Guo Fengjin

Effects of icariin on expression of OPN mRNA and type I collegen in rat osteoblastsin vitro

SummaryTo study the effects of Icariin on expression of osteopontin (OPN) mRNA and type I collagen in rat osteoblastsin vitro and to explore its possible mechanisms in preventing osteoporosis. OB was isolated from calvaria of new-born new-born fetal Sprague-Dawley (SD) rats by means of modified sequential collagenase digestion and incubated in MEM medium and the cell morphology was observed under inverted phase contrast microscope, OB was identified by alkaline phosphatase (ALP) staining. Different concentration (0.1 μg/mL, 1.0 μg/mL, 10 μg/mL) of Icariin was added to the OB and incubated. The effect of Icariin on the proliferation and osteogenesis of OB was monitored by MTT analysis. The expression of type I collagen was estimated with immunohistochemistry techniques. The expression levels of mRNA of OPN in the cells in every group were examined by reverse-transcriptase polymerase chain reaction (RT-PCR). The expression of OPN mRNA and type I collagen was strengthened gradually with the increase of Icariin concentration and peaked with 10 μg/mL Icariin on the 5th day. Icariin could significantly promote the expression of OPN mRNA and type I collagen in rat osteoblastsin vitro. The levels of expression of OPN mRNA and type I collagen were changed with different concentration of Icariin. Icariin could effectively prevent and treat osteoporosis and promote the bone formation.

Inhibition of Proliferation of Human Osteosarcoma Cells Transfected with PIN1 Antisense Gene

Objective: To evaluate the inhibition of proliferation of human osteosarcoma cells transfected with Pin1 anti-sense gene. Methods: Different doses of antisense Pin1 gene (0, 20, 50, 100, 200, 250 µL) were transfected into osteosarcoma MG-63 cells. The cells and culture supernatant before and after transfection were collected. The curve of cell growth was made by MTT method. The cell growth cycle and apoptosis were detected by FCM. The expression of Pin1 was detected by Western-blot and that of Pin1 mRNA by polymerase chain reaction (RT-PCR) respectively. Results: MTT and FCM assays indicated that the transfection by antisense Pin1 gene could inhibit MG-63 proliferation and induce apoptosis. Western-blot assays revealed that the antisense Pin1 gene-transfected MG-63 cells had weaker staining than those without transfected with antisense Pin1 gene, and staining intensity was negatively related with doses. The cells transfected by different doses of gene (0, 20, 50, 100, 200, 250 µL) had different absorbance rate: 0.854±0.136, 0.866±0.138, 0.732±0.154, 0.611±0.121, 0.547±0.109, 0.398±0.113, 0.320±0.151 respectively, with the difference being significant by F and q test (P<0.05). The expression of Pin1 mRNA had the similar results and its absorbance rate was 0.983±0.125, 0.988±0.127, 0.915±0.157, 0.786±0.125, 0.608±0.124, 0.433±0.130, 0.410±0.158 respectively (P <0.05). Conclusion: The expression of Pin1 mRNA in MG-63 cells could be inhibited by antisense Pin1 gene, so to reduce the expression of Pin1 and depress the proliferation of human osteosarcoma cells MG-63.

Relationship between the Expression of MTA1 Gene and Invasion and Metastasis of Human Osteosarcoma Cells

AbstractObjectiveTo compare the expression level of metastasis associated-1 (MTA1) in the higher and lower metastasis sublines of human osteosarcoma cells (MG63), and to investigate the relationship between the expression level of MTA1-EGFP and in vitro invasion and metastasis of human osteosarcoma cells.MethodsThe expression level of MTA1 in two sublines of MG63 cells was detected by semi-quantitative RT-PCR, and cell invasion assay and cell proliferation assay were used to evaluate the invasive capacity in vitro in two sublines. The lower metastasis line of MG-63 cells were transfected with MTA1-EGFP full-length cDNA expression plasmid by lipofectamine. The changes of the MTA1-EGFP expression and in vitro invasion potential were measured after transfection.ResultsM8 subline expressed significantly higher level of MTA1 than that of M6 subline by RT-PCR. The invasive potentials of low metastasis MG63 cell line were increased after MTA1 gene transfection.ConclusionThere may be a relationship between MTA1 and invasive potentials of human osteosarcoma cells, and MTA1 may play a role in the molecular mechanism of tumor metastases and be a potential target for gene therapy of osteosarcoma. Further studies of MTA1 in human ostersarcoma cell metastasis are needed.

Expression of Integrin α4 in Osteosarcoma and Significance

AbstractObjective: To investigate the expression of integrin α4 in osteosarcoma and significance. Methods: Forty-six patients with osteosarcoma (Enneking I-III) were analyzed for the expression of α4 integrin subunit using immunohistochemical method. Results: Twenty-nine (63.04%) of 46 samples demonstrated positive (+-++) integrin α4 expression. Loss expression of integrin α4 was observed in the patients with advanced Enneking stage (P=0.0040) and with metastatic disease at presentation (P=0.0158). Integrin α4 expression correlated with cell differentiation, the level of malignancy and the invasive behavior of osteosarcoma. Conclusion: The loss expression of integrin α4 subunit might be a predictor indicating the invasive potential of osteosarcoma and play a role in metastasis of osteosarcoma patients.