一氧化氮外源性供体L-arginine对破骨细胞骨吸收功能的影响  

Effect of L-arginine as an exogenous donator of nitrogen monoxidum on osteoclast resorptive activity

作　　者：镐英杰[1] 王珍[2] 李秀群[2] 裴福兴[3] 

机构地区：[1]郑州大学第一附属医院骨科,河南省郑州市410052 [2]四川大学华西医院,生物治疗国家重点实验室,干细胞与组织工程研究室,四川省成都市610041 [3]四川大学华西医院骨科

出　　处：《中国组织工程研究与临床康复》2007年第45期9031-9034,共4页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金资助项目(30571871)~~

摘　　要：目的:一氧化氮在维持机体多个系统的生理功能中起重要作用,许多慢性疾病可造成一氧化氮产生减少,此时一氧化氮供体是一种必要的补充。观察一氧化氮外源性供体L-arginine对体外培养破骨细胞增殖及骨吸收功能的影响。方法:实验于2005-06/2006-05在四川大学华西医院生物治疗国家重点实验室干细胞与组织工程研究室完成。选择出生1d的清洁级SD大鼠乳鼠,采用骨髓诱导法体外培养破骨细胞,培养液内分别加入0.3,0.6,1.0g/L不同浓度的L-arginine,并以等体积三蒸水作为对照。培养7d后,以抗酒石酸酸性磷酸酶染色观察破骨细胞数目、形态,MIAS-2000图像分析仪检测骨片上骨吸收陷窝的数目和面积,并用扫描电镜观察不同浓度L-arginine对骨吸收陷窝的影响。结果:①破骨细胞的一般形态:破骨细胞较其他细胞大,形态不规则,呈油煎蛋形、长条形、腊肠形或漏斗形等,细胞内可见几个至几十个核不等。抗酒石酸酸性磷酸酶染色酶活性部分酒红色,颗粒状。②抗酒石酸酸性磷酸酶阳性细胞数:各组破骨细胞数目随着L-arginine浓度增加而减少(P<0.05)。③骨吸收陷窝的面积和数目:骨片培养7d,吸收陷窝计数的结果显示,0.3g/L以上浓度L-arginine对破骨细胞吸收功能均有明显抑制作用,并呈剂量相关性。0.3g/LL-arginine组陷窝面积为对照组的91%(P<0.05),0.6g/LL-arginine组为对照组的80%(P<0.05),1.0g/LL-arginine组为对照组的69%(P<0.01)。结论:采用骨髓诱导法培养的破骨细胞数量多、纯度高,且具有明显的骨吸收功能。L-arginine抑制破骨细胞增殖,抑制破骨细胞骨吸收功能,并呈剂量相关性。AIM： Nitrogen monoxidum plays an important role in maintaining physiological function of multiple systems, but many chronic diseases can cause reduction of nitrogen monoxidum, so nitrogen monoxidum supplement is necessary. This study was aimed to observe the effects of L-arginine on osteoclast proliferation differentiation and resorptive activity. METHODS： The experiment was carried out at Division of Stem Cells and Tissue Engineering, State Key Laboratory of Biotherapy, West China Hospital, Sichuan University from June 2005 to May 2006. Osteoclasts were cultured from clean-grade neonatal SD rats aged 1 day by marrow induction method and incubated on bone sheet in MEM with different concentration of L-arginine （0.3,0.6,1.0 g/L）, and distilled water as control. After 7 days, number and morphology of the osteoclasts were detected with tartrate-resistance acid phosphatase （TRAP） staining. Image MIAS-2000 analysator was used to count the resorption pits and areas on slices. The effect of different concentrations of L-arginine on the resorption pits was analyzed under scanning electron microscope. RESULTS：①Osteoclasts were bigger than other cells and had irregular shapes, showing frying egg, sliver, allanto-shape or infundibulate. Several or decades of nucleus were seen in cells. The cells were stained in red by TRAP. ②Number of osteoclasts was decreased with increasing of L-arginine concentrations （P 〈 0.05）. ③After incubation for 7 days, resorption pits were decreased above the 0.3 g/L of L-argnine and with a concentration dependent manner. Areas of the resorption pits at 0.3 g/L, 0.6 g/L and 1.0 g/L were 91% （P 〈 0.05）, 80% （P 〈 0.05） and 69% （P 〈 0.01） of those of control group respectively. CONCLUSION： Many osteoclasts with high purity are cultured by marrow induction method, and osteoclasts have significant bone absorptive function. L-arginine inhibits osteoclast proliferation differentiation and resorptive activity in a concentration dependent manner.

关 键 词：L-ARGININE 破骨细胞 一氧化氮 骨吸收 

分 类 号：R681[医药卫生—骨科学]