DE102018129658A1 - Medical implant comprising magnesium and fibroin - Google Patents

Abstract

The invention relates to a biodegradable medical implant device for tissue reconstruction comprising fibroin and a magnesium source, the implant device having a three-dimensional (3D) tissue support structure. The invention also includes this implant device which is filled with a cell population. Ultimately, the invention relates to a method for producing the implant according to the invention.

Description

Field of the Invention:

The invention relates to a biodegradable medical implant device for tissue reconstruction comprising fibroin and a magnesium source, the implant device having a three-dimensional (3D) tissue support structure. The invention also includes this implant device which is filled with a cell population. Ultimately, the invention relates to a method for producing the implant according to the invention.

Background of the Invention

In an aging society, regeneration and tissue reconstruction are becoming increasingly important. In Germany, for example, around 9 million people suffer from cartilage damage or arthrosis and 2.5 million diabetics need tissue grafts. Although plastic surgery has developed improved surgical techniques, the limited availability of endogenous grafts is the main bottleneck for the use of these techniques. A promising approach to tissue reconstruction is the processing of materials into complex support structures that enable specific generation of different tissue types based on stem cells or progenitor cells. The processing of animal tissue, which is used as an alternative strategy, on the other hand, is very expensive and burdened with ethical objections.

In addition, many materials used in tissue reconstruction interfere with the rapid vascularization of the implanted cells or tissue, causing the newly generated tissue to die before the new vascular structures allow adequate nutrient transport and the physiological integrity of the tissue.

Current implant devices are limited to certain tissues. There is no synthetic support structure available specifically for muscle tissue that allows the production of such complex tissue.

The design of optimized implants for tissue reconstruction is hampered by the fact that the implant should combine different complex requirements. First, it should have a sufficiently stable support structure at the time of implantation, but should ideally be biodegradable, so that the implant structures, which are becoming smaller, are replaced by the endogenously regenerated tissue structures. Secondly, the materials and the structures built from them should enable a good substrate for the colonization, growth and / or differentiation of biological cells. Third, the materials should be non-toxic and non-immunogenic. Fourth, the implant should enable the regeneration of different tissues to be used in different pathological conditions. The prior art implants only partially meet these requirements and have disadvantages with respect to one or more of these requirements.

Therefore, there is still a need for improved medical implant devices for tissue reconstruction. The aim of the present invention is therefore to provide an implant device which overcomes at least one of the disadvantages mentioned above.

This problem is solved by the provision of a biodegradable implant device according to claim 1. Specific embodiments are the subject of further independent claims.

Summary of the invention

In a first aspect, the present invention provides a biodegradable medical implant device for tissue reconstruction which comprises a three-dimensional (3D) tissue support structure comprising (i) fibroin and (ii) magnesium, a magnesium salt or a magnesium alloy.

The medical implant device of the present invention has several advantages over conventional implant devices.

Fibroin and magnesium are both non-toxic and non-immunogenic materials, which means that they have an adequate safety profile.

Due to its unique properties, Fibroin is particularly suitable as a support structure for tissue regeneration and tissue reconstruction. Its strength, which is comparable to that of Kevlar, and its high elasticity together with its excellent biocompatibility and low immunogenicity are the key properties for its use as an implant material. In addition, Fibroin can self-assemble into mechanically robust material structures that are also biodegradable.

In addition, fibroin and magnesium as biodegradable materials can be degraded by the organism at the same time as the tissue is reconstructed at the implantation site. This is an important clinical advantage since the second surgery to remove the implant is not necessary after the operated tissue regions have healed. Because the use of the resorbable implant of this invention avoids implant removal surgery, it helps reduce the cost and burden on the patient.

By using magnesium, it is not necessary to use a non-absorbable tissue support structure e.g. B. made of titanium to use.

Another advantage of magnesium as an implant material is the fact that magnesium is a natural part of the body within the body or after implantation and also has many important functions in the body. As a result, its biodegradation leads to the formation of Mg ²⁺ cations, which are useful for several cell types, in particular for nerve cells and bone cells.

The fibroin-Mg-based composite structure of the present implant device enables human stem cells, in particular mesenchymal stem cells, to be differentiated into different tissue types and to grow into a large-volume tissue replacement within the 3D support structure. There are also new approaches to making fibroin e.g. B. in genetically modified plants such as the potato.

As a by-product of commercial silk production in the textile industry, Fibroin has unlimited availability and reduced production costs compared to other synthetic materials, making it a superior material in the context of life cycle management.

However, the implant of the invention can be provided with additional active substances such as cytokines, growth factors or trophic factors, which further promote tissue formation and the healing process.

Through an individual selection of the fibroin structure and the magnesium material, the implant can be easily adapted to the specific requirements of the disease to be treated. It therefore offers enormous flexibility in use.

Since the implant device of the invention is based on known components, it can easily be produced in a cost-effective manner.

Detailed description of the invention:

The implant has structural and compositional requirements. As a structural requirement, the implant includes a 3-dimensional support structure, which enables the formation of biological tissue and is therefore referred to in the context of the present invention as a “tissue support structure”.

In relation to the invention, the aforementioned support structure contains fibroin as the first component and magnesium, a magnesium salt or a magnesium alloy as the second component.

Fibroin tissue support structure

In a first embodiment of the invention, the 3D tissue support structure is mainly made of fibroin or consists of it with enclosed particles of magnesium, magnesium salt or magnesium alloy.

In this embodiment, the fibroin component is the structure-determining component of the 3D tissue support structure. As a consequence, fibroin is processed in such a way that a 3D structure with sufficient stability and rigidity is created, which provides cavities for the colonization of cells. A corresponding support structure is referred to in the present invention as a “fibroin 3D tissue support structure”.

Accordingly, in a preferred embodiment, the 3D support structure of the invention can contain 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 95% or more or 98% or more than percent by weight fibroin.

Fibroin is a fibrous silk-based protein, the hierarchical structure of which consists of fibrils and microfibrils in which the fibers are arranged in a highly oriented crystalline form. The silk fibers made by the silkworm, Bombyx mori, have a diameter of 10-25 mm and contain a light (approx. 26 kDa) and a heavy chain (approx. 390 kDa) in a ratio of 1: 1, connected by disulfide bridging. The heavy chains are amphiphilic and consist of hydrophobic and hydrophilic sections. The hydrophobic domains contain repeating sequences of Gly-Ala-Gly-Ala-Gly-Ser, which form crystalline structures, while the hydrophilic sequences contain shorter, non-repetitive structures with polar and bulky side chains.

The fibroin obtained from silk can be obtained from any silk-producing animal. In a preferred embodiment, the fibroin is obtained from the Lepidoptera, such as the Bombyx mori, Anteraea pernyi, Antheraea yamamai or Philosoamia ricini, from the arachnids such as the Nephila clavipes or other insects.

In a further preferred embodiment of the invention, the fibroin is obtained from the silkworm Bombyx mori.

In an alternative embodiment, the protein fibroin can be produced recombinantly by expression in genetically modified cells. Genes that can be used to express recombinant fibroin include weaver ant WAF-1 (Oecophylla smaragdina), fibroin (WAF) or SF (Bombyx mori silk fibroin). The protein fibroin can also be produced by expressing genetically modified plants, for example in potatoes.

In the context of the invention, the term "fibroin" includes fragments or domains of fibroin that are capable of forming fibroin filaments through fibroin-fibroin compounds. As an example, the recombinant expression of the fibroin protein of the spider silk AvMaSp-R, which consists of the 240 amino acid repetitive domain of the spider silk fibroin protein (Araneus ventricosus), leads to a fibroin protein without cytotoxicity or immunogenicity.

In another embodiment, the recombinant fibroin consists only of the heavy chain or a fragment thereof. The (GAGAGS) hexapeptide is the core unit of the H chain and plays an important role in the formation of crystalline domains of fibroin. Fibroin-protein-like multiblock polymers from the repetitive domains of the silk of B. mori and the silk of the spider dragline aggregate spontaneously to form beta-sheet structures that are similar to natural silk. A model polypeptide (Ala-Gly) _n (n = 12 and 5-9) was synthesized and its structure determined, the structure influencing the degree of polymerization. A block copolymer containing elastin (GVGVP) and silk (GAGAGS) motifs was biosynthesized and showed silk-like properties. A full-length recombinant protein derived from the hornet was also produced by E. coli and shows high cell adhesion activity. A type of a silk-elastin-like polymer consisting of repetitive silk-fibroin (GAGAGS) and mammalian-elastin (VPGVG) motifs was expressed in E. coli and slightly improved cell viability. A recombinant spider silk protein with the cell-binding Arg-Gly-Asp motif (RGD) was produced in E. coli and showed sufficient cell adhesion properties.

In one embodiment of the invention, the Fibroin 3D tissue support structure comprises or consists of one of the following structures: Fibroin sponge, Fibroin textile z. B. fleece, woven, braided, knitted, crocheted, felted; Fibroin tubes; Fibroin filaments or yarns; Fibroin hydrogel, Fibroin films or a combination thereof.

In a preferred embodiment of the invention, the Fibroin-3D tissue support structure comprises or consists of a three-dimensional grid of rod-like or beam-like Fibroin structures, which form channels in all three orthogonal directions and which preferably have a distance of 50 to 800 μm and / or have a dimension between 50 and 800 µm.

In a further embodiment of the invention, the structure has a dimension between the rod-like or bar-like structures of 50-500 μm and / or a dimensioning of the rod-like or bar-like structures between 50 and 500 μm.

According to the invention, particles of magnesium, magnesium salt or magnesium alloy are embedded within the Fibroin-3D tissue support structure.

The particles of magnesium or a magnesium alloy can be of any granular structure suitable for metallic materials or metal alloys. This includes metal powder, metal rods, spherical particles, metal chips, metal wires or profile cuts.

In a preferred embodiment, the particles of Mg (alloy) have an average diameter between 0.1 and 250 μm, more preferably between 1 and 100 μm and most preferably between 5 and 50 μm.

In a preferred embodiment, the magnesium alloy is selected from the group consisting of magnesium silver alloys (Mg-Ag), Mg alloys which contain rare earths and magnesium alloys which contain calcium and / or zinc.

In a preferred embodiment, the magnesium alloy contains one or more rare earths, which are preferably selected from a list consisting of yttrium, neodymium, zirconium, gadolinium and dysprodium or any combination thereof. In a more preferred embodiment, the alloy is a Mg-Y-RE-Zr alloy and in particular a Mg-Y-Nd-Zr alloy, which is also known as a WE43 alloy. The mentioned rare earths (RE) Dy, Y, Nd and Gd have a low toxicity and improve the mechanical and corrosion properties. Because of their excellent properties, e.g. B. a relatively slow degradation in aqueous solution and good electrochemical properties in addition to excellent mechanical properties, the WE43 alloy is the most preferred magnesium alloy.

In a preferred embodiment, the Mg alloy contains Ca and / or Zn. In a more preferred embodiment, the Mg alloy contains zirconium, zinc and / or calcium.

In one embodiment, the particles of magnesium or a magnesium alloy are suspended in a non-aqueous solvent to add them to the Fibroin-3D tissue support structure. The use of a non-aqueous solution has the advantage that the corrosion of the Mg (alloy) particles can be greatly reduced or even prevented.

In a further embodiment, the particles of Mg or Mg alloy are coated or surface-treated in order to improve the biophysical properties such as biodegradability or the tissue reaction. B. Modify cytotoxicity.

Said coating can also be a conversion coating. In one embodiment, the conversion coating is carried out by surface treatment with an inert gas such as carbon dioxide.

In a preferred embodiment of the invention, the surface treatment of the particles of Mg or a Mg alloy from a list of methods, consisting of plasma electrolytic oxidation (PEO), also known as MAO (micro arc oxidation), thermal oxidation, carbonate production with CO ₂ inflow , phosphating, anodizing, galvanic and non-galvanic coating.

Any pharmaceutically acceptable magnesium salt can be used in combination with the Fibroin 3D tissue support structure. In a preferred embodiment, the salt anion is obtained from one of the following acids: 1-hydroxy-2-naphtaoic acid, 2,2-dichloroacetic acid, 2-hydroxyethanesulfonic acid (isethionic acid), 2-oxoglutaric acid (α-ketoglutaric acid), 2-acetamidobenzoic acid (acedoben) , 4-aminosalicylic acid, acetic acid, adipic acid, Ascobrinsäure (vitamin C), aspartic acid, benzenesulfonic acid, benzoic acid, camphoric acid, camphorsulfonic acid, capric acid (decanoic acid), caproic acid (hexanoic acid), caprylic acid (octanoic acid), carbonic acid, cinnamic acid, citric acid, cyclamic acid, dodecylsulfuric acid, Ethane-1,2-disulfonic acid, ethanesulfonic acid, formic acid, fumaric acid, galactaric acid (mucic acid), gentisic acid, glucoheptonic acid, gluconic acid, glucuronic acid, glutamate acid, glutaric acid, glycerin-phosphoric acid, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, isobutyric acid, isobutyric acid , Malic acid, malonic acid, mandelic acid, methanesulfonic acid, naphthalene-1,5-disulfonic acid (Armstrong acid), naphtha halin-2-sulfonic acid, nicotinic acid, nitric acid, oleic acid, oxalic acid, palmitic acid, pamoic acid, phosphoric acid, propionic acid (propanoic acid), pyroglutamic acid (pidolic acid), salicylic acid, sebacic acid, stearic acid, succinyl acid (succinic acid), sulfuric acid, tartaric acid, tartaric acid (tartaric acid), Toluenesulfonic acid, undecylenic acid (undecenoic acid).

In a preferred embodiment, the magnesium salt is obtained from a group consisting of magnesium acetate, magnesium amino ethyl phosphate, magnesium aspartate, magnesium bishydrogen aspartate, magnesium carbonate, magnesium chloride, magnesium citrate, magnesium gluconate, magnesium hydrogen citrate, magnesium hydrogen glutamate, magnesium hydrogen phosphate, magnesium hydroxide, magnesium orotate, magnesium oxide and magnesium trisulfate sulfate sulfate.

Ma (alloy) fabric support structure

In an alternative embodiment, the 3D tissue support structure is essentially made of or consists of magnesium or a magnesium alloy and is filled with or coated with a fibroin-containing composition.

In this embodiment, the magnesium or the magnesium alloy is the structure-determining component of the 3D tissue support structure. As a result, the magnesium (or the magnesium alloy) is processed in such a way that a three-dimensional structure with sufficient stability and rigidity is created, which creates cavities for the colonization of cells. A corresponding support structure is referred to in the present invention as “3D Mg (alloy) tissue support structure”.

Magnesium or a magnesium alloy as previously disclosed may be used in the 3D Mg (alloy) tissue support structure, which are preferably coated or surface treated as previously disclosed.

In a further preferred embodiment, magnesium or a magnesium alloy is subjected to selective laser melting (SLM), also known as laser powder bed fusion (LPBF), in order to produce a porous structure. The porous structure is also referred to as a lattice, scaffold, 3D support structure or support structure. In a special version, the magnesium alloy generated by SLM has pores with a diameter of approx. 800 µm. As such, the porous Mg alloy structure is very similar to the structure of cancellous bone, also called trabecular or cancellous bone, which represents the inner tissue of skeletal bones and has an open cellular-porous network structure with pores with a diameter of 200-1000 μm, preferably of 400-800 µm and more preferably 200-650 µm.

Accordingly, the 3D support structure of the invention can, in a preferred embodiment, 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 95% or more or 98% or more as a weight percent of magnesium or magnesium alloy contain.

According to the invention, the Mg (alloy) tissue support structure is filled with a fibroin hydrogel or coated with a layer comprising or consisting of fibroin.

In a preferred embodiment of the invention, the fibroin layer as the coating of the magnesium (the magnesium alloy) has a thickness between 0.1-100 μm, preferably between 1-75 μm and most preferably between 5-30 μm.

In a preferred embodiment of the invention, the magnesium (alloy) 3D tissue support structure comprises or consists of a three-dimensional grid of rod-like or beam-like structures made of magnesium or a magnesium alloy, which channels form in all three orthogonal directions and which preferably have a distance of 50 to 800 microns between the rod or bar-like structures and / or have a dimension of the bar or bar-like structures between 50 and 800 microns.

In a further preferred embodiment of the invention, the distance between the rod-like or bar-like structures is 50-500 μm and / or the dimension of the rod-like or bar-like structures is between 50-500 μm.

In a preferred embodiment of the invention, the Magneisum 3D fabric support structure comprises or consists of magnesium / Mg alloy wires which, by processing into a textile structure, form a 3D structure such as a fleece, woven, braided, knitted, crocheted or felted textiles create three-dimensional structure.

In a particularly preferred embodiment, the magnesium 3D fabric support structure is made of a woven textile made of magnesium or magnesium alloy wires.

Implant properties

For proper vascularization in the recipient's organism, it is preferred that the medical implant device have a tubular shape that is open at both ends, or connections or uncovered areas at both ends for ingrowth or attachment of vascular structures.

The biodegradable medical implant device preferably further comprises an outer shell, which partially or completely surrounds the implant device with an outer shell.

In a preferred embodiment, the outer shell is a membrane which comprises or consists of fibroin. This fibroin membrane preferably has a thickness between 1 and 1000 μm, preferably between 10 and 800 μm, more preferably between 20 and 600 μm, particularly preferably between 50 and 500 μm, in particular between 100 and 300 μm and finally between 150 and 250 μm. The Fibroin membrane enables small molecules such as salts and micronutrients to be transported into the inner volume of the tube, but prevents the undesired ingrowth of cells such as connective tissue cells with a harmful effect on the functionality of the graft.

1. (a) Extrazelluläre Matrix;
2. (b) Matrizelluläre Protein Zytokine;
3. (c) Hormone;
4. (d) Wachstumsfaktoren;
5. (e) Glykosaminoglykane;
6. (f) Proteoglykane;
7. (g) Heparansulfat;
8. (h) Knochenmorphogenetische Proteine.

In a further embodiment, the 3D tissue support structure also contains or is coated with one or more of the following components:

1. (a) extracellular matrix;
2. (b) Matricellular protein cytokines;
3. (c) hormones;
4. (d) growth factors;
5. (e) glycosaminoglycans;
6. (f) proteoglycans;
7. (g) heparan sulfate;
8. (h) Bone Morphogenetic Proteins.

The additional bioactive substances support the perfect colonization, vascularization, proliferation or differentiation of the cells or the tissue that is formed by them.

1. 1. ein Fibroin-Schwamm, der eine längliche Röhre als 3D-Gewebestützstruktur für zu Fettgewebe differenzierende mesenchymale Stammzellen ausformt, bevorzugt umhüllt mit einer Fibroin-Membran, wobei die umhüllte längliche Röhre offen an beiden Enden ist oder Anschlüsse oder unbedeckte Bereiche an beiden Enden für das Einwachsen oder die Anheftung von vaskulären Strukturen aufweist. Diese Komposit-Struktur wird bevorzugt für die Erzeugung von Fettgewebe genutzt. Es wurde herausgefunden, dass die poröse Struktur des Fibroin-Schwamms die Differenzierung in Fettgewebe unterstützt und den Transport von Nährstoffen und Wachstumsfaktoren erleichtert.
2. 2. ein Bündel paralleler, hohler Fibroin-Röhren oder Fibroin-Garne als 3D-Gewebestützstruktur für zu Muskelgewebe differenzierende Stammzellen, bevorzugt umhüllt mit einer Fibroin-Membran und offen an beiden Enden oder aufweisend Anschlüsse oder unbedeckte Bereiche an beiden Enden für das Einwachsen oder die Anheftung von vaskulären Strukturen. Durch die Bereitstellung von parallel orientierten Fibroin-Strukturen, wie hohle Röhren, Filamente oder Garne, ist ein zelltypischer Stimulus für die Bildung von Muskelgewebe gegeben. Wenn Fibroin-Filamente verwendet werden, ist es bevorzugt, Fasern mit einer vergrößerten Oberfläche zu verwenden, wie sie durch Filamente bereitgestellt werden, die Querschnitte haben, die ausgewählt sind aus der Liste bestehend aus trilobal, dogbone, hantelförmig, 4-lappig, oktolobal, unregelmäßig, schneeflockenförmig, limabohnenförmig, polygonal, gezahnt limabohnenförmig, sternförmig, ziehharmonikaförmig, dreieckig, gezahnt kreisförmig, lobular mit Längsstreifung.
3. 3. ein dreidimensionales Gitter gemacht aus stangen- oder balkenförmigen Strukturen aus Magnesium- oder Mg-Legierung, welche Kanäle in alle drei orthogonalen Richtungen bilden, um ein Fibroin-Hydrogel aufzunehmen, welches zu Knochengewebe differenzierende Stammzellen enthält, wobei das Gitter eine tubuläre Form hat, bevorzugt umhüllt mit einer Fibroin-Membran und an beiden Enden offen oder aufweisend Anschlüsse oder unbedeckte Bereiche an beiden Enden für das Einwachsen oder die Anheftung von vaskulären Strukturen.

In a specific embodiment of the invention, the biodegradable medical implant device preferably has one of the following three composite structures:

1. 1. a fibroin sponge, which forms an elongated tube as a 3D tissue support structure for mesenchymal stem cells differentiating into adipose tissue, preferably covered with a fibroin membrane, the covered elongated tube being open at both ends or connections or uncovered areas at both ends for shows the ingrowth or attachment of vascular structures. This composite structure is preferred for the production of adipose tissue. It has been found that the porous structure of the fibroin sponge supports differentiation in adipose tissue and facilitates the transport of nutrients and growth factors.
2. 2. a bundle of parallel, hollow Fibroin tubes or Fibroin yarns as a 3D tissue support structure for stem cells that differentiate into muscle tissue, preferably covered with a Fibroin membrane and open at both ends or having connections or uncovered areas at both ends for ingrowth or the Attachment of vascular structures. The provision of parallel-oriented fibroin structures, such as hollow tubes, filaments or yarns, provides a cell-typical stimulus for the formation of muscle tissue. When using fibroin filaments, it is preferred to use fibers with an increased surface area as provided by filaments that have cross sections selected from the list consisting of trilobal, dogbone, dumbbell-shaped, 4-lobed, octolobal, irregular, snowflake-shaped, lima-bean-shaped, polygonal, serrated lima-bean-shaped, star-shaped, accordion-shaped, triangular, serrated circular, lobular with longitudinal striation.
3. 3. a three-dimensional lattice made of rod or bar-shaped structures made of magnesium or Mg alloy, which form channels in all three orthogonal directions, around a fibroin hydrogel which contains stem cells which differentiate into bone tissue, the lattice having a tubular shape, preferably encased in a fibroin membrane and open or having both ends with connections or uncovered areas at both ends for the ingrowth or attachment of vascular structures.

In a preferred embodiment, the medical implant device of the invention is used as a bone implant because of the presence of magnesium. Especially since magnesium is the fourth most common cation in the human body, with an estimated 1 mol of magnesium stored in the body of an average 70 kg adult, with half of the total physiological magnesium stored in the bone tissue. The presence of magnesium in the bone system offers advantages in terms of bone strength and bone growth. Magnesium alloys have a specific density (1.74-2 g / cm ³ ) and modulus of elasticity (41-45 GPa), which are comparable to those of bones in the human body (1.8-2.1 g / cm ³ ; 3-20 GPa) come closest. Therefore, magnesium alloys in orthopedics and bone repair or replacement are superior to other metallic or polymer implants in terms of physical and mechanical properties, since the differences in the modulus of elasticity between the implant and natural bone can result in stress-shielding effects, the stress peaks at the interface between the implant and the bone, thereby reducing the stimulation to form new bone tissue and reducing the stability of the implant.

1. (a) eine filamentöse Fibroin-3D-Gewebestützstruktur ausgestattet mit darin enthaltenen Partikeln aus Magnesium oder Magnesiumlegierung zur Ausformung einer länglichen Röhre als 3D-Gewebestützstruktur für das Einwachsen von neuralen Strukturen, bevorzugt umhüllt mit einer Fibroin-Membran, wobei die umhüllte längliche Röhre offen an beiden Enden ist oder Anschlüsse oder unbedeckte Bereiche an beiden Enden für das Einwachsen oder die Anheftung von vaskulären Strukturen aufweist; oder
2. (b) ein Bündel paralleler hohler Fibroin-Röhren oder Fibroin-Garne als dreidimensionale Stützstruktur zur Differenzierung mesenchymaler Stammzellen in Muskelgewebe, bevorzugt mit einer äußeren Fibroin-Membran umhüllt ist und an beiden Enden offen ist oder Anschlüsse oder nichtabgedeckte Bereiche an beiden Enden aufweist, zur Aufnahme oder Anheftung vaskulärer Strukturen.

In one embodiment, the biodegradable medical implant device is a neural implant device and is preferred

1. (a) a filamentous Fibroin-3D tissue support structure equipped with particles of magnesium or magnesium alloy contained therein to form an elongated tube as a 3D tissue support structure for the ingrowth of neural structures, preferably covered with a fibroin membrane, the covered elongated tube being open is either end or has connections or uncovered areas at both ends for ingrowth or attachment of vascular structures; or
2. (b) a bundle of parallel hollow fibroin tubes or fibroin yarns as a three-dimensional support structure for differentiating mesenchymal stem cells into muscle tissue, preferably with an outer fibroin membrane and which is open at both ends or has connections or uncovered areas at both ends Admission or attachment of vascular structures.

In a further aspect, the invention provides a biodegradable medical implant device, the 3D tissue support matrix being filled with stem cells or progenitor cells. In a preferred embodiment, hematopoietic stem cells or mesenchymal stem cells are used.

In an alternative embodiment, the biodegradable medical implant device is implanted without prior cell colonization and is filled after the implantation by ingrowth of cells and / or tissue structures of the recipient.

In another aspect, the invention provides a method of making a biodegradable medical implant that includes a three-dimensional fibroin tissue support structure, the method including the following steps:

Fibroin sponge:

• ○ 6 ml Fibroin solution and 2 ml of a 4 vol% ethanol solution in water are mixed and frozen
• ○ After 36 hours, the solution is thawed for 1.5 hours and then frozen again
• ○ The freeze-thaw cycle is repeated five times to create a sponge-like structure
• ○ The Fibroin sponge can be removed from the solution and stored in a refrigerator.

Fibroin membrane:

• ○ A petri dish is filled with 15 ml of fibroin solution
• ○ This dries horizontally on a leveling plate for 36 hours
• ○ The membrane is dissolved by filling the Petri dish with 40 ml of a 35 vol% ethanol solution in water
• ○ The membrane is removed and stored in distilled water in the refrigerator.

Fibroin fleece produced by the electrospinning process

The Fibroin solution is processed in a spinning machine using a spinneret or a needleless spinning system with the following parameters:

Parameters for spiders with a spinneret

Fibroin concentration Syringes volume Cannulae diameter temperature Relative humidity Flow rate 30-60% 1.5-100 ml 0.2-1.08 mm 10-80 ° C 5-40% 0.1-1.0 ml / h

Parameters for needleless spinning

Fibroin concentration revolutions per minute Distance between the collector and the top of the spinning apparatus temperature Relative humidity tension 25-65% 2-15 8-20 cm 10-90 ° C 5-60% 30-60 kV

In order to prepare the fibroin support structure, the trained person can use various manufacturing methods as are established for polymeric proteins. For example, the fibroin structure can be produced by one or more processes, selected from a group consisting of solvent casting particle leaching, gas foaming, melt molding, freeze drying, rapid prototyping, production of solid free-form surfaces, electrohydrodynamic process (EHD), EHD direct jet process (EHD -JD), EHD direct writing, the electrospinning process and three-dimensional printing.

EXAMPLES

Preparation of a three-dimensional fibroin support structure as the basis for an implant

Fibroin sponge

• ◯ 6 ml of fibroin solution and 2 ml of a 4 vol% ethanol solution in water are mixed and frozen
• ◯ After 36 hours, the solution is thawed for 1.5 hours and then frozen again.
• ◯ The freeze-thaw cycle is repeated five times to create a sponge-like structure
• ◯ The fibroin sponge can be removed from the solution and stored in a refrigerator.

Fibroin membrane

• ◯ A petri dish is filled with 15 ml of fibroin solution
• ◯ This dries horizontally on a height adjustment plate for 36 hours
• ◯ The membrane is released by filling the Petri dish with 40 ml of a 35 vol% ethanol-water solution
• ◯ The membrane is detached and stored in distilled water in the refrigerator.

Addition of particles from a magnesium alloy

• Die Partikel aus der Magnesiumlegierung können zwischen zwei Vliesmembranen aus Fibroin-Fasern eingebettet werden, welche vorzugsweise durch das Elektrospinning-Verfahren hergestellt werden.
• Die Partikel aus der Magnesiumlegierung können im Herstellungsprozess der Vliese zugegeben werden, wobei die Fibroin-Fasern durch mechanische, thermische oder chemische Verwicklungen miteinander verbunden werden. Als Ergebnis sind die Partikel aus der Magnesiumlegierung innerhalb des Vlieses verteilt.

The magnesium alloy particles can be added by the following methods:

• The magnesium alloy particles can be embedded between two fibroin fiber fleece membranes, which are preferably produced by the electrospinning process.
• The magnesium alloy particles can be added to the nonwovens during the manufacturing process, the fibroin fibers being connected to one another by mechanical, thermal or chemical entanglements. As a result, the magnesium alloy particles are dispersed within the fleece.

For the manufacture of sponge-like products, the magnesium alloy particles can be added to a fibroin-containing solution, followed by a subsequent dynamic freeze to produce a fibroin sponge with intermediate magnesium alloy particles.

For the production of film-like products, the particles from the magnesium alloy can be added to the film-forming solution, so that after drying the film contains embedded particles from the magnesium alloy. Alternatively, a sandwich structure can be produced with a first fibroin film on which particles made of the magnesium alloy are placed and a further fibroin film which is applied or cast onto the particles made of the magnesium alloy.

Enveloping a tubular implant structure with a fibroin membrane

The fibroin sheath that covers the tubular implant structure can be produced by the electrospinning process. For this purpose, a tubular implant structure is coated with a three-dimensional fibroin support structure, preferably a fibroin sponge, by rotating the support structure in the coating process with fibroin fibers to form a fleece. Alternatively, a fibroin membrane can be produced as a cover by dip coating with subsequent drying of the tubular implant structure.

Figure list

These and other aspects of the invention will be apparent and explained with reference to the embodiments described hereinafter.

The invention will now be described based on embodiments by way of example with reference to the accompanying drawings.

• zeigt schematische Zeichnungen von drei röhrenförmigen Implantatstrukturen, umhüllt von einer Fibroin-Membran, mit einer 3D-Stützstruktur gebildet aus einem Fibroin-Schwamm mit eingestreuten Magnesiumpartikeln in (A), einer 3D-Stützstruktur gebildet aus einem Bündel von parallelen Fibroin-Filamenten oder Garnen mit eingestreuten Magnesiumpartikeln in (B) und einer 3D-Stützstruktur aus einer Mg 3D-Stützstruktur gefüllt mit einem Fibroin-Hydrogel in (C).
• zeigt schematische Zeichnungen von Querschnitten von zwei zylinderförmigen Nervenimplantaten gemäß der vorliegenden Erfindung mit einer filamentösen Fibroin-Stützstruktur in (A) und einem Fibroin-Vlies in (B).

In the pictures:

• shows schematic drawings of three tubular implant structures, encased by a fibroin membrane, with a 3D support structure formed from a fibroin sponge with interspersed magnesium particles in (A), a 3D support structure formed from a bundle of parallel fibroin filaments or yarns interspersed magnesium particles in (B) and a 3D support structure made of a Mg 3D support structure filled with a fibroin hydrogel in (C).
• shows schematic drawings of cross sections of two cylindrical nerve implants according to the present invention with a filamentous fibroin support structure in (A) and a fibroin fleece in (B).

In the figures, the same numbers refer to the same objects throughout. Objects in the illustrations are not necessarily drawn to scale.

Detailed description of the embodiments:

Various embodiments of the invention will now be described with the aid of the figures.

shows schematic drawings of three tubular implant structures. In (A), the tubular implant structure consists of a fibroin sponge 3D support structure 1 with embedded particles made of a magnesium alloy 6 . The support structure 1 is from a fibroin membrane 5 covered. In the implanted state, the tubular structure is supported by an arteriovenous vascular loop (AV loop) 8th pulled through. In the implant structure of (B) is the 3D support structure made of fibroin filaments 2nd with adhered particles from a magnesium alloy 6 made and made of a fibroin membrane 5 covered. In the implant structure of (C), the 3D support structure is made of a magnesium alloy 3D support structure 4, filled with a fibroin hydrogel 3rd and covered by a fibroin membrane 5 . When implanted, as shown in to , the tubular structure of an arteriovenous vascular loop (AV loop) 8th pulled through.

shows a schematic drawing of the cross section of two cylindrical nerve implants according to the present invention. In (A) is a filamentous fibroin support structure 2nd with particles of a Mg alloy 6 filled and covered by a cylindrical fibroin membrane 5 . In (B) is the filamentous fibroin support structure 2nd with particles of a Mg alloy 6 filled and covered by a cylindrical fibroin membrane 5 . In the implanted state, the implants (A) or (B) are located between two nerve segments in order to bridge a defect, so that the axons of the injured nerve grow into the implant structure and the fibroin support structure is finally infiltrated with further supporting components, for example glial cells and blood vessels can.

Reference symbol list

1

3D support structure in the form of a fibroin sponge

2nd

3D filamentous fibroin support structure

3rd

Fibroin hydrogel

4th

3D support structure made of magnesium (or a magnesium alloy)

5

Fibroin membrane cover

6

Magnesium (or magnesium alloy) particles

7

Nerve segments

8th

Arteriovenous vascular loop (A-V loop)

9

Tubular implant structure

Definitions

The term “biodegradable” in the context of the present invention refers to a device that is degraded under physiological conditions.

According to the invention, the “three-dimensional tissue support matrix” is defined as a three-dimensional matrix which serves to support viability, proliferation and / or differentiation of living cells. As a result, these cells, which colonize the support matrix in vitro, ex vivo or in vivo, form a “tissue”. The term "tissue" as used herein refers to a cellular level of organization between cells and a whole organ. As a result, the cells that are enclosed in the three-dimensional support structure form a cellular ensemble in which the cells interact in order to perform a specific function. In a preferred embodiment of the invention, the “tissue” is an ensemble of similar cells and extracellular matrix of the same origin, which together perform a specific function.

As used in the context of the invention, the term “biocompatible” refers to a device that is essentially non-toxic in an environment in vivo and is essentially not rejected by the recipient's physiological system.

The term “hydrogel” refers to a three-dimensional (3D), preferably cross-linked network of hydrophilic polymers that swell in water or other polar solvents. In general, hydrogels consist mostly of water. In some embodiments, the hydrogel of the invention may e.g. B. consist of up to 50% or more, 60% or more, 70% or more, 80% or more, 90% or more, 95% or more or 98% or more by weight water. In some embodiments, other components such as proteins, carbohydrates or salts and preferably magnesium particles can be included in the hydrogel. Typically, these additional components are included in the hydrogel by mixing a suspension or solution containing these additional components with a precursor solution (ie, a solution consisting of fibroin proteins suitable for hydrogel production) and the resulting one Network is networked using conventional means of networking. In some embodiments, a variety of hydrogels can be combined into a hydrogel composition.

Claims (15)

A biodegradable medical implant device comprising a three-dimensional (3D) tissue support structure (i) fibroin, and (ii) Magnesium, a magnesium salt or a magnesium alloy. A biodegradable medical implant device according to Claim 1 , wherein the 3D tissue support structure is essentially made of fibroin or consists of fibroin with embedded magnesium, magnesium salt or magnesium alloy particles. A biodegradable medical implant device according to Claim 2 , wherein the fibroin-containing 3D fabric support structure comprises or consists of one of the following components: (a) fibroin sponge, (b) fibroin textile such as fleece, fabric, braid, knitted fabric, crocheted textile or felt, (c) fibroin Tubes, (d) fibroin filaments or yarns, or (e) fibroin hydrogel, (f) fibroin textile made by needleless e-spinning or spinneret e-spinning processes; (g) fibroin film; or a combination of these. A biodegradable medical implant device according to Claim 3 , wherein the 3D tissue support structure comprises a three-dimensional grid of rod-shaped or beam-shaped fibroin structures or consists of which channels form in all three orthogonal directions and which preferably have a distance between the rod-shaped or beam-shaped structures of 50 to 800 μm and / or dimension of the rod-shaped or bar-shaped structures between 50 and 800 microns. A biodegradable medical implant device according to Claim 1 , wherein the 3D tissue support structure is essentially made of magnesium or a magnesium alloy or consists of a magnesium or a magnesium alloy, which comprises a silk or fibroin-containing component, which is preferably used as a filling or as a coating of the structure of magnesium or magnesium alloy with a Silk or fibroin-containing composition is used. A biodegradable medical implant device according to Claim 5 , wherein the 3D-Mg / Mg alloy support structure is a three-dimensional lattice made of rod-shaped or bar-shaped structures made of magnesium or magnesium alloy, which form channels in at least two and preferably in all three orthogonal directions and which are a distance between the rod- or bar-shaped structures from 50 to 800 microns and / or a dimension of the rod-shaped or bar-shaped structures between 50 and 800 microns. A biodegradable medical implant device according to one of the preceding claims, wherein the medical implant device has a tubular shape which is open at both ends or which have connections or uncovered areas at both ends for the ingrowth or attachment of vascular and / or neuronal structures. A biodegradable medical implant device according to any one of the preceding claims, further comprising an outer shell which partially or completely envelops the medical device. A biodegradable medical implant device according to one of the preceding claims, wherein the outer shell is a membrane which comprises or consists of fibroin and which preferably has a thickness between 1 and 1000 µm, particularly preferably between 10 and 800 µm and particularly preferably between 20 and 600 µm. A biodegradable medical implant device according to one of the preceding claims, wherein the 3D tissue support matrix additionally comprises one or more of the following components: (a) extracellular matrix; (b) matricellular protein cytokines; (c) hormones; (d) growth factors; (e) glycosaminocanes; (f) proteoglycans; (g) heparan sulfate; (h) Bone Morphogenetic Proteins. A biodegradable medical implant device according to one of the preceding claims, wherein the medical implant device has one of the following composite structures: (a) a fibroin sponge that forms an elongated tube as a 3D tissue support structure for mesenchymal stem cells that differentiate into adipose tissue, preferably encased with a fibroin membrane, the encased elongated tube being open at both ends or connections or uncovered areas at both ends for ingrowth or attachment of vascular structures; (b) a bundle of parallel, hollow fibroin tubes or fibroin yarns as a 3D tissue support structure for stem cells differentiating into muscle tissue, preferably covered with a fibroin membrane and open at both ends or having connections or uncovered areas at both ends for ingrowth or the attachment of vascular structures; (c) a three-dimensional lattice made of rod or bar-shaped structures made of magnesium or Mg alloy which form channels in all three orthogonal directions to accommodate a fibroin hydrogel containing stem cells differentiating into bone tissue, the lattice being tubular in shape has, preferably covered with a fibroin membrane and open at both ends or having connections or uncovered areas at both ends for ingrowth or attachment of vascular structures. A biodegradable medical implant device according to one of the preceding claims, wherein the 3D tissue support matrix is filled in vivo or ex vivo with a cell population, preferably with a stem cell population and particularly preferably with hematopoietic or mesenchymal stem cells. A biodegradable medical implant device according to one of the Claims 1 to 10th , wherein the device is a neural implant device and preferably (a) a filamentous Fibroin-3D tissue support structure equipped with particles of magnesium or magnesium alloy contained therein to form an elongated tube as a 3D tissue support structure for the ingrowth of neural structures, preferably covered with a fibroin Membrane, wherein the covered elongate tube is open at both ends or has connections or uncovered areas at both ends for the ingrowth or attachment of vascular structures; or (b) a bundle of parallel hollow fibroin tubes or fibroin yarns as a 3D tissue support structure for differentiating mesenchymal stem cells into muscle tissue, which is preferably covered with an outer fibroin membrane and is open at both ends or connections or uncovered areas at both ends has for the absorption or attachment of vascular structures. A method of manufacturing a biodegradable medical implant Claim 2 comprising the following steps: (a) providing a fibroin solution (b) producing a 3D support structure from fibroin, preferably a fibroin fleece, a fibroin sponge or a bundle; (c) providing the fibroin structure with magnesium, at least one magnesium salt or with magnesium alloy particles; (d) shaping the 3D support structure from step (c) into the desired implant shape, which is preferably a tubular shape; (e) coating the structure from step (d) with a membrane, which is preferably a fibroin membrane. A method for producing a biodegradable medical implant accordingly Claim 5 comprising the steps of: (a) providing a powder of magnesium or magnesium alloy; (b) production of a 3D magnesium (alloy) support structure by processing the Mg (alloy) powder from step (a), preferably by using the laser powder bed fusion method (LPBF); (c) providing a 3D magnesium (alloy) framework with a fibroin coating or filling; (d) shaping the 3D framework from step (c) into the desired implant shape, which is preferably a tubular shape; (e) coating the structure from step (d) with a membrane, which is preferably a fibroin membrane.

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