阿仑膦酸钠对兔膝骨关节炎关节软骨及软骨下骨保护的作用

Protective effect of alendronate on articular cartilage and subchondral bone in osteoarthritis rabbit model

目的研究阿仑膦酸钠(ALN)对兔膝骨关节炎关节软骨及软骨下骨的保护作用。方法健康3月龄雄性日本大耳白兔30只,随机分为对照组(Sham)、盐水组(ACLT+NS)和ALN(ACLT+ALN),每组10只。盐水组和ALN行右侧膝关节前交叉韧带切断术(ACLT)造膝骨关节炎模型。术后ACLT+ALN组皮下注射ALN20μg·kg-1·2 d-1,连续6 w;ACLT+NS组则给予等剂量盐水,给药后6 w处死动物,在处死前10d和4d分别皮下注射盐酸四环素和钙黄绿素行荧光双标记。取右侧膝胫骨近端包埋制成硬组织切片,用于软骨下骨小梁形态计量学测量;取股骨远端行骨密度测量,脱钙制成切片,行HE染色和Mankin评分,免疫组化染色切片观察MMP-13表达。结果①Mankin评分:ACLT+NS组分数显著高于Sham组和ACLT+ALN组(P<0.05)。②股骨远端骨密度:ACLT+ALN组均显著高于ACLT+NS组。③骨组织形态计量学结果:ACLT+ALN组骨小梁相对体积、厚度、数量等均高于ACLT+NS组(P<0.05);而ACLT+ALN组骨小梁分离度低于ACLT+NS组(P<0.05)。④免疫组化:ACLT+ALN组和Sham组MMP-13的表达显著低于ACLT+NS组。结论皮下注射ALN 20μg·kg-1·2d-1能够保护关节软骨,维持骨量,改善软骨下骨小梁微观结构。

Objective To access the effect of alendronate（ALN） on cartilage and subchondral bone in osteoarthritis rabbit induced by anterior cruciate ligament transaction（ACLT）.Methods Thirty 3-month-old male Japanese white rabbits were randomly divided into sham,ACLT＋NS,ACLT＋ALN groups.Rabbits in ACLT＋ALN and ACLT＋NS groups were undergone ACLT on the right knee.The rabbits of sham group were undergone sham operation.Every rabbit was placed in a cage（60 cm×60 cm×80 cm） without any further immobilization.Then the rabbits of ACLT ＋ ALN group were given subcutaneous injection of ALN at a dosage of 20 μg·kg-1·2 d-1 of body weight for 6 weeks.In contrast,ACLT ＋NS group was given NS at the same dose.All rabbits were sacrificed at 6 weeks post surgery,and injected with Tetracycline（30 mg/kg） and Calcein（10 mg/kg） 10 days and 4 days before sacrificed respectively.The macro-pathologic changes were observed and stained by HE and graded by Mankin＇s scale.The bone mineral density（BMD） was measured,the level of MMP-13 was detected by immunohistochemistry.The bone histomorphometry analysis was done in subchondral bone of proximal tibia.Results Mankin＇s scores in ACLT＋ALN and sham groups were significantly lower than that in ACLT＋ NS group（P0.05）.The BMD of distal femora was significantly higher than that in ACLT＋NS group（P0.05）.Compared with that of ACLT＋NS group,the rabbits in ACLT＋ALN group significantly increased the trabecular bone relative volume（BV/TV）,trabecular bone thickness（Tb.Th） trabecular bone number（Tb.N）,and significantly decreased trabecular bone separation（Tb.Sp）（P0.05）.Immunohistochemistry staining showed that the expression of MMP-13 in ACLT＋ALN group was significantly lower than that in ACLT＋NS group（P0.05）.Conclusions ALN could inhibit the development of osteoarthritis from modulating subchondral bone metabolism and thereby preventing bone loss and improving microarchitecture of subchondral bone.