Objective A new type of bone graft material was constructed by combining calcium phosphate cement(CPC) with bone morphogenetic protein(BMP). In order to evaluate the feasibility to use this material to repair the segmental bone defect in clinic, the ability of CPC was compared with CPC/BMP in bone defects reconstruction by animal experiments. Methods The model of 15 mm bone defect was established in the middle shaft of the radius in 60 rabbits, of which 30 defects were implanted with CPC/BMP composites, 22 implanted with CPC, and the other 8 rabbits served as control group. The specimens were harvested separately at the end of 2, 4, 8, 16 and 24 weeks after operation. In order to observe the formation of new bone and the degradation process of the material, a series of examinations were carried out including of radiography, histomorphology, serum detection, energy dispersion analysis X-ray(EDAX), scanning electron microscope(SEM),bone density detection, mechanical measurement and inorganic substance detection. The results of CPC group and CPC/BMP group were compared on the same condition. Results All the animals survived after operation, and no reactions of toxicity were found. New bone formation was observed to be increasing significantly in CPC/BMP group with the time of implantation. Only little new bone formed in CPC group and no healing was found in the control group. By the end of 24 weeks, new bone had bridged the gap between the proximal and distal fragments in CPC/BMP group. In histomorphological detection, chondrocytes were found at the 2nd week, and woven bone at 4th week in CPC/BMP group. Remodeling of new lamellar bone and absorption of the composite material were observed at the 16th week, and the mechanical strength of the composite material reached almost to normal level at the 24th week. Calcification was significantly higher in CPC/BMP group than that in CPC group examined by EDAX, new bone density detection and measurement of inorganic substance in specimens. During the repairing process of bone defect, the material degraded while new bone formed, the speed of degradation of CPC/BMP was evidently higher than that of CPC group. Moreover, in the process of CPC degradation, the concentration of calcium in serum increased, and the concentration of phosphate in serum kept unchanged. Conclusion The CPC/BMP composite has great potential in bone defects repairing and could be used as a material for bone graft substitute in clinical patients.

To develop a biodegradable implant with both the ossific and anti infectious abilities. Antigen extracted bovine cancellous bone combined with bBMP as the center carrier was coated with either gelatin or poly ? caprolaton (PCL). They were then compounded with gentamicin. Their in vivo gentamycin releasing characteristics were assayed. There was an initial phase of rapid release of gentamycin within 24 hours, followed by a second phase of sustained but gradual diminution of drug release. Effective release of gentamicin from the PCL encapsulated RBX G LDDS may last 30 days in vivo . There was no significant difference in releasing rate between 30% gelatin and 40% gelatin in statistic. The results showed that RBX gentamycin possessed a fine in vivo gradual release property. Collaborated with its ossification activity, it could be considered as an ideal repair material for open fracture with bone defect.

BACKGROUND: Scaffolds are an important part in bone tissue engineering. However, no perfect scaffolds have been developed for bone tissue engineering yet.OBJECTIVE: To evaluate the repair of rabbit radial defects by poly (L-lactic acid)/tricalcium phosphate(PLLA/TCP) scaffolds prepared by rapid prototyping(RP) technology so as to find a new carrier for growth factors.DESIGN: A completely randomized controlled study was conducted. SETTING: Orthopaedic institute of a military medical university.MATERIALS: The study was conducted in the General Orthopedic Institute,Fourth Military Medical University of Chinese PLA, from May 2001 to February 2002. Twenty clean New Zealand rabbits with body mass of(2.5 ±0. 5) kg for this study were obtained from the Experiment Animal Center of Fourth Military Medical University of Chinese PLA. The animals were divided into experiment group and control group with 10 rabbits in each group.INTERVETIONS: PLLA/TCP scaffolds prepared by RP technology and loaded with or without bovine bone morphogenetic protein (BMP) were used to repair the rabbit radial defects of 15 mm.MAIN OUTCOME MEASURES: Main outcomes: ① microscopic observation results of transplanted materials of the two groups; ② degradation rate of scaffolds. Secondary outcomes: ① gross observation; ② radiographic results; ③ bone density.RESULTS: At week 12, bone defect healing in experiment group was good. X-ray examination showed continuous bone callus and partial molding of different degrees. Degradation rate of scaffolds was 39.6%, and bone density in the defected part reached 70% of the normal level. All the indexes of experiment group were superior to those of control group, and no healing was found in the defected area in control group.CONCLUSION: PLLA/TCP scaffolds prepared by RP technology and loaded with bovine BMP can repair radial defects of 15mm in rabbits.

Myelodysplastic syndromes (MDS) are a heterogenous group of hematologic malignancies characterized by clonal expansion of BM myeloid cells with impaired differentiation. Of particular interest mutations is the recent recognition that genes involved in the regulation of histone function (EZH2, ASXL1,and UTX) and DNA methylation (DNMT3A,IDH 1/IDH2,TET2) are recurrently mutated in MDS,providing an important link between genetic and epigenetic alterations in this disease. Ongoing analysis of the seminal AZA-001study has taught many important lessons in the use of DNA methyltransferase (DNMT) inhibitors.Improved survival in patients with high-risk MDS treated with azacitidine extends to patients with any International Working Group-defined hematologic response.New information on the impact of DNMT inhibitors on the immune system and on stem cells will likely lead to novel uses of these drugs in MDS and other hematologic and nonhematologic malignancies. The immunomodulating drug thalidomide and its derivative lenalidomide have been used in the treatment of MDS,principally in lower-risk MDS.

BACKGROUND:The structure of tissue engineering carrier affects the bio-action of cells greatly.OBJECTIVE: To investigate the biological characteristics of bone marrow stem cells (MSCs) in different concentrations of alginate combined with de-antigen bone xenograft (DBX).DESIGN: Observational trial.SETTING: PLA Institute of Trauma and Orthopedics, the Fourth Military Medical University of Chinese PLA.MATERIALS: Alginate, calcium chloride, MSCs, bone xenograft.METHODS: Bovine cancellous bone was out into cubes, which were degreased, deproteinized and then lyophilized.Cubes in pore size within 300-500 μm were selected for use after ethylene oxide sterilization. The purified sodium alginate was dissolved in DMEM cell culture medium of concentrations as different as 0.5%, 2%, 8% and 16%; 1×1012 L-1 induced MSCs were blended with isopyknic alginate-DMEM and compounded with DBX at a status of 0.5 Mpa negative pressure for 5 minutes in order to make a cell suspension fully fill into the pores of the cancellous bone. Then alginate was crosslinked with 50 g/L calcium gluconic acid for 30 seconds. The complex was put into a CO2 incubator and cultured for 4 days. The gel compound and cell growth in the pores of the complex were grossly observed with an inverted microscope. Status of cell growth in the complex with different concentrations of alginate was observed with scanning electron microscopeMAIN OUTCOME MEASURES: Compound status of alginate and bone xenograft, cell growth status and matrix secretion in compound carries.RESULTS: When the concentration of alginate was 0.25% or 1%, alginate was equally combined in DBX, while that of 4% and 8% only combined on the surface of cancellous bone. After in vitro cultured for 4 days, alginate of 0.25% were broken off from DBX surface. But alginate of 1% was equally combined with DBX pores with cells secreting well in alginate. Development of cells in alginate of 4% was restricted and no cells were seen in alginate of 8%.CONCLUSION: Alginate of 1% is suitable for constructing the carrier of bone tissue engineering with bone xenograft.

Objective To explore the incidence and risk factors of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Methods The clinical data of 72 cases allo-HSCT from Oct 2004 to Dec 2008 were analyzed. Thirteen factors possibly correlated with the development of aGVHD were analyzed. Results aGVHD was developed in 32 cases (44.4 %), in which grades Ⅰ aGVHD was 11.1%, gradesⅡaGVHD was 18.1%, and grades Ⅲ-Ⅳ aGVHD was 15.3 %. The univariate analysis showed that diagnosis, the status of disease, use ATG, conditioning regimen, donor type,ABO blood group disparity between donor and recipient, CD34+ cell number, early engraftment and neutropenic infection, HLA locus were associated with the occurence of aGVHD (P ＜0.1). On the COX regression mode, an increased risk of aGVHD was associated with HLA mismatch (HR =2.58, P ＜0.005), GVHD prophylaxis without ATG (HR =2.94, P ＜ 0.001), and unrelated donor (HR =1.97, P ＜0.01). Conclusion aGVHD is a common complication after allo-HSCT, and HLA mismatch and unrelated donor are independent risk factors for aGVHD.

Objective To evaluate the relationship between the immune rejection and the osteoinductive potential of bone allograft.Methods Allogeneic and syngeneic fresh bone, autolyzed antigen-extracted bone, bone matrix gelatin and demineralized bone matrix were implanted into the muscle of mice, and immunological tests, histological observation and alkaline phosphatase assay were performed.Results Three and 6 weeks after implantation, all kinds of allogeneic implants activated immune rejection, among them, fresh bone induced the most vigorous immune rejection and bone matrix gelatin caused the weakest response. Allogeneic autolyzed antigen-extracted bone, bone matrix gelatin and demineralized bone matrix inhibited proliferation of the lymphocytes in vitro and bone matrix gelatin had the most powerful inhibiting effect. Both allogeneic and syngeneic autolyzed antigen-extracted bone, bone matrix gelatin, and demineralized bone matrix induced heterotopic osteogenesis in vivo and bone matrix gelatin had the best osteoinductive capacity.Conclusion There is a negative correlation between immune rejection to bone allograft and osteoinductive capacity of the graft.

OBJECTIVESTo construct new type of bone graft material by combining calcium phosphate cement (CPC) with bone morphogenetic protein (BMP), and then to detect its osteogenic activity.

METHODSThe surface of CPC and CPC/BMP composite were observed by scanning electron microscope (SEM). CPC and CPC/BMP pellets were separately implanted into the thigh muscle pouches of mice. Samples obtained at different times were tested by histological analysis, SEM, organic substance detection, and alkaline phosphatase (ALP) measurement to observe the induced ectopic bone formation.

RESULTSUnder SEM, the CPC and CPC/BMP composite was found to consist primarily of platy crystals, granular crystals and some small rod-like crystals with micropores about 10-50 microm in size. BMP about 1-5 microm in size was seen like micro globules distributing evenly in the micropores. Newly formed cartilage or bone was not found in the CPC group. In the CPC/BMP group, mesenchymal cells were proliferated and abundant cartilage was found in one week. Woven bone appeared at 2 weeks. New bone formation increased with bone marrow at 4 weeks. At 8 weeks, the implanted CPC/BMP became heterogeneous and a lot of collapsed granules were observed. At the end of 16 weeks, mature lamellar bone appeared and the volume of the implanted CPC/BMP became smaller. One week after implantation, the ALP increased evidently in the CPC/BMP groups and reached the highest level at the 4th week, which was about 168 U/L. The content of organic substance in specimens increased from 22% to 39% by the end of the 16th week, showing the continuous calcification and formation of new bone. SEM also showed that the CPC/BMP composite had good potentiality of ectopic bone induction, and the new bone formed accompanied by the slow degradation of the material.

CONCLUSIONThe results of this study suggested that the CPC/BMP composite could be used as material for bone graft substitute.

Animals ; Bone Cements ; Bone Morphogenetic Proteins ; pharmacology ; Bone Substitutes ; chemical synthesis ; pharmacology ; Calcium Phosphates ; Male ; Materials Testing ; Mice ; Mice, Inbred Strains ; Osteogenesis ; drug effects ; Prosthesis Implantation

OBJECTIVETo assess possible beneficial effect in prevention of osteomyelitis by anti-infective reconstituted bone xenograft (ARBX) in the rabbit.

METHODSA proximal tibia osteomyelitis rabbit model was used in which staphylococcus aureus was injected through a bony window, followed by immediate implantation of three ARBX pellets containing 30 mg of slowly-delivered gentamicin in group A, three pellets of RBX in conjunction with intramuscular gentamicin (30 mg) for 5 days in group B, three pellets of RBX without antibiotic in group C. Specimens were harvested 8 weeks postoperatively for gross observation, radiological, histological and bacteriological evaluation, comparing the three groups with regard to the beneficial effect of the above procedures in preventing osteomyelitis.

RESULTS(1) Bacteria counting, modified Norden scoring, and gross and microscopic evidence for osteomyelitis in group B were less than those in group C (P < 0.01). (2) In group A, bacteria culture and counting yielded 0 values at 8 weeks, while radiologically modified Norden scoring for osteomyelitis gave by far the smallest values among all three groups (P < 0.01) with no evidence of osteomyelitis found in gross and histological examinations.

CONCLUSIONS(1) Conventional systemic administration of antibiotics are reasonably effective in prevention of infection, but the anti-infective effect usually is not strong enough to prevent osteomyelitis when used along with primary bone grafting. (2) Apart from its osteoconductive and osteoinductive effects, ARBX is capable of slowly delivering antibiotics, thus being highly anti-infective when administered locally, so it could be used for primary grafting to repair a contaminated bone defect for effective prevention of osteomyelitis.

Animals ; Bone Transplantation ; methods ; Cattle ; Colony Count, Microbial ; Female ; Gentamicins ; administration & dosage ; Male ; Osteomyelitis ; prevention & control ; Postoperative Complications ; prevention & control ; Rabbits ; Staphylococcal Infections ; prevention & control ; Tibia ; surgery ; Transplantation, Heterologous

BACKGROUND: Alginic acid has a relatively mild gel condition and good biocompatibility, and it has been widely used in bio-tissue engineering.OBJECTIVE: To construct bone tissue engineering scaffolds using alginate gel composite bone xenograft approach, and to observe the cell biological properties and in vivo osteogenic potential in scaffolds.METHODS: The bone marrow was harvested from two 2-week-old New Zealand rabbits, 1 ×10~(-8)mol/L recombinant human bone morphogenetic protein-2 was used to induce bone marrow mesenchymal stem cells. The induced bone marrow mesenchymal stem cells at the second generation were incubated into 1% sodium alginate gel, after cultured for 4 days, the cell morphology in gel was observed by hematoxylin-eosin staining. Bone marrow mesenchymal stem cells at the second generation were divided into simple DMEM gel group and DMEM containing 1% sodium alginate gel group, followed by a culture of 7 days. Then bone morphogenic protein-2 immunohistochemical staining was performed. A total of 24 nude mice were randomly divided into two groups, both sides of the thigh muscle pockets were implanted with bone marrow-derived mesenchymal stem cells/alginate gel/bovine cancellous bone complex as an experimental group, with bone marrow-derived mesenchymal stem cells/bovine cancellous bone as a control group. At 2 and 4 weeks post-operation, the osteogenesis in the composite was observed by histological examination, the percentage area of new bone or cartilage was determined using image analysis system.RESULTS AND CONCLUSION: The bone marrow-derived mesenchymal stern cells in the sodium alginate gel exhibited a well-stacked morphology, they suspended in a gel, showing cell division and mitosis phase. In the simple DMEM gel group and DMEM gel containing 1% sodium alginate group, the immunohistochemical results showed that, cell division and proliferation were normal, with prominence at a variety of forms, large nucleus, and clear nucleolus. The bone morphogenetic protein-2 expression had no significant difference between the simple DMEM gel group and DMEM gel containing 1% sodium alginate group (P>0.05).Scanning electron microscopy revealed that, the alginate gel evenly composited in bovine cancellous bone micropores, cell grew at different planes. Animal experiments showed that there were significant differences regarding the percentage of new bone or cartilage area between the experimental group and control group at 2 and 4 weeks postoperation (P< 0.05). It is indicated that constructing bone tissue engineering scaffolds by using alginate gel/bovine cancellous bone, complies with the ultra-structural principle of tissue engineering scaffolds, can maximize the cell loads, achieve good bio-performance, without adverse affects on the proliferation, osteogenic phenotype and related biological properties of bone marrow-derived mesenchymal stem calls, the in vivo osteogenic efficiency was high.