复合HRZ/PLGA缓释抗结核药涂层材料在兔脊柱结核病灶释药研究

Drug-release properties of sustained-release of anti-tuberculosis drugs composite HRZ/PLGA bone graft material in spinal tuberculosis in rabbits

目的 :观察复合HRZ/PLGA缓释抗结核药涂层材料在兔脊柱结核病灶的释药特性。方法 :应用兔脊柱结核模型120只,按三联抗痨药异烟肼(INH,H)、利福平(RFP,R)和吡嗪酰胺(PZA,Z)给药剂型及途径不同,随机分为4组(各30只):A组(复合HRZ/PLGA缓释材料局部给药组);B组(局部给药组);C组(灌胃给药组);D组(假手术灌药组)。其中A、B、C三组行L4/5脊柱结核病灶清除及自体髂骨植骨术,D组行假手术。采用高效液相色谱(HPLC)检测术后3d、7d、14d、28d、56d、84d各组病灶骨组织中H、R及Z的浓度,绘制药物浓度-时间曲线;使用DAS 3.2.1统计软件对药动学参数进行分析。结果:A组病灶骨组织中H、R及Z的浓度均在术后3d时达最高,此后随检测时间点的后移三种药物浓度均逐渐下降,至给药后84d时最低(H为6.69±1.42μg/ml;R为6.28±0.77μg/ml;Z为19.88±0.90μg/ml),三种药物浓度在各检测时间点均≥10倍的MIC(即杀菌浓度),对同种药物在不同时间检测的浓度进行两两比较,均有统计学意义(P<0.05);B组病灶骨组织中H、R及Z的浓度在术后3d时均明显增高,此后较快衰减,至给药后14d时三种药物浓度均检测不到,对同种药物在3d与7d检测的浓度进行比较,差异有统计学意义(P<0.05);C组及D组随检测时间点的后移,病灶骨组织中H、R及Z的药物浓度均维持在相对平稳的较低水平,对同组中同种药物在不同时间检测的浓度进行两两比较均无统计学意义(P>0.05),术后84d检测时C组H为3.21±0.32μg/ml、R为3.68±0.42μg/ml、Z为6.68±0.25μg/ml,D组H为3.24±0.33μg/ml、R为3.53±0.44μg/ml、Z为6.39±0.45μg/ml。对A、B、C、D四组同时间点同种药物浓度进行比较:C、D两组间同时间点同种药物浓度比较均无统计学差异(P>0.05);A组病灶骨组织中H、R及Z的浓度均高于C组及D组的浓度,84d检测时A组中的H、R及Z的浓度分别为C组的3.02、1.70及2.97倍,差异有统计学意义(P<0.05);A、C、D三组药物浓度-时间曲线平缓,均未见突释现象;B组存在明显突释现象,药物浓度衰减较快,术后14d后检测不到H、R及Z的浓度。药动学数据分析:A组的曲线下面积、生物半衰期均显著高于B组及C组,差异有统计学意义(P<0.05)。结论:复合HRZ/PLGA缓释抗结核药涂层材料在兔脊柱结核病灶局部可实现三药同时长程、高效、平缓释药。

Objectives： To observe the drug-release properties of sustained-release of anti-tuberculosis drugs and bone graft complex in spinal tuberculosis in rabbits. Methods： Spinal tuberculosis model was introduced in 120 rabbits. The models were randomly divided into sustained-release material group（group A）, local ad-ministration group（group B）, oral gavage group（group C） and sham operation group（group D） based on the dosage and administration of isoniazid（INH, H）, rifampicin（RFP, R） and pyrazinamide（PZA, Z）. Among them,the rabbits of group A, B and C underwent L4/5 spinal tuberculosis debridement and autologous iliac bonegraft surgery, group D underwent sham surgery. The concentration of H, R and Z in the focal bone tissue was detected by the method of high performance liquid chromatography（HPLC） on the 3rd, 7th, 14 th, 28 th,56th and 84 th day after surgery. Then the drug concentration-time curve was drawn, followed by analyzing the pharmacokinetic parameters by DAS software 3.2.1. Results： The highest concentration of H, R and Z in the focal bone tissue was detected on the 3rd day in group A, thereafter the concentration decreased gradually over time. By the 84 th day, the concentration reached the lowest point（H, 6.69±1.42μg/ml; R, 6.28±0.77μg/ml; Z, 19.88 ±0.90μg/ml）. The concentration of all the three drugs was 10 times higher than MIC in each detecting time point, which was known as minimal inhibitory concentration. Comparing the concentration of each drug at different detecting time points, a significant level was set at P〈0.05. In group B, the detect-ing concentration of H, R and Z in the focal bone tissue significantly increased on the 3rd day, then dropped sharply over time, which became undetectable on the 14th day. While the difference between concentration of each drug on the 3rd day and that on the 7th day was statistically significant （P〈0.05）. The concentration of H, R and Z in the focal bone tissue was kept in a relatively stable low level over time in group C and group D. In group C, on the 84th day after surgery, the concentration of H was 3.21＋0.32μg/ml, R was 3.68_＋0.42μg/ml, and Z was 6.68_＋0.25μg/ml. In group D, on the 84th day after surgery, the concentration of H was 3.24±0.33μg/ml, R was 3.53±0.44μg/ml, and Z was 6.39±0.45μg/ml. Pairwise comparisons of concen-tration of each drug detected at different time points did not show statistically significant difference （P〉0.05）. Comparing the concentration of each drug at the same time point among group A, B, C and D, there was no significant difference in the same time between group C and D （P〉0.05）. The drug concentration-time curves in group A, C and D were flat, without burst release, and the concentration of H, R and Z in group A was higher than that in group C and group D. On the 84th day, the concentration of H, R and Z in group A was 3.02, 1.7 and 2.97 times as high as that in group C, respectively, and a significant level was set at P〈 0.05. In group B, there was a significant burst release phenomenon. The drug concentration dropped sharply, and on the 14th day, there was no H, R or Z detected. Pharmacokinetic data analysis： the area under the curve, the biological haft-life in group A were significantly higher than those in group B and group C, and the difference was statistically significant（P〈0.05）. Conclusions： The three drugs in slow-releasing anti- tuberculosis drugs hone graft complex is sustained, efficient and gentle release locally in the model of spinal tuberculosis in rabbits.