Transfection of adeno-associated virus encoding beta-nerve growth factor into endothelial progenitor cells / 中国组织工程研究

BACKGROUND:Nowadays, gene therapy has become a new trend for disease therapy and brought promise for some refractory diseases. Its key is to choose proper cel s, genes and vectors. OBJECTIVE:To use recombinant adeno-associated virus mediatedβ-nerve growth factor (β-NGF) to transfect rat bone marrow-derived endothelial progenitor cel s in vitro, and to investigate the effect ofβ-NGF expression on the proliferation of endothelial progenitor cel s. METHODS:The endothelial progenitor cel s were isolated, cultured and identified from the bone marrow of rats. Empty vector or recombinant adenovirus-associated virus containingβ-NGF gene was transferred into endothelial progenitor cel s. We examined the transfection efficiency by fluorescence expression of green fluorescent protein. Expression ofβ-NGF protein was detected using ELISA, and its effect on the proliferation of endothelial progenitor cel s was determined using MTT method. RESULTS AND CONCLUSION:Rat endothelial progenitor cel s were isolated and cultured successful y in vitro and were identified positive by the function of cel s and immunofluorescence staining. The endothelial progenitor cel s were infected directly by the recombinant adenovirus-associated virus containingβ-NGF gene with an efficiency of 65.3%.β-NGF protein was detected in the culture supernatant of transfected endothelial progenitor cel s, which reached a high level at 10 days after gene transfection. Furthermore, there was noβ-NGF protein in the blank and empty vector groups. After transfection, the proliferative ability of endothelial progenitor cel s was increased, which was significantly higher than the blank and empty vector groups (P0.05). These findings suggest that recombinant adenovirus-associated virus containingβ-NGF gene can be successful y transferred into rat bone marrow-derived endothelial progenitor cel s and promote the proliferation of endothelial progenitor cel s.

Differential Expression of Sox9 in Conventional Chondrosarcoma and Dedifferentiated Chondrosarcoma / 中国肿瘤临床

Objective: To investigate the differential expression of Sox9 in conventional chondrosarcoma,dedifferentiated chondrosarcoma and normal cartilage. Methods: We reported 12 cases of chondrosarcomas,which were initially diagnosed as chondrosarcomas(6 cases of conventional chondrosarcoma and 6 cases of dedifferentiated chondrosarcoma)at Peking University People's Hospital between January 2003 and January 2007.We used genechip method to identify difierentially expressed genes involved in conventional chondrosarcoma,dedifferentiated chondrosarcoma and in normal cartilage(6 cases)and found thousands of differentially expressed genes after extensive statistical analysis.With Sox9 which played crucial roles in the process of both differentiation and maturation of chondrocyte as a candidate,we used Real-time PCR,Westem blot and immunohistochemistry to confirm the results found by gene chip. Results: DNA microarray results showed that Sox9 was up-regulated about 1.6 times in conventional chondrosarcoma compared with that in normal cartilage.But in dedifferentiated chondrosarcoma,the expression level of Sox9 was significantly down-regulated,0.082 times of that in normal cartilage.Real-time PCR results showed that the expression levels of Sox9 mRNA in conventional chondrosarcomas and dedifferentiated chondrosarcomas were 1.68±0.119 and 0.088±0.017,respectively.Sox9 protein level was significantly higher in humen conventional chondrosarcomas than that in normal cartilage.Sox9 protein level in dedifferentiated chondrosarcomas was significantly lower than that in normal cartilage tissue.All of the 6 cases of conventional chondrosarcomas showed diffuse and strong staining of Sox9.However,Only scattered staining was observed in dedifferentiated chondrosarcomas. Conclusion: Compared with that in normal cartilage,Sox9 expression is up-regulated in conventional chondrosarcomas and down-regulated in dedifferentiated chondrosarcomas.Decrease of Sox9 expression in dedifferentiated chondrosarcoma is correlated with poor survival,indicating that Sox9 may serve as a molecular prognostic marker for chondrosarcomas and disease progression.

Lethal effects of nanoliposome encapsulated cisplatin on Saos-2 cells and its distribution in nude mice bearing human osteosarcoma xenografts / 北京大学学报(医学版)

Objective:To investigate the killing effect of nanoliposome encapsulated cisplatin(NLE-CDDP) on human osteosarcoma cell line Saos-2 and explore the distribution of platinum(Pt) in tumor-bearing mice.Methods: Saos-2 cells were cultured at different concentrations of NLE-CDDP.MTT assay,inverted microscopic observation and flow cytometry assay(FCM)were used to observe the antiproli-ferative effect of NLE-CDDP on the human osteosarcoma cells.Antitumor effect of NLE-CDDP was determined using the xenografts models of human osteosarcoma cell Saos-2 in nude mice.The Pt concentration in the tissues of tumor-transplanted mice was determined by atomic spectrophotometer.Results: When treated at different concentrations of NLE-CDDP for 24-96 hours,the survival rate of Saos-2 cells decreased significantly(P