Functionalized guided tissue regeneration membrane as well as preparation method and application thereof
The amino-functionalized black phosphorus/titanium oxide nanosheet composite GTR membrane constructed through layer-by-layer self-assembly technology solves the problems of single function and unstable performance of existing GTR membranes, realizes intelligent synergy of antibacterial and osteogenic functions, adapts to the dynamic needs of periodontal tissue regeneration process, and improves treatment efficacy and safety.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2026-01-16
- Publication Date
- 2026-04-14
AI Technical Summary
Existing GTR membranes have limited functionality, making it difficult to meet both antibacterial and osteogenic requirements. Their release behavior is uncontrollable, and their simple surface modification methods lead to unstable performance. They also lack the ability to precisely control their spatiotemporal properties, which limits their efficacy in the complex microenvironment of periodontitis.
A layer-by-layer self-assembly technique was used to construct an amino-functionalized black phosphorus/titanium oxide nanosheet composite GTR membrane. The antibacterial and osteogenic functions were intelligently synergistically achieved through external light modulation. The functional release was precisely regulated by photocatalysis and photothermal effects, and the number of nanosheet layers and the amount of amino modification were precisely controlled.
It achieves integrated antibacterial and osteogenic functions, possesses precise designability and individualized adaptation, and can dynamically adjust functions according to the treatment stage, thereby improving efficacy and reducing the risk of bacterial resistance, and ensuring functional stability and reliability.
Smart Images
Figure CN121846385A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of biomaterials technology, and in particular to a functionalized tissue regeneration membrane, its preparation method, and its application. Background Technology
[0002] Guided tissue regeneration (GTR) membranes are biomaterials that act as a physical barrier to prevent the migration of cells such as gingival epithelium, providing space for periodontal or bone tissue regeneration. They are primarily used in periodontal disease and dental implant bone defect repair, guiding the regeneration of periodontal ligaments, alveolar bone, and other tissues. Materials are classified as non-absorbable (such as titanium mesh) and absorbable (such as collagen membranes and chitosan membranes), with absorbable membranes being more favored because they do not require secondary surgery. Recent developments, such as asymmetric porous membranes and tea polyphenol-regulated hydrogel membranes, optimize mechanical properties and degradation rates, improving regeneration efficacy.
[0003] Although GTR membranes have achieved some success in isolating soft tissues, existing technologies still have significant limitations, mainly in the following aspects: (1) Single function, making it difficult to meet both antibacterial and osteogenic needs: Traditional GTR membranes mainly play the role of physical barriers, and the material itself usually does not have active biological functions. In order to enhance the therapeutic effect, existing improvement technologies mostly focus on giving GTR membranes a single function; (2) Lack of precise regulation and individualized treatment capabilities: The release behavior of functional molecules (such as rate and dose) of existing functional GTR membranes is mostly passively controlled by the degradation or diffusion process of the material itself, making it difficult to achieve programmed and intelligent precise regulation; (3) Simple surface modification technology, insufficient interfacial bonding and stability: At present, the functional modification of GTR membranes mostly adopts simple processes such as bulk blending or surface physical adsorption. The functional molecules loaded by these methods are unevenly distributed, have weak bonding with the membrane matrix, and are easily lost rapidly under the flushing of body fluids, resulting in a short and uncontrollable functional period.
[0004] Existing GTR membrane technologies suffer from several core drawbacks, including limited functionality (antibacterial and osteogenic effects are mutually exclusive), uncontrollable release behavior, lack of precise spatiotemporal regulation, and unstable performance due to simplistic surface modification methods. These limitations severely restrict the efficacy of GTR membranes in achieving efficient tissue regeneration within the complex microenvironment of periodontitis. Therefore, there is an urgent need to develop a novel GTR membrane modification strategy that simultaneously endows the membrane with durable and highly effective antibacterial capabilities and active osteogenic activity, providing a stable and controllable surface construction method to ensure durable and reliable function. Summary of the Invention
[0005] The purpose of this invention is to provide a functionalized guided tissue regeneration membrane, its preparation method, and its application. Based on a layer-by-layer self-assembly technology, an amino-functionalized black phosphorus / titanium oxide nanosheet composite GTR membrane achieves integrated and intelligent synergy of antibacterial and osteogenic functions. It possesses precise designability and individualized adaptation potential, aligning with the temporal treatment concept of the dynamic process of tissue regeneration. Precise temporal treatment is achieved through external light modulation: in the initial treatment phase, visible light parameters are used to stimulate a strong antibacterial effect, while in the restorative phase, the near-infrared light mode is adjusted to continuously promote osteolysis, thereby meeting the dynamic needs of different stages of periodontal tissue regeneration.
[0006] To achieve the above objectives, the present invention provides a functionalized guided tissue regeneration membrane with dual functions of antibacterial and osteogenic properties. The membrane comprises a base layer and a multilayer composite coating. The multilayer composite coating is formed by alternating deposition on the surface of the base layer through a layer-by-layer self-assembly technique. It is composed of alternating layers of positively charged amino-functionalized black phosphorus nanosheets and negatively charged two-dimensional titanium dioxide nanosheets, with the structure (BP-NH2 / TNs)n, where n is a value between 1 and 20.
[0007] The present invention also provides a method for preparing a functionalized guided tissue regeneration membrane, comprising the following steps.
[0008] Step 1, Substrate pretreatment: After cleaning and drying, the substrate layer is subjected to plasma treatment to make the substrate surface negatively charged; Step 2, Preparation of assembly unit dispersion: Prepare positively charged amino-functionalized black phosphorus nanosheet dispersion and negatively charged two-dimensional titanium dioxide nanosheet dispersion; wherein, the number of amino groups on the surface of amino-functionalized black phosphorus nanosheets is controlled by adjusting the amination reaction conditions, and amino-functionalized black phosphorus nanosheets with different numbers of layers are obtained by adjusting the exfoliation process. Step 3, Layer-by-layer self-assembly: The substrate pretreated in Step 1 is sequentially and alternately immersed in amino-functionalized black phosphorus nanosheet dispersion and two-dimensional titanium dioxide nanosheet dispersion. After each immersion, it is cleaned and dried. This process is repeated n times to form a multilayer composite coating. Step 4, Post-processing: The assembled composite membrane is finally cleaned and cured to obtain a functionalized tissue regeneration guided membrane.
[0009] Preferably, in step 2, amino-functionalized black phosphorus nanosheets are prepared by controlling the degree of exfoliation and amination of black phosphorus through plasma to obtain amino-functionalized black phosphorus nanosheets with different number of layers and different amino modification densities; two-dimensional titanium dioxide nanosheets are prepared by exfoliating layered titanates to obtain two-dimensional titanium dioxide nanosheets, ensuring that they are stably dispersed in the aqueous phase and have a negatively charged surface.
[0010] The present invention also provides an application of a functionalized guided tissue regeneration membrane for the preparation of medical devices for treating periodontitis.
[0011] Preferably, the guided tissue regeneration membrane is used to regulate the sequential antibacterial and osteogenic functions through external light irradiation parameters, in order to prepare periodontal tissue regeneration materials with sequential therapeutic effects.
[0012] Preferably, the guided tissue regeneration membrane is used to kill periodontal pathogens and promote alveolar bone regeneration.
[0013] Preferably, periodontal pathogens include Porphyromonas gingivalis, Aggregobacter actinomycetes, and Streptococcus mutans.
[0014] The advantages and beneficial effects of the above-mentioned functionalized guided tissue regeneration membrane, its preparation method, and its application are as follows: 1. This invention utilizes layer-by-layer self-assembly (LBL) technology to orderly assemble amino-functionalized black phosphorus nanosheets (BP-NH2) with photocatalytic-photothermal antibacterial properties and two-dimensional titanium dioxide nanosheets with good biocompatibility and photocatalytic activity at the molecular / nanoscale. This achieves a high degree of integration of photocatalytic-photothermal synergistic antibacterial action and photothermal / bio-ion dual-mode osteogenic action on a single GTR membrane. Both functions are uniformly triggered and regulated by the non-invasive physical signal of external light irradiation. Under light irradiation, antibacterial and osteogenic processes are simultaneously initiated. Furthermore, by adjusting different light sources and irradiation parameters (wavelength, time), the intensity and timing of the two functions can be precisely controlled in a "switching" or "gradient" manner, intelligently matching treatment needs.
[0015] 2. This invention allows for precise control of the thickness and loading of the functional coating by adjusting the number of assembled LBL layers, thereby macroscopically regulating the strength and durability of the function. By changing the amount of amino modification in BP-NH2 and the number (thickness) of nanosheets, the surface chemistry, band structure, photothermal conversion efficiency, and degradation rate of the composite material can be controlled at the molecular level, thereby microscopically regulating antibacterial activity, phosphorus release kinetics, and cellular response behavior. This enables physicians to tailor the most suitable GTR membrane for different patients based on preoperative assessments.
[0016] 3. The LBL technology of this invention is based on electrostatic interaction. The nanocomposite coating constructed on the surface of the GTR membrane is firmly bonded, significantly improving the stability of the functional layer, preventing the rapid loss of functional components, and ensuring long-term efficacy. Relying on photocatalytically generated reactive oxygen species (ROS) and photothermal effects to kill bacteria is a physicochemical mechanism that is less likely to induce bacterial resistance, solving the clinical problems caused by antibiotic overuse. The degradation products of black phosphorus are biocompatible phosphates, a natural component of bone; titanium dioxide is bio-inert and stable. The combination of these two technologies ensures both functionality and enhanced biosafety.
[0017] 4. This invention conforms to the time-sequential treatment concept of the dynamic process of tissue regeneration, and its design concept perfectly matches the periodontal tissue regeneration process: in the early stage of implantation, visible light fluctuations exert a strong antibacterial effect and purify the wound; during the tissue repair period, the near-infrared light irradiation mode is adjusted to focus on using gentle photothermal stimulation and slow release of phosphorus ions to continuously promote bone formation. This time-varying, programmable functional output mode is not available in existing GTR membrane technology.
[0018] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. Attached Figure Description
[0019] Figure 1 The in vitro photocatalytic antibacterial activity test of the functionalized guided tissue regeneration membrane (GTR) of the present invention is shown in (a) as Escherichia coli colony images on GTRs with different modification layers and (b) as the antibacterial rate of GTRs with different modification layers. Figure 2 The in vitro photothermal antibacterial activity test of the functionalized guided tissue regeneration membrane (GTR) of the present invention is shown in (a) as Escherichia coli colony images on GTRs with different modified layers and (b) as the antibacterial rate of GTRs with different modified layers. Figure 3 The in vitro photothermal performance test of the functionalized guided tissue regeneration membrane (GTR) of the present invention is shown in (a) as the temperature rise curves of the unmodified and modified GTR membranes under near-infrared light, and (b) as the thermal images of the unmodified and modified GTR membranes under near-infrared light. Figure 4 This invention functionalizes the expression of osteogenic genes in the tissue regeneration membrane (GTR), wherein (a) represents the expression of the osteogenic gene OCN, and (b) represents the expression of the osteogenic gene Runx2. p<0.05, p<0.01, p<0.001); Figure 5 (a) represents the antibacterial rate of Streptococcus mutans under visible light irradiation, and (b) represents the antibacterial rate of Streptococcus mutans under near-infrared light irradiation. Figure 6 (a) shows the rat body weight after 6 weeks of treatment, and (b) shows the blood biochemical analysis after 6 weeks of treatment. Figure 7 In the middle, (a) is the vertical distance between the alveolar bone ridge (ABC) and the cementum-enamel junction (CEJ), and (b) is the relative bone volume; Figure 8The electron paramagnetic resonance spectra of free radicals are used to detect the free radicals, where (a) represents superoxide radicals and (b) represents hydroxyl radicals. Detailed Implementation
[0020] The technical solution of the present invention will be further described below with reference to the accompanying drawings and embodiments.
[0021] Unless otherwise defined, the technical or scientific terms used in this invention shall have the ordinary meaning as understood by one of ordinary skill in the art to which this invention pertains.
[0022] The following examples are not intended to limit the invention, but are only for illustration. Unless otherwise specified, the experimental methods used in the following examples are generally performed under conventional conditions. Unless otherwise specified, the materials and reagents used in the following examples are commercially available.
[0023] Example 1 A functionalized tissue regeneration guiding membrane includes a base layer and a multilayer composite coating. The base layer includes a collagen membrane, a polylactic acid membrane, and an oral repair membrane. The base layer can be any tissue regeneration guiding membrane, including but not limited to collagen membranes, polylactic acid membranes, and oral repair membranes. The multilayer composite coating is formed by alternating deposition on the surface of the base layer using a layer-by-layer self-assembly technique. It consists of alternating layers of positively charged amino-functionalized black phosphorus nanosheets and negatively charged two-dimensional titanium dioxide nanosheets, with the structure (BP-NH2 / TNs)n, where n is a value between 1 and 20. The composite coating consists of functionalized black phosphorus nanosheets and titanium dioxide nanosheets. For example, n=1 indicates a single-layer composite coating, n=2 indicates a two-layer composite coating, and n=3.5 indicates a three-layer composite coating or a half-layer composite coating (i.e., a single functionalized black phosphorus nanosheet layer or a single titanium dioxide nanosheet layer).
[0024] A method for preparing a functionalized tissue regeneration membrane includes the following steps.
[0025] Step 1, Substrate Pretreatment: After cleaning and drying, the substrate layer is subjected to plasma treatment to make the substrate surface negatively charged. Cleaning the surface, introducing active groups, and adjusting their surface charge (usually making them negatively charged) enhances the adhesion to subsequent nanosheets.
[0026] Step 2, Preparation of assembly unit dispersions: Preparation of positively charged amino-functionalized black phosphorus nanosheet dispersions and negatively charged two-dimensional titanium dioxide nanosheet dispersions; wherein, the number of amino groups on the surface of the amino-functionalized black phosphorus nanosheets is controlled by adjusting the amination reaction conditions, and amino-functionalized black phosphorus nanosheets with different numbers of layers are obtained by adjusting the exfoliation process.
[0027] Amino-functionalized black phosphorus nanosheets (BP-NH2) were prepared. By controlling the degree of exfoliation and amination of black phosphorus using plasma, BP-NH2 with different numbers of layers (e.g., monolayer, few layers) and different amino modification densities could be obtained.
[0028] Two-dimensional titanium dioxide nanosheets were prepared. Two-dimensional titanium dioxide nanosheets were obtained by exfoliating layered titanates, ensuring their stable dispersion in the aqueous phase and their negatively charged surface.
[0029] Step 3, Layer-by-layer self-assembly: The substrate pretreated in Step 1 is sequentially and alternately immersed in amino-functionalized black phosphorus nanosheet dispersion and two-dimensional titanium dioxide nanosheet dispersion. After each immersion, it is cleaned and dried. This process is repeated n times to form a multilayer composite coating.
[0030] Layer-by-layer self-assembly (LBL) process: ① First layer adsorption: The pretreated GTR membrane was immersed in a positively charged BP-NH2 dispersion for 15 minutes, so that BP-NH2 was adsorbed on the membrane surface by electrostatic interaction. After removal, it was cleaned and dried.
[0031] ② Second layer adsorption: Immerse the membrane that has adsorbed BP-NH2 into a negatively charged TNs dispersion, so that TNSs adsorb on the BP-NH2 layer to form a bilayer structure (BP-NH2 / TNs). After removal, clean and dry.
[0032] ③ Cyclic assembly: Repeat steps ① and ② to obtain composite coatings with different numbers of layers (e.g., n=1, 3, 5, 6, 6.5, 7…...), denoted as (BP-NH2 / TNs)n.
[0033] Step 4, Post-processing: The assembled composite membrane is finally cleaned and cured to obtain a functionalized tissue regeneration guided membrane.
[0034] Regulation of functional strength: The total loading of functional nanomaterials on the membrane surface can be directly controlled by changing the number of assembled layers (n) of the LBL. The number of layers affects its antibacterial and osteogenic activities.
[0035] Functional timing and balance regulation: achieved by changing the intrinsic properties of the building blocks.
[0036] Controlling the amount of amino modification in BP-NH2: By changing the amination conditions such as plasma treatment, BP-NH2 with different amino densities can be prepared. The amino density affects its surface charge, the interfacial bonding strength with TNs titanium dioxide nanosheets, and the separation efficiency of photogenerated carriers, thereby finely controlling the strength of photocatalytic antibacterial activity.
[0037] Controlling the number (thickness) of BP-NH2 layers: The band gap of black phosphorus exhibits a thickness dependence (quantum confinement effect). By using BP-NH2 with different numbers of layers, the band structure, light absorption range, and generation and separation efficiency of photogenerated electron-hole pairs in the composite material can be controlled. This directly determines the response wavelength and efficiency of photocatalytic antibacterial activity.
[0038] Dual-function principle.
[0039] (1) Photocatalytic-photothermal synergistic antibacterial mechanism.
[0040] The composite GTR membrane of this invention can achieve a synergistic antibacterial effect of photocatalysis and photothermal effect under irradiation of specific wavelengths (such as visible light or near-infrared light), producing a highly efficient and broad-spectrum bactericidal effect.
[0041] Photocatalytic antibacterial pathway: Amino-functionalized black phosphorus nanosheets (BP-NH2), acting as a photosensitive material, are excited under light to generate electron-hole pairs. Due to the tight heterojunction interface formed between BP-NH2 and two-dimensional titanium dioxide nanosheets (TNs), photogenerated electrons can efficiently migrate from the conduction band of BP-NH2 to the conduction band of TNs. This process greatly suppresses the recombination of photogenerated carriers and extends their lifetime.
[0042] Electrons migrating to the surface of titanium dioxide nanosheets (TNs) can react with dissolved oxygen (O2) in the environment to generate superoxide radicals (·O2). - ).
[0043] Holes (h) remaining in the BP-NH2 valence band + It has a strong oxidizing ability and can directly attack the cell wall / membrane of bacteria, or react with water molecules (H2O) to generate hydroxyl radicals (·OH).
[0044] These reactive oxygen species (ROS, including O2) - ·OH and h + These free radicals can act together on bacteria, damaging their cell membrane structure, proteins, enzymes, and genetic material (DNA / RNA) through oxidation, leading to bacterial death. The presence of these free radicals and their main target species can be verified using electron paramagnetic resonance (ESR) spectroscopy. Figure 8 (a) and Figure 8 (b)
[0045] Photothermal-assisted antibacterial pathway: Black phosphorus nanosheets possess excellent photothermal conversion properties. Under the same illumination, BP-NH2 can efficiently convert absorbed light energy into heat energy, causing a rapid and controllable increase in the local temperature of the composite material. A moderate photothermal effect can: exacerbate the fluidity of the bacterial cell membrane phospholipid bilayer, disrupting its integrity and making it more sensitive to ROS attack; cause reversible or irreversible denaturation of bacterial proteins, interfering with their normal physiological functions; and create a synergistic effect with ROS generated by photocatalysis, jointly disrupting the structure of bacterial biofilms and significantly improving the bactericidal efficiency.
[0046] (2) Mechanism of promoting osteolysis.
[0047] The osteogenic function of the composite GTR membrane is mainly achieved through the mechanism of photothermal stimulation and synergistic release of bioactive ions, providing multiple beneficial signals for alveolar bone regeneration.
[0048] Controllable photothermal stimulation promotes bone growth: Utilizing the photothermal effect of BP-NH2, external near-infrared light can be used to perform non-contact, spatiotemporally precise thermal stimulation of the local microenvironment at the implantation site.
[0049] Moderate heat stimulation (mild warming) has been shown to promote osteoblast proliferation, differentiation, and extracellular matrix secretion. Heat stimulation can upregulate the expression of osteogenic-related marker genes (such as Runx2, ALP, and OCN) and enhance alkaline phosphatase (ALP) activity, thereby accelerating the osteogenic process.
[0050] By adjusting light parameters (such as intensity, time, and interval), precise control over the intensity and duration of thermal stimulation can be achieved, simulating a physiological thermal environment conducive to bone repair.
[0051] Slow-release of bioactive phosphorus: Black phosphorus nanosheets undergo mild, controllable oxidative degradation in physiological environments, gradually releasing phosphate (PO4) ions. 3- Phosphorus exists in the form of ).
[0052] Phosphorus is a key component of the inorganic mineral phase (hydroxyapatite) of human bones and teeth. Locally released PO4... 3- It can directly participate in and promote the deposition of a bone-like apatite layer on the surface of implant materials, enhancing the osteoconductivity of the materials. PO4 3- Ions themselves are also important biological signaling molecules, capable of activating intracellular signaling pathways related to osteogenic differentiation, and stimulating mesenchymal stem cells to differentiate into osteoblasts from a biological perspective. By regulating the number of layers, size, and assembly structure of black phosphorus nanosheets, their degradation rate and phosphorus ion release kinetics can be precisely controlled, matching them to different stages of bone regeneration.
[0053] The performance of the functionalized tissue regeneration membrane prepared in Example 1 was tested.
[0054] 1. Photocatalytic antibacterial properties.
[0055] The photocatalytic antibacterial effect of GTR with different modification layers was estimated using *E. coli* (ATCC25922). *E. coli* suspensions were diluted to 10-1 7 CFU / mL was added to sterile PBS buffer. GTRs with different modification layers were then placed in the solution and irradiated with a 300W xenon lamp (100mW·cm²). -2 The cultured cells were cultured in a wavelength range of 320-780 nm for 0.5 h. The diluted cultured cells were plated on LB agar plates and co-incubated at 37°C for 24 h, and the resulting colonies were counted. Unmodified GTR membranes were used as a control. All treatments were prepared in triplicate, and each experiment was repeated at least three times. Inhibition of bacterial growth was quantified by antibacterial rate: antibacterial rate (%) = (AB) / A × 100%.
[0056] Where A represents the colony count in the untreated group and B represents the colony count in the experimental group.
[0057] like Figure 1 (a) and Figure 1 As shown in Figure (b), the number of E. coli colonies decreased with increasing GTR modification layers, indicating that GTR modification can inhibit the growth of E. coli through photocatalysis. The more modification layers, the better the photocatalytic antibacterial effect; after fifteen modification layers, the inhibition rate reached 94.4%.
[0058] 2. Photothermal antibacterial properties.
[0059] The photothermal antibacterial effect of GTR with different modification layers was estimated using *E. coli* (ATCC25922). *E. coli* suspensions were diluted to 10-1 7 CFU / mL was placed in sterile PBS buffer. Different modified GTR layers were added and irradiated with an 808nm NIR laser at a power density of 1.0W / cm² for 10 min. The cultured cell dilutions were plated on LB agar plates and co-incubated at 37°C for 24 hours, and colonies were counted. Unmodified GTR membranes were used as a control. All treatments were prepared in triplicate, and each experiment was repeated at least three times. Bacterial growth inhibition was quantified by antibacterial rate: antibacterial rate (%) = (AB) / A × 100%.
[0060] like Figure 2 (a) and Figure 2As shown in Figure (b), with the increase of the number of GTR modification layers, excellent photothermal performance was observed, and the number of E. coli colonies decreased, indicating that the modified GTR can inhibit the growth of E. coli through photothermal action. The more modification layers, the better the photothermal antibacterial effect; after fifteen modification layers, the antibacterial rate can reach 90.1%.
[0061] Resuspend the *Streptococcus mutans* in PBS solution and determine the bacterial concentration using a spectrophotometer or turbidimeter, adjusting the concentration to 10⁻⁶. 7 CFU / mL. The prepared BP-NH2 / TNs modified GTR membranes with different numbers of layers (5, 10, and 15 layers) were cut into small pieces of the same size and placed in containers containing bacterial suspensions, ensuring that the bacteria could fully contact the GTR membrane surface. Visible light irradiation ( Figure 5 (a) The sample containing bacteria and the GTR membrane is irradiated under a visible light source, and the irradiation time, light intensity, and other conditions should be kept consistent. Near-infrared photothermal conditions ( Figure 5 (b) The sample containing bacteria and the GTR membrane was irradiated under a near-infrared light source. The irradiation time, near-infrared wavelength, and light intensity were kept consistent. The treated bacterial suspension was spread onto a plate and incubated in a constant temperature incubator for 24 hours. The antibacterial rate was calculated using the equation: Antibacterial rate (%) = (AB) / A × 100%.
[0062] Comparing the antibacterial rates of BP-NH2 / TNs modified GTR membranes with different numbers of layers under visible light irradiation ( Figure 5 (a) and the antibacterial rate under near-infrared photothermal conditions ( Figure 5 In (b), under near-infrared photothermal conditions, the antibacterial rate against Streptococcus mutans increased with the increase of the number of GTR membrane layers. After reaching 15 layers, the antibacterial rate reached 95% and 90% under visible light and near-infrared photothermal conditions, respectively.
[0063] 3. Changes in light, heat, and temperature.
[0064] To evaluate the photothermal performance of the modified GTR, an 808nm NIR laser with a power density of 1.0W / cm was used, and temperature changes were recorded using an infrared thermal imager.
[0065] like Figure 3 (a) and Figure 3 As shown in (b), the unmodified GTR film did not exhibit a significant photothermal heating effect under 808 nm near-infrared irradiation. After modification, the temperature of BP-NH2 / TNS increased from 25°C to 50°C within 15 minutes of 0.5 W near-infrared irradiation, demonstrating excellent photothermal properties. This photothermal property provides both antibacterial and bone-promoting effects.
[0066] 4. Osteoblast gene expression.
[0067] like Figure 4 (a) and Figure 4 As shown in (b), the bar chart illustrates the expression levels of two key osteogenic marker genes, OCN and Runx2, under different treatment conditions. OCN and Runx2 are early and late markers in osteogenic differentiation, respectively; elevated expression levels typically indicate cell differentiation into osteoblasts or enhanced osteogenic activity. The results in the figure demonstrate that the modified GTR membrane exhibits significantly higher expression levels of these osteogenic markers. This clearly reveals that the modified GTR membrane possesses a stronger osteogenic-promoting capacity under near-infrared light.
[0068] For in vivo osteogenic experiments, a typical bacterial-induced rat model of periodontitis was established. Fifteen male SD rats were randomly divided into five groups: untreated group (Control), periodontitis (inflammation group), periodontitis + commercial GTR membrane (GTR), periodontitis + BP-NH2 / TNs group (BP-NH2 / TNs), and periodontitis + BP-NH2 / TNs group irradiated with NIR light (BP-NH2 / TNs+NIR). The surgical area was irradiated with NIR light daily. After four weeks, the maxillae with teeth were collected from the sacrificed rats and analyzed using micro-computed tomography (Micro-CT). In the Micro-CT analysis, the vertical distance between the alveolar ridge (ABC) and the enamel-osseointegration junction (CEJ) was measured using three-dimensional reconstruction technology, and the relative bone volume was used to assess osteogenic capacity. To verify the long-term biocompatibility of BP-NH2 / TNs, the weight of the animals in the BP-NH2 / TNs+NIR group was recorded, and blood biochemical analysis was performed.
[0069] To assess the biosafety of BP-NH2 / TNs-modified GTR membranes, whole blood and serum were collected from rats for routine blood tests. The untreated group (Control) and the periodontitis + BP-NH2 / TNs group were irradiated with NIR light (BP-NH2 / TNs+NIR), and blood biochemical indicators, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP), were analyzed.
[0070] from Figure 6 In (a), a slight decrease in body weight was observed in the experimental group rats, possibly due to difficulty eating caused by oral surgery, but there was no significant difference compared to the normal group. In blood biochemical analysis, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and serum alkaline phosphatase (ALP) levels were not significantly different from the normal group. Figure 6 (b) The above results confirm that BP-NH2 / TNs have good biocompatibility in vivo.
[0071] This invention uses amino-functionalized black phosphorus nanosheets (BP-NH2, positively charged) and two-dimensional titanium dioxide nanosheets (TNs, negatively charged) as basic building blocks. A nanocomposite coating with a (BP-NH2 / TNs)n periodic structure is assembled on a GTR film substrate using a precisely controlled LBL process. The key to this structure lies in the molecular-level close contact heterogeneous interface formed between the two two-dimensional materials, which is the core structural basis for achieving efficient photogenerated charge separation, energy transfer, and synergistic biological functions.
[0072] like Figure 7 (a) and Figure 7 As shown in (b), in vivo osteogenic properties were analyzed by micro-CT 15 days after treatment. The degree of alveolar bone loss was quantified by the vertical distance between the alveolar ridge (ABC) and cementum-enamel junction (CEJ) of the maxillary second molar. The results showed that the surface-modified GTR membrane exhibited good osteogenic activity under near-infrared light therapy.
[0073] This invention goes beyond the simplistic approach of merely controlling thickness. It establishes a method for multi-dimensional and precise control of the final product's performance by regulating the number of assembled layers (n), the degree of amination of BP-NH2, and the number (thickness) of black phosphorus nanosheets. This achieves comprehensive control from macroscopic loading to microscopic electronic structure, from surface chemistry to degradation kinetics, giving the GTR film's key performance indicators such as antibacterial strength, action period, and osteoproliferative rate designability and customization potential.
[0074] Utilizing an external light source as a non-invasive intelligent switch and regulator, two core functions are simultaneously triggered and controlled. This achieves a synergistic antibacterial mechanism through photocatalytic generation of reactive oxygen species (ROS) and photothermal generation of localized temperature increases. These two mechanisms mutually enhance each other, improving sterilization efficiency and reducing the risk of drug resistance. The gentle thermal stimulation generated by photothermal generation is combined with the slow-release of bioactive phosphorus (PO4) from the degradation of black phosphorus. 3- This combination provides osteoblasts with both physical and chemical beneficial signals.
[0075] This mechanism allows doctors to dynamically adjust the focus of function at different stages of treatment by adjusting the light therapy program, achieving a sequential treatment from "powerful debridement" to "gentle regeneration promotion".
[0076] Therefore, this invention employs the above-mentioned functionalized guided tissue regeneration membrane, its preparation method, and its application. Based on the layer-by-layer self-assembly technology, an amino-functionalized black phosphorus / titanium oxide nanosheet composite GTR membrane is constructed, achieving integrated and intelligent synergy of antibacterial and osteogenic functions. It possesses precise designability and potential for individualized adaptation, conforming to the temporal treatment concept of the dynamic process of tissue regeneration.
[0077] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. 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 still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.
Claims
1. A functionalized tissue regeneration guiding membrane, characterized in that: The tissue regeneration membrane has dual functions of antibacterial and osteogenic properties. It includes a base layer and a multilayer composite coating. The multilayer composite coating is formed by alternating deposition on the surface of the base layer through layer-by-layer self-assembly technology. It is composed of alternating layers of positively charged amino-functionalized black phosphorus nanosheets and negatively charged two-dimensional titanium dioxide nanosheets. Its structure is (BP-NH2 / TNs)n, where n is a value between 1 and 20.
2. The method for preparing a functionalized guided tissue regeneration membrane according to claim 1, characterized in that, Includes the following steps: Step 1, Substrate pretreatment: After cleaning and drying, the substrate layer is subjected to plasma treatment to make the substrate surface negatively charged; Step 2, Preparation of assembly unit dispersion: Prepare positively charged amino-functionalized black phosphorus nanosheet dispersion and negatively charged two-dimensional titanium dioxide nanosheet dispersion; wherein, the number of amino groups on the surface of amino-functionalized black phosphorus nanosheets is controlled by adjusting the amination reaction conditions, and amino-functionalized black phosphorus nanosheets with different numbers of layers are obtained by adjusting the exfoliation process. Step 3, Layer-by-layer self-assembly: The substrate pretreated in Step 1 is sequentially and alternately immersed in amino-functionalized black phosphorus nanosheet dispersion and two-dimensional titanium dioxide nanosheet dispersion. After each immersion, it is cleaned and dried. This process is repeated n times to form a multilayer composite coating. Step 4, Post-processing: The assembled composite membrane is finally cleaned and cured to obtain a functionalized tissue regeneration guided membrane.
3. The method for preparing a functionalized guided tissue regeneration membrane according to claim 2, characterized in that: In step 2, amino-functionalized black phosphorus nanosheets are prepared by controlling the degree of exfoliation and amination of black phosphorus through plasma to obtain amino-functionalized black phosphorus nanosheets with different number of layers and different amino modification densities; two-dimensional titanium dioxide nanosheets are prepared by exfoliating layered titanates to obtain two-dimensional titanium dioxide nanosheets, ensuring that they are stably dispersed in the aqueous phase and have a negatively charged surface.
4. The application of the functionalized tissue regeneration membrane according to claim 1, characterized in that: Used in the manufacture of medical devices for the treatment of periodontitis.
5. The application of the functionalized tissue regeneration membrane according to claim 4, characterized in that: Guided tissue regeneration membranes, through the regulation of external light parameters, exhibit temporal antibacterial and osteogenic functions, and are used to prepare periodontal tissue regeneration materials with temporal therapeutic effects.
6. The application of the functionalized tissue regeneration membrane according to claim 4, characterized in that: The tissue regeneration membrane is used to kill periodontal pathogens and promote alveolar bone regeneration.
7. The application of a functionalized tissue regeneration membrane according to claim 6, characterized in that: Periodontal pathogens include Porphyromonas gingivalis, Aggregobacter actinomycetes, and Streptococcus mutans.