TWI555841B - Carbonized materials and methods of promoting cell differentiation using carbonized materials - Google Patents

Description

Carbonized material and method for promoting cell differentiation by using carbonized material

The present invention relates to cell differentiation; in particular, to a carbonized material and a method for promoting cell differentiation using a carbonized material.

In the modern advancement of neuroscience medical research, a variety of causes of cranial neuropathy have been discovered. In the case of Parkinson's disease, the cause is due to damage or death of neurons in the human central nervous system that should release neurotransmitter-dopamine. Dopamine is indispensable for the regulation of motor function in the basal ganglia of the brain. Therefore, the reduction in the number of dopamine will impair human motor coordination.

In the early stage of rickets, dopamine can be used to make up for the lack of dopamine in patients, and it can effectively control the problem of impaired motor function. However, drug treatment cannot control the symptoms of serious patients, and even only causes side effects. At this time, surgical intervention is required. Among them, cell replacement therapy replaces damaged cells by transplanting normal cells, or provides a bridge for the regeneration of damaged nerve cells, and completes or regenerates the conduction pathway of the nervous system to restore nerve conduction function. .

The normal neural cell line that is often obtained clinically is differentiated from human neural stem cells (hNSC). In the past, human neural stem cell lines were cultured on flat culture dishes with a differentiation rate of only about 2%; even after the addition of differentiation factor (DF), the differentiation rate was only increased to about 18%, differentiation. Efficiency is not high. Therefore, although cell replacement therapy avoids the side effects of drug treatment, the efficiency of cell differentiation does need to be improved.

In view of the above, an object of the present invention is to provide a carbonized material and a method for promoting cell differentiation by using a carbonized material, which can simulate the environment in a living body by the three-dimensional stacked structure of the carbonized material, that is, form a stereo scaffold to promote cell differentiation. .

In order to achieve the above object, the method for promoting cell differentiation by using a carbonized material according to the present invention comprises the following steps: A, providing a fiber woven fabric, wherein the fiber woven fabric comprises a polyacrylonitrile fiber; B, high temperature carbonizing the fiber woven fabric Clothing to produce a carbonized material; wherein the fiber woven fabric is carbonized at a pressure greater than 100 Torr and less than one atmosphere and at a temperature between 500 and 1200 ° C; the high temperature carbonization time is between 1 and 25 minutes, and An inert gas is supplied to the carbonization environment during the carbonization process; the carbonization material comprises a plurality of carbon fibers, wherein each of the carbon fibers comprises a body and a plurality of carbon columns, the carbon columns are formed on the surface of the body, and the carbon columns are Various directions are stacked on each other; and C, a cell is placed on the carbonized material, wherein the cell has the ability to differentiate and can differentiate into a differentiated cell having a specific function; thereby, the cell is attached to the differentiation process during differentiation A carbon column is used to promote its differentiation function, accelerating the differentiation of the cell into the differentiated cells having this specific function.

The present invention further provides a carbonized material as a substrate for promoting cell differentiation, which is produced by the following steps: A, providing a fiber woven fabric, wherein the fiber woven fabric comprises a polyacrylonitrile fiber; B, high temperature carbonizing the fiber Woven to produce a carbonized material; wherein the fiber woven fabric is carbonized at a pressure greater than 100 Torr and less than one atmosphere and at a temperature between 500 and 1200 ° C; the carbonization time is between 1 and 25 minutes. And supplying an inert gas to the carbonization environment during the carbonization process; wherein the carbonization material comprises a plurality of carbon fibers, and each of the carbon fibers comprises a body and a plurality of carbon columns, and the carbon columns are formed on the surface of the body, and the The carbon columns are stacked on each other in various directions.

The effect of the present invention is that the carbonized material formed by the carbonization of the fiber woven fabric provides a new medical use and can effectively improve the efficiency of cell differentiation.

100‧‧‧Carbide

10‧‧‧Carbon fiber

12‧‧‧Ontology

14‧‧‧ carbon column

20‧‧‧ cells

22‧‧‧ axons

24‧‧‧Dopamine

D1‧‧‧ Height

D2‧‧‧ diameter

Θ‧‧‧contact angle

1 is a flow chart of a method for promoting cell differentiation using a carbonized material in accordance with a preferred embodiment of the present invention.

2 is a perspective view of a carbonized material of the above preferred embodiment of the present invention, showing the imaging of the carbonized material under an electron microscope.

Figure 3 is a perspective view of the carbonized material of the above preferred embodiment of the present invention, revealing the imaging of the carbon column under an electron microscope.

Figure 4 is a schematic illustration of the carbonized material of the above preferred embodiment of the present invention.

Fig. 5 is a perspective view showing the differentiation of neural stem cells attached to a conventional cell culture medium.

Fig. 6 is a schematic view showing the carbonized material of the above preferred embodiment of the present invention, revealing that the three-dimensional structure of the carbon column promotes differentiation of neural stem cells.

In order to explain the present invention more clearly, the preferred embodiment will be described in detail with reference to the accompanying drawings. Referring to FIG. 1 , a method for promoting cell differentiation using a carbonized material 100 is illustrated in a preferred embodiment of the present invention. flow chart.

The carbonized material 100 is a carbon fiber fabric. The carbonization method and the carbonization device used by the applicant have been disclosed in the Chinese Patent Publication No. I445661. The process is roughly as follows: First, a fiber woven fabric is provided, wherein The fiber woven fabric comprises a polyacrylonitrile fiber; then, the fiber woven fabric is carbonized at a high temperature to produce the carbonized material 100. Wherein the fiber woven fabric is carbonized under a pressure of more than 100 Torr and less than one atmosphere, and the temperature is between 500 and 1200 ° C; the time of high temperature carbonization is between 1 and 25 minute. In addition, an inert gas (such as nitrogen) is supplied to the carbonization environment during the carbonization process to prevent the fiber woven fabric from coming into contact with oxygen in the air to generate an oxidation reaction. Other detailed procedures are not mentioned here.

As shown in FIG. 2, the carbonized material 100 includes a plurality of cross-hatched carbon fiber bundles, and each of the carbon fiber bundles is assembled from a plurality of linear carbon fibers 10. It is characterized in that, when the carbon fibers 10 are observed at a high magnification by an electron microscope, it can be observed that each of the carbon fibers 10 includes a plurality of carbon columns 14 in addition to a linear body 12; the carbon columns 14 are located therein. The body 12 is surfaced and stacked in various directions (see Figure 3 for reference).

Please refer to FIG. 4, in order to more clearly define the size of the carbon columns 14, and the relative relationship of the carbon columns 14 to the cells, the carbon columns 14 in the figure are represented by a matrix arrangement of axial vertical surfaces. The height D1 of each of the carbon columns 14 is about 0.001 to 5 mm; and the diameter D2 of each of the carbon columns 14 is about 1 to 4 μm.

Finally, a cell 20 having differentiation ability is placed on the carbonized material 100. In this embodiment, the cell 20 is a neural stem cell, but is not limited by a neural stem cell, but may also be a neuron-like cell. Differentiate into a precursor cell of a nerve cell. The specific function of the nerve cells grows at least one axon and releases dopamine through the axon to transfer information from the cell to other cells.

As shown in Fig. 5, neural stem cells have been cultured on a flat cell culture substrate 1 for differentiation in the past, but the differentiation rate is low. In other words, the cell 20 develops a slower rate of axonal 22 and releases dopamine 24 to deliver a lower level of function. In addition, since the cell 20 is attached to the substrate 1 as a whole, the contact angle θ formed by the surface of the substrate 1 to which the cell 20 is attached and the highest point of the cell 20 is small, which is not conducive to cell growth and movement. Differentiation.

Referring to FIG. 6, the carbon column 14 of the carbonized material 100 of the present embodiment is attached to the cell 20 during the differentiation process, and the axon of the cell 20 can be favored. The formation of 22 simultaneously promotes the release of dopamine 24 at the ends of axons 22. Further, the carbonized material 100 of the present invention has an electrically conductive property. In detail, the carbon atoms in the carbonized material 100 are bonded to each other by the sp ² mixed orbital region, and thus have characteristics of easily attracting electrons. Since the dopamine 24 releases electrons, the axon 22 releasing the dopamine 24 is slightly attracted to the carbon column 14 and can be firmly attached to the carbon column 14. The cells 20 are attached to the carbon columns 14 by the axons 22, so that the cells not only attach to the surface of the body 12, but also increase the contact angle θ, which is favorable for cell growth, movement and differentiation.

In addition, the cell 20 has a higher concentration of tyrosine hydroxylase in the cell 20 in addition to a change in morphology and release of dopamine during differentiation. Tyrosine hydroxylase is an important enzyme for converting tyrosine in cell 20 into dopamine 24. Therefore, the concentration and activity of tyrosine hydroxylase in cells can be used as an indicator of neural stem cell differentiation. The cells 20 adhering to the carbonized material 100 for differentiation have higher tyrosine hydroxylase concentration and activity than the cells 20 attached to the matrix 1. It can be seen that the carbonized material 100 does promote the type change of the cells 20 and the intracellular signal transmission associated with differentiation, and facilitates the differentiation of the cells 20.

In summary, the present invention utilizes a carbonized material as a matrix for promoting cell differentiation, and utilizes the three-dimensional structure and conductive properties of the carbonized material 100 to promote growth and differentiation of the cell 20 thereon.

The carbonized material 100 of the present embodiment can be further applied to a culture dish, wherein the culture dish has a bottom plate and an inner space, and the bottom plate has an upper surface facing the inner space. The carbonized material 100 is bonded to the upper surface, so that when the cells 20 are placed on the upper surface of the bottom plate of the culture dish, they can be attached to the carbonized material 100 to grow, thereby promoting differentiation of the cells 20.

The inclusion of the carbonized material 100 described above can also be used to reconstruct a nerve defect caused by a disease or injury. There are stem cells required for tissue regeneration in the tissue and blood at the wound. Therefore, if nerve defects are to be repaired, it is necessary to construct a biological scaffold at the nerve defect for cell attachment, proliferation and differentiation. However, the use of conventional biological scaffolds can only support a certain degree of tissue regeneration, and the bioscaffold itself does not have the ability to induce tissue regeneration and differentiation. If the carbonized material 100 of the present invention is used as a biological stent built on a nerve defect, the growth and differentiation of stem cells at the wound can be accelerated, and a better nerve reconstruction effect can be achieved.

The above is only a preferred embodiment of the present invention, and equivalent changes to the scope of the present invention and the scope of the patent application are intended to be included in the scope of the present invention.

Claims (5)

A method for promoting cell differentiation by using a carbonized material, comprising the steps of: A, providing a fiber woven fabric, wherein the fiber woven fabric comprises a polyacrylonitrile fiber; B, carbonizing the fiber woven fabric at a high temperature to produce a carbonized material; The fiber woven fabric is carbonized under a pressure of more than 100 Torr and less than one atmosphere, and the temperature is between 500 and 1200 ° C; the carbonization time is between 1 and 25 minutes, and an inert gas is supplied during the carbonization process. a carbonization environment; the carbonized material comprises a plurality of carbon fibers, wherein each of the carbon fibers comprises a body and a plurality of carbon columns formed on the surface of the body and stacked in various directions; and C, placing a cell The carbonized material, wherein the cell has a differentiation ability, can be differentiated into a differentiated cell having a specific function; thereby, the cell is adhered to the carbon column during differentiation to promote its differentiation function and accelerate the differentiation of the cell. It is the differentiated cell having this specific function. A method for promoting cell differentiation using a carbonized material as described in claim 1, wherein each of the carbon columns has a diameter of about 1 to 4 μm. A method for promoting cell differentiation using a carbonized material as described in claim 1, wherein each of the carbon columns has a height of about 0.001 to 5 mm. A method for promoting cell differentiation using a carbonized material according to claim 1, wherein the cell of the step C comprises a neural stem cell. A method for promoting cell differentiation using a carbonized material according to claim 1, wherein the differentiated cell of step C comprises a nerve cell, wherein the specific function of the nerve cell comprises releasing dopamine.