Technical Field of the Invention

The invention relates to an exosome-based biocompatible implant obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials. Said exosomes are attached to the implant material at a level of 1 ng-1 pgr/cm.

State of the Art

Implants refer to inanimate materials placed inside the body and living tissues. The types of implants applied to various parts of the human body, such as the arm or knee, are called orthopaedic implants, and the implants applied to the tooth area are called dental implants. Implants are applied to provide osseointegration between living tissue and non-living tissue, regardless of the areas where they are applied. The unity between the implant and living bone tissue is called osseointegration. In ensuring osseointegration, tissue compatibility and surface properties of the implant material are of great importance, as well as the quality of the bone where the implant will be placed [1 ].

In the state of the art, implants consist of metal alloys and several types of plastic. However, these implants are inadequate in terms of biocompatibility, prevention of inflammation, and cell stress, especially in ensuring osseointegration. Osseointegration is one of the most important factors in long-term implant success, and the reason for this is that the success of the implant depends on osseointegration following placement.

The implant surfaces used in the state of the art are titanium plasma spray-coated surfaces, sandblasted and acidified surfaces, sandblasted surfaces, acidified surfaces, hydroxyapatite-coated surfaces, titanium oxide (TiO2) surfaces, oxidized surfaces, and machine-prepared surfaces, which are designed to ensure osseointegration. Although these surfaces are effective in osseointegration because they positively affect the healing processes of the bones to which the implant is applied [1] due to their rough structure, these surfaces are insufficient to imitate the cellular matrix, they bring costs and are also insufficient in terms of effects such as prevention of inflammation and cellular stress, increasing the biocompatibility of implants, increasing cell adhesion, changing the surface modification in a way that favours the cell, providing a long-lasting implant, providing an immune system-supporting implant, providing an implant- supported by cells, and ensuring that the implant adheres to the applied area. Since the implant surfaces in the state of the art are insufficient to imitate the cellular matrix, prevent inflammation and cellular stress, ensure biocompatibility, and increase cell adhesion, these implant surfaces do not last long in the areas where they are applied and require a new treatment process. Therefore, extra costs and loss of time are inevitable.

Although today, studies on the use of biological agents and biological polymers in implants for purposes such as increasing biocompatibility in implants and preventing inflammation are focused, it is not yet possible to find a material that can completely imitate the cellular matrix of the bone structure. Therefore, the need for an implant material that is biocompatible, prevents inflammation and cell stress, is long-lasting, supports the immune system, increases cell adhesion, and is low-cost, is increasing day by day, while ensuring osseointegration and at the same time imitating the cell matrix that forms the bone structure almost exactly. The reason why the need for implant materials with these properties is increasing day by day is that the implants in the state of the art, which are inadequate in terms of said technical effects, bring about a second surgical intervention process after the implant is applied. In addition, a second surgical intervention greatly increases the possibility of re-infection of the implanted area.

For reasons such as limitations and inadequacies of the implants in the state of the art, implants consisting of metal alloys and a few types of plastic in the state of the art being insufficient in ensuring biocompatibility, especially in ensuring osseointegration, and preventing inflammation and cell stress, implants with titanium plasma spray coated surfaces, sandblasted and acidified surfaces, sandblasted surfaces, acidified surfaces, hydroxyapatite coated surfaces, titanium oxide (TiO2) surfaces, oxidized surfaces and machine-prepared surfaces, which are designed to ensure osseointegration, bringing high costs, in addition, implants with these surfaces being insufficient in preventing inflammation and cellular stress, increasing the biocompatibility of implants, increasing cell adhesion, changing the surface modification in favour of the cell, providing a long-lasting implant, providing an implant that supports the immune system, providing an implant supported by cells, and ensuring the retention of the implant in the applied area and the studies on the use of biological agents and biological polymers in implants for purposes such as increasing biocompatibility in implants and preventing inflammation being carried out today being ineffective in fully mimicking the cellular matrix of the bone structure, it has become necessary to introduce an implant that eliminates all these problems.

Brief Description and Aims of the Invention

In the invention, an exosome-based biocompatible implant obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials is described. Said exosomes are attached to the implant material at a level of 1 ng-1 pgr/cm. By means of the exosome-based biocompatible implant of the invention, the desired osseointegration is achieved, the adhesion of surface proteins is ensured, the lipid structure can completely imitate the normal matrix of the cells as a surface covering, inflammation in the area where the implant is applied is prevented, cellular stress is prevented, and retention of the implant is ensured in the area where it is applied. Additionally, increasing the surface roughness of implants obtained by spraying exosomes increases cell integration (cell adhesion) on the implant surface. In addition to all these, the implant of the invention is long-lasting and low-cost.

The main aim of the invention is to provide a low-cost implant that provides biocompatibility, prevents inflammation and cell stress, increases cell adhesion, supports the immune system, and can imitate the cellular matrix of the bone structure with osseointegration between the implant and living bone tissue. These effects are achieved by the exosome-based biocompatible implant of the invention, which is obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials. Additionally, increasing the surface roughness of implants obtained by spraying exosomes increases cell integration (cell adhesion) on the implant surface. Exosomes play an important role in the transport of bioactive substances and are also rich in surface proteins and markers. By means of the richness of exosomes in surface proteins and markers, sufficient roughness is provided for the retention of cells and the formation of cell feet (pods) is also ensured. Exosomes used in the implant of the invention play an important role in increasing cell adhesion, changing the surface modification in favour of the cell, providing long-lasting implants, supporting the immune system, creating the desired cell matrix on the implant, supporting the implant by the cell and increasing implant durability.

The aim of the invention is to provide an implant in which osseointegration between the implant and living bone tissue is increased. Increasing osseointegration, which is the unity between the implant and living bone tissue, is achieved by the exosome- based biocompatible implant of the invention, which is obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials. Increasing the surface roughness of implants obtained by spraying exosomes increases cell integration (cell adhesion) on the implant surface. Increasing cell integration also means increasing osseointegration. Exosomes used in the implant of the invention have an important place in the transport of bioactive substances and are also rich in surface proteins and markers. By means of these exosomes, cell adhesion is increased and the formation of cell-matrix on the implant is encouraged, therefore osseointegration is increased with the implant of the invention.

Another aim of the invention is to provide a biocompatible implant. In the invention, biocompatibility is achieved by spraying exosomes obtained from stem cells onto implant materials consisting of different materials.

Exosomes used in the implant of the invention have an important place in the transport of bioactive substances and are also rich in surface proteins and markers. Thanks to these exosomes, cell adhesion is increased and the formation of the cell matrix on the implant is encouraged, thus biocompatibility is increased with the implant of the invention. Stem cells are cells that carry common and general substances throughout the body and where the immune response cannot be developed. Biocompatible means that the immune system does not respond. By means of the exosome used in the implant of the invention, an implant in which the immune response cannot be developed is provided.

Another aim of the invention is to provide an implant that can mimic the cellular matrix of the bone structure. Providing an implant that can mimic the cellular matrix of the bone structure is achieved by the exosome-based biocompatible implant of the invention, which is obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials. Since the exosomes used in the implant of the invention are effective in transporting bioactive substances and are rich in surface proteins and markers, a matrix that can mimic the cellular matrix of the bone structure is provided with the implant of the invention. The bone matrix is covered with porous and rough active components. The roughness on the surface of the exosome applied by spraying to the surface of the implant material of the invention is nanometer in diameter and resembles a bone matrix.

Another aim of the invention is to provide an implant that prevents inflammation in the area where the implant is applied. Since the exosomes used in the implant of the invention greatly support the immune system, inflammation that may occur in the area where the implant of the invention is applied is prevented. Inflammation occurs when microorganisms or foreign objects adhere to the surface. At this stage, the exosome imitates the bone structure by increasing cell adhesion on the implant surface and also eliminates foreign body interaction by covering the surface of the implant. As a result, inflammation in the implant of the invention is prevented by the exosome sprayed onto the surface of the implant material.

With the invention, cellular stress in implant applications is prevented. In this invention, cellular stress is prevented by means of the use of exosomes. Cellular stress begins with the arrival of the foreign environment and white blood cells to the area where the implant is applied. Said cellular stress formation is prevented by the exosome used in the implant of the invention creating a biocompatible environment on the implant.

Another aim of the invention is to provide an implant that provides adhesion in the area where it is applied. Adhesion of the implant to the applied area is achieved by applying exosomes obtained from stem cells to the implant. By means of the exosomes, osseointegration in the implants obtained is increased and with the increase in osseointegration, the fracture in the implant is delayed and the structural strength of the implant is increased.

Another aim of the invention is to provide the implant having significant effects such as osseointegration and biocompatibility at a low cost. The implant that is the subject of the invention is low cost is ensured by the fact that exosomes based on human stem cell-derived multicentric mesenchymal stromal cells (MSC) or dental pulp-derived stem cells (DPSC) are used instead of high-cost matrix materials, metals, and peptides to ensure osseointegration between living tissue and non-living tissue. In addition, since the increase in osseointegration in the implants of the invention comprising exosomes will also increase the structural strength of the implant, a longer-lasting implant is provided and therefore the cost of a second implant procedure is prevented.

Detailed Description of the Invention

The invention relates to an exosome-based biocompatible implant obtained by spraying exosomes obtained from stem cells onto implant materials consisting of different materials. Said exosomes are exosomes based on human stem cell-derived multicentric mesenchymal stromal cells (MSC) or dental pulp-derived stem cells (DPSC). The implant that is the subject of the invention can be used both dentally and orthopedically.

The implant that is the subject of the invention is attached to implant types containing an alloy of 0.01 -1 % exosome and 99% serum physiology. Said alloys are platinumzirconium, titanium-zirconium, titanium-iron, copper-titanium, titanium-platinum- zirconium or titanium-steel alloys. In the implant of the invention, plastic materials can also be used instead of said alloys. Said plastic materials are selected from polyvinyl alcohol (PVA), polystyrene (PS), polyethylene (PE), or mixtures thereof.

Exosomes are applied on implant material comprising platinum zirconium alloy, titanium zirconium alloy, or titanium iron alloy with the help of serum physiology. A serum-based exosome solution of 1 mg/100 mL is prepared and this solution is attached to the implant material by spraying at a pressure of 1 -15 bar, 1 ng-1 pgr/cm. The given dose is the exosome dose required for each implant surface (cm ² ). For example, a nail and a femur implant have different volumes. The serum-based solution comprising exosomes must be applied to the surface of the implant in said proportions. In the spray application method, there is generally no need to dry the implant. If drying is done, this period should be 30 minutes at room temperature and in a sterile environment. The thickness of the exosome applied on the implant is in the range of 10-30 nm.

The method of preparing the biocompatible implant that is the subject of the invention comprises the process steps of; i. obtaining pure exosomes by precipitation process, ii. mixing the obtained exosomes with sterile water as serum physiology and obtaining a solution, iii. attaching the prepared solution to the implant material made of alloys or plastic materials by spraying, and iv. drying the prepared implants in a sterile environment and at room temperature and obtaining the implant with exosomes on its surface.

In one embodiment of the invention, the method of preparing said biocompatible implant comprises the process steps of: i. obtaining pure exosomes after precipitation at 100 thousand-200 thousand rpm for 1 hour, ii. mixing the obtained exosomes with 0.9 gram/1 Litre of sterile water as serum physiology at a rate of 1 mg/100 ml and obtaining a solution, iii. attaching the prepared solution onto the implant material made of alloys or plastic material by spraying at a pressure of 1 -15 bars at a level of 1 ng-1 pgr/cm, and iv. drying the prepared implants for 25-30 minutes in a sterile environment and at room temperature and obtaining an implant with 10-30 nm thick exosomes on its surface.

The serum physiology-based exosome solution obtained in step (ii) can also be stored in kit form for later use. The reason for using serum physiology in the method is to prevent exosomes from bursting and possible deformations. Serum physiology is 0.9 g/1 litre of sterile water.

By means of the exosome-based biocompatible implant of the invention, the desired osseointegration is achieved, the cellular matrix of the bone structure can be completely imitated, inflammation in the area where the implant is applied is prevented, cellular stress is prevented, and the implant is retained in the area where it is applied. Additionally, increasing the surface roughness of implants obtained by spraying exosomes increases cell integration (cell adhesion) on the implant surface. In addition to all these, the implant of the invention is long-lasting and low-cost. The exosome-based biocompatible implant, obtained by spraying exosomes obtained from the stem cell that is the subject of the invention onto implant materials consisting of different materials, comprises said exosomes, which are vesicles with a diameter of 30-150 nm. Exosomes carry bioactive substances and are rich in surface proteins and markers. Said surface proteins are tetraspanin, integrin, actin, and clathrin. Tetraspanins are a family of membrane proteins found in all multicellular eukaryotes. By means of tetraspanins, it becomes easier for the implant to be recognized by the cells. Integrins are transmembrane receptors that facilitate cell-cell and cell- extracellular matrix adhesion. Actine is a round-shaped, structural protein that polymerizes in a helix shape to form actin filaments, also known as microfilaments. Actine filaments form the cellular skeleton. Clathrins are basic proteins that play a role in extracellular vesicles taking a certain shape and facilitating the formation of small vesicles in the cytoplasm.

Exosomes increase cell adhesion and change surface modifications in favour of the cell in the implant of the invention in which they are used. In addition, the implant that is the subject of the invention is long-lasting by means of the exosome it comprises in its structure. Since exosomes comprise a structure that supports the immune system, when used in the implant of the invention, they prevent any inflammation that may occur after implant application. In addition to all these, said exosomes are encouraged to form the cell matrix on the implant of the invention. It should also be noted that the exosomes in the implant of the invention ensure that the implant is supported by the cell and increases implant durability.

REFERENCES

[1] Uzun, G., & Keyf, F. (2007). SURFACE CHARACTERISTICS OF THE IMPLANT SYSTEMS AND OSSEOINTEGRATION, 43-50.