System and Method for Collagen Isolation

Abstract

System and methods for isolation of collagen and other fibrous tissue from adipose tissue are described herein. The method of the present invention isolates the collagen from adipose tissue by sonication. The tissue to be sonicated is placed in a container or a flow cell transparent to ultrasound waves. After sonication the sonicated material is filtered out through the bottom of the flow cell and the sonicated collagen is trapped in the filter, which may be taken for further processing. The isolated collagen can then be combined with a suitable carrier for re-injection to correct various tissue defects such as wrinkles, to form a carrier for the stem cells, a filler, and matrix for new collagen production by injecting into the desired area of the host.

1 . A method for isolation of one or more connective tissue proteins from one or more fat containing tissues comprising the steps of:
 providing or flowing the fat containing tissue harvested by liposuction, abdominoplasty, or any other fat removal techniques in a container or a flow cell that is jacketed, wherein the container or the flow cell is an acoustically permeable closed system and further comprises a filter;   exposing the fat containing tissue to one or more acoustic pulses to separate the one or more connective tissue proteins from a stromal vascular fraction (SVF), wherein the SVF comprises lipids, cells, or any other extracellular elements; and   isolating the separated connective tissue proteins.   
     
     
         2 . The method of  claim 1 , wherein the isolated connective tissue protein comprises at least one of collagen type I, collagen type II, collagen type III, collagen type IV, collagen type V, collagen type VI, collagen type VII, collagen type VIII, collagen type IX, collagen type X, collagen type XI, collagen type XII, collagen type XIII, collagen type XIV, collagen type XV, collagen type XVI, collagen type XVII, collagen type XVIII, collagen type XIX, collagen type XX, collagen type XXI, collagen type XXII, collagen type XXIII, collagen type XXIV, collagen type XXV, collagen type XXVI, collagen type XXVII, collagen type XXVIII, collagen type XXIX, or elastin. 
     
     
         3 . The method of  claim 1 , further comprising the steps of:
 processing the one or more isolated connective tissue proteins for long-term storage using one or more cryogenic processes with one or more optional preservatives;   reconstituting the one or more isolated connective tissue proteins in vehicle or diluent, wherein the vehicle or diluent is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof;   combining the one or more isolated connective tissue proteins with one or more pharmacologically active or inactive additives selected from at least one of epidermal growth factor, fibroblast growth factor (FGF), nerve growth factor, keratinocyte growth factor, platelet derived growth factor (PDGF), vasoactive intestinal peptide, vascular endothelial growth factor (VEGF), stem cell factor, bone morphogenic proteins, chondrocyte growth factor, analgesic drugs, hemostatic drugs, antibiotic drugs, antioxidants, heparin, anti-inflammatory drugs, local anesthetics, tissue formation agents, adipose tissue formation agents, transforming growth factor, transforming growth factor-β, connective tissue activating peptides, β-thromboglobulin, insulin-like growth factors, tumor necrosis factors, interleukins, colony stimulating factors, erythropoietin, nerve growth factors, interferons, mesenchymal stem cells, epidermal stem cells, cartilage stem cells, adipose derived stem cells (ASCs), hematopoietic stem cells, and any combinations thereof; and   injecting the one or more isolated connective tissue proteins to perform or enhance effectiveness of one or more cosmetic surgery procedures, wherein the one or more cosmetic surgery procedures selected from the group consisting of treatment of wrinkles, deep folds, frown lines, deep scars, lip tissue augmentation, wound healing, facial rejuvenation, regeneration and tissue engineering, cleft palate surgery, carrier for stem cells, filler and matrix for new collagen production, and other combinations and modifications thereof.   
     
     
         4 . A composition for use in one or more cosmetic surgery procedures in a subject comprising:
 one or more connective tissue proteins derived or isolated from one or more fat containing tissues, wherein the connective tissue proteins are isolated by a method comprising the steps of:
 providing or flowing the fat containing tissue harvested by liposuction, abdominoplasty, or any other fat removal techniques in a container or a flow cell that is jacketed, wherein the container or the flow cell is an acoustically transparent closed system and further comprises a mesh filter; 
 exposing the fat containing tissue to one or more acoustic pulses to separate the one or more connective tissue proteins from a stromal vascular fraction (SVF), wherein the SVF comprises lipids, cells, or any other extracellular elements; and 
 isolating the separated connective tissue proteins from the SVF; and 
   a vehicle or medium for dissolving, dispersing, or suspending the one or more tissue proteins, wherein the vehicle or medium is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof.   
     
     
         5 . The composition of  claim 4 , wherein the composition may comprise one or more pharmacologically active or inactive additives selected from the group consisting of epidermal growth factor, fibroblast growth factor (FGF), nerve growth factor, keratinocyte growth factor, platelet derived growth factor (PDGF), vasoactive intestinal peptide, vascular endothelial growth factor (VEGF), stem cell factor, bone morphogenic proteins, chondrocyte growth factor, analgesic drugs, hemostatic drugs, antibiotic drugs, antioxidants, heparin, anti-inflammatory drugs, local anesthetics, tissue formation agents, adipose tissue formation agents, transforming growth factor, transforming growth factor-β, connective tissue activating peptides, β-thromboglobulin, insulin-like growth factors, tumor necrosis factors, interleukins, colony stimulating factors, erythropoietin, nerve growth factors, interferons, mesenchymal stem cells, epidermal stem cells, cartilage stem cells, adipose derived stem cells (ASCs), hematopoietic stem cells, and any combinations thereof. 
     
     
         6 . The composition of  claim 4 , wherein the composition is adapted for use in one or more cosmetic surgery procedures selected from the group consisting of treatment of wrinkles, deep folds, frown lines, deep scars, lip tissue augmentation, wound healing, facial rejuvenation, regeneration and tissue engineering, cleft palate surgery, dermal fillers, carrier for stem cells, filler and matrix for new collagen production, and other combinations and modifications thereof. 
     
     
         7 . The composition of  claim 4 , wherein the acoustic pulses comprise frequencies ranging from subaudio to ultrasonic waves. 
     
     
         8 . The composition of  claim 4 , wherein a source of the one or more acoustic pulses comprises acoustic horns, transducers, piezo-electric crystals, a sonicating probe, or any combinations thereof. 
     
     
         9 . The composition of  claim 4 , wherein the method of isolating the connective tissue proteins is performed under sterile conditions. 
     
     
         10 . The composition of  claim 4 , wherein a frequency for the one or more ultrasonic pulses ranges from 40-500 kHz. 
     
     
         11 . The composition of  claim 4 , wherein a pore size of the mesh filter ranges from 5 microns to 300 microns. 
     
     
         12 . The composition of  claim 4 , wherein a concentration of the vehicle or medium in the composition ranges from 10% to 60%. 
     
     
         13 . The composition of  claim 4 , wherein the composition is formed into a scaffold for seeding of cells into the scaffold. 
     
     
         14 . A method for performing, enhancing effectiveness of a cosmetic surgery procedure, or both in a subject comprising the steps of:
 identifying the subject in need of the cosmetic surgery procedure, enhanced effectiveness of the cosmetic surgery procedure, or both; and   injecting an effective amount of a composition comprising one or more connective tissue proteins derived or isolated from one or more fat containing tissues, wherein the connective tissue proteins are isolated by a method comprising the steps of:
 providing or flowing the fat containing tissue harvested by liposuction, abdominoplasty, or any other fat removal techniques in a container or a flow cell that is jacketed, wherein the container or the flow cell is a transparent closed system and further comprises a mesh filter; 
 exposing the fat containing tissue to one or more acoustic pulses to separate the one or more connective tissue proteins from a stromal vascular fraction (SVF), wherein the SVF comprises lipids, cells, or any other extracellular elements; and 
 isolating the separated connective tissue proteins from the SVF; and 
   a vehicle or medium for dissolving, dispersing, or suspending the one or more tissue proteins, wherein the vehicle or medium is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof.   
     
     
         15 . The method of  claim 14 , wherein the composition may comprise one or more pharmacologically active or inactive additives selected from at least one of epidermal growth factor, fibroblast growth factor (FGF), nerve growth factor, keratinocyte growth factor, platelet derived growth factor (PDGF), vasoactive intestinal peptide, vascular endothelial growth factor (VEGF), stem cell factor, bone morphogenic proteins, chondrocyte growth factor, analgesic drugs, hemostatic drugs, antibiotic drugs, antioxidants, heparin, anti-inflammatory drugs, local anesthetics, tissue formation agents, adipose tissue formation agents, transforming growth factor, transforming growth factor-β, connective tissue activating peptides, β-thromboglobulin, insulin-like growth factors, tumor necrosis factors, interleukins, colony stimulating factors, erythropoietin, nerve growth factors, interferons, mesenchymal stem cells, epidermal stem cells, cartilage stem cells, adipose derived stem cells (ASCs), hematopoietic stem cells, and any combinations thereof. 
     
     
         16 . The method of  claim 14 , wherein the composition is adapted for use in one or more cosmetic surgery procedures selected from the group consisting of treatment of wrinkles, deep folds, frown lines, deep scars, lip tissue augmentation, wound healing, facial rejuvenation, regeneration and tissue engineering, cleft palate surgery, carrier for stem cells, filler and matrix for new collagen production, and other combinations and modifications thereof. 
     
     
         17 . The method of  claim 14 , wherein the one or more fat containing tissues comprises adipose tissue, dermis, collagen, other microvascular tissues including placenta or muscle. 
     
     
         18 . The method of  claim 14 , wherein the acoustic pulses comprise frequencies ranging from subaudio to ultrasonic waves. 
     
     
         19 . The method of  claim 14 , wherein the acoustic pulses comprise ultrasonic waves. 
     
     
         20 . The method of  claim 14 , wherein a source of the one or more acoustic pulses comprises acoustic horns, transducers, piezo-electric crystals, a sonicating probe, or any combinations thereof. 
     
     
         21 . The method of  claim 14 , wherein the method of isolating the connective tissue proteins is performed under sterile conditions. 
     
     
         22 . The method of  claim 14 , wherein a frequency for the one or more ultrasonic pulses ranges from 40-500 kHz. 
     
     
         23 . The method of  claim 14 , wherein a pore size of the filter ranges from 5 microns-300 microns. 
     
     
         24 . The method of  claim 14 , wherein a concentration of the optional vehicle or medium in the composition ranges from 10% to 60%. 
     
     
         25 . The method of  claim 14 , wherein the fat containing tissue is heterologous tissue. 
     
     
         26 . The method of  claim 14 , further comprising the step of morselizing the tissue prior to reinjection into a host or use in vitro. 
     
     
         27 . A sterile process for collagen isolation from adipose tissue comprising the steps of:
 providing fat containing adipose tissue harvested by liposuction, abdominoplasty, or any other fat removal techniques in a jacketed container, wherein the jacketed container comprises a mesh filter and is transparent to permit a passage of one or more ultrasonic waves or pulses and may be a closed system;   exposing the fat containing tissue to one or more ultrasonic waves or pulses to separate the collagen and an extracellular matrix (ECM) from a stromal vascular fraction (SVF), wherein the SVF comprises lipids, cells, or any other extracellular elements; and   isolating the separated collagen and ECM from the SVF;   separating the isolated collagen from the ECM by one or more physical separation techniques selected from the group consisting of centrifugation, filtration, ultrasonic separation, or any combinations thereof;   processing the isolated collagen for long-term storage using one or more cryogenic methods with one or more optional preservatives;   reconstituting the isolated collagen in a vehicle or diluent, wherein the vehicle or diluent is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof;   combining the isolated collagen with one or more pharmacologically active or inactive additives selected from at least one of epidermal growth factor, fibroblast growth factor (FGF), nerve growth factor, keratinocyte growth factor, platelet derived growth factor (PDGF), vasoactive intestinal peptide, vascular endothelial growth factor (VEGF), stem cell factor, bone morphogenic proteins, chondrocyte growth factor, analgesic drugs, hemostatic drugs, antibiotic drugs, antioxidants, heparin, anti-inflammatory drugs, local anesthetics, tissue formation agents, adipose tissue formation agents, transforming growth factor, transforming growth factor-β, connective tissue activating peptides, β-thromboglobulin, insulin-like growth factors, tumor necrosis factors, interleukins, colony stimulating factors, erythropoietin, nerve growth factors, interferons, mesenchymal stem cells, epidermal stem cells, cartilage stem cells, adipose derived stem cells (ASCs), hematopoietic stem cells, and any combinations thereof; and   injecting the isolated collagen to perform or enhance effectiveness of one or more cosmetic surgery procedures, wherein the one or more cosmetic surgery procedures selected from the group consisting of treatment of wrinkles, deep folds, frown lines, deep scars, lip tissue augmentation, wound healing, facial rejuvenation, regeneration and tissue engineering, cleft palate surgery, carrier for stem cells, filler and matrix for new collagen production, and other combinations and modifications thereof.   
     
     
         28 . A composition for use in one or more cosmetic surgery procedures in a subject comprising: collagen derived or isolated from one or more adipose tissues optionally dissolved, dispersed, or suspended in a suitable vehicle or diluent, wherein the vehicle or diluent is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof. 
     
     
         29 . A method for performing, enhancing effectiveness of a cosmetic surgery procedure, or both in a subject comprising the steps of:
 identifying the subject in need of the cosmetic surgery procedure, enhanced effectiveness of the cosmetic surgery procedure, or both;   obtaining a tissue for injection by:   providing or flowing the fat containing adipose tissue in a container or a flow cell, wherein the container or the flow cell is an acoustically transparent closed system and further comprises a filter;   exposing the adipose tissue to one or more ultrasonic waves or pulses to separate the collagen and an extra cellular matrix (ECM) from a stromal vascular fraction (SVF), wherein the SVF comprises lipids, cells, or any other extracellular elements; and   isolating the separated collagen and ECM from the SVF; and   injecting an effective amount of a composition comprising collagen derived or isolated from one or more adipose tissues optionally dissolved, dispersed, or suspended in a suitable vehicle or diluent, wherein the vehicle or diluent is selected from the group consisting of water, buffer, saline, serum and other body fluids, and any combinations thereof.