BACKGROUND:Preliminary studies have showed that vacuum sealing drainage based on wound surface dressing biomaterials is a good method to cover the wound as succedaneous peau when the soft tissuedefects along with open fracture cannot be completely repaired during the first operation. OBJECTIVE:To explore the efficacy of vacuum sealing drainage based on wound surface dressing biomaterials in repair of soft tissue defects of foot and ankle. METHODS:Fourteen patients with soft tissue defects of foot and ankle were treated using free skin graft combined with vacuum sealing drainage technique. Meanwhile, the traditional skin graft after wound dressing changes was applied in another 11 patients. The clinical outcomes were compared between two groups. RESULTS AND CONCLUSION:The transplanted skin in 10 cases of the vacuum sealing drainage group survived. The total survival rate was 71%, and surgical dressing change was applied in the left four patients to finaly cover the wound. By comparison, the transplanted skin in four cases of the traditional group survived. The total survival rate was 54%. To finish the wound, three of the left patients were turned to surgical dressing change and two of them stil needed skin graft operation once more. The total survival rate between the two groups has no statistical significance(P > 0.05). The therapeutic procedure noted that the time waiting for the secondary surgical visit, times for dressing change before the second intervention and the time for final union between the two groups were statisticaly different (P < 0.05). So the vacuum sealing drainage based on wound surface dressing biomaterials may accelerate the repair of soft tissue defects of foot and ankle when using the free skin graft operation.

BACKGROUND:Single-row and double-row suture method are commonly used in the rotator cuff repair. Previous studies have shown that, double-row suture is not better than single-row suture in clinics.
OBJECTIVE:To compare clinical outcomes of single-row suture and double-row suture for rotator cuff repair, and evaluate the difference of therapeutic efficacy between two methods.
METHODS:A computer-based search was performed in the Medline (from January 2003 to February 2014), EMBASE (from January 2003 to February 2014) and Cochrane library (February 2014). According to the inclusion and exclusion criteria, al the randomized control ed studies addressing the outcome of single-row repair and double-row repair techniques were included in this meta-analysis. The methodological quality of each study was judged and a meta-analysis was conducted using Revman5.0. The preoperative and postoperative differences between the Constant score, American Shoulder and Elbow Surgeons (ASES) score, University of California, Los Angeles (UCLA) score, the re-rupture rate and the muscle strength were compared. The forest chart was used to compare the data between two groups, and the funnel plot was finished to detect the publication bias.
RESULTS AND CONCLUSION:A total of 10 randomized control ed trials (Levels I, II) were included. Meta-analysis showed that, there was no statistical y significant difference in the Constant, ASES and UCLA scores in the double-row group and the single-row group before and after treatment. In the postoperative fol ow-up, double-row group had a lower re-rupture rate and a higher abductor muscle strength than single-row group. When the rotator cuff tear was less than 3 cm, double-row group had no significant difference with the single-row suture group. While in the over 3-cm tear group, double-row group showed better results than the single-row suture group on the Constant scpre, ASES score and UCLA score. Double-row suture has a low re-rupture rate than single-row suture in rotator cuff injury, and could achieve better abduction muscle strength. There is no significant difference in the functional score between double-row suture and single-row suture in the rotator cuff tear of less than 3 cm, while in the over 3-cm tear, double-row suture could achieve better functional score.

BACKGROUND:Chitosanase is an enzyme for efficient and special hydrolysis of chitoan, and hence its effective and stable expression with enzymatic activity wil contribute to improving gene therapeutic effect.
OBJECTIVE: To construct a chitosanase gene for the efficient and specifical hydrolysis of chitosan, and to investigate its expression inEscherichia coli and the main influencing factors of enzymatic activity.
METHODS:According to the sequences of endo-chitosanase ofAspergilus sp. CJ22-326 provided in Genbank (EU302818), primers were designed and synthesized. The Asperguilus endo-chitosanase gene was amplified by successive extension PCR. And then the recombinant pET28a-His6-CSN was constructed and expressed in
Escherichia coli BL21. Finaly the recombinant His6-CSN fusion protein was analyzed by sodium dodecyl sulfate polyacrylamide gelelectrophoresis (SDS-PAGE), the western blot and dinitrosalicylic acid assay for detecting the enzyme activity of eluted His6-CSN fusion protein. The influence of different pH value and temperature on the enzyme activity of the recombinant chitosanase was investigated.
RESULTS AND CONCLUSION: SDS-PAGE showed that 29 kDa proteins were expressed and the western blot assay showed that His6-CSN expressed successfuly in the host. Dinitrosalicylic acid assay determined the
enzymatic activity of His6-CSN was significantly higher than that of lysozyme, but lower than that of chitosanase from Streptomyces griseus (P < 0.05). The recombinant chitosanase displayed the maximal activity at temperature of 50℃ and pH value of 6.0. There were a higher enzymatic activity remaining at pH value of 4.0-7.0 and
temperature of 30-50℃.

BACKGROUND:Chitosan is wel known as good biocompatibility and biodegradability;however, its extensive use in biomedical applications is restricted due to its poor transfection efficiency. OBJECTIVE:To prepare the polyethyleneimine-chitosan/DNA nanoparticles loading enhanced green fluorescent protein gene, and to detect their physicochemical properties and gene transfection efficiency towards chondrocytes in vitro.
METHODS:Low molecular weight polyethyleneimine was covalently linked to chitosan backbone to construct chitosan-graft-polyethyleneimine;then the chitosan-graft-polyethyleneimine was mixed with DNA nanoparticles, which loaded enhanced green fluorescent protein gene, by a complex coacervation method. The nanoparticle morphology was observed under a scanning electron microscopy. The sizes and zeta-potentials of the
nanoparticles were measured by a Marven-nano laser diffractometer. The binding capacity of plasmid DNA was evaluated by agarose gel electrophoresis analysis. The gene transfection experiments in vitro were performed towards rabbit’s chondrocytes. The gene transfection efficiency was measured with flow cytometry and under fluorescence microscope. How marked DNA entered into the nucleus of chondrocytes mediated by the nanoparticles was detected by laser scanning confocal microscopy.
RESULTS AND CONCLUSION:The prepared nanoparticles were mainly spherical, with an average size of (154.6±18.6) nm, and zeta-potential of (24.68±6.82) mV. The agarose gel electrophoresis analysis confirmed that the nanoparticles could effectively protect plasmid DNA from DNase Ⅰ-induced degradation. Gene transfection in vitro proved that the nanoparticles were efficient in transfecting rabbit’s chondrocytes and the expression of green fluorescent proteins was observed under fluorescent microscope, with a transfection efficiency of (23.80±1.74)%that was significantly higher than that of the naked plasmid DNA and chitosan/DNA nanoparticles (P<0.05). But no significant differences were observed between polyethyleneimine-chitosan/DNA nanoparticles and LipofectamineTM 2000. These findings indicate that the polyethyleneimine-chitosan/DNA nanoparticles can effectively protect plasmid DNA from nuclease degradation, and exhibit the favorable transfection ability towards articular chondrocytes.

Objective To explore the method and effect of transinfection of rabbit early knee osteoarthritis models via chitosan microsphere with gene of recombined human IL-1Ra gene and TGF-β1 gene. Methods Chitosan microspheres with plasmids of IL-1Ra gene and TGF-β1 gene, and rabbit early knee osteoarthritis models were prepared. Rabbits in different groups had intra-articular injections of chitosan microsphere containing IL-1Ra gene and / or TGF-β1 gene, and chitosan solution as control group before being executed regularly and randomly. The joint specimens were evaluated by HE staining, lycopene red O staining and immunohistochemical analysis and Mankin's score. ELISA was used for detection of IL-IRa and TGF-β1 concentration of articular cavity fluid in each group. Results The control group was consistent with the pathological changes of early OA. In co-transinfection group, judging from the appearance and staining of cartilage,the OA damage of the specimens was less serious than other groups'. Its Mankin's score was significantly lower than single-gene transinfection group (P ＜ 0.05), and the latters Mankin's score were significantly lower than control group (P ＜ 0.05). Conclusion Intra-articular injection of chitosan microspheres containing both IL-1Ra gene and TGF-β1 gene could inhibit the degeneration of cartilage and promote cartilage repair.

Objective To investigate and compare the clinical effects of intraarticular injection of sodium hyaluronate and diprospan on knee osteoarthritis. Methods 94 patients with knee osteoarthritis were divided into two groups, the HA group and Cortieosteroid group. Each patient in the HA group was treated with intra-articular injection of sodium hyaluronate at 2.5 ml every week for 5 weeks, and each patient in the Corticosteroid group was treated with intra-articular injection of diprospan at 1ml on the first and fourth week. The clinical assessments included pain,joint effusion,and Lequesne Index. Assessments were done at baseline, at week 4, and week 12. Results 88 cases were followed up for 3 months. A significant decrease in VAS scores for pain and in Lequesne Index was found in both groups at week 4 when compared to baseline and there were no significant differences between the two groups. However,at 12 week improvement in pain score and Lequesne Index was found in favour of hyaluronic acid. In addition,diprospan seemed to have preferable short-term effect on patient with joint effusion. Conclusion Both intra-articular injection of sodium hyaluronate and diprospan provided clinically significant improvement in short-term and demonstrated that hyaluronic acid had a more long-term beneficial effect in patients with knee osteoarthritis.

Objective To introduce experience of using the new AO anterolateral distal tibia locking com-pression plate (LCP) for treatment of Pilon fractures. Methods Between February and August of 2009,8 closed Pi-lon fractures were treated by open reduction and internal fixation. The distal fibula was fixed with a one-third tubular plate or an recontruction plate via a straight incision posterior to the fibular crest. The distal tibia was approched by a straight incision over the ankle joint, and the fracture was stabilized using an anterolateral distal tibia LCP. Regular follow up was made to observe and evaluate the preliminary clinical outcomes. Results Seven of the 8 patients were availabe for follow up for 3 ～ 6 months (average 4.5 months). All incisions obtained primary healing, though one ex-perienced mild superficial inflammation,and none developed deep infections. Based on the Burwell and Charnley radi-ographic criteria,anatomical reduction was obtained in 5 cases,good in 1 ,and fair in 1. Among the 5 cases exceeding 5 months postsurgery,4 were evaluated as excellent and 1 us good according to Tometta' s clinically based criteria for Pilon fractures. Conclusion With good surgical timing,internal fixation with anterolateral LCP for Pilon fractures is reliable and warrants less complications.

Objective To explore transinfection of rabbit chondrocytes via chitosan microsphere with human IL-1Ra and TGF-β1 gene. Methods Chitosan-DNA microspheres carrying plasmids with IL-1 Ra and TGF-β1 genes were prepared.The encapsulation efficiency,DNA-released kinetics and lysozyme degradation in vitro were performed.Articular rabbit chondrocytes were co-transinfected with the plasmids with IL-1Ra and TGF-β1 genes via chitosan-DNA mierosphere,evaluated by fluorescence microscope,TaqMan real-time PCR assay and MTF test. Results The chitosan microspheres with IL-1Ra and TGF-β1 genes were(2.8±0.2)μm and(2.6±0.1)μm in diameters respectively.The encapsulation efficiency were(88.3±4.1)%and(87.2±2.6)%.During the degradation,significant morphological changes were noticed.The plasmids could be released in a multiphasic fashion.Enhanced green fluorescent protein and Real-Time PCR analysis showed that genes were expressed in chondrocytes.lasting near 30 days.MTT indicated that the cotransinfection promoted the chondrocytes'proliferation. Conclusion IL-1Ra and TGF-β1 genes cotransfected into chondrocytes via chitosan-DNA microsphere could be expressed in a long term and cotransinfection promoted the chondroeytes'proliferation,which is the base of inhibiting the degeneration of cartilage and promote cartilage repair.

BACKGROUND: Currently, the materials used in clinical practice to repair cruciate ligament of knee joint contain auto-graft bone- mid 1/3 patella tendon-bone (B-T-B), auto-semitendinous muscle, gracilis muscle and allogenic tissue graft. All of them are limited to a certain degree in clinical application. Therefore, people hope to consistently develop artificial ligaments to take the place of auto- and allografts. OBJECTIVE: To investigate the feasibility to construct artificial biological ligament (ABL) by applying a novel biochemical technique using porcine tendon as the raw material. DESIGN: Research of new biological material. SETTING: Department of Orthopedics, Third Affiliated Hospital of Sun Yat-sen University. MATERIALS: Adult pigs of either gender were provided by the Animal Center of Sun Yat-sen University. Scanning electron microscope (SEM, S-520) was provided by Hitachi, Japan, and micro-controlled electron tension-testing device (Model LWK-10B) by Guangzhou Experimental Devices Factory. METHODS: The experiment was performed at the Animal Center of Sun Yat-sen University from January 2004 to June 2005. ABL was established by means of treating porcine tendon with epoxy cross-linking fixation, diversified antigen minimization process, mechanic enhancement modification and surface activating process. Under aseptic condition, a 6-month-old goat's bone marrow was abstracted, and then the bone marrow matrix stem cells were cultured in ABL stent for 3 weeks. Scanning electron microscope was used to observe structure and compatibility of artificial ligament, and mechanics test was used to analyze biomechanics characteristics of ABL. MAIN OUTCOME MEASURES: Structural features, cell compatibility and biomechanics characteristics of ABL.RESULTS: ① Structural features of ABL: The appearance of ABL was similar to that of the normal human ligament. Histological examination showed that the ABL was collagen fibers with no cells. Electron microscope examination revealed that the ABL was composed of hair-looking and fiber-like objects running uniformly in a certain direction and closely parallel-arranged. ② Cell compatibility: Three weeks after xenogenic marrow matrix cells were cultured on the surface of the ABL, it was noted that cells adhered and the matrix secreted by the cells precipitated around the cells. There were no cells found inside the ABL. ③ Mechanical strength of the ligament: The average diameter of ABL was 5 mm and the mechanical test at a speed of 100 mm/min showed that its averaged tensile limit was 927.19 N and the tension-resistant strength was 47.22 N/mm those were close to the corresponding parameters of the normal goat's ACL. The normal goat's ACL was 5 mm. The greatest tensile load was 807.50 N and the tension-resistant strength was 41.13 N/mm.CONCLUSION:As we used the unique biochemical technique and minimized the xenogenic protein immunogenicity of the porcine tendon, ABL has acceptable biomechanical properties and superior biocompatibility. As a substitute of the ligament in the reconstruction of the ACL, ABL has a promising prospect in clinical applications.

BACKGROUND: A new composite scaffold for cartilage tissue engineer ing has been employed to culture chondrocytes and overcome many limits related to traditional scaffolds, such as poor biocompatibility, inferior mechanical property, inappropriate biodegradability, and simplex structure which can not match layered structure of articular cartilage, etc. The new composite scaffolds provided a new approach for the research of cartilage tissue engineering.OBJECTIVE: To evaluate the feasibility and value of layered cylindrical collagen/hydroxyapatite (HA) composite for cartilage tissue engineering by observing how it absorbs chondrocytes and affects its cellular characteristics.DESIGN: Completely randomized design and controlled experimental study.SETTING: Department of Orthopaedics, Third Hospital Affiliated to Sun Yat-sen University, and College of Material Science, South China University of Technology.MATERIALS: The experiment was conducted at the central experimental laboratory of the Third Hospital Affiliated to Sun Yat-sen University from June to November 2004. One two-week-old male healthy New Zealand rabbit,which was bred in 20 ℃ and 40% humidity, was used in this experiment.METHODS: ①Right amount of deionized water was added into HA, collagen I solution was added to disperse HA, then carbodiimide was added in the mixture at a proportion for getting the collagen/HA composite at different ratios. Pour to the certain mould in successive layers. The upper layer was pure collagen and the bottom was pure HA. The prepared layered cylindrical collagen/HA composite was put into the ultra low temperature freezer, lyophilized, and sterilized by ethylene oxide for the following procedures. ② Chondrocytes of juvenal rabbit were isolated and multiplied in vitro, then chondrocytes were seeded onto porous collagen/HA composite scaffold and cultured. The effects of composite scaffold on chondrocytes'morphological changes, proliferation, and function were evaluated through a series of examinations such as inverted phase-contrast microscopy, histological, scanning electron microscopy and immunohistochemistry.MAIN OUTCOME MEASURES: ①Morphology of collagen/HA porousscaffold. ②The characteristics of chondrocytes cultured in vitro. ③ The chondrocytes' compatibility with the collagen/HA porous scaffold.RESULTS: ①Morphology of collagen/HA porous scaffold: Collagen/ HA was a cylindrical three-dimensional porous scaffold in layer structure with certain mechanical strength. The top layer was pure collagen, the bottom was pure HA, and the intermediate layer was collagen and HA composite.Under scanning electron microscope, the scaffold had irregular blowhole structure, the apertures were even and pores were communicated each other. The average pore diameter was about 147 μm. ②The characteristics of chondrocytes cultured in vitro: The chodrocytes that had just been isolated were spherical; rate of living cells was 95%. 24 hours later, the cells were going to attach to the wall, extend to be triangle or polygon. The cytoplasms were abundant with secretary granules inside. The cells kept on growing in monolayer and the multiply rate accelerated after passage, and chondrocytes overspreaded the culture flask in 4 days. After passaged for several times, the multiply rate decreased. In passage Ⅵ, the ability of cell division was decreased obviously, the cytomorphosis was clear. Cellular sizes were larger. Most of cells were shuttle-shape, less refracted with blur edges, which trended to age and dedifferentiate to fibroblast. ③The chondrocytes' compatibility with the collagen/HA porous scaffold: Cylindrical collagen/ HA composite scaffold in layer structure had good hydrophilicity. The chondrocytes adhered to the surface of the scaffold, proliferated and migrated toward the scaffold through the pore. Chondrocytes attached to wall of microholes of the scaffold had largely maintained a rounded morphology and could secrete the extracellular matrix on the porous scaffold.CONCLUSION: Cylindrical collagen/ HA composite scaffold in layer structure has good cellular compatibility.It is stronger in mechanical property than pure collagen, is anideal scaffold for cartilage tissue engineering.