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Composite for Thermo-Sensitive Cell-Tissue Transplanted Scaffold and Use thereof

a technology of thermosensitive cells and scaffolds, applied in the field of composites for thermosensitive celltissue transplanted scaffolds, can solve the problems of unsatisfactory commercialization of the above-mentioned technology, the limitation of the amount of available skin, and the possibility of donor site morbidity and other problems, to achieve the effect of convenient stripping

Inactive Publication Date: 2008-11-06
VETERANS GEN HOSPITAL TAIPEI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0007]The present invention provides a composite comprising (a) a stem cell which differentiates into mature normal cell, osteocyte, chondrocyte, adipocyte, epithelium cell, epidermis-related cell, keratocyte, neuron, neural cell, insulin-positive cell, glucagons-positive cell, or tissues thereof; (b) a biodegradable layer for the stem cell to grow and differentiation, wherein the layer contains material selected from the group consisting of gelatin, fibronectin, collagen, laminin, bFGF, EGF, insulin, progesterone, glucose, SDF and thymosin beta-4; and (c) a N-isopropylacrylamide (NIPAAm), which polymerizes with the biodegradable layer to provide the feature of thermo-sensitive response for easy stripping.

Problems solved by technology

The disadvantages of an autologous skin graft are possible donor site morbidity and the limitation of available skin amount especially in the case of subjects with extensive burns.
One key barrier is how to manufacture scaffolds that exhibit spatially controlled distributions of cells, growth factors, and scaffold materials and microstructures.
Scaffold-based processes have limitations in addressing these needs due, in part, to restrictions of the manufacturing methods used to synthesize and seed scaffolds.
However, the development of the above is still unsatisfied in commercialization due to their undesirable properties.
Furthermore, where a definition or use of a term in a reference, which is incorporated by reference herein is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

Method used

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  • Composite for Thermo-Sensitive Cell-Tissue Transplanted Scaffold and Use thereof
  • Composite for Thermo-Sensitive Cell-Tissue Transplanted Scaffold and Use thereof
  • Composite for Thermo-Sensitive Cell-Tissue Transplanted Scaffold and Use thereof

Examples

Experimental program
Comparison scheme
Effect test

example 1

Materials and Methods

[0054]Preparation of Easy-Peeling Scaffold

[0055]Polypropylene (PP) non-woven was cut into strips (2×3 cm2and washed in distilled water and the 95% ethanol solution. The N-isopropylacrylamide (NIPAAm) was supplied by Eastman Kodack Co. Ltd USA; the ammonium persulfate (APS) was purchased from Wako Pure Chemical Industry Co., Ltd. The N,N,N′,N′-tetra-methylethylene-diamine (TEMED) was purchased from Fluka and N,N′-methylene-bis-acrylamide (NMBA) from Sigma. Other agents or drugs were all of chemical grade. The PP nonwoven was placed into a plasma treatment system with a rotating substrate holder. Then the graft polymerization of the 10 wt % NIPAAm solution was carried out with a 1000 W UV light irradiated for 30 minutes. The NIPAAm-grafted PP non-woven was chemically crosslinked in gelatin solution by the GA crosslinking agent. The NIPAAm-grafted PP non-woven was placed into a −80° C. refrigerator for three hours and further placed into a freeze-dryer for 1-2 days...

example 2

Easy-Peeling Scaffold Property Evaluation

[0071]As showed in FIG. 1(b), the composite of the trilayer (gelatin / pNIPAAm / PP non-woven) cell-transferred scaffold was demonstrated by scanning electrical microscope. As showed in FIG. 2, the thermo-sensitive response behavior and swelling ratio variation between the pure pNIPAAm hydrogel and the trilayer scaffold showed a similar trend. Furthermore, they all revealed the equivalent lower critical solution temperature (LCST) point at around 32° C. It demonstrated that the pNIPAAm hydrogel exhibited its temperature-sensitive property even if it was grafted onto the surface of different substrates. The trilayer scaffold after the freeze-dried treatment exhibited the higher force (57 Newton) for PP mesh being separated from the scaffold. When immersed this composite in room-temperature water for 2-3 hours to the swelling state, the strip-off force decreased to 1 Newton. However, when it was put in hot water (>34° C.), the force increased to 7 ...

example 3

Bone Marrow Stem Cells (BMSCs) Culture

[0072]As showed in FIG. 3(a), the protocol for isolating the hBMSCs from fresh human BM of five donors was used. hBMSCs usually appeared as spindle-shaped cells with scant cytoplasm and had granules around the nuclei. Within 7 days, the cultures of hBMSCs were nearly confluent and were passaged at 1:3. As showed in FIG. 3(b), the flow cytometry analysis revealed that the hBMSCs were strongly positive for CD13, CD49b, CD105, CD81, SH3, and SH4, but negative for CD45, CD34, and AC133. As showed in FIG. 4, the hBMSC morphology and phenotype remained unchanged for more than 30 cell passages [n=5]. To further monitor the tumor cell proliferation in vitro and in vivo, GFP gene was transduced into hBMSCs using MSCV. As showed in FIG. 5(a), the GFP-positive hBMSCs (hBMSCs-GFP) were then sorted by flow cytometry. The hBMSCs-GFP stably passaged and expressed the markers of mesenchymal stem cells to 30th passage (Table III).

TABLE IIIThe percentage of immun...

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Abstract

A composite comprising a stem cell; a biodegradable layer, which can provide an environment for the stem cell to grow and to differentiate, and; a N-isopropylacrylamide (NIPAAm), which can polymerize with the biodegradable layer and possess the temperature-responsive character for easy stripping. This invention also provides a method of treating a subject with a skin defect by covering the composite of the present invention on the skin defect of the subject in need of such treatment. Furthermore, using this composite with different growth factors, stem cells can be induced to differentiate into skin-related, neuronal cells, neuron, and insulin-positive cells in biodegradable scaffolds as well as transplanted graft. Finally, this invention also provides a quick and convenient method of monitoring cell growth or tissue engineering in an animal.

Description

FIELD OF THE INVENTION[0001]This invention relates to a composite for thermo-sensitive cell-tissue transplanted scaffold and use thereof.DESCRIPTION OF PRIOR ART[0002]Skin is not just a passive barrier for fluid loss and mechanical injury but a complex organ in the human body. Skin injuries and defects may be caused by burn, trauma, cancer, or other diseases. For a small skin defect, primary or secondary healing is typically the simplest and most effective management. Autologous skin graft, however, is the standard treatment for large skin defects such as a major burn or trauma. The disadvantages of an autologous skin graft are possible donor site morbidity and the limitation of available skin amount especially in the case of subjects with extensive burns.[0003]Skin tissue engineering is a possible solution for treating extensive skin defects. The ultimate goal of skin tissue engineering is to restore the complete functions of native skin, but until now the structures and functions ...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): A61K35/12A61K49/00A61P17/02C12Q1/02
CPCA61K35/12A61L27/3834A61L27/50A61F2/105C12N5/0068C12N2533/52C12N2539/10A61L27/60Y10T442/2926Y10T442/2525A61P17/00A61P17/02
Inventor CHIOU, SHIH-HWAPERNG, CHERNG-KANGLIN, HAN-TZOYANG, YI-PING
Owner VETERANS GEN HOSPITAL TAIPEI
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