摘要
研究背景:
自然铜是我国中医骨伤科接骨要药,为我国历版药典所收载。中药自然铜具有散瘀止痛,接骨续筋的功效,临床用于治疗跌打损伤,筋断骨折,血瘀疼痛等。自然铜是接骨续骨最常用的药物,其在骨科方剂中的地位不可替代。
临床上自然铜的使用绝大多数是与其他诸位中药配伍使用,并且多以口服方剂为主,有少部分使用浸泡疗法和膏剂案例。骨折发生大多为局部病灶,面对自然铜这样一个对于骨折疗效显著的药物,如果单独使用应选择一种直接在病灶处发挥作用的方式,因此自然铜的局部给药可能是一种理想方式。局部给药使局部组织的药物浓度高,药物主要在局部发挥作用,进入全身血循环的药物较少,对全身的影响较小。皮下注射是其中方法之一,但是局部注射有疼痛、需要多点、多部位注射和注射层次掌握有一定难度,药物分布不均匀,药物在缺血组织中渗透差等不足。低频超声介导下的经皮给药是近年来兴起的局部给药方法之一,是继注射和口服给药后又一种新的给药方式。在一定能量的低频超声介导下,蛋白质等大分子药物可以经局部皮肤被吸收,达到经皮给药的目的,而且这种给药方式不受局部血循环的影响,对缺血组织也同样能够实现局部经皮给药,药物分布均匀。这样局部小剂量的经皮给药,药物就可以在局部形成一个相对的高浓度,发挥其重要的效应。同时由于经皮给药后,药物具有缓释作用,药物进入全身血循环的剂量很小,用药安全,能减少全身的副作用。
研究目的:
通过应用低频超声透入中药自然铜,观察给药后有效成分组织浓度,自然铜对骨折愈合中BMP-2基因表达的影响以及低频超声透入自然铜对兔肢体骨折愈合的影响。
研究方法:
1.按随机分组的原则将75只新西兰大耳家兔分成五组,每组15只。治疗方法:治疗1组:浓度为0.5g/ml自然铜煎出液,治疗2组:浓度为1.0g/ml自然铜煎出液,治疗3组:浓度为2.0g/ml自然铜煎出液。治疗1-3组均采用低频超声透入给药,频率为20KHz,强度为100mW/cm~2,1次/日,每次20min。对照组:单纯采用超声照射,频率为20KHz,强度为100mW/cm~2,1次/日,每次20min。模型组:生理盐水20ml灌胃,1次/日。
2.自然铜制备生品自然铜,严格挑选出杂质,将矿石捣碎,用火锻醋淬法:将净自然铜锻至暗红,醋淬至表面呈黑褐色,光泽消失并酥松(如不酥,可反复锻淬至酥),每100kg自然铜用醋30kg(国家药典委员会.中华人民共和国药典2005年版(一部):95)。
3.X光透视
4.骨痂干燥
5.骨痂中Fe、Cu、Mn、Zn的测定
6.杂交
7.洗片
8.血清Ca~(2+)、P离子浓度、ALP活性检测
9.结果统计
研究结果:
自然铜能够明显提高家兔骨痂中微量元素Fe、Cu的含量(P＜0.05),从而增加骨痂的生长,表现在X光透视时,试验组家兔骨痂量显著多于对照组。从本次试验所测定的4种微量元素来看,自然铜治疗3组中铁高达1186μg/ml,家兔骨痂铁的含量达(172.68±14.32μg/g)之多,骨痂铜亦明显高于对照组。
在BMP-2表达方面,自然铜各治疗组均为棕色,BMP-2表达阳性。对照组及模型组片所有组织均未显色。模型组及对照组所有标本的外骨膜均增厚,治疗组1,2组绝大多数标本骨折断端成纤维细胞和纤维组织及类骨质增多,纤维排列较规则;所有标本外骨膜成骨细胞显著增生,增生细胞融合成一片,骨基质沉着,形成骨样组织、软骨及骨小梁,并向骨折间隙延伸,出现不完全的软骨与骨小梁骨痂相连,在成纤维细胞、成软骨细胞、软骨细胞、成骨细胞、骨细胞等表达阳性。治疗3组编织骨形成,骨髓腔内的骨小梁变细,骨髓腔再通。实验表明自然铜低频超声透入给药能促进血Ca、P、ALP含量增加兔肢体骨折愈合能力,能明显提高家兔骨痂中微量元素的含量(p＜0.05),增加骨痂生长量,从而促进骨修复过程。
结论:
1.低频(20KHz)超声药物导入是一种安全、有效的经皮局部给药的方式。
2.在低频超声介导下,煅自然铜可以经皮给药,在组织局部发挥其生物效应。
3.在低频超声介导下,随给药浓度提高,可以增加局部组织的药物浓度,且单纯的低频超声具有一定的促进愈合功效。
4.自然铜在超声介导下可以经皮给药能够诱导BMP-2合成加速骨折愈合;提高血Ca、P、ALP含量增加兔肢体骨折愈合能力;自然铜能明显提高家兔骨痂中微量元素的含量(p＜0.05),增加骨痂生长量,从而促进骨修复过程。
Background:Pyritum is important Chinese medicine for bone fracture and recorded in all ages in our national pharmacopoeia codex.It can help improving blood circulation and relieving bruises and pains and muscle and ligament reunion of bone.It is clinically used for injuries from falls,muscle and bone broken,gore and pains etc.Pyritum is the most common used medicine for bone-knitting and symplectic bone which there is no other orthopaedics prescription to replace.
Pyritum is always used matching with other Chinese medicine clinically and in multi-oral prescription.A few of them are used in marinate and electuary ointment.Bone broken always occurs locally.As Pyritum has such an effection in treatment of fracture,it must he a ideal method to administrate locally with Pyritum which can induce a high local drug concentration and drug plays a role locally that there is little drug concentration in body plasma.Hyodermic is one method of it.However it is painful,multi-point needed,difficult to master the level,not well-distrbuted.The disadvantage is low drug concentration in less plasma position.Through using a certain level low frequency ultrasound, giant molecule such as protein can be obtained across the skin arriving on target which will not be affected by local blood circle.It can realize the acquisition of local body insufficient of plasma and drug well-distribute.So local small dose across skin can make a relatively high concentration and has an important effects;meanwhile it has a sustained release effective.There is a little dosage in plasma and it is safe.Also it reduces secondary reaction.
Object:
After administrating Chinese medicine Pyritum with low frequency ultrasound,we observed local effective agent concentration and the effects of BMp-2's expression on fracture restitution when using Pyritum and low frequency ultrasound's administration effects on rabbit's bone restitution.
Method:
1.Put 75 New Zealand white big ears rabbits into 5 groups randomly,15 each.Method:one treatment group:0.5g/ml Pyritum decoction;two treatment groups:1.0g/ml Pyritum decoction;three treatment groups: 2.0g/ml Pyritum decoction.Group 1-3 with low frequency ultrasound, once per day.Control group:ultrasound with frequency of 20KHz, intensity of 100mw/cm~2,low frequency in vitro irradiation,once per day,20min once.Model group:normal saline 20ml lavage,once per day.
2.Pyritum preparation:Pyritum pick up impurity strictly,smash the ore, heat it in fire and dip it into vinegar:heat it to dark red,dip it into vinegar to pitchy,luster disappears and loose(if not loose over and over again till loose )vinegar 30kg per 100kg Pyritum(NF. Pharmacopoeia of PRC2005 edition(the lst):95)
3.X-ray transmission
4.poroma dry
5.determination of Fe,Cu,Mn,Zn in poroma
6.hybrid
7.develop a film
8.determination of Ca~(2+),P ion concentration and Alp activity in plasma
9.statistics results
Results
Pyritum can obviously increase Fe,Cu contents in poroma of rabbits.(P＜0.05),and then increase poroma's growth.We can observe that poroma in experimental group is more than it in control group when using X-ray.Observed from 4 trace elements from the testing,among 3 Pyritum experimental groups Fe concentration reaches 1186μg/ml,while it reaches(172.68±14.32μg/ml)in rabbit poroma.Cu in poroma in experimental group is higher than it in control group.
All Pyritum experimental groups are b(?)n,BMP-2 to express masculine. There is no color development in all control groups and model groups. All samples outside periosteum of model and control group grew thicker. Most samples of fracture mechanocyte,nerve structure and osteoid increased and fiber arrayed regularly in experimental group 1,2.All samples outside periosteum osteoplast got hyperplasia.The hyperplasia got together,bone matrix adipopexised and informed osteoid tissue,cartilage and bone trabecula,and expanded towards fracture clearance.Lame cartilage and bone trabecula's poroma connected and expressed masculine in mechanocyte,chondroblast cartilage,osteoplast and osteocyte.In 3 experimental groups woven bone got grow and medullary space bone trabecula got thinner and medullary space got open.It improved Ca,P,ALP concentration that union got increased.Pyritum can increase microelement in rabbit poroma obviously(p＜0.05),increase poroma growth and then improve bone repairing.
Conclusion:
1.Low frequency(20KHz)ultrasound administration is a safe,effective local percutaneous administration method.
2.With low frequency ultrasound,heated Pyritum can be administrated across skin and has bioeffects on local body.
3.With low frequency ultrasound local concentration will increase as dosage increase and single low frequency has effects on wound healing.
4.With low frequency ultrasound Pyritum can induce BMP-2 to make broken healing and improve Ca,P,ALP concentration to heal rabbit broken bone.Pyritum can increase trace element in rabbit poroma and then improve course of repair.
引文
[1]侯树勋.现代创伤骨科学[M].北京:人民军医出版社,2008.8:29-33.
[2]何洪阳,邓友章.现代骨伤诊断与治疗[M].北京:人民卫生出版社,2002.10:152.
[3]Bruce D.Browner主编.王学谦,娄思权等主译.创伤骨科学[M].天津:天津科技翻译出版公司出版,2007.1:49-50.
[4]陈昱,陈宗雄,徐皓等.骨折愈合机制的现代研究[J].中国组织工程研究与临床康复,12,50:9951-9956.
[5]张建军,李兵仓.骨折愈合的现代研究与局部基因治疗[J].创伤外科杂志,2003.5:230-232.
[6]Sampath TK,Coughlin JE,Whetstone RM,et al.Bovine osteogenic prot ein is composed of dimers of DP-1 and BMP-2A,two members of the transforming growth factor-beta superfamily.J Biol Chem,1990,265(3):13198.
[7]Rundle CH,Miyakoshi N,Ramirez E,et al.Expression of the fibroblast growth factor receptor genes in fracture repair[J].Clin Orthop,2002,(403):253-263.
[8]陆敏安.骨折愈合因素分子水平的研究进展[J].右江民族医学院学报,2004.6:418-420.
[9]Tarkka T,Sipola A,Jamsa T,et al.Adenoviral VEGF A gen etransfer inducesan giogenesisand promotes bone formation in healing osseous tissues[J].J Crene Med,2003,5(7):560-566.
[10]丁真奇,郭延杰.胶原与骨折愈合[J].骨与关节损伤,1997,12(2):125.
[11]胡蕴玉.骨折愈合的分子生物学进展[J].中国现代医学,2002,1:27-32.
[12]王立平,党耕町.骨连接素在骨愈合中作用的探讨[J].中华骨科杂志,1998,8:497-499.
[13]王立平,党耕町.骨桥蛋白在骨愈合过程中的基因表达[J].北京医科大学学报,1998,8:355-357.
[14]刘道德.骨折愈合过程及相关细胞因子[J].四川解剖学杂志, 2006,14:46-48.
[15]Joseph A,Buckwalter,MD,MS,et al.陈启明,梁国穗等译.骨科基础科等(第二版)[M].人民卫生出版社,2002:326-328.
[16]裘法祖.外科学(第四版)[M].人民卫生出版社,1995:712-714.
[17]曾敬,徐栋梁等.成骨细胞移植促进骨质疏松性骨折愈合过程中相关因子的动态表达[J].中国临床康复,2003.7(3):448-450.
[18]卢世璧主译.坎贝尔骨科手术学(第九版)[M].济南.山东科学技术出版社,2001:2
[19]秦书俭等.影响骨折愈合的化学因素[J].解剖科学进展,2000(6):14-17.
[20]黄遂柱,赵永强,陈风苞等.雌激素对实验性骨质疏松症骨折愈合的影响[J].中国骨质疏松杂志,2000.6(1):17.
[21]袁凌伟,夏亚一,鲁茂森等.骨折愈合应力机制研究进展[J].国际骨科学杂志,2007.3:84-86.
[22]苏佳灿,任可,张春才.应力对骨折愈合的影响及其作用机制[J].第二军医大学学报,2001.10:988-990.
[23]陈建平,谭润初,李济时.He-Ne激光加速兔骨折愈合的生物学效应及机理探讨[J].中山医科大学学报,1991(12):265-270.
[24]Diwan AD,Wang MX,Jang D,et al.Nitric oxide modliates fracture healing[J].J Bone Miner Res,2000,15(2):342-351.
[25]Baldik,Y,Talu U,Altinel L,et al.Healing regulated by nitric oxide;an expermental study in rat[J].Clin Orthop Relat Res,2002,(404):343-352
[26]乔林.微动对骨愈合的影响及作用机理.中国人民解放军军医进修学院博士学位论文.2004.4
[27]赵小纲.创伤性脑损伤影响骨折愈合速度机制的研究.浙江大学博士论文.2006.3
[28]王丹,胡蕴玉.低强度超声促进骨折愈合[J].国外医学生物医学工程分册,2000(23):247-251
[29]王和鸣,严孟宁.调频脉冲电磁场影响骨折愈合的实验研究总结[J].福建 中医学院学报,2003,5:23-25.
[30]李晓林,杜靖远,饶振玉等.高能震波进骨折愈合机理的探讨[J].中华实验外科杂志,2003,3:128-129.
[31]吴颖,谭日,姜群英等.高压氧治疗骨折愈合障碍临床疗效及机理分析[J].中华医药学杂志,2004(3):5-6.
[32]高春锦,杨捷云编著.实用高压氧学[M].北京:学苑出版社,1997,8:352-353.
[33]黄志强.高压氧在骨科的运用[M].北京:人民军医出版社,1989,7:44.
[34]马瑞山.航空航天生理学[M].西安:陕西科学技术出版社,1999,8:142-143.
[35]刘子藩,易治.实用高压氧医学[M.广州:广东科技出版社,1990,5:43.
[36]叶俊材,张俐.基因治疗在骨折愈合中的研究[J].中国中医骨伤科杂志,2003,12:54-58.
[37]高铁军.左旋多巴促进骨折愈合的作用机理[J].中华骨科杂志,1990(10):213-216.
[38]汤文辉,罗亦雄,黄慧姬.左旋多巴促进骨折早期愈合的临床研究[J].广州医药,2005(36):37-39.
[39]续力民,张世华,崔瑞祥等.体内磁场促进骨折愈合的实验研究[J].北京军区医药,1994(6):329-331.
[40]王占朝.外固定架结合低频脉冲电刺激仪促进骨折愈合的实验研究.中国人民解放军军医进修学院博士学位论文,2005.5
[41]唐广应,黄海,胡建山.促进骨折愈合方法的研究进展[J].贵阳中医学院学报,2007,7:51-54.
[42]李展振,危立军,应文世等.接骨系列方促进骨折愈合疗效观察[J].江西中医药,1999,39(5):36.
[43]荷维英.接骨汤促进骨折愈合的临床观察[J].浙江中医学院学报,1998,22(5):19-20.
[44]汪光晔,黄家伦.全龟胶囊促进骨折愈合疗效观察[J].基层中药杂志,1998,12(4):63-64.
[45]李明,李日才,刘方田.调理脾胃促进骨折愈合—附210例疗效观察[J].山东中医杂志,1994,13(4):156.
[46]谭德雄.金锁固精丸治疗骨折迟缓愈合22例报告[J].中国中医骨伤科杂志,1991,7(2):44-47.
[47]任桂兰.阳和汤合剂治疗骨折延迟连接疗效观察[J].中医正骨,1999,11(2):11-12.
[48]杜洪刚.六味地黄汤合八珍汤加味治疗骨折延期愈合[J].贵阳中医学院学报,2001,23(4):17-18.
[49]朱海,李全,黄涛等.漏天机接骨丹促进胫腓骨中下1/3骨折愈合的临床研究[J].中医正骨,2002,14(1):13-14.
[50]顾科民.关于中医伤科“接骨方药”的文献整理[J].上海中医药杂志,1963,8:31.
[51]杨益,李宇明,翁凤全.驳骨丸促进骨折愈合的临床研究[J].中国中医骨伤科杂志,1998,6(5):19-23.
[52]张三康.张氏接骨散对骨折愈合影响的x线和血尿生化学的临床研究[J].中医正骨,1995,7(1):9-11.
[53]海孝忠.续骨糖蟹膏促进骨折愈合的疗效观察[J].河南中医药学刊,1997,12(5):54-55.
[54]刘献祥,许书亮,王和鸣.麦饭石促进家兔骨盐沉积的实验研究[J].福建中医药,1991,22(1):321.
[55]刘献祥,许书亮,王和鸣.麦饭石对骨折愈合过程影响的实验研究[J].中医正骨,1992,4(1):4.
[56]刘献祥,许书亮,郭文贵.麦饭石对骨折愈合作用的骨痂强度微量元素研究[J].福建中医学院学报,1994,4(2):251.
[57]刘献祥,许书亮,王和鸣.麦饭石促进骨折愈合的实验研究[J].中国中医骨伤科杂志,1995,3(2):51.
[58]洪定钢,时光达.续断对实验性骨折愈合影响的骨组织形态计量学研究[J].中国中医骨伤科杂志,1999,7(3):41.
[59]洪定钢,时光达.淫羊藿影响骨折愈合的骨组织学及骨组织形态计量学研 究[J].中医正骨,1999,11(2):31.
[60]罗佩强.土鳖虫促进骨折愈合的实验研究[J].中国骨伤,1992,5(6):61.
[61]夏志道,房世源,常超英等.壮骨素促进骨折愈合的实验研究[J],中国骨伤,1994,7(1):7.1
[62]邱桐,林维君,王慎.“接骨一粒丹”对家兔尺骨骨折X线片表现的影响[J].甘肃中医学院学报,1992,9(2):351.
[63]熊芳丽,肖亚平.针灸促进骨折愈合的临床观察[J].中国针灸,1997,17(11):6851.
[64]阮朝阳.中药促进骨折愈合作用机制的实验研究进展[J].湖南中医药导报,2003,9(2):49.
[65]张晓燕,吴铁.针灸促进骨折愈台的研究现状[j].河北中医,2005,27(7):548
[66]王文远.针刺臀痛穴对大白兔下肢软组织损伤修复的影响[J].中国矫形外科杂志,1997,4(4):312.
[67]李澎涛.针刺对家兔超负荷运动骨骼肌细胞内钙离子活度的影响[J].中国针灸,1994,14(6):30.
[68]胡美珠,张菊英,刘季兰等.丹参注射液对家兔桡骨骨折以后肢体血容量的影响[J].中华外科杂志,1979,17(1):33.
[69]周重建,王绪辉,朱显华等.益气化瘀剂在骨折愈合中的生化和病理观察[J].中国中医骨伤科杂志,1989,5(3):7.
[70]侯筱魁,石关桐,王公林等.外敷中药对家兔桡骨骨折愈合的影响[J].中国中医骨伤科杂志,1992,8(2):10.
[71]马克昌,朱太永,刘鲜茹等.益气化瘀汤对实验性骨折的影响[J].中医正骨,1995,7(1):15.
[72]侯晓峰,刘景生.中药促进骨折愈合机理.中国中医骨伤科杂志,2003,2(11):61.
[73]孙炳烈,徐今宇,赵然.平乐内服接骨丹对骨折愈合作用的实验研究[J].中西医结合杂志,1987,7(9):544.
[74]贵阳中医学院骨折科研协作组.中药“接骨Ⅰ号”促进骨折愈合作用的实验 观察[J].中华医学杂志,1978,11:670.
[75]曲克服,王智兴,王以进等.理气活血剂在骨折愈合过程生化平”生物力学观察[J].上海中医药杂志,1982,5:42.
[76]林燕萍,王和鸣,陈奕权.煅狗骨对实验性骨折愈合作用的组织学和组织化学研究[J].中国中医骨伤科杂志,1991,17(6):4.
[77]刘季兰,张菊英,柴本甫.丹参注射液对骨折愈合过程中钙吸收的影响[J].中西医结合杂志,1985,5(7):418.
[78]刘毓顺,周金水,林葆凯等.接骨散对家兔骨折的实验研究[J].福建中医药,1994,25(3):19.
[79]夏志道,房世源,常超英等.壮骨素促进骨折愈合的实验研究[J].中国骨伤,1994,7(1):7.
[80]王军跃,成扶真,度元斌等.接骨七厘片对实验性骨折愈合的影响[J].中国中医骨伤科杂志,1997,5(2):1.
[81]秦军志,王贤淑.丹参对大鼠胫骨骨折早期愈合过程中血清、骨痂及骨组织中钙、锌、铜的影响[J].中国中西医结合杂志,1992,12(6):354.
[82]柴本甫,汤雪明.活血化瘀药丹参治疗骨折的超微结构研究[J].中西医结合杂志,1987,7(7):417.
[83]王宝泉,姚树元,徐尔真.生肌象皮药膏在感染性开放骨折中的应用及实验研究[J].中医杂志,1981,1:24.
[84]林润台,王忠,孙冀等.丹参促进下颌骨骨折愈合的超微结构研究[J].中华口腔医学杂志,1992,27(4):215.
[85]杨淮纭,沈志祥,董福慧.中草药常绿油麻藤种子提前左旋多巴促进骨折愈合的临床及实验研究[J].中西医结合杂志,1985,5(7):398.
[86]林燕萍,王和鸣,周瑞祥.煅狗骨对实验性骨折愈合作用的垂体TSH,GH细胞免疫细胞化学研究[J].中国骨伤,1993,6(6)53.
[87]李也白,温宏,徐华梓等.复合中药制剂促进骨折愈合的实验研究[J].骨与关节损伤杂志,1999,14(2):108.
[88]姜欣,胡兴山.整骨煎对小鼠粉碎性骨折应激的免疫调节实验研究[J].辽宁中医杂志,1994,10(10):476.
[89]魏玉玲,晏雷生,粱克玉.伤科接骨片促进骨折愈合作用的实验研究[J].中医正骨,1996,8(4):3.
[90]刘艺,顾国奎,王和鸣.海螵哨接骨动物实验的组织学观察[J].中国中医骨伤科杂志,1995,3(5):6.
[91]张俐.针刺对骨折家兔微量元素的影响[J].中国中医骨伤科杂志,1999,7(5):1.
[92]汤耿民,沈霖,涂意辉,等.补肾活血方对成骨细胞生长因子TGF—Ⅰ mRNA 表达的影响[J].中国中医骨伤科杂志,1999,7(5):5.
[93]张志杰,蔡宝昌.中药自然铜研究概况[J].南京中医药大学学报,2005,21(1):66-68.
[94]明李时珍.本草纲目[M].校点本 上册 人民卫生出版社1985:466.
[95]高天爱.矿物药及其应用[M].中国中医药出版社,1997,12:120.
[96]国家药典委员会.中华人民共和国药典[S],2005年版(一部):95
[97]太医院编.圣济总录[M].北京:人民卫生出版社,1962:879
[98]雷.雷公炮炙论[M].北京:人民卫生出版社,1957.5.
[99]陈师文.太平惠民和剂局方[M].北京:人民卫生出版社,1985:269.
[100]张子和.儒门事亲[M].上海:上海卫生出版社,1958.213.
[101]朱震亨.丹溪心法[M].上海:上海科学技术出版社,1959.175.
[102]朱棣.普济方[M].北京:人民卫生出版社,1959.325.
[103]张景岳.景岳全书[M].上海:上海科学技术出版社,1959.949.
[104]汪昂.本草备要[M].上海:商务印书馆铅印,1954.96.
[105]曾白林,朱云昌.自然铜不同炮制品中金属元素的含量测定[J].基层中药杂志,2001,15(2):29.
[106]张亚敏.自然铜的质量探讨[J].中成药,1995,17(10):21.
[107]李轩贞,张世臣,刘智伟等.不同来源自然铜炮制的研究[J].中国中药杂志,1995,20(8):467.
[108]彭怀仁.中医方剂大辞典[M].第9册 北京:人民卫生出版社,1996,383-400.
[109]赵利平,房少新.自然铜对家兔骨痂中微量元素的影响[J].中兽医医药杂 志,2003,(3):39-40.
[110]何赞厚,刘庆思.中药驳骨煎剂对骨折小鼠45Ca和32P水平的影响[J].广州中医药大学学报,1998,15(4):278.
[111]刘进,张雪华.氟化钠、自然铜、维生素C成骨效果的实验研究[J].临床口腔医学杂志,1997,13(4):224-226.
[112]王智兴,过邦辅,柴本甫等.自然铜在骨折愈合过程中对总胶原、不溶性胶原、钙、磷和生物力学的影响[J].中华骨科杂志,1986,(4):305.
[113]刘海成,朱显,赵士林等.磁与自然铜促进骨折愈合的初步实验研究[J].中华外科杂志,1983,21(1):1.
[114]徐爱贤,高学媛.磁与自然铜促进骨折愈合的实验研究[J].山东医学杂志2008,28(8):558-560.
[115]张克勤,刘湘秀,李瑞宗.七厘散对骨折愈合作用的初步实验研究[J].中华外科杂志,1962,10(5):305.
[116]徐经采.微量元素与骨骼[J].中国中医骨伤科杂志,1987,3(1):55.
[117]陈荣三,张祖暄译.微量元素与人[M].北京:科学出版社,1979:34.
[118]李静均,李舜贤.矿物药“自然铜”.有色金属[J],1990,42(2):77.
[119]叶定江.中药炮制学[M].上海:上海科学技术出版社,1996:270.
[120]张克勤,刘湘秀,李瑞宗等.自然铜在骨折愈合中的作用[J].天津医药(骨科附刊),1962,4(2):72.
[121]张惠法.自然铜在伤科的应用史及其现代研究[J].南京中医药大学学报,1996,12(6).
[122]神农本草经(M).人民卫生出版社,1963:7.
[123]丁继华,等.中医骨伤科荟萃[M].中医古籍出版社,1986:23.
[124]王焘.外台秘要[M].人民卫生出版社,1955:(29):787.
[125]唐慎微.重修政和经史证类备用本草[M].人民卫生出版社,1957:127.
[126]蔺道人.仙授理伤续秘方[M].人民卫生出版社.1954:8,9,14,17,18.
[127]韦以宗.中国骨科技术史[M].上海:上海科技出版社,1983:300.
[128]顾科民.关于中医伤科接骨方药的文献整理[J].上海中医药杂志1963,8:31.
[129]陈馥馨,等.接骨丹文献研究275方剂的方药分析[J].河南中医,1981(4):44.
[130]彭怀仁.中医方剂大辞典[M].第9册.北京:人民卫生出版社,1996:383-400.
[131]北京医学院学报[J].1959:(1):149.
[132]陆炳全,黄素碧,贺景国等.自然铜促进骨折愈合实验研究[J].World Health Digest,2006,9(3):76-77.
[133]冯伟,傅文,朱雅萍等.接骨中药对培养成骨细胞增殖、分化和矿化功能的影响[J].浙江中医学院学,2004,28(2):65-67.
[134]张军.自然铜的鉴定与接骨侔用[J].医函授通讯,1998,17(5):53.
[135]王景祥等.骨愈片促进骨折愈合作用的研究[J].中草药,1988,19(8):17.
[136]赵建等.促骨愈胶囊治疗骨折延迟愈合临床观察[J].河北中医,2005,27(8)592-593.
[137]高振强等.复方芪铜胶囊的制备及临床疗效观察[J].医院制剂,1999,8(9):49.
[138]刘又文,三七接骨丸治疗创伤骨折202例[J].新中医,2004,36(9):60-61.
[139]陆炳全.生骨丸与续骨长泡剂促进骨折早期愈合的临床观察[J].西部医学,2003,1(1):33-34.
[140]周文敏,林熙文.林如高跌打补骨丸治疗骨折188例[J].福建中医药,2004,35(3):16.
[141]张士勇.舒筋接骨膏的制备与临床应用[J].基层中药杂志,2001,15(2):59.
[142]郑俊民.《经皮给药新剂型》.2006,5:25.
[143]王军红,王松俊,刁天喜.中国人民解放军军事医学科学院.2005,5:27.
[144]Mitragotri S,Kost J.Low-frequency sonophoresis:a noninvasive method of drug delivery and diagnostics[J].Biothechnol,2000,16:488-492.
[145] Zhang F, Oswald T, Lin S et al. Vascular endothelial growth factor(VEGF)expression and the effect of exogenous VEGF on survival of a random flap in therat[J]. Br J Plast Surg, 2003, 56:653-659.
[146] Leduc S. Indroduction of medicinal substances into the depth of tissues by electric current[J]. Ann d' Electrobiol,1900, 3:545.
[147] Rein H. Experimental electroendosmotic studies on licing human skin[J]. Z Biol, 1924, 81:124.
[148] Blank IH. Penetration of low-molecular-weight alcohols into skin. I. Effect of concentration of alcohol and type of vehicle[J]. J Invest Dermatiol, 1964, 43:415.
[149] Michaels AS, Chandrasekaran SK, Shaw JE. Drug permeation through human skin: theory and in vitro experimental measurement [J]. AlChE J, 1975,21:985.
[150] Mitragotri S, Blankschtein D, Langer R. Ultrasound-mediated transdermal protein delivery[J]. Science, 1995,269:850.
[151] Lee S, McAuliffe DJ, Flotte TJ, et al. Photomechanical transcutaneous delicery of macromolecules[J]. J Inverst Dermatol ,1998,111:925.
[152] Bramson J, Dayball K, Evelegh C. et al. Enabling topical immunization via microporation: a novel method to for pain-free and needle-free delivery of adenovirus-based vaccines[J]. Gene Ther,2003, 10:251.
[153] Mitragotri S, Kost J. Low-frequency sonophoresis: q review[J]. Sdv Drug Deliv Rev, 2004, 5(5):589.
[154] Prausnitz MR, Mitragotri S, Langer R. Current status and future potential of Transdermal drug delivery[J]. Nat Rev Drug Discov,2004,3(2): 115.
[155] Lavon I, Kost J. Ultrasound and transdermal drug delivery. Drug Discov Today,2004,9(15):670 Merino G,Kalia YN,Guy RH.Ultasound and transdermal drug delivery[J].J Pharm Sci,2003,92(6):1125.
[156]Machet L,Boucaud A.Phonophoresis:efficiency,mechanisms and skin toleranc[J].Int J Phram,2002,243:1.
[157]Joshi A,Raje J.Sonicated transdermal drug transport[J].J Controlled Release,2002,83:13.
[158]Meidan VM,et al.Phonophoresis-is it reality?[J].Int J Phram,1995,118:129.
[159]周万松.超声药物透入疗法的临床研究进展.应用声学,2001,20(1):6.
[160]Viak T.Comparative study of the efficacy of ultrasound and sonophoresis in the treatment of painful shouler syndrome[J].Reumatizam,1999,46:5-11.
[161]高建青,梁文权.促进药物经皮渗透的新方法.国外医学药学分册,1997,23(17):42.
[162]Fellinger K,Schmid J.Klinik an Therapie des Chronischen[J].Maudrich,Vienna,Austria:Gelenkreumatismus,1954:549-552.
[163]梁炳文,梁文权,平其能等.经皮给药制剂.第1版.北京:中国医药出版社1992:180.
[164]郑俊民,李英健,丁平田.药物的经皮吸收途径的初步探讨[J].首届全国新型给药技术与中药现代化学术研讨会论文集,2002:11-14.
[165]Tachibana K,Tachibana S.Transdermal delivery of insulin by ultrasound vibration[J].J Pharm Pharmacol,1991,43:270.
[166]Tezel A,Sens A,Wuchscherer J et al.Frequency dependence of sonophoresis[J].Pharmaceu Res,2001,18:694-1700.
[167]Mitragotri,S.;Blankschtein,D.;Langer,R.Ultrasound-mediated transversal protein delivery[J].Science,1995,269:850-853.
[168]S.E.Cross and M.S.Roberts.Physical Enhancement of Transversal Drug Application:Is Delivery Technology Keeping up with Pharmaceutical Development Current Drug Delivery[J], 2004,1:81-91.
[169] Zhdanov.-v-plaster non-contact drug delivery methods:mathematical and experimental substantiation.Crypt-Rev-Biomed-Eng[J], 2001,29(1):142-156.
[170] Chang-SL: The effect of electro oration on iontophoretic transdermal delivery of calcium regulating hormones.J-Control-Release[J]. 2000 May 15; 66(2-3):127-33.
[171] Wallace,-M-S;Topical delivery of lidocaine in healthy volunteers by electroporation, electroicorporation, or iontophoresis: an evaluation of skin anesthesia[J]. Reg-Anesth-Pain-Med, 2001 May-Jun; 26(3):229-38.
[172] Mir, -L-M. Therapertic perspectives of in vivo cell electropermeabilization[J]. Bioelectrocheimstry. 2001 Jan;53(1):1-10.
[173] Cemazarm, el. Changed delivery of boron to tumours using electroporation for boron neuture therapy with BSH[J].Br-T-Kadiol. 2000, Feb;73(866): 195-200.
[174] Terry O Herndon,Salvador Gonzalez, TR Gowrishankar, R Rox Anderson, James C Weaver, Transdermal microconduits by microscission for drug delivery and sample acquisition[J]. BMC Medicine, 2004, 2:12.
[175] Allegretti, -J-P. Electroporation therapy for head and neck cancer including carotidartery involvement[J]. Laryngoscope, 2001, Jan;111(1): 52-6.
[176] Chien YW. Reansdermal controlled-release drug administration In:Chien YW, ed. Novel Drug Kelivery Systems[J]. Vol.4, New York:Marcel Dekker, 1982:155-157.
[177] Viak T. Comparative study of the efficacy of ultrasound and sonophoresis in the treatment of painful shouler syndrome[J].Reumatizam, 1999,46:5-11.
[178] Pauza KJ, Howell S Dreyfuss P. A randomized, placebo-controlled trialof intradiscal lectrothermal therapy for the treatment of discogenic low back pain[J]. Spine J, 2004;4: 27-35.
[179] Zhu, -H; Quantification of pore induction in human epidermal membrane during iontophoresis: the importance of background electrolyte selection .J-Pharm-Sci.2001 Jul; 90(7):932-42
[180] Barry,-B-W. Novel mechanism and devices to enable successful transdermai drug delivery[J]. Eur-J-Pharm-Sci. 2001 Sep, 14(2):101-14.
[181] Saal JA, Saal JS Intradiscal electrothermal thera-py for the treatment of chroniv discogenic low back pain[J]. Clin SportsMed,2002; 21: 167-187.
[182] Mechael D, Martin MD, Cristopher M, et al. Pathophysiology of Lumbar Disc Degeneration: A Review of the Literature[J]. Neurosurg Focus, 2002, 13:198-200.
[183] Byl N. The use of ultrasound as an enhancer for transcutaneous drug delivery: phonophoresis[J]. Phys Ther, 1995, 75:539-553.
[184] Ballerini R, Casini A, Chinol M, et al. Study on the absorption of ketoprofen topically administered in man: comparison between tissue and plasma levels[J]. Int J Clin Pharm Res, 1986,6:69-72.
[185] Erkan Kozanoglu, Sibel Basaran, Rengin Guzel. Fusun Guler-Uysal Short term, efficacy of ibuprofen phonophoresis versus continuous ultrasound therapy in knee osteoarthritis[J]. Swiss Med Wkly,2003,133:333- 338.
[186] Tang,H; Theoretical description of transdermai transport of hydropbilic permeants: application to low-frequency sonophoresis[J]. J-Pharm-Sci, 2001 May, 90(5):545-68.
[187]Tomizawa,-M.Irradiation with ultrasound of low output intensity increased chemosensitivity of subcutaneous solid tumors to an anti-cancer agen[J]t.Cancer-Lett,2001 Nov 8:173(1)31-5.
[188]Cuschieri.In situablation of hepatic tumors[J].Semin -Laparosc-Surg.2001 May,8(1):25-41.
[189]Yamashita,-Y-I.Electroporation-mediated interleukin- 12 gene therapy for hepatocellular carcinoma in the mice model[J].Cancer-Res.2001 Feb,1,61(3):1005-12.
[190]Montorsi-F.Transdermal electromotive multi-drug administration for Peyronie disease:preliminary results[J].J-Androl.2000Jan-Fed,21(1):85-90.
[191]Mitragotri S,Blankschtein D,Langer R.Transdermal drug delivery using low-frequency sonophoresis[J].Pharm Res,1996,13:411.
[192]张风兰等.超声药物透入治疗狭窄性腱鞘炎20例[J].中华理疗杂志,1987,10(3):185.29
[193]苟建平等.超声氢化可的松透入治疗健鞘炎的观察[J].第三届全国超声治疗学术会议论文集,1995:24.
[194]许建丽.超声中频电同步叠加治疗软组织损伤[J].第一届全国超声治疗学术会议论文集,1991:20.
[195]骆正良.第四届全国超声治疗学术会议论文集[J],1997:21.
[196]第一军医大学附一院理疗科.超声波跌打膏治疗扭挫伤疗效观察[J].中华理疗杂志,1987,1(2):104.
[197]李莉等.超声利多卡因透入治疗慢性软组织疼痛的疗效观察[J].第三届全国超声治疗学术会议论文集,1995:18.
[198]杨小霞等.超声氢化可的松透入治疗肋软骨炎的疗效观察[J].第三届全国超声治疗学术会议论文集,1995:26.
[199]许宝霞等.超声波玻璃酸酶透入治疗瘫痕硬结的观察[J].第三届全国超声治疗学术会议论文集,1995:1.
[200]张文琦等.第一届全国超声治疗学术会议论文集,1991:69.
[201]曾立源.超声治疗前列腺疾病的观察[J].中华理疗杂志,1991,14(1):32-33.
[202]刘莉等,扶他林乳胶耦合剂超声治疗网球肘疗效观察[J].中华理疗杂志,1998,21(3):183.
[203]结克.醋酸可的松超声透入治疗颞颌关节功能紊乱40例[J].中华理疗杂志,1986,9(3):138.
[204]吴弥群等.超声药物透入治疗手足癣观察[J].中华理疗杂志,1998,21(2):86-87.
[205]Tezel A,Mitiagotri S.Interactions of inertial cavitation bubbles with stratum corneum lipid bilayers during low2frequency sonophoresis[J].Biophys,2003,85:3502.
[206]Zhang GL,Shi ZD.Progress on research of low2frequency sonophoresis[J].Journal of Huaxi Medical,2001,16(5):367.
[207]Tezel A,Sens A,Mitragotri S.Investigations of the role of cavita2 tion in low2frequency sonophoresis using acoustic spectroscop[J]y.Pharm Sci,2002,91:444.
[208]Tong H,Hu J H.The Development of research on using low-frequency sonophoresis to drug transdermal delivery.Medicine Abroad —Synthetic Medicine[J].Biochemical,Agents Volumes,2002,23(6):366.
[209]itragotri S,Kost J.Low2frequency sonophoresis a review[J].Ad2vanced Drug Deliwry Reviews,2004,56:589.
[210]FangJ,Fang C,Sung KC,et al.Effect of low2frequency ultra2 1478
[211]Boucaud A,Machet L,Arebeille B,et al In vitro study of low2frequency ultrasound2enhanced transdermal transport of fentany and caffoine across human and hairless rat skin[J].Int J Pharm,2001,228(122):69.
[212]Cheng C,Sun XF.The Development of research on transdermal delivery of insulin using ultrasound[J].Journal of Gilin Medical, 2007,28(8):963.
[213]杨中林,邢桂兰.超声促透对洋金花膜中东莨菪碱透过量的影响研究[J].中国中医基础医学杂志,1999,5(12):34.
[214]杨中林,韦英杰,张继全.超声促透对丹参中丹参素透过量的影响[J].中国中医基础医学杂志,2000,6(11):53.
[215]林海.超声中药透入治疗颈肩腰腿软组织疼痛的疗效观察[J].第一届全国超声治疗学术会议论文集,1991:65.
[216]张淑惠等,超声中药透入治疗血栓闭塞性脉管炎12例[J].中华理疗杂志,1987,1(1):51.
[217]张吉武等.第一届全国超声治疗学术会议论文集,1991:60.
[218]刘延春等.超声中药透入治疗颈椎病对颅内动脉血液动力学变化的影响[J]..中国超声医学杂志,1994,10(1):70-72.
[219]贺燕生等,超声中药透入治疗膝盖关节病临床观察[J].中国超声医学杂志,1992,8(5):359-360.
[220]胡中等.超声药物透入治疗椎动脉供血不足血流动力学研究[J].中华理疗杂志,2000,23(2):86-88.
[221]童华娅.第一届全国超声治疗学术会议论文集,1995:15.
[222]张玉英等.超声波对组织硬结的动物实验和临床疗效观察[J].中国超声医学杂志,1994,10(4):38-39.
[223]胥方元等.中药超声透入防治瘢痕增生[J].中华理疗杂志,1998,21(3):181.
[224]Mitragotri S,Blankchtein D,Langer R.Transdermal drug delivery using low-frequency sonophoresi[J]s.Pharm Res,1996,13(3):411.
[225]Boucaud A,Montharu J,Machet L,Arbeille B,mahet MC,Patat F.Vaillant L.Clinical histologic,and electron microscopy study of skin exposed to low-frequency ultrasound[J].Anat Rec,2001,246:114.
[226]Kost J,Mitragori S,Gabbay R,et al.Transdermal extraction of glucose and other analyses using ultrasoun[J]d.Nat Med,2000, 6:347.
[227]李相平等.超声透入丙酸睾丸酮治疗骨折观察及临床应用[J].中华理疗杂志,1999,22(1):36-37.
[228]Rediske AM,Roeder BL,Brown MK,Nelson JL,Robison RL,Drper DO,Schaalje GB,Robison RA,Pitt WG.Ulrasonic enhancement of antibiotic action on Escherichia coli biofilms:an in vivo model[J].Antimicrob Agents Chemother,1999 May,43(5);1211-4.
[229]Grabois,Martin,Managemt of Chronic Low Back Pain.American journal of Ph(?)cal Medicine(?) Rebobilitation[J].34(3)Supplement:S29-S41.March 2005.
[230]Barbara Cagnie,Elke Vinck,Steven Rimbaut,Guy Vanderstraeten.Phonophoresis Versus TopicalApplication of Ketoprofen:Comparison Between Tissue and Plasma Levels[J].Phys Ther,2003,83(8):707-712.
[231]Rathbone.M.J.Hadgraft,J.and Roberts,M.S.eds.Modified release drug delivery technology(Marcel Dekker)[J].Cevc,G.2003:533-560.
[232]Kost J,Pliquen U,Mitragotri S,et al.Synergistic effect of electric field and ultrasound on transdermal transport[J].Pharm Res,1996,13(4);633.
[233]Kunio Awazu,Takeyuki Uchizono,Sachiko Suzuki,Therapeutic Laser Applications and Laser-Tissue Interactions Ⅱ,Hubert van den Bergh,Alfred Vogel;Eds,Aug 2005,p.81-88.
[234]Lee WR,Shen SC,Lai HH.Transdermal drug delivery enhanced and controlled by erbium:YAG laser:a comparative study of lipophilic and hydrophilic drugs[J].J Control Release,2001 Jul 10,75(1-2):155-66.
[235]郑俊民.经皮给药新剂型.北京:人民卫生出版社,1997:23.
[236]Wallace,-M-S;Topical delivery of lidocaine in healthy volunteers by electroporation,electroicorporation,or iontophoresis:an evaluation of skin anesthesia[J].Reg-Anesth-Pain-Med.2001May-Jun,26(3):229-38.
[237]朱于春.促进透皮给药的物理和化学方(M).国外医药学分册,1993,12:15.
[238]Matsubara,-H.Combinatory anti-tumor effects of electroporation -mediated chemotherapy and wild-type p53 gene transfer to human esophageal cancer cells[J].Int-J-Oncol,2001 APT,18(4):825-9.
[239]马世勇等.电超导药物组织浸润治疗中晚期肝癌68例分析[J].中医肿瘤杂志,1999,21(5):216.
[240]孙孝祥,徐凯健,张亚平.中药透皮吸收研究概况[J].中国药学杂志,1996,1(31):163.
[241]刘文惠等.红花提取液超声电导体外透皮实验研究[J].中国中药杂志,1997,23(7):234.
[242]Doan N,Reher P,Meghji S,Harris M.In vitro effects of therapeutic ultrasound on cell proliferation,protein synthesis,and cytokine production by human fibroblasts,osteoblasts,andmonocytes[J].J Oral maxillofac.Surg,1999 Apr,57(4):409-19.
[243]Ng K Y,Liu Y.Therapeutic ulrasound:its application in drug delicery[J].Med Res Rev,2002 Mar,22(2):204-23.
[244]Schlafer O,Soevers M,Klotzbucher H,Onyeche TI.Improvement of biological activity by low energy ultrasound assisted bioteactors[J].Ultrasonics,2000 Mar,38(l-8):711-6.
[245]Boucaud A,Machet L,Arbeille B,Machet MC,Sournac M,Macon A,Patat F Vaillant L.Invitro study of low- frequency ultrasound-enhanced transdermal transport of fentanyl and caffeine across human and hairless rat skin[J].Int J Pharm,2001Oct 9,228(1-2):69-77.
[246]Nakano N,Nakano T.Surgicai technique of anterior extroperitoneal lumbar discetomy to the lumar disk hemiation Operation[J].Acat-Orthop-Bely, 1987, 41:1.
[247] Kaepp. VW. Experieat of Iontophoresis Controlled Release[J]Acat-Orthop-Bely, 1990,12:15.
[248] DoanN, Reher P, Meghji S, Harris M. In vitro effects of therapeutic ultrasound on cell proliferation, protein synthesis, and cytokine production by human fibroblasts, osteoblasts, andmonocytes[J]. J Oral maxillofac Surg, 1999 Apr, 57(4):409-19.
[249] Y. W Chen. Tranedermal Controlled Svstemic Medications, Marcei Dekker in New York and Basei[J]. Int J Pharm, 1997, 31(5) :675.
[250] Keyhani K, Guzman HR, Parsons A.Lewis TN. Intracellular drug delivery using low-frequency ultrasound: quantification of molecular uptake and cell viability. Prausnitz MR[J].Pharm Res, 2001Nov, 18(11): 14-20.
[251] Erkan Kozanoglu, Sibel Basaran, Rengin Guzel, Fusun Guler- Uysal.Short term, efficacy of ibuprofen phonophoresis versus continuous ultrasound therapy in knee osteoarthritis[J].Swiss Med Wkly,2003,133:333-338.
[252] Grabois, Martin. Management of chronic Low Back Pain. American Journa] of Physical Medicine & Rehabilitation. 84(3)Supplement:S29-S41,March 2005.
[253] Becker B M, Helfrich S, Baker E, Lovgren K, Minugh P A, Machan J T. Ultrasound with topical anesthetic rapidly decreasespain of intravenous sticks, Academic Emergency Medicine[J]. Swiss Med Wkly,2005,12:pp289-295.
[254] Barbara Cagnie, Elke Vinck, Steven Rimbaut, Guy Vanderstraeten.Phonophoresis Versus Topical Application of Ketoprofen:Comparison Between Tissue and Plasma Levels[J]. Phys Ther Vol.83,No.8,August, 2003:707-712.
[255]Lee WR,Shen SC,Lai HH.Transdermal drug delivery enhanced and controlled by erbium;YAG laser:a comparative study of lipophilic and hydrophilic drugs[J].J Control Release,2001 Jul 10,75(1-2);155-66.
[256]Longbridge,DJ,Sweeney,PA,Burkoth,TL.Effects of particle siae and cylinder pressure on dermal powderject delivery of testosteronetoconscious rabbits.Proc.Int.Symp.1998.Control.Rel Bioact.Mat.25,964.Tezel A,Sens A,Tuchscherer J,Mitragotri S.Frequency dependence of sonophoresis[J].Pharm.Res.,2001Dec,18(12):1694-700.
[257]Ng K Y,Liu Y.Therapeutic ultrasound:its application in drug.delicery[J].Med Res Rev,2002 Mar,22(2)'204-23.
[258]房少新,赵利平,吕红.山羊骨痂中微量元素与骨折愈合的实验研究(J).中兽医医药杂志,2001,3:17-19.
[259]初同伟,王正国,朱佩芳等.骨折愈合过程中vEGF及其受体的表达[J].中华创伤杂志,2001,17(6):66-69.