Vascularized osteomyocutaneous allografts are permissive to tolerance by induction-based immunomodulatory therapy.

Vascularized composite allografts (VCAs) are unique among transplanted organs in that they are composed of multiple tissues with disparate antigenic and immunologic properties. As the predominant indications for VCAs are non-life-threatening conditions, there is an immediate need to develop tolerance induction strategies and to elucidate the mechanisms of VCA rejection and tolerance using VCA-specific animal models. In this study, we explore the effects of in vitro induced donor antigen-specific CD4(-) CD8(-) double negative (DN) Treg-based therapy, in a fully MHC mismatched mouse VCA such as a vascularized osteomyocutaneous as compared to a non-VCA such as a full thickness skin (FTS) transplantation model to elucidate the unique features of VCA rejection and tolerance. We demonstrate that combined therapy with antigen-induced CD4 derived DN Tregs and a short course of anti-lymphocyte serum, rapamycin and IL-2/Fc fusion protein results in donor-specific tolerance to VCA, but not FTS allografts. Macrochimerism was detected in VCA but not FTS allograft recipients up to >60 days after transplantation. Moreover, a significant increase of CD4(+) Foxp3(+) Tregs was found in the peripheral blood of tolerant VCA recipients. These data suggest that VCA are permissive to tolerance induced by DN Treg-based induction therapy.

No red cell alloimmunization or change of clinical outcome after using fresh frozen cancellous allograft bone for acetabular reconstruction in revision hip arthroplasty: a follow up study.

BACKGROUND: Possible immunization to blood group or other antigens and subsequent inhibition of remodeling or incorporation after use of untreated human bone allograft was described previously. This study presents the immunological, clinical and radiological results of 30 patients with acetabular revisions using fresh frozen non-irradiated bone allograft. METHODS: AB0-incompatible (donor-recipient) bone transplantation was performed in 22 cases, Rh(D) incompatible transplantation in 6 cases. The mean follow up of 23 months included measuring Harris hip score and radiological examination with evaluation of remodeling of the bone graft, implant migration and heterotopic ossification. In addition, all patients were screened for alloimmunization to Rh blood group antigens. RESULTS: Compared to the whole study group, there were no differences in clinical or radiological measurements for the groups with AB0- or Rh(D)-incompatible bone transplantation. The mean Harris Hip Score was 80.6. X-rays confirmed total remodeling of all allografts with no acetabular loosening. At follow up, blood tests revealed no alloimmunization to Rh blood group donor antigens. CONCLUSIONS: The use of fresh frozen non-irradiated bone allograft in acetabular revision is a reliable supplement to reconstruction. The risk of alloimmunization to donor-blood group antigens after AB0- or Rh-incompatible allograft transplantation with a negative long-term influence on bone-remodeling or the clinical outcome is negligible.

The expression and distribution of xenogeneic targeted antigens on porcine bone tissue.

OBJECTIVE: The objective of this study was to investigate the distribution of α-galactosyl (α-Gal), major histocompatibility complex (MHC)-I, and MHC-II antigens on adult porcine bone tissue. METHODS: Distribution of α-Gal, MHC-I, and MHC-II antigens on porcine bone tissue were observed using immunohistochemistry. RESULTS: α-Gal, MHC-I xenogeneic antigens were extensively observed on the surface of bone marrow cells, osteocytes, osteoblasts, and Harversian canals; MHC-II antigens were mainly expressed on bone marrow cells. CONCLUSION: α-Gal, MHC-I, and MHC-II are the main xenogeneic antigens that must be deleted to avoid xenogeneic immune reactions against bone xenografts.

Evaluation of a novel reconstituted bone xenograft using processed bovine cancellous bone in combination with purified bovine bone morphogenetic protein.

BACKGROUND: Xenogeneic grafting represents an alternative to autogenous grafting in osseous reconstruction and exhibits many beneficial properties. However, the usefulness of xenogeneic bone relies on necessary processing procedures for removing antigens and viruses, and preserving biological activities simultaneously. By chemical treatment of bovine cancellous bone to make it an antigen-free scaffold, and extraction of bone morphogenetic protein (BMP) from bovine cortical bone, followed by recombination of the scaffold with the BMP, we developed a new grafting material, reconstituted bone xenograft (RBX). METHODS: In this study, scanning electron microscope and energy dispersive X-ray were first employed to observe the structure and components of RBX. Then the biomechanical property was evaluated by applying compression in a materials testing machine. Subsequently, the immunologic evaluation was performed by measuring galactose-alpha-1,3-galactose (α-gal) epitope in vivo and proinflammatory cytokine (TNF-α) secreted by human monocytic cell line (THP-1) in vitro. Finally, this RBX was implanted into segmental radial defects in a rabbit model, and its ability to treat large bone defects was specifically evaluated. RESULTS: Although the compressive strength of RBX was 10% lower than that of unprocessed bovine cancellous bone (UBCB), the basic porous structure and natural components were still kept in this composite. The α-gal xenoantigen level was significantly lower in RBX (P < 0.05) compared with UBCB. Moreover, the TNF-α level was significantly (P < 0.05) reduced compared with UBCB when THP-1 was exposed to RBX. On the other hand, RBX appeared to induce cartilage formation from immature cell populations and resulted in osteogenesis through endochondral-like ossification from 4 to 12 weeks in repairing segmental bone defects. CONCLUSIONS: These results demonstrate that RBX, with its natural microstructure and components, certain mechanical strength and strong osteoinductivity without evoking immune rejection, has significant potential for the treatment of bone defects.

Mecanismos envolvidos na resposta imune e inflamatória frente à implantação de membrana de cortical óssea bovina no tecido subcutâneo de camundongos: caracterização histomorfométrica, imunoenzimática e molecular / Mechanisms involved in immune and inflammatory response to implantation of bovine bone cortical membrane in subcutaneous tissue of mice: characterization histomorphometric, imunoenzimatic and molecular

Os avanços relacionados à ciência dos biomateriais e engenharia tecidual buscam esclarecer os mecanismos envolvidos na resposta biológica associada ao uso desses dispositivos e sua interação com o sistema imune. Neste estudo, avaliou-se a resposta imune e inflamatória desenvolvida em camundongos frente à implantação de membrana de cortical óssea bovina no tecido subcutâneo, em implantação única e sequencial de duas membranas, assim como os possíveis mecanismos envolvidos no processo de reconhecimento e reabsorção desse biomaterial, de acordo com análise histomorfométrica, enzimática e molecular. Após a implantação da membrana, sinais de reabsorção que antes eram notados em pontos isolados, aos poucos se unem até sua completa degradação, observada somente após 15 dias. Todo o processo de reabsorção da membrana é acompanhado por uma reação inflamatória de magnitude moderada, seguida pelo declínio do número de leucócitos, surgimento de células gigantes multinucleadas e formação de uma cápsula de tecido conjuntivo fibroso. A cinética de TNF-α e MMP-2, MMP-8, MMP-9 e MMP-13 apresentou um padrão de produção decrescente, entretanto os níveis dos inibidores de metaloproteinases (TIMPs) e TGF-β parecem atuar de forma inversa. A velocidade de reabsorção após duas implantações consecutivas da membrana foi maior quando comparada ao grupo de animais que sofreu apenas uma implantação, porém os resultados do teste de hipersensibilidade do tipo tardia (DTH) demonstraram que a membrana é biocompatível, pois não elicitou resposta imunológica exacerbada após uma segunda implantação, confirmando então a natureza não imunogênica desse biomaterial. Finalmente, os animais CD14KO e MyD88KO apresentaram uma reabsorção mais lenta da membrana implantada, quando comparados aos animais C57Bl/6 (WT)...

Advances related to the biomaterials science and tissue engineering seek to clarify the mechanisms involved in the biological response associated with the use of those devices and their interaction with the immune system. This study evaluated the inflammatory and immune response developed in mice after implantation bovine bone cortical membrane in subcutaneous tissue, in both, unique and sequential implantation of 2 membranes, as the mechanisms involved in this biomaterial recognition and resorption process, on regards to histomorphometric, enzymatic and molecular analysis. After membrane implantation, previously observed signs of resorption in isolated spots, gradually unite until their complete degradation after 15 days. The whole membrane resorption process is accompanied by a moderate inflammatory reaction, followed by a decline in the leukocytes number, appearance of multinucleated giant cells and formation of a capsule of fibrous connective tissue. The kinetics of TNF-α, MMP-2, MMP-8, MMP-9 e MMP-13 showed a pattern of decreasing production, however, levels of metalloproteinases inhibitors (TIMPs) and TGF-β seem to act in reverse way. The resorption rate after two successive membrane implantations was higher when compared to the group which suffered only one implantation, however, the results of delayed test hypersensity (DTH) demonstrated that the membrane is biocompatible, that is, it does not elicited too high immune response after a second position, confirming the non immunogenic nature of this biomaterial. Eventually, CD14KO and MyD88KO strains showed a slower membrane resorption when compared to animals C57Bl/6, demonstrating that the CD14 and MyD88 molecules are involved in biomaterial recognition and play an important role in bovine cortical bone membrane resorption process, indicating that PAMPs and/or DAMPs are involved in biological response generated by this biomaterial.

[Bone allografts and the functions of bone banks]. / Aloinjertos óseos y la función del banco de huesos.

The use of bone grafts is a common practice in musculoskeletal surgery to provide mechanical stability where there is a defect and it allows skeletal reconstruction. Classically auto and allografts have been used. The latter are the choice in large, complex defects. Allografts can be transplanted despite cell death, have osteoconduction and osteoinduction capacity, low antigenicity and biomechanical properties similar to the original bone. They can be obtained from living and death donors. They are stored by cryopreservation and lyophilization in entities called bone banks. This is a review about bone allografts and the organization and function of the bone banks.

Aloinjertos óseos y la función del banco de huesos / Bone allografts and the functions of bone banks

The use of bone grafts is a common practice in musculoskeletal surgery to provide mechanical stability where there is a defect and it allows skeletal reconstruction. Classically auto and allografts have been used. The latter are the choice in large, complex defects. Allografts can be transplanted despite cell death, have osteoconduction and osteoinduction capacity, low antigenicity and biomechanica lproperties similar to the original bone. They can be obtained from living and death donors. They are stored by cryopreservation and lyophilization in entities called bone banks. This is a review about bone allografts and the organization and function of the bone banks.

Avaliação clínica, histológica e imunológica de enxertos ósseos alógenos fresco-congelados utilizados como técnica na preservação de rebordo alveolar pós-extração / Preservation of alveolar bone in extraction sockets using fresh-frozen bone allograft: a clinical, histological and immunological study

O osso alógeno fresco-congelado (FFBA, do inglês fresh-frozen bone allograft) é uma alternativa para os procedimentos cirúrgicos de enxerto ósseo, principalmente na preparação do rebordo alveolar para a instalação de implantes osseointegráveis. No entanto, existem alguns paradigmas que envolvem a relação entre resposta do sistema imunológico à aloantígenos presentes no enxerto e o seu comportamento clínico. Procurando entender essa relação, o FFBA foi avaliado como enxerto para preservar o rebordo alveolar pós-extração. Os resultados mostraram que embora tenha ocorrido uma redução estatisticamente significante na altura, espessura e volume do rebordo entre a avaliação inicial e final, essa redução não foi clinicamente significante, permitindo a instalação de implantes osseointegráveis. Em adição, as análises histológicas sugerem um bom comportamento do enxerto, com ausência de reação do tipo corpo estranho e formação de novo osso em todos os sítios analisados. Ao analisar o comportamento da resposta imune, os resultados mostraram que a injeção intradérmica de aloantígenos presentes no FFBA, não induziu uma reação de hipersensibilidade tardia nos pacientes após 4 meses do enxerto. Além disso, os monócitos do sangue periférico (PBMCs) dos pacientes não proliferaram frente aos aloantígenos in vitro. No entanto, os dados também demonstraram que os aloantígenos aumentam a produção de IL-2 e IFN-, mas não alteram a produção de IL-4 e IL-10, por PBMCs dos pacientes. Ao avaliar a relação entre a produção dessas citocinas e o comportamento clínico do enxerto, os dados mostram que existe uma correlação estatisticamente significante entre a produção de IL-2 in vitro e a redução (em %) da altura do rebordo alveolar, embora essa redução não tenha sido clinicamente significante. De fato, a presença de aloantígenos no FFBA não é suficiente para sua contraindicação como material de enxertia.

The fresh-frozen bone allograft (FFBA) is an alternative to surgical procedures of bone grafts, mainly in the preservation of alveolar ridge prior the installation of osseointegrated implants. However there are paradigms that surround the relation between immune response to alloantigens present inside the graft and the clinical response of the graft. An attempt to understand this relationship, the FFBA was evaluated as a graft to preserve the alveolar ridge post-extraction. The results show a statistically significant reduction in height, thickness and volume of the ridge between the initial and final examination, however this reduction was not clinically significant. The ridge preservation allowed implant installation and osseointegration. In addition, histologic analysis suggests a good performance of the graft with no foreign body reaction and formation of new bone at all sites. In analyzing the behavior immune response, the results showed that stimulation with alloantigens present in bone allograft induced no delayed hypersensitivity reaction in vivo. Additionally, periphery blood mononuclear cells (PBMC) from patients no proliferate in response to alloantigens in vitro. However, the data also demonstrated that the alloantigens increase IL-2 and IFN- production, but no IL-4 and IL-10 production, by PBMCs from patients. When evaluate the relation between the cytokines production and clinical parameters, the results demonstrate that there statistically significant correlation between IL-2 production in vitro and ridge height changes (%), although this clinical parameter is not clinically significant. In fact, the alloantigens in FFBA are not sufficient for its contraindications as grafting material.

Particulate gold as an anti-inflammatory mediator in bone allograft---an animal study.

Morselized allograft is widely used when increased bone stock is needed in implant surgery. Gold ions liberated from metallic gold surfaces act in an anti-inflammatory manner by inhibiting cellular NF-κB-DNA binding and suppressing I-κ B-kinase activation. This study investigated the effect of 45-63 µm sized gold particles mixed in morselized allograft. It was hypothesized that bio-released gold ions would influence allograft reabsorption, increase mechanical stability, and further stimulate osseointegration. A pair of 10 mm long implants surrounded by a 2.5-mm gap was inserted in proximal part of each humerus in 10 sheep. Each gap was filled with morselized allograft with 1.29 mg gold particles or nothing. Observation time was 12 weeks. The gold ion liberation was visualized by autometallographic tracing and showed liberation of gold ions. Biomechanical push-out tests and stereological histomorphometric analyses showed no statistically significant differences in the two groups. Although particulate gold was primarily observed surrounded by bone marrow tissue, no obvious clinically relevant short-term effects could be measured using gold as an anti-inflammatory mediator. These findings show that the released gold ions have only influenced cells adjacent to the particles without influencing the fixation and illustrates gold ions' limited field of effect. We suggest a new design for orthopedic implants by introducing gold dots on the prosthesis surface. This aims at suppressing the inflammatory foci along the implant-bone zone and reduces the risk of chronic inflammation before aseptic loosening without affecting bone remodeling. This implant model will be investigated in further studies.

Biological therapy of bone defects: the immunology of bone allo-transplantation.

IMPORTANCE OF THE FIELD: Bone is one of the most transplanted tissues worldwide. Autograft is the ideal bone graft but is not widely used because of donor site morbidity and restricted availability. Allograft is easily accessible but can transmit infections and elicit an immune response. AREAS COVERED IN THIS REVIEW: This review identifies all in vitro and in vivo evidence of immune responses following bone transplantation and highlights methods of improving host tolerance to bone allotransplantation. WHAT THE READER WILL GAIN: In humans, the presence of anti-HLA specific antibodies against freeze-dried and fresh-frozen bone allografts has been demonstrated. Fresh-frozen bone allograft can still generate immune reactions whilst freeze-dried bone allografts present with less immunogenicity but have less structural integrity. This immune response can have an adverse effect on the graft's incorporation and increase the incidence of rejection. Decreasing the immune reaction against the allograft by lowering the immunogenic load of the graft or lowering the host immune response, would result in improved bone incorporation. TAKE HOME MESSAGE: It is essential that the complex biological processes related to bone immunogenicity are understood, since this may allow the development of safer and more successful ways of controlling the outcome of bone allografting.

Impact of freezing on immunology and incorporation of bone allograft.

With an increasing clinical use of deep frozen allograft for bone reconstruction, it is important to understand the immunological and biological events of allograft incorporation. In this study, we have investigated the impact of deep freezing on immunology and biopotency for incorporation of bone allografts. Deep frozen bone grafts matched or mismatched for major histoscompatibilty complex (MHC) were implanted in an 8-mm segmental defect in the tibia in rats. The construct was stabilized with intramedullary nailing. The immune response was evaluated by determination of serum antibody against the grafts MHC molecules at day 1 and after 2 and 4 months. Incorporation of the graft was compared with fresh syngeneic grafts and assessed with the use of conventional radiography, biomechanical testing and measurement of bone mineral content and density after 4 months. The analyses revealed no antibody responses in the rats that received grafts from donors differing at histocompatibility loci, and at 4 months the frozen grafts showed an overall reconstruction that was not significantly different from the fresh grafts. This study indicates that in the long run there are no significant consequences; either immunological or biomechanical, of the use of deep frozen allogenous bone as compared to fresh autogenous bone grafts in this animal model.

Vascularized bone grafts within composite tissue allotransplants can autocreate tolerance through mixed chimerism with partial myeloablative conditioning: an experimental study in rats.

BACKGROUND: Certain composite tissue allotransplants contain vascularized bone grafts (e.g., hand/forearm composite tissue allotransplants). The authors investigated the role of the vascularized bone graft within the hind-limb osteomyocutaneous flap in inducing tolerance. METHODS: Brown Norway and Lewis rats were used as composite tissue allotransplant donors and recipients, respectively. Experimental groups were as follows: group I, syngeneic controls (Lewis to Lewis); group II, allogeneic controls (both received no pretransplantation total body irradiation); and groups III, IV, and V, which received 5 mg of antilymphocyte globulin administered intraperitoneally (on days -1 and 10), 1 mg/kg of tacrolimus administered intraperitoneally (on days 0 to 10), and total body irradiation (600, 400, and 200 cGy, respectively, 1 day before composite tissue allotransplantation). Each Lewis rat in groups II, III, IV, and V received a composite tissue allotransplant on day 0 in the form of a Brown Norway hind-limb osteomyocutaneous flap. Different donor hematopoietic cell lineages in recipients' peripheral blood were assessed by flow cytometry on posttransplantation days 15, 30, 60, 90, 120, and 150. Secondary Brown Norway skin grafts in recipients with composite tissue allotransplant acceptance were performed at 150 days after transplantation. RESULTS: Allotransplanted hind-limb osteomyocutaneous flaps produced chimerism and donor multilineage hematopoietic cells in groups III, IV, and V; composite tissue allotransplant acceptance rates in these groups were 37.5, 16.7, and 0 percent, respectively. Graft-versus-host disease occurred in 62.5 percent of group III recipients. Recipients with acute graft-versus-host disease produced a higher percentage of donor T cells compared with composite tissue allotransplant-accepting recipients (p < 0.05). Secondary Brown Norway skin graft acceptance in composite tissue allotransplant-accepting recipients confirmed durable tolerance. CONCLUSION: Vascularized bone grafts contained within composite tissue allotransplants can autocreate chimerism and tolerance in rats with partial myeloablative tacrolimus-based conditioning.

Dissociation between peripheral blood chimerism and tolerance to hindlimb composite tissue transplants: preferential localization of chimerism in donor bone.

BACKGROUND: Mixed chimerism induces donor-specific tolerance to composite tissue allotransplants (CTAs). In the present studies, we used a nonmyeloablative conditioning approach to establish chimerism and promote CTA acceptance. METHODS: Wistar Furth (RT1A(u)) rats were conditioned with 600 to 300 cGy total body irradiation (TBI, day-1), and 100 x 10(6) T-cell-depleted ACI (RT1A(abl)) bone marrow cells were transplanted on day 0, followed by a 11-day course of tacrolimus and one dose of antilymphocyte serum (day 10). Heterotopic osteomyocutaneous flap transplantation was performed 4 to 6 weeks after bone marrow transplantation. RESULTS: Mixed chimerism was initially achieved in almost all recipients, but long-term acceptance of CTA was only achieved in rats treated with 600 cGy TBI. When anti-alphabeta-T-cell receptor (TCR) monoclonal antibody (mAb) (day-3) was added into the regimens, donor chimerism was similar to recipients preconditioned without anti-alphabeta-TCR mAb. However, the long-term CTA survival was significantly improved in chimeras receiving more than or equal to 300 cGy TBI plus anti-alphabeta-TCR mAb. Higher levels of donor chimerism were associated with CTA acceptance. The majority of flap acceptors lost peripheral blood chimerism within 6 months. However, donor chimerism persisted in the transplanted bone at significantly higher levels compared with other hematopoietic compartments. The compartment donor chimerism may be responsible for the maintenance of tolerance to CTA. Long-term acceptors were tolerant to a donor skin graft challenge even in the absence of peripheral blood chimerism. CONCLUSIONS: Mixed chimerism established by nonmyeloablative conditioning induces long-term acceptance of CTA, which is associated with persistent chimerism preferentially in the transplanted donor bone.

Review article: bone transplantation and immune response.

Bone is the second most common transplant tissue after blood, with the iliac crest autologous graft being most used. Bone transplantation induces osteogenesis to repair bone defects. Despite being the most efficient, autogenous bone requires an additional incision and its supply may be inadequate. Deep-frozen allogeneic bone can be an alternative, but is at risk of microbiological contamination, transmission of unrecognised germs, delayed incorporation, and cellular and humoral immune reactions. Synthetic graft substitutes combine scaffolding properties with biological elements to stimulate cell proliferation and differentiation and eventually osteogenesis. However, they generally lack osteoinductive or osteogenic properties and have various effects on bone healing. We present an overview of bone grafts and graft substitutes in clinical use, and the immune responses to allogeneic bone.

Long-term follow-up after non-myeloablative transplant of bone and marrow in BXSB mice.

We present long-term outcomes of BXSB mice after non-myeloablative bone marrow transplants using major histocompatability complex (MHC)-matched cells. Groups differed in sources of donor lymphocytes or mesenchymal stromal cells (MSC). Unfractionated marrow cells from green fluorescent protein (GFP) transgenic (Tg) mice (BMT group) or from RAG1-/- B6 mice (RAG group) were injected intravenously (i.v.) into irradiated (550 cGy) hosts. As a source of mesenchymal cells, bone chips from GFP-Tg were injected intraperitoneally alone (MSC group) or along with i.v. bone marrow cells (BMT + MSC group). Controls were untreated mice (UnTx) or mice exposed to radiation only (Rad Cont). At 62 weeks post-transplant, surviving mice were harvested for histopathology, flow cytometry and real time polymerase chain reaction (RT-PCR). The mice from BMT + MSC group had the best outcomes for survival rates (71.4% vs. 43.8%), renal scores (2.9% vs. 28.8% glomerular sclerosis) and percent splenic monocytes (4.2 vs. 11.3%) compared with mice from Rad Cont. Improvement in RAG and BMT groups was less prominent but were comparable with one another. Although MSC alone were not sufficient to control the renal pathology, it limited the expansion of CD4(-)CD8(-) T cell populations without a change in Foxp3 expression. The results suggest the importance of the innate immune system in disease pathogenesis and a role for MSC in immunomodulation.

Free bone graft attenuates acute rejection and in combination with cyclosporin a leads to indefinite cardiac allograft survival.

We report on a novel approach aimed at preventing acute vascular rejection (AVR), one of the major unresolved hurdles of clinical transplantation. In a C3H-to-BALB/c heterotopic heart transplant model, we demonstrate that free bone transplantation combined with cyclosporin A suppresses antidonor Ab responses, induces indefinite cardiac allograft survival (>100 days), and preserves graft architecture. In contrast, untreated- or cyclosporin A alone-treated recipients rejected their cardiac grafts on days 7.7 +/- 0.6 and 15.5 +/- 1.1, respectively, with graft histology indicative of AVR. Splenic dendritic cells from nonrejecting recipients expressed low levels of MHC II, CD40, and CD86, reduced ability to stimulate donor cell proliferation, and augmented IL-10 production of responding T cells in vitro. Adoptive transfer of dendritic cells from long-term surviving recipients 1 day before cardiac grafting was able to confer hyporesponsiveness to naive BALB/c recipients of cardiac allografts. To determine whether graft survival was associated with hematopoietic or stromal elements of the transplanted free bone, we administered isolated bone marrow mononuclear cells or free bone that was irradiated to deplete hematopoietic elements. Although bone marrow mononuclear cells had no effect on cardiac graft survival, irradiated free bone transplantation was capable of prolonging graft survival. Most interestingly, the prolongation effect was Ag nonspecific, because third party irradiated bone graft was also effective. Due to the fact that current immunosuppressive approaches are clinically ineffective at preventing AVR, this study provides promise for further investigations of BM components as a means of addressing a currently unmet medical need.

Human tissue allograft processing: impact on in vitro and in vivo biocompatibility.

This work investigates the impact of chemical and physical treatments on biocompatibility for human bone/tendon tissues. Nontreated and treated tissues were compared. In vitro testing assessed indirect and direct cytotoxicity. Tissues were subcutaneously implanted in rats to assess the immunological, recolonization, and revascularization processes at 2-4 weeks postimplantation. No significant cytotoxicity was found for freeze-dried treated bones and tendons in comparison to control. The cellular adhesion was significantly reduced for cells seeded on these treated tissues after 24 h of direct contact. A significant cytotoxicity was found for frozen treated bones in comparison to freeze-dried treated bones. Tissue remodeling with graft stability, no harmful inflammation, and neo-vascularization was observed for freeze-dried chemically treated bones and tendons. Frozen-treated bones were characterized by a lack of matrix recolonization at 4 weeks postimplantation. In conclusion, chemical processing with freeze-drying of human tissues maintains in vitro biocompatibility and in vivo tissue remodeling for clinical application.

[Clinical study of nanometer calcium phosphate ceramic artificial bone].

OBJECTIVE: To study the clinical effects and security of nanometer ceramics artificial bone transplantation to treat the bone defect. METHODS: From March 2005 to November 2007, 32 patients (artificial bone group) with extremity bone defects applied nanometer ceramics artificial bone transplantations, included 19 males and 13 females, aged from 17 to 63 years old (averaged 31.4 years). The other 36 patients (internal fixation group) with extremity bone defects were treated by the internal fixation in the same period, included 21 males and 15 females, aged from 16 to 65 years old (averaged 32.6 years). Ca, P, B-ALP, IgG, IgA, IgM, CIC, C3, SL-2R and CD4+/CD8+ in the peripheral venous blood were measured in the 1st and 2th week and 1st, 3rd, 6th month after operation. All patients were followed up and the limb function was evaluated according to Enneking standard. RESULTS: The wounds of all patients smoothly healed after operation. Every immunological indicators had no significant difference between two groups. Serum calcium and phosphorus content did not significantly increased. Serum B-ALP of all patients were increased after operation, fell to normal levels in the internal fixation group, but remained at a relatively high level in the artificial bone group. All patients were followed-up for from 9 to 24 months (averaged 15 months). All patients get the excellent physical function. CONCLUSION: The artificial bone has no immunogenicity, no rejection,does not affect the blood calcium and phosphorus content, and has higher osteogenic activity. It is affirmed that nanometer ceramics artificial bone is used to treat the smaller bone defect on clinical.

Incorporation and immunogenicity of cleaned bovine bone in a sheep model.

This study was conducted to determine if a novel cleaning process could extract antigenic material from bovine bone thereby improving incorporation. Cleaned bovine xenograft, untreated bovine xenograft and sheep allograft were implanted into the tibia of mature sheep for 12 and 24 weeks. Inflammation, bone integration and immunological reactions were evaluated via standardized assays. Cleaned bovine bone dowels induced significantly lower inflammatory responses (p < 50.05) when compared to traditionally processed xenograft. Bone integration, measured by in situ biomechanics, was not different between cleaned bovine bone and allograft controls (p = 0.96). A transient antibody response was observed for non-treated xenografts although this response abated by 3 months.