牙周缓释凝胶治疗牙周炎的实验研究
摘要
牙周炎是口腔临床上常见的疾病之一,是中老年人失牙的主要原因,到目前为止,尚无有效的治疗方法。它是一种病因复杂的炎症破坏性疾病,由牙周袋内革兰氏阴性厌氧菌引起,已知硝基咪唑类和四环素类抗菌药物对其有较强的杀灭作用,但由于药物浓度持续时间短、不良反应大,疗效并不理想。而缓释制剂能较好的解决这些问题,是目前牙周炎治疗的研究方向之一。
牙周缓释制剂有两个关键因素,有效药物和缓释辅料。多西环素除了具有较强的厌氧菌杀灭作用外,还能通过多种途径抑制MMPs的活性,减轻结缔组织的破坏,达到牙周病的阻断治疗。rhBMP-2具有骨诱导活性,可以促进牙周韧带细胞(PDLC)向成骨细胞表型分化,促进牙槽骨和牙骨质的再生,并能诱导牙周膜的生长。PLGA是可完全降解吸收的高分子材料,随着材料的缓慢崩解,复合在载体材料上的药物缓慢释放,起到持续作用的目的,根据乙交酯(LA)与丙交酯(GA)的配比不同和分子量的大小,PLGA崩解的速度也不同,是目前的研究热门。
作者用PLGA作缓释辅料,以多西环素和rhBMP-2作有效治疗药物,以1MP和GTA作混合溶剂制成凝胶,测定凝胶的性能和凝结时间,并作体外释放试验,筛选出最佳配方,并以动物试验验证该制剂的有效性。
本研究分成两部分:
第一部分 牙周缓释凝胶的研制及体外释放度试验
Periodontitis is one of the familiar dental diseases and it is a main cause which leads to tooth loss .There is no efficient method to cure it at present. it is a complicated inflammation which is caused by gram-negative anaerobic bacteria . It has been known that tinidazole and tetracycline can kill these bacterias , but the effect is not very good owing to the short sustained time and large side effect . The sustained-release formula can solve these problems and it is a pathway of periodontitis' s studying.Sustained-release gel have two key elements : efficient drug and sustained-release meterial. Besides the effect of killing bacteria, doxycycline can also inhibit the activity of matrix metalloproteinase(MMPs) by many ways such as reducing the breakage of conjunctive tissue and stoping the progress of periodontitis. Recombinant human bone morphogenetic protein-2(rhBMP-2) has bone inductive activity and can promote periodontal ligament cell(PDLC)to be differentiated into osteoblast. It can also promote the regeneration of alveolar bone and cementum which can induce the growth of parodontal membrance . Poly lactic and glycollic acid(PLGA) is a kind of biodegradable material . The efficient drug which was mixed in the gel will release slowly alone with the degradation of PLGA , so it can reach the aid of continuous action . Because of the different molecular weight and different proportion between lactic acid (LA) and glycollic acid(GA) , the degradation speed of PLGA is also different .
The gel was made using PLGA as sustained material, including doxycycline and rhBMP-2 as treating drugs. The gel' s performance and coagulate time were measured. The best formula and the efficient of sustained-release gel were tested .This study consists of two sectionsSection one preparation of sustained-release gel and in vitro testThe aid of this study is to develop a good sustained-release gel which can be used in practice , it should accord the following characteristics:1. PLGA will become gel when it is dissolved in mixed solvent and it has the characteristic of fluidity and stickiness.2. The gel coagulate within 10-15 seconds after touching water.3. The gel dissolve within 7-10 days, the drug mixed in the gel release slowly alone with the breakdown of gel.4. rhBMP-2 which was mixed in the gel has biologic activity.In order to reach these aids ,this study use PLGA75:25 ( average molecular weight is below 10 thousand)and PLGA80:20(average molecular weight is near 20 thousand) as the sustained-release material.The concentration of PLGA75:25 are 25w/w% , 30w/w% & 35w/w% separately, the concertration of PLGA80:20 are 9w/w% , 12w/w% & 15w/w% separately , the mixed solvent consist of 2 kinds of solvent: IMP and GTA,the proportion of the 2 solvent is 6:4 ,7:3 ,8:2 separately , and the concentration of doxcycline is 6w/w% ,9w/w% & 12w/w% prepare 18 kinds of gel following the method of orthogonal design .Measure the concentration of doxycycline in water and PBS at the different times , filter out the best formula according with its releasing.
The result showed: for the PLGA80:20 , the gel don' t coagulate when the concentration of PLGA below 4w/w% .According to the orthogonal score , we get the best formula:concentration of PLGA80:20 is 14w/w% , 1MP:GTA=7:3 , concentration of doxycycline is 6w/w%.For the PLGA 75:25, the gel don' t coagulate when the concentration of PLGA below 10w/w% . According to the orthogonal score , we get the best formula : concentration of PLGA is 30w/w% ,1MP:GTA=7:3 , the concentration of doxycycline is 6w/w% .In addition , we found that the gel' s stickiness of PLGA75:25 is better than PLGA80:20' s.Integrate all these charactristics,we choose PLGA75:25 as the best formula . We add the rhBMP-2 to the gel .validate its activity by animal test.Section two Effect of sustained-release gel on experimental periodontitis of ratsIn order to validate the effect of sustained-release gel on periodontitis, the study do the reserch on periodontitis animal model of rats. We firstly created experimental animal mod
牙周缓释制剂有两个关键因素,有效药物和缓释辅料。多西环素除了具有较强的厌氧菌杀灭作用外,还能通过多种途径抑制MMPs的活性,减轻结缔组织的破坏,达到牙周病的阻断治疗。rhBMP-2具有骨诱导活性,可以促进牙周韧带细胞(PDLC)向成骨细胞表型分化,促进牙槽骨和牙骨质的再生,并能诱导牙周膜的生长。PLGA是可完全降解吸收的高分子材料,随着材料的缓慢崩解,复合在载体材料上的药物缓慢释放,起到持续作用的目的,根据乙交酯(LA)与丙交酯(GA)的配比不同和分子量的大小,PLGA崩解的速度也不同,是目前的研究热门。
作者用PLGA作缓释辅料,以多西环素和rhBMP-2作有效治疗药物,以1MP和GTA作混合溶剂制成凝胶,测定凝胶的性能和凝结时间,并作体外释放试验,筛选出最佳配方,并以动物试验验证该制剂的有效性。
本研究分成两部分:
第一部分 牙周缓释凝胶的研制及体外释放度试验
Periodontitis is one of the familiar dental diseases and it is a main cause which leads to tooth loss .There is no efficient method to cure it at present. it is a complicated inflammation which is caused by gram-negative anaerobic bacteria . It has been known that tinidazole and tetracycline can kill these bacterias , but the effect is not very good owing to the short sustained time and large side effect . The sustained-release formula can solve these problems and it is a pathway of periodontitis' s studying.Sustained-release gel have two key elements : efficient drug and sustained-release meterial. Besides the effect of killing bacteria, doxycycline can also inhibit the activity of matrix metalloproteinase(MMPs) by many ways such as reducing the breakage of conjunctive tissue and stoping the progress of periodontitis. Recombinant human bone morphogenetic protein-2(rhBMP-2) has bone inductive activity and can promote periodontal ligament cell(PDLC)to be differentiated into osteoblast. It can also promote the regeneration of alveolar bone and cementum which can induce the growth of parodontal membrance . Poly lactic and glycollic acid(PLGA) is a kind of biodegradable material . The efficient drug which was mixed in the gel will release slowly alone with the degradation of PLGA , so it can reach the aid of continuous action . Because of the different molecular weight and different proportion between lactic acid (LA) and glycollic acid(GA) , the degradation speed of PLGA is also different .
The gel was made using PLGA as sustained material, including doxycycline and rhBMP-2 as treating drugs. The gel' s performance and coagulate time were measured. The best formula and the efficient of sustained-release gel were tested .This study consists of two sectionsSection one preparation of sustained-release gel and in vitro testThe aid of this study is to develop a good sustained-release gel which can be used in practice , it should accord the following characteristics:1. PLGA will become gel when it is dissolved in mixed solvent and it has the characteristic of fluidity and stickiness.2. The gel coagulate within 10-15 seconds after touching water.3. The gel dissolve within 7-10 days, the drug mixed in the gel release slowly alone with the breakdown of gel.4. rhBMP-2 which was mixed in the gel has biologic activity.In order to reach these aids ,this study use PLGA75:25 ( average molecular weight is below 10 thousand)and PLGA80:20(average molecular weight is near 20 thousand) as the sustained-release material.The concentration of PLGA75:25 are 25w/w% , 30w/w% & 35w/w% separately, the concertration of PLGA80:20 are 9w/w% , 12w/w% & 15w/w% separately , the mixed solvent consist of 2 kinds of solvent: IMP and GTA,the proportion of the 2 solvent is 6:4 ,7:3 ,8:2 separately , and the concentration of doxcycline is 6w/w% ,9w/w% & 12w/w% prepare 18 kinds of gel following the method of orthogonal design .Measure the concentration of doxycycline in water and PBS at the different times , filter out the best formula according with its releasing.
The result showed: for the PLGA80:20 , the gel don' t coagulate when the concentration of PLGA below 4w/w% .According to the orthogonal score , we get the best formula:concentration of PLGA80:20 is 14w/w% , 1MP:GTA=7:3 , concentration of doxycycline is 6w/w%.For the PLGA 75:25, the gel don' t coagulate when the concentration of PLGA below 10w/w% . According to the orthogonal score , we get the best formula : concentration of PLGA is 30w/w% ,1MP:GTA=7:3 , the concentration of doxycycline is 6w/w% .In addition , we found that the gel' s stickiness of PLGA75:25 is better than PLGA80:20' s.Integrate all these charactristics,we choose PLGA75:25 as the best formula . We add the rhBMP-2 to the gel .validate its activity by animal test.Section two Effect of sustained-release gel on experimental periodontitis of ratsIn order to validate the effect of sustained-release gel on periodontitis, the study do the reserch on periodontitis animal model of rats. We firstly created experimental animal mod
引文
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