BACKGROUND: Scar hypertrophy is always followed by the wound healing in burn and trauma. Endothelial cells play a key role in scar hypertrophy, so inhibitory growth of endothelial cells can relieve scar hypertrophy to a certain degree. OBJECTIVE: To construct a recombinant adenovirus vector expressing human endostatin (Ad/hEnd), and to investigate the cooperative effect of Ad/hEnd and keratinocyte on endothelial cell proliferation. DESIGN, TIME AND SETTING: Observational study, which was performed in the State Key Laboratory of Trauma, Burn and Combined Injury, Institute of Burn Research, Southwest Hospital of the Third Military Medical University of Chinese PLA between September 2006 and May 2007. MATERIALS: pAdTrack-CMV and pAdEasy-1 were obtained from Stratagene Company, USA; 293 cell and Ecoli.DH5α were stored in our laboratory. METHODS: The endostatin gene sequence was obtained by reverse transcription-polymerase chain reaction (RT-PCR) and polymerase chain reaction (PCR) based on mRNA of human fetal hepatic tissue and inserted into the adenovims shuttle plasmid pAdTrack-CMV to obtain recombinant plasmid pAdTrack-ES. After identification, positive recon was transformed into pAdeasy 1 recipient virus to screen positive clones. The adenovirus Ad/hEnd was generated from 293 cells and identified by PCR and fluorescence microscope. Then the keratinocytes were infected with Ad/hEnd, and co-cultured with endothelial cells by nest dish culture method. The content of endostatin was detected, and the non-transfection keratinocytes were used as the controls. MAIN OUTCOME MEASURES: Homologous recombination and identification of pAd/hEnd; generation and identification of Ad/hEnd; endostatin expression after 293 cell transfection; purification and titer measurement of Ad/hEnd; content of endostatin in culture solution; apoptotic percentage of endothelial cells; inhibitory ratio of endothelial cells. RESULTS: Ad/hEnd was constructed and the virus titer was generally up to 1.65×1012 PFU/L. Ad/hEnd-infected keratinocytes could effectively express and secrete endostatin of which the content reached 226 μg/L after 3 days of co-culture. The apoptotic percentage and inhibitory ratio of the endothelial cells co-cultured with Ad/hEnd-infected keratinocytes were significantly higher than those in control group (P<0.05). CONCLUSION: Ad/hEnd-infected keratinocytes co-cultured with endothelial cells can promote apoptosis and inhibit proliferation of endothelial cells through excretion of endostatin.

Objective To establish the quality standard for Tiaojing Pills. Methods Radix Angelicae Sinensis and Rhizoma Chuanxiong in Tiaojing Pills were identified by TLC and the content of paeoniflorin was determined by HPLC. Results Radix Angelicae Sinensis and Rhizoma Chuanxiong could be identified by TLC. Paeoniflorin showed a good linearity in the range of 0.086 88～0.868 8 ?g,r=0.999 6.The average recovery was 101.28 %,and RSD was 1.31 %. Conclusion The established methods are simple,convenient and reproducible,and can be used for the quality control of Tiaojing Pills.

Intervertebral disc (IVD) degeneration is one of the major causes of low back pain. As current clinical treatments are aimed at restoring biomechanical function and providing symptomatic relief, the methods focused on biological repair have aroused interest and several tissue engineering approaches using different cell types have been proposed. Owing to the unsuitable nature of degenerate cells for tissue engineering, attention has been given to the use of mesenchymal stem cells (MSCs). In this connection, we have made a study on the characteristics of MSCs derived from adult bone marrow and on the feasibility of constructing IVD tissue-engineering cell under a Three-Dimensional Pellet Culture System. The human bone marrow MSCs were isolated and purified with density gradient solution and attachment-independent culture system. MSCs isolated using this method are a homogeneous population as indicated by morphology and other criteria. They have the capacity for self-renewal and proliferation, and the multilineage potential to differentiate.
Adolescent ; Adult ; Bone Marrow Cells ; cytology ; Cell Culture Techniques ; methods ; Cells, Cultured ; Chondrogenesis ; physiology ; Humans ; Intervertebral Disc ; Intervertebral Disc Degeneration ; therapy ; Mesenchymal Stromal Cells ; cytology ; Tissue Engineering ; methods ; Young Adult