Objective To compare the effect between vascularization osteogenesis and membrane guided osteogenesis in the bone repair by the tissue engineered bone with pedicled fascial flap packing autologous red bone marrow (ARBM), so as to provide a reference for the bone defect repair in cl inic. Methods The tissue engineered bone was constructed with ARBM and the osteoinductive absorbing recombinant human materials with recombinant human bone morphogenetic protein 2. Sixty New Zealand rabbits (aged 4-5 months, weighing 2.0-2.5 kg) were randomly divided into group A (n=16), group B (n=22), and group C (n=22). The complete periosteum defect model of 1.5 cm in length was prepared in right ulnar bone, then the tissue engineered bone was implanted in the bone defect area in group A, the tissue engineered bonewith free fascial flap in group B, and the tissue engineered bone with pedicled fascial flap in group C. At 4, 8, 12, and 16 weeks, the tissue of bone defect area was harvested from 4 rabbits of each group for the general, histological, and immunohistochemical staining observations; at 8, 12, and 16 weeks, 2 rabbits of groups B and C, respectively were selected to perform ink perfusion experiment by axillary artery. Results The general observation showed that the periosteum-l ike tissues formed in the fascial flap of groups B and C, chondroid tissues formed in group B, new bone formed in group C, and the fibrous and connective tissues in group A at 4 and 8 weeks; a few porosis was seen in group A, more new bone in group B, and bone stump formation in group C at 12 and 16 weeks. Histological observation showed that there were few new blood vessels and new bone trabeculae in groups A and B, while there were large amounts of new blood vessels and mature bone trabeculae in group C at 4 and 8 weeks. There were a few new blood vessels and new bone trabeculae in group A; more blood vessels, significantly increased mature trabeculae, and the medullary cavity formation in group B; and gradually decreased blood vessels, the mature bone structure formation, and the re-opened medullary cavity in group C at 12 and 16 weeks. The immunohistochemical staining observation showed that the levels of CD105, CD34, and factor VIII were higher in group C than in groups A and B at different time points.The bone morphometry analysis showed that the trabecular volume increased gradually with time in 3 groups after operation; the trabecular volume in group C was significantly more than those in groups A and B at different time points (P lt; 0.05); and there was significant difference between groups A and B (P lt; 0.05) except the volume at 4 weeks (P gt; 0.05). The vascular image analysis showed that the vascular regenerative area ratio in group C was significantly higher than those in groups A and B at different time points (P lt; 0.05). The ink perfusion experiment showed that the osteogenic zone had sparse ink area with no obvious change in group B, while the osteogenic zone had more intensive ink area and reached the peak at 8 weeks, then decreased in group C. Conclusion The tissue engineered bone with pedicled fascial flap packing ARBM has the vascularization osteogenesis effect at early stage, but the effect disappears at late stage gradually when the membrane guided osteogenesis is main.
Objective To construct a new type of self-assembling peptide nanofiber scaffolds—RGDmx, and to study the cell compatibility of the new scaffolds and the proliferation and chondrogenic differentiation of precartilaginous stem cells(PSCs) in scaffolds. Methods PSCs were separated and purified from newborn Sprague Dawley rats by magnetic activated cell sorting and indentified by immunohistochemistry and immunofluorescent staining. The RGDmx were constructed by mixing KLD-12 and KLD-12-PRG at volume ratio of 1 ∶ 1. PSCs at passage 3 were seeded into the KLD-12 scaffold (control group) and RGDmx scaffold (experimental group). The proliferation of PSCs in 2 groups were observed with the method of cell counting kit (CCK) -8 after 1, 3, 7, and 14 days after culture. The RGDmx were constructed by mixing KLD-12-PRG and KLD-12 at different volume ratios of 0, 20%, 40%, 60%, 80%, and 100% and the prol iferation of PSCs was also observed. The complete chondrogenic medium (CCM) was used to induce chondrogenic differentiation of PSCs in different scaffolds. The differentiation of PSCs was observed by toluidine blue staining and RT-PCR assay. Results PSCs were separated and purified successfully, which were identified by immunohistochemistry and immunofluorescent staining methods. The results of CCK-8 showed that the absorbance (A) value in the experimental group increased gradually and reached the highest at 7 days; the A value in the experimental group was significantly higher than that in the control group at 7 days and 14 days (P lt; 0.05). Meanwhile, the A value in the RGDmx scaffold with a volume ratio of 40% was significantly higher than those in others (P lt; 0.05). After 14 days of induction culture with CCM, the toluidine blue staining results were positive in 2 groups; the results of RT-PCR showedthat the expression levels of collagen type II and the aggrecan in the experimental group were significantly higher than those in the control group (P lt; 0.05). Conclusion The self-assembling peptide nanofiber scaffold—RGDmx is an ideal scaffold for tissue engineer because it has good cell compatibility and more effective properties of promoting the differentiation of PSCs to chondrocytes.
ObjectiveTo explore the reliability and effectiveness of soft tissue defect reconstruction using the lower trapezius musculocutaneous flap carrying a portion of the latissimus dorsi muscle.MethodsBetween December 2014 and December 2019, 13 patients underwent the reconstruction of soft tissue defects in various sites using the lower trapezius musculocutaneous flap carrying a portion of the latissimus dorsi muscle. There were 10 males and 3 females, with an average age of 52.1 years (range, 29-83 years). Twelve wounds were resulted from resection of various malignant tumor, including 6 cases of head and neck tumors, 5 cases of shoulder and back tumors, 1 case of chest and back tumors. Among the 12 cases, 4 cases were complicated with wound infection and bone exposure, 1 case with skull defect and cerebral dura exposure, and 1 case with wound infection, skull necrosis, and cerebrospinal fluid leakage. One case was injured in a traffic accident, which resulted in infection wound in the back and shoulder and bone exposure. The sizes of the defect and musculocutaneous flap ranged from 11 cm×7 cm to 23 cm×15 cm and 25 cm×8 cm to 40 cm×14 cm, respectively. According to the spatial relationship between the donor and recipient sites, propeller flaps (8 cases) or percutaneous tunnel island flaps (5 cases) were used to transfer the myocutaneous flap to the recipient area to repair the wound. The donor site was directly closed and sutured in 9 cases, and those with excessive tension were repaired with free skin grafts in 2 cases or transferred flaps in 2 cases.ResultsAfter the operation, necrosis of the distal 4-cm of the musculocutaneous flap occurred in 2 cases. After debridement, the resultant wounds were reconstructed using a local flap and a posterior intercostal artery perforator flap, respectively. The remaining 11 myocutaneous flaps survived completely without arteries and veins crisis. The wounds in the donor and recipient areas healed by first intention. All the patients were followed up 1 to 48 months (mean, 7.4 months). The color and texture of the flap were good. During the follow-up, 1 patient underwent tumor resection again due to tumor recurrence, and 1 patient with a scalp hemangiosarcoma died due to unexplained thoracic hemorrhage. Tumor recurrence was not found in the remaining patients. The musculocutaneous flap coverage was stable and the infection was controlled.ConclusionThe lower trapezius musculocutaneous flap carrying a portion of the latissimus dorsi muscle can be an alternation option to reconstruct refractory wounds with exposure of vital structures and organs and infection.
ObjectiveTo investigate the effects of micro-fracture and insul in-l ike growth factor 1 (IGF-1) in treatment of articular cartilage defect in rabbits. MethodsTwenty-four New Zealand white rabbits (aged, 4-6 months; weighing, 2.5-3.5 kg) were randomly divided into 4 groups (n=6):micro-fractures and recombinant human IGF-1 (rhIGF-1) treatment group (group A), micro-fracture control group (group B), rhIGF-1 treatment control group (group C), and blank control group (group D). Full thickness articular cartilage defects of 8 mm×6 mm in size were created in the bilateral femoral condyles of all rabbits. The micro-fracture surgery was performed in groups A and B. The 0.1 mL rhIGF-1 (0.01 μg/μL) was injected into the knee cavity in groups A and C at 3 times a week for 4 weeks after operation, while 0.1 mL sal ine was injected in groups B and D at the same time points. At 4, 12, and 24 weeks, the gross, histological, and immunohistochemical observations were performed, and histological score also was processed according to Wakitani's score criteria. The collagen contents in the repair tissues and normal patellofemoral cartilage were detected by the improved hydroxyproline (HPR) method at 24 weeks. Electron microscope was used to observe repair tissues of groups A and B at 24 weeks. Results All animals were survival at the end of experiment. At 24 weeks after operation, defect was repaired with time, and the repair tissue was similar to normal cartilage in group A; the repair tissue was even without boundary with normal cartilage in group B; and the repair tissue was uneven with clear boundary with normal cartilage in groups C and D. Histological staining showed that the repair tissues had no difference with normal cartilage in group A; many oval chondrocytes-l ike cells and l ight-colored matrix were seen in the repair tissues of group B; only a few small spindle-shaped fibroblasts were seen in groups C and D. Moreover, histological scores of group A were significantly better than those of groups B, C, and D (P<0.05) at 4, 12, and 24 weeks. Electron microscope observation showed that a large number of lacuna were seen on the surface of repair tissue in group A, and chondrocytes contained glycogen granules were located in lacunae, and were surrounded with the collagen fibers, which was better than that in group B. Collagen content of the repair tissue in group A was significantly higher than that in groups B, C, and D (P<0.05), but it was significantly lower than that of normal cartilage (P<0.05). Conclusion Combination of micro-fracture and rhIGF-1 for the treatment of full thickness articular cartilage defects could promote the repair of defects by hyaline cartilage.
Objective To explore the effect of short-term low-frequency electrical stimulation (SLES) during operation on nerve regeneration in delayed peripheral nerve injury with long gap. Methods Thirty female adult Sprague Dawley rats, weighing 160-180 g, were used to prepare 13-mm defect model by trimming the nerve stumps. Then all rats were randomly divided into 2 groups, 15 rats in each group. After nerve defect was bridged by the contralateral normal sciatic nerve, SLES was applied in the experimental group, but was not in the control group. The spinal cords and dorsal root ganglions (DRGs) were harvested to carry out immunofluorescence histochemistry double staining for growth-associated proteins 43 (GAP-43) and brain-derived neurotrophic factor (BDNF) at 1, 2, and 7 days after repair. Fluorogold (FG) retrograde tracing was performed at 3 months after repair. The mid-portion regenerated segments were harvested to perform Meyer’s trichrome staining, immunofluorescence double staining for neurofilament (NF) and soluble protein 100 (S-100) on the transversely or longitudinal sections at 3 months after repair. The segment of the distal sciatic nerve trunk was harvested for electron microscopy and morphometric analyses to measure the diameter of the myelinated axons, thickness of myelin sheaths, the G ratio, and the density of the myelinated nerve fibers. The gastrocnemius muscles of the operated sides were harvested to measure the relative wet weight ratios. Karnovsky-Root cholinesterase staining of the motor endplate was carried out. Results In the experimental group, the expressions of GAP-43 and BDNF were higher than those in the control group at 1 and 2 days after repair. The number of labeled neurons in the anterior horn of gray matter in the spinal cord and DRGs at the operated side from the experimental group was more than that from the control group. Meyer’s trichrome staining, immunofluorescence double staining, and the electron microscopy observation showed that the regenerated nerves were observed to develop better in the experimental group than the control group. The relative wet weight ratio of experimental group was significantly higher than that of the control group (t=4.633,P=0.000). The size and the shape of the motor endplates in the experimental group were better than those in the control group. Conclusion SLES can promote the regeneration ability of the short-term (1 month) delayed nerve injury with long gap to a certain extent.
ObjectiveTo objectively evaluate the effectiveness of the ventricular fold pull-down combined with strip myofascial flap to repair laryngeal defect after early glottic carcinoma operation with glottic morphological parameters and voice parameters. MethodsBetween January 2008 and December 2012, 47 patients with early glottic carcinoma and anterior commissure involvement underwent partial laryngectomy. All patients were male, aged from 60 to 75 years (mean, 68.5 years). The disease duration was 4-11 months (mean, 7.2 months). According to American Joint Committee on Cancer (AJCC) TNM criteria, 28 cases were classified as T1aN0M0, 14 cases as T1bN0M0, and 5 cases as T2N0M0. Laryngeal defect after resection of tumor was repaired by ventricular fold pull-down combined with strip myofascial flap. At 1 day before operation and at 1 year after operation, multilayer spiral CT was used to scan larynx, to measure and compare the anteroposterior diameter of vocal area, the distance between both sides of the vocal process, and the thickness of soft tissue of vocal area, and the effect of combined soft tissue flap was objectively assessed in laryngeal morphology reconstruction. The actual voice parameters[including F0, Jitter, Shimmer, normalized noise energy (NNE), and maximum phonatory time (MPT)] were tested and compared, and the effect of the combined soft tissue flap on postoperative laryngeal pronunciation was evaluated. ResultsPostoperative pathological examination revealed well-differentiated squamous cell carcinoma in 38 cases, and moderately-differentiated squamous cell carcinoma in 9 cases; no tumor was found in the resection margin. Healing of neck incision was obtained in all patients at 7-9 days after operation. Forty-four cases were decannulated at 9-11 days after operation and the remaining 3 cases were decannulated at 3 weeks after operation. Oral feeding usually started in all cases at 3-4 days after operation. All patients were followed up 1 year. At 1 year after operation, the anteroposterior diameter of vocal area was significantly reduced when compared with preoperative one (t=15.161, P=0.000); the distance between both sides of the vocal process and the thickness of soft tissue of vocal area had no significant changes (P > 0.05). Compared with preoperative ones, there were significant differences in Shimmer, NNE, and MPT (P < 0.05), but no significant difference was found in F0 and Jitter (P > 0.05) at 1 year after operation. ConclusionVentricular fold pull-down combined with strip myofascial flap can repair laryngeal defect effectively after partial laryngectomy and maintain the effective airway after operation. It not only has no effect on postoperative laryngeal morphology, but also can be used as new laryngeal voice vibration body.
ObjectiveTo investigate the feasibility of tissue engineered periosteum (TEP) constructed by porcine small intestinal submucosa (SIS) and bone marrow mesenchymal stem cells (BMSCs) of rabbit to repair the large irregular bone defects in allogenic rabbits. MethodsThe BMSCs were cultivated from the bone marrow of New Zealand white rabbits (aged, 2 weeks-1 month). SIS was fabricated by porcine proximal jejunum. The TEP constructed by SIS scaffold and BMSCs was prepared in vitro. Eighteen 6-month-old New Zealand white rabbits whose scapula was incompletely resected to establish one side large irregular bone defects (3 cm×3 cm) model. The bone defects were repaired with TEP (experimental group,n=9) and SIS (control group,n=9), respectively. At 8 weeks after operation, the rabbits were sacrificed, and the implants were harvested. The general condition of the rabbits was observed; X-ray radiography and score according to Lane-Sandhu criteria, and histological examination (HE staining and Masson staining) were performed. ResultsAfter operation, all animals had normal behavior and diet; the incision healed normally. The X-ray results showed new bone formation with normal bone density in the defect area of experimental group; but no bone formation was observed in control group. The X-ray score was 6.67±0.32 in experimental group and was 0.32±0.04 in control group, showing significant difference (t=19.871,P=0.001). The general observation of the specimens showed bone healing at both ends of the defect, and the defect was filled by new bone in experimental group; no new bone formed in the control group. The histological staining showed new bone tissue where there were a lot of new vessels and medullary cavity, and no macrophages or lymphocytes infiltration was observed in the defect area of experimental group; only some connective tissue was found in the control group. ConclusionTEP constructed by porcine SIS and BMSCs of rabbit can form new bone in allogenic rabbit and has the feasibility to repair the large irregular bone defects.
ObjectiveTo explore the application of three-dimensional (3-D) printing technique in repair and reconstruction of maxillofacial bone defect. MethodsThe related literature on the recent advance in the application of 3-D printing technique for repair and reconstructing maxillofacial bone defect was reviewed and summarized in the following aspects:3-D models for teaching, preoperative planning, and practicing; surgical templates for accurate positioning during operation; individual implantable prosthetics for repair and reconstructing the maxillofacial bone defect. Results3-D printing technique is profoundly affecting the treatment level in repair and reconstruction of maxillofacial bone defect. Conclusion3-D printing technique will promote the development of the repair and reconstructing maxillofacial bone defect toward more accurate, personalized, and safer surgery.
Objective To discuss the role of heparan sulfate (HS) in bone formation and bone remodeling and summarize the research progress in the osteogenic mechanism of HS. Methods The domestic and abroad related literature about HS acting on osteoblast cell line in vitro, HS and HS composite scaffold materials acting on the ani-mal bone defect models, and the effect of HS proteoglycans on bone development were summarized and analyzed. Results Many growth factors involved in fracture healing especially heparin-binding growth factors, such as fibroblast growth factors, bone morphogenetic protein, and transforming growth factor β, are connected noncovalently with long HS chains. HS proteoglycans protect these proteins from protease degradation and are directly involved in the regulation of growth factors signaling and bone cell function. HS can promote the differentiation of stem cells into osteoblasts and enhance the differentiation of osteoblasts. In bone matrix, HS plays a significant role in promoting the formation, maintaining the stability, and accelerating the mineralization. Conclusion The osteogenesis of HS is pronounced. HS is likely to become the clinical treatment measures of fracture nonunion or delayed union, and is expected to provide more choices for bone tissue engineering with identification of its long-term safety.
ObjectiveTo review the methods of improving the mechanical properties of hydrogels and the research progress in bone tissue engineering. MethodsThe recent domestic and foreign literature on hydrogels in bone tissue engineering was reviewed, and the methods of improving the mechanical properties of hydrogels and the effect of bone repair in vivo and in vitro were summarized. ResultsHydrogels are widely used in bone tissue engineering, but their mechanical properties are poor. Improving the mechanical properties of hydrogels can enhance bone repair. The methods of improving the mechanical properties of hydrogels include the construction of dual network structures, inorganic nanoparticle composites, introduction of conductive materials, and fiber network reinforcement. These methods can improve the mechanical properties of hydrogels to various degrees while also demonstrating a significant bone repair impact. ConclusionThe mechanical properties of hydrogels can be effectively improved by modifying the system, components, and fiber structure, and bone repair can be effectively promoted.