Feasibility of percutaneous gene transfer to an atrophic nonunion in a rabbit

Treatment of atrophic nonunions is a challenge to orthopaedic surgeons. Growth factors potentially are valuable factors for improvement of tissue healing. The use of growth factors, however, is limited by their short half-lives. Gene therapy has the potential to improve the treatment. This study aimed to establish and validate an atrophic nonunion model in a rabbit for the use of a percutaneous in vivo gene therapy protocol. An atrophic tibial nonunion was established in 24 New Zealand White rabbits. Radiologic and histologic followup was for 64 weeks. The rabbit tibias showed no radiologic or histologic signs of healing. In addition, an adenoviral vector carrying a marker gene was injected percutaneously into the nonunion site in 12 rabbits. Expression of the marker gene was assessed for as many as 4 weeks. The percutaneous gene delivery resulted in transgene expression in the nonunion site for as many as 4 weeks. The described model reliably leads to an atrophic tibial nonunion in rabbits. Adenoviral percutaneous gene delivery into the nonunion site is feasible and leads to transgene expression locally for at least 1 month. This study provides investigators with a reliable and reproducible model of an atrophic nonunion.     

VSD结合外固定支架在西藏高原地区胫腓骨开放性骨折分期治疗中应用

目的：探讨负压吸引敷料结合外固定支架技术在西藏高原地区胫腓骨开放性骨折分期治疗中的应用及临床疗效。方法2014年8月-2015年8月收治16胫腓骨开放性骨折患者,其中男性12例,女性4例,年龄19~66岁,平均年龄39.4岁(19~66岁);胫腓骨中上段骨折4例,下段骨折12例。待患者病情稳定后患者生命体征稳定后,无继发性损伤,予行清创、外固定支架固定骨折端,并采用负压吸引敷料覆盖创面或创腔。7 d后更换VSD或予行二期缝合或软组织覆盖。 X线检查明确骨折愈合时间并记录相关并发症。结果本组16例创面均二期愈合,未见感染、皮肤或皮瓣坏死等软组织并发症。本组共随访患者12例(电话通知来院复诊),4例失访,平均随访18个月(12~24个月)。影像学检查明确平均骨折愈合时间为5.5个月(3~7个月),其中骨折一期临床愈合9例(75%),延迟愈合3例(25%)。随访期间除2例出现钉道感染,经保守治疗后治愈,余患者未见深部感染、植皮或皮瓣坏死、畸形愈合、骨不连或骨髓炎发生。结论负压吸引敷料结合外固定支架治疗西藏高原地区胫腓骨开放性骨折,在迅速有效地稳定骨折的同时,能安全有效地封闭创面,缩短二期创面修复时间,促进骨折愈合,减少并发症。     

The influence of fracture etiology and type on fracture healing: a review of 104 consecutive tibial shaft fractures

This study was conducted to determine retrospectively the factors which influence fracture healing and risk for nonunion in patients with tibial shaft fractures. One hundred consecutive patients with 104 tibial shaft fractures and a mean age of 40 (14-85) years were reviewed. Fractures were classified according to the AO classification system. There were 22 open fractures and 52 comminuted fractures. Thirty-eight fractures were caused by high-energy trauma. Fracture pattern, soft-tissue condition, level of energy of the trauma, malalignment, and treatment methods were identified. The influence of these factors on the time of hospital stay and sick-leave, delayed union, and nonunion were calculated. Normal healing occurred in 61 fractures with a mean healing time of 17 weeks, delayed union in 27 with a mean of 35 weeks, and nonunion in 16 with a mean of 69 weeks. The relative risk of developing nonunion in open fractures was 8.2 (CI = 2.9-10.5) and 2.9 (CI = 1.2-3.2) in fractures exposed to high-energy trauma. This study showed that the soft-tissue condition and level of trauma energy are good predictors for the development of complications. Considering these risk factors at an early stage in the planning of treatment might reduce the risk of nonunion. ergy level of the trauma, fracture comminution, initial fracture displacement, treatment method, contamination, and associated injuries will influence fracture healing. Conservative treatment has in general been recommended for undisplaced closed or grade I open fractures caused by low-energy trauma [15, 18, 21]. However, conservative treatment of a tibial shaft fracture means immobilization of the lower leg for a long period of time, especially if healing is delayed. Thus, it is important to consider the risk of healing complications when planning the appropriate treatment method in the early stage after the injury. The purpose of this study was to analyze complications such as delayed union and nonunion and to identify factors which affect the healing of tibial shaft fractures.     

Wound repair by bone marrow stromal cells through growth factor production

We have previously shown that treatment with bone marrow stromal cells (BMSCs) augments the healing of fascial wounds in the rat. However, the biochemical mechanism by which BMSCs improve wound healing was not investigated. Growth factors have been shown to play a key role in repairing damaged tissue. In this study, we investigated whether BMSCs are capable of producing growth factors that play a critical role in healing of the damaged tissue. Growth factor expression in BMSCs stimulated with pro-inflammatory cytokines or wound superfusate was measured by RT-PCR and growth factor-specific quantitative sandwich enzyme-linked immunosorbent assay (ELISA). RT-PCR analysis demonstrated that BMSCs are capable of expressing transforming growth factor beta-1 (TGF-beta1), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF) platelet-derived growth factor (PDGF), keratinocyte growth factor (KGF), fibroblast growth factor (FGF), and hepatocyte growth factor (HGF) constitutively or upon stimulation with LPS, IL-1alpha, or TNF-alpha. Quantitative analysis of growth factor production by ELISA showed that BMSCs do not secrete TGF-beta1, EGF or VEGF in response to uninjured fascia tissue superfusate; however, production of these growth factors is significantly increased when cells were stimulated with wound tissue superfusate. The ability of wound to stimulate growth factor production in BMSCs could be detected as early as day 1 and lasted through day 7 after wounding. Thus, growth factor production by BMSCs in response to wound microenvionment suggests that BMSCs might augment wound healing through the responsive secretion of growth factors that enhance angiogenesis and promote wound repair.     

Repairing of goat Tibial Bone Defects with BMP-2 Gene–Modified Tissue-Engineered Bone

Bone defects larger than a critical size are major challenges in orthopedic medicine. We combined tissue-engineered bone and gene therapy to provide osteoprogenitor cells, osteoinductive factors, and osteoconductive carrier for ideal bone regeneration in critical-sized bone defects. Goat diaphyseal bone defects were repaired with tissue and genetically engineered bone implants, composed of biphasic calcined bone (BCB) and autologous bone marrow derived mesenchymal stem cells (BMSC) transduced with human bone morphogenetic protein-2 (hBMP-2). Twenty six goats with tibial bone defects were divided into groups receiving implants by using a combination of BCB and BMSCs with or without the hBMP-2 gene. In eight goats that were treated with BCB that contained hBMP-2 transduced BMSC, five had complete healing and three showed partial healing. Goats in other experimental groups had only slight or no healing. Furthermore, the area and biochemical strength of the callus in the bone defects were significantly better in animals treated with genetically engineered implants. We concluded that the combination of genetic and tissue engineering provides an innovative way for treating critical-sized bone defects.     

体外冲击波在促进骨愈合中的应用

目的:观察体外冲击波促进骨折愈合、治疗骨不连的疗效。方法:选取22例外伤性骨折愈合延迟、骨不连患者进行体外冲击波冲击治疗,治疗后每4～6周进行X线片复查,观察骨折愈合情况。首次冲击波治疗12周后如骨折处无明显骨痂形成,则进行第2次冲击波治疗。冲击波能量为0.4mJ/mm2,每次脉冲总数为1600～2400次。结果:22例下肢外伤性骨折愈合延迟及骨不连经体外冲击波治疗后,骨性骨痂形成的时间为4～12周,20例有明显骨痂形成,其中17例在治疗后3～6个月内骨折完全愈合。2例胫骨下段骨折冲击波治疗无效。结论:体外冲击波在治疗骨不连,促进骨折愈合方面效果满意,有广阔的应用前景。     

Treatment of tibial defect and bone nonunion with limb shortening with external fixator and reconstituted bone xenograft

Objective: To explore the effect of external fixator and reconstituted bone xenograft (RBX) in the treatment of tibial bone defect, tibial bone nonunion and congenital pseudarthrosis of the tibia with limb shortening. Methods : Twenty patients ( 13 males and 7 females)with tibial bone defect, tibial bone nonunion or congenital pseudarthrosis of the tibia with limb shortening were treated with external fixation, Two kinds of external fixators were used: a half ring sulcated external fixator used in 13 patients and a combined external fixator in 7 patients.Foot-drop was corrected at the same time with external fixation in 4 patients. The shortened length of the tibia was in the range of 2-9 cm, with an average of 4.8 cm. For bone grafting, RBX was used in 12 patients, autogenous ilium was used in 3 patients and autogenous fibula was implanted as a bone plug into the medullary canal in 1 case,and no bone graft was used in 4 patients. Results: All the 20 patients were followed-up for 8 months to 7 years, averaging 51 months. Satisfactory function of the affected extremities was obtained. All the shortened extremities were lengthened to the expected length. For all the lengthening area and the fracture sites,bone union was obtained at the last. The average healing time of 12 patients treated with RBX was 4.8 months. Conclusions: Both the half ring sulcated external fixator and the combined external fixator have the advantages of small trauma, simple operation, elastic fixation without stress shielding and non-limitation from local soft tissue conditions, and there is satisfactory functional recovery of affected extremities in the treatment of tibial bone defects, tibial bone nonunion and congenital pseudarthrosis of the tibia combined with limb shortening.RBX has good biocompatibility and does not cause immunological rejections. It can also be safely used in treatment of bone nonunion and has reliable effect to promote bone healing.     

硫酸钙对小鼠骨髓间充质干细胞分化和迁移的影响

目的 分析不同浓度的硫酸钙（calcium sulfate , CaSO4）对小鼠骨髓间充质干细胞（mesenchymal stem cells , MSCs）分化和迁移的影响,探讨CaSO4促进骨修复的细胞机制。方法 分离小鼠骨髓MSCs,绘制细胞生长曲线,评价不同浓度的CaSO4对细胞生长的影响,用qRT-PCR方法评价CaSO4作用后的成骨相关基因Osterix因子和骨钙素的表达,通过刮痕合拢实验和琼脂糖斑点实验,评估CaSO4对MSCs迁移能力的影响。结果 中浓度（0.8 g/L）和低浓度（0.1 g/L）的CaSO4对细胞生长无明显影响,高浓度（1.5 g/L）的CaSO4抑制了细胞增殖;中浓度的CaSO4在作用了1~4 d后,显著提高了MSCs成骨基因Osterix和骨钙素的表达,也明显促进了细胞迁移;高浓度（1.5 g/L）的CaSO4反而抑制了MSCs分化和迁移。结论 一定浓度的CaSO4能够诱导MSCs成骨分化,使MSCs细胞迁移能力增强;成骨细胞的出现可能是细胞迁移能力提高的原因之一。 关键词： 硫酸钙;间充质干细胞;迁移;骨再生     

加压交锁髓内钉固定加植骨治疗胫骨骨不连的临床研究

目的:探讨加压交锁髓内钉内固定加交锁髓内钉开口处取骨植骨治疗胫骨骨不连的临床疗效。方法:回顾性分析自2008年2月至2010年10月采用加压交锁髓内定内固定加髓内针开口处取骨植骨治疗18例胫骨骨干骨不连,男12例,女6例;年龄31～67岁,平均42岁。受伤至手术时间6～18个月,平均8个月。骨折不愈合11例,延迟愈合7例。术后根据HSS评分系统评价膝功能,采用Tenny和Wiss评分系统评估疗效。结果:术后随访12～36个月,平均18个月,患者切口愈合良好,无感染,无皮肤坏死。全部患者未见骨不愈合、感染、畸形及再骨折发生。骨性愈合时间4～8个月,平均6个月。患者术后1年膝关节功能HSS评分平均(89.97±3.21)分。术后根据Tenny和Wiss评分系统评估疗效,优16例,良2例。结论:采用加压交锁髓内钉内固定加交锁髓内钉开口处取骨植骨治疗胫骨骨折不愈合及延迟愈合,能提高骨折愈合率,避免髂骨取骨带来的并发症,减少患者医疗费用。     

The effects of growth and differentiation factor 5 on bone marrow stromal cell transplants in an in vitro tendon healing model

The effects of growth differentiation factor-5 (GDF-5) and bone marrow stromal cells (BMSCs) on tendon healing were investigated under in?vitro tissue culture conditions. BMSCs and GDF-5 placed in a collagen gel were interpositioned between the cut ends of dog flexor digitorum profundus tendons. The tendons were randomly assigned into four groups: 1) repaired tendon without gel; 2) repaired tendon with BMSC-seeded gel; 3) repaired tendon with GDF-5 gel without cells; and 4) repaired tendon with GDF-5 treated BMSC-seeded gel. At 2 and 4 weeks, the maximal strength of repaired tendons with GDF-5 treated BMSCs-seeded gel was significantly higher than in tendons without gel interposition. However, neither BMSCs nor GDF-5 alone significantly increased the maximal strength of healing tendons at 2 or 4 weeks. These results suggest that the combination of BMSCs and GDF-5 accelerates tendon healing, but either BMSCs or GDF-5 alone are not effective in this model.     

椎间盘镜下微创植骨治疗下肢骨折术后骨不连

目的探讨在椎间盘镜下微创植骨治疗股骨、胫骨骨折术后骨不连的疗效。方法 2004年8月-2008年8月,采用椎间盘镜下植骨治疗股骨、胫骨骨折术后骨不连患者40例。男23例,女17例;年龄20～63岁,平均41.5岁。骨折后初次手术采用股骨带锁髓内钉固定治疗12例,钢板固定16例;胫骨带锁髓内钉固定9例,钢板固定3例。内固定术后10～16个月发生骨不连。X线片示肥大型骨不连24例,正常型3例,萎缩型13例。结果患者手术时间40～80 min,平均61min;出血量80～130 mL,平均100 mL。住院时间6～11 d,平均8.1 d。患者切口均Ⅰ期愈合,无感染及血管、神经损伤等并发症发生。40例均获随访,随访时间10～16个月,平均12.3个月。X线片示40例骨折均愈合,愈合时间4～10个月,平均6.8个月。无患肢疼痛及功能障碍,无内固定失效。结论经椎间盘镜下微创植骨治疗股骨、胫骨骨折内固定术后骨不连是一种微创、有效的治疗方法。     

股骨、胫骨骨折交锁髓内钉固定后骨不连的诊治

目的探讨股骨、胫骨骨折应用交锁髓内钉固定后骨不连的诊断及应用微创内固定系统(LISS)或锁定加压钢板(LCP)治疗其骨不连的临床疗效。方法2003年2月～2004年12月,对7例股骨和胫骨骨折患者髓内钉固定后应用X线或CT扫描观察骨不连情况,并应用LISS或LCP固定 植骨治疗,病程10～49个月,平均23.3个月。结果7例患者获4～16个月(平均9.1个月)随访;骨折均在术后4～6个月牢固连接,平均愈合时间4.7个月,无植入物松动等并发症发生。结论对骨折端较长时间存在骨折线、且骨折局部伴有疼痛症状者要果断进行手术干预,消除骨折端的微动和消灭骨缺损。LISS或LCP因其先进的锁定设计,可有效治疗股骨和胫骨骨不连。     

The role of low-grade infection in the pathogenesis of apparently aseptic tibial shaft nonunion

PurposeAim of this study was to determine the rate of low-grade infection in patients with primarily as aseptic categorized tibial shaft nonunion and lack of clinical signs of infection.MethodsIn a retrospective study between 2006 and 2013, all patients who underwent revision surgery for treatment of tibial shaft nonunion without clinical evidence of infection were assessed. Bacterial cultures harvested during nonunion revision, C-reactive protein (CRP) and/or white blood cell (WBC) values at hospital admission, outcome, and epidemiological data were analyzed.ResultsIn 88 patients with tibial shaft nonunion without any clinical signs of infection, bacterial samples remained negative in 51 patients. In 37 patients, microbiological diagnostic studies after long-term culturing demonstrated positive bacterial cultures whereas after short-term culturing for 2 days only 17 positive cultures were observed. In 12 cases a mixed culture with 2.3 different bacteria on average was detected. Among patients with negative bacterial cultures bone healing was achieved after 13.2 months. Nonunion with positive bacterial cultures required 19 (range 2-42) months until osseous healing (p = 0.009). Furthermore, nonunion with positive bacterial cultures require statistically more surgical revisions to achieve healing (2.9 ± 0.5 vs. 1.3 ± 0.1 additional procedure; (p = 0.003). Hematological studies carried out before surgical intervention did not demonstrate significant differences in CRP values (negative vs. positive cultures: 0.3 (range 0.3-2.8) mg/dl vs. 0.5 (range 0.3-5.7) mg/dl (p = 0.181) and in WBC values (negative vs. positive cultures: 7.4 (range 3.5-11.9) /nl vs. 7.3 (range 3.7-11.1) /nl (p = 0.723). Limitations of this study may include the varying amount of the at least four samples for microbiological diagnostics as well as the circumstance that for diagnosing low-grade infection swabs and tissue samples were included in this evaluation as being equivalent.ConclusionThe pathogenesis of nonunion may originate from low-grade infection even in patients without clinical signs of infection. In addition, nonunion with positive bacterial cultures require statistically more surgical revisions to achieve healing. Therefore, during any revision surgery, multiple bacterial samples are intended to be harvested for long-term culturing. Particularly, in tibial shaft nonunion following Gustilo-Anderson type III open fractures, low-grade infection should be suspected.Trial registration numberDRKS00014657.Date of registration04/26/2018 retrospectively registered     

SDF-1促进BMSCs迁移的研究进展

骨髓中主要包括两种多能干细胞,即造血干细胞(hematopoietic stem cells,HSCs)和骨髓间充质干细胞(bone marrow derived mesenchymal stem cells,BMSCs)。BMSCs具有多种分化潜能,在一定条件下可向多种细胞方向分化,并且具有免疫调节能力。由于取材方便,不会违背伦理问题。因此近年来,其在免疫调节及组织工程学方面的应用有着越来越重要的作用。基质细胞衍生因子-1(stromal cell-derived factor 1,SDF-1)属于趋化因子家族,与其受体CXCR4(C-X-C Chemokine Receptor 4)相互作用后对BMSCs具有明显定向趋化作用,虽然近年来有很多研究对其作用机制进行了探讨,但具体机制仍不十分清楚。本文对近年来SDF-1及其受体对BMSCs的趋化作用机制进行总结,旨在为后续进一步研究提供参考。     

经皮微创锁定加压钢板内固定术的临床应用

目的探讨经皮微创锁定加压钢板内固定术的临床应用及其疗效。方法应用间接复位技术，通过建立骨折两端皮下隧道，采用锁定加压钢板内固定治疗胫腓骨干骨折15例，锁骨骨折12例，肱骨骨折4例。结果所有患随访4～18个月，平均10个月。骨折愈合时间：胫骨10～20周(平均12周)，1例骨折延迟愈合(20周)，无骨不愈合；锁骨6～10周(平均8周)，肱骨8～13周(平均11周)，均无骨折延迟愈合及不愈合。所有病例无感染及内固定失败等并发症。结论经皮微创锁定加压钢板内固定符合生物学固定(BO)原则，有利于骨折的愈合及软组织的修复。     

膜诱导技术结合内固定治疗胫骨感染性骨不连

目的探讨膜诱导技术结合内固定治疗胫骨感染性骨不连患者的疗效。方法回顾性分析2016年1月至2016年12月陆军军医大学西南医院骨科收治的21例胫骨感染性骨不连患者的临床资料。其中,男15例,女6例;平均年龄41(22~58)岁。所有患者均采用膜诱导技术结合内固定治疗。手术分两期进行:一期彻底清创,抗生素骨水泥填塞,钢板内固定;6~8周感染控制后行诱导膜内植骨重建骨缺损,同时更换内固定。记录随访期间患者的骨愈合时间及并发症发生情况。结果平均随访27(24~30)个月。所有患者均获得影像学及临床愈合,影像学愈合时间平均为4.5(4~6)个月,临床愈合时间平均为6(5~8)个月。随访期间有1例患者出现一期清创后感染复发,经过再次清创感染控制后二期重建达到骨愈合。末次随访时,患者下肢功能量表(LEFS)评分平均为70(59~80)分。结论膜诱导技术结合内固定治疗胫骨感染性骨不连患者的临床疗效显著,具有感染复发率低、骨愈合快、愈合率高、功能恢复好等优势。     

Functional bracing in the treatment of delayed union and nonunion of the tibia

We treated 16 delayed unions and 57 nonunions of the tibial diaphysis with a below-the-knee functional brace. In 48 cases, bracing was preceded by fibular ostectomy, and ten patients had an additional bone graft. We were able to follow 67 patients, of whom six (8.7%) failed to respond to treatment. In patients with nonunion, bony healing occurred with a median of 4 months. There was no difference in the speed of healing according to the level of the defect. Shortening of the limb following ostectomy of the fibula had a mean of 3 mm in the delayed-union group and 5 mm in the nonunion group.     

带监测皮岛的胫前动脉穿支骨膜瓣治疗骨不连的临床应用

目的 探讨带监测皮岛的胫前动脉穿支骨膜瓣治疗骨不连的临床效果.方法 应用带监测皮岛的胫前动脉穿支骨膜瓣治疗骨不连25例,总结手术方法和操作要点,观察临床效果.结果 术中切取的骨膜瓣大小适宜,监测皮岛血运良好,术后伤口大部分一期愈合,2例皮岛边缘少许坏死,换药后愈合.25例术后随访10～24个月,平均15.1个月,原骨不连处骨痂生长良好,髓腔再通,骨折线在3～6个月消失,骨折愈合后关节功能恢复均较满意.结论 带监测皮岛的胫前动脉穿支骨膜瓣治疗骨不连,手术成功率高,是治疗骨不连的一种有效可行的方法.     

局部肌瓣转移和自体红骨髓注射治疗胫骨骨折骨不连

目的报告局部转移肌瓣和注射自体红骨髓治疗胫骨骨折不连接的效果。方法对15例伤后10～42个月,平均22个月仍不连接的胫骨骨折实施切开复位、带锁髓内钉固定、髂骨植骨和局部转移肌瓣覆盖,术后12～15d局部注射自体红骨髓治疗。结果随访表明所有病例骨折均得到连接,术后3～11个月(平均4.8个月)愈合,除2例术后出现切口皮缘小块坏死经换药愈合外,创口均一期愈合,没有发生感染和内固定失败。结论在稳固内固定和植骨以外,局部转移肌瓣和注射自体红骨髓治疗胫骨骨折不愈合能取得满意疗效。