Method for Preparing Porous Scaffold for Tissue Engineering, Cell Culture and Cell Delivery

Abstract

The present invention relates to a method for preparing a porous scaffold for tissue engineering. It is another object of the present invention to provide a porous scaffold obtainable by the method as above described, and its use for tissue engineering, cell culture and cell delivery. The method of the invention comprises the steps consisting of: a) preparing an alkaline aqueous solution comprising an amount of at least one polysaccharide, an amount of a cross-linking agent and an amount of a porogen agent b) transforming the solution into a hydrogel by placing said solution at a temperature from about 4° C. to about 80° C. for a sufficient time to allow the cross-linking of said amount of polysaccharide and c) submerging said hydrogel into an aqueous solution d) washing the porous scaffold obtained at step c).

1 .- 19 . (canceled) 
     
     
         20 . A porous scaffold obtainable by a method comprising the following steps:
 a) preparing an alkaline aqueous solution comprising an amount of at least one polysaccharide selected from the group consisting of dextran, agar, pullulan, inulin, heparin, fucoidan, and mixtures thereof; an amount of a covalent cross-linking agent selected from the group consisting of trisodium trimetaphosphate (STMP), phosphorus oxychloride (POCl 3 ), epichlorohydrin, formaldehydes, and glutaraldehydes; and an amount of a porogen agent selected from the group consisting of ammonium carbonate, ammonium bicarbonate, calcium carbonate, sodium carbonate, and sodium bicarbonate and mixtures thereof;   b) transforming the solution into a hydrogel by placing said solution at a temperature from about 4° C. to about 80° C. for a sufficient time to allow the cross-linking of said amount of polysaccharide;   c) submerging said hydrogel into an aqueous acidic solution to form pores; and   d) washing the porous scaffold obtained at step c),   wherein   the porous scaffold is a hydrogel;   the porogen agent is dissolved in step a);   the weight ratio of the polysaccharide to the porogen agent is in the range from 6:1 to 1:1; and   wherein the alkaline aqueous solution in a) does not include a surfactant.   
     
     
         21 . The porous scaffold according to  claim 20  wherein the weight ratio of the polysaccharide to the cross-linking agent is in the range from 15:1 to 1:1. 
     
     
         22 . The porous scaffold according to  claim 20  wherein the solution of step a) is poured in a mould before step b). 
     
     
         23 . The porous scaffold according to  claim 20  wherein said scaffold is shaped. 
     
     
         24 . The porous scaffold according to  claim 20  wherein the porogen agent used in step a) is selected from the group consisting of ammonium carbonate, calcium carbonate, sodium carbonate, sodium bicarbonate, and mixtures thereof. 
     
     
         25 . The porous scaffold according to  claim 20  wherein the size of the pores is comprised between 1 μm and 500 μm. 
     
     
         26 . The porous scaffold according to  claim 20  wherein the porosity is in the range from 4% to 50%. 
     
     
         27 . The porous scaffold according to  claim 20  loaded with an amount of cells. 
     
     
         28 . The porous scaffold according to  claim 27  wherein the cells are selected in the group consisting of yeast cells, mammalian cells, insect cells, and plant cells. 
     
     
         29 . The porous scaffold according to  claim 28  wherein mammalian cells are selected from the group consisting of chondrocytes; fibrochondrocytes; osteocytes; osteoblasts; osteoclasts; synoviocytes; bone marrow cells; epithelial cells, hepatocytes, mesenchymal cells; stromal cells; muscle cells, stem cells; embryonic stem cells; precursor cells derived from adipose tissue; peripheral blood progenitor cells; stem cells isolated from adult tissue; and genetically transformed cells. 
     
     
         30 . The porous scaffold according to  claim 20  for tissue engineering, cell culture and cell delivery. 
     
     
         31 . A vascular substitute made with a scaffold of  claim 20 . 
     
     
         32 . Cartilage or bone implants made with a scaffold of  claim 20 . 
     
     
         33 . A method for evaluating the toxicity and/or pharmacology of a product comprising the use of a scaffold of  claim 20 . 
     
     
         34 . A controlled release system of an active agent made with a scaffold of  claim 20 .