Tissue engineering

From Freepedia

Tissue engineering can perhaps be best defined as the use of a combination of cells, engineering materials, and suitable biochemical factors to improve or replace biological functions in an effort to effect the advancement of medicine. Probably the first definition of tissue engineering was by Langer and Vacanti who stated it to be "an interdisciplinary field that applies the principles of engineering and life sciences toward the development of biological substitutes that restore, maintain, or improve tissue function”. MacArthur and Oreffo (as cited in "References") defined tissue engineering as "understanding the principles of tissue growth, and applying this to produce functional replacement tissue for clinical use." A further description goes on to say that an "underlying supposition of tissue engineering is that the employment of natural biology of the system will allow for greater success in developing therapeutic strategies aimed at the replacement, repair, maintenance, and/or enhancement of tissue function." These more general definitions are driven in part by recent scientific progress with completely autologous approaches. That is, many groups (Nicolas L'Heureux at Cytograft Tissue Engineering, Julie Campbell at University of Queensland etc, Loex laboratories at the Universite of Laval etc.) are demonstrating functional tissue engineered devices/organs without using synthetic biomaterials/scaffolds. These recent approaches are clearly based more on an understanding of cell biology than materials science.

In 2003, the NSF published a report titled "The Emergence of Tissue Engineering as a Research Field", which gives a thorough description of the history of this field.

A typical tissue engineering solution consists of a number of parts as alluded to above. This article will discuss each part in turn, along with its implications.

Contents 1 Cells 2 Engineering materials 3 Synthesis of tissue engineering scaffolds 4 Assembly methods 5 Related topics 6 External links 7 Agencies that Support Tissue Engineering Research 8 References

Cells

Tissue engineering solves problems by using living cells as engineering materials. These could be artificial skin that includes living fibroblasts, cartilage repaired with living chondrocytes, or other types of cells used in other ways.

Cells became available as engineering materials when scientists at Geron Corp. discovered how to extend telomeres in 1998. Before this, laboratory cultures of healthy, noncancerous mammalian cells would only divide a fixed number of times, up to the Hayflick limit.

The cells are often categorized by their source. "Autologous" cells come from the same body as that to which they will be reimplanted. "Allogenic" cells come from another body. "Xenogenic" cells come from another species. "Primary" cells are from an organism. "Secondary" cells are from a cell bank.

Autologous cells have the fewest problems with rejection and pathogen transmission—however in genetic disease, suitable autologous cells are not available. In severe burns, autologous cells will not be available in sufficient quantities. Autologous cells also must be cultured from samples before they can be used. This takes time, so autologous solutions are not very quick.

From fluid tissues such as blood, cells are extracted by bulk methods, usually centrifugation or aspheresis.

From solid tissues, extraction is more difficult. Usually the tissue is minced, and then digested with the enzymes trypsin or collagenase to remove the cellular matrix that holds the cells. After that, the cells are free floating, and extracted using centrifugation or aspheresis.

Digestion with trypsin is very dependent on temperature. Higher temperatures digest the matrix faster, but create more damage. Collagenase is less temperature dependent, and damages fewer cells, but takes longer and is a more expensive reagent.

Engineering materials

Cells as found above are generally implanted or 'seeded' into an artificial structure capable of supporting three-dimensional tissue formation. Such devices, usually referred to as scaffolds, serve at least one of the following purposes:

• Enhance structural properties
• Deliver biochemical factors
• Deliver or allow delivery of vital cell nutrients
• Exert certain mechanical and biological influences to modify the behaviour of the cell phase

To achieve the goal of tissue reconstruction, scaffolds must meet some specific requirements. A high porosity and an adequate pore size are necessary to facilitate cell seeding and diffusion throughout the whole structure of both cells and nutrients. Biodegradability is essential since scaffolds need to be absorbed by the surrounding tissues without the necessity of a surgical removal. The rate at which degradation occurs has to coincide as much as possible with the rate of tissue formation: this means that while cells are fabricating their own natural matrix structure around themselves, the scaffold is able to provide structural integrity within the body and eventually it will break down leaving the neotissue, newly formed tissue which will take over the mechanical load.

Many different materials (natural and synthetic, biodegradable and permanent) have been investigated. Most of these materials have been known in the medical field before the advent of tissue engineering as a research topic, being already employed as bioresorbable sutures. Examples of these materials are collagen or some linear aliphatic polyesters.

A commonly used synthetic material is PLA - polylactic acid. This is a polyester which degrades within the human body to form lactic acid, a naturally occurring chemical which is easily removed from the body. Similar materials are polyglycolic acic (PGA) and polycaprolactone (PCL): their degradation mechanism is similar to that of PLA, but they exhibit respectively a faster and a slower rate of degradation compared to PLA.

Synthesis of tissue engineering scaffolds

A number of different methods has been described in literature for preparing porous structures to be employed as tissue engineering scaffolds. Each of these techniques presents a its own advantages, but none is devoid of drawbacks.

• Textile technologies: these techniques include all the approaches that have been succesfully employed for the preparation of non-woven meshes of different polymers. In particular non-woven polyglycolide structures have been tested for tissue engineering applications: such fibrous structures have been found useful to grow different types of cells. The principal drawbacks are related to the difficulties of obtaining high porosity and regular pore size.

• Solvent Casting & Particulate Leaching (SCPL): this approach allows the preparation of porous structures with regular porosity, but with a limited thickness. First the polymer is dissolved into a suitable organic solvent (e.g. polylactic acid could be dissolved into dichloromethane), then the solution is cast into a mold filled with porogen particles. Such porogen can be an inorganic salt like sodium chloride, crystals of saccharose, gelatin spheres or paraffin spheres. The size of the porogen particles will affect the size of the scaffold pores, while the polymer to porogen ratio is directly correlated to the amount of porosity of the final structure. After the polymer solution has been cast the solvent is allowed to fully evaporate, then the composite structure in the mold is immersed in a bath of a liquid suitable for dissolving the porogen: water in case of sodium chloride, saccharose and gelatin or an aliphatic solvent like hexane for paraffin. Once the porogen has been fully dissolved a porous structure is obtained. Other than the small thickness range that can be obtained, another drawback of SCPL lies in its use of organic solvents which must be fully removed to avoid any possible damage to the cells seeded on the scaffold.

• Gas Foaming: to overcome the necessity to use organic solvents and solid porogens a technique using gas as a porogen has been developed. First disc shaped structures made of the desired polymer are prepared by meas of compression molding using a heated mold. The discs are then placed in a chamber where are exposed to high pressure CO₂ for several days. The pressure inside the chamber is gradually restored to atmospheric levels. During this procedure the pores are formed by the carbon dioxide molecules that abandon the polymer, resulting in a sponge like structure. The main problems related to such a technique are caused by the excessive heat used during compression molding (which prohibits the incorporation of any temperature labile material into the polymer matrix) and by the fact that the pores do not form an interconnected structure.

• Emulsification/Freeze-drying: this technique does not require the use of a solid porogen like SCPL. First a synthetic polymer is dissolved into a suitable solvent (e.g. polylactic acid in dichloromethane) then water is added to the polymeric solution and the two liquids are mixed in order to obtain an emulsion. Before the two phases can separate, the emulsion is cast into a mold and quickly frozen by means of immersion into liquid nitrogen. The frozen emulsion is subsequently freeze-dried to remove the dispersed water and the solvent, thus leaving a solidified, porous polymeric structure. While emulsification and freeze-drying allows a faster preparation if compared to SCPL, since it does not require a time consuming leaching step, it still requires the use of solvents, moreover pore size is relatively small and porosity is often irregular. Freeze-drying by itself is also a commonly employed technique for the fabrication of scaffolds. In particular it is used to prepare collagen sponges: collagen is dissolved into acidic solutions of acetic acid or hydrochloric acid that are cast into a mold, frozen with liquid nitrogen then liophylized.

• Liquid-liquid phase separation: similar to the previous technique, this procedure requires the use of a solvent with a low melting point that is easy to sublime. For example dioxane could be used to dissolve polylactic acid, then phase separation is induced through the addition of a small quantity of water: a polymer-rich and a polymer-poor phase are formed. Following cooling below the solvent melting point and some days of vacuum-drying to sublime the solvent a porous scaffold is obtained. Liquid-liquid phase separation presents the same drawbacks of emulsification/freeze-drying.

• CAD/CAM Technologies: since most of the above described approaches are limited when it comes to the control of porosity and pore size, computer assisted design and manifacture techniques have been introduced to tissue engineering. First a three-dimensional structure is designed using CAD software, then the scaffold is realized by using ink-jet printing of polymer powders or through Fused Deposition Modeling of a polymer solution.

Assembly methods

One of the continuing, persistent problems with tissue engineering is to grow or construct a system of blood vessels to feed an organ. Each layer of cells must be no more than two cell-thicknesses from a source of oxygen. Use of growth factors has been ineffective.

It might be possible to print organs, or possibly entire organisms. A recent innovative method of construction uses an inkjet mechanism to print precise layers of cells in a matrix of thermoreversable gel. Endothelial cells, the cells that line blood vessels, have been printed in a set of stacked rings. When incubated, these fused into a tube.

Related topics

• Biomedical engineering is a closely related (and often regarded as parental) field.
• Biological engineering is a broader field that generally encompasses tissue engineering and related fields (eg biomaterials).

External links

• Tissue engineering
• Pittsburgh Tissue Engineering Initiative
• Tissue Engineering Pages
• Dentigenix - Dental Tissue Engineering
• Cytograft - Tissue Engineered Blood Vessels

Agencies that Support Tissue Engineering Research

• National Institutes of Health
• National Science Foundation

References

• Langer, R & Vacanti JP, Tissue Enginering, Science 260(5110), 920-6, 1993 (more than 1000 quotations!)
• MacArthur, B. D. & Oreffo, R. O. C. (6 January 2005). Bridging the gap. In Nature, 433, 19.
• R.M. Nerem (from Principles of Tissue Engineering. Lanza, Langer and J Vacanti (eds), 2000)
• http://www.nsf.gov/pubs/2004/nsf0450/start.htm "The Emergence of Tissue Engineering as a Research Field"
• cell selection for tissue engineering.
• Peter X. Ma: Scaffolds for tissue fabrication - Materials Today, May 2004, 30-40
• Mikos A. G. & Temenoff J. S., "Formation of highly porous biodegradable scaffolds for tissue engineering" Electronic Journal of Biotechnology Vol.3 No.2, August 2000, 114-119 [1]