Vascularization is the basis of tissue regeneration. To get engineered tissue, it is necessary that timely and enough nutrition be available to the seed cells, especially those within the biomaterials. Therefore reconstruction of blood supply is the key step leading from basic research to clinical application. Thee major methods have been used to reconstruct blood supply: promotion of vessel growth by means of VEGF, combined grafting of vascular endothelial cells and osteoblasts and microsurgical techniques. The microsurgical techniques, such as wrapping around with pedicled fascia flap, wrapping around with vascularized muscle flap and blood vessel implanting, can help reconstruct blood supply when engineered tissue is fabricated. When engineered tissue is used in clinic to repair defects in vicinity, vascularization can be done in the defect area. But when no fascia flap, muscle flap or vessel can be used in the defect area, the local defects can be repaired through reconstruction of vascularized engineered tissue in other areas and through second stage transplantation with microsurgical techniques.

BACKGROUND: The main aspect of the study in the bone histological engineering is how to maintain and improve theosteogenesis of the osteoblasts in vivo and in vitro. The gene transference may provide a new effective method to deal with theproblem.OBJECTIVE: To discuss the effect of the reverse transcription virus mediated human bone morphogenetic protein7(hBMP-7) gene transfection on the proliferation and osteogenetic differentiation of the bone marrow mesenehymal stemcells (BMSCs) of the rabbits.DESIGN:Cells taken as the study object, grouping control, repeat observation andmeasurement.SETTING: Traumatological and othopaedic lab of a medical university hospital.PARTICIPANTS: The study wascompleted in the Traumatological and Othopaedic Lab in the Affiliated Nanfang Hospital of the Southern Medical University from July 2001 to July 2003. Four New Zealand rabbits,whose weights varied from 1.0 to 1.5 kg, were provided without sexlimit by the Animal Experiment Center of the First Military Medical University of Chinese PLA.METHODS:The reverse transcription virus carriersof the hBMP-7 were constructed,and then the BMSCs were transfected by the virus containing target genes. The expression of the hBMP-7 protein was detected with the immunohistochemical method. The cell proliferation, cycle and ALP synthesis were respectively detected with the MTT method,flow cytometer and NPP method.MAIN OUTCOME MEASURES: Primary results: ① the detection results of the cell proliferation. ② the detection results of the ALP.Secondary results: ① the expression of the hBMP-7 protein in the transfected BMSCs. ② the detection results of the cell cycle.RESULTS: After the BMP-7 gene transfection, there was hBMP-7 positive expression in the BMSCs of the rabbits,using the immunohistochemical detection. There was no significant change in the BMSCs proliferation of the rabbits after the hBMP-7 gene transfection ( P ＞ 0.05). Compared with the ALP synthesis of the transfected BMSCs(294. 592 ± 86. 567) nkat/L, there was significant difference in the ALP synthesis of the empty carrier transfected BMSCs(155. 231 ±86.567) nkat/L and the un-transfected BMSCs (160. 866 ±91. 585)nkat/L( F =5. 660, P ＜ 0. 05).CONCLUSION: After the BMP-7 gene transfection, the BMSCs can synthesize and express the extragenous BMP-7. The hBMP gene transfection can promote the differentiation of the BMSCs cultured in vitro into the osteoblasts and can be used as the seed cells in the construction of the histological en gineering bone tissues and in further application.

BACKGROUND: It is a key point to revascularize the tissue-engineered bone during the repairing of large bone defect. Fascia flap is commonly used in clinic to accelerate the blood supply of implant.OBJECTIVE: To observe the feasibility of repairing goat tibia defects with tissue-engineered bone and accelerating revascularization with fascia flaps.DESIGN: Randomized and controlled animal experiment SETTING: Department of Traumatic Orthopaedics, Nanfang Hospital,Southern Medical University.MATERIALS: Totally 36 goats with the body mass of 14.5-15.5 kg of either gender were enrolled.METHODS: This experiment was conducted at the Department of Traumatic Orthopaedics, Nanfang Hospital, formerly the First Military Medical University of Chinese PLA from December 1999 and December 2003.Bone and periosteum defects 20 mm long were made and fixed with plate of left tibia in 36 goats. They were randomly divided into four groups: Group A in which the defects were filled with coral hydroxyapatite (CHAP), Group B I CHAP+ bone marrow stroma cells (BMSc); Group C with fascia flaps;Group D with nothing. Next, the bone regeneration and the revasculariza tion were evaluated. Radionuclide bone imaging was done 2, 4, 8 weeks after operation. After X-ray examination, the index of optical density of Xray films and histology of the implants were analyzed at 4, 8, 12 weeks after operation, and the biomechanical characters were studied 12 weeks postoperatively.MAIN OUTCOME MEASURES: Gross observation and X-ray, radionuclide bone imaging, biomechanical and histological observation RESULTS: Totally 36 goats entered the stage of result analysis. ① Gross observation of the repair sample of bone defects of the animals in each group: there was no osteogenesis postoperatively at each time point in the blank control group . In Group B, at week 8 to 12, there was no obvious osteogenesis and callus formation on the surface of the materials. In Group C,At weeks 8 to 12, bone defects were filled gradually, many bone callus processes were seen on the surface of the materials , centralizing and enwrapping the materials. The osteogenetic process in the Group C were superior to that of theGroup B. ②Examination result with -901/SA PET-CT scanners: It was seen by naked eyes that at weeks 2 to 8 in the Group A,the radioactivity concentration at region of interesting (ROI) of the operation side had obvious increasing trend, and similar trend of changing appeared in the Group B and Group C, but the ROI counts and T/NT value in the Group B were both lower than those in the Group C. The decreasing trend in the Group A was lower than that in the Group B. ③) Radiological results: the osteogenesis volume through measuring absorbance in the order from large to small was Group C, Group B, and Group A[At week 12, they were (4.180±0.192), (3.480±0.453), (2.959±0.682)respectively ].④Biomechanical results: there were significant difference of loading and bending stress in the Group C, Group B and Group A [ The loading was (758.333±88.754), (530.214±65.297), (359.667±60.715)N , respectively; and the bending stress was (13.937±2.199), (10.123±1.243),(6.223±0.945)N/mm2, respectively ].⑤)Histological results: Slices at various time points in the blank control group showed no bone tissue. In the other three 3 groups, with the prolongation of time, the osteagenetic range and quality were in the order of Group C, Group B and Group A.CONCLUSION: The fascia flaps can accelerate the revascularization process in the formation of tissue-engineered bone so that the capability of tissue engineered bone to repair the large bone defects may be enhanced.

Objective To compare four methods of monitoring vascularization of tissue engineered bone in the rhesus so as to find our the best. Methods Twenty-five lower limbs of 13 rhesuses were used in this study to make models of tibial diaphyseal defect of 20mm which were to be fixed with an AO reconstruction plate of 7 holes. The monkeys were randomly divided into five groups according to defect filling materials: group A:?-tricalcium phosphate (?-TCP) and bone marrow stromal cells (BMSCs) and blood vessel bundles; group B:?-TCP and blood vessel bundles; group C:?-TCP and BMSCs; group D:?-TCP; group E: blank. Perfusion weighted MR imaging (PWMR), X-ray, radionuclide imaging and histological examinations were carried out at weeks 4, 8, 12 postop- eratively. The maximum slope rates of the single intensity-time curve (SS_(max)) and values of baseline (Sl_(?))were calculated at the same time points. Transmittances of the X-ray films were assessed. Ratios between isotope counts in region of interest (ROI) were calculated. Chinese ink perfusion and calculation of blood vessel areas were done for histological examinations, Results Compared with other groups, the SS_(max) in group A was the highest at weeks 4, 8, 12 postoperatively. In group A, the SS_(max) at week eight was significantly higher than that at week four (P= 0. 003), and the SS_(max) and transmittance of X-ray were negatively related at week 12 after operation (rs=-0. 892, P=0. 042), but the SS_(max) and blood vessel area were positively related (rs=0. 894, P=0.041)Conclusions PWMR can be a sensitive, quantitative, noninvasive and non-radiant method to monitor vascularization of tissue engineered bone, because SS_(max) of the single intensity-time curve of PWMR can reflect the most accurately the process of vascularization of tissue engineered bone.

BACKGROUND:The poly-lactic acid and its ramifications have many advantages, such as eligible biocompatibility,nontoxicity of degradation product, easy procession and suitable intensity. Thus, they have been widely used in bone tissue engineering.OBJECTIVE: To study the cellular biocompatibility and in vitro adhesion of poly(lactic-co-glycolic acid) (PLGA) scaffold and bone marrow stromal stem cells (BMSCs) so as to provide a basis for preparation a PLGA scaffold that can load many cytokines.DESIGN: Contrast observation.SETTING: Department of Orthopaedics and Traumatology in Nanfang Hospital of Southern Medical University.MATERIALS: The experiment was carried out in Key Laboratory of Tissue Construction and Detection of Guangdong Province from September 2004 to June 2005. One New Zealand healthy male rabbit (2 months old, 1.5-2.0 kg weight)was adopted in this study. Experimental materials: PLGA scaffold was obtained from Institute of Polymer Science in Sun Yat-sen University); beta-tricalcium phosphate (β-TCP) was supplied by AO Company (Switzerland).METHODS: Bone marrow was aspirated from the New Zealand rabbit. Mononuclear cells were harvested using whole bone marrow culturing, then were induced and amplified in the conditional culture medium. BMSCs were inoculated onto the PLGA and β-TCP at a concentration of 1 ×109 L-1, while those in the medium without materials were taken as blank control group. The development of implanted cells and the adhesion between cells and materials were observed with phase contrast microscope and scanning electron microscope. The proliferation and cycle of cells were tested with MTT method and flow cytometry.MATN OUTCOME MEASURES: ①Phase contrast microscope was used to observe the development of cells and the adherence between cells and materials at a fixed time every day. ②Cellular development on days 1, 3, 6 was observed by scanning electron microscope. ③Cellular proliferation was detected by MTT method. ④Alkaline phosphatase (ALP) activity was determined by chemical colorimetry.⑤Flow cytometer test: The effects of PLGA and β-TCP on the cellular cycle, content of DNA and polyploid levels of BMSCs were investigated. The DNA index of the candidate cells was also calculated.RESULTS: ①Phase contrast microscope observation: In the blank control group, cells culture for 7-10 days presented a larger number of shuttle-shaped, and no contact inhibition effect was observed. The time of cells beginning adherence in PLGA group was obviously later than that in β-TCP group. Cells began to develop on the circumambience and surface of the materials, with the prolonging of culture time. Most of the cells were multangular. The cells in both PLGA and β-TCP groups were similar to those in control group in items of cellular development and shape. ②Scanning electron microscope observation: On the seventh day of culture, the cells of control group remained a monolayer and amalgamated to be a patch with multangular-shaped ones increasing. There were substances in granule shape on the surface of the cells and micro-silk links between cells. In PLGA group, the cells After 7 days' cultivation proliferated sharply, and the compressed-shaped ones were inosculated to be a patch through integrations among cells, resulting in a large number of matrixes. While in β-TCP group, the number of cells increased from the 7th day. The cells were combined together to be a monolayer and moved to pores with matrixes creating out of cells.③Cellular proliferation: The number of the cells in each group all increased to some extents. However, there was no significant difference between the PLGA group and the control group, as well as the PLGA group and the β-TCP group (P ＞ 0.05).④ALP activity: The　content of ALP in all the groups enhanced without exception, while the activities between the PLGA group and the Both control group, as well as the PLGA group and the β-TCP group had no significant difference (P ＞ 0.05).⑤Cellular cycle:Both PLGA and β-TCP had slight effects on cellular cycle of BMSCs. The cells in each group were all normal diploid,and no heteroploid cells were discovered.CONCLUSION: This type of PLGA scaffold possesses good cellular biocompatibility, and can be used as a carrier of many factors in bone tissue engineering.

Objective To introduce the transfer of anterolateral thigh flap with iliotibial tract in repair of massive skin and soft tissue defects of the forearm and reconstruction of extension or flexion of the forearm. Methods Eight cases of complicated raw wounds of the forearm were repaired with transfer of anterolateral thigh flap with iliotibial tract. Flexor tendons of 3 cases and extensor tendons of 5 cases were repaired with iliotibial tract. The axial vessel of the flap was used to rebuild blood supply of the hand. Results All the flaps survived completely. A follow-up ranging from 3 months to 2.5 years re- vealed that the reconstructed forearms were good in appearance and soft in texture and restored protective sensation. Affected limbs could perform extension or flexion. Total range of motion (TRM) of the hands was excellent in 6 cases and poor in 2. Conclusion Anterolateral thigh flap with iliotibial tract can not only repair soft tissue defects of the forearm, but also reconstruct the main extension or flexion of the forearm simultaneously.

Objective To discuss an initial clinical application of three-dimensional reconstruction of ilium bone flap. Methods From December 2006 to June 2008, bone defects of 6 patients with frac-tures of femoral shaft and tibia were repaired using vascularized iliac bone flap. After injection of contrast a-gent, CT scan was performed. Amira 4.0 Software was applied to reconstruct personalized three-dimensional structures of the iliac bone flap. The points, lines, and surfaces were marked in the personalized three-dimensional images reconstructed to provide guidance for the actual surgery. Results The person-alized three-dimensional reconstruction of iliac bone flap for the 6 individual patients were successfully used in the actual surgery. The three-dimensional structures of blood vessels, bone and adjacent relationship which had been clearly shown in the reconstructed flaps were confirmed by the actual surgical findings. All the 6 lilac bone flaps survived uneventfully. Conclusion The preoperative three-dimensional reconstruction of lilac bone flap by CT scan, angiography and digital technology can provide a useful aid for actual surgical design and harvest of the flap, minimizing intraoperative injury to blood vessels and enhancing flap survival.

Objective To explore the clinical effects of variable angle locking plate ( VLP ) in the treatment of pediatric subtrochanteric femoral fractures. Methods Between January 2012 and November 2014, 9 pre-school children were treated at our department for subtrochanteric femoral fractures. They were 6 boys and 3 girls, with an average age of 4. 8 years ( from 4 to 6 years ) . By the Seinsheimer classification, 5 cases were of typeⅡB and 2 of typeⅡC and 2 of typeⅢA. The intervals between injury and surgery averaged 3 days ( from 2 to 5 days ) . All of them were treated with open reduction and VLP internal fixation. Results All the wounds healed well without any infection. All the stitches were removed within 12 days. They were followed up for 8 to 26 months ( average, 16 months ) . All the fractures united within 3 months after operation. Follow-ups revealed no plate or screw loosening, or refracture at the same site. According to the Beaty imaging criteria, the early outcomes were all satisfactory. At the final follow-ups, all the children gained normal gait after full-weight rehabilitation. The affected and normal hips are nearly identical in range of motion and muscle strength. All the children recovered their pre-injury status. By the Sanders scoring for function of the affected hip, 7 cases were rated as excellent and 2 as good. Conclusion VLP can be an effective option for treatment of subtrochanteric femoral fractures in preschool children patients.

The process of tissue repair involves a complex tissue response to injury in which growth factors, playing a major role in this process, trigger, control and terminate soakage of inflammatory cells, cells proliferation, secretion of matrix and scars formation by autocrine, paracrine or both. Thus, growth factors can be used to alter the microenvironment of the wounded tissues and to promote their repair. But, there are notable disadvantages in using purified recombination growth factors, 1) the source is so limited that their prices are expensive; 2) the ir half-lives are short and easy to be destroyed by wound proteases; 3) there is no perfect carrier; 4) high initial doses are required but easy to bring toxicity; 5) it is difficult to apply growth factors in deep wounded tissues again and again, their function cannot be played enough accordingly; 6) most of growth factors are the products of recombination. All above-mentioned disadvantages result in a low activity.

Objective:To investigate the clinical significance of preserving supraclavicular nerve in the reduction and fixation of clavicular fracture.Methods:A retrospective study was conducted of the 68 pa-tients with clavicular fracture who had been treated surgically from October 2016 to April 2018 at Department of Orthopedics and Traumatology, Heyuan Hospital of Traditional Chinese Medicine.They were 35 males and 33 females, aged from 25 to 45 years (average, 34.8 years).The supraclavicular nerve was preserved in the re-duction and fixation of clavicular fracture in 32 patients (reservation group) but not in the other 36 ones (control group).The 2 groups were compared in terms of operation time, intraoperative blood loss, fracture healing time; postoperative pain tolerance, feeling (by the British Medical Research Council assessment), shoulder function (by the Constant-Murley scoring) and impact of numbness on life.Results:The 2 groups were compatible due to insignificant differences in the general clinical data between them ( P>0.05).All the 68 patients were followed up for 12 to 26 months (average, 15 months).There was no significant difference between the 2 groups either in intraoperative blood loss or fracture healing time ( P>0.05).The preservation group had significant longer operation time (72.6 min±7.2 min) than the control group (47.3 min±7.4 min), but a significantly lower rate of analgesic usage on the postoperative day [6.3%(2/32)] than the control group [91.7%(33/36)] (both P<0.05).By the British Medical Research Council assessment, the postoperative feeling was rated as S3 or S4 in 93.8%(30/32) of the patients in the preservation group, significantly higher than the 72.2% (26/36) of the control group ( P<0.05).The preservation group was also significantly better than the control group in Constant-Murley scores of the injured shoulder [100.0 (93.5, 100.0) versus 87.0 (81.0, 89.0)] and impact of numbness on life [0 versus 50.0%(18/36)] (both P<0.05). Conclusion:In surgical treatment of clavicular fracture, preservation of the supraclavicular nerve can have a positive effect on reducing postoperative pain in the operative area and impact of numbness on life.