Allogenic grafting of vascularized bone segments under immunosuppression. Clinical results in the transplantation of femoral diaphyses

Trauma surgery lack, substitute, for the reconstruction of large defects of the long bones. Encouraged by the promising results of bone allotransplantation in animal models, we successfully performed vascularized bone transplantation in humans. Vascularized femoral diaphyses were allogenically transplanted into three patients suffering from chondrosarcoma or post-traumatic osteomyelitis with postoperative immunosuppression. The bone segments were harvested from multi-organ donors and perfused with UW solution. After back-table preparation, the grafts were transplanted into the defect zone. Interlocking devices were used in these operations. Vascular anastomoses were performed in end-to-side technique. The early clinical course of the patients was not free of anatomical, technical, or immunological complications. However, all patients are currently free of malignancy and infection. They are also free of pain and full weight bearing. We conclude that allogenic grafting of vascularized bone segments has the potential to become an alternative for the replacement of large bone defects. Received: 21 July 1997 Received after revision: 17 December 1997 Accepted: 9 January 1998     

异体胎骨移植治疗小儿四肢肿瘤性骨缺损

胎儿骨移植是近几年国内采用的一种新的植骨方法，我院采用胎儿骨移植治疗小儿四肢肿瘤性骨缺损４２例，取得了较好的效果，治疗结果表明胎儿骨免疫排斥反应弱，骨诱导能力强，来源丰富，取材容易，能有效的弥补小儿肿瘤性骨缺损范围大，需要大量植骨，而自体骨可供移植用的骨量较少，取材困难的缺陷。     

异体骨移植治疗骨肿瘤

作者统计了３５年中应用大块异体骨移植的方法治疗骨肿瘤外科手术切除后遗留的大范围缺损１２３例，并对其中资料齐全的６４例病人从疾病种类，发病部位、是否化疗与术后并发症和预后关系方面进行了阐述。该组６４例病人中，平均年龄２９．１岁。疾病种类以骨巨细胞瘤和骨肉瘤为主。手术切除范围以Ｅｎｎｅｋｉｎｇ分期标准为原则。     

同种异体骨移植的研究进展

同种异体骨移植是目前临床广泛使用于治疗各种骨缺损和促进脊柱融合的方法。根据处理方式的不同,同种异体骨可以分为：新鲜异体骨、深冻骨、冷冻干燥骨（fresh frozen allograft, FFA）和脱钙骨基质（demineralized bone matrix, DBM）。同种异体骨与新鲜的自体骨的愈合机制有着本质不同,其主要依靠“爬行替代”实现与宿主骨的融合,因此愈合速度慢、愈合质量差,容易发生骨不连和移植骨骨折,故存在较高的失败率。而复合生长因子、复合间充质干细胞以及红骨髓等方法可以提高异体骨愈合的速度。利用组织工程的原理与方法,采用复合移植的方法使异体骨得到活化,是今后异体骨移植研究与应用的发展方向。     

Tolerance induction following allogeneic vascularized bone marrow transplantation — the possible role of microchimerism

Abstract We have noticed that bone marrow transplanted in a vascularized limb graft, providing a continuous supply of donor bone marrow cells (BMC), may prolong the survival time of a skin graft from the same donor. The question arises whether the microchimerism raised plays a role in the prolonged survival of skin allografts. The aim of the study was to follow the development of microchimerism after allogeneic vascularized bone marrow transplantation (VBMTx) concomitantly with the rejection process of transplanted skin. Brown Norway (BN) rats served as donors and Lewis rats as recipients of VBMTx and free skin flap allografts. A hind limb was transplanted, followed by a full-thickness skin graft on the dorsum. Cellular microchimerism was investigated in recipients of VBMTx and skin grafts in blood, spleen, mesenteric lymph node, and bone marrow with the monoclonal antibody OX27 directed against MHC class I polymorphic RT1 on BN cells and quantitatively analyzed in a FACStar. In the VBMTx group, the free skin flap survived 70 days after weaning off cyclosporine A (CsA). An intravenous infusion of BMC in suspension equivalent to that grafted in the hind limb did not prolong skin graft survival after cessation of CsA therapy. Donor-derived cells could be detected in VBMTx recipients as long 70 days after weaning off CsA but not in recipients of i. v. suspension BMC grafting.     

羟基磷灰石多孔层修饰镍钛合金的组织相容性评价

目的 评价羟基磷灰石多孔层修饰镍钛合会的组织相容性.方法 选取健康成年雄兔24只.随机分为实验组(C组)和对照组(R组).选取左后肢股骨下1/3外侧为植入区,C组植入羟基磷灰石多孔层修饰镍钛合金.R组植入镍钛合金.分别于术后1、2、4、8周处死动物.观察各期骨组织局部反应、骨缺损修复情况.检测肝脏镍离子含量.并用免疫组化染色法测定TGF-β1表达阳性细胞的百分比.结果 R组各期肝脏镍离子含量测定均明显高于C组,且差异有统计学意义(P<(0.05);C组术后1、2、4周TGF-β1表达阳性细胞的百分比明显高于R组,且差异有统计学意义(P<0.05).结论 羟基磷灰石涂层能显著抑制镍钛合金中镍离子的释放;羟基磷灰石涂层能刺激骨组织中TGF-β1的表达.促进骨缺损的修复;羟基磷灰石多孔层修饰镍钛合金具有很好的组织相容性.     

Half-joint allograft transplantation in human bone tumours

Summary Seven patients with malignant or aggressive bone tumours involving the end of a long bone have been treated by wide resection and an allograft providing a half-joint transplantation. The observation period is from 2 to 9 years (average 5 years). The allografts were preserved at –70 °C for a period ranging from 1 week to more than 2 years. Computed tomography was very useful in the preoperative determination of the grade of infiltration tumour growth in bone. The host's own ligaments were used in the reconstruction. Immunosuppression was not used.The fate and metabolism of the allografts were followed by clinical examination, conventional radiographs, angiography, bone scintigraphy, cell-mediated immunity tests, biopsies for histology and by determining the excretion of the urinary products of bone metabolism (duHYPro, dUCa, dUPi). Positive isotope labelling was demonstrated as early as 2 weeks after transplantation and this increased and stabilised at about 6 months. Simultaneously, the collagen/bone matrix turnover (HYPro excretion) showed a slightly raised level for up to 3–6 months, stabilising thereafter indicating a moderate, but prolonged, regenerative activity of the graft. On the basis of these and the histological studies, weight-bearing was gradually started at 6 months, which is earlier than described in previous reports. The patients had knee flexion up to 90° and walked well. Studies on whole-blood cell-mediated immunity showed only slight non-significant changes. The results indicate that the grafts incorporated well with early, and subsequently prolonged, evidence of metabolic activity.

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Résumé 7 malades atteints de tumeurs osseuses malignes ou aggressives situées au niveau de l'extrémité d'un os long ont été traités par résection large et allogreffe, réalisant la transplantation d'une hémi-articulation. La durée d'observation est de 2 à 9 ans (en moyenne 5 ans). Les allogreffes ont été conservées à –70° pendant une période allant d'une semaine à plus de 2 ans. La tomographie computerisée a été très utile pour la détermination pré-opératoire du degré d'infiltration de la croissance tumorale osseuse. Les ligaments du sujet receveur ont été utilisés pour la reconstruction. On n'a pas eu recours à l'immuno-suppression.L'évolution et le métabolisme des allogreffes ont été suivis grâce à l'examen clinique, aux radiographies standard, à l'angiographie, à la scintigraphie osseuse, aux tests d'immunité cellulaire, aux biopsies pour examen histologique et en déterminant l'excrétion urinaire des produits du métabolisme osseux (hydroxyproline) calcium et phosphates). Des réactions isotopiques positives se sont manifestées dès la deuxième semaine après la transplantation, puis ont augmenté et se sont stabilisées vers le 6ème mois. Simultanément, le »turnover« collagène/matrice osseuse (excrétion de l'hydroxyproline) a montré une légère augmentation pendant 3 à 6 mois, indiquant une activité de régénération de la greffe, modérée mais prolongée. Du fait de ces constatations et des résultats des examens histologiques, la reprise de l'appui a été progressivement autorisée à partir du 6ème mois, c'est à dire plus tôt qu'il n'est dit dans les publications plus anciennes. Les malades ont récupéré une flexion du genou atteignant 90° et marchent correctement. L'étude de l'immunité cellulaire sanguine n'a montré que de minimes changements, non significatifs. Ces résultats prouvent que les greffes s'incorporent bien, avec des signes d'activité métabolique précoce et longtemps prolongée.

     

Bone graft substitutes for spine fusion: A brief review

Bone graft substitutes are widely used in the field of orthopedics and are extensively used to promote vertebral fusion. Fusion is the most common technique in spine surgery and is used to treat morbidities and relieve discomfort. Allograft and autograft bone substitutes are currently the most commonly used bone grafts to promote fusion. These approaches pose limitations and present complications to the patient. Numerous alternative bone graft substitutes are on the market or have been developed and proposed for application. These options have attempted to promote spine fusion by enhancing osteogenic properties. In this review, we reviewed biology of spine fusion and the current advances in biomedical materials and biological strategies for application in surgical spine fusion. Our findings illustrate that, while many bone graft substitutes perform well as bone graft extenders, only osteoinductive proteins (recombinant bone morphogenetic proteins-2 and osteogenic protein-1) provide evidence for use as both bone enhancers and bone substitutes for specific types of spinal fusion. Tissue engineered hydrogels, synthetic polymer composites and viral based gene therapy also holds the potential to be used for spine fusion in future, though warrants further investigation to be used in clinical practice.     

生物衍生骨支架材料的组织相容性研究

目的 了解不同处理方法制得的3种生物衍生骨支架材料的组织相容性。方法 将物理化学方法处理制得的复合型完全脱蛋白骨（composite fully deproteinized bone,CFDB)、部分脱蛋白骨（partially deproteinized bone,PDPB)、部分脱钙骨（partially decalcified bone,PDCB)3种材料各10块分别植入兔肌肉内及骨膜下，进行一般观察、血清抗体检测、局部细胞免疫评估、常规HE染色，观测3种材料对机体的毒性作用、免疫效应及骨膜成骨的影响。结果 3种材料均无毒性作用，且能引导周围组织长入材料内；3种材料对骨膜成骨无不良影响，且能促进骨膜形成的软骨或类骨组织钙化形成新骨，并有一定的骨结合能力；3种材料引起机体免疫反应强弱为：PDCB＞PDPB＞CFDB。结论 CFDB、PDPB、PDCB3种天然生物衍生骨材料皆有良好的生物相容性。     

异体骨段复合人工股骨头置换治疗股骨近端骨转移癌

目的：总结股骨近端骨转移癌患者行异体骨段复合人工股骨头置换的手术效果。方法：13例患者术后平均随访11．5个月。结果：术后患者肢体功能改善,手术优良率为84．6％;术后骨不连、假体松动、围手术期死亡各发生1例。结论：异体骨段复合人工股骨头置换术利于软组织修复重建和肢体功能康复,术前需综合评价患者的全身条件及局部转移灶状况。     

Bone graft tuberculosis outbreak in USA: Is it a concern in India?

Globally, 25% of the population is infected with tuberculosis, which poses a leading cause of death worldwide. The transmission of tuberculosis (TB) during organ transplant is reported in the literature whereas only one report has been published on the transmission of TB, during bone allograft transplantation. In the US, in May 2021, an outbreak of TB occurred in patients undergoing spine surgery with bone allograft. This bone graft was retrieved from 80 years deceased donor with latent TB, which was not diagnosed earlier. The recipients were started with a long course of anti-tuberculous drugs. This review narrates the pathway of TB spread among transplant recipients and the strategies to be followed while performing organ or tissue transplantation.     

大段异体骨移植后骨端不愈合的外科治疗

目的 探讨应用自体骨植骨再固定的方法治疗大段同种异体骨移植后骨端不愈合的手术方法和疗效.方法 1994年1月至2006年12月北京积水潭医院所行大段同种异体骨移植患者176例中41例不愈合,22例进行了骨接触端的重新植骨治疗,其中15例临床资料完整.再植骨治疗时年龄15～34岁,平均为24岁.初始疾病为:骨肉瘤5例、骨巨细胞瘤4例、皮质旁骨肉瘤2例、血管内皮瘤2例、骨原始神经外胚瘤(PNET)2例.发病部位:股骨下端7例、肱骨中段3例、股骨中段2例、胫骨上端2例、肱骨上端1例.15例患者中,8例单纯不愈合行植骨术,另7例不愈合并发原内固定断裂行植骨和再固定手术.结果 随访时间18～148个月,平均47个月.15例中13例骨愈合,占86.7%.愈合时间5～20个月,平均13个月.其中8例无原内固定失败者均重新愈合,愈合时间平均为14个月.另7例原内固定失败者5例重新愈合,愈合时间平均为12个月,与上述8例比较愈合时间相似.无感染等并发症发生.2例仍不愈合,最终行人工假体置换术.13例骨愈合的患者MSTS评分平均25.1分,8例无原内固定失败者25.4分,5例原内固定失败者24.6分,评分基本相同.结论 应用自体骨植骨再固定的方法治疗大段异体骨移植后骨端不愈合手术简单,并发症少,愈合率高,再手术后功能影响小,较人工假体置换相比有明显优势,对于大段异体骨移植后骨端不愈合患者,应为首选治疗方法.     

骨髓移植诱导特异性免疫耐受同种皮肤移植的动物实验研究

目的　证实通过动物实验模型的骨髓移植可以诱导同种皮肤移植的免疫耐受。方法　将 114只日本白色家兔和Dutch家兔分为对照组和实验组 ,日本白色家兔作为供体 ,Dutch家兔作为受体。对照组 ,在不使用免疫抑制剂的情况下 ,将 12只日本白色家兔与 12只Dutch家兔行相同面积的背部全厚皮肤互换移植 ,观察其成活时间。实验组 ,将 4 5只日本白色家兔和 4 5只Dutch家兔行全厚皮肤移植的同时行骨髓移植 ,然后将作为受体的Dutch家兔分为A ,B ,C ,D四组 ,分别行非致死量的γ射线全身照射的骨髓细胞移植及同种皮肤移植 ,观察移植皮肤的成活时间。结果　对照组 ,供体与受体移植皮肤的平均成活时间分别为 (12 .0± 1.7)天和 (10 .3± 1.3)天。实验组 ,A ,B ,C ,D四组移植皮肤的平均成活时间分别为 (6 1.0± 7.2 )、(80 .7± 10 .4 )、(78.8± 12 .7)、(88.0± 6 .0 )天。结论　通过骨髓移植导特异性免疫耐受同种皮肤移植的动物实验 ,旨在为临床应用提供了理论基础及可靠依据 ,为同种组织重建提供一个新方法     

Study of Cell Viability and Etiology of Contamination in Decalcified Bone Allograft: A Pilot Study

BackgroundBone allografts can elicit immune responses which is correlated with the presence of Human Leukocyte Antigen (HLA) and cellular DNA. It also has risk of causing occult infection arising out of contamination during its processing and storage. The presence of immunogenic materials like cells, cellular remnants and DNA in a decalcified bone allograft during different phases of processing has never been studied. Present study was conducted to explore- the cell viability using routine Hematoxylin and Eosin, presence of DNA using Feulgen staining and etiology of contamination in decalcified bone allograft during procurement, demineralization and ethanol preservation.MethodsThe harvested bones from patients undergoing hemireplacement/THR/TKR were processed to prepare decalcified bone allografts. The samples during procurement (A), HCL treatment (B) and ethanol preservation (C) were sent for histopathological analysis (number of osteocytes in the maximum density field under 40x and the cells demonstrating presence of DNA on feulgen stain) and microbiological assessment (aerobic/anaerobic/fungal cultures).ResultsHistopathological study demonstrated the presence of osteocytes and other cells like bone marrow, adipocytes, endothelial cells in the decal bone allograft. The average number of osteocytes gradually decreased from 55.47, 9.6, 0.86 in sample A, B, C, respectively. Feulgen staining confirmed the presence of DNA in osteocytes and other cells which decreased both qualitatively and quantitatively in subsequent stages of processing. Rate of contamination demonstrated at the procurement was 6.67% (Staphylococcus aureus). After treatment with HCl (demineralisation), 7.14% of non-contaminated allografts were found contaminated (Staphylococcus epidermidis). None of the remaining 13 non-contaminated allografts showed contamination after storage in ethanol. Overall 13% of the patients had positive cultures on microbiological assessment.ConclusionThe population of osteocytes in the harvested bone reduced significantly after processing with HCl and ethanol preservation. Presence of DNA, demonstrated by using Feulgen staining, was observed in bone marrow cells, adipocytes along with osteocytes which showed quantitative reduction on processing. Hence, antigenicity, conferred by cells and their DNA, reduced significantly after processing of decal bone. Contamination rate of banked decalcified allograft was 13%. Thus, culture and sensitivity tests should be carried out at each step of processing of decal bone allograft.     

Donor skin allograft survival after bone marrow transplantation: Case report and systematic review of the literature☆

Background

We present a case of skin allograft survival in a patient who previously received a bone marrow transplant from the same HLA-matched donor. DNA fingerprinting of skin biopsies showed mixed cellularity originating from the donor and recipient (68% and 32% donor DNA in the allograft skin and the native recipient's skin, respectively). Histologic sections demonstrated both grade 3/4 rejection and graft-versus-host-disease. We have conducted a systematic review in search for other cases of donor skin allograft survival after a bone marrow or hematopoietic stem cell transplantation.

Cardiac allograft vasculopathy: current concepts and treatment

Cardiac allograft vasculopathy (CAV) remains the leading limiting factor of patient and graft survival after the first post-operative year. The pathogenesis involves both immunological and non-immunological factors. Here, we present recent advances and discuss potential preventative and treatment regimens. A review of the current literature of heart transplantation, detailing molecular mechanisms, pharmacological risk factors and novel immunosuppression regimens was performed. Recent findings demonstrate the pivotal role of the endothelium, resulting in release of pro-fibrotic cytokines, recruitment of circulating leucocytes, proliferation of vascular smooth muscle cells, and deposition of extracellular matrix proteins (ECMs). The role of HMG-CoA reductase inhibitors and anti-hypertensives remains controversial, but there is increasing evidence advocating their prophylactic use. We can conclude that novel immunosuppressive agents such as rapamycin, mycophenolate mofetil and FTY-720 are experimental immunosuppressive agents that are undergoing evaluation in clinical trials. The prophylactic use of statins and anti-hypertensive drugs needs to be defined but needs to suggest potential strategies to prolong cardiac allograft survival.     

异体大鼠骨髓单个核细胞胰岛细胞移植治疗糖尿病

目的 观察异体骨髓单个核细胞和胰岛细胞通过肝脏和静脉途径移植后对糖尿病大鼠的治疗作用.方法 密度梯度离心法分离胰岛细胞,淋巴细胞分离液分离骨髓单个核细胞,28只糖尿病大鼠模型随机分为A、B、C、D组,A组在肝脏被膜下多点注射1000个胰岛细胞,B组在体外将1000个胰岛细胞和1×107个骨髓单个核细胞混合后在肝脏被膜下多点注射,C组通过尾静脉注射1000个胰岛细胞,D组在体外将1000个胰岛细胞和1×107个骨髓单个核细胞混合后通过尾静脉注射,移植后于不同时间点尾静脉测定随机血糖,比较不同细胞组合和移植途径之间对糖尿病的治疗作用.结果 A、B组血糖于术后3 d内开始下降,A组血糖可降至正常水平(7.98±2.28)mmol/L,血糖维持正常水平(3.71±0.95)d,B组降至(7.72±1. 75)mmol/L可维持(4.86±1.06)d,静脉移植组血糖于术后4 d内降至正常(7.35±1.40)mmol/L,可维持(7.85±1.46)d,D组静脉注射胰岛于4 d起效(7.00±0.83)mmol/L,血糖可降至正常水平可维持(14.10±1.21)d,各组间血糖随时间变化的趋势及维持正常水平的时间具有统计学意义(P＜0.05).结论 骨髓单个核细胞和胰岛混合细胞通过尾静脉移植对大鼠血糖维持正常时间最长,血糖控制水平最理想.