Reconstruction of Large Area of Deep Wound in the Foot and Ankle with Chimeric Anterolateral Thigh Perforator Flap.

OBJECTIVE: To evaluate the clinical application and surgical efficacy of the chimeric perforator flap pedicled with the descending branch of the lateral circumflex femoral artery and the lateral thigh muscle flap for the reconstruction of the large area of deep wound in foot and ankle. METHODS: Clinical data of 32 cases who underwent chimeric anterolateral thigh perforator flap to repair the large area of deep wound of the foot and ankle from January 2015 to December 2018 were retrospectively analyzed. The sizes of the defects ranged from 18 cm × 10 cm to 35 cm × 20 cm, with exposed tendon and bone and/or partial defects and necrosis, contaminations, accompanied by different degrees of infection. Following the radical debridement and VSD, chimeric anterolateral thigh perforator flap was employed to repair the deep wounds according to the position, site and deep-tissue injury of the soft-tissue defects. The skin flap and muscle flap were fanned out on the wound, and single- or two-staged split-thickness skin grafting was performed on the muscle flap. The operation time and blood loss were recorded. The survival and healing conditions of the operational site with chimeric anterolateral thigh perforator flap were evaluated post-operationally. Complications at both recipient site and donor site were carefully recorded. RESULTS: The mean time of the operation was 325.5 min and average blood loss was 424.8 mL. Among the 32 cases, two cases developed vascular crisis, which were alleviated with intensive investigation and treatment; Four cases suffered from partial necrosis of the flap or skin graft on the muscle flap or on the residual local wound, which were improved after treatment of further dressing change and skin grafting. Another four cases experienced post-traumatic osteomyelitis accompanied by bone defect were treated with simple bone grafting or Mesquelet bone grafting at 6-8 months after wound healing. Postoperatively, the wounds were properly healed, and the infection was effectively controlled without sinus tract forming. Overall, all 32 cases received satisfactory efficacy, without influencing subsequent functional reconstruction, and observed infection during the 12-36 months post-operational follow-up. CONCLUSION: The chimeric perforator flap pedicled with the descending branch of the lateral circumflex femoral artery and the lateral thigh muscle flap provides an effective and relative safe procedure for the repair of a large area of deep wound in the foot and ankle, particularly with irregular defect or deep dead space.

Free flap transplantation combined with Ilizarov bone transport for the treatment of severe composite tibial and soft tissue defects.

OBJECTIVE: To evaluate the clinical efficacy of free flap transplantation combined with Ilizarov bone transport in the treatment of severe composite tibial and soft tissue defects. METHODS: We retrospectively analyzed the clinical data of 40 patients with severe composite tibial and soft tissue defects who underwent free flap transplantation combined with Ilizarov bone transport. The clinical efficacy was evaluated according to the following criteria: success rate of wound repair by free flap transplantation, incidence or recurrence rate of deep infection, healing rate of bone defects and external fixation index, incidence of complications, and functional score of affected extremities. RESULTS: All infections were generally well controlled by radical debridement and negative-pressure therapy, and all 40 patients' wounds healed after repair and reconstruction of the tibia and soft tissues. Postoperative complications were alleviated by active treatment. The mean external fixation time was 12.83 ± 2.85 months, and the external fixation index was 1.55 m/cm. According to the Association for the Study and Application of Methods of Ilizarov (ASAMI) score, an excellent or good functional outcome was attained in 85% of patients. CONCLUSION: Free flap transplantation combined with Ilizarov bone transport is an effective treatment for severe composite tibial and soft tissue defects.

A neglected problem in the utilization of free anterolateral thigh flap toward reconstructing complicated wounds of extremities: the obliteration of deep dead space.

BACKGROUND: Deep dead space may be thought as an independent risk factor of the poor infection control after flap reconstruction in complex limb wounds. But it can be easily neglected. The conventional skin flap and musculocutaneous flap are difficult to obliterate the deep dead space in irregular shape effectively. It was investigated that the clinical application of chimeric anterolateral thigh perforator flap in the treatment of complex wounds complicated with deep dead space of the extremities in the paper. METHODS: Fifty-six cases complicated with deep dead space wounds were registered in group. Following thorough debridement and treatment with VSD, the granulation tissues grew with well-controlled infection. And then the chimeric anterolateral thigh perforator flap was used to obliterate the deep dead space and repair the wounds. The postoperative flap survival and infection conditions were evaluated. RESULTS: Overall, the infection was effectively controlled, without persistent exudation or sinus tract formation after wound healing. While 5 cases lost to follow-up, the remaining 51 cases were followed up until 15 months on average. Generally, the affected extremities recovered satisfactorily with normal appearances and texture of the flaps, along with normal functions. Importantly, no recurrence of infection was observed. CONCLUSION: During the grafting of chimeric perforator flap pedicled with lateral thigh muscle flap, the muscle flap is recommended to obliterate the deep dead space while the skin flap is being used to cover the wound. The combination of these two technologies performed well in the repair and reconstruction of the complex wounds of the extremities, possessing potential for broader clinical application.

Treatment options for infected bone defects in the lower extremities: free vascularized fibular graft or Ilizarov bone transport?

OBJECTIVE: The objective was to explore the relative indications of free vascularized fibular graft (FVFG) and Ilizarov bone transport (IBT) in the treatment of infected bone defects of lower extremities via comparative analysis on the clinical characteristics and efficacies. METHODS: The clinical data of 66 cases with post-traumatic infected bone defects of the lower extremities who underwent FVFG (n = 23) or IBT (n = 43) from July 2014 to June 2018 were retrieved and retrospectively analyzed. Clinical characteristics, operation time, and intraoperative blood loss were statistically compared between two groups. Specifically, the clinical efficacies of two methods were statistically evaluated according to the external fixation time/index, recurrence rate of deep infection, incidence of complications, the times of reoperation, and final functional score of the affected extremities. RESULTS: Gender, age, cause of injury, Gustilo grade of initial injury, proportion of complicated injuries in other parts of the affected extremities, and numbers of femoral/tibial defect cases did not differ significantly between treatment groups, while infection site distribution after debridement (shaft/metaphysis) differed moderately, with metaphysis infection little more frequent in the FVFG group (P = 0.068). Femoral/tibial defect length was longer in the FVFG group (9.96 ± 2.27 vs. 8.74 ± 2.52 cm, P = 0.014). More patients in the FVFG group presented with moderate or complex wounds with soft-tissue defects. FVFG treatment required a longer surgical time (6.60 ± 1.34 vs. 3.12 ± 0.99 h) and resulted in greater intraoperative blood loss (873.91 ± 183.94 vs. 386.08 ± 131.98 ml; both P < 0.05) than the IBT group, while average follow-up time, recurrence rate of postoperative osteomyelitis, degree of bony union, and final functional scores did not differ between treatment groups. However, FVFG required a shorter external fixation time (7.04 ± 1.72 vs. 13.16 ± 2.92 months), yielded a lower external fixation index (0.73 ± 0.28 vs. 1.55 ± 0.28), and resulted in a lower incidence of postoperative complications (0.87 ± 0.76 vs. 2.21±1.78, times/case, P < 0.05). The times of reoperation in the two groups did not differ (0.78 ± 0.60 vs. 0.98 ± 0.99 times/case, P = 0.615). CONCLUSION: Both FVFG and IBT are effective methods for repairing and reconstructing infected bone defects of the lower extremities, with unique advantages and limitations. Generally, FVFG is recommended for patients with soft tissue defects, bone defects adjacent to joints, large bone defects (particularly monocortical defects), and those who can tolerate microsurgery.

Free flap transplantation combined with skin grafting and vacuum sealing drainage for repair of circumferential or sub-circumferential soft-tissue wounds of the lower leg.

BACKGROUND: This study is aimed at evaluating the operation techniques and clinical significance of free flap transplantation combined with skin grafting and vacuum sealing drainage (VSD) in repairing severe traumatic extensive circumferential or semi-circumferential soft-tissue defects of the lower leg. MATERIAL AND METHODS: Thirty patients with severe lower leg injuries were treated by free flap transplantation combined with skin grafting and VSD from January 2008 to June 2011. The size of the wounds ranged from 23×8 cm to 44×28 cm and all affected more 70% of the low leg circumferential area. Wounds were complicated by exposure, necrosis, or infection of deep tissues. The wounds were first debrided and covered by VSD. When the condition of the wound had improved (5 to 7 days later), free flaps were harvested to reconstruct damaged tissue and skin grafts and VSD was used to cover granulation tissues around the transplanted flap. RESULTS: Granulation tissues developed and the area requiring flap cover decreased in all 30 patients after debridement and VSD. In 28 of 30 cases, the transplanted flaps grew well without complication. Peripheral necrosis was observed in only 2 cases, which required a second debridement and skin graft. Ten wound areas covered by grafts were left with scattered peripheral wounds, which healed with the help of 1 more skin graft or dressing change. Morphological appearance and functional recovery were satisfactory in all 30 cases. CONCLUSIONS: Initial debridement and the temporary VSD cover followed after several days by free flap transplantation combined with skin grafting and VSD protection is a reliable treatment regimen for traumatic large circumferential or sub-circumferential soft tissue wounds of the lower leg with deep tissue exposure.

Reconstruction of shoulder abduction by multiple nerve fascicle transfer through posterior approach.

PURPOSE: To evaluate the feasibility and clinical efficacy of multiple nerve fascicle transfer through posterior approach for reconstruction of shoulder abduction in patients with C5 or upper brachial plexus injury. METHODS: 11 patients (aged between 17 and 56 years) with dysfunction of shoulder abduction post C5 or upper brachial plexus injury were recruited in this study. Among them, four out of 11 patients also had dysfunction of elbow flexion simultaneously. The duration from injury to the surgery ranged from 4 to 12 months, with an average of 6.7 months. The affected shoulder joints showed abduction, extension and elevation dysfunction, but the muscle strength of shoulder shrugging and elbow extension was graded to M4 or higher. Accessory nerve was transferred to the suprascapular nerve and triceps muscle was branched to the axillary nerve through posterior approach. Ulnar fascicle was transferred to the motor branches of biceps for the 4 patients involved with elbow flexion dysfunction. RESULTS: Ten out of 11 cases were followed-up for 15-36 months. Neo-potential of deltoid and supraspinatus/infraspinatus was documented at 4-5 months post surgery. Shoulder abduction (and elbow flexion) was reanimated at 4-8 months post surgery. Significant improvement was observed at 15-36 months post surgery, shoulder abduction regained to 40-160° (mean: 92.5°), muscle strength of supraspinatus/infraspinatus and deltoid were graded to M3-M5 (mean: 4.0 and 4.1); 3 cases muscle strength of elbow flexion was graded from M4 to M5- (mean: 4.4) with 1 case loss. Shoulder shrugging of trapezius was graded to M5 in 5 cases, M5- in 2 cases, M4 in 2 cases and M3 in 1 case (mean: 4.5). All cases showed normal elbow extension and muscle strength of triceps (M5). CONCLUSION: It is feasible to carry out multiple nerve fascicle transfers for early reconstruction of shoulder abduction by posterior approach. Patients who received this procedure achieved good functional recovery and their donor site morbidity/injury was minimal.

[Treatment of large circular soft tissue defect in lower extremities with a combination of bridge flaps and free skin graft covered by vacuum sealing drainage].

OBJECTIVE: To explore the surgical technique and clinical value of treatment for posttraumatic large circular soft tissue defect in the lower extremities using a combination of posterior tibial vascular bridge flap in the unaffected leg and skin graft covered by vacuum sealing drainage (VSD). METHODS: From January 2008 to June 2010, 11 cases with posttraumatic large circular soft tissue defects with deep tissue exposed or partial necrosis in the lower extremities were treated by bridge flaps and combined with free skin graft covered by VSD. There are 7 males and 4 females, with an average age of 32.5 years (range from 15 to 52 years). The size of wound varied from 24 cm × 13 cm to 45 cm × 24 cm. After the wound were completely debrided, the external fixation or internal fixation was conducted for the patients with unstable fracture. Then VSD were used to covered the wound for 1-2 times with a period of 5 to 7 days according to the wound condition. After granulation tissue grew, bridge flap transplantation was performed to repair tissue defect and cover the exposed bone, which combined with skin graft covered by VSD was used to cover the residual wound. Survival rate and quality healing of the flaps were followed up postoperatively. RESULTS: The granulation growth of 11 cases with large circular soft tissue defect in the lower extremities associated tissue exposure was good after the application of VSD. All the cases were covered by free flap transplantation and skin grafting except for 6 cases due to large defect. The wound was covered by skin re-grafting in 4 cases and frequently dressing change in 2 cases. All the flaps were successful with good infection control and no sinus. The average period of follow-up was 10.6 months (5 - 24 months). All the patients were satisfied with the good outline and good function of the affected limb. CONCLUSION: Patients with posttraumatic large circular soft tissue defect in the lower extremities can be effectively treated with a combination of bridge flaps and free skin graft covered by VSD, which can shorten the course of treatment, and restore the function of affected extremities as much as possible.

Sequential therapy of vacuum sealing drainage and free-flap transplantation for children with extensive soft-tissue defects below the knee in the extremities.

PURPOSE: The aim of the study is to evaluate the surgical technique and clinical significance of the sequential therapy of vacuum sealing drainage (VSD) and free-flap transplantation for children with extensive soft-tissue defects below the knee in the extremities. METHODS: Twenty-two children (aged from 3 to 10 years) received sequential therapy of VSD and free-flap transplantation. All cases suffered from extensive area soft-tissue defects and exposure or partial defects of bones, tendons and other deep tissues. The wound sizes varied from 10 cm × 6 cm to 30 cm × 22 cm. Amongst 22 cases, 12 cases had fresh wounds and the remaining 10 children had necrotising infection. After complete debridement, the wounds were covered by VSD. External fixation or Kirschner-wire fixation should be performed for the cases complicated by unsteady fractures. After the removal of negative pressure VSD devices, free-flap transplantations were performed in 8 cases after debridement, and 14 cases received combined therapy of free-flap transplantation and skin grafting depending upon the severity of soft-tissue and deep-tissue defects. The flap survival and wound healing were followed up postoperatively. RESULTS: After VSD treatment, the infection of deep-tissue exposure was effectively prevented, and granulation tissues surrounding the exposed areas of tendons and bones grew well. All patients who received free-flap transplantation at the second stage survived without the occurrence of vascular crisis, infection or sinus formation. During follow-up ranging from 6 to 24 months, all the patients were satisfied with the morphological appearance and functional recovery of the affected limbs. CONCLUSION: Sequential therapy of VSD and free-flap transplantation can serve as a reliable option for children with extensive soft-tissue defects below the knee in the extremities and exposed deep tissues, after complete debridement, which significantly shortens remedy period, enhances success rate for surgery and achieves maximal restoration of limb function.

[Three-dimensional reconstruction technique in gastrocnemius flap surgery: initial clinical application].

OBJECTIVE: To discuss the experience with three-dimensional reconstruction technique in initial clinical application in gastrocnemius muscle flap surgery. METHOD: From 2007 to 2008, 7 patients received gastrocnemius muscle flap surgeries to repair the wounds. Preoperative CT angiography or magnetic resonance imaging (MRI) was performed after injection of the contrast media for individualized three-dimensional gastrocnemius muscle flap reconstruction using Amira4.1 software. According to the size of the defect in the wound, individualized three-dimensional gastrocnemius muscle flap was designed and harvested from the posterior leg. RESULTS: Individualized three-dimensional reconstruction of the gastrocnemius flap was performed in 7 cases, and the reconstructed flaps clearly displayed the blood vessels, skin and the adjacent three-dimensional structures. In 6 cases the main perforating branched and trunk of the blood vessels in the designed flap were consistent with the surgical findings; in 1 case, the perforating branches failed to be clearly displayed in the designed flap, and surgical examination identified perforating branches with an average diameter of 0.5 mm (minimally 0.3 mm). The flaps survived in all the 7 cases. CONCLUSIONS: Three-dimensional reconstruction of the gastrocnemius flap based on the lower limb CT angiography or MRI allows three-dimensional observation of the anatomy of the flap and accurate marking of the extent of the flap to be harvested, therefore avoiding intraoperative injuries to the blood vessels to better survival of the flaps.

Reconstruction of phalangeal articulations of the hand with vascularised phalangeal articulations of foot.

SUMMARY: Since arthroplasty, prosthetic replacement and non-vascularised articulation autografting do not normally produce very satisfactory results for ankylosis of metacarpophalangeal and interphalangeal joints, the authors performed reconstruction of phalangeal articulations of the hand using vascularised phalangeal articulations of the foot in 11 patients with ankylosis of the metacarpophalangeal and interphalangeal joints of hand due by trauma. Procedures included reconstruction of 9 hand metacarpophalangeal joints with vascularised grafting of pedal metatarsophalangeal joints in six patients, reconstruction of the hand metacarpophalangeal joints with grafting of vascularised proximal interphalangeal joints of foot in two patients and reconstruction of the hand proximal interphalangeal joints with grafting of vascularised proximal interphalangeal joints of foot in three patients. Early functional exercise was encouraged in all cases post-operatively. Follow-up ranged from 3 to 10 years and revealed that 9 cases had normal appearance and length of recipient area, 1 had slightly clumsy dorsal skin in the hand and 1 had slight dorsal angulation of a metacarpal bone. Recovery of joint range of motion was satisfactory. Radiographic, gross and sensation examinations also showed good operative outcomes. The authors believe that vascularised pedal metatarsophalangeal joints, with a rich blood supply, can be grafted to effectively reconstruct the finger joints with good function. A low rate of degeneration results because pedal and hand metatarsophalangeal joints are similar in anatomy and physiological function.

Replantation of articular composite tissue masses severed from extremities.

SUMMARY: The techniques and outcomes of in situ replantation are discussed for managing 5 cases of articular composite tissue masses severed from an extremity (digit). All 5 cases treated with in situ replantation survived. Rehabilitation was performed after surgery. Follow-ups of 2-5 years showed good appearance, satisfactory functional and sensory recovery. In situ replantation is indicated for an articular composite tissue masses severed from an extremity (digit), if its structure is complete and a blood supply vessel in the mass is available for anastomosis. Replantation can achieve better outcomes than transfer or grafting of adjacent skin or osteocutaneous flaps, or transplantation of a metatarsophalangeal or interphalangeal joint.

[Green fluorescent protein as a tracer of bone marrow stromal cells in bone tissue engineering in rhesus].

OBJECTIVE: To observe the role of green fluorescent protein (GFP) in tracing rhesus bone marrow stromal cells (rBMSCs) during tissue-engineered bone formation in vivo. METHODS: Ad5.CMV-GFP was amplified by infecting QBI-293A cells, and the bone marrow was harvested from the ilium of adult male rhesus to obtain rBMSCs, which were cultured and passaged in vitro. GFP was transfected into the third-passage rBMSCs via adenovirus vector and the labeled cells were inoculated into absorbable HA scaffold and cultured for 3 days, with untransfected rBMSCs as control, before the cell-matrix compounds were implanted into the latissimus dorsi muscles of rhesus. Samples were harvested at 6 week and embedded in paraform, and ground sections of the bone tissue were prepared to observe green fluorescence under laser scanning confocal microscope. Propidium iodide staining of the sections was also performed for observation. RESULTS: The rBMSCs grew well after GFP transfection, and green fluorescence could be seen 24 h after the transfection and became stronger till 48 h, with a positive transfection rate beyond 80%. Six weeks after cell implantation, the rBMSCs labeled by GFP-emitted green fluorescence were detected in the bone tissue under laser scanning confocal microscope. CONCLUSION: GFP can effectively trace BMSCs during bone tissue engineering, and the transplanted BMSCs constitute the main source of bone-forming cells in bone tissue engineering.

[Fingertip replantation after amputation: report of 32 fingers].

OBJECTIVE: To describe the surgical techniques and our experiences in fingertip replantation after amputation. METHODS: On the basis of examination of the anatomic features and the degree of fingertip vascular injury, 32 amputated fingertips in 26 cases were replanted, and flexible revascularization procedures of both artery and vein anastomoses, artery-only anastomosis, arterialized vein and arteriovenous anastomosis were adopted. All the replanted fingertips were trained with comprehensive rehabilitation program. RESULTS: Twenty-nine replanted fingertips survived but 3 failed, and the overall survival rate was 90.06%. During the follow-up lasting from 4 months to 5 years, the 29 replanted fingertips survived with excellent blood supply, good sensory functions, satisfactory shape and functions according to the criteria by Society of Hand Surgery of Chinese Medical Association. CONCLUSIONS: Fingertip replantation after amputation can achieve not only high survival rate but also satisfactory appearance and functions as long as appropriate operative procedures are adopted with comprehensive rehabilitation therapy.

[Induced differentiation of adult human bone marrow derived mesenchymal stem cells in vitro toward osteoblasts].

OBJECTIVE: To investigate the feasibility of inducing in vitro mesenchymal stem cells (MSCs) derived from adult human bone marrow differentiate into osteoblasts and potential applicability of the MSCs as the seed cells in tissue engineering. METHODS: Adult human bone marrow was collected from the healthy adult volunteers to obtain the MSCs, which, after in vitro culture in DMEM supplemented with 10% fetal bovine serum and incubation under standard condition, were induced to differentiate into osteoblasts in DMEM containing dexamethasone (1x10(-8) mol/L), beta-sodium glycerophosphate (10 mmol/L) and ascorbic acid (50 mg L). Proliferation and differentiation of the MSCs were observed continually under inverted phase-contrast microscope and transmission electron microscope. The collagen typeI was detected by immunohistochemistry, alkaline phosphatase (AP) in the MSCs stained by Gomori, the calcified nodules were stained by von Kossa method, and the changes in the content of AP were measured. RESULTS: The MSCs proliferated rapidly in in vitro culture and after a 2- to 3-week induction, the cells began to generate large amount of enlarged endoplasmic reticulum, Golgi complexes and mitochondria, with immature cell nuclei. Positive staining for collagen typeIand strong reaction for AP and calcified nodules were observed. Increasing AP secretion by the MSCs was seen as the time of induction prolonged (P<0.01). CONCLUSIONS: Human bone marrow-derived MSCs can be induced to differentiate into osteoblasts through relatively simple procedures, which provide ideal autogenous source of seed cells for bone tissue engineering. The method adopted in this experiment may be used for routine culture of the seed cells for bone tissue engineering.

[Fabrication of laryngeal cartilage by means of tissue engineering technique].

OBJECTIVE: To explore the method of fabricating tissue engineered laryngeal cartilage. METHODS: The rib and articular cartilage of infant New Zealand white rabbits were harvested in sterile condition. The chondrocytes were separated by collagenase digestion and cultured in vitro for 3 passage. Serial steps of solution casting, extrusion molding and particulate leaching were used to make larynx-shaped biomaterial models with poly(3-hydroxybutyrate-co-3-hydroxyhexanoate, PHBHH). The chondrocytes were seeded onto PHBHH scaffolds to form cell-PHBHH composites, which were subsequently in vitro for one week. After that, the measure of filling inner space of cell-PHBHH composites together with wrapping total composites using either greater omentum (n = 9) or fascia flap and muscle (also n = 9) in experimental groups was taken to implant the larynx-shaped biomaterial models seeded with chondrocytes into the belly and the back of adult New Zealand white rabbits. Control groups (every group n = 3) were the same measure as experimental groups but without chondrocyte on PHBHH scaffolds. Finally, morphological observation, HE staining & special staining and immunohistochemical test were conducted to assess cartilage regeneration and its shape at different period following implantation. RESULTS: The rate of viable cell in the final cell suspension was (93 +/- 2)% after well-controlled prolongation of digestion trypsin. Similar to that by traditional procedures (94 +/- 2)% (P > 0.05). The larynx-shaped PHBHH models with edges and corners of laryngeal cartilage made by us appeared to be hollow half-trumpet shape and its porosity was more than 90%. It showed that chondrocytes equally attached to the surface of porous PHBHH and filled within porousness with scanning electron microscopic examination. Tissue engineered larynx-shaped specimens could alternatively be harvested with the above mentioned two different implantation measures. The specimens presented to be similar to that before implantation in gross shape. It was demonstrated to be cartilaginous tissue through histological and immunohistochemical examination. Furthermore, There was nearly no difference between two kinds of tissue engineered laryngeal cartilage with two measures of implantation in morphology and histology. CONCLUSIONS: The regeneration of tissue engineered cartilage in vivo is not influenced by the chondrocytes harvested by improvement of well-controlled prolonged digestion with trypsin during in vitro cell culture. It seems that PHBHH may be used as scaffold in cartilage tissue engineering and wrapping together with filling method with either greater omentum or fascia flap and muscle is appropriate for fabricating tissue engineered laryngeal cartilage.

[Study of osteoblasts transfected with gfp in vitro and traced in vivo].

OBJECTIVE: To observe the osteoblasts transfected with green fluorescent protein (GFP)by adenovirus vector expressed in vitro and traced it in vivo in order to research the feasibility of GFP as a tracer of seeding cells for tissue engineering. METHODS: CFP were transfected into the osteoblasts which derived from adult human bone marrow stromal cell (hBMSc) by adenovirus vector after being packed in 293A cells. The nontransfected hBMSc was used as the control group. The osteoblasts in each group were observed under an inverted phase contrast microscope and fluorescence microscope. The expressive efficiency of GFP was examined by flow cytometry,and alkaline phosphatase (ALP) activities and osteocalcin (OCN) synthesis. After eight days of the transfection,the osteoblasts were implanted into the muscle of nude mouse thigh while the non-transfected osteoblasts were also implanted as a control. Four and eight weeks after the operation, the nude rats were killed and the continuous tissue sections were examined using fluorescence microscopy after adjacent sections were performed by immunohistochemistry or routine HE staining. RESULTS: The green fluorescence was shown the transfected osteoblasts which derived from bone marrow. The rate of positive expression was over 75%. After eight days of the transfection, the marker proteins of the surface of the osteoblasts showed extremely efficient expression of CD29 and CD44, but the CD34 expressed negative. Either ALP or OCN of the osteoblasts was no significant difference between the two groups (P > 0.05) four and eight days after the transfection. The GFP were obviously expressed in nude mouse both at four and eight weeks, meanwhile it did not harm on the morphology and function of the transfected osteoblasts whose immunohistochemistry examination showed positive reaction. CONCLUSIONS: GFP could be transfected osteoblasts effectively in vitro and traced in vivo in nude mouse. It may be an optimal tracer for living cells on tissue engineering research.

[The method of accelerating osteanagenesis and revascularization of tissue engineered bone in big animal in vivo].

OBJECTIVE: To study whether tissue engineered bone can repair the large segment bone defect of large animal or not. To observe what character the fascia flap played during the osteanagenesis and revascularization process of tissue engineered bone. METHODS: 9 Chinese goats were made 2 cm left tibia diaphyseal defect. The repairing effect of the defects was evaluated by ECT, X-ray and histology. 27 goats were divided into three groups: group of CHAP, the defect was filled with coral hydroxyapatite (CHAP); group of tissue engineered bone, the defect was filled with CHAP + bone marrow stroma cells (BMSc); group of fascia flap, the defect was filled with CHAP + BMSc + fascia flap. After finished culturing and inducing the BMSc, CHAP of group of tissue engineered bone and of fascia flap was combined with it. Making fascia flap, different materials as described above were then implanted separately into the defects. Radionuclide bone imaging was used to monitor the revascularization of the implants at 2, 4, 8 weeks after operation. X-ray examination, optical density index of X-ray film, V-G staining of tissue slice of the implants were used at 4, 8, 12 weeks after operation, and the biomechanical character of the specimens were tested at 12 weeks post operation. RESULTS: In the first study, the defect showed no bone regeneration phenomenon. 2 cm tibia defect was an ideal animal model. In the second study, group of CHAP manifested a little trace of bone regeneration, as to group of tissue engineered bone, the defect was almost repaired totally. In group of fascia flap, with the assistance of fascia flap which gave more chance to making implants to get more nutrient, the repair was quite complete. CONCLUSIONS: The model of 2 cm caprine tibia diaphyseal defect cannot be repaired by goat itself and can satisfy the tissue engineering's demands. Tissue engineered bone had good ability to repair large segment tibia defect of goat. Fascia flap can accelerate the revascularization process of tissue engineered bone. And by this way, it augment the ability of tissue engineered bone to repair the large bone defect of goat.

[An experimental study of treatment of comminuted distal femur fracture with locked double- plating].

OBJECTIVE: To explore new techniques for the treatment of comminuted distal femoral fractures to promote functional restoration and reduce the complications of the fracture. METHODS: Sixteen adult canines were selected for this study, and the experimental and control hindlimb for each canine were randomly determined. Distal comminuted fracture (AO/ASIF Type C2) was induced in both sides of the canine's femurs, and one side was fixed with the double plates (steel plate and polyethylene plate), whereas the other with a single steel plate. By the end of 3, 6, 9 and 12 weeks respectively after surgery, 4 of the dogs were used for weight-bearing observation and radiological and histological examination of the limbs. RESULTS: In comparison with dogs in single plate group, double plate fixation resulted in earlier recovery of weight bearing, more abundant and better callus, finer callus moulding and quicker healing. There were slightly more inflammatory cells, however, in the tissues under the polyethylene than those under the steel plate. CONCLUSION: Locked double-plate internal fixation is an excellent alternative for treating comminuted fractures of the distal femur, and may have its potential use in clinical settings.

[Biocompatibility of adult human osteoblasts with coral-derived hydroxyapatite in vitro].

OBJECTIVE: To study the biocompatibility of the osteoblasts from adult human bone marrow with coral-derived hydroxyapatite (CHA) in in vitro culture. METHODS: Bone marrow was obtained from healthy adult subjects and cultured in Dulbecco's modified Eagle's medium (DMEM) containing 10 % fetal bovine serum. The subsequent cell passaging was conducted in conditioned medium containing dexamethasone, beta-sodium glycerophosphate and ascorbic acid, with the osteoblasts in culture then divided into CHA group (in which the cells were cultured with CHA) and osteoblasts group (without CHA). The proliferation and differentiation of all the cultured cells were observed at different time points under inverted phase contrast microscope, optical microscope with HE staining and scanning electron microscope respectively. Proliferation of the cultured cells were evaluated by MTT assay, and the activity of alkaline phosphatase and total micro-protein contents in these cultured osteoblasts were quantitatively detected. RESULTS: The osteoblasts from adult human bone marrow grow well in vitro, regardless of the presence of CHA, with biological and morphological characteristics similar to those of normal osteoblasts. CHA improved the adhesion, growth and proliferation of the cultured cells, showing no adverse effects on the cell functions. CONCLUSION: CHA is an optimal scaffold material for bone tissue engineering, which may potentially find clinical application for bone defect repair.

[Biomechanical study of double-plate fixation for comminuted fractures of the distal femur].

OBJECTIVE: To explore the new treatment approach for comminuted fractures of the distal femur. METHODS: Supra- and intercondylar comminuted fractures were induced in the femurs of 20 fresh adult cadavers, 10 of which received fixation with lateral steel plates and medial polyethylenes plates while the rest with single plate in the lateral only. Biomechanical tests of bending, torsion and compression were performed. RESULTS: Biomechanical test demonstrated that double-plate fixation provided better resistance to compression (P<0.05) as well as bending and torsion (P<0.01) than single-plate fixation did. CONCLUSION: Double-plate fixation is effective and may have potential clinical value in treating comminuted fractures of the distal femur.