A periodontal regeneration material with the functions of regulating periodontal coagulation immune microenvironment and releasing trace elements for promoting periodontal regeneration

Abstract

The application discloses a periodontal regeneration material with the functions of regulating a periodontal coagulation immune microenvironment and releasing trace elements for promoting periodontal regeneration, and belongs to the technical field of medical materials. The periodontal regeneration material prepared by the preparation method of the periodontal regeneration material has the functions of loading trace elements for promoting periodontal regeneration and regulating a fibrin assembly process in a periodontal coagulation process, and can construct a mesoporous silica with an ideal periodontal blood coagulation microstructure. The fibrin network structure of the blood coagulation produced after the action can reduce an inflammatory reaction, induce a regenerative immune microenvironment, and reduce periodontal tissue regeneration failure caused by fiber wrapping. The periodontal regeneration material can promote periodontal multi-tissue collaborative regeneration, so as to regenerate a functional structure of a cementum-periodontal ligament-alveolar bone functional complex. Meanwhile, the periodontal regeneration material provided by the application is an acidic compound, and can be slowly degraded under an alkaline condition in a body, so as to achieve the purpose of releasing trace elements.

Description

Technical Field

[0001] This invention belongs to the field of medical materials technology, and in particular relates to a periodontal regeneration material that combines the functions of regulating the periodontal coagulation immune microenvironment and slow-release trace elements that promote periodontal regeneration. Background Technology

[0002] The periodontal tissue structure is complex, containing cementum, periodontal ligament, alveolar bone, and gingival connective tissue, among other composite structures. These tissues are closely interconnected, forming the "cementum-periodontal ligament-alveolar bone functional complex," which collectively performs oral functions. Periodontal tissue regeneration involves the regeneration of soft and hard tissues from different sources, as well as the functional integration between these tissues. Therefore, periodontal regeneration is a distinct field from bone regeneration, and its difficulty far exceeds that of simple bone regeneration. However, currently, there are no materials specifically designed for periodontal tissue regeneration in clinical practice. Often, bone regeneration materials are directly applied to periodontal tissues, frequently leading to clinical problems such as epithelial integration and fibrous encapsulation, resulting in a lack of breakthroughs in periodontal treatment in clinical dentistry. Therefore, it is essential to develop biomaterials specifically designed for periodontal tissue regeneration, addressing its physiological characteristics, to achieve complex periodontal tissue regeneration and overcome current bottlenecks in periodontal tissue regeneration. This is of significant strategic importance and a consensus within the current dental field. Summary of the Invention

[0003] The purpose of this invention is to overcome the shortcomings of the prior art and provide a periodontal regeneration material, its preparation method, and its application.

[0004] To achieve the above objectives, in a first aspect, the present invention provides a periodontal regeneration material, wherein the periodontal regeneration material is a mesoporous silica microsphere having a regulatory periodontal coagulation immune microenvironment, and the mesoporous silica microsphere is loaded with trace elements that promote periodontal regeneration; the trace elements include at least one selected from copper, strontium, magnesium, lithium, iron, zinc, manganese, selenium, molybdenum, and cobalt.

[0005] Blood clotting is a crucial initiating factor in periodontal regeneration. Periodontal blood clots, by creating a favorable local immune microenvironment, mediate the migration, proliferation, and differentiation of stem cells derived from periodontal soft and hard tissues, effectively inducing the regeneration of periodontal complex tissues. An ideal periodontal blood clot is characterized by fine fiber diameter, numerous branching points, high density, and low porosity. Previous studies have confirmed that controlling the fiber diameter of the periodontal blood clot to 0.1-0.15 μm and the porosity to 20-25%, with a multi-branched clot, can effectively promote periodontal tissue regeneration. This structure reduces the contact area of ​​immune cells, lowers macrophage inflammation levels, promotes the formation of a regenerative and repairing immune microenvironment, and prevents periodontal tissue regeneration failure caused by excessive inflammatory responses, such as fiber encapsulation and epithelial integration. Furthermore, the fibrin network within the blood clot acts as a scaffold structure, guiding the migration of osteoblasts and periodontal ligament fibroblasts from the alveolar bone and periodontal ligament, promoting early in situ tissue regeneration. Meanwhile, the blood clot exhibits good stability, which helps prevent premature ingrowth of cells from epithelial tissue, thus preventing periodontal complex regeneration failure. Therefore, high-performance periodontal regeneration materials need to form a good periodontal blood clot microstructure, create a regenerative immune microenvironment, avoid periodontal tissue regeneration failure caused by fibrous encapsulation, and promote periodontal tissue regeneration.

[0006] However, while creating a favorable periodontal coagulation and immune microenvironment by regulating blood clot parameters can prevent periodontal regeneration failure caused by excessive fibrosis, it is still difficult to completely rebuild the complex structure of cementum-periodontal ligament-alveolar bone. Therefore, it is necessary to further introduce trace elements that promote the synergistic regeneration of multiple periodontal tissues to synergistically promote the functional integration of periodontal soft and hard tissues and achieve functional regeneration of periodontal tissues. Previous studies have shown that the levels of copper, lithium, and strontium ions in the crevicular fluid of patients with periodontitis are significantly lower than normal. Introducing trace elements that promote the synergistic regeneration of multiple periodontal tissues during periodontal regeneration can stimulate the regeneration of cementum-periodontal ligament-alveolar bone tissue and the functional integration of soft and hard tissues by activating the Wnt and PI3K-Akt signaling pathways. Therefore, high-performance periodontal regeneration materials need to create a favorable periodontal coagulation and immune microenvironment while simultaneously achieving controlled-release of trace elements that promote the synergistic regeneration of multiple periodontal tissues to rebuild the "cementum-periodontal ligament-alveolar bone functional complex" structure and achieve functional regeneration of periodontal tissues.

[0007] Therefore, given the current clinical lack and significant need for materials specifically designed for periodontal tissue regeneration, and addressing the complex regeneration requirements of the periodontal tissue structure (cementum-periodontal ligament-alveolar bone), this invention proposes a periodontal regeneration material. Customized mesoporous silica microspheres can adsorb fibrinogen during coagulation, forming heterogeneous fibrinogen nuclei that regulate the fibrin assembly process in periodontal coagulation, creating periodontal blood clots adapted for periodontal regeneration. This fosters a regenerative immune microenvironment and reduces periodontal tissue regeneration failure caused by fibrin encapsulation. Simultaneously, the periodontal regeneration-promoting trace elements loaded in the mesoporous silica microspheres are chemically bonded to the silica, rather than adsorbed by ions. This avoids poor coagulation caused by early burst release of trace elements. Furthermore, the amorphous mesoporous silica microspheres are acidic compounds that can slowly degrade under specific alkaline conditions in vivo, achieving controlled-release of trace elements and regenerating the functional complex of cementum-periodontal ligament-alveolar bone, thus realizing periodontal regeneration.

[0008] In a preferred embodiment of the periodontal regeneration material of the present invention, the mesoporous silica microspheres have an average particle size of 50-260 nm and an average specific surface area > 500 m². 2 / g.

[0009] The periodontal regeneration material provided by this invention not only has a special mesoporous structure, but also has a small particle size and a large specific surface area. Furthermore, the mesoporous size can be adjusted, thereby enabling it to load different types of trace elements and achieve superior results.

[0010] In a second aspect, the present invention provides a method for preparing the periodontal regeneration material, the method comprising the following steps:

[0011] (1) Mix the aqueous solution of the compound containing trace elements with the silica precursor solution and add anhydrous ethanol. Then adjust the pH of the system to 6-7, followed by gelation reaction and aging reaction to obtain the aged gel.

[0012] (2) After the aged gel is broken up, it is replaced with hexane to obtain a solvent-replaced gel.

[0013] (3) The gel after solvent replacement was modified with hexamethyldisilazane to obtain the modified gel;

[0014] (4) Wash the modified gel and mix it with anhydrous ethanol, break it up and disperse it to obtain an emulsion solution;

[0015] (5) The emulsion solution is dried and then calcined to obtain periodontal regeneration material.

[0016] The periodontal regeneration material prepared by the method provided in this invention has an average particle size of 50-260 nm and an average specific surface area >500 m². 2 / g can effectively promote periodontal regeneration.

[0017] As a preferred embodiment of the preparation method of the present invention, in step (1), the method for preparing the silica precursor solution is as follows: tetraethyl orthosilicate, water, anhydrous ethanol and hydrochloric acid are mixed and hydrolyzed to obtain the silica precursor solution.

[0018] Preferably, in the preparation of the silica precursor solution, the mass ratio of tetraethyl orthosilicate, anhydrous ethanol, and water is tetraethyl orthosilicate: anhydrous ethanol: water = 1:(3.5-4.5):0.5; the hydrochloric acid is used to adjust the pH of the silica precursor solution to 6.2-6.5, and the molar concentration of the hydrochloric acid is 0.8-1.2 mol / L.

[0019] Preferably, the hydrolysis time is 3.5-4.5 hours.

[0020] In a preferred embodiment of the preparation method of the present invention, in step (1), the molar volume ratio of trace elements to the silica precursor solution is (1-2)×10⁻⁶. -4 mol: 1L;

[0021] The aqueous solution containing trace elements has a molar concentration of 0.8-1.2 mol / L for the trace elements.

[0022] For example, the compound containing trace elements may be copper nitrate, copper sulfate, magnesium sulfate, zinc chloride, etc.

[0023] In a preferred embodiment of the preparation method of the present invention, in step (1), the gelation reaction time is 0.5-1.5h and the aging reaction time is 20-28h.

[0024] Preferably, in step (1), the gelation reaction time is 1 hour and the aging reaction time is 24 hours.

[0025] In a preferred embodiment of the preparation method described in this invention, in step (2), the volume of n-hexane is 2-4 times the volume of the aged gel, and the replacement time is 10-14 h.

[0026] Preferably, in step (2), the replacement time is 12 hours.

[0027] Preferably, in step (2), the fragments of the aged gel after breakage have a volume of (4-6)mm × (4-6)mm × (4-6)mm.

[0028] As a preferred embodiment of the preparation method of the present invention, in step (3), the modification process is as follows: hexamethyldisilazane is injected into the replaced gel at a rate of 0.3-1.5 mL / h; the volume of hexamethyldisilazane is 0.5-2 times the volume of the replaced gel, and the modification time is 72-120 h.

[0029] Preferably, in step (3), hexamethyldisilazane is injected into the replaced gel at a rate of 0.5-0.8 mL / h; the volume of hexamethyldisilazane is 1-1.5 times the volume of the replaced gel, and the modification time is 96 h.

[0030] In a preferred embodiment of the preparation method of the present invention, in step (4), the washing is performed using anhydrous ethanol.

[0031] In a preferred embodiment of the preparation method described in this invention, in step (4), the volume of anhydrous ethanol is 4-6 times the volume of the modified gel.

[0032] As a preferred embodiment of the preparation method of the present invention, in step (4), the crushing and dispersion are carried out by a cell-breaking probe ultrasonic pulverizer with a power of 120-170w and a time of 3-6min.

[0033] In a preferred embodiment of the preparation method of the present invention, in step (5), the heating rate of calcination is 110-130℃ / h, the calcination temperature is 550-650℃, and the calcination time is 3-6h.

[0034] Preferably, in step (5), the calcination heating rate is 120℃ / h, the calcination temperature is 600℃, and the calcination time is 4h.

[0035] In a preferred embodiment of the preparation method of the present invention, in step (5), the drying temperature is 75-85℃ and the drying time is 22-26h.

[0036] In a third aspect, the present invention provides the application of the periodontal regeneration material in the preparation of materials that regulate the periodontal coagulation immune microenvironment.

[0037] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0038] The periodontal regeneration material prepared by the method of this invention is a mesoporous silica loaded with periodontal regeneration-promoting trace elements and capable of regulating the periodontal coagulation immune microenvironment. Firstly, the periodontal regeneration material provided by this invention can construct an ideal periodontal blood clot microstructure by regulating the fibrin assembly process during periodontal coagulation, thereby guiding periodontal tissue regeneration. Secondly, addressing the specific needs of the periodontal blood clot immune microenvironment, the fibrin network structure of the blood clot generated after using the periodontal regeneration material provided by this invention can reduce inflammatory responses, induce a regenerative immune microenvironment, and reduce fibrin... The invention addresses several issues: firstly, the failure of periodontal tissue regeneration due to encapsulation; secondly, addressing the need for periodontal complex tissue regeneration, the trace elements in the periodontal regeneration material provided by this invention can promote the synergistic regeneration of multiple periodontal tissues, thereby regenerating the functional structure of the "cementum-periodontal ligament-alveolar bone functional complex"; thirdly, in the periodontal regeneration material provided by this invention, the trace elements form chemical bonds with mesoporous silica microspheres, preventing early ion burst release from adversely affecting coagulation; and fourthly, the periodontal regeneration material provided by this invention is an acidic compound that can be slowly degraded under alkaline conditions in vivo, achieving the purpose of releasing trace elements. Furthermore, the preparation method of the periodontal regeneration material provided by this invention is simple to operate and beneficial for practical applications. Attached Figure Description

[0039] Figure 1 The image shows a scanning electron microscope image of the periodontal regeneration material prepared in Example 1.

[0040] Figure 2 Selected area electron diffraction pattern of the periodontal regeneration material prepared in Example 1;

[0041] Figure 3 The Fourier transform infrared spectrum of the periodontal regeneration material prepared in Example 1 is shown below.

[0042] Figure 4 The image shows the contact angle test results of the periodontal regeneration material prepared in Example 1;

[0043] Figure 5 The figure shows the nitrogen adsorption-desorption test results of the periodontal regeneration material prepared in Example 1;

[0044] Figure 6 The graph shows the Cu release curve of the periodontal regeneration material prepared in Example 1.

[0045] Figure 7 This is a transmission electron microscope image of the periodontal regeneration material prepared in Example 1 after 8 days of degradation;

[0046] Figure 8 The image shows the results of regulating the blood clot structure using the periodontal regeneration material prepared in Example 1:

[0047] Figure 9 This is a diagram showing the results of regulating the immune microenvironment with the periodontal regeneration material prepared in Example 1;

[0048] Figure 10 The image shows the results of periodontal regeneration material prepared in Example 1 promoting the regeneration of periodontal composite tissue.

[0049] Figure 11 This is a schematic diagram illustrating the preparation and action process of periodontal regeneration materials. Detailed Implementation

[0050] To better illustrate the purpose, technical solution, and advantages of the present invention, the present invention will be further described below in conjunction with specific embodiments.

[0051] Unless otherwise specified, the reagents, methods and equipment used in this invention are all conventional reagents, methods and equipment in the field.

[0052] Example 1

[0053] This invention provides a periodontal regeneration material, the preparation method of which includes the following steps:

[0054] (1) Preparation of silica precursor solution: 5 mL tetraethyl orthosilicate, 2.5 mL ultrapure water, 10 mL anhydrous ethanol and 10 μL 1 mol / L hydrochloric acid were mixed and hydrolyzed for 4 h to obtain silica precursor solution with pH value of 6.2.

[0055] (2) Add 2.5 μL of 1 mol / L copper nitrate solution to the silica precursor solution in step (1) and mix well;

[0056] (3) Add 7.5 mL of anhydrous ethanol to the system after mixing in step (2), and adjust the pH of the system to 6.5 with 0.15 mol / L ammonia. Then gel for 1 h, and then let it stand for 24 h to age, and obtain the aged gel.

[0057] (4) The aged gel was broken into 5mm×5mm×5mm pieces, and then 30mL of n-hexane (3 times the volume of the aged gel) was added to replace it for 12h to obtain the solvent-replaced gel.

[0058] (5) Inject 40 mL of hexamethyldisilazane (1 times the volume of the replaced gel) into the replaced gel at a rate of 0.5 mL / h. Start timing from the beginning of adding hexamethyldisilazane and modify for a total of 96 h to obtain the modified gel.

[0059] (6) Wash the modified gel with anhydrous ethanol, then mix it with 5 times the volume of anhydrous ethanol and add it to a cell disruption probe ultrasonic grinder. Disperse it at 150w for 5 minutes to obtain an emulsion solution.

[0060] (7) The emulsion solution was dried at 80°C for 24 hours, and then calcined in a muffle furnace at 600°C for 4 hours. The heating rate during the calcination process was 120°C / h. After calcination, periodontal regeneration material was obtained.

[0061] Example 2

[0062] This invention provides a periodontal regeneration material. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (5). In this example, step (5) is as follows: 40 mL of hexamethyldisilazane (1 times the volume of the replaced gel) is injected into the replaced gel at a rate of 1.5 mL / h. The modification is carried out for a total of 72 h from the start of the addition of hexamethyldisilazane to obtain the modified gel.

[0063] Example 3

[0064] This invention provides a periodontal regeneration material. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (2). In this embodiment, step (2) is: adding 2.5 μL of 1 mol / L copper sulfate solution to the silica precursor solution in step (1) and mixing them evenly.

[0065] Example 4

[0066] This invention provides a periodontal regeneration material. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (2). In this embodiment, step (2) is: adding 2.5 μL of 1 mol / L magnesium sulfate solution to the silica precursor solution in step (1) and mixing them evenly.

[0067] Example 5

[0068] This invention provides a periodontal regeneration material. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (2). In this embodiment, step (2) is: adding 2.5 μL of 1 mol / L zinc chloride solution to the silica precursor solution in step (1) and mixing them evenly.

[0069] Comparative Example 1

[0070] The present invention provides a periodontal regeneration material in a comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is that step (2) is not included, that is, the periodontal regeneration material obtained does not contain trace elements.

[0071] Comparative Example 2

[0072] The present invention provides a periodontal regeneration material in a comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (2). Step (2) of this comparative example is: adding 25 μL of 1 mol / L copper nitrate solution to the silica precursor solution in step (1) and mixing evenly.

[0073] Comparative Example 3

[0074] The present invention provides a periodontal regeneration material in a comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (3). Step (3) of this comparative example is as follows: 7.5 mL of anhydrous ethanol is added to the system after uniform mixing in step (2), and the pH value of the system is adjusted to 7.8 with 0.15 mol / L ammonia water. Then gel for 1 h, and then stand for aging for 24 h to obtain the aged gel.

[0075] Comparative Example 4

[0076] The present invention provides a periodontal regeneration material in a comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (3). Step (3) of this comparative example is as follows: 7.5 mL of anhydrous ethanol is added to the system after uniform mixing in step (2), and the pH value of the system is adjusted to 8.0 with 0.15 mol / L ammonia water. Then gel for 1 h, and then stand for aging for 24 h to obtain the aged gel.

[0077] Comparative Example 5

[0078] The present invention provides a periodontal regeneration material in a comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (5). Step (5) of this comparative example is as follows: 40 mL of hexamethyldisilazane (1 times the volume of the replaced gel) is injected into the replaced gel at a rate of 2.5 mL / h. The modification is carried out for a total of 96 h from the start of the addition of hexamethyldisilazane to obtain the modified gel.

[0079] Comparative Example 6

[0080] The present invention provides a periodontal regeneration material in comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (7). Step (7) of this comparative example is: the emulsion solution is dried at 80°C for 24 hours, and then calcined in a muffle furnace at 450°C for 4 hours, wherein the heating rate during the calcination process is 120°C / h.

[0081] Comparative Example 7

[0082] The present invention provides a periodontal regeneration material in comparative example. The only difference between the preparation method of the periodontal regeneration material and that of Example 1 is step (7). Step (7) of this comparative example is: the emulsion solution is dried at 80°C for 24 hours, and then calcined in a muffle furnace at 750°C for 4 hours, wherein the heating rate during the calcination process is 120°C / h.

[0083] Example of effect

[0084] The performance of the periodontal regeneration materials prepared in the examples and comparative examples of this invention is verified by the effect examples, wherein the preparation and action process of the periodontal regeneration materials are shown in the schematic diagram. Figure 11 As shown; specifically including the following aspects:

[0085] 1. Physicochemical property testing

[0086] The prepared periodontal regeneration material was subjected to physicochemical property tests. Specifically, its morphology and structure were characterized by scanning electron microscopy and transmission electron microscopy, its chemical groups were characterized by Fourier transform infrared spectroscopy, its hydrophilicity was characterized by contact angle test, its specific surface area was characterized by nitrogen adsorption-desorption test, its trace element slow release ability was characterized by inductively coupled plasma atomic emission spectrometry, and the periodontal regeneration material after 8 days of degradation was further observed by transmission electron microscopy.

[0087] Data on average particle size, contact angle, average specific surface area, and trace element release capacity were recorded, and the results are shown in Table 1.

[0088] Table 1

[0089]

[0090]

[0091] As shown in Table 1, the average particle size of the product prepared using the technical solution of this invention is between 212.81 and 236.45 nm, and the average specific surface area is between 667.70 and 834.52 m². 2 The particle size is between 0.25 and 0.32 mg / L, and the trace element release capacity over 8 days is between 0.25 and 0.32 mg / L. That is, the product prepared by the technical solution of this invention has a small particle size, a large average specific surface area, excellent hydrophilicity, and good trace element sustained release capacity.

[0092] As can be seen from Example 1 and Comparative Example 1, the particle size and pore size of the material did not change significantly compared with the material without trace elements. As can be seen from Example 1 and Comparative Examples 3-4, when the pH value during gelation is too high, the diameter of the mesopores formed will also be too high or too low, and the pore size can be adjusted according to specific needs. As can be seen from Example 1 and Comparative Example 5, when the rate of adding the modifier during modification is too fast, the specific surface area will change significantly, and the specific surface area can be adjusted according to specific needs. As can be seen from Example 1 and Comparative Examples 6-7, when the calcination temperature is too low or too high, the element release rate changes significantly, and the specific release rate can be adjusted according to specific needs.

[0093] Furthermore, images of the periodontal regeneration material prepared in Example 1 are attached. Specifically, its morphology and structure were characterized using scanning electron microscopy, and the results are as follows. Figure 1 As shown, the chemical groups were characterized using Fourier transform infrared spectroscopy, and the results are as follows. Figure 3 As shown, its hydrophilicity was characterized using a contact angle test, and the results are as follows. Figure 4 As shown, its specific surface area was characterized using a nitrogen adsorption-desorption test, and the results are as follows. Figure 5 As shown, the slow-release capability of Cu element was characterized using inductively coupled plasma atomic emission spectrometry, and the results are as follows. Figure 6 As shown, the periodontal regeneration material was further observed using transmission electron microscopy after 8 days of degradation, and the results are as follows. Figure 7 As shown;

[0094] Among them, by Figure 1 It is evident that the periodontal regeneration material prepared in Example 1 has a size between 50-260 nm and exhibits a unique mesoporous structure; Figure 2 It can be seen that the periodontal regeneration material prepared in Example 1 has an amorphous structure; Figure 3 It can be seen that the periodontal regeneration material prepared in Example 1 contains only Si-O bonds and -OH groups, with no other compound residues; Figure 4 It can be seen that the periodontal regeneration material prepared in Example 1 is a hydrophilic material, and due to its good water absorption, no water droplets are visible; Figure 5 It can be seen that the periodontal regeneration material prepared in Example 1 has a typical isothermal ring caused by mesoporous structure, and the specific surface area test value is greater than 700m². 2 / g; by Figure 6 The Cu release curve in the sample shows that the Cu in the periodontal regeneration material prepared in Example 1 is released stably and has a significant sustained-release effect, avoiding the adverse effects of early ion burst release on coagulation; Figure 7It can be seen that the periodontal regeneration material prepared in Example 1 has obvious degradability, and its original morphology is no longer visible after 8 days of degradation.

[0095] 2. Blood clot regulation performance

[0096] Test Procedure: A 24-well plate was divided into five groups: a blank control and a material group. Each group had at least three biological replicates. The blank control group was treated with 22 μl of pure water and 200 μl of whole rat blood, while the material group was treated with 22 μl of the material solution and 200 μl of whole rat blood. The plates were shaken, and the blood clotting time was recorded. Blood clots were fixed with paraformaldehyde, dehydrated, embedded, sectioned, and stained with hematoxylin and eosin (HE) to observe the clot structure.

[0097] The results are shown in Table 2;

[0098] Table 2

[0099]

[0100]

[0101] As can be seen from Table 2, when the technical solution of the present invention is adopted, the obtained product has good blood clot regulation performance, and the result is that it can reduce the blood coagulation time. As can be seen from Examples 1-5 and the blank control group, when various trace elements are loaded, they can all reduce the blood coagulation time. As can be seen from Examples 1 and Comparative Example 2, when the amount of loaded trace elements is too large, the coagulation time increases, which is positively correlated with the amount of loaded elements. As can be seen from Examples 1 and Comparative Examples 3-4, when the pH value during gelation increases, the coagulation time increases, which is positively correlated with excessively high pH. As can be seen from Examples 1 and Comparative Examples 6-7, when the calcination temperature is too low, the material is still in a hydrophobic state and cannot be integrated with the blood, thus prolonging the coagulation time.

[0102] The periodontal regeneration material prepared in Example 1 and the blood clot section results obtained from the Blank group ( Figure 8 The results show that the periodontal regeneration material prepared in Example 1 forms a dense, fine-diameter, and multi-branched blood clot network structure after application. The prepared periodontal regeneration material can adsorb fibrinogen during the coagulation process, forming "heterogeneous fibrinogen nuclei" to regulate the fibrin assembly process in periodontal coagulation.

[0103] 3. The immune microenvironment regulates periodontal regeneration performance.

[0104] 1) Immune microenvironment regulation performance

[0105] Test Procedure: The test consisted of a blank control group and a material group, with at least three biological replicates in each group. In 24-well plates, 22 μl of purified water and 200 μl of whole rat blood were added to the blank control group, while 22 μl of material solution and 200 μl of whole rat blood were added to the material group, resulting in blank and material blood clots. Red blood cells were lysed using 1X erythrocyte lysis buffer and seeded into the RAW264.7 macrophage cell line. Total RNA was extracted after 24 h, and RT-qPCR was used to detect the expression of genes related to the macrophage immune microenvironment.

[0106] Depend on Figure 9 The results of the periodontal regeneration material prepared in Example 1 regulating the immune microenvironment show that after implantation of the material group in Example 1, the expression of anti-inflammatory genes in the periodontal defect was upregulated, such as CD206, Tgfb1, and IL4, while the expression of pro-inflammatory genes was downregulated, such as TNF and IL1. This indicates that the periodontal regeneration material prepared in this invention can effectively regulate the periodontal regeneration immune microenvironment.

[0107] 2) Periodontal regeneration performance

[0108] Test Procedure: The test consisted of a blank control group and a material group, with at least three biological replicates in each group. The blank control group was treated with 22 μl of pure water and 200 μl of whole rat blood, while the material group was treated with 22 μl of the material solution and 200 μl of whole rat blood. Blank blood clots and material blood clots were obtained. Defects were prepared in the mandibular premolar region of rats, and blood clots were implanted. Samples were taken at 28 days to observe the periodontal regeneration of the rats.

[0109] Depend on Figure 10 The results of the periodontal regeneration material prepared in Example 1 promoting the regeneration of periodontal composite tissue can be seen in the figure:

[0110] in, Figure 10 The black arrow points to newly formed cementum, the asterisk indicates widened newly formed periodontal ligament fibers, and NB indicates newly formed alveolar bone tissue. Figure 10 As can be seen from the results, the periodontal regeneration material prepared in Example 1 showed local regeneration of alveolar bone-periodontal ligament-cementum, while only the regeneration of fibers and a small amount of new bone was observed in the Control group, suggesting that the periodontal regeneration material prepared in this invention can achieve good periodontal multi-tissue regeneration.

[0111] Finally, it should be noted that the above embodiments are used to illustrate the technical solutions of the present invention and not to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A periodontal regeneration material, characterized in that, The periodontal regeneration material is a mesoporous silica microsphere that regulates the periodontal coagulation immune microenvironment. The mesoporous silica microsphere is loaded with trace elements that promote periodontal regeneration. The trace elements include at least one of copper, strontium, magnesium, lithium, iron, zinc, manganese, selenium, molybdenum, and cobalt. The preparation method includes the following steps: (1) Mix the aqueous solution of the compound containing trace elements with the silica precursor solution and add anhydrous ethanol. Then adjust the pH of the system to 6-7, followed by gelation reaction and aging reaction to obtain the aged gel. (2) After the aged gel is broken up, it is replaced with hexane to obtain a solvent-replaced gel; (3) Modify the solvent-displaced gel using hexamethyldisilazane to obtain the modified gel; (4) Wash the modified gel and mix it with anhydrous ethanol, break it up and disperse it to obtain an emulsion solution; (5) The emulsion solution is dried and then calcined to obtain periodontal regeneration material; In step (1), the molar volume ratio of trace elements to the silica precursor solution is (1-2)×10. -4 mol: 1L; The modification process is as follows: hexamethyldisilazane is injected into the replaced gel at a rate of 0.3-1.5 mL / h; The calcination temperature is 550-650℃.

2. The periodontal regeneration material according to claim 1, characterized in that, The mesoporous silica microspheres have an average particle size of 50-260 nm and an average specific surface area >500 m². 2 / g.

3. The periodontal regeneration material according to claim 1, characterized in that, In step (1), the method for preparing the silica precursor solution is as follows: mix tetraethyl orthosilicate, water, anhydrous ethanol and hydrochloric acid and hydrolyze to obtain the silica precursor solution.

4. The periodontal regeneration material according to claim 1, characterized in that, In step (1), the gelation reaction time is 0.5-1.5h, and the aging reaction time is 20-28h.

5. The periodontal regeneration material according to claim 1, characterized in that, In step (2), the volume of n-hexane is 2-4 times the volume of the aged gel, and the replacement time is 10-14h.

6. The periodontal regeneration material according to claim 1, characterized in that, In step (3), the volume of hexamethyldisilazane is 0.5-2 times the volume of the replaced gel, and the modification time is 72-120h.

7. The periodontal regeneration material according to claim 1, characterized in that, In step (5), the heating rate of calcination is 110-130℃ / h, and the calcination time is 3-6h.

8. The application of the periodontal regeneration material as described in claim 1 or 2 in the preparation of materials that regulate the periodontal coagulation immune microenvironment.

Images