摘要
一研究背景
骨髓炎骨缺损(Osteomyelitic bone defection,OBD)对于病人来说是相当痛苦的,其治疗对于医生来说也是非常棘手的,有很多患有此病的患者最后不得不截肢。其治疗主要是抗感染和骨缺损的修复,近年来,出现了大量有关转移局部带血供肌瓣(Muscleflap with vessel pedicle,MFVP)修复骨缺损的报道,也有转移以腺病毒为载体表达的骨形态发生蛋白-2(Adenovirus Recombined Human Bone Morphogenetic Protein-2,Ad-rhBMP-2)基因诱导肌细胞成骨的报道,但尚未发现有关于转移肌瓣骨化过程及转移Ad-rhBMP-2基因修复OBD的报道。我们应用显微外科技术,转移局部带血管蒂肌瓣骨化修复OBD,并联合应用基因技术,缩短肌瓣骨化修复OBD时间,提出治疗OBD有效的新方法。
二实验目的
观察、比较两种治疗方案诱导肌瓣骨化修复OBD的效果。①应用显微外科技术,转移MFVP填充清创术后的OBD,保证肌瓣的血供。②联合应用显微外科技术与基因技术,转移MFVP填充清创术后的OBD,并转移Ad-rhBMP-2基因至肌瓣。
三实验方法
(一)大鼠OBD模型制备:
向胫骨上段骨缺损腔内塞入蘸有3×10~6CFU金葡菌0.3 ml的纱布及注射5%鱼肝油酸钠(Sodium Morrhuate SM)注射液(含有2%苯甲醇),以骨蜡封闭骨面,14天后,造成骨髓炎骨缺损模型。
(二)治疗OBD
1分组:
将120只Wistar大鼠OBD模型随机分为3组,每组40只:第1组,对照组;第2组,肌瓣组;第3组,复合转移Ad-rhBMP-2组。
2清创:
清除不健康的软组织及坏死的骨组织,保留感染骨质,钻通封闭的髓腔。
3实验过程:
第1组:膝下愿切口进入,适当延长,清创后,缝合创口。
第2组:取膝下切口,局部清创。更换手套及操作器械,自腹股沟中点向上方做“S”形切口,长约4cm,应用显微外科技术,自股动脉起始处找到向上走行的腹壁浅血管神经束,保留肌肉深层,游离血管神经蒂,宽4-5mm,及以腹壁浅血管、神经为蒂的肌瓣(Muscle flap with epigastria superficialis vessel pedicle,MFSAVP),并在与股动脉神经束交界处切断神经束。上下口之间做皮下潜行隧道,将肌瓣经隧道穿至下方的清创口,将肌瓣填充残腔,缝合创口。
第3组:第一步:清创及转移肌瓣,同第2组;第二步:在肌瓣上分3点局部注射Ad-rhBMP-2,注意不损害肌瓣血供,缝合创口。
4术后处理:
3天内给予肌肉注射盐酸罂粟碱预防血管痉挛,按(10mg/kg/d),肌肉注射注射用头孢呋辛(按100mg/kg/d)抗感染知道伤口愈合。并分别在术后7、14、28、42、56天时,测量肛温、检验白细胞分析,以断髓法处死8只大鼠,观察大鼠肌瓣,检查胫骨CR,光镜下观察肌瓣组织病理切片,观测成骨情况。
四实验结果
(一)模型
体重、肛温、局部表现、CR及光镜下所见病理图片示:7天时,骨髓炎急性期,7天后,症状逐渐缓解,14天时,骨髓炎转为慢性期,制造出OBD模型。
(二)治疗
第1组伤口愈合时间,比第2、3组时间长。第1组,术后56天时,残腔变小为原来体积的1/2,瘢痕填充缺损腔。第2组:术后56天时,肌瓣骨化,修复OBD。第3组,术后42天时,肌瓣骨化,修复OBD。
五结论
(一)将蘸有3×10~6CFU金黄色葡萄球菌的棉条植入Wistar大鼠胫骨上段骨缺损腔内,并腔内注射5%鱼肝油酸钠,骨蜡封闭,可以造成OBD模型。
(二)将蘸有3×10~6CFU金黄色葡萄球菌的棉条植入Wistar大鼠胫骨上段骨缺损腔内,并腔内注射5%鱼肝油酸钠,骨蜡封闭,7天时,骨髓炎处于急性期;经过应用抗生素7天,骨髓炎转为慢性。
(三)骨髓炎骨缺损区骨质直径(d),骨缺损面积(m),当m＞d×1.5d时,骨髓炎骨缺损很难自愈。
(四)转移局部带血管蒂肌瓣,肌瓣可以骨化修复骨髓炎骨缺损。
(五)Ⅳ途径转移5×10~8PFU/ml滴度的Ad-rhBMP-2诱导肌瓣骨化过程中,无明显排异反应。
(六)Ⅳ途径转移Ad-rhBMP-2至肌瓣,可以诱导肌瓣骨化修复骨髓炎骨缺损。
(七)当转移带血管蒂肌瓣修复骨髓炎骨缺损时,Ad-rhBMP-2可以缩短肌瓣骨化时间。
一Background
Osteomyelitic bone defection(OBD) is very suffering to patients and formidable to doctors.Many patients with the OBDs have to receive amputation.The therapis for the OBD are anti-infection and repair bone defection(BD).These years,there are many articles about repairing the BD by Muscle flap with vessel pedicle(MFVP) and inducing muscle cells to bone cells by Ad-rhBMP-2.There was no article about the course of muscle flap ossification and transferring Ad-rhBMP-2 for curing the OBD.My study is to cure the OBD by the ossification of the MFVP,then transferring Ad-rhBMP-2 accelerating inducing muscle flap ossification.Provide a new effect therapy for the CO.
二Objective
Evaluate and compare the therapy effect of the two ways:①using the technology of microsurgery transferring the MFVP,②combining the technology of microsurgery transferring the MFVP to OBD with geneⅣtransferring the Ad-rhBMP-2 to MFVP.
三Method
(一) Make OBD modals of the Wistar rats.
We put a carbasus with 0.3 ml Staphylococcus aureus(SA)(3×106CFU) and injected 5%SM in the bone defection,and covered by bone wax.14 days later,we got the modals.
(二) The treatment of OBD
1 Grouping
The 120 Wistar rats modals were meanly divided into 3 groups randomly.The 1st group:Control group.The 2nd group:Muscle flap group.The 3rd group:with transferring Ad-rhBMP-2 group.
2 Debriding
Morbid soft tissue and necrotic bone were debrided until we can see the normal hemodiapedesis and infected bone was remained.The containing cavum medulla was penetrated by high speed electrical drill.
3 Operations
The 1st group:we had not any proposal after debriding.
The 2nd group:Before debriding,We did an incision about 4cm from inguina on, finding the superficial abdominal vessel and take the adaptive MFSAVP through a latent tunnel to fill the OBD,then sutured skin.
The 3rd group:The 1st step was transferring muscle flap.It is the same as the 2nd group.The 2nd step was that we injected Ad-rhBMP-2 in the muscle flap from 4 points, and then sutured skin.
4 Post-operation treatments
Viewed the rats condition of all over the body and vulnus timedly everyday.Muscle injection the Papaverine(10mg/kg/d) avoids artery spasm in 3 days and the Cefuroxime (100mg/kg/d) for anti-infection until intention.We executed 8 rats respectively at 7,14,28, 42,56day after theoperation to get the appearance of local tissue condition,the rectal temperature,the leuk-analysis,the tibias DR and pathological section for evaluating the course of ossification.
四Result
(一) Modal
Temperature,lencocyte count,weight,CR and pathological section under light microscope showed at the 7 day after modal making,it is acute osteomyelitis stage,and 14 day after modal making,it is chronic osteomyelitis stage,so we got the modal.
(二) Therapy
The intention time of the 1 st group was longer than that of the 2nd and 3rd.The 1st group:56 days later,bone defection is clear with its verge smooth and bone density higher while its area diminished to 1/2 of it was.The 2nd group at the 56 day and the 3rd group at the 42 day after operation:the OBDs were repaired by the ossification of the muscle flap.
五Conclusion
(一) The Wistar rats tibia acute osteomyelitic bone defection modal was made by injecting Staphylococcus aureus(3×10~6CFU),sodium morrhuate(5%) in bone defection then which was covered by absorbing film 7 days later.
(二) The acute stage of osteomyelitis turned to chronic stage after 7 days effective antibiotic therapy.
(三) The letter "d" means the diameter of the bone which the OBD occurred.The letter "m" means the OBD area.The natural cure is impossible if m＞d×2d.
(四) muscle flap ossification repair OBD after transferred to it with vessel pedicle.
(五) There is no obvious rejection during the ossification course sinceⅣtransferring Ad-rhBMP-2(5×10~8PFU/ml) in muscle flap byⅣway.
(六) Transferring Ad-rhBMP-2 in muscle flap byⅣway,induce it ossification to repair the OBD.
(七) Ad-rhBMP-2 shorten the time of ossification when use the muscle flap with vessel pedicle repairing the OBD.
引文
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[3]侯春林。自体组织移植进展中国修复重建外科杂志。2006;20(4):309-312
[4]于明琨,王永谦。自体颅骨瓣修复颅骨缺损研究进展。中国现代神经疾病杂志。2006;6(6)3:168-173
[5]Mbalaviele G;Sheikh S;Stains JP,et al.Beta-catenin and BMP-2 synergize to promote osteoblast differentiation and new bone formation.Journal of Cellular Biochemistry.2005;94:403-418
[6]Li C;Vepari C;Jin HJ;et al.Electrospun silk-BMP-2 scaffolds for bone tissue engineering.Biomaterials.2006;27:3115-3124
[7]吴宗键,王继芳,卢世璧。诱导成骨的局部基因治疗。中华骨科杂志。2001;12(21)12:760-762
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