Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Solid support and method of culturing cells on said solid...
Reexamination Certificate
2001-03-16
2003-11-04
Naff, David M. (Department: 1651)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Solid support and method of culturing cells on said solid...
C424S093700, C435S178000, C435S325000, C435S397000
Reexamination Certificate
active
06642050
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a three-dimensional cell culture material made into a three-dimensional shape whereby proliferation, morphology and function of cells can be expressed in the same manner as in the cells in vivo and also to a method for the incubation of cells using the cell culture material.
2. Description of the Related Art
Progress in glyco-engineering in recent years is remarkable. For example, proteoglycan, glycolipid, glycoprotein, etc. in cell walls of vegetable cells are exemplified as biopolymers having sugar chains. Each of them is believed to participate in (1) stabilization of cells, (2) differentiation, proliferation, adhesion and migration of cells and (3) interaction among the cells and cell recognition and various reports have been available. In addition, a mechanism where the sugar chains of those polymers act for another, assist, amplify, adjust or inhibit their functions of each other whereby they control high and precise bioreactions has been gradually clarified.
Further, such sugar chains differentiate and proliferate the cells and participate in adhesion of cells and their relation with immune and malignant alteration of cells has been made clear. Thus, development of various industries can be expected by attempting a new development by a close connection of the glyco-engineering with medicine, cell engineering or tissue engineering.
An example is that there have been brisk studies for onset of diseases due to abnormal interaction among sugar chains on cell surfaces and among sugar chain receptors or for role of sugar chain in viral infection such as AIDS (Takehiro Kawano,
Bio Industry
, 14, 22-30(1997)). In addition, studies on protein which recognizes the sugar chain such as secretin, contactin and contactinhibin as a molecule for mediating the adhesion between cells have become important in understanding the bioreactions (Tuneshige Takeuchi and Naoki Takahashi,
Saibo Kogaku
, 16, 801-812(1997)).
The present inventors already carried out an intensive investigation paying their attention to a cell-specific interaction of sugar chain and, as a model for ligand against asialoglycoprotein receptors, they designed and synthesized poly(N-p-vinylbenzyl-[O-&bgr;-D-galactopyranosyl-(1→4)-D-gluconamide]) (abbreviated as PVLA) which was a polystyrene having galactose in a side chain. For example, in a hepatocyte incubation experiment on a culture dish coated with this PVLA as a cell culture material, it was found that parenchymal hepatocytes are selectively incubated via a specific affinity of PVLA with asialoglycoprotein receptor on the surface of parenchymal hepatocytes and that, even under a two-dimensional culture material environment, parenchymal hepatocytes per se were made into a three-dimensional shape and specifically existed as spheroids (A. Kobayashi, M. Goto, K. Kobayashi, T. Akaike, J. Biomater.
Sci. Polymer Edn
., 6, 325-342 (1994)). However, this PVLA has a structure having polystyrene which is a synthetic polymer as a main chain and, therefore, it has disadvantages that no biodegradability is available and there are possibilities of expression of antigenicity and of having a toxicity.
In the meanwhile, incubation of cells using a culture dish coated with an adhesion protein such as collagen, fibronectin or laminin which is a natural protein has been also carried out. However, that is a two-dimensional culture and, therefore, proliferation of cells, expression of cell functions, etc. are not sufficiently achieved.
Recently, there has been an attempt where natural or synthetic polymer such as collagen, alginic acid, hyaluronic acid and polyisopropylacrylamide is cross-linked using ultraviolet ray, radioactive ray or chemical cross-linking agent to give a culture material in a three-dimensional form and cells are incubated using the same. However, there are problems that cell adhesion is not good whereby proliferation of the cells is often suppressed, cell function lowers, cell form is extended unlike that in vivo, etc. and no system for a good incubation of cells has been available.
SUMMARY OF THE INVENTION
An object of the present invention is therefore to provide a three-dimensional cell culture material made into a three-dimensional form whereby adhesion and proliferation of cells are promoted, cell functions are maintained and improved and the cell form can be retained in a form near that in vivo, and also to provide a method for the incubation of cells using the three-dimensional cell culture material.
Thus, the present invention is a three-dimensional cell culture material, characterized in that a cell culture material comprising a sugar containing polymer in which at least one kind of sugar chain is bonded as a side chain via a spacer molecule is made into a three-dimensional shape.
Further, the present invention is a method for the incubation of cells, characterized in that, cells are incubated using the three-dimensional cell culture material whereby proliferation, morphology and function of the cells are maintained and improved.
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Akaike Toshihiro
Goto Mitsuaki
Yang Jun
Amcite Research, Ltd.
Naff David M.
Wenderoth , Lind & Ponack, L.L.P.
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