Artificial Skin – Culturing of Different Skin Cell Lines for Generating an Artificial Skin Substitute on Cross-Weaved Spider Silk Fibres

Background: In the field of Plastic Reconstructive Surgery the development of new innovative matrices for skin repair is in urgent need. The ideal biomaterial should promote attachment, proliferation and growth of cells. Additionally, it should degrade in an appropriate time period without releasing harmful substances, but not exert a pathological immune response. Spider dragline silk from Nephila spp meets these demands to a large extent. Methodology/Principal Findings: Native spider dragline silk, harvested directly out of Nephila spp spiders, was woven on steel frames. Constructs were sterilized and seeded with fibroblasts. After two weeks of cultivating single fibroblasts, keratinocytes were added to generate a bilayered skin model, consisting of dermis and epidermis equivalents. For the next three weeks, constructs in co-culture were lifted on an originally designed setup for air/liquid interface cultivation. After the culturing period, constructs were embedded in paraffin with an especially developed program for spidersilk to avoid supercontraction. Paraffin cross-sections were stained in Haematoxylin & Eosin (H&E) for microscopic analyses. Conclusion/Significance: Native spider dragline silk woven on steel frames provides a suitable matrix for 3 dimensional skin cell culturing. Both fibroblasts and keratinocytes cell lines adhere to the spider silk fibres and proliferate. Guided by the spider silk fibres, they sprout into the meshes and reach confluence in at most one week. A well-balanced, bilayered cocultivation in two continuously separated strata can be achieved by serum reduction, changing the medium conditions and the cultivation period at the air/liquid interphase. Therefore spider silk appears to be a promising biomaterial for the enhancement of skin regeneration

Interactions between Spider Silk and Cells – NIH/3T3 Fibroblasts Seeded on Miniature Weaving Frames

Native spider silk does not require any modification to its application as a biomaterial that can rival any artificial material in terms of cell growth promoting properties. We could show adhesion mechanics on intracellular level. Additionally, proliferation kinetics were higher than in enzymatically digested controls, indicating that spider silk does not require modification. Recent findings concerning reduction of cell proliferation after exposure could not be met. As biotechnological production of the hierarchical composition of native spider silk fibres is still a challenge, our study has a pioneer role in researching cellular mechanics on native spider silk fibres

Bundles of Spider Silk, Braided into Sutures, Resist Basic Cyclic Tests: Potential Use for Flexor Tendon Repair

<div><p>Repair success for injuries to the flexor tendon in the hand is often limited by the in vivo behaviour of the suture used for repair. Common problems associated with the choice of suture material include increased risk of infection, foreign body reactions, and inappropriate mechanical responses, particularly decreases in mechanical properties over time. Improved suture materials are therefore needed. As high-performance materials with excellent tensile strength, spider silk fibres are an extremely promising candidate for use in surgical sutures. However, the mechanical behaviour of sutures comprised of individual silk fibres braided together has not been thoroughly investigated. In the present study, we characterise the maximum tensile strength, stress, strain, elastic modulus, and fatigue response of silk sutures produced using different braiding methods to investigate the influence of braiding on the tensile properties of the sutures. The mechanical properties of conventional surgical sutures are also characterised to assess whether silk offers any advantages over conventional suture materials. The results demonstrate that braiding single spider silk fibres together produces strong sutures with excellent fatigue behaviour; the braided silk sutures exhibited tensile strengths comparable to those of conventional sutures and no loss of strength over 1000 fatigue cycles. In addition, the braiding technique had a significant influence on the tensile properties of the braided silk sutures. These results suggest that braided spider silk could be suitable for use as sutures in flexor tendon repair, providing similar tensile behaviour and improved fatigue properties compared with conventional suture materials.</p> </div

Tensile test results of spider silk sutures of 3×60–3×120 single fibres and 1×360 native spider silk fibres.

<p>Tensile test results of spider silk sutures of 3×60–3×120 single fibres and 1×360 native spider silk fibres.</p

Supercontraction of braided sutures.

<p>Braided sutures were measured under wet conditions after being moistened with 0.9% NaCl. The resulting supercontraction (reduction of length) of the braided sutures is clearly visible. The results are highly significant, with p<0.0001.</p

Maximum load before and after cyclic testing for spider silk sutures comprised of 3×120 single fibres with an increase in strain between the first and last cycle.

<p>Maximum load before and after cyclic testing for spider silk sutures comprised of 3×120 single fibres with an increase in strain between the first and last cycle.</p

Results for cyclic tests of Prolene® 6-0 and 3×120 spider silk.

<p>A shows the cyclic testing results for Prolene® 6-0. The suture was subjected to over 1000 cycles between 0 <i>N</i> and 2.3 <i>N</i>, which was half of the mean failure load. After these 1000 cycles, Prolene® was stretched up to breaking point. The failure load after cyclic testing was compared with the previous single tensile testing (C). The results of biomechanical tests of 3×120 spider silk are shown in B. The first 100 to 300 cycles of testing did not remain under the limit of half of the failure load of 2.9 <i>N</i>. No difference in failure load before and after cyclic use was noticed.</p

Definition of the nomenclature of sutures.

<p>Definition of the nomenclature of sutures.</p

SEM of single spider silk fibres and braided spider silk sutures.

<p>A shows a braided suture of 4×60 single spider silk fibres. Fibres are clearly visible in their corresponding strands. Strands are at an angle of approximately 30° from horizontal. B shows a braided suture of 3×120 single silk fibres. C depicts a large section of harmonic braiding. D presents a typical example of the SEM images used for diameter measurements.</p