透明质酸钠对兔创伤性骨关节炎软骨诱导型一氧化氮合酶mRNA表达的干预(英文)

Interventional effect of sodium hyaluronate on inducible nitric oxide synthase mRNA expression in cartilage of rabbits with traumatic osteoarthritis

背景:各种关节损伤、关节手术后导致的创伤性骨关节炎较为常见,关节腔注射透明质酸钠已被视为一种治疗骨关节炎的有效手段。目的:以前交叉韧带切断的方法建立兔创伤性骨关节炎模型,观察关节腔注射透明质酸钠对其关节软骨中诱导型一氧化氮合酶mRNA表达水平的影响。设计:随机对照动物实验。单位:武汉大学人民医院骨科。材料:实验于2003-04/12在武汉大学人民医院骨科实验室完成。选取清洁级5～6月龄大耳白兔16只,随机数字表法分为透明质酸钠注射组、生理盐水对照组,8只/组。透明质酸钠(上海建华精细生物制品有限公司提供,国药准字2000第366095号)。方法:①两组均建立创伤性骨关节炎模型。每只兔以氯胺酮1.0mg/kg体质量麻醉,单侧膝关节行前交叉韧带切断造模。②术后第5周,透明质酸钠注射组患膝关节腔注射质量浓度为10g/L的透明质酸钠0.3mL,1次/周,连续5周。生理盐水对照组注射等量生理盐水。③术后10周处死,观察两组股骨内髁关节软骨的大体形态学(0分:关节面光滑,色泽正常;1分:关节面粗糙,有小的裂隙,色泽灰暗;2分:关节面糜烂,软骨缺损达软骨表层或中层;3分:关节面溃疡形成,缺损达软骨深层;4分:软骨剥脱,软骨下骨质暴露)和组织学病理变化,采用反转录聚合酶链反应方法检测软骨诱导型一氧化氮合酶mRNA的表达情况。主要观察指标:①两组股骨髁关节面标本大体观察结果。②两组股骨内髁软骨光镜组织学观察结果。③两组软骨诱导型一氧化氮合酶mRNA的表达。结果:实验选取清洁级大耳白兔16只,全部进入结果分析。①两组股骨髁关节面标本大体观察结果:解剖显微镜下观察股骨髁关节面病理变化,透明质酸钠注射组软骨退变程度较生理盐水对照组明显减轻。②两组股骨内髁软骨光镜组织学观察结果:透明质酸钠注射组见软骨膜变性脱落,表层软骨细胞变性、坏死、排列紊乱,形成糜烂;生理盐水对照组见软骨细胞变性、坏死,排列紊乱,溃疡病变达软骨深层,并见新生的毛细血管和成纤维细胞增生,溃疡底部见纤维组织增生。③两组软骨诱导型一氧化氮合酶mRNA的表达:透明质酸钠注射组软骨诱导型一氧化氮合酶mRNA表达量均值为1.09±0.18,生理盐水对照组的表达量均值为1.26±0.23,两组基本相似(P>0.05)。结论:关节腔注射透明质酸钠对早期骨关节炎软骨具有修复和保护作用,能有效减轻早期创伤性骨关节炎关节软骨的退变,对诱导型一氧化氮合酶的表达没有下调作用。

BACKGROUND： Traumatic osteoarthritis （0A） resulted from the injury of joints and postoperation of joints is commonly observed, lntra-articular injection of sodium hyaluronate （Na-HA） has been considered as effective method for OA. OBJECTIVE： To observe the influence of intra-articular injection of Na-HA on mRNA expression of inducible nitric oxide synthase （iNOS） in eartilage of traumatic OA induced by transection of anterior cruciate ligament. DESIGN： Randomized controlled animal experiment. SETTING： Department of Orthopaedics, Renmin Hospital, Wuhan U ni versity. MATERLALS： The experiment was performed in Laboratory of Department of Orthopaedics, Renmin Hospital, Wuhan University from April to December 2003, in which, 16 clean healthy flat-eared white rabbits, aged 5-6 months were employed. The rabbits were randomly divided into Na-HA injection group and saline control group with 8 rabbits in each group. Na-HA （2000, No 366095） was provided by Shanghai Jiahua Fine Biological Products Co. METHODS： （1）OA model was established in rabbits of the two groups. Each rabbit was anesthetized intravenously with 1.0 mg/kg ketamine hydrochloride and underwent unilateral anterior cruciate ligament transection. （3）5 weeks after transection, Na-HA injection group rabbits received 0.3 mL of intra-articular 10 g/L Na-HA injection, once a week for 5 weeks. Animals in saline controlled group were treated with saline of the same volume. （3）The rabbits were killed at week 10 after operation, general morphology and histopathological changes of articular cartilage degeneration of medial femoral condyle were evaluated （0 points as smooth articular surface with normal color and luster; 1 point as minimal fibrillation or a slight crevice and dark grey color of the surface; 2 points as erosion extending into superficial or middle layers of cartilage, 3 points as ulceration and erosion extending into the deep layers, and 4 points as denudation of cartilage, erosion extending to the sub-chondral bone）. The mRNA expression of iNOS in cartilages was examined with reverse transcription-polymerase chain reaction （RT-PCR）. MAIN OUTCOME MEASURES： （1）Observation of articular cartilage degeneration of medial femoral condyle, （2）observation of articular cartilage degeneration of medial femoral condyle at light microscopic level, （3）expression of iNOS in cartilages of each group. RESULTS： A total of 16 clean healthy rabbits entered the result analysis. （1） Result of articular cartilage degeneration of medial femoral condyle： Pathological change of articular surface of femoral condyle was observed under anatomic microscope. Cartilage degradation in experimental group was significantly less severe than that in saline control group. （2）Histological changes of articular cartilage degeneration of medial femoral condyle at light microscopic level： The Na-HA injection group showed cartilage changes： Membrane of cartilage presented denaturalization and abscission. Chondrocytes of superficial zone presented denaturalization, necrosis, turbulence and erosion. Animals treated with saline showed denaturalization, necrosis and disorder of chondrocytes, ulceration penetrating into the middie or deep zone of the cartilage. New hyperplasia of capillary vessels and fibroblasts were more obvious. Proliferation of fibrous tissue appeared at the bottom of ulcer. （3）Expression of iNOS in cartilages of two group： The gene expression of iNOS in cartilage of Na-HA injection group was （1.09±0.18） and the expression of saline control group was （1.26±0.23）. No significant difference of iNOS expression was found between the two groups （P 〉 0.05）. CONCLUSION： lntra-articular injection of Na-HA has protective and repairing effect on cartilage with early OA and can significantly reduce the severity of cartilage degradation during early stage of traumatic OA. Intraarticular injection of Na-HA does not down-regulated iNOS expression in cartilage