US20110201117A1 - Forming porous scaffold from cellulose derivatives - Google Patents

Forming porous scaffold from cellulose derivatives Download PDF

Info

Publication number
US20110201117A1
US20110201117A1 US12/809,534 US80953408A US2011201117A1 US 20110201117 A1 US20110201117 A1 US 20110201117A1 US 80953408 A US80953408 A US 80953408A US 2011201117 A1 US2011201117 A1 US 2011201117A1
Authority
US
United States
Prior art keywords
polymer
scaffold
self
substituent
partially substituted
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Abandoned
Application number
US12/809,534
Other languages
English (en)
Inventor
Zhilian Yue
Feng Wen
Hanry Yu
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Agency for Science Technology and Research Singapore
Original Assignee
Agency for Science Technology and Research Singapore
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Agency for Science Technology and Research Singapore filed Critical Agency for Science Technology and Research Singapore
Priority to US12/809,534 priority Critical patent/US20110201117A1/en
Priority to US12/824,105 priority patent/US8283028B2/en
Publication of US20110201117A1 publication Critical patent/US20110201117A1/en
Abandoned legal-status Critical Current

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/56—Porous materials, e.g. foams or sponges
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B32—LAYERED PRODUCTS
    • B32B—LAYERED PRODUCTS, i.e. PRODUCTS BUILT-UP OF STRATA OF FLAT OR NON-FLAT, e.g. CELLULAR OR HONEYCOMB, FORM
    • B32B3/00—Layered products comprising a layer with external or internal discontinuities or unevennesses, or a layer of non-planar shape; Layered products comprising a layer having particular features of form
    • B32B3/26—Layered products comprising a layer with external or internal discontinuities or unevennesses, or a layer of non-planar shape; Layered products comprising a layer having particular features of form characterised by a particular shape of the outline of the cross-section of a continuous layer; characterised by a layer with cavities or internal voids ; characterised by an apertured layer
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/14—Macromolecular materials
    • A61L27/20—Polysaccharides
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
    • C08B11/00—Preparation of cellulose ethers
    • C08B11/20—Post-etherification treatments of chemical or physical type, e.g. mixed etherification in two steps, including purification
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J9/00—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof
    • C08J9/28—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof by elimination of a liquid phase from a macromolecular composition or article, e.g. drying of coagulum
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08L—COMPOSITIONS OF MACROMOLECULAR COMPOUNDS
    • C08L1/00—Compositions of cellulose, modified cellulose or cellulose derivatives
    • C08L1/08—Cellulose derivatives
    • C08L1/10—Esters of organic acids, i.e. acylates
    • C08L1/14—Mixed esters, e.g. cellulose acetate-butyrate
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2201/00—Foams characterised by the foaming process
    • C08J2201/02—Foams characterised by the foaming process characterised by mechanical pre- or post-treatments
    • C08J2201/024—Preparation or use of a blowing agent concentrate, i.e. masterbatch in a foamable composition
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2201/00—Foams characterised by the foaming process
    • C08J2201/02—Foams characterised by the foaming process characterised by mechanical pre- or post-treatments
    • C08J2201/026—Crosslinking before of after foaming
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2201/00—Foams characterised by the foaming process
    • C08J2201/04—Foams characterised by the foaming process characterised by the elimination of a liquid or solid component, e.g. precipitation, leaching out, evaporation
    • C08J2201/048—Elimination of a frozen liquid phase
    • C08J2201/0484—Elimination of a frozen liquid phase the liquid phase being aqueous
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2201/00—Foams characterised by the foaming process
    • C08J2201/04—Foams characterised by the foaming process characterised by the elimination of a liquid or solid component, e.g. precipitation, leaching out, evaporation
    • C08J2201/052—Inducing phase separation by thermal treatment, e.g. cooling a solution
    • C08J2201/0524—Inducing phase separation by thermal treatment, e.g. cooling a solution the liquid phase being aqueous
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2205/00—Foams characterised by their properties
    • C08J2205/04—Foams characterised by their properties characterised by the foam pores
    • C08J2205/044—Micropores, i.e. average diameter being between 0,1 micrometer and 0,1 millimeter
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2207/00—Foams characterised by their intended use
    • C08J2207/10—Medical applications, e.g. biocompatible scaffolds
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2301/00—Characterised by the use of cellulose, modified cellulose or cellulose derivatives
    • C08J2301/08—Cellulose derivatives
    • C08J2301/26—Cellulose ethers
    • C08J2301/28—Alkyl ethers
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T428/00—Stock material or miscellaneous articles
    • Y10T428/249921—Web or sheet containing structurally defined element or component
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T428/00—Stock material or miscellaneous articles
    • Y10T428/249921—Web or sheet containing structurally defined element or component
    • Y10T428/249953—Composite having voids in a component [e.g., porous, cellular, etc.]
    • Y10T428/249978—Voids specified as micro
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T428/00—Stock material or miscellaneous articles
    • Y10T428/249921—Web or sheet containing structurally defined element or component
    • Y10T428/249953—Composite having voids in a component [e.g., porous, cellular, etc.]
    • Y10T428/249978—Voids specified as micro
    • Y10T428/249979—Specified thickness of void-containing component [absolute or relative] or numerical cell dimension

Definitions

  • the present invention relates to porous scaffolds and methods of forming such scaffolds.
  • Macroporous scaffolds are useful for supporting cells as the large pores can facilitate migration and growth of the cells inside the pores and allow sufficient nutrient access and removal of metabolites. It has been reported that when hydroxypropylcellulose (HPC) is crosslinked at different temperatures, both non-porous and microporous HPC hydrogels can be formed. The average pore sizes in reported microporous HPC hydrogels are less than 10 microns. Generally, micropores refer to pores that have an average pore size in the range of 2 to 50 microns, and macropores refer to pores that have an average pore size of larger than 50 microns.
  • HPC hydroxypropylcellulose
  • macroporous scaffolds can be formed from hydroxypropylcellulose partially substituted by a substituent, where the substituent comprises one or more self-crosslinkable groups.
  • the substituent can be allyl isocyanate.
  • the macroporous scaffold formed from such partially substituted hydroxypropylcellulose can also have a relatively high interconnected porosity, such as about 50% or higher.
  • the partially substituted hydroxypropylcellulose may be replaced by another thermo-sensitive polymer precursor, such as methylcellulose partially substituted by a substituent that comprises a self-linkable group, or a pH-sensitive polymer precursor partially substituted by a substituent that comprises a self-linkable group.
  • a scaffold comprising a polymer defining macropores and comprising hydroxypropylcellulose partially substituted by a substituent, the substituent comprising a self-crosslinkable group, the partially substituted hydroxypropylcellulose being crosslinked through the self-crosslinkable group, the macropores having an average pore size of larger than 50 microns and being at least partially interconnected.
  • the polymer may have an interconnected porosity of about 50% or higher.
  • the polymer may have a total porosity of about 80% or higher.
  • the macropores may have a pore size distribution peaking at above 50 microns, such as at about 90 or about 100 microns.
  • the polymer may have an equilibrium water content of about 85%.
  • the polymer may have a Young's modulus of about 10 to about 20 kPa in a hydrated state.
  • the self-crosslinkable group may comprise an unsaturated double carbon-carbon bond.
  • the substituent may comprise allyl isocyanate, methacrylic acid, acrylic acid, or glycidyl methacrylate.
  • the partially substituted hydroxypropylcellulose may have a degree of substitution of less than about 2.5, such as about 2.1.
  • the polymer may be a gel, such as when in a hydrated state.
  • a method of forming a scaffold comprising forming a bicontinuous emulsion comprising a continuous aqueous phase and a continuous polymer phase, the polymer phase comprising hydroxypropylcellulose partially substituted by a substituent, the substituent comprising a self-crosslinkable group; crosslinking the partially substituted hydroxypropylcellulose through the self-crosslinkable group to form a polymer defining at least partially interconnected pores.
  • the substituent may comprise allyl isocyanate, methacrylic acid, acrylic acid, or glycidyl methacrylate.
  • the pores may comprise macropores.
  • the crosslinking may comprise irradiating the emulsion with ⁇ -ray.
  • the crosslinking may comprise crosslinking at least about 90 wt % of the partially substituted hydroxypropylcellulose in the emulsion.
  • Water may be removed from the pores by freeze-drying the polymer. After the freeze-drying, the polymer may have an interconnected porosity of about 50% or higher, and the pores may have an average pore size of larger than 50 microns.
  • the emulsion may comprise about 80 to about 90 wt % of the aqueous phase and about 10 to about 20 wt % of the polymer phase.
  • the partially substituted hydroxypropylcellulose may have a degree of substitution of about 2.5 or less, such as about 2.1.
  • the polymer may be a gel, such as when in a hydrated state.
  • the emulsion may be formed by subjecting a solution comprising water and the partially substituted hydroxypropylcellulose to heat treatment.
  • the heat treatment may comprise heat treatment at a temperature of about 313 K for about 5 minutes.
  • a method of forming a scaffold comprising inducing phase separation in a solution comprising a polymer precursor and water, to form a bicontinuous emulsion comprising a continuous polymer phase and a continuous aqueous phase, the polymer precursor comprising a self-crosslinkable group; crosslinking the polymer precursor through the self-crosslinkable group in the emulsion to form a polymer defining at least partially interconnected macropores.
  • the polymer precursor may be a cellulose derivative.
  • the cellulose derivative may be a methylcellulose derivative, or a hydroxypropylcellulose derivative, such as hydroxypropylcellulose partially substituted by allyl isocyanate.
  • the cellulose derivative may be partially substituted by a substituent that comprises a self-linkable group.
  • the self-crosslinkable group may comprise an unsaturated double carbon-carbon bond.
  • the substituent may comprise allyl isocyanate, methacrylic acid, acrylic acid, or glycidyl methacrylate.
  • the polymer precursor may be thermo-sensitive, and the inducing phase separation may comprise heating the solution.
  • the polymer precursor may be pH-sensitive, and the inducing phase separation may comprise changing pH of the solution.
  • the crosslinking may comprise irradiating the emulsion with ⁇ -ray.
  • the polymer may be a gel, such as when in a hydrated state.
  • a scaffold which comprises a crosslinked polymer that defines macropores.
  • the macropores are at least partially interconnected and have an average pore size of larger than 50 microns.
  • the interconnected porosity may be about 50% or higher.
  • the polymer is formed from a polymer precursor that is responsive to a phase separation stimuli to undergo phase separation in an aqueous solution.
  • the stimuli may be heat or pH change in the solution.
  • the polymer precursor also comprises a self-crosslinkable group so that the polymer molecules are crosslinked through the self-crosslinkable group.
  • the crosslinkable group may be selected so that it will crosslink with each other when irradiated with ⁇ -ray.
  • the polymer precursor may be a cellulose derivative.
  • the cellulose derivative may be a methylcellulose derivative, or a hydroxypropylcellulose derivative, such as hydroxypropylcellulose partially substituted by allyl isocyanate.
  • the cellulose derivative may be partially substituted by a substituent that comprises a self-linkable group.
  • the self-crosslinkable group may comprise an unsaturated double carbon-carbon bond.
  • the substituent may comprise allyl isocyanate, methacrylic acid, acrylic acid, or glycidyl methacrylate.
  • the polymer may have a total porosity of about 80% or higher.
  • the macropores may have a pore size distribution peaking at above 50 microns, such as at about 90 or about 100 microns.
  • the polymer may have an equilibrium water content of about 85%.
  • the polymer may have a Young's modulus of about 10 to about 20 kPa in a hydrated state.
  • the polymer may have a degree of substitution of less than about 2.5, such as about 2.1.
  • the polymer may be a gel, such as when in a hydrated state.
  • FIG. 1 is a data graph showing the dependence of UV absorbance on solution temperature for different sample solutions
  • FIG. 2 is a scanning electron microscopy (SEM) image of a cross-section of a sample scaffold
  • FIGS. 3 and 4 are SEM images of portions of the sample scaffold shown in FIG. 2 , with increased magnification;
  • FIG. 5 is a line graph showing the pore size distribution in two sample scaffolds
  • FIG. 6 is a confocal micrograph of a cross-section of a sample scaffold
  • FIG. 7 is a confocal micrograph of a cross-section of a comparison scaffold
  • FIG. 8 is a confocal micrograph of the sample scaffold of FIG. 6 but stained with a different marker
  • FIG. 9 is a confocal micrograph of the sample scaffold of FIG. 8 but with higher magnification
  • FIG. 10 is a data graph showing the dependence of cell numbers in a sample scaffold on culture time
  • FIGS. 11 and 12 are confocal micrographs of cells cultured in a sample scaffold stained with different markers
  • FIG. 13 is a superposition of FIGS. 11 and 12 ;
  • FIGS. 14 , 15 , 16 and 17 are SEM images of cells cultured in a sample scaffold taken at different times with different magnification, respectively;
  • FIG. 18 is a bar graph showing measured urea synthesis data
  • FIGS. 19 and 20 are bar graphs showing measured comparison data
  • FIGS. 21 , 22 , 23 and 24 are SEM images of cells cultured in a sample scaffold
  • FIG. 25 is a confocal micrograph (transmitted image) of cells cultured in a sample scaffold
  • FIG. 26 is a confocal micrograph of the cells cultured in the sample scaffold of FIG. 25 ;
  • FIG. 27 is an SEM image of the cells cultured in the sample scaffold of FIG. 25 at an earlier time
  • FIG. 28 is a confocal micrograph (transmitted image) of the cells of FIG. 25 at a later time;
  • FIG. 29 is a confocal micrograph of the cells of FIG. 28 ;
  • FIGS. 30 , 31 , 32 and 33 are confocal micrographs of different cells cultured in different sample scaffolds.
  • An exemplary embodiment of the present invention relates to a scaffold that is formed of crosslinked hydroxypropylcellulose partially substituted by allyl isocyanate (H-A).
  • a scaffold is any porous material or structure that can be used to provide a supporting framework to support something else, such as cells or tissues.
  • the pores of the H-A polymer are macropores of an average pore size of larger than 50 microns. The H-A polymer when hydrated may form a gel.
  • the H-A polymer has an interconnected porosity of about 50% or higher, and may have a total porosity of about 80% or higher.
  • the interconnected porosity refers to the extent of connection between adjacent pores, and can be determined by mercury intrusion porosimetry.
  • the macropores may have a pore size distribution peaking at above 50 microns, such as at about 90 or about 100 microns.
  • the peak may be determined from the pore size distribution curves obtained by mercury intrusion porosimetry.
  • the pore sizes and their distribution in a scaffold may be determined using a porosimeter, such as a PASCAL 140 mercury porosimeter, available from Thermo Finnigan®.
  • the pore sizes may be measured when the H-A polymer is in a hydrated state or dehydrated state, depending on the technique used.
  • the H-A polymer may have an equilibrium water content (EWC) of about 85%, and a Young's modulus of about 10 to about 20 kPa in a hydrated state.
  • the H-A molecules may have a degree of substitution (DS) of about 2.5 or less, such as about 2.1.
  • the DS may be adjusted by varying the molar ratio of —OH in HPC to —NCO in allyl isocyanate.
  • the DS may be determined using nuclear magnetic resonance (NMR) spectroscopy, as can be understood by persons skilled in the art.
  • a bicontinuous emulsion is prepared, which contains a continuous aqueous phase and a continuous polymer phase.
  • the polymer phase contains H-A molecules as a polymer precursor.
  • the DS of H-A may be about 2.5 or less, such as about 2.1.
  • the H-A molecules are crosslinked to form a polymer with at least partially interconnected macropores.
  • the emulsion may be irradiated with ⁇ -ray to induce crosslinking between the H-A molecules.
  • the emulsion may be selected and irradiated such that at least about 90 wt % of the H-A molecules (polymer precursor) are crosslinked (i.e. the degree of crosslink is 90 wt %).
  • the H-A polymer will initially form a gel containing water or an aqueous phase in the macropores after crosslinking, which may be removed such as by freeze-drying the gel.
  • the contents of the emulsion may be adjusted so that, after freeze-drying, the resulting polymer has an interconnected porosity of about 50% or higher, and the macropores have an average pore size of larger than 50 microns.
  • the pore size distribution may peak at above about 50 microns, such as at about 90 or about 100 microns.
  • the emulsion may include about 80 to about 90 wt % of the aqueous phase and about 10 to about 20 wt % of the polymer phase (or H-A)
  • the bi-phase emulsion may be formed by subjecting an aqueous solution of H-A to heat treatment at a temperature of about 313 K for about 5 minutes to effect phase separation in the solution.
  • the scaffolds prepared as described herein can have certain benefits and advantages and can be used in various applications such as various medical or biological applications.
  • macroporous hydrogels are prepared in complex procedures and their pores are not interconnected.
  • the pores in these conventional hydrogels also lack depth distribution and tend to stay close to the surface.
  • embodiments of the present invention can provide 3D scaffolds where the interconnected macropores are distributed substantially uniformly in the entire 3D material.
  • the scaffold disclosed herein can be prepared using relatively simple chemistry in a relatively simple fabrication procedure.
  • the process can be performed in a simple one step chemistry under mild temperature conditions, without using any organic solvent or additional agent (such as surfactant or crosslinking agent).
  • cellulose poly (1,4′-anhydro- ⁇ -D-glucopyranose)
  • HPC HPC
  • HPC Hydroxypropylcellulose
  • FDA United State Food and Drug Administration
  • HPC and H-A have high solubility in both water and a wide range of organic solvents. They are relatively easy to functionalize to meet various specific requirements in particular applications.
  • An aqueous solution of HPC or H-A can undergo a unique low critical solution temperature (LCST) phase transition from an isotropic aqueous phase to a metastable bi-phase emulsion. This phase separation allows the formation of macroporous polymers and gels.
  • LCST critical solution temperature
  • microporous hydrogels and nanostructured hydrogels can be formed using other HPC derivatives, it has been recognized that a macroporous hydrogel would have certain benefits over hydrogels with smaller pores and it has been surprisingly discovered that a bi-phase emulsion containing a H-A phase can be conveniently utilized to form macroporous hydrogels.
  • HPC may be partially substituted with another suitable substituent, instead of ally isocyanate.
  • ally isocyanate For example, methacrylic acid, acrylic acid, glycidyl methacrylate, or the like may be a suitable substituent in some applications.
  • thermo-sensitive phase behavior of the H-A polymer precursor makes it convenient to form bicontinuous polymer-rich and water-rich phases in the solution by subjecting the solution to heat treatment. Conveniently, it is not necessary to add any chemical solvent or agent (e.g. surfactant) to form the bicontinuous emulsion.
  • any chemical solvent or agent e.g. surfactant
  • the liquid biphasic structure can be stabilized (solidified) by crosslinking, which in turn may be efficiently effected with ⁇ -ray irradiation.
  • crosslinking of H-A may be effected using other techniques
  • ⁇ -Ray irradiation may be advantageous in some applications. For instance, ⁇ -ray can penetrate deeper into a target material than some other types of curing light.
  • crosslinking in the solution can be activated uniformly at different depths with ⁇ -ray.
  • ⁇ -ray irradiation does not give rise to many undesired side chemical effects and do not require any chemical initiator. Thus, it is a chemically “clean” technique.
  • H-A with a degree of substitution of about 2.5 or less, such as about 2.1 may be advantageous in some applications.
  • H-A polymer precursors with a higher DS tend to have a lower solubility in water but crosslinked H-A polymer with a lower DS may have inferior mechanical properties as compared to H-A polymer with a higher DS.
  • a balance between different desirable properties may be achieved with a DS of about 2.1 in some applications.
  • the mechanical properties such as mechanical strength of the H-A scaffolds may be controlled by varying the degree of crosslinking therein.
  • the degree of crosslinking may be controlled by varying H-A concentration in the precursor solution and the crosslinking time.
  • the ability to control mechanical properties of the H-A scaffolds can be advantageous in some applications. For example, it may be desirable to be able to adjust the mechanical properties to provide suitable mechanical stability and structural integrity for supporting cells.
  • the H-A polymer described herein can form three-dimensional (3D) scaffolds for use as soft tissues.
  • the scaffolds can be made to have various desirable physiochemical, mechanical and bio-interfacial properties.
  • the scaffold may have properties that are suitable for promoting cell adhesion.
  • the scaffolds can be made relatively soft and are inherently hydrophilic.
  • the interconnected macropores can facilitate cell growth into the scaffold with cell densities similar to biological tissues. Due to the 3D interconnected macropores, the scaffolds not only can have relatively high equilibrium water content (EWC), be biocompatible, and sustain gradual release of bioactive molecules, but can also support cell growth, migration, and ultimately, tissue formation.
  • EWC equilibrium water content
  • the scaffolds also allow ready incorporation of extracellular matrix (ECM) cues to regulate cell and tissue functions.
  • ECM extracellular matrix
  • the choices of material and the fabrication procedure can be flexible so that it is possible to introduce 3D ECM cues for regulation of cellular functions.
  • a water soluble precursor can be used for preparing the scaffold, it may be possible to adjust the degradation profile of the scaffold by modifying the side chain chemistry of the polymer.
  • Embodiments of the present invention can also provide 3D scaffolds that have integrated interconnected macroporosity, nanofeatures (surface structures having a nanometer scale dimension, see Examples below), high water content and mechanical integrity suitable for soft tissue engineering.
  • Such scaffolds can exhibit controllable, moderate elasticity, and hydrophilicity, which can be advantageous in providing mechanical stability and structural integrity to cells and tissues.
  • the scaffolds can also have nano-features in the pores, which may play a role in the regulation of cell behavior.
  • the nanofeatures may also be useful for loading of bioactive molecules such as adhesion proteins and growth factors etc to control cell behavior and functions.
  • 3D H-A scaffolds as described herein may be suitable for a wide range of applications in soft tissue engineering and regenerative medicine.
  • Some embodiments of the present invention may also be suitable for application in in vitro substrates for cell culture and studies of cellular behavior in a 3D environment.
  • 3D cell culture environments more closely mimic in vivo microenvironments than 2D culture environments.
  • 3D scaffolds can therefore serve as ECM analogues to provide biomimicry 3D substrates for cell attachment, growth, migration, function and differentiation, or the like.
  • Some embodiments of the present invention may also be suitable for use in in vitro 3D tissue models that are based upon human cells for pathological study and drug testing.
  • Some embodiments of the present invention can be used for in vitro cultivation of cells for the creation of external support organs.
  • the highly interconnected macroporous architecture can provide temporary support to the cultivation of a sufficient cell mass with adequate cell-cell contact.
  • Preliminary results have shown that the primary rat hepatocytes cultured in the scaffolds described herein can maintain their liver-specific functions over a period of one week (see Examples below).
  • Other types of cells can also be cultivated in these scaffolds. Examples for relevant applications include inartificial liver assisted devices (BLAD) to provide essential liver functions for the patients with acute liver failure.
  • BLAD inartificial liver assisted devices
  • a soft, hydrophilic, and macroporous scaffold may be suitable for cell transplantation in soft tissue and organ repair or regeneration applications.
  • Some embodiments of the present invention can be used for high content screening for drug discovery and screening.
  • a cellulosic scaffold can be easily fabricated into microplate configuration to provide uniform 3D microenvironments for various cellular assays.
  • Some embodiments of the present invention can be used for localized and sustained delivery of biologically or pharmaceutically active compounds.
  • the scaffolds are hydrophilic and have a high water retention ability, they can be conveniently used to incorporate and provide sustained release of bioactive molecules or other substances.
  • HPC may be replaced with another thermo-sensitive polymer precursor that can form a stable bicontinuous emulsion in an aqueous solution in response to heat.
  • methylcellulose may be used to replace HPC.
  • a polymer precursor that can form a stable bicontinuous emulsion in an aqueous solution in response to another type of stimuli, such as a change in pH may also be used to replace HPC.
  • a suitable pH-sensitive polymer precursor may be used.
  • allyl isocyanate may be replaced with another suitable substituent that includes a self-crosslinkable group.
  • Self-crosslinkable groups refer to functional groups that can be crosslinked between themselves.
  • a suitable substituent or the crosslinkable group may have an unsaturated C ⁇ C bond for providing a crosslinkable site.
  • a suitable substituent should also have a functional group for bonding to HPC, so that the HPC can be partially substituted by the substituent.
  • Suitable replacement for allyl isocyanate may include methacrylic acid, acrylic acid, glycidyl methacrylate, or the like.
  • the substituent may include a multi-functional monomer.
  • the degree of substitution should be adjusted to retain sufficient thermo-sensitivity in the substituted HPC to allow convenient phase separation, and to allow sufficient crosslinking of the modified HPC during phase separation.
  • a scaffold which comprises a crosslinked polymer that defines macropores.
  • the macropores are at least partially interconnected and have an average pore size of larger than 50 microns.
  • the interconnected porosity may be about 50% or higher.
  • the polymer is formed from a polymer precursor that is responsive to a phase separation stimuli to undergo phase separation in an aqueous solution. The stimuli may be heat or pH change in the solution.
  • the polymer precursor also comprises a self-crosslinkable group so that the polymer are crosslinked through the self-crosslinkable group.
  • the crosslinkable group may be selected so that it will crosslink with each other when irradiated with ⁇ -ray.
  • the polymer precursor may be a cellulose derivative.
  • the cellulose derivative may be a methylcellulose derivative, or a hydroxypropylcellulose derivative, such as hydroxypropylcellulose partially substituted by allyl isocyanate.
  • the cellulose derivative may be partially substituted by a substituent that comprises a self-linkable group.
  • the self-crosslinkable group may comprise an unsaturated double carbon-carbon bond.
  • the substituent may comprise allyl isocyanate, methacrylic acid, acrylic acid, or glycidyl methacrylate.
  • the polymer may have a total porosity of about 80% or higher.
  • the macropores may have a pore size distribution peaking at above 50 microns, such as at about 90 or about 100 microns.
  • the polymer may have an equilibrium water content of about 85%.
  • the polymer may have a Young's modulus of about 10 to about 20 kPa in a hydrated state.
  • the polymer may have a degree of substitution of less than about 2.5, such as about 21.
  • the polymer may be a gel, such as when in a hydrated state.
  • HPC (M n ⁇ 10,000; degree of etherification was about 3.4, as determined by 1 H NMR) was dehydrated by azeotropic distillation in toluene.
  • the dehydrated HPC (2.0 g, 6.0 mmol [OH]) was dissolved in chloroform (100 ml), to which a solution of allyl isocyanate (1.83 ml, 3.5 molar equivalents) in chloroform (10 ml) was added dropwise. After one drop of dibutyltin dilaurate was added as a catalyst. the reaction mixture was stirred at room temperature for about 48 hours, then concentrated using a rotatory evaporator and precipitated into diethyl ether.
  • reaction product was collected by vacuum filtration, and purified by re-dissolution in chloroform and precipitation into diethyl ether.
  • the residual impurities were removed by Soxhlet extraction from diethyl ether.
  • the degree of substitution was 2.1, as calculated by 1 H NMR recorded in CDCl 3 .
  • the temperature-mediated phase behavior of the sample H-A solutions at 10 wt % and 20 wt % (weight percentage of H-A in the solution) was investigated on a UV/VIS/NIR spectrophotometer (JascoTM, V-570, Japan), by measuring the optical densities at 480 nm as a function of temperature.
  • the temperatures of sample holders were controlled using a Jasco PSC-498 temperature controller.
  • the samples were allowed to reach equilibrium at each temperature for 10 min before the readings were taken. Representative measurement results are shown in FIG. 1 .
  • the occurrence of phase separation was indicated by reduction in optical density.
  • the temperature at which the reduced optical density reached a plateau was selected as the operating temperature for inducing phase separation of H-A solutions and subsequent crosslinking.
  • FIG. 1 shows representative measured data indicating the temperature-dependences of normalized (normalized to the absorption intensity at 298 K) UV absorption of sample solutions containing different concentrations of HPC (diamonds: 10 wt %-solid, 20 wt %-hollow) and H-A (triangles: 10 wt %-solid, 20 wt %-hollow).
  • HPC diamonds: 10 wt %-solid, 20 wt %-hollow
  • H-A triangles: 10 wt %-solid, 20 wt %-hollow
  • the H-A samples became white and opaque when the temperature was increased, but without any noticeable sedimentation over the temperature range studied, indicating the formation of stable colloidal systems in the H-A solutions.
  • Stable colloidal formation can be advantageous for subsequent generation of 3D open porous structures.
  • 313 K was selected as a suitable temperature for induction of phase separation and crosslinking of H-A solutions.
  • Sample solutions of H-A and water were prepared.
  • the solutions contained about 10 or about 20 wt % of H-A respectively.
  • the water was degassed and deionised before mixed with H-A.
  • a glass vials (10 mm in diameter ⁇ 50 mm height) containing a sample H-A solution was placed in a water bath at 313 K for 5 min to induce phase separation in the solution to form a bicontinuous emulsion.
  • the emulsion was transferred, in a beaker containing water at 313 K, to a gamma irradiator (Gammacell 220, MDS NordionTM, Canada), and subjected to ⁇ -ray irradiation at a dose rate of 10 kGy h ⁇ 1 for 30 min, to crosslink the polymeric phase in the emulsion to form a gel.
  • a gamma irradiator Gammacell 220, MDS NordionTM, Canada
  • the crosslinked gel was freeze-dried, washed with deionised water for one week with daily water change, and lyophilized for storage.
  • Sample I The sample polymer (scaffold) formed with 10 wt % H-A is referred to as Sample I, and the sample polymer (scaffold) formed with 20 wt % H-A is referred to as Sample II.
  • H-A solutions as prepared in Example II were also crosslinked at temperatures from about 273 to about 277 K, without phase separation, by irradiation as described in Example II.
  • the resulting products were homogenous gels and were subjected to similar post-irradiation treatment as described in Example II. These gels were referred to as Sample IC (for 10 wt % H-A) and Sample IIC (for 20 wt % H-A).
  • the degree of crosslink in the samples was expressed as the weight percentage of the crosslinked fraction of the scaffold.
  • Residual soluble polymers were removed by Soxhlet extraction with acetone for 4 h.
  • the lyophilized scaffolds were allowed to swell in water at room temperature for 48 h. The samples were weighed before and after this hydration process. The EWC of the sample scaffolds were determined based on their dry and hydrated weights, according to the equation discussed above.
  • the Mechanical properties of hydrated scaffolds were evaluated by compression tests on an InstronTM Micro-Tester 5848 (Instron Co., Canton, Mass., U.S.A.), at a speed of 0.5 mm/min and a temperature of 25 ⁇ 2° C.
  • the compression moduli were calculated from the slopes of the initial linear portion of the stress-strain curves.
  • the pore size distribution in the sample scaffolds was determined using a PASCAL 140 mercury porosimeter (Thermo Finnigan, Italy, S.p.A.) with S-CD6 dilatometer.
  • Cross-sectional surfaces of sample lyophilized H-A scaffolds were sputter coated with gold and examined on a field-emission scanning electron microscope (FESEM) (JEOLTM, JSM-7400M, Japan) at an accelerating voltage of 20 kV.
  • FESEM field-emission scanning electron microscope
  • the morphology of the hydrated H-A scaffolds was investigated by laser confocal fluorescence microscopy.
  • the freeze-dried scaffolds were stained overnight with fluorescein isothiocyanate conjugated dextran (FITC-dextran) (1.0 mg ml ⁇ 1 ) or propdium iodide (PI) (50 ⁇ g ml ⁇ 1 ) in the phosphate-buffered saline (PBS), respectively.
  • FITC-dextran fluorescein isothiocyanate conjugated dextran
  • PI propdium iodide
  • peak pore size refers to the location of the highest peak in the pore distribution, and is the pore size that occurs most frequently in a porous material.
  • FIG. 2 is a representative SEM image of Sample II at a magnification factor of 90. Further magnified SEM images of a portion of the image of FIG. 2 are shown in FIGS. 3 (500 magnification) and 4 (15000 magnification). Similar interior morphology of interconnected macroporosity were also observed in Sample I. Nano-scale features and structures on the pore surfaces, such as edges, spikes in the regions connecting the macropores, were also visible in the images.
  • the sample scaffolds formed of hydrophobically modified H-A had interconnected macropores.
  • the H-A samples also retained the phase behavior characteristics of HPC, as indicated by the formation of stable, opaque colloidal system over the temperature range studied.
  • Both samples had a broad and bimodal distribution of pore sizes, as shown in FIG. 5 (Sample I—solid; Sample II—hollow), with 50% to 60% interconnected porosity respectively.
  • the pore size distribution had two peaks, a taller and sharper peak at a smaller pore size and a shorter and broader peak at a larger pore size.
  • the taller peak in the pore size distribution curve was shifted to the left (smaller pore size) and the higher peak was shifted to the right (larger pore size); and the taller peak became taller and the shorter peak became shorter.
  • PI-stained Sample II The highly macroporous architecture in hydrated Sample II was also confirmed in another staining study using propidium iodide (PI). Representative images of PI-stained Sample II are shown in FIGS. 8 and 9 .
  • the sizes of water-filled pores varied broadly from tens to hundreds microns and the thickness of water-swollen struts ranged from about 5 to about 20 microns.
  • Samples I and II were cut into discs of about 10 mm in diameter and about 2 mm in thickness.
  • NIH 3T3, HepG-2 and MCF-7 cells were cultured according to the standard procedure in high glucose Dulbecco's modified Eagle's medium (DMEM) (InvitrogenTM, Singapore) supplemented with 10% fetal bovine serum, 100 unit/ml penicillin and 100 ⁇ g/ml streptomycin in a humid incubator at 37° C. and 5% CO 2 , respectively.
  • DMEM high glucose Dulbecco's modified Eagle's medium
  • Hepatocytes were harvested from male Wistar rats weighing 250-300 g by a two-step in situ collagenase perfusion method, as described in P. O, Seglen, Methds Cell Biol., 1976, vol. 13, p. 29. Viability of the hepatocytes was ⁇ 90%, as determined by Trypan Blue Exclusion assay.
  • Hepatocytes cultured in the scaffolds were maintained in William's E supplemented with 1 mg/ml bovine serum albumin (BSA), 10 ng/ml epidermal growth factor (EGF), 0.5 ⁇ g/ml insulin, 5 nM dexamethasone, 50 ng/ml linoleic acid, 100 unit/ml penicillin and 100 ⁇ g/ml streptomycin.
  • BSA bovine serum albumin
  • EGF epidermal growth factor
  • 0.5 ⁇ g/ml insulin 0.5 ⁇ g/ml insulin
  • 5 nM dexamethasone 50 ng/ml linoleic acid
  • penicillin 100 ⁇ g/ml streptomycin.
  • the sample scaffold discs were sterilized in a 12 well plate by gamma irradiation at a dose rate of 10 kGy/h for 4 h.
  • Cell seeding was conducted at a density of 0.5 ⁇ 1.0 ⁇ 10 6 cells per scaffold, by adding 20 ⁇ l of concentrated cell suspension into each scaffold followed by 200 ⁇ l of cell medium.
  • Cell-free scaffold was used as control for all the following work.
  • the scaffolds were incubated for 3 h, after which time 1 ml of cell medium was added, respectively.
  • the viability of cells cultivated in the scaffolds was assessed by fluorescence live/dead staining.
  • the scaffolds were incubated for 30 min with 5 ⁇ M Calcein AM (Molecular probe, USA) and 25 ⁇ g/ml of propdium iodide in Dulbecco's modified essential medium (DMEM) at 37° C., 5% CO 2 . Images of live (green) and dead (red) cells were then acquired by laser confocal fluorescence microscopy. Optical sectioning with a Z-resolution of 2 ⁇ m was used to obtain a 3D image stack of scaffolds.
  • DMEM Dulbecco's modified essential medium
  • Proliferation of NIH 3T3 seeded in sample scaffolds were assessed by monitoring their metabolic activities using almarBlueTM assay.
  • the cell-seeded and cell-free scaffolds were incubated for 4 hours with 10% (v/v) almarBlue(Biosource) in phenol-red free, supplemented DMEM medium.
  • 200 ⁇ l of the media from each sample was transferred to a 96-well plate, and the absorbance at 570 nm and 600 nm were measured using a SunriseTM microplate reader (Tecan, Switzerland).
  • the reduction of almarBlue of each sample was calculated, and converted to cell numbers based upon a standard curve constructed from 2D cell culture with known cell numbers.
  • Sample II was selected for cellular compatibility tests. A detailed cell viability and proliferation test was conducted on the mouse fibroblast NIH 3T3 cultivated in the scaffold, by measuring the metabolic activity using Alamar Blue assay. The cells were found to be not only viable but also to proliferate well over prolonged culture. The number of cells over time in Sample II was plotted in the graph of FIG. 10 .
  • FIG. 11 is a fluorescence microscopy image of NIH 3T3 cells cultured in Sample II after 4 weeks of cultivation. The cells were stained with Calcein AM (shown as bright regions in FIG. 11 ) for live cells.
  • FIG. 12 is a similar image but for cells stained with PI (shown as lighter spots in FIG. 12 ) for dead cells.
  • FIG. 13 is a superposition of the images of FIGS. 11 and 12 .
  • FIGS. 14 and 15 show SEM images of NIH 3T3 cultured in Sample II for one day at different magnifications.
  • FIGS. 16 and 17 show SEM images of NIH 3T3 cultured in Sample II for five days at different magnifications.
  • the scale bars represent 100 microns in FIGS. 14 and 16 , and 10 microns in FIGS. 15 and 17 .
  • the cells cultured in the sample scaffolds were incubated in a culture medium containing 1.0 mM NH 4 Cl for 90 min; the medium was analyzed by the Urea Nitrogen Kit (Sigma Diagnostics), as described in S, Ng et al., Biomaterials, 2005, vol. 26, p. 3163. The data was normalized by the number of cells seeded in the scaffolds that was quantified using Quan-iTTM PicoGreen dsDNA Assay Kit (Invitrogen, Singapore).
  • FIG. 18 shows the measured result.
  • FIGS. 19 and 20 show albulmin secretion and urea synthesis of the primary rat hepatocytes cultured in Sample II, in comparison to 2D collagen monolayer culture.
  • 2D collagen monolayer has been conventionally used for hepatocyte culture.
  • the sustained liver-specific functions of primary rat hepatocytes cultured in the scaffolds were different in the two tested scaffolds.
  • the albumin secretion and urea products of the hepatocytes cultivated in the 3D Sample II were higher than those cultured in the 2D collagen monolayer.
  • FIGS. 21 , 22 , 23 and 24 show SEM images of primary rat hepatocytes cultured in Sample II, taken over a period of 7 days, and the morphologies of these cells in Sample II. The cells appeared round and tended to form cellular aggregates in the scaffold, well supported by cell-cell and cell-matrix interactions.
  • Fluorescence viability staining was also carried out on human MCF-7 breast cancer cells (MCF-7), human hepatoblastoma cells (C3A), showing good cellular biocompatibility.
  • FIG. 25 is a transmitted image of C3A cells cultured in sample 3D scaffolds for two days.
  • the C3A cells formed spheroids in the scaffolds.
  • FIG. 26 is a fluorescence image of live/dead staining of the C3A spheroids at day 2 .
  • FIG. 27 is an SEM image of the C3A spheroids at day 1 and
  • FIG. 28 is a transmitted image of the C3A spheroids at day 7 .
  • FIG. 29 is a fluorescent image of live/dead staining of C3A spheroids at day 7 .
  • FIG. 30 shows an image of live/dead staining of human foreskin fibroblasts cultured in an unmodified sample scaffold at day 1 ;
  • FIG. 31 shows an image of live/dead staining of human foreskin fibroblasts cultured in an collagen conjugated sample scaffold at day 1 ;
  • FIG. 32 shows an image of live/dead staining of human umbilical vein endothelial cells cultured in the unmodified scaffold at day 1 ;
  • FIG. 33 shows an image of live/dead staining of human umbilical vein endothelial cells cultured in the collagen conjugated sample scaffold at day 1 .

Landscapes

  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Organic Chemistry (AREA)
  • Polymers & Plastics (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Oral & Maxillofacial Surgery (AREA)
  • Transplantation (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Dermatology (AREA)
  • Epidemiology (AREA)
  • Materials Engineering (AREA)
  • Engineering & Computer Science (AREA)
  • Dispersion Chemistry (AREA)
  • Biochemistry (AREA)
  • Manufacture Of Porous Articles, And Recovery And Treatment Of Waste Products (AREA)
  • Materials For Medical Uses (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
  • Immobilizing And Processing Of Enzymes And Microorganisms (AREA)
US12/809,534 2007-12-18 2008-12-18 Forming porous scaffold from cellulose derivatives Abandoned US20110201117A1 (en)

Priority Applications (2)

Application Number Priority Date Filing Date Title
US12/809,534 US20110201117A1 (en) 2007-12-18 2008-12-18 Forming porous scaffold from cellulose derivatives
US12/824,105 US8283028B2 (en) 2007-12-18 2010-06-25 Forming porous scaffold from cellulose derivatives

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US609007P 2007-12-18 2007-12-18
PCT/SG2008/000491 WO2009078819A1 (fr) 2007-12-18 2008-12-18 Formation d'un bâti poreux à partir de dérivés de cellulose
US12/809,534 US20110201117A1 (en) 2007-12-18 2008-12-18 Forming porous scaffold from cellulose derivatives

Related Parent Applications (1)

Application Number Title Priority Date Filing Date
PCT/SG2008/000491 A-371-Of-International WO2009078819A1 (fr) 2007-12-18 2008-12-18 Formation d'un bâti poreux à partir de dérivés de cellulose

Related Child Applications (1)

Application Number Title Priority Date Filing Date
US12/824,105 Continuation-In-Part US8283028B2 (en) 2007-12-18 2010-06-25 Forming porous scaffold from cellulose derivatives

Publications (1)

Publication Number Publication Date
US20110201117A1 true US20110201117A1 (en) 2011-08-18

Family

ID=40795781

Family Applications (3)

Application Number Title Priority Date Filing Date
US12/809,534 Abandoned US20110201117A1 (en) 2007-12-18 2008-12-18 Forming porous scaffold from cellulose derivatives
US12/824,105 Active 2029-06-29 US8283028B2 (en) 2007-12-18 2010-06-25 Forming porous scaffold from cellulose derivatives
US13/611,339 Active 2029-03-08 US8993537B2 (en) 2007-12-18 2012-09-12 Forming porous scaffold from cellulose derivatives

Family Applications After (2)

Application Number Title Priority Date Filing Date
US12/824,105 Active 2029-06-29 US8283028B2 (en) 2007-12-18 2010-06-25 Forming porous scaffold from cellulose derivatives
US13/611,339 Active 2029-03-08 US8993537B2 (en) 2007-12-18 2012-09-12 Forming porous scaffold from cellulose derivatives

Country Status (4)

Country Link
US (3) US20110201117A1 (fr)
EP (1) EP2234655B1 (fr)
JP (1) JP5420565B2 (fr)
WO (1) WO2009078819A1 (fr)

Families Citing this family (15)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20130142763A1 (en) * 2010-10-27 2013-06-06 Empire Technology Development Llc Crosslinked cellulosic polymers
US11284676B2 (en) 2012-06-13 2022-03-29 John C. S. Koo Shoe having a partially coated upper
EP2882392B1 (fr) 2012-08-08 2019-03-20 Smith & Nephew PLC Appareils et procédés de traitement de plaie sur mesure destinés à être utilisés dans une thérapie de plaie par pression négative
US9182420B2 (en) * 2013-03-15 2015-11-10 Palo Alto Research Center Incorporated Phase-change enabled flow field visualization
US10150258B2 (en) 2013-07-29 2018-12-11 Carnegie Mellon University Additive manufacturing of embedded materials
CN106456376B (zh) 2014-01-21 2020-12-15 史密夫及内修公开有限公司 伤口治疗设备
EP3023455A1 (fr) * 2014-11-21 2016-05-25 Fundacíon Tecnalia Research & Innovation Matériaux cellulosiques poreux et leur procédé de préparation
US11220597B2 (en) 2015-06-01 2022-01-11 The Trustees Of The University Of Pennsylvania Bijels and methods of making the same
MX2016013830A (es) * 2015-10-27 2017-05-10 Shinetsu Chemical Co Eter de celulosa porosa no ionico soluble en agua que tiene excelente solubilidad y metodo para producir el mismo.
CN108102129B (zh) * 2016-11-25 2021-01-01 山西加乐医疗科技有限责任公司 一种纤维素海绵的制备方法
US10928383B2 (en) * 2017-06-03 2021-02-23 Shanxi Pishon Biomedical Technology Co., Ltd Method for predicting effect of drug
IT201700087978A1 (it) 2017-07-31 2019-01-31 Univ Degli Studi Genova Scaffold di idrogel tridimensionale per colture cellulari e metodo per la sua produzione
CN110433784A (zh) * 2019-08-15 2019-11-12 毛艳艳 一种工厂污水用环保处理功能材料的制备方法
US11786879B2 (en) 2019-11-11 2023-10-17 The Trustees Of The University Of Pennsylvania Three dimensional multiphasic structures via vaporization induced phase separation (VIPS)
CN111840653B (zh) * 2020-05-27 2021-07-20 华南理工大学 能包埋亲水性功能因子的甘草酸基缓释多孔泡沫材料及其制备方法与应用

Family Cites Families (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4964989A (en) * 1986-12-23 1990-10-23 Pall Corporation Membranes with low protein adsorbability
JP2802103B2 (ja) * 1989-07-03 1998-09-24 アラコ株式会社 板状装飾部材の製造方法
GB9006726D0 (en) * 1990-03-26 1990-05-23 Ici Plc Micro-emulsions
US5573994A (en) * 1994-05-13 1996-11-12 University Of Cincinnati Superabsorbent foams, and method for producing the same
AU5236400A (en) * 1999-06-22 2001-01-09 Avecia Limited Ink receptive substrates
CA2440844A1 (fr) 2001-03-20 2002-09-26 Universitat Zurich Traitement en deux phases de polymeres thermosensibles destines a etre utilises comme biomateriaux
WO2003022910A1 (fr) 2001-09-08 2003-03-20 Access Pharmaceuticals, Inc. Synthese et utilisations de reseaux de nanoparticules de gel polymere

Also Published As

Publication number Publication date
EP2234655A1 (fr) 2010-10-06
WO2009078819A1 (fr) 2009-06-25
US20130005945A1 (en) 2013-01-03
JP5420565B2 (ja) 2014-02-19
US20110159272A1 (en) 2011-06-30
US8283028B2 (en) 2012-10-09
EP2234655A4 (fr) 2013-10-16
EP2234655B1 (fr) 2018-12-12
US8993537B2 (en) 2015-03-31
JP2011505871A (ja) 2011-03-03

Similar Documents

Publication Publication Date Title
EP2234655B1 (fr) Formation d'un bâti poreux à partir de dérivés de cellulose
US11511016B2 (en) Method for preparing porous scaffold for tissue engineering, cell culture and cell delivery
Aki et al. 3D printing of PVA/hexagonal boron nitride/bacterial cellulose composite scaffolds for bone tissue engineering
Reverchon et al. Supercritical fluids in 3-D tissue engineering
Salerno et al. Bio-safe fabrication of PLA scaffolds for bone tissue engineering by combining phase separation, porogen leaching and scCO2 drying
Yue et al. Preparation of three-dimensional interconnected macroporous cellulosic hydrogels for soft tissue engineering
Ma et al. Collagen/chitosan porous scaffolds with improved biostability for skin tissue engineering
KR100664772B1 (ko) 생분해성 중합체 스캐폴드
DE69917339T2 (de) Poröse polymere stützgewebe für gewebetechnologie
Lv et al. Three-dimensional fibroin/collagen scaffolds derived from aqueous solution and the use for HepG2 culture
Panjapheree et al. Biphasic scaffolds of silk fibroin film affixed to silk fibroin/chitosan sponge based on surgical design for cartilage defect in osteoarthritis
Zawko et al. Crystal templating dendritic pore networks and fibrillar microstructure into hydrogels
Rakin et al. Tunable metacrylated hyaluronic acid-based hybrid bioinks for stereolithography 3D bioprinting
Eigel et al. Cryogel biomaterials for neuroscience applications
Agostino et al. Semi-interpenetrated hydrogels composed of PVA and hyaluronan or chondroitin sulphate: chemico-physical and biological characterization
Zhang et al. A highly transparent, elastic, injectable sericin hydrogel induced by ultrasound
Li et al. Fabrication, characterization, and in vitro evaluation of biomimetic silk fibroin porous scaffolds via supercritical CO2 technology
Silvestro et al. Chitosan scaffolds with enhanced mechanical strength and elastic response by combination of freeze gelation, photo-crosslinking and freeze-drying
Zhao et al. Chitosan/silk fibroin biomimic scaffolds reinforced by cellulose acetate nanofibers for smooth muscle tissue engineering
Nizan et al. Reinforcement of hydroxyethyl cellulose/poly (vinyl alcohol) with cellulose nanocrystal as a bone tissue engineering scaffold
Wang et al. Synthesis of a photocurable acrylated poly (ethylene glycol)-co-poly (xylitol sebacate) copolymers hydrogel 3D printing ink for tissue engineering
Budharaju et al. Efficient dual crosslinking of protein–in–polysaccharide bioink for biofabrication of cardiac tissue constructs
Kang et al. In situ crosslinkable collagen-based hydrogels for 3D printing of dermis-mimetic constructs
Rzhepakovsky et al. Composite of bacterial cellulose and gelatin: A versatile biocompatible scaffold for tissue engineering
Shi et al. Activated carbon fibers/poly (lactic-co-glycolic) acid composite scaffolds: preparation and characterizations

Legal Events

Date Code Title Description
STCB Information on status: application discontinuation

Free format text: ABANDONED -- INCOMPLETE APPLICATION (PRE-EXAMINATION)