Ca_3SiO_5及Ca_3SiO_5/CaCl_2复合牙科材料的制备与性能研究
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摘要
在吉林大学纳米基金重点项目的支持下,针对牙脱敏材料和根管充填材料存在问题,根据硅酸三钙自固化材料具有的良好的生物相容性、降解性和生物活性等特性,通过对应用硅酸三钙糊剂后的牙本质进行表征发现,硅酸三钙不仅通过自固化过程与牙本质紧密结合,而且在生理环境下能诱导牙本质再矿化,有效阻塞牙本质小管,可能作为脱敏治疗的有效药物。结合氯化钙的促凝作用,首次制备了硅酸三钙和氯化钙复合材料,并证明氯化钙可缩短硅酸三钙的凝固时间,提高其抗压强度,降低其pH值;体外实验研究表明,Ca3SiO5/CaCl2复合材料具有优良的生物相容性和诱导羟基磷灰石沉积的能力,根管充填后的密封性也优于传统的氧化锌丁香油水门汀,为进一步应用于根管治疗奠定了基础。
Dentin hypersensitivity, pulpitis and periradicular inflammation are common clinical conditions of stomatology, the therapy of which is closely related to the dental materials. At present, the materials of desensitizing and root canal filling have some shortcomings. Chemical and physical materials have been reported effective in reducing dentine hypersensitivity. However, these materials were not components of natural dentine, and may diminish during teeth brushing and food chewing. An ideal root canal filling material should have several properties, such as being harmless to the periapical tissues and permanent tooth germs; resorbing readily if pressed beyond the apex; easily filling the root canals; adhering to the walls of the canal; not being susceptible to shrinkage; being easily removed if necessary; being radiopaque and not discolouring the tooth. At present, none of root canal filling materials is able to meet all the above requirements. Zinc oxide eugenol cement (ZOE) was the first root canal filling material to be recommended for permanent teeth, as described by Sweet in 1930. However, later studies showed that this cement has a slow rate of resorption in the canals, limited antibacterial action and cytotoxic effects possibly causing by the release of eugenol and formaldehyde. In recent years, the introduction of calcium hydroxide in dental therapy has received much attention for its biological activity. It is antibacterial and anti-inflammatory primarily due to the high pH value of the surrounding environment (approximately 12.5) after its dissolution. In addition, calcium hydroxide can induce mineralization resulting in a repair of damaged dentin matrix. Although calcium hydroxide has some advantages compared with ZnO eugenol cement and has already been used in the root canal filling system, it has also shown disadvantages such as the low hardening rate and high dissolution, which resulting in a loose filling. So it can only be used as temporary filling materials in indications for which hermeticity is a priority. A new class of restorative material called mineral trioxide aggregate (MTA) was recently introduced as dental materials. In vitro and in vivo studies have demonstrated that the bioactivity and biocompatibility of MTA are superior to those of the traditional materials, but the main disadvantage of it is its long setting time. In addition, the complex of the constitution makes it difficult to prepare, which in turn lead to the high cost. Therefore, it is of important significance for the fundamental theory and clinical application to prepare the desensitizing materials and root canal filling materials with outstanding properties and low cost.
Since Hench invented 45S5 Bioglass? and put forward the concept of bioactivity, many scholars have been interested in CaO-SiO2 materials. Tricalcium silicate was one of the main components of Portland cement and MTA. In our earlier studies, phase pure Ca3SiO5 self-setting materials have been successfully prepared by sol-gel method, which were reported to not only have good bioactivity, degradability and compatibility, but also have self-setting properties. The hydration reaction of Ca3SiO5 leaded to the formation of calcium silicate hydrate(CSH) with size about 100nm, which maybe penetrate into dentinal tubules. The aim of this investigation was to study whether the bioactive tricalcium silicate paste could cover the dentine surface during the hydration and intimately bond to tooth, which would remineralize the tooth surface due to contact with oral fluids and significantly occlude the opening of dentinal tubules.
Meanwhile, the initial setting time of Ca3SiO5 seems too long to meet the need of clinical applications, while calcium chloride is one of the most effective accelerators of hydration and setting in Portland cement pastes. According to this, Ca3SiO5/CaCl2 composite cements were developed to examine the changes in mechanical properties, bioactivities and compatibility. In addition, the sealing ability and antimicrobial activity of the composite cements were also studied to explore the possibility for using it as a root canal filling material. The results are described as follows:
1. Ca3SiO5 paste as a desensitizing agent
The in vitro dentin hypersensitivity model was prepared and the tricalcium silicate paste was applied to the experimental surface of dentine samples with a cotton swab. Then, the samples were air-dried and the dentine surface was wiped with a dry swab after drying. This process was repeated 3 times before soaking the samples in 20ml artificial saliva at 37℃under continuous shaking. After soaking in the artificial saliva for 7 days, the SEM analysis showed that the dentine tubules were occluded completely, and the higher magnification SEM image showed that the particles were consist of round and strip-like nanocrystals with 100nm in diameters and 300nm in length, and many of these particles formed agglomerates, which were identified by XRD as hydroxyapatite (HA). EDS results showed that the ratio of Ca/P was 1.62, which was close to that for the intertubular dentine but higher than that for the peritubular dentine. In conclusion, tricalcium silicate could not only occlude the dentinal tubules by its self-setting process, but also induce the re-mineralization of the dentine in a physiological environment. It may be used for treatment of dentine hypersensitivity.
2. Preparation and properties of Ca3SiO5/CaCl2 composite cements
Composite cements were obtained by mixing Ca3SiO5 and different amount of CaCl2 (0, 5%, 10% and 15%) with deionized water as liquid phase. The purpose of this study was to investigate the effect of CaCl2 on the setting time, pH values and compressive strength of tricalcium silicate, and the in vitro bioactivity of Ca3SiO5/CaCl2 composite cements was also studied. The results showed that with the addition of CaCl2 from 0 to 15%, the initial setting time time decreased obviously from 90 to 50min and the final setting time from 180 to 85min. The compressive strength of the Ca3SiO5/CaCl2 composite cements after setting for 7 days increased obviously from 5.28 to 23.46MPa as the content of CaCl2 increased from 0 to 10%. It reached the maximum at the 10%CaCl2, and then decreased with the further increased of CaCl2. When the content of CaCl2 was same, the compressive strength of composite cement increased with prolonged time. The variation in the pH values for the Ca3SiO5/CaCl2 composite cements showed a similar trend with a rapid increase of the pH value in the first 3 hours followed by a rapid decrease in 5-8 hours, and stabilized thereafter. The higher the content of CaCl2 is, the lower is the maximum and minimum pH value. Furthermore, the paste with up to 15% of CaCl2 showed good ability to induce the formation of hydroxyapatite (HA).
3. Compatibility of Ca3SiO5/CaCl2 composite cement
The investigation of cell proliferation was conducted using extraction method with mouse fibroblast cell line L929 and osteoblast cell. The MTT method was used to assess the cell proliferation levels. Ca3SiO5/CaCl2 composite cements could release different ionic products in culture medium, and the ionic products at a certain concentration range could stimulate proliferation of L929 fibroblast cells and osteoblast cells. Ca2+, Si4+ ions play a main role in the stimulation of cell proliferation. Cl- ions may also contribute to the that. However, more detailed investigation is required to confirm the role of the Cl- ions.
4. Properties of Ca3SiO5/CaCl2 composite cement as root canal filling materials
4.1 Sealing ability of Ca3SiO5/CaCl2 composite cement Sealing ability was a very important index to evaluate the root canal filling materials. Inadequate obturation could make saliva or periodontal tissue liquid penetrating into the root canal, which became irritants after degradation leading to the periradicular inflammation. The objective of this study was to introduce a glucose fluid transport model for quantitative analysis of endodontic microleakage of Ca3SiO5/CaCl2 composite cement, ZOE and Cortisomol paste. Fifty single-rooted human maxillary anterior teeth were collected and divided into 3 experimental groups and 2 control groups randomly. The canals were prepared with the step-back technique and obtrurated with gutta-percha and 3 experimental materials. The glucose fluid transport model was used to test the glucose leakage after 1, 3, 5, 7, 14, 20 and 30days. The results showed that Ca3SiO5/CaCl2 composite cement and Cortisomol paste had a better sealing ability than ZOE(P<0.05).
4.2 Antibacterial ability of Ca3SiO5/CaCl2 composite cement The antibacterial ability of root canal filling materials was the important factor on evaluating quality of root canal treatment. Agar diffusion method was used to evaluate the antibacterial ability of Ca3SiO5/CaCl2 composite cement, ZOE and calcium hydroxide paste on the predominant bacteria of infected root canal, which included Mutans streptococci, Porphyromonas gingivalis, Prevotalla intermedius and Actinomyces viscosus. The results showed that Ca3SiO5/CaCl2 composite cement had no effect on Porphyromonas gingivalis, Prevotalla intermedius and Actinomyces viscosus. The effect of antibacterial on Mutans streptococci was Ca(OH)2 paste> ZOE> Ca3SiO5/CaCl2 composite cement ( P<0.05 ) . Therefore, Ca3SiO5/CaCl2 composite cement should be compounded with other materials with good antibacterial ability to use as root canal filling materials according to the optimum design rule in the next study.
In conclusion, the results of this study indicated that tricalcium silicate paste could not only occlude the dentinal tubules by its self-setting process, but also induce the re-mineralization of the dentine in physiological environment. It may be used for treatment of dentine hypersensitivity. Meanwhile, the setting speed and compressive strength of Ca3SiO5 have been improved by adding CaCl2, and Ca3SiO5/CaCl2 composite cements showed excellent bioactivity, cell compatibility and sealing ability as compared with traditional ZOE. These in vitro studies provided foundations for Ca3SiO5/CaCl2 composite cements using as root canal filling materials clinically. The innovation of this study was that tricalcium silicate paste was first applied to occlude the dentinal tubules and obtained good results, which may be used for treatment of dentine hypersensitivity. Moreover, Ca3SiO5/CaCl2 composite cement was first prepared and proved to possess outstanding properties, which may be used as root canal filling materials for clinical application in the future.
Dentin hypersensitivity, pulpitis and periradicular inflammation are common clinical conditions of stomatology, the therapy of which is closely related to the dental materials. At present, the materials of desensitizing and root canal filling have some shortcomings. Chemical and physical materials have been reported effective in reducing dentine hypersensitivity. However, these materials were not components of natural dentine, and may diminish during teeth brushing and food chewing. An ideal root canal filling material should have several properties, such as being harmless to the periapical tissues and permanent tooth germs; resorbing readily if pressed beyond the apex; easily filling the root canals; adhering to the walls of the canal; not being susceptible to shrinkage; being easily removed if necessary; being radiopaque and not discolouring the tooth. At present, none of root canal filling materials is able to meet all the above requirements. Zinc oxide eugenol cement (ZOE) was the first root canal filling material to be recommended for permanent teeth, as described by Sweet in 1930. However, later studies showed that this cement has a slow rate of resorption in the canals, limited antibacterial action and cytotoxic effects possibly causing by the release of eugenol and formaldehyde. In recent years, the introduction of calcium hydroxide in dental therapy has received much attention for its biological activity. It is antibacterial and anti-inflammatory primarily due to the high pH value of the surrounding environment (approximately 12.5) after its dissolution. In addition, calcium hydroxide can induce mineralization resulting in a repair of damaged dentin matrix. Although calcium hydroxide has some advantages compared with ZnO eugenol cement and has already been used in the root canal filling system, it has also shown disadvantages such as the low hardening rate and high dissolution, which resulting in a loose filling. So it can only be used as temporary filling materials in indications for which hermeticity is a priority. A new class of restorative material called mineral trioxide aggregate (MTA) was recently introduced as dental materials. In vitro and in vivo studies have demonstrated that the bioactivity and biocompatibility of MTA are superior to those of the traditional materials, but the main disadvantage of it is its long setting time. In addition, the complex of the constitution makes it difficult to prepare, which in turn lead to the high cost. Therefore, it is of important significance for the fundamental theory and clinical application to prepare the desensitizing materials and root canal filling materials with outstanding properties and low cost.
Since Hench invented 45S5 Bioglass? and put forward the concept of bioactivity, many scholars have been interested in CaO-SiO2 materials. Tricalcium silicate was one of the main components of Portland cement and MTA. In our earlier studies, phase pure Ca3SiO5 self-setting materials have been successfully prepared by sol-gel method, which were reported to not only have good bioactivity, degradability and compatibility, but also have self-setting properties. The hydration reaction of Ca3SiO5 leaded to the formation of calcium silicate hydrate(CSH) with size about 100nm, which maybe penetrate into dentinal tubules. The aim of this investigation was to study whether the bioactive tricalcium silicate paste could cover the dentine surface during the hydration and intimately bond to tooth, which would remineralize the tooth surface due to contact with oral fluids and significantly occlude the opening of dentinal tubules.
Meanwhile, the initial setting time of Ca3SiO5 seems too long to meet the need of clinical applications, while calcium chloride is one of the most effective accelerators of hydration and setting in Portland cement pastes. According to this, Ca3SiO5/CaCl2 composite cements were developed to examine the changes in mechanical properties, bioactivities and compatibility. In addition, the sealing ability and antimicrobial activity of the composite cements were also studied to explore the possibility for using it as a root canal filling material. The results are described as follows:
1. Ca3SiO5 paste as a desensitizing agent
The in vitro dentin hypersensitivity model was prepared and the tricalcium silicate paste was applied to the experimental surface of dentine samples with a cotton swab. Then, the samples were air-dried and the dentine surface was wiped with a dry swab after drying. This process was repeated 3 times before soaking the samples in 20ml artificial saliva at 37℃under continuous shaking. After soaking in the artificial saliva for 7 days, the SEM analysis showed that the dentine tubules were occluded completely, and the higher magnification SEM image showed that the particles were consist of round and strip-like nanocrystals with 100nm in diameters and 300nm in length, and many of these particles formed agglomerates, which were identified by XRD as hydroxyapatite (HA). EDS results showed that the ratio of Ca/P was 1.62, which was close to that for the intertubular dentine but higher than that for the peritubular dentine. In conclusion, tricalcium silicate could not only occlude the dentinal tubules by its self-setting process, but also induce the re-mineralization of the dentine in a physiological environment. It may be used for treatment of dentine hypersensitivity.
2. Preparation and properties of Ca3SiO5/CaCl2 composite cements
Composite cements were obtained by mixing Ca3SiO5 and different amount of CaCl2 (0, 5%, 10% and 15%) with deionized water as liquid phase. The purpose of this study was to investigate the effect of CaCl2 on the setting time, pH values and compressive strength of tricalcium silicate, and the in vitro bioactivity of Ca3SiO5/CaCl2 composite cements was also studied. The results showed that with the addition of CaCl2 from 0 to 15%, the initial setting time time decreased obviously from 90 to 50min and the final setting time from 180 to 85min. The compressive strength of the Ca3SiO5/CaCl2 composite cements after setting for 7 days increased obviously from 5.28 to 23.46MPa as the content of CaCl2 increased from 0 to 10%. It reached the maximum at the 10%CaCl2, and then decreased with the further increased of CaCl2. When the content of CaCl2 was same, the compressive strength of composite cement increased with prolonged time. The variation in the pH values for the Ca3SiO5/CaCl2 composite cements showed a similar trend with a rapid increase of the pH value in the first 3 hours followed by a rapid decrease in 5-8 hours, and stabilized thereafter. The higher the content of CaCl2 is, the lower is the maximum and minimum pH value. Furthermore, the paste with up to 15% of CaCl2 showed good ability to induce the formation of hydroxyapatite (HA).
3. Compatibility of Ca3SiO5/CaCl2 composite cement
The investigation of cell proliferation was conducted using extraction method with mouse fibroblast cell line L929 and osteoblast cell. The MTT method was used to assess the cell proliferation levels. Ca3SiO5/CaCl2 composite cements could release different ionic products in culture medium, and the ionic products at a certain concentration range could stimulate proliferation of L929 fibroblast cells and osteoblast cells. Ca2+, Si4+ ions play a main role in the stimulation of cell proliferation. Cl- ions may also contribute to the that. However, more detailed investigation is required to confirm the role of the Cl- ions.
4. Properties of Ca3SiO5/CaCl2 composite cement as root canal filling materials
4.1 Sealing ability of Ca3SiO5/CaCl2 composite cement Sealing ability was a very important index to evaluate the root canal filling materials. Inadequate obturation could make saliva or periodontal tissue liquid penetrating into the root canal, which became irritants after degradation leading to the periradicular inflammation. The objective of this study was to introduce a glucose fluid transport model for quantitative analysis of endodontic microleakage of Ca3SiO5/CaCl2 composite cement, ZOE and Cortisomol paste. Fifty single-rooted human maxillary anterior teeth were collected and divided into 3 experimental groups and 2 control groups randomly. The canals were prepared with the step-back technique and obtrurated with gutta-percha and 3 experimental materials. The glucose fluid transport model was used to test the glucose leakage after 1, 3, 5, 7, 14, 20 and 30days. The results showed that Ca3SiO5/CaCl2 composite cement and Cortisomol paste had a better sealing ability than ZOE(P<0.05).
4.2 Antibacterial ability of Ca3SiO5/CaCl2 composite cement The antibacterial ability of root canal filling materials was the important factor on evaluating quality of root canal treatment. Agar diffusion method was used to evaluate the antibacterial ability of Ca3SiO5/CaCl2 composite cement, ZOE and calcium hydroxide paste on the predominant bacteria of infected root canal, which included Mutans streptococci, Porphyromonas gingivalis, Prevotalla intermedius and Actinomyces viscosus. The results showed that Ca3SiO5/CaCl2 composite cement had no effect on Porphyromonas gingivalis, Prevotalla intermedius and Actinomyces viscosus. The effect of antibacterial on Mutans streptococci was Ca(OH)2 paste> ZOE> Ca3SiO5/CaCl2 composite cement ( P<0.05 ) . Therefore, Ca3SiO5/CaCl2 composite cement should be compounded with other materials with good antibacterial ability to use as root canal filling materials according to the optimum design rule in the next study.
In conclusion, the results of this study indicated that tricalcium silicate paste could not only occlude the dentinal tubules by its self-setting process, but also induce the re-mineralization of the dentine in physiological environment. It may be used for treatment of dentine hypersensitivity. Meanwhile, the setting speed and compressive strength of Ca3SiO5 have been improved by adding CaCl2, and Ca3SiO5/CaCl2 composite cements showed excellent bioactivity, cell compatibility and sealing ability as compared with traditional ZOE. These in vitro studies provided foundations for Ca3SiO5/CaCl2 composite cements using as root canal filling materials clinically. The innovation of this study was that tricalcium silicate paste was first applied to occlude the dentinal tubules and obtained good results, which may be used for treatment of dentine hypersensitivity. Moreover, Ca3SiO5/CaCl2 composite cement was first prepared and proved to possess outstanding properties, which may be used as root canal filling materials for clinical application in the future.
引文
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