand for organ transplantation. We have combined this technology with that of gene therapy to create genetically modified cartilage that is resistant to the destructive processes of arthritis. Such tissue may be more effective in repairing cartilage in these patients. Based on this technique, but incorporating new technology known as 'RNA interference' we are now in a unique position to create human tissue in which any specific protein can be 'knocked-out'. This tissue has the potential to be a powerful research tool that could be used to determine the role of any molecule in normal or diseased human tissues. In recent years this task has, to a large extent, been fulfilled by genetically engineered mice. However production of such 'transgenic' animals can be controversial and is also expensive and time consuming. More importantly, animal models do not always correctly reflect the situation in humans. The aim of this research proposal is to create novel human transgenic tissue engineered cartilage and demonstrate that it can be used as a model to determine which specific molecules are involved in tissue destruction in arthritis. This should be of great benefit in designing specific drugs for the treatment of this disease. Successful application of this technology in cartilage should be equally applicable to other engineered tissues such as bone and skin and could be of great scientific and therapeutic interest.

Technical Summary

Intensive research effort is being focussed on tissue engineering as a means of repairing damaged or diseased organs and tissues. We have extended this process, using gene therapy based technology, to create stable transgenic tissue engineered cartilage that over-expresses a protease inhibitor, enabling the tissue to resists inflammatory damage. We are now in a unique position to combine this technique with stable RNA interference, to create a powerful new research tool for determining the normal and pathological role of specific proteins in human tissues. Such tissue would provide a much needed human alternative to transgenic mice which, although having proved highly effective in this regard in recent years, are expensive and time consuming to create and may sometimes display species specific differences that render results obtained misleading. As proof of concept, we aim to create transgenic 'knock-down' human tissue engineered cartilage to verify recent results obtained using transgenic mice that have identified one specific member of the ADAMTS family of proteinases, ADAMTS5, as being critical in the breakdown of aggrecan in arthritis. Aggrecan is one of two key structural components of articular cartilage and targeting this enzyme therapeutically may, therefore, be of great value in treating or preventing this disease. However several other members of this family have also been shown to possess aggrecanase activity and evidence from other animal models and human cell cultures that ADAMTS5 is the key enzyme in arthritis is more ambiguous, suggesting there may be species specific variation. Before large scale investment in ADAMTS5 targeting is contemplated, it would be of great value to demonstrate that this enzyme is indeed the culprit in human arthritis. Successful application of this technology could pave the way for the production of various transgenic human tissues for both academic and pharmacological research.

Publications

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Coughlan T (2010) Lentiviral shRNA knock-down of ADAMTS-5 and -9 restores matrix deposition in 3D chondrocyte culture in Journal of Tissue Engineering and Regenerative Medicine