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From 2013.igem.org

Silk Silk

There are many anthropoids that have the ability to produce silk, most popular being the Bombyx mori silk worm used in the production of silk for clothing. However the spider easily takes the crown in terms of applications. The Spider uses its silk as their Swiss Army Knife,some of these uses include, spinnig webs for catching preys, Dragline's for better movement, and for reproduction. Over the 400 million years of evolution the silk is optimized in many aspects. Like with many fascinating phenomena from nature, humans have learned to utilize the spider webs. Giant spiders() in tropical rain forests() have powerful webs to catch flying birds, and old cultures have used these webs for fishing purposes.

Spider silk as biomaterial

This makes spider silk a biomaterial with great potential. On one hand it has incredible high tensile strength combined with elasticity, which makes spider silk as strong as steel and tougher than kevlar. On the other hand spider silk also has some peculiar medical properties. Foremost the fact that spider silk (and silk in general) doesn’t cause a immune response in the human body. Immune response obviously being one of the major concerns when placing implants inside patients, so our iGEM team saw a great opportunity to combine the spider silk with the implants.

History

While modern day researchers are very interested in spider silk, it should be noted that since ancient times uses (and fascination) of spider silk have been recorded trough history.
The oldest known application of a spider web dates back to from Ancient Greece were it was used as wound dressing . Some other historical mentions date from around 1600. In the Polish book ‘With Fire and Sword’:

‘ “This is nothing, nothing at all” said he, feeling the wounds with his fingers. “He will be well to-morrow. I will take care of him. Mix up bread and spider-webs for me! ’

And in the famous Shakespearian comedy ‘Midsummer Night’s Dream’:

“I shall desire you of more acquaintance, good master cobweb, If I cut my finger, I shall make bold of you.”

Apparently the medical properties of spider webs have been known for a long time. Its use as wound cover improves the recovery without giving problems. What is it that gives the spider silk such excellent healing properties?

Biomedical properties

Normal cells need a scaffold to adhere to. If there is a wound the tissue is repaired very slowly, and only after that the cells from the wound are able to completely recover. When a wound is covered with a good biomedical material, the cells will start to spread, proliferate and differentiate more quickly. On an unsuitable material, the interaction can lead to immune responses and cell death.

Spider silk is suitable as biomedical material because:

1. It has no toxicity[1][2][3]. Short-term and long-term test with various cell lines have given no indication of toxicity.
2. No/low immune response[5][6][7]. Different types of cells were examined for immunological response to silk. One study compared collagen, which is used as biomaterial for coating, to silk. The results favoured silk, and deemed silk as being biocompatible[5].
3. Cell adherence. This depends on the processing of the silk. With small modifications (for instance, genetic addition of an RGD motif) cell adhesion can be improved, to stimulate tissue recovery of a wound. However, for the use of implants, cell adhesion is undesired, as fibrosis can occur.
4. Biodegradable. Spider silk has a low biodegradability, which is useful for medical applications.
5. Elasticity and mechanical properties. The spider silk material is strong and stable material. The elasticity of a biomedical material has shown to influence cell growth and proliferation, and therefore the flexible and adjustable spider silk is very potent.
6. Does not swell. Due to the nature of the protein it is both hydrophilic and hydrophobic, the latter preventing any major uptake of water.
7. No bacterial or fungal degradation[5]; this is credited to the long evolution. One can imagine that if the base of a spiders nature, its web, would be easily torn apart by the hundreds of bacteria and fungi around, then they would not have gotten this far. The silk typically has a very flat surface that does not allow cells to grow on it very easily, nor be identified by antibodies.

Studies performed with coated implants in mice and pigs support these medical properties. Also spider silk coated breast implants are tested in preclinical trials, and have so far shown promising results (AMsilk).

Spider silk is our choice to approach the problems of medical implants. However, so far no large scale production of spider silk is achieved, and production, purification and processing are challenging. Why is the production of the spider silk protein so difficult? How can we make a coating from the spider silk?

Silk protein

Spider silk protein repetitive blabla

Silk

Silk is a natural protein fibre that is known for its use in textiles. The best known silk comes from the silk moth pupa but a lot of animals such as arthropods are also capable of producing silk. One of the arthropods well known for its silk is the spider and spider silk is our material of choice for our implant coating.

The spider silk we used is the so called dragline silk from the Argiope aurantia (MaSp2) from Brooks et al (2008). It is the strongest silk produced inside the spider. The spider silk consists out of a big repetitive domain (around 2500 base pairs) with an N and a C terminus (figure 1 b).

The properties of silk

The unique properties of silk are a result of its highly constant and repetitive amino-acid structure. The sequence of amino-acids determines what secondary structures will arise, and thus the final preferred protein conformation. The secondary structures may be beta sheets, beta-spirals, and beta-helices, of which the sheets realize the silk's amazing tensile strength, and the spirals and helices its elongation.

In the figure 2 below a stress-strain diagram can be found. where Clavipus

A final general property of silk it that it can be integrated with the human body, it will not induce an immune response, making it an ideal choice for many biomedical applications. Its compatibility extends to the gastrointestinal tract, that is, it is even safe to eat.

Figure 2: Properties of spidersilk versus other materials (Frank K. Ko, et at. 2001). Spider silk is compared to, Kevlar 29, normal silkworm silk, polyethylene terephthalate (PET), Nylon 6, and Merino wool. The stress-strain diagram relates the degree of deformation to the amount of energy absorbed.

Silk as a coating material

Silk can be regarded as a superb version of collagen. Collagen is a material used a lot for the coating of implants. Although collagen is a naturally occurring protein found inside the human body, the implants are coated with proteins harvested from cows, which can give an immune response. On the other hand spider silk is an in allergic material and will never give an immune response. Spider Silk is biodegradable, has a low bacterial adherence and can rival any artificial material with the respect of cell growth (1-3). Therefore spider silk is a good candidate for the coating of medical implants.

For example one can think of a cartilage implant. The implant consists of a biodegradable polymer coated with spider silk. The human cells will grow into the implant, slowly degrading the implant. After a certain amount of time the implant is fully degraded, leaving only a new piece of cartilage in for example a knee.

References

(1) Silk-based materials for biomedical applications, A.Leal-Egana (2010)
(2) Silk as biomaterial, C. Vepari (2007)
(3) Interactions between spider silk and cells, J.W. Kuhlier (2010)