Development of silkworm ( Bombyx mori ) as a platform for producing - PowerPoint PPT Presentation

Development of silkworm ( Bombyx mori ) as a platform for producing biomaterials and growth factors for Tissue Engineering José Luis Cenis (IMIDA, La Alberca, Murcia, Spain)

THE SILK IN SPAIN

THE SILK IN SPAIN SERICICULTURE ACTIVITY IN SPAIN IN XX CENTURY • Establishment of the Sericicultural Station of Murcia in 1892 to provide technology, on the model of the Padova Sericicultural Station Production of raw cocoon in Spain 1.500 Tm Peak 1925: 1.170 Tm Low 1930s: 100 Tm End of activity: 1976 1.000 Tm 500 Tm 1900 1910 1920 1930 1940 1950 1960 1970 1980

NON TEXTILE USES OF SILK: BIOMATERIALS FOR BIOMEDICINE David L. Kaplan, Tufts University, Medford, MA, USA.

NON TEXTILE USES OF SILK: EXPRESSION OF RECOMBINANT PROTEINS WITH BACULOVIRUS VECTORS IN BOMBYX MORI BODY Through the innoculation of insect larvae w ith a recom binant baculovirus vector it is possible to get protein expression levels higher than in an insect cell culture at a low er cost

PREPARATION OF AQUEOUS SOLUTION OF FIBROIN Rearing of Silkworms Washing of sericin Disolution of fibroin Dialysis in water. Presentations of fibroin: Aqueous solution of fibroin -Films -Porous 3D Sponges -Electrospun nanofibers -Hydrogels -Micro and nanoparticles -Tubular structures -Extruded Fibers

PRESENTATION OF FIBROIN BIOMATERIALS POROUS SPONGES 3D • Scaffold for growing mesenchymal stem cells for reparation of bone Inmersion in Addition de ClNa, methanol for Fibroin in 10% 800 micr., in a insolubilizing the solution mold fibroin. Washing of salt with water Bar: 500 Μμ.

1 . PROJECT I N THE I MI DA W I TH POROUS SPONGES 3 D 1 . PROJECT I N THE I MI DA W I TH POROUS SPONGES 3 D Developm ent of scaffolds of fibroin prem ineralized w ith tricalcium Developm ent of scaffolds of fibroin prem ineralized w ith tricalcium phosphate and seeded w ith m esenchym al cells for bone reparation phosphate and seeded w ith m esenchym al cells for bone reparation Seeding with adult mesenchymal cells. 72 hours/ 1 week

PRESENTATION OF FIBROIN BIOMATERIALS NANOFIBER MATS OBTAINED BY ELECTROSPINNING • 100 to 400 nanometers of diameter • 5 micrometers of pore • Similar configuration than extracellular matrix Growth of human cells on a non-woven silk fibroin net: a potential for use in tissue engineering. R. Unger et al. (2004). Biomaterials 25: 1069-1075.

2 . PROJECT I N THE I MI DA W I TH ELECTROSPUN NANOFI BERS 2 . PROJECT I N THE I MI DA W I TH ELECTROSPUN NANOFI BERS Developm ent of nanofiber m ats for grow ing keratynocytes and Developm ent of nanofiber m ats for grow ing keratynocytes and fibroblasts for skin substitutes fibroblasts for skin substitutes Seeding with cells CCMv1Lu

Seeding with adult mesenchymal cells. 72 hours Seeding with adult mesenchymal cells. 1 week

PROTEIN EXPRESSION IN BOMBYX MORI BODY

3 . PROJECT I N THE I MI DA W I TH EXPRESSI ON OF PROTEI N I N 3 . PROJECT I N THE I MI DA W I TH EXPRESSI ON OF PROTEI N I N BOMBYX MORI W I TH BACULOVI RUS VECTORS BOMBYX MORI W I TH BACULOVI RUS VECTORS Four proteins are currently expressed with the silkworm/baculovirus platform: • Canine Interferon-alpha • Dihydrofolate Reductase • Fibroblast Growth Factor basic (bFGF) • Vascular Endothelial Growth Factor (VEGF)

DIHYDROFOLATE REDUCTASE

PACKAGE OF APPLI CATI ONS OF SI LK PROTEI NS I N TI SSUE ENGI NEERI NG PACKAGE OF APPLI CATI ONS OF SI LK PROTEI NS I N TI SSUE ENGI NEERI NG TISSUES AND ORGANS STEM CELLS SCAFFOLDS Factores de crecimiento PUPA FIBROIN SERICIN • Used as a biorreactor • Used for the stimulation of the • Used for the fabrication for the expression of of at least three cellular proliferation in vitro. It is two growth factors: used as a component in cell culture configurations of bFGF and VEGF scaffolds: film, porous and cryopreservation media as a sponges and electrospun substitute of FBS (fetal bovine serum) at 1/200 of the cost. nanofiber mats.

The production of proteins for Tissue Engineering provides a potential source of income for silkworm rearing The proof of concept described is made in cooperation with a network of Institutes of Regenerative Medicine A new facility has been built at the IMIDA for this specific development to produce silkworm, silk and scaffolds under GMPs conditions Josel.cenis@carm .es w w w .im ida.es I MI DA Estación Sericicola c/ Mayor, 1 3 0 1 5 0 La Alberca ( Murcia) Spain