WO2019135237A1

WO2019135237A1 - Scaffold for tissue welding

Info

Publication number: WO2019135237A1
Application number: PCT/IL2019/050023
Authority: WO
Inventors: Tal Dvir; Maayan MALKI; Sharon FLEISCHER
Original assignee: Ramot At Tel-Aviv University Ltd.
Priority date: 2018-01-04
Filing date: 2019-01-03
Publication date: 2019-07-11

Abstract

Provided herein is a composition-of-matter, comprising a porous scaffold that comprises a polymer; and a plurality of particles attached to said polymer, wherein the scaffold is having at least one region that is translucent to low-energy radiation, the particles are capable of generating heat upon interacting with the radiation, and the generated heat renders the polymer reactive to bonding with a tissue in a subject.

Description

SCAFFOLD FOR TISSUE WELDING

RELATED APPLICATION/S

This application claims the benefit of priority from U.S. Provisional Patent Application No. 62/613,445 filed on 4 January 2018, the contents of which are incorporated herein by reference in their entirety.

FIELD AND BACKGROUND OF THE INVENTION

The present invention, in some embodiments thereof, relates to tissue engineering and, more particularly, but not exclusively, to an artificial tissues designed for tissue welding.

Engineered tissues are considered a promising approach for regenerating infarcted organs. For example, cardiac patches are prepared by seeding cardiac cells within a 3D biomaterial scaffold, which provide a physical, structural and biochemical supporting microenvironment. The scaffolds encourage cell-cell and cell-matrix interactions, which lead to the formation of a functioning tissue. Cardiac patches are usually attached to the scar tissue of the heart by a surgical operation, involving synthetic sutures or staples. Although these procedures can attach a patch to a desired site, this surgical approach suffers from several limitations, which include the blockage of blood supply to the patch, bleeding, injury to a healthy tissue and risk of infection. Such trauma may cause an additional deterioration of the ventricle’s function, expanding the damage.

In recent years, in order to attach tissues, several alternatives to surgical procedures were used. For example, biological glues, such as medical grade cyanoacrylates and coupling agent were used to adhere tissues in subjects. Although, the materials strongly adhere to tissues, some are associated with toxicity. In addition, the strong adhesion leads to an area with stiff mechanical properties, which do not match the elastic properties of the surrounding tissue (e.g., myocardium), impeding proper contractile function and provoking inflammation. To address this limitation several biological glues were developed. For example, an elegant elastomeric biodegradable and biocompatible, gecko-inspired tissue adhesive tape was developed. The glue relied on the synergy between chemistry and topography, showing effective sealing of surgical colon and stomach defects. In another study, nanoparticle solution was used to glue two hydrogels by connecting to their polymer chains and reorganizing their structure. Other approaches include neat glues that do not contain any solvents and their mechanical properties can be tuned. However, although all these materials exhibit excellent adhesive properties, several properties limit their application for engineering thick tissues. For example, they are not porous, do not supply a supporting fibrous biomimetic microenvironment for tissue assembly, and in some cases their water-free environment does not allow cell penetration into their core.

Gold nanostructures, such as spheres or particles with higher aspect ratio can be incorporated into 3D scaffolds to increase the transfer of the electrical signal between electrogenic cells. Gold nanostructures can also be used for tissue welding and wound sealing. For example, there were some proposals to use gold nanoshells or particles in solutions that can absorb light, heat up, and locally melt tissues and fuse them. However, such techniques of fusion of tissues by heating might be destructive for viability of cells. In addition, using of particles in a solution in combination with a three dimensional assembly of a tissue is limited.

Additional prior art documents include, for example, studies by Lauto, A. et al. [“ Adhesive biomaterials for tissue reconstruction”, J. Chem. Technol. Biotechnol., 2008, 83(4), pp. 464-472], Lee Y. et al. [" Bioinspired Nanoparticulate Medical Glues for Minimally Invasive Tissue Repair", Adv Healthc Mater., 2015, 4(16), pp. 2587-96], Fleischer, S. et al. [" Coiled fiber scaffolds embedded with gold nanoparticles improve the performance of engineered cardiac tissues", Nanoscale, 2014, 6(16), pp. 9410-4], Cao, H. et al. ["The topographical effect of electrospun nanofibrous scaffolds on the in vivo and in vitro foreign body reaction", J Biomed Mater Res A., 2010, 93(3), pp. 1151-9], Shevach, M. et al. [ "Gold nanoparticle-decellularized matrix hybrids for cardiac tissue engineering" , Nano Lett., 2014, 14(10), pp. 5792-6], Baranes, K. et al. [ "Gold Nanoparticle-Decorated Scaffolds Promote Neuronal Differentiation and Maturation" , Nano Lett., 2016, 16(5), pp. 2916-20], Annabi, N. et al. [" Surgical Materials: Current Challenges and Nano-enabled Solutions", Nano Today, 2014, 9(5), pp. 574-589], Li, J. et al. ["Tough adhesives for diverse wet surfaces", Science, 2017, 357(6349), pp. 378-381], Wu, J. et al. [" Enhancing cell infiltration of electrospun fibrous scaffolds in tissue regeneration", Bioactive Materials, 2016, 1(1), pp. 56-64] and U.S. Patent Application Publication No. 2012/0220991.

SUMMARY OF THE INVENTION

Aspects and embodiments of the present invention relate to an artificial scaffold which can be safely attached to an organ or extracellular matrix in a subject by using low-energy radiation welding.

Thus, according to an aspect of some embodiments of the present invention there is provided a composition-of-matter that includes a porous scaffold that includes a polymer; and a plurality of particles attached to the polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat upon interacting with the radiation, and the heat renders the polymer reactive to bonding with a tissue in a subject

According to some embodiments of the invention, the region is located at a perimeter, an edge, a frame, and/or an end of the scaffold.

According to some embodiments of the invention, the region is located where the scaffold is making a close intimate contact with the tissue.

According to some embodiments of the invention, the amount of the particles ranges from 0.001 mg to 10 mg per 1 gram of the scaffold.

According to some embodiments of the invention, the radiation is near infrared radiation having a wavelength that ranges from 700 nm to 950 nm.

According to some embodiments of the invention, the region admits at least 20 % of the radiation.

According to some embodiments of the invention, the particles comprise a substance and/or have a size and/or have a shape that is conducive to the generation of heat upon exposure to NIR.

According to some embodiments of the invention, the shape of the particles is an anisotropic shape.

According to some embodiments of the invention, the size of the particles ranges from 0.5 nm to 2 pm.

According to some embodiments of the invention, the substance of the particles is having a physical property selected from the group consisting of electric conductance, anisotropy, ferromagnetism, ferrimagnetism, diamagnetism, superparamagnetism or paramagnetism.

According to some embodiments of the invention, the particles comprise a metal and/or a metal oxide.

According to some embodiments of the invention, the metal is selected from the group consisting of gold, silver, aluminum, iron, nickel, copper, platinum, titanium, and any blends and alloys thereof.

According to some embodiments of the invention, the particles comprise an organic material.

According to some embodiments of the invention, the particles comprise a substance selected from the group consisting of a carbon allotrope, a composite material, polyaniline, polypyrrole, and blends thereof.

According to some embodiments of the invention, the particles have a core-coat structure. According to some embodiments of the invention, the substance of the core and/or the coat is selected from the group consisting of a conductive substance, a dielectric substance, a metal, an organic substance, an inorganic substance, and any mixture thereof.

According to some embodiments of the invention, the substance of the core and/or the coat is selected from the group consisting of gold, silver, an organic polymer, a ceramic and a metal oxide.

According to some embodiments of the invention, the metal is gold.

According to some embodiments of the invention, the particles are having an aspect ratio that ranges from 2 to 10.

According to some embodiments of the invention, the radiation is near IR characterized by a wavelength that ranges from 750 nm to 850 nm.

According to some embodiments of the invention, the translucent region in the scaffold is having a thickness that ranges from 50 pm to 100 pm.

According to some embodiments of the invention, the scaffold is having a form selected from the group consisting of a patch, a sheet, a mesh, a rod, a tube, a ring, and a three- dimensional object.

According to some embodiments of the invention, the polymer is having a shape selected from the group consisting of fibers, ribbons, flakes, spheres, coils, helicoils, beads-on-a-string, and any combination thereof.

According to some embodiments of the invention, the ribbons are having a mean width that ranges from 100 nm to 10 pm, and mean thickness that ranges from 100 nm to 10 pm.

According to some embodiments of the invention, the fibers are having a thickness that ranges from 1 pm to 0.1 pm.

According to some embodiments of the invention, the polymer is in a form of electrospun fibers or ribbons.

According to some embodiments of the invention, the polymer exhibits a plurality of functional groups capable of binding the particles.

According to some embodiments of the invention, the polymer exhibits a plurality of functional groups capable of the bonding upon exposure to the heat.

According to some embodiments of the invention, the polymer comprises a protein, a polysaccharide, alginate, chitosan, a polycaprolactone, polyglycolic acid, poly(L-lactide), a poly(ester urethane), polydioxanone, polyethylene glycol, and any copolymer thereof. According to some embodiments of the invention, the protein is selected from the group consisting of albumin, collagen, fibrin, elastin, fibronectin, glycoprotein, laminin, silk, and any combination thereof.

According to some embodiments of the invention, the polymer includes albumin.

According to some embodiments of the invention, the polymer includes electrospun albumin fibers.

According to some embodiments of the invention, the porous scaffold is having an average pore size that ranges from 50 nm to 2 mm.

According to some embodiments of the invention, the composition-of-matter presented herein further includes a plurality of cells distributed within and/or on the scaffold.

According to some embodiments of the invention, the cells are viable cells selected from the group consisting of seed cells, stem cells, mature cells, and a tissue.

According to some embodiments of the invention, the cells correspond to the tissue in the subject.

According to some embodiments of the invention, the polymer includes electrospun albumin fibers, the plurality of particles includes gold particles having an anisotropic shape characterized by an aspect ratio of at least 2, and further includes a plurality of viable cells distributed within and/or on the scaffold.

According to some embodiments of the invention, the tissue to which the composition- of-matter presented herein is attached, is selected from the group consisting of a connective tissue, a muscle tissue, a nervous tissue and an epithelial tissue.

According to some embodiments of the invention, the muscle tissue is selected from the group consisting of a smooth muscle, a visceral muscle, a skeletal muscle, and cardiac muscle.

According to some embodiments of the invention, the connective tissue is selected from the group consisting of an extracellular matrix, a fibrous connective tissue, a skeletal connective tissue, a fascia, a bone, a tendon, a ligament, an adipose tissue and an areolar tissue.

According to an aspect of some embodiments of the present invention there is provided a method of attaching/fusing/welding the composition-of-matter presented herein to a tissue in a subject in need thereof, the method includes:

placing the composition on the tissue; and

irradiating at least the translucent region(s) in the composition with radiation suitable for absorption by the particles.

According to some embodiments of the invention, the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 1 W/cm². According to some embodiments of the invention, the irradiating is effected for at least 60 seconds.

According to some embodiments of the invention, the composition-of-matter presented herein is used as a patch and the tissue in the subject is selected from the group consisting of a cardiac muscle, a visceral muscle, a skeletal muscle, a fascia, and an abdominal wall.

According to an aspect of some embodiments of the present invention there is provided a process of manufacturing the composition-of-matter presented herein, the process includes:

obtaining the porous scaffold as described herein; and

contacting the porous scaffold with a dispersion/suspension of the plurality of particles, to thereby obtain the porous scaffold having the plurality of particles attached thereto.

According to some embodiments of the invention, the process further includes, prior or subsequent to the contacting, introducing viable cells into the porous scaffold under conditions suitable for maintaining the viable cells.

According to an aspect of some embodiments of the present invention, there is provided a method of treating a tissue defect in a subject in need thereof, the method is effected by:

contacting the composition-of-matter presented herein with the tissue in the subject such that at least the radiation-translucent region(s) overlap with healthy parts of the tissue surrounding the tissue defect, and the composition is covering the tissue defect; and

irradiating at least the radiation-translucent region(s) in the composition with the radiation, thereby fusing the composition to the tissue.

According to some embodiments of the invention, the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 1 W/cm2.

According to some embodiments of the invention, the irradiating is effected for at least 60 seconds.

According to some embodiments of the invention, the tissue defect is selected from the group consisting of a damaged cardiac muscle, an inguinal hernia, an incisional hernia, a femoral hernia, a skeletal muscle tear, and a damaged skin.

Unless otherwise defined, all technical and/or scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of the invention, exemplary methods and/or materials are described below. In case of conflict, the patent specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and are not intended to be necessarily limiting. BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS

Some embodiments of the invention are herein described, by way of example only, with reference to the accompanying images drawings and charts. With specific reference now to the images drawing and charts in detail, it is stressed that the particulars shown are by way of example and for purposes of illustrative discussion of embodiments of the invention. In this regard, the description taken with the images and charts makes apparent to those skilled in the art how embodiments of the invention may be practiced.

FIGs. 1A-D present schematic illustrations of the process of preparing a composition-of- matter, according to some embodiments of the present invention, designed as a cardiac patch, and using the same in a tissue welding process, showing the adsorption of gold nanorods (FIG.

IA) followed by seeding of an electrospun albumin fiber scaffold with viable cardiac cells (FIG.

IB), to afford a cardiac patch ready for use (FIG. 1C), followed by patching the injured heart on a rat by the patch using a NIR laser source (FIG. 1D);

FIGs. 2A-H illustrate the fabrication and feature characterization of an engineered tissue, according to some embodiments of the present invention, wherein FIG. 2A shows HRTEM micrographs of AuNRs (scale bar: 50 nm), FIG. 2B shows the UV-visible spectra wavescan of AuNRs, FIG. 2C shows SEM micrographs of electrospun albumin fiber scaffolds (scale bar: 100 pm), FIG. 2D shows SEM micrographs of electrospun albumin fiber scaffolds (scale bar: 10 pm), FIG. 2E shows AuNRs adsorption to electrospun albumin scaffolds at t=0 (upper panel) and t=60 min (lower panel), FIG. 2F shows ESEM micrographs of AuNRs adsorbed to electrospun albumin fiber scaffold (scale bar: 2 pm), FIG. 2G shows ESEM micrographs AuNRs adsorbed to electrospun albumin fiber scaffold (scale bar: 500 nm), and FIG. 2H shows XRD spectroscopy confirming the existence of Au on the albumin fibers;

FIGs. 3A-F present mechanical properties and viability of the engineered tissues, according to some embodiments of the present invention, welded to a porcine heart, wherein FIG. 3A shows a stress versus laser power flux plot, FIG. 3B shows a stress versus time of NIR irradiation plot, FIG. 3C shows schematics of the geometry of the albumin fiber scaffold used for tissue welding, FIG. 3D shows a stress versus thickness of albumin scaffold plot, FIG. 3E shows viability of cardiomyocyte on electrospun albumin fiber scaffolds before and after irradiation, and FIG. 3F shows immunofluorescence image of cardiac a-sarcomeric actinin (pink), connexin- 43 (green) and cell nuclei (blue) (scale bar: 20 pm); and

FIGs. 4A-E present illustrations and characterization of the cardiac patch welded to a heart, wherein FIG. 4A shows a schematic representation of the welding/integration process, FIG. 4B shows the cardiac patch welded to rat heart, FIG. 4C is a SEM micrograph of cross section of the welded heart showing the interaction between the patch and the heart (scale bar: 200 pm), FIG. 4D is a larger magnification SEM micrograph of a cross section of the welded heart showing the interaction between the patch and the heart (scale bar: 50 pm), and FIG. 4E shows the H&E staining of interaction between the patch and the heart (scale bar: 500 pm).

DESCRIPTION OF SOME SPECIFIC EMBODIMENTS OF THE INVENTION

The principles and operation of the present invention may be better understood with reference to the figures and accompanying descriptions.

Before explaining at least one embodiment of the invention in detail, it is to be understood that the invention is not necessarily limited in its application to the details set forth in the following description or exemplified by the Examples. The invention is capable of other embodiments or of being practiced or carried out in various ways.

As discussed hereinabove, although cardiac patches hold a promise for repairing the infarcted heart, their integration with the myocardium by sutures may cause further damage to the diseased organ. To address this issue and solve other problems associated with the currently known methodologies and technologies, a facile and safe, suture-free technology for attachment of engineered tissues to living organs is provided herein. According to an embodiment of the present invention, a composite scaffold (a composition-of-matter), comprising albumin electrospun fibers and gold nanorods (AuNRs) is provided herein. As demonstrated in the Examples section that follows below, viable cardiac cells were successfully seeded within the electrospun albumin scaffolds and assembled into a functioning patch, or engineered tissue. The engineered tissue was then positioned on the myocardium and irradiated with near IR laser (808 nm). Due to the definitive and purposeful design of the patch, the AuNRs were able to absorb the low-energy NIR radiation sufficiently to convert it into thermal energy, which locally changed the molecular structure of the fibrous scaffold, allowing it to attach itself to the wall of the heart tissue strongly but safely. Such hybrid biomaterials can be used to integrate any engineered tissue with any defected organs, while minimizing the risk of additional injury for the subject, caused by the conventional stitching methods.

Aspects of the present invention are drawn to a scaffold which can be safely attached to a living tissue, an organ or an extracellular matrix in a live subject by using near infrared (NIR) radiation. According to an aspect of some embodiments of the present invention there is provided a composition-of-matter, which is based on a porous scaffold made of a polymer; and a plurality of particles attached to the polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat upon interacting with the radiation, and the heat renders the polymer in the scaffold reactive to bonding with a tissue in a subject.

In the context of embodiments of the present invention, bonding is also referred to as “fusing”, welding” and“merging” between the scaffold and the tissue. Bonding is formed upon irradiation of the particles-containing scaffold and the tissue, whereas the radiation is converted into heat by the particles, and the heat renders the polymer in the scaffold and the parts of the tissue that are in close intimate contact with the scaffold reactive towards one-another, thereby effecting the bonding.

In the context of embodiments of the present invention, the term“bonding” refers to any form of chemical, physical or mechanical attachment of the at least part of the polymer in the scaffold to the tissue in the subject. Bonding, in the sense of embodiments of the present invention, includes forming hydrogen bonds, covalent bonds, salt-bridge bonds, disulfide bonds, aromatic interactions and the likes, between functional groups in the scaffold and corresponding functional groups in the tissue. The term“bonding” also encompasses entanglement of filament, strands and chains in the scaffold and the tissue.

According to some embodiments of the invention, the radiation-translucent region(s) is/are located at a perimeter, an edge, a frame, and/or an end of the scaffold, or in the alternative, the translucent region is located where the scaffold is intended for attachment with the tissue, or intended for making a close intimate contact with the tissue.

According to some embodiments of the invention, the amount of the particles per 1 gram of the scaffold ranges from 0.001 mg to 10 mg, or at least 0.001 mg, 0.01 mg, 0.1 mg, 1 mg, or at least 10 mg per 1 gram of the scaffold.

According to some embodiments of the invention, the radiation is a low-energy radiation which is substantially non-deleterious to living cells, such as near infrared radiation having a wavelength that ranges from 700 nm to 950 nm.

According to some embodiments of the invention, the radiation-translucent region(s) in the scaffold admit at least 5, 10, 15, 20, 30, 40 or 50 percent of the radiation.

According to some embodiments of the invention, the particles are selected, manufactured and designed to interact with the low-energy radiation and generate heat upon exposure to the radiation; hence, the particles comprise a certain substance and/or have a certain size and/or have a certain shape that is conducive to the generation of heat upon exposure to low- energy radiation such as NIR.

According to some embodiments of the invention, the shape of the particles is an anisotropic shape (non-spherical, elongated, elliptical, or oddly/irregularly shaped.

According to some embodiments of the invention, the size of the particles ranges from 0.5 nm to 2 pm, or less than 100 pm, less than 50 pm, less than 10 pm, less than 10 pm, less than 1 pm, less than 0.1 pm, less than 50 nm, less than 10 nm, less than 5 nm, or less than 1 nm.

According to some embodiments of the invention, the substance of the particles is having a physical property selected from the group consisting of electric conductance, anisotropy, ferromagnetism, ferrimagnetism, diamagnetism, superparamagnetism or paramagnetism. Such physical property may also be conducive to interaction with electric, magnetic or electromagnetic radiation that is harmless to cells and can further be employed by the generated tissue. For example, electrically conductive particles may assist in the process of forming nerve cell connections.

According to some embodiments of the invention, the particles comprise a metal and/or a metal oxide, wherein the metal may be, without limitation, gold, silver, aluminum, iron, nickel, copper, platinum, titanium, and any blends and alloys thereof.

According to some embodiments of the invention, the particles have a core-coat structure, as these radiation-responsive particles are known in the art.

According to some embodiments of the invention, the substance of the core and/or the coat is selected from the group consisting of a conductive substance, a dielectric substance, a metal, an organic substance, an inorganic substance, and any mixture thereof.

According to some embodiments of the invention, the metal is gold.

According to some embodiments of the invention, the particles are having an aspect ratio that ranges from 2 to 10. According to some embodiments of the invention, the radiation is near IR characterized by a wavelength that ranges from 750 nm to 850 nm.

According to some embodiments of the invention, the translucent region in the scaffold is having a thickness that is configured to admit sufficient flux of radiation to allow the particles adsorbed thereon/therein to generate sufficient heat to case welding of the scaffold to the tissue. The optimal thickness depends on the type of polymer used in the scaffold, the amount and type of particles adsorbed therein, and other practical considerations. Hence, the thickness of the region is about 40 pm, 50 pm, 60 pm, 70 pm, 80 pm, 90 pm, 100 pm, 110 pm, 120 pm, 150 pm, 200 pm, or ranges from 50 to 100 pm.

According to some embodiments of the invention, the polymer exhibits a plurality of functional groups capable of binding the particles. For example, a protein-based scaffold may exhibit a plurality of thiol functional groups, to which gold nanoparticles can bind spontaneously.

According to some embodiments of the invention, the polymer exhibits a plurality of functional groups capable of the bonding to a tissue upon exposure to the heat generated locally by the interaction of the radiation with the particles. For example, hydrogen-bond donor/acceptor functional groups may be rendered free for inter-molecular bonding between the polymer of the scaffold and the tissue.

According to some embodiments of the invention, the polymer includes albumin.

According to some embodiments of the invention, the cells correspond to the tissue in the subject, namely the type of cells that is being introduced into the scaffold corresponds to the type of cells of the tissue, to which the composition-of-matter presented herein is intended to be fused. In some embodiments, the cells are harvested from the subject, and in some embodiments the cells are harvested from a healthy section of the tissue that is being treated.

According to some embodiments of the invention, the tissue to which the composition- of-matter presented herein can be attached/fused/welded, includes without limitation, a connective tissue, a muscle tissue, a nervous tissue and an epithelial tissue.

According to some embodiments of the invention, the muscle tissue includes without limitation, a smooth muscle, a visceral muscle, a skeletal muscle, and cardiac muscle.

According to some embodiments of the invention, the connective tissue includes without limitation, an extracellular matrix, a fibrous connective tissue, a skeletal connective tissue, a fascia, a bone, a tendon, a ligament, an adipose tissue and an areolar tissue.

placing the composition on the tissue; and irradiating at least the translucent region(s) in the composition with radiation suitable for absorption by the particles.

According to some embodiments of the invention, the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 0.5, 1, 2, 3, 4, 5, 6, 7, 8,

9, 10, 15, 20, or 25 W/cm².

According to some embodiments of the invention, the irradiating is effected for at least 5,

10, 20, 30, 40, 50, 60, 70, 80, 90, or 100 seconds.

obtaining the porous scaffold as described herein; and

According to another aspect of some embodiments of the present invention, there is provided a method of treating a tissue defect in a subject in need thereof, the method is effected by:

In some embodiments, the tissue defect is a necrotic or gangrenous tissue, and the composition-of-matter presented herein is fabricated as patch designed to reconstruct the tissue after the damaged part of the tissue had been surgically removed or cleared. In some embodiments the patch is an engineered tissue containing viable cells that correspond to the type of cells in the treated tissue in the subject.

According to some embodiments of the invention, the tissue defect is selected from the group consisting of a damaged cardiac muscle, an inguinal hernia, an incisional hernia, a femoral hernia, a skeletal muscle tear, and a damaged skin. Hence, the composition-of-matter presented herein is useful in treating cardiac muscle defects, hernia and muscle tear or rupture. Unless otherwise defined, all technical and/or scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of the invention, exemplary methods and/or materials are described below. In case of conflict, the patent specification, including definitions, will control. In addition, the materials, methods, and examples are illustrative only and are not intended to be necessarily limiting

It is expected that during the life of a patent maturing from this application many scaffolds which can be safely attached with an organ or extracellular matrix in a subject by using radiation will be developed and the scope of the term composition-of-matter, comprising a porous scaffold that comprises a polymer; and a plurality of particles attached to said polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat from said radiation, and the heat renders the polymer reactive to bonding with an extracellular matrix in a subject, is intended to include all such new technologies a priori.

As used herein the term“about” refers to ± 10 %.

The terms "comprises", "comprising", "includes", "including", “having” and their conjugates mean "including but not limited to".

The term“consisting of’ means“including and limited to”.

The term "consisting essentially of" means that the composition, method or structure may include additional ingredients, steps and/or parts, but only if the additional ingredients, steps and/or parts do not materially alter the basic and novel characteristics of the claimed composition, method or structure.

As used herein, the phrases "substantially devoid of" and/or "essentially devoid of" in the context of a certain substance, refer to a composition that is totally devoid of this substance or includes less than about 5, 1, 0.5 or 0.1 percent of the substance by total weight or volume of the composition. Alternatively, the phrases "substantially devoid of" and/or "essentially devoid of" in the context of a process, a method, a property or a characteristic, refer to a process, a composition, a structure or an article that is totally devoid of a certain process/method step, or a certain property or a certain characteristic, or a process/method wherein the certain process/method step is effected at less than about 5, 1, 0.5 or 0.1 percent compared to a given standard process/method, or property or a characteristic characterized by less than about 5, 1, 0.5 or 0.1 percent of the property or characteristic, compared to a given standard. The term“exemplary” is used herein to mean“serving as an example, instance or illustration”. Any embodiment described as“exemplary” is not necessarily to be construed as preferred or advantageous over other embodiments and/or to exclude the incorporation of features from other embodiments.

The words“optionally” or“alternatively” are used herein to mean“is provided in some embodiments and not provided in other embodiments”. Any particular embodiment of the invention may include a plurality of“optional” features unless such features conflict.

As used herein, the singular form "a", "an" and "the" include plural references unless the context clearly dictates otherwise. For example, the term "a compound" or "at least one compound" may include a plurality of compounds, including mixtures thereof.

Throughout this application, various embodiments of this invention may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

Whenever a numerical range is indicated herein, it is meant to include any cited numeral (fractional or integral) within the indicated range. The phrases“ranging/ranges between” a first indicate number and a second indicate number and“ranging/ranges from” a first indicate number “to” a second indicate number are used herein interchangeably and are meant to include the first and second indicated numbers and all the fractional and integral numerals therebetween.

As used herein the terms“process” and "method" refer to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, material, mechanical, computational and digital arts.

As used herein, the term“treating” includes abrogating, substantially inhibiting, slowing or reversing the progression of a condition, substantially ameliorating clinical or aesthetical symptoms of a condition or substantially preventing the appearance of clinical or aesthetical symptoms of a condition. It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination or as suitable in any other described embodiment of the invention. Certain features described in the context of various embodiments are not to be considered essential features of those embodiments, unless the embodiment is inoperative without those elements.

Various embodiments and aspects of the present invention as delineated hereinabove and as claimed in the claims section below find experimental and/or calculated support in the following examples.

EXAMPLES

Reference is now made to the following examples, which together with the above descriptions illustrate some embodiments of the invention in a non-limiting fashion.

Materials and Methods

Preparation of Au nanorods:

A cetyl trimethylammonium bromide solution (CTAB, 15.3 mL, 0.20 M) was mixed with 1.7 mL of 1.016 mM HAuCL. The solution was separated into two solutions, seed and growth (2 ml and 16 ml, respectively). Approximately 0.12 mL of ice-cold 0.01 M NaBH₄ was added to the seed solution, resulting in the formation of a brownish-yellow solution. The seed solution was vigorously stirred for 3 minutes. Following, it was kept at 25 °C for 1 hour to ensure total decomposition of the borohydride. Ascorbic acid (0.15 ml, 0.115 M) and of AgN0₃ (0.15 ml, 20 mM) were added to the growth solution. The seed solution was diluted in distilled deionized water at a ratio of 1 to 10. Finally, 0.18 mL of the seed solution was added to the growth solution. The final solution was kept for at least 4 hours at 32 °C, reaching a concentration of 20 mg/ml. The AUNRs exhibited strong absorbance at approximately 808 nm.

Electrospinning of albumin fiber scaffolds

Bovine serum albumin [BSA; Fraction V, MP Biomedicals, Aurora, OH; 14 % (w/v)] was dissolved in tetrafluoroethylene (TFE) and distilled water (9:1, respectively), following, excess of b-mercaptoethanol (Merck, Darmstadt, Germany) was added for overnight reaction. The solution was electrospun at room temperature, using a syringe pump (Harvard Apparatus) delivered at a rate of 2 mL/h. A high voltage supply (Glassman High Voltage, NJ, US) was used to apply a 12 kV potential between the capillary tip and the grounded aluminum collector placed at a distance of 14 cm. Following, 100-120 mih thick electrospun mats were collected, cut into 8 mm in diameter circles and the perimeter was trimmed to form a circular“frame” with thickness of 60-80 pm, or cut into a rectangular shape (gauge length: 10 mm, and width: 4 mm, thickness: l00-l20pm) and the edges was trimmed to thickness of 60-80 pm.

High resolution transmission electron microscopy (HRTEM):

AuNRs with concentration of 50 mg/ml were deposited on carbon coated copper grids (SPI) for transmission electron microscopy (TEM) analysis. Images were captured using FEI™ F20 Philip s-Tecnai equipped with FEG (Schottky type).

Scanning electron microscopy (SEM) of albumin fiber scaffolds:

Samples were mounted onto aluminum stubs with conductive paint and sputter-coated with an ultrathin (150 A) layer of gold in a Polaron E 5100 coating apparatus (Quorum technologies, Laughton, UK). The samples were viewed under JCM-6000PLUS NeoScope Benchtop (JEOL USA Inc., Peabody, MA).

Albumin fiber scaffold integrated to rat hearts:

The constructs were fixed with 2.5 % glutaraldehyde (24 h at 4 °C), followed by graded series of ethanol-water solutions for dehydration (25-100 %). All samples were critical point dried, sputter-coated with gold in a Polaron E 5100 coating apparatus (Quorum technologies, Lewis, UK) and observed under JSM-840A SEM (JEOL, Tokyo, Japan).

Environmental scanning electron microscope (ESEM):

AuNRs solution were suspended on albumin scaffold and imaged without additional coating using a Quanta 200 FEG Environmental Scanning Electron Microscope (ESEM) with a field-emission gun (FEG) electron source. Imaging was carried out under low vacuum with a high tension of 20 kV and a working distance of 10 mm. Elemental composition was performed using X-Max 20, Oxford EDX and acquired using INCA Software.

Cardiac cell isolation, seeding and cultivation:

Cardiac cells were isolated according to Tel Aviv University ethical use protocols. Briefly, left ventricles of 0-3-day-old neonatal Sprague-Dawley rats (Envigo Laboratories, Israel) were harvested, and cells were isolated using six cycles (30 min each at 37 °C) of enzyme digestion with collagenase type II (95 U/mL; Worthington, Lakewood, NJ) and pancreatin (0.6 mg/mL; Sigma- Aldrich) in Dulbecco’s modified Eagle Medium (DMEM, CaCl₂-2H₂0 (1.8 mM), KC1 (5.36 mM), MgS0₄-7H₂0 (0.81 mM), NaCl (0.1 M), NaHCOs (0.44 mM), NaH₂P0₄ (0.9 mM)). After each round of digestion cells were centrifuged (600 G, 5 minutes) and resuspended in culture medium composed of M-199 supplemented with 0.6 mM CuS0₄-5H₂0, 0.5mM ZnS0₄-7H₂0, L5mM vitamin B12, 500 U/mL Penicillin and 100 mg/mL streptomycin, and 0.5 % (v/v) FBS. To enrich the cardiomyocytes population, cells were suspended in culture medium with 5 % FBS and pre-plated twice (45 minutes). Cell number and viability were determined by a hemocytometer and trypan blue exclusion assay. Cardiac cells (8-10⁵) were seeded onto the scaffolds by adding 10 pL of the suspended cells followed by a 45 minutes incubation period (Humidified incubator, 37 °C, 5 % C0₂). Following, cell constructs were supplemented with culture medium (5 % FBS) and further incubated.

Cardiac tissue integration to heart tissues by laser irradiation:

AuNRs solution (3.5 pl, 20 mg/ml) was suspended on the edges of albumin scaffolds. Following, the scaffolds were placed on the rat heart tissue and irradiated with ranging laser power fluxes (1, 1.2, and 1.5 W/cm²) and ranging time (60, 90, 120 sec). An 808 nm diode laser was used as a radiation source (MDL-III-808, 0-2.5W continuous wave output; Optoengine, Midvale, UT) and a 400 pm fiber optic cable was used. The fiber optic was attached to a silica- lens collimator with a 22.2-mm aperture (CeramOptec Industries, Changchun, China). The adhesion was tested using a Lloyd tensile testing instrument (model LS1) with a 20 N load cell at a rate of 5 mm per minute.

Cell viability:

Compositions-of-matter, in the form of cardiac tissues, prepared according to some embodiments of the present invention, were integrated to porcine heart as previously described. Following, 6 replicates of the samples were cultured immediately after irradiation in Presto Blue viability assay (Life Technologies, NY). As a control group 6 replicates of cardiac tissues with AuNRs, but no irradiation was cultured in Presto Blue as well.

Immuno staining:

Cell constructs were fixed and permeabilized in 100 % cold methanol for 10 minutes, washed three times in DMEM-based buffer and then blocked for 1 hour at room temperature in DMEM -based buffer containing 2 % FBS, after which the samples were washed three times. Cardiac tissues were incubated with primary mouse anti a-sarcomeric actinin antibody (1:750, Sigma- Aldrich) and connexin 43 (1:250, Invitrogen, Carlsbad, CA), washed three times and incubated for 1 hour with Alexa Fluor 647 conjugated goat anti-mouse antibody (1:500; Jackson, West Grove, PA) and Alexa Fluor 488 conjugated goat anti-rabbit antibody (1:500; Jackson). For nuclei detection, the cells were incubated for 3 minutes with Hoechst 33258 (1:100; Sigma) and washed three times. Samples were visualized using a scanning laser confocal microscope (Nikon Eclipse Ni). Haematoxylin and eosin stain (H&E):

Cardiac tissues were soldered to rat tissue as previously described. Following constructs were fixed in 4% formalin (24 h at 40° C) and embedded in OCT. Sections (6 pm thick) were prepared using a cryotome™ FSE (Thermo scientific) and affixed to X-tra® adhesive glass slides (Leica Biosystems, Wetzler, Germany), dehydrated in graduated ethanol steps (70-100%), and stained with hematoxylin and eosin. Samples were visualized using an inverted fluorescence microscope (Nikon Eclipse TI).

Statistical analysis:

Data are presented as means ± SEM. Differences between samples were assessed by a Student’s t-test. All analyses were performed using GraphPad Prism version 6.00 for Windows (GraphPad Software), whereas p < 0.05 was considered significant.

IB), to afford a cardiac patch ready for use (FIG. 1C), followed by patching the injured heart on a rat by the patch using a NIR laser source (FIG. 1D).

Example 1

Preparing Scaffolds Attachable by NIR Radiation

A proof of concept of some embodiments of the present invention was carried out by preparing engineered tissue comprising electrospun albumin scaffolds and gold nanorods, of which their attachment to tissues is made by NIR radiation.

FIGs. 2A-D illustrate an example of a 3D porous scaffold of albumin electrospun fiber, according to an embodiment of the present invention, where mono-dispersed AuNRs with a mean size of about 60 nm x 20 nm were incorporated. The AuNRs, which are shown in FIG. 2A, were synthesized so as to allow rapid and efficient conversion of a laser radiation with a wavelength of about 800 nm to thermal energy. Such AuNRs can be characterized by TEM and UV-vis-NIR. The UV-vis spectrum of those AuNRs showed a sharp and strong absorption centered at about 810 nm as shown in FIG. 2B.

The preparing of the structure of the scaffolds included dissolving albumin in trifluoroethanol and distilled water, and adding b-mercaptoethanol after 24 hours. The solution was electrospun at room temperature at a flow rate of 2 mL/hour under an electrical field of 12.5 kV. The resulting scaffolds were composed of ribbon-like fibers with a mean width of 2.4 pm pm and thickness of 0.5 pm (FIG. 2C and FIG. 2D). Next, the AuNRs were integrated into the albumin scaffolds by soaking the scaffolds in an AuNRs solution during 60 min to allow quick adsorption. The spontaneous attachment of the AuNRs onto the structures of the scaffolds turned the color of the scaffolds into brown, while leaving the solution transparent (FIG. 2E). Surface analysis by scanning electron microscope images and EDX revealed a homogenous distribution of the AuNRs on the surface of the scaffolds fibers (FIGs. 2F-H).

Example 2

Demonstration of an attachment of a scaffold to an Organ

Pieces of pig’s myocardiums were used to demonstrate attaching the composite scaffolds to an organ, by irradiating near IR. Fluxes of 1, 1.2 and 1.5 W/cm2 were applied on thin scaffolds and a mechanical tester was used to evaluate the adhesion strength between the layers. A flux of 1.5 W/cm2 enabled to attach the scaffolds significantly stronger than the 1 and 1.2 W/cm² fluxes (p= 0.0009 and p= 0.0008, respectively; FIG. 3A). Attaching of the scaffolds to the tissues in three irradiation durations of 60, 90, and 120 sec, at flux of 1.5 W/cm² are shown in FIG. 3B. As shown, a time-dependent effect was observed with significantly stronger attachment after 120 seconds.

Example 3

Functionality of Different Thicknesses

Shaping thin edges at the perimeters of thick scaffolds facilitated the penetration of sufficient radiation through the scaffolds (FIG. 3C). The functionality of different thicknesses, irradiated with 1.5 W/m² during 120 seconds, is shown in FIG. 3D. As shown, a thickness of 60- 80 pm facilitates significantly stronger attachment to a tissue, as compared to 100-120 pm probably due to better penetration of the light.

Example 4

Demonstration of Cells’ Viability

Non-destructiveness for cells of the thermal energy generated by the AuNRs after irradiated with near IR for 120 seconds, at 1.5 W/cm² was demonstrated by isolating cardiac cells from the left ventricles of neonatal rat hearts and seeding them within the hybrid scaffolds. After 7 days of incubation, which is considered sufficient period for cells’ organization and tissue assembly, the cardiac patches prepared as presented herein, were irradiated. As shown in FIG. 4E, exposure to near IR did not affect cells’ viability. Following tissue irradiation, the patches were fixed and immunostained for cardiac sarcomeric actinin and connexin 43 (Cx43), proteins associated with cell contraction and electrical coupling, respectively. As shown by confocal microscope images, the cardiac cells exhibited massive actinin staining, indicating on the contraction ability of the assembled tissue. Furthermore, pronounced Cx43 expression was observed between the cardiomyocytes, indicating on the ability of the cells to function synchronously. Such hallmarks further indicated that the exposure to near IR and the local heating by the AuNRs did not harm the engineered tissue. Demonstration for attachment to a heart in wet and dynamic conditions was carried by opening the chest of Sprague Dawley rats, exposing the heart and placing a patch prepared as presented herein on the left ventricle. Then the patch was irradiated with near IR for 120 seconds as illustrated in FIG. 1A. The adhesion of the thin edges of the patch to the heart can be well appreciated from the image in FIG. 1B. Scanning electron microscope images of the sliced heart reveal a tight interaction between the heart and the patch (FIG. 1C). Furthermore, the thin edges were able to strongly secure the thick tissue layer on the heart, ensuring a strong fixation (FIG. 4C-E). As shown, no changes in the scaffolds’ structure could be observed. This further indicates on the safety of the procedure.

In conclusion, the physical properties of the AuNRs were utilized to provide a suture- free engraftment of a cardiac patch to the heart. Such technology embodying aspects of the present invention, may be implemented to integrate various engineered tissues or pure biomaterials with other defected organ, minimizing the risk of additional injury for the patient.

Although the invention has been described in conjunction with specific embodiments thereof, it is evident that many alternatives, modifications and variations will be apparent to those skilled in the art. Accordingly, it is intended to embrace all such alternatives, modifications and variations that fall within the spirit and broad scope of the appended claims.

All publications, patents and patent applications mentioned in this specification are herein incorporated in their entirety by reference into the specification, to the same extent as if each individual publication, patent or patent application was specifically and individually indicated to be incorporated herein by reference. In addition, citation or identification of any reference in this application shall not be construed as an admission that such reference is available as prior art to the present invention. To the extent that section headings are used, they should not be construed as necessarily limiting.

Claims

WHAT IS CLAIMED IS:

1. A composition-of-matter, comprising:

a porous scaffold that comprises a polymer; and

a plurality of particles attached to said polymer,

wherein said scaffold is having at least one region that is translucent to radiation suitable for absorption by said particles, said particles are capable of absorbing said radiation and generate heat upon said absorbing, and said heat renders said scaffold reactive to bonding with a tissue in a subject.

2. The composition of claim 1, wherein said at least one region is located at a perimeter, an edge, a frame, and/or an end of said scaffold.

3. The composition of claim 1, wherein said at least one region is located where said scaffold is making a close intimate contact with said tissue.

4. The composition of any one of claims 1-3, wherein an amount of said particles ranges from 0.001 mg to 10 mg per 1 gram of said scaffold.

5. The composition of any one of claims 1-4, wherein said radiation is near infrared radiation having a wavelength that ranges from 700 nm to 950 nm.

6. The composition of any one of claims 1-5, wherein said region admits at least 20 % of said radiation.

7. The composition of any one of claims 1-6, wherein said particles comprise a substance and/or characterized by a size and/or have a shape that promotes said radiation absorption said and said heat generation.

8. The composition of claim 7, wherein said shape is an anisotropic shape.

9. The composition of claim 7, wherein said size ranges from 0.5 nm to 2 pm.

10. The composition of claim 7, wherein said substance is having a physical property selected from the group consisting of electric conductance, anisotropy, ferromagnetism, ferrimagnetism, diamagnetism, superparamagnetism or paramagnetism.

11. The composition of claim 10, wherein said particles comprise a metal and/or a metal oxide.

12. The composition of claim 11, wherein said metal is selected from the group consisting of gold, silver, aluminum, iron, nickel, copper, platinum, titanium, and any blends and alloys thereof.

13. The composition of claim 10, wherein said particles comprise an organic material.

14. The composition of claim 10, wherein said particles comprise a substance selected from the group consisting of a carbon allotrope, a composite material, polyaniline, polypyrrole, and blends thereof.

15. The composition of any one of claims 7-12, wherein said particles have a core coat structure.

16. The composition of claim 15, wherein a substance of said core and/or said coat is selected from the group consisting of a conductive substance, a dielectric substance, a metal, an organic substance, an inorganic substance, and any mixture thereof.

17. The composition of claim 15, wherein a substance of said core and/or said coat is selected from the group consisting of gold, silver, an organic polymer, a ceramic and a metal oxide.

18. The composition of claim 12, wherein said metal is gold.

19. The composition of claim 18, wherein said particles are having an aspect ratio that ranges from 2 to 7.

20. The composition of claim 19, wherein said radiation is near IR characterized by a wavelength that ranges from 750 nm to 850 nm.

21. The composition of claim 20, wherein at said at least one region said scaffold is having a thickness that ranges from 50 pm to 100 pm.

22. The composition of any one of claims 1-21, wherein said scaffold is having a form selected from the group consisting of a patch, a sheet, a mesh, a rod, a tube, a ring, and a three-dimensional object.

23. The composition of any one of claims 1-22, wherein said polymer is having a shape selected from the group consisting of fibers, ribbons, flakes, spheres, coils, helicoils, beads-on-a-string, and any combination thereof.

24. The composition of claim 23, wherein said ribbons are having a mean width that ranges from 100 nm to 10 pm, and mean thickness that ranges from 100 nm to 10 pm.

25. The composition of claim 23, wherein said fibers are having a thickness that ranges from 1 pm to 0.1 pm.

26. The composition of any one of claims 24 or 25 wherein said polymer is in a form of electrospun fibers or ribbons.

27. The composition of any one of claims 1-26, wherein said polymer exhibits a plurality of functional groups capable of binding said particles.

28. The composition of any one of claims 1-27, wherein said polymer exhibits a plurality of functional groups capable of said bonding upon exposure to said heat.

29. The composition of any one of claims 1-28, wherein said polymer comprises a protein, a polysaccharide, alginate, chitosan, a polycaprolactone, polyglycolic acid, poly(L- lactide), a poly(ester urethane), polydioxanone, polyethylene glycol, and any copolymer thereof.

30. The composition of claim 29, wherein said protein is selected from the group consisting of albumin, collagen, fibrin, elastin, fibronectin, glycoprotein, laminin, silk, and any combination thereof.

31. The composition of any one of claims 1-30, wherein said polymer comprises albumin.

32. The composition of claim 31, wherein said polymer comprises electrospun albumin fibers.

33. The composition of any one of claims 1-32, wherein said porous scaffold is having an average pore size that ranges from 50 nm to 2 mm.

34. The composition of any one of claims 1-33, further comprising a plurality of cells distributed within and/or on said scaffold.

35. The composition of claim 34, wherein said cells are viable cells selected from the group consisting of seed cells, stem cells, mature cells, and a reconstructed tissue.

36. The composition of claim 35, wherein said cells correspond to said tissue in said subject.

37. The composition of any one of claims 1-36, wherein said polymer comprises electrospun albumin fibers, said plurality of particles comprises gold particles having an anisotropic shape characterized by an aspect ratio of at least 2, and further comprising a plurality of viable cells distributed within and/or on said scaffold.

38. The composition of any one of claims 1-37, wherein said tissue in said subject is selected from the group consisting of a connective tissue, a muscle tissue, a nervous tissue and an epithelial tissue.

39. The composition of claim 38, wherein said muscle tissue is selected from the group consisting of a smooth muscle, a visceral muscle, a skeletal muscle, and cardiac muscle.

40. The composition of claim 38, wherein said connective tissue is selected from the group consisting of an extracellular matrix, a fibrous connective tissue, a skeletal connective tissue, a fascia, a bone, a tendon, a ligament, an adipose tissue and an areolar tissue.

41. A method of fusing the composition-of-matter of any one of claims 1-40 to a tissue in a subject in need thereof, the method comprising:

placing the composition on said tissue; and

irradiating at least said at least one region in the composition with said radiation.

42. The method of claim 41, wherein said radiation is emitted on said composition by a radiation source capable of emitting a flux of at least 1 W/cm².

43. The method of claim 42, wherein said irradiating is effected for at least 60 seconds.

44. The method of claim 42, wherein the composition-of-matter is used as a patch and said tissue in said subject is selected from the group consisting of a cardiac muscle, a visceral muscle, a skeletal muscle, a fascia, and an abdominal wall.

45. A process of manufacturing the composition-of-matter of any one of claims 1-40, the process comprising:

obtaining said porous scaffold; and

contacting said porous scaffold with a dispersion/suspension of said plurality of particles, to thereby obtain said porous scaffold having said plurality of particles attached thereto.

46. The process of claim 45, further comprising, prior or subsequent to said contacting, introducing viable cells into said porous scaffold under conditions suitable for maintaining said viable cells.

47. A method of treating a tissue defect in a subject in need thereof, the method comprising:

contacting the composition-of-matter of any one of claims 1-40 with said tissue in the subject such that at least said at least one region overlaps with a healthy part of said tissue surrounding the tissue defect, and the composition is covering the tissue defect; and irradiating at least said at least one region in the composition with said radiation, thereby fusing the composition to said tissue.

48. The method of claim 47, wherein said radiation is emitted on said composition by a radiation source capable of emitting a flux of at least 1 W/cm².

49. The method of claim 48, wherein said irradiating is effected for at least 60 seconds.

50. The method of any one of claims 47-49, wherein the tissue defect is selected from the group consisting of a damaged cardiac muscle, an inguinal hernia, an incisional hernia, a femoral hernia, a skeletal muscle tear, and a damaged skin.