In the eighth of our weekly series of articles by NUI Galway researchers, Dr. Oliver Carroll, Research Technical Officer with the Network of Excellence for Functional Biomaterials, writes about the power of tissue engineering to help the body to repair injured or degenerated tissue.

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Various scaffolds used to promote cell growth and tissue repair: Osteoblast cultured on calcium phosphate scaffold. Collagen sponge for bone regeneration. Collagen sponge for wound healing. Nerve cells cultured on non-aligned collagen scaffold. Aligned electrospun nanofibres (Images courtesy NFB http://www.nfb.ie/research)

Cell regeneration therapy is a developing technology to meet the increasing demand to treat injured or degenerated tissue. Organ transplants are the ideal treatment for many patients with tissue damage, but the demand of organs surpasses available organs for transplantation. There are several types of cell-based regenerative therapies currently being applied, including injection of isolated cells, scaffold engineering, and cell sheet tissue engineering. 

Injecting cells to treat injured or degenerated tissues can be used to create tissue-specific extracellular matrix (ECM), i.e. the collection of extracellular molecules secreted by cells that provides structural and biochemical support to the surrounding cells. However, isolated cell injection has a low cell survival rate once transplanted into a host, which reduces their therapeutic benefits. Scaffolds made from various materials (e.g collagen, titanium or a synthetic polymer such as polylactic acid) have been used to help control the retention and distribution of transplanted cells, but can have damaging affects in sensitive organs.

Scaffold-free tissue engineering techniques such as cell sheet production have been developed, whereby an intact layer of cells is grown for eventual implantation. A variety of cell types may be used to produce a suitable cell sheet, depending on which type of tissue is damaged.

How to prepare a cell sheet

Cell sheets have been grown at 370C, a physiological temperature, and the developed cell-sheet can be detached from petri dished by lowering temperature to 200 C (author's own image)
Cell sheets have been grown at 37˚C, a physiological temperature, and the developed cell-sheet can be detached from petri dished by lowering temperature to 20˚C (Images courtesy of Abhigyan Satyam, NUI Galway)

Cell sheets are generally engineered by growing the desired cells on a thermo-responsive polymer surface. Cells proliferate on this polymer surface, creating cell-cell junctions from multi-protein complexes, which hold the sheet together. Cell sheets are typically cultured at 37˚C, which is ideal for adhesion to the polymer surface. Once the cells have proliferated sufficiently, the temperature is lowered to 20˚C, producing a water layer between the cell sheet and the polymer, allowing the intact cell sheet to dissociate from the polymer without damage.

The sheet maintains its ECM when detached from the culture surface, which is beneficial for cell adhesion to the host. Cell sheets are approximately half a millimetre in width and can be layered by placing the sheets on top of one another in order to produce a more cell-dense, three dimensional structure. The detached sheet can then be implanted into a patient and help regenerate the damaged tissue.

Faster production of engineered tissues

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Macromolecular crowding: diagram of inert macromolecules added to tissue culture flask emulating the dense in vivo extracellular space

Despite the promising outcomes for various tissue types, considerable time (up to 196 days) is required for the cells to create ECM once they are transplanted, and this often results in loss of cell function. Macromolecular crowding (MMC) for tissue engineering is an approach proposed by researchers at the Network of Excellence for Functional Biomaterials (NFB) at NUI Galway, as a means to engineer tissues faster for transplantation.

Abhigyan Satyam from the research team led by Principal Investigator Dr. Dimitrios Zeugolis is investigating MMC to increase cellular activities, thereby creating ECM-rich tissue equivalents faster. Cells in the body (in vivo) are in a naturally crowded space causing rapid collagen synthesis. However, when cells are placed in a less dense population area, such as culture conditions in a lab (in vitro), collagen production is much slower. Dr Zeugolis’s team are adding inert polydispersed macromolecules to their growing cell populations, with the goal of increasing the density of the growth area, which will, in turn, increase ECM production.

Applications of cell sheets

Applications of cell sheets (image credit: Tokyo Women's Medical University http://twins.twmu.ac.jp/first/en/technology_base.html)
Applications of cell sheets (image credit: Tokyo Women’s Medical University http://twins.twmu.ac.jp/first/en/technology_base.html)

Two of the most prominent areas of research where cell sheets have been used are in corneal reconstruction and cardiac tissue regeneration. Clinical trials prove that this technology can produce long term benefits for patients with a corneal condition known as Limbal stem-cell deficiency, where normal corneal regeneration does not occur. Furthermore, when cardiomyocyte (heart muscle) cell sheets are implanted into a host, they are able to mimic the activity of the host’s own cells, assisting in the cardiac function. Long term goals will see cell sheets being used in the construction of organ-like structures.

Pramod Kumar, a PhD student at NUI Galway, is researching the use of cell sheet technology to develop ECM-rich corneal stromal tissue, which is the most important factor in corneal engineering. He hopes to produce stromal cell sheets quickly by employing macromolecular crowding. He has previously found that cell sheet technology combined with macromolecular crowding helps in the protection and deposition of tissue-specific ECM secreted from the corneal fibroblast. He says “we can develop the corneal stromal tissue like structure in the form of ECM rich corneal fibroblast cell sheets within 4-6 days of culture”. Due to the ECM rich nature, the developed cell sheet will be used to make a full thickness cornea in combination with other corneal cells such as epithelium and endothelium.

Diana Gaspar, a PhD student at NUI Galway, is using cell sheet technology to grow sheets of cells that will be used for tendon regeneration. Tendon injuries affect around 33 million people annually worldwide and the currently available therapies do not completely restore functionality. Diana hopes to provide sheets which can be given to patients to repair tendon injuries by regenerating the damaged tissue without using foreign materials. She has already developed sheets of tendon cells and is currently investigating the use of more easily obtainable, alternative cell sources, such as skin.

Dr Sarah Gundy, Professor Abhay Pandit and An Taoiseach Enda Kenny at the official opening of the €30 million NUI Galway Biosciences Building which hosts the NFB
Dr Sarah Gundy, Professor Abhay Pandit and An Taoiseach Enda Kenny at the official opening of the €30 million NUI Galway Biosciences Building which hosts the NFB

To find out more about the research being carried out at the NFB, visit http://www.nfb.ie/research

The theme of this year’s Science Week, coordinated by SFI Discover, is ‘The Power of Science‘. Check out http://www.science.ie/ to find out what Science Week events are happening in your area between November 9-16, or why not organise your own!

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