Orthopedic trauma of limbs associated with vascular injuries / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To evaluate the treatment for patients with major vascular injuries associated with traumatic orthopedic injuries.</p><p><b>METHODS</b>A total of 196 patients, aged from 4-67 years with the mean age of 29.88 years, had major vascular injuries associated with traumatic orthopedic injuries and were treated in our hospital in a period of 44 years. The most common mechanism of trauma was blunt trauma (67.3%), open injuries accounted for 32.4% and 54.5% of the injuries were located in the lower extremities. The vascular injury frequently happened in the femoral artery (26.7%) and popliteal artery (20.3%). The treatment principle consisted of aggressive resuscitation, Doppler imaging and stable bone internal fixation with subsequent vascular repair and debridement. The vascular repair for injuries included end-to-end anastomosis (80 cases, 39.6%), interpositional vein graft (94, 46.5%), vascular decompression through fracture distraction (18, 8.9%), arterial ligation (6, 3.0%), vein patch (2, 1.0%), bypass graft (2, 1.0%), venous repair including autogenous vein graft (9, 24.3%) and ligation (28, 75.7%) and prophylactical fasciotomy (15, 7.4%). Postoperative amputation was performed in 16 cases (16.3%).</p><p><b>RESULTS</b>No intraoperative death was observed and all fractures united within 6 months. Limbs were salvaged in 180 patients (91.8%). Among these patients, early complications were found in 19 patients (9.7%) and late complications were observed in 8 patients (4.1%).</p><p><b>CONCLUSIONS</b>A well-organized approach, based on a specific treatment principle, not only improves clinical outcome but also does good to excellent functional recovery for patients with severe orthopedic injuries and concomitant vascular lesion.</p>

Three dimensional induction of autologous mesenchymal stem cell and the effects on depressing long-term degeneration of tissue-engineering cartilage / 中华外科杂志

<p><b>OBJECTIVES</b>To induce autologous bone marrow derived mesenchymal stem cell (aMSC) into chondrocyte, and to confirm the effects of 3 dimensional (3D) dynamic inducing in vitro and their long-term animal model repairing in vivo.</p><p><b>METHODS</b>aMSC were separated from rabbits bone marrow aspirates, then respectively experienced 3D dynamic inducing in alginate drops in modified rotating wall bioreactor culture or in two dimensional (2D) inducing (culture flask) for 10 d. The induced cells were harvest and then mixed with fibrin sealant (FS) to repair rabbit knee femoral trochlea cartilage defects model. After 8, 12, 24, 48 weeks animals were euthanized. Gross appearance, histological appearances were examined.</p><p><b>RESULTS</b>Flask culture groups showed a little chondrocyte differentiation, 3D inducing group showed obviously chondrocyte differentiation, improved collagen II and proteoglycan production. For 3D inducing ones in vivo, the cartilage defects were smoothly repaired by white translucent hard tissue with obvious hyaline-like cartilage histological appearance after 8, 12 weeks, and the defects boundary were hard to be identified with hyaline like cartilage with sustained histological appearance and score after 24, 48 weeks. For 2D ones in vivo, the cartilage defects were smoothly repaired after 8 weeks by hyaline like cartilage which showed accelerated degeneration after 24 weeks and lose cartilage performance completely after 48 weeks.</p><p><b>CONCLUSIONS</b>3D dynamic inducing may assist aMSC on differentiating into chondrocyte, improve its long-term in vivo repairing effects, and enlighten its further applications in tissue engineering cartilage.</p>

Repair of the radial defect of rabbit with polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To evaluate the effects of repairing rabbit radial defects with polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with bovine bone morphogenetic protein (bBMP), and find new carriers for growth factors.</p><p><b>METHODS</b>Polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with and without bovine BMP were used to repair the 15 mm radial defect in rabbit. Then the results of radiography, histology, scaffolds degrade rates and bone mineral density (BMD) were appraised to examine the effects at the 12th week.</p><p><b>RESULTS</b>At the 12th week postoperatively, all defects treated with bBMP were radiographically repaired. No radius implanted polyester/tricalcium phosphate scaffolds without bBMP showed radiographic and histological union. At experimental groups, longitudinal alignment of lamellar structure was observed histologically at the 12th week, indicating that remodeling of regenerated bone was complete in different degree. Of the three experimental groups, the bony regeneration and remodeling of callus in poly lactide-co-glycolide/tricalcium phosphate (PLGA/TCP) group was the best. The BMD values were beyond 70% of normal value at the 12th week while the PLGA/TCP scaffolds group was the highest, and no abnormalities were observed in the surrounding soft tissue in all groups.</p><p><b>CONCLUSIONS</b>Polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with bovine BMP can repair a 15 mm radial defect of rabbit. As for the results, the PLGA/TCP scaffold is ideal and better than poly L-lactide-co-D, L-lactide (PDLLA/TCP) scaffold, but the ploy L-lactic acid (PLLA/TCP) is not so good for its low degradation rates.</p>

Compositional variation of fibrous callus and joint cartilage in different internal environments / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To evaluate the compositional variation of fibrous callus in the fracture site and the joint cavity and joint cartilage after being transplanted in the muscle pouch.</p><p><b>METHODS</b>Thirty 2 month old New Zealand white rabbits (weighing 1-1.5 kg) were randomly divided into two groups: a callus transplantation group (Group A, n=15) and a cartilage transplantation group (Group B, n=15). In Group A, closed radius fracture was made and the autologous fibrous callus was transplanted in the right knee joint cavity at 12 days postoperatively. In Group B, the right knee joint cartilage of the animals was transplanted in the autologous back muscle pouches under anesthesia. Then all the animals were killed by overdose anesthetic 3 weeks after transplantation. And the transplanted fibrous callus, the healed bones in the fracture sites and the transplanted joint cartilage were obtained for assessment of compositional variation.</p><p><b>RESULTS</b>Pure fibrous composition was found in the callus at the fracture sites in Group A at 12 days postoperatively. And for 11 out of the 15 animals, the fibrous callus was transformed into cartilaginous tissues after 3 weeks of transplantation, but the fibrous callus was absent in the other 4 animals. The fibrous calluses at the original site and the fracture locus were differentiated into bony tissues. Bony tissue transformation was found in the transplanted joint cartilages in the muscle pouch of all the animals in Group B.</p><p><b>CONCLUSIONS</b>The fracture sites or joint cavity may facilitate callus differentiation in different ways: the former is helpful for osteogenesis while the latter for the development and maintenance of cartilages, and the muscle pouch is inclined to induce the osteogenic phenotype for cartilages.</p>

Morphology research of the rat sciatic nerve bridged by collage-heparin sulfate scaffold / 中华外科杂志

<p><b>OBJECTIVE</b>To observe the treating effect of collage-heparin sulfate after the 10 mm rat sciatic nerve defect was bridged by it.</p><p><b>METHODS</b>A new kind of nervous tissue engineering scaffold was produced by freeze-drying technique from collagen-heparin sulfate. Thirty-two SD rats were randomly divided into A, B, C and D groups. Sciatic nerve defect in group A was bridged by collagen-heparin sulfate. In group B, sciatic nerve was bridged by auto-nerve transplantation. Group C was the blank control group. Animals in group D were normal. And 10 mm sciatic nerve defect was bridged in the experiment. Thirty-six weeks after the operation, the experimental animals were detected by HRP labeled retrograde trace, HE staining, toluidine staining, silvering staining, S100, GAP-43 and NF immunohistological staining, MBP immunofluorescence staining and transmission electron microscope to observe the nerve regeneration inducing effect of this new scaffold.</p><p><b>RESULTS</b>Nine months after operation, the collage-heparin sulfate scaffold was replaced by newly regenerated nerve. The number of HRP labeled spinal cord anterior horn cells and the area of sensation nerve fiber at the posterior horn were similar with that was repaired by auto-nerve. GAP-43, NF and S100 labeled regenerated nerve fiber had passed the total scaffold and entered the distal terminal. The regenerated nerve fibers were paralleled, lineage arranged, coincide with the prearranged regenerating "channel" in the collagen-heparin sulfate scaffold. MBP immunofluorescence staining also proved that the newly regenerated nerve fiber could be ensheathed. In the experimental group, the area of myelinated nerve fiber and the thickness of the myelin sheath had no obvious difference with that of the group repaired by auto-nerve, except that the density of the regenerated myelinated sheath fiber was lower than that of the control group.</p><p><b>CONCLUSION</b>Nervous tissue engineering scaffold produced by collagen-heparin sulfate can guide the regeneration of nerve fibers. The nerve function recovers fine. This kind of material has great application potential.</p>

Repair of the radial defect of rabbit by polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology / 中华外科杂志

<p><b>OBJECTIVE</b>To evaluate the repairing effect of the rabbits radial defects of by polyester/tricalcium phosphate scaffolds prepared by rapid forming technology loaded with bovine BMP, and find a new carrier for growth factor.</p><p><b>METHODS</b>Polyester/Tricalcium phosphate scaffolds prepared by rapid prototyping (RP) technology loaded with and without bovine BMP were used to repair the 15 mm radial defect of rabbit. Then results of radiography, histology, scaffolds degrade rates and bone density were appraised to examine the repairing effects of the scaffolds at 12 weeks.</p><p><b>RESULTS</b>At 12 weeks, all defects treated with bBMP were radiographically repaired. No radii implanted polyester/tricalcium phosphate scaffolds alone showed radiographic and historical union. At experimental groups, longitudinal alignment of lamellar structure was observed histologically at 12 weeks, indicating that remodeling of regenerated bone almost completed, the scaffolds degradation rates were different by 12 weeks, and no abnormalities were observed in the surrounding soft tissue in all groups.</p><p><b>CONCLUSION</b>Polyester/tricalcium phosphate scaffolds prepared by rapid prototyping technology loaded with bovine BMP can repair the rabbits radical defects. As for the effects, the poly (L-lactic-co-glycolide)/tricalcium phosphate (PLGA/TCP) scaffold are ideal and better than poly (L-lacide-co-D, L-lactide)/tricalcium phosphate (PDLLA/TCP) scaffold, but the poly (L-lactic acid)/tricalcium phosphate (PLLA/TCP) is not so good for its low degradation rates.</p>

Effect of type I collagen on the adhesion, proliferation, and osteoblastic gene expression of bone marrow-derived mesenchymal stem cells / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the effects of porous poly lactide-co-glycolide (PLGA) modified by type I collagen on the adhesion, proliferation, and differentiation of rabbit marrow-derived mesenchymal stem cells (MSCs).</p><p><b>METHODS</b>The third generation MSCs isolated from mature rabbits by density gradient centrifugation were cultured at different initial concentrations on 0.3 cm x 1.2 cm x 2.0 cm 3-D porous PLGA coated by type I collagen in RPMI 1640 containing 10% fetal calf serum, while cultured on PLGA without type I collagen as control. The cells adhesive and proliferative behavior at 7, 14, and 21 days after inoculation was assessed by determining the incorporation rate of [(3)H]-TdR. In order to examine MSCs differentiation, the expression of osteoblasts marker genes, osteocalcin (OCN), alkaline phosphatase (ALP), osteopontin (OPN) mRNA, were evaluated by reverse transcription-polymerase chain reaction (RT-PCR), and further more, the cell morphology at 21 days was also observed by scanning electron microscope (SEM).</p><p><b>RESULTS</b>Type I collagen promoted cell adhesion on PLGA. The valve was significantly higher than controls (6 h, 2144 cpm+/-141 cpm vs. 1797 cpm+/-118 cpm, P=0.017; 8 h, 2311 cpm+/-113 cpm vs. 1891 cpm+/-103 cpm, P=0.01). The cells which cultured on PLGA coated with type I collagen showed significantly higher cell proliferation than controls on the 7 th day (1021 cpm+/-159 cpm vs. 451 cpm+/-67 cpm, P=0.002), the 14th day (1472 cpm+/-82 cpm vs. 583 cpm+/-67 cpm, P<0.001) and 21 th day (1728 cpm+/-78 cpm vs. 632 cpm+/-55 cpm, P<0.001). Osteoblasts markers, OCN, ALP, OPN mRNA, were all detected on PLGA coated by type I collagen on the 21 th day, but OCN, OPN mRNA could not be found in controls. Spindle and polygonal cells well distributed on the polymer coated by type I collagen while cylindric or round cells in controls.</p><p><b>CONCLUSIONS</b>Type I collagen is effective in promoting the adhesion, proliferation and differentiation of MSCs on PLGA.</p>

The cellular mechanism of stimulative effects of CollagenⅠon rabbit bone marrow stromal cells(BMSCs) adhesion / 中华创伤骨科杂志

Objective To investigate the stimulative effects of CollagenⅠon the increased adhesion of rabbit bone marrow stromal cells (BMSCs), cytoskeleton actin organization and intracellular free Ca~(2+) concentration. Methods The third generation BMSCs isolated from mature rabbits were cultured at different initial concentrations on cover-slice coated by collagenⅠin RPMI1640 containing 10% fetal calf serum, and cultured on the same kind of cover-slice untreated with collagenⅠas control. The cells adhesive behavior at different times was assessed. Cellular actin organization was described as either typeⅠor typeⅡcells. In general, typeⅠcells are round and represent a preliminary stage of actin assembly, while typeⅡcells are elongated with organized actin fiber network. At the same time intracellular free Ca~(2+) concentration was measured by using calcium fluorescent probe Fluo-3/AM and laser confocal microscope. Results We found more typeⅡcells in BMSCs cultured with collagen typeⅠsix hours after culture than in the control group. At 12 hours 89% of the BMSCs were typeⅡcells, while only 55% were typeⅡcells in the control group. This indicated active cellular actin organization after being modified by collagen typeⅠ. We also found that the BMSCs cultured with collagen typeⅠincreased intracellular free Ca~(2+) concentration in monolayer culture. Conclusions CollagenⅠis effective in promoting the cellular adhesion, which suggests that a kind of internal relationship or cross-talk may exist between cellular actin organization, intracellular free Ca~(2+) concentration and cell adhesion. Further study, however, is needed.

Construction and biological activity study of human osteoprotegerin expressing adenoviral system / 生物工程学报

Using the isolated total RNA from osteosacoma cell line MG63, the cDNA encoding human OPG was amplified by RT-PCR. A recombinant adenoviral vector carrying cDNA of OPG was constructed and OPG expression in mouse myoblast C2C12 cells was confirmed by Western blot and ELISA. The secreted expression of OPG protein persisted more than 6 weeks in vitro, and the growth of C2C12 cells infected by recombinant adenoviral were in good state. Osteoclasts derived from mouse bone marrow cells infected with recombinant adenoviral made less number of TRAP positive cells and resorption pits formed on dentine slices.

Expression of human osteoprotegerin gene in E. Coli and bioactivity analysis of expression product / 中华外科杂志

<p><b>OBJECTIVE</b>To express human osteoprotegerin (OPG) in E. Coli and analyze its bioactivity in vitro.</p><p><b>METHODS</b>Synthetic oligonucleotides were used to amplify human OPG gene by RT-PCR from total RNA of human osteosarcoma cell line MG63. The OPG cDNA coding for 380 amino acid residues was inserted into prokaryotic expression vector pRSET-A, transformed into competent E. Coli BL21, and induced by 0.1 mmol/l IPTG. SDS-PAGE and Western blot were performed to identify OPG-6His fusion protein. After purified by affinity chromatography, 1,000 microg/L or 1,500 microg/L of OPG-6His were added into the mouse bone marrow cells culture medium. The number of tartrate-resistant acid phophatase (TRAP)-positive multinucleated cells and resorption pits were counted to assess the bioactivity of expression products.</p><p><b>RESULTS</b>The sequence of OPG mature peptide encoding cDNA obtained in this experiment was as same as reported. SDS-PAGE showed 24% of total bacterial protein was of OPG-6His fusion protein. Western blot assay demonstrated that the molecular weight of recombinant protein was about 46 KD and could react specifically with human anti-OPG antibody. The mouse bone marrow cells were induced by 1alpha, 25-dihydroxyvitaminD3 (10(-8) mol/L) and Dexamethasone (10(-7) mol/L) to form osteoclastic-like multinucleated cells. 1,500 microg/L of purified OPG-6His protein could decrease the number of resorption pits and TRAP-positive multinucleated cells in vitro (P < 0.05), but it didn't show the same effects when the concentration of OPG-6His fusion protein was of 1,000 microg/L.</p><p><b>CONCLUSIONS</b>Human OPG-6His fusion protein is expressed and purified in E. Coli. The expression products have moderate inhibitory effects on osteoclast differentiation and bone resorption in vitro only when excessive amount of proteins are added into the culture medium, indicating that prokaryotic expression of fuctionalal OPG protein awaits further investigation.</p>

Reciprocal action between BMP-2 and BMP-3 in cultured fibroblast in vitro / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To explore reciprocal action between BMP-2 (bone morphogenetic protein-2) and BMP-3 for better understanding of the mechanism of BMP during bone fracture union.</p><p><b>METHODS</b>rhBMP-2 was added into the cultured fibroblasts with the concentration of 1,200 ng/ml. The expression of BMP-3 in fibroblasts was detected by immunohistochemistry. Eukaryotic expression vector pcDNA3-BMP-3 was transfected into the fibroblasts. After the effective expression of BMP-3 was identified, BMP-2 was also detected by immunohistochemistry in BMP-3 expression cells. The fibroblasts transfected with empty vector pcDNA3 were used as the control.</p><p><b>RESULTS</b>Exogenous rhBMP-2 could promote the expression of BMP-3 in fibroblasts. BMP-3 also could be detected in these cells.</p><p><b>CONCLUSIONS</b>BMP-2 and BMP-3 could reciprocally adjust the expression in fibroblasts.</p>

Repair of radius defect with bone-morphogenetic-protein loaded hydroxyapatite/collagen-poly(L-lactic acid) composite / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To explore the method to repair bone defect with bone-morphogenetic-protein loaded hydroxyapatite/collagen-poly(L-lactic acid) composite.</p><p><b>METHODS</b>18 adult beagle dogs were randomly divided into 3 groups. In Group A, bone-morphogenetic-protein (BMP) loaded hydroxyapatite/collagen-poly(L-lactic acid) (HAC-PLA) scaffold was implanted in a 2 cm diaphyseal defect in the radius. In Group B, unloaded pure HAC-PLA scaffold was implanted in the defects. No material was implanted in Group C (control group). The dogs were sacrificed 6 months postoperatively. Features of biocompatibility, biodegradability and osteoinduction were evaluated with histological, radiological examinations and bone mineral density (BMD) measurements.</p><p><b>RESULTS</b>In Group A, the radius defect healed after the treatment with BMP loaded HAC-PLA. BMD at the site of the defect was higher than that of the contralateral radius. Fibrous union developed in the animals of the control group.</p><p><b>CONCLUSIONS</b>BMP not only promotes osteogenesis but also accelerates degradation of the biomaterials. Optimized design parameters of a three-dimensional porous biomaterial would give full scope to the role of BMP as an osteoinductive growth factor.</p>

Anti-infective reconstituted bone xenograft used for primary bone grafting to repair contaminated defect in the radius in dogs / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>To investigate the effect of anti-infective reconstituted bone xenograft as a primary graft to repair a segmental with severe contamination.</p><p><b>METHODS</b>A canine model of contaminated defect of 1.5 cm in size in the radius was used, in which anti-infective reconstituted bone xenograft or reconstituted bone xenograft was implanted as a primary graft followed by internal fixation. The effectiveness of the two grafting materials in repairing a contaminated segmental defect was compared.</p><p><b>RESULTS</b>The animals which had received implant of anti-infective reconstituted bone xenograft should largely healed defects 6 months after operation while the defects implanted with reconstituted bone xenograft remained unrepaired with bone infection.</p><p><b>CONCLUSIONS</b>Besides its strong osteoinductive and osteoconductive activity, anti-infective reconstituted bone xenograft is highly antibacterial and can be used as a primary graft to repair the severely contaminated segmental defect.</p>

Treatment of tibial defect and bone nonunion with limb shortening with external fixator and reconstituted bone xenograft / 中华创伤杂志(英文版)

<p><b>OBJECTIVE</b>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.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSIONS</b>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.</p>

Biomechanic results of PLGA/TCP scaffold with bovine BMP in treating peri-porous-titanium bone defects in femur of adult rabbits / 中华创伤杂志

Objective To evaluate the feasibility of composite material of bone morphogenetic protein (BMP) and PLGA/TCP in repair of peri-porous-titanium bone defects in adult rabbits.Meth- otis The composite of PLGA/TCP scaffold with bovine BMP was made and implanted in distal bone de- fects peri-porous-titanium in femur of adult rabbits.The effect of BMP with PLGA/TCP on peri-prosthesis was evaluated by means of scanning electron microscope (SEM),energy dispersive X-ray analysis (EDX) and biomechanics.Results BMP with PLGA/TCP showed obvious peak of Ca and P of EDX in the pores of titanium in experiment group six weeks after operation and higher than that in control group (P＜0.05).Push-out test demonstrated that the bonding strength between the composite of HA coating/ porous titanium and bone was increased significantly with time (P＜0.05).In experiment group,at 6 and 12 weeks,peri-porous-titanium had higher shearing force compared with control group (P＜0.05). Conclusion BMP loaded with PLGA/TCP is a promising bone graft for bone defects in revision arthro- plasty,as indicates that bone induction of BMP plays an important role in biological stabilizaiton.