Application of temperature-sensitive cellulose derivative as nervous tissue repair and regeneration material
By preparing the temperature-sensing cellulose derivative with hydrophilic and hydrophobic carbonate groups, the problem of poor repair of peripheral nerve damage in the prior art is solved, and a solid support structure is formed in the body and nerve cell regeneration is achieved. Effect.
Patent Information
- Application Number
- CN202411100967.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-08-12
- Publication Date
- 2024-12-10
AI Technical Summary
The prior art has poor effect on repairing peripheral nerve damage, especially large, long-term nerve defects and multiple nerve damage, and the matrix material cannot effectively induce the regeneration of host tissue in tissue engineering.
Using temperature-sensitive cellulose derivatives, the esterification reaction between hydroxypropyl cellulose and carbonate under a catalyst is carried out to prepare cellulose derivatives with hydrophilic and hydrophobic carbonate groups. This material can form solid support in the body. structure and are slowly degraded to achieve slow release of effective substances, promoting nerve cell attachment and regeneration.
This cellulose derivative has excellent nerve cell adhesion in the body, can regulate the formation of solid support structures near 37°C, promote the regeneration and repair of neural tissue, and achieve the slow release of effective substances through slow degradation, providing neural tissue engineering Effective application of brackets.
Smart Images
Figure CN119097763A_ABST
Abstract
Description
Technical Field
[0001] The invention relates to the technical field of nerve damage repair, and in particular to application of a temperature-sensitive cellulose derivative as a nerve tissue repair and regeneration material. Background Art
[0002] Neurons are neuronal cells, which are the most basic structural and functional units of the nervous system. They are divided into two parts: the cell body and the processes. The cell body is composed of the nucleus, cell membrane, and cytoplasm, and has the function of connecting and integrating input information and transmitting information. There are two types of processes: dendrites and axons. Dendrites are short and have many branches. They extend and protrude directly from the cell body to form branches. Their function is to receive impulses from the axons of other neurons and transmit them to the cell body. Axons are long and have few branches. They are slender processes with uniform thickness. They often originate from the axon hillock. Their function is to receive external stimuli and then transmit them from the cell body. In addition to branching out side branches, the ends of axons form branch-like nerve endings. The endings are distributed in certain tissues and organs, forming various nerve ending devices. Sensory nerve endings form various receptors; motor nerve endings are distributed in skeletal muscles to form motor end plates.
[0003] At present, the clinical repair of peripheral nerve injury is still not ideal, and there is no effective treatment for thick, long-segment nerve defects and multiple nerve injuries. With the development of tissue engineering and neuroscience, the application of tissue engineering principles to prepare nerve tissue grafts to replace autologous nerves to bridge long segments of damaged nerves has achieved better therapeutic effects. Therefore, nerve tissue engineering grafts have received more and more attention, and it is expected that tissue engineering will solve the problem of nerve tissue repair. In recent years, people have gradually realized that matrix materials play an important role in tissue engineering. With the in-depth study of the interaction mechanism between materials and organisms, matrix materials are required to induce favorable responses of the host, such as inducing the regeneration of host tissues.
[0004] In view of this, the present invention is proposed. Summary of the invention
[0005] The purpose of the present invention is to provide a thermosensitive cellulose derivative for use as a nerve tissue repair and regeneration material. The thermosensitive cellulose derivative has hydrophilic and hydrophobic carbonate groups, has excellent adhesion to nerve cells and can promote nerve regeneration. In addition, the LCST of the cellulose derivative can be regulated at around 37°C, can form a solid state in the body, has a certain supporting effect, and can be slowly degraded by the body to achieve slow release of effective substances.
[0006] In order to achieve the above-mentioned purpose of the present invention, the following technical solutions are particularly adopted:
[0007] The first aspect of the present invention provides an application of a temperature-sensitive cellulose derivative as a nerve tissue repair and regeneration material, characterized in that the preparation method of the temperature-sensitive cellulose derivative comprises the following steps:
[0008] (a) adding hydroxypropyl cellulose and carbonate into a solvent in the presence of a catalyst to carry out an esterification reaction;
[0009] (b) adding an extractant to the reaction product after the esterification reaction is completed to extract the product, and collecting the extract layer;
[0010] (c) subjecting the extraction layer to rotary evaporation to remove the solvent, and then subjecting the extraction layer to vacuum drying to obtain the thermosensitive cellulose derivative.
[0011] Preferably, in step (a), the mass ratio of hydroxypropyl cellulose to carbonate is 1:(0.01-10).
[0012] Preferably, in step (a), the carbonate is an alkyl chloroformate or an aromatic chloroformate;
[0013] The alkyl chloroformate includes methyl chloroformate, ethyl chloroformate and tert-butyl chloroformate.
[0014] Preferably, in step (a), the catalyst is selected from at least one of triethylamine, tributylamine, pyridine, 4-diaminopyridine, N,N-dimethylaniline and N,N-diisopropylethylamine.
[0015] Preferably, in step (a), the esterification reaction temperature is 0 to 100° C., and the reaction time is 1 to 24 hours.
[0016] Preferably, in step (a), the solvent is selected from at least one of cyclohexanone, isopropanol, dichloromethane, chloroform and THF.
[0017] Preferably, in step (b), the extractant is selected from at least one of toluene, ether, methanol and ethanol.
[0018] Preferably, the neural tissue repair and regeneration material is in the form of an injectable / coatable temperature-sensitive hydrogel, neural tissue engineering scaffold or microsphere. Compared with the prior art, the beneficial effects of the present invention include at least:
[0019] The present invention successfully prepares carbonated hydroxypropyl cellulose by utilizing a certain proportion of hydroxyl molecules on the main chain of hydroxypropyl cellulose for a carbonation reaction; the cellulose derivative has excellent adhesion to nerve cells and can promote nerve tissue regeneration due to the introduction of carbonate groups that are both hydrophilic and hydrophobic. In addition, the LCST of the cellulose derivative of the present invention can be regulated at around 37°C, can form a solid state in the body, has a certain supporting effect, and can be slowly degraded by the body to achieve the slow release of effective substances. Therefore, the temperature-sensitive cellulose derivative of the present application can be used as a scaffold material for nerve tissue engineering and is beneficial to promoting the regeneration and repair of nerve tissue. BRIEF DESCRIPTION OF THE DRAWINGS
[0020] In order to more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the following is a brief introduction to the drawings required for the specific embodiments or the description of the prior art. In all the drawings, similar elements or parts are generally identified by similar reference numerals. In the drawings, the elements or parts are not necessarily drawn according to the actual scale.
[0021] Figure 1 This is a microscope photo of mouse hippocampal neuronal cells after 21 days of culture. DETAILED DESCRIPTION
[0022] The following embodiments of the technical solution of the present invention are described in detail in conjunction with the embodiments. The following embodiments are only used to more clearly illustrate the technical solution of the present invention, and are therefore only used as examples, and cannot be used to limit the protection scope of the present invention.
[0023] It should be noted that, unless otherwise specified, the technical terms or scientific terms used in this application should have the common meanings understood by those skilled in the art to which the present invention belongs.
[0024] The embodiment of the present invention provides an application of a temperature-sensitive cellulose derivative as a nerve tissue repair and regeneration material. The preparation method of the temperature-sensitive cellulose derivative comprises the following steps:
[0025] (a) adding hydroxypropyl cellulose and carbonate into a solvent in the presence of a catalyst to carry out an esterification reaction;
[0026] (b) adding an extractant to the reaction product after the esterification reaction is completed to extract the product, and collecting the extract layer;
[0027] (c) subjecting the extraction layer to rotary evaporation to remove the solvent, and then subjecting the extraction layer to vacuum drying to obtain the thermosensitive cellulose derivative.
[0028] The present invention successfully prepares carbonated hydroxypropyl cellulose by utilizing a certain proportion of hydroxyl molecules on the main chain of hydroxypropyl cellulose for a carbonation reaction; the cellulose derivative has excellent adhesion to nerve cells and can promote nerve regeneration due to the introduction of carbonate groups that are both hydrophilic and hydrophobic. In addition, the LCST of the cellulose derivative of the present invention can be regulated at around 37°C, can form a solid state in the body, has a certain supporting effect, and can be slowly degraded by the body to achieve the slow release of effective substances. Therefore, the temperature-sensitive cellulose derivative of the present application can be used as a scaffold material for neural tissue engineering and is beneficial to promoting the regeneration and repair of neural tissue.
[0029] In one embodiment, in the step (a), the mass ratio of hydroxypropyl cellulose to carbonate is 1:(0.01-10).
[0030] In one embodiment, in step (a), the carbonate is an alkyl chloroformate or an aromatic chloroformate;
[0031] The alkyl chloroformate includes methyl chloroformate, ethyl chloroformate and tert-butyl chloroformate.
[0032] In one embodiment, in step (a), the catalyst is selected from at least one of triethylamine, tributylamine, pyridine, 4-diaminopyridine, N,N-dimethylaniline and N,N-diisopropylethylamine.
[0033] In one embodiment, in the step (a), the esterification reaction temperature is 0 to 100° C., and the reaction time is 1 to 24 hours.
[0034] In one embodiment, in step (a), the solvent is selected from at least one of cyclohexanone, isopropanol, dichloromethane, chloroform and THF.
[0035] In one embodiment, in step (b), the extractant is selected from at least one of toluene, diethyl ether, methanol and ethanol.
[0036] In one embodiment, the neural tissue repair and regeneration material is in the form of an injectable / coatable thermosensitive hydrogel, a neural tissue engineering scaffold or a microsphere.
[0037] The technical solution of the present invention is further described in detail below through specific embodiments.
[0038] Example 1
[0039] This embodiment is a method for preparing a temperature-sensitive cellulose derivative, and the method for preparing the temperature-sensitive cellulose derivative comprises the following steps:
[0040] (a) 10 g of hydroxypropyl cellulose was dissolved in cyclohexanone (500 ml), and then 9.5 g of triethylamine and 5 g of methyl chloroformate were added and reacted at room temperature for 5 h;
[0041] (b) after the esterification reaction is completed, 200 ml of toluene is added to the reaction product for extraction, and the extraction layer is collected;
[0042] (c) The extraction layer is subjected to rotary evaporation to remove the solvent, and then vacuum dried to obtain a thermosensitive cellulose derivative.
[0043] Example 2
[0044] This embodiment is a method for preparing a temperature-sensitive cellulose derivative, and the method for preparing the temperature-sensitive cellulose derivative comprises the following steps:
[0045] (a) 10 g of hydroxypropyl cellulose was dissolved in cyclohexanone (500 ml), and then 9.5 g of triethylamine and 5 g of ethyl chloroformate were added and reacted at room temperature for 5 h;
[0046] (b) after the esterification reaction is completed, 200 ml of toluene is added to the reaction product for extraction, and the extraction layer is collected;
[0047] (c) The extraction layer is subjected to rotary evaporation to remove the solvent, and then vacuum dried to obtain a thermosensitive cellulose derivative.
[0048] Example 3
[0049] This embodiment is a method for preparing a temperature-sensitive cellulose derivative, and the method for preparing the temperature-sensitive cellulose derivative comprises the following steps:
[0050] (a) 10 g of hydroxypropyl cellulose was dissolved in cyclohexanone (500 ml), and then 9.5 g of triethylamine and 5 g of tert-butyl chloroformate were added and reacted at room temperature for 5 h;
[0051] (b) after the esterification reaction is completed, 200 ml of toluene is added to the reaction product for extraction, and the extraction layer is collected;
[0052] (c) The extraction layer is subjected to rotary evaporation to remove the solvent, and then vacuum dried to obtain a thermosensitive cellulose derivative.
[0053] Example 4
[0054] This embodiment is a neural tissue engineering scaffold, and the neural tissue engineering scaffold is prepared by the following method:
[0055] The thermosensitive cellulose derivative of Example 1 was dissolved in 4° C. pure water to prepare a 5% aqueous solution, and then the aqueous solution was heated to 37° C. to form a gel, thereby obtaining the scaffold material for nerve tissue repair and regeneration engineering.
[0056] The above gel material can be used for the regeneration and culture of mouse hippocampal neuronal cell tissue. Under a sterilized environment, the 18th day embryo is taken out from the abdomen of the pregnant mouse, quickly placed in a pre-cooled phosphate buffered saline (PBS), and then the hippocampal tissue is separated on ice and placed in a pre-cooled PBS. After all the samples are taken, the supernatant is discarded and washed once with PBS. Then each hippocampal tissue is placed in a neuronal cell culture containing DNase and Trypsin, and placed in a 37°C incubator for digestion for 15-30 minutes. During this period, a 1ml pipette is used to aspirate / discharge the liquid several times to disperse the cells. After the time is up, put it in a centrifuge, centrifuge at 7000rpm for 1 minute, then discard the supernatant, add 1ml of neuronal cell culture solution and mix evenly, count with a cell counter, sow it in a culture vessel coated with a thermosensitive cellulose derivative gel, and then place it in a 37°C incubator for 21 days. After culturing, the neuronal cells grow more mature, the protrusions between neurons are more closely connected, and a dense network of neurons can be observed under a microscope ( Figure 1 ).
[0057] Example 5
[0058] This embodiment is a neural tissue engineering scaffold, and the neural tissue engineering scaffold is prepared by the following method:
[0059] The temperature-sensitive cellulose derivative of Example 2 is heated to a molten state (250° C.), and then poured into a mold for molding to obtain the neural tissue engineering scaffold.
[0060] Example 6
[0061] This embodiment is a neural tissue engineering scaffold, and the neural tissue engineering scaffold is prepared by the following method:
[0062] The temperature-sensitive cellulose derivative of Example 3 is heated to a molten state (250° C.), and then poured into a mold for molding to obtain the neural tissue engineering scaffold.
[0063] Experimental example
[0064] Thermosensitive cellulose derivatives were prepared according to Examples 1 to 3 respectively;
[0065] The yield, degree of carbonation and LCST of the temperature-sensitive cellulose derivative prepared in step (c) of each embodiment were tested, and the test results are shown in Table 1;
[0066] Table 1
[0067]
[0068] From Table 1, we can see that:
[0069] The LCST of the thermosensitive cellulose derivative prepared in the examples of the present application can be controlled at around 37° C. and can be used for nerve tissue repair and regeneration.
[0070] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein by equivalents. These modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention, and they should all be included in the scope of the claims and specification of the present invention.
Claims
1. Application of thermosensitive cellulose derivatives as nerve tissue repair and regeneration materials, characterized in that: The preparation method of the temperature-sensitive cellulose derivative comprises the following steps: (a) adding hydroxypropyl cellulose and carbonate into a solvent in the presence of a catalyst to carry out an esterification reaction; (b) adding an extractant to the reaction product after the esterification reaction is completed to extract the product, and collecting the extract layer; (c) subjecting the extraction layer to rotary evaporation to remove the solvent, and then subjecting the extraction layer to vacuum drying to obtain the thermosensitive cellulose derivative.
2. The use according to claim 1, characterized in that: In the step (a), the mass ratio of hydroxypropyl cellulose to carbonate is 1:(0.01-10).
3. The use according to claim 1, characterized in that: In the step (a), the carbonate is an alkyl chloroformate or an aromatic chloroformate; The alkyl chloroformate includes methyl chloroformate, ethyl chloroformate and tert-butyl chloroformate.
4. The use according to claim 1, characterized in that: In the step (a), the catalyst is selected from at least one of triethylamine, tributylamine, pyridine, 4-diaminopyridine, N,N-dimethylaniline and N,N-diisopropylethylamine.
5. The use according to claim 1, characterized in that: In the step (a), the esterification reaction temperature is 0 to 100° C., and the reaction time is 1 to 24 hours.
6. The use according to claim 1, characterized in that: In the step (a), the solvent is selected from at least one of cyclohexanone, isopropanol, dichloromethane, chloroform and THF.
7. The use according to claim 1, characterized in that: In the step (b), the extractant is selected from at least one of toluene, ether, methanol and ethanol.
8. The use according to claim 1, characterized in that: The nerve tissue repair and regeneration material is in the form of an injectable / coatable temperature-sensitive hydrogel, a nerve tissue engineering scaffold or a microsphere.