Development of a centrally vascularized tissue engineering bone graft with the unique core-shell composite structure for large femoral bone defect treatment.

Great effort has been spent to promote the vascularization of tissue engineering bone grafts (TEBG) for improved therapeutic outcome. However, the thorough vascularization especially in the central region still remained as a major challenge for the clinical translation of TEBG. Here, we developed a new strategy to construct a centrally vascularized TEBG (CV-TEBG) with unique core-shell composite structure, which is consisted of an angiogenic core and an osteogenic shell. The in vivo evaluation in rabbit critical sized femoral defect was conducted to meticulously compare CV-TEBG to other TEBG designs (TEBG with osteogenic shell alone, or angiogenic core alone or angiogenic core+shell). Microfil-enhanced micro-CT analysis has been shown that CV-TEBG could outperform TEBG with pure osteogenic or angiogenic component for neo-vascularization. CV-TEBG achieved a much higher and more homogenous vascularization throughout the whole scaffold (1.52-38.91 folds, p < 0.01), and generated a unique burrito-like vascular network structure to perfuse both the central and peripheral regions of TEBG, indicating a potential synergistic effect between the osteogenic shell and angiogenic core in CV-TEBG to enhance neo-vascularization. Moreover, CV-TEBG has generated more new bone tissue than other groups (1.99-83.50 folds, p < 0.01), achieved successful bridging defect with the formation of both cortical bone like tissue externally and cancellous bone like tissue internally, and restored approximately 80% of the stiffness of the defected femur (benchmarked to the intact femur). It has been further observed that different bone regeneration patterns occurred in different TEBG implants and closely related to their vascularization patterns, revealing the potential profound influence of vascularization patterns on the osteogenesis pattern during defect healing.

[Clinical significance of PCT, CRP, ESR, WBC count as predictors in postoperative early infectious complications with fever after posterior lumbar internal fixation].

OBJECTIVE: To evaluate the role of serum level of procalcitonin (PCT), C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and white blood cell count (WBC) as predictors in postoperative early infectious complications with fever after posterior lumbar internal fixation (PLIF). METHODS: A retrospective study was conducted from January 2012 to January 2014. Fifty-two patients with fever in the early stage(within 10 days) after the PLIF were collected in the study. They were divided into infection group and non-infection group (group A and group B) according to the results of postoperative blood culture. There were 26 patients in group A and 32 patients in group B. The values of PCT, CRP, ESR, and WBC count were compared and analyzed between two groups. RESULTS: The values of PCT, CRP, and ESR in group A were higher than those of group B. Meanwhile, CRP and ESR in group B were still higher than the normal range. Among the 26 patients with infections (group A), PCT was superior to CRP and ESR, had a good ability in discriminating different kinds of postoperative infections. The area under the ROC curve of serum PCT levels was the largest (CI 95% was 0.81 to 0.98) in the indexs; and ROC curve of WBC count was no statistically significant. When the cut off points of each predictors were evaluated, the higher sensitive was CRP and reached at 90.27% and the higher specific was ESR and reached at 88.50%. CONCLUSION: For the patients with fever at the early stage after the PLIF should be paid attention and reasonable choosing predictors are helpful to identify postoperative infection in the early stage. The CRP and ESR may be influenced by the surgery, and the PCT level is helpful to differentiate infection type.

Advanced oxidation protein products induce inflammatory response and insulin resistance in cultured adipocytes via induction of endoplasmic reticulum stress.

Accumulation of advanced oxidation protein products (AOPPs) is prevalent in metabolic syndrome and type 2 diabetes. Adipocyte dysfunction has been recognized as a link between these conditions. To examine the effect of AOPPs on adipocyte perturbation, 3T3-L1 adipocytes were treated with increased levels of AOPPs as seen in these conditions. Exposure of adipocytes to AOPPs induced overexpression of tumor necrosis factor α and interleukin-6. This inflammatory response was completely blocked by nuclear factor-κB inhibitor SN50. AOPPs challenge also impaired insulin signaling, which was partly prevented by SN50. Treatment with AOPPs triggered endoplasmic reticulum (ER) stress, revealed by phosphorylation of PKR-like eukaryotic initiation factor 2α kinase, eukaryotic translational initiation factor 2α, inositol-requiring enzyme 1 and c-jun N-terminal kinase, and by overexpression of glucose regulated protein 78. AOPPs-induced ER stress was mediated by reactive oxygen species (ROS) generated by activation of NADPH oxidase since it was prevented by NADPH oxidase inhibitors or ROS scavenger. Treating the cells with inhibitors of NADPH oxidase or ER stress could completely abolish AOPPs-induced overexpression of adipocytokines and insulin resistance, suggesting that AOPPs induced adipocyte perturbation probably through induction of ROS-dependent ER stress. Our data identified AOPPs as a class of important mediator of adipocyte perturbation. Accumulation of AOPPs might be involved in adipocyte dysfunction as seen in metabolic syndrome and type 2 diabetes.

[Tissue engineering vascularized bone repairing segmental femoral bone defects in rabbits].

OBJECTIVE: To investigate the effectiveness and mechanism of tissue engineering vascularized bone in repairing segmental femoral bone defects in rabbits. METHODS: Thirty-two rabbits were randomized into two groups (n = 16 each). A segmental and critical bone defect of 15 mm in length was made at left femur. In experimental group, the tissue engineering bone constructed from autologous bone marrow mesenchymal stem cells plus beta-tricalcium phosphate (beta-TCP) and vascular bundle was implanted into bony defect. In control group, there was no implantation of vascular bundle. Animals were sacrificed at 2, 4, 8 and 12 weeks post-implantation respectively. Histological observation was conducted to determine the process of new bone formation and remodeling. The expression of vascular endothelial growth factor (VEGF) in new bone was measured by immunohistochemistry, real-time PCR and Western blot. RESULTS: As indicated by histological observations over time, new bone formation increased in both groups. It was better in the experimental group than the control group at the beginning of 4 weeks. The expression level of VEGF gradually decreased in each group after an initial rise. And the expression of VEGF was significantly higher than the control group after implantation at all time points and peaked at 4 weeks. CONCLUSION: Tissue engineering vascularized bone accelerates bone repair in critical size defect model of femur in rabbit. Implantation of vascular bundle can promote the secretion of VEGF. And VEGF is an essential mediator of both angiogenesis and ossification.

Osteogenesis and angiogenesis of tissue-engineered bone constructed by prevascularized ß-tricalcium phosphate scaffold and mesenchymal stem cells.

Although vascularized tissue-engineered bone grafts (TEBG) have been generated ectopically in several studies, the use of prevascularized TEBG for segmental bone defect repair are rarely reported. In current study, we investigated the efficacy of prevascularized TEBG for segmental defect repair. The segmental defects of 15 mm in length were created in the femurs of rabbits bilaterally. In treatment group, the osteotomy site of femur was implanted with prevascularized TEBG, which is generated by seeding mesenchymal stem cells (MSCs) into ß-TCP scaffold, and prevascularization with the insertion of femoral vascular bundle into the side groove of scaffold; whereas in the control group, only MSC mediated scaffolds (TEBG) were implanted. The new bone formation and vascularization were investigated and furthermore, the expression of endogenous vascular endothelial growth factor (VEGF) which might express during defect healing was evaluated, as well. At 4, 8, and 12 weeks postoperatively, the treatment of prevascularized TEBG led to significantly higher volume of regenerated bone and larger amount of capillary infiltration compared to non-vascularized TEBG. The expression of VEGF in mRNA and protein levels increased with implantation time and peaked at 4 weeks postoperatively, followed by a slow decrease, however, treatment group expressed a significant higher level of VEGF than control group throughout the whole study. In conclusion, this study demonstrated that prevascularized TEBG by insertion of vascular bundle could significantly promote the new bone regeneration and vascularization compared to non-vascularized TEBG, which could be partially explained by the up-regulated expression of VEGF.