ABSTRACT
BACKGROUND: Choriocarcinoma is a kind of trophoblastic neoplasm with highly aggressive phenotypes. Forkhead box D3 (FoxD3) is an embryonic and trophoblastic stem cel transcription factor. It plays important roles in different physical and pathological situations such as embryogenesis, carcinogenesis and tumor progression. OBJECTIVE:To investigate the role of FoxD3 in choriocarcinoma malignancy and the possible mechanism. METHODS:The human choriocarcinoma JAR cel line was employed in this study. The mRNA and protein expressions of genes were measured by quantitative RT-PCR (qRT-PCR) and western blot, respectively. The FoxD3 specific short hair RNA was applied to down-regulate gene expression. The cel proliferation was evaluatedin vitro by cel counting assay andin vivo by tumor growth. The migration/invasion was determined by transwel assay. The profile of FoxD3 targeted genes was investigated with an Agilent microarray and verified by qRT-PCR. RESULTS AND CONCLUSION: The FoxD3 mRNA and protein expressions in JAR cells were significantly higher than those in primarily cultured normal trophoblastic cells. Knockdown of FoxD3 by short hair RNA in JAR cells could inhibit cell proliferation and migration/invasionin vitro, and suppress thetumor growth with decreased β-human chorionic gonadotropin secretionin vivo. A profile of seven focal adhesion molecules (ITGA5, ITGB6, THBS4, COL6A3, VTN, NRXN3 and NLGN1) was verified to be targeted by FoxD3. Furthermore, knockdown of FoxD3 by short hair RNA could decrease the activation of focal adhesion kinase. All these findings suggest the overexpression of FoxD3 can contribute to the aggressive phenotype of choriocarcinoma JAR cells by regulating the profile of focal adhesion molecules and focal adhesion kinase.
ABSTRACT
BACKGROUND:Fetal liver stem cel s have the potential to differentiate into hepatocytes and bile duct cel s, and participate in the repair and reconstruction of the liver, which is an important source of hepatocytes. But there are a little amount of fetal liver stem cel s in human body, and how to obtain a certain number of high-purity fetal liver stem cel s is currently a hot research. OBJECTIVE:To construct a siRNA carrier that can effectively inhibit the expression of connexin 43 (Cx43) in rat fetal liver stem cel s, and to investigate the effect of Cx43 inhibition on the proliferation and cel cycle of fetal liver stem cel s cultured in vitro. METHODS:Fetal liver stem cel s were cultured by the suspension culture in vitro, siRNA sequences targeting Cx43 (Cx43-siRNA) and negative control sequence (NC-siRNA) were designed and synthesized. Then, rat fetal liver stem cel s were transferred electrophoretical y and divided into three groups:blank group, NC-siRNA group, Cx43-siRNA group. Real-time PCR and western blot were used to assess the knockdown efficiency. Cel ular proliferation was determined by cel growth curve and cel counting kit-8 assay. The cel cycle was analyzed by flow cytometry. RESULTS AND CONCLUSION:After transfection, the Cx43 gene and protein expression levels were declined dramatical y in the Cx43-siRNA, NC-siRNA and blank groups, and the cel s grew faster. The number of cel s at G0/G1 phase decrease, but the number of cel s in S phase increased. There were significant differences between the groups (P<0.05). Electrophoretic transfer of Cx43-siRNA can promote the proliferation of cultured fetal liver stem cel s and optimize the cel culture.
ABSTRACT
BACKGROUND:The surface antigen CD80/CD86 on mature dendritic cells can activate helper T (Th) cells, reduce the differentiation of Th cells toward Th1 cells, and promote the differentiation of Th cells toward Th2 cells. OBJECTIVE:To investigate the effect of smal interfering RNA (siRNA) inhibiting the expression of surface antigens CD80/CD86 from asthmatic murine mature dendritic cells on Th1/Th2 type cytokines, interferon-γand interleukin-4. METHODS:Asthmatic model of mice was established;then bone marrow-derived mature dendritic cells were separated and cultured. The expression of CD11c, CD80 and CD86 on mature dendritic cells were examined by flow cytometry. The siRNA was transferred into mature dendritic cells of asthmatic mice, and the CD80/CD86 mRNA and protein expression before and after interference were determined by fluorescent quantitative PCR and flow cytometry. The mature dendritic cells in non-siRNA group, siRNA group and negative siRNA group were co-cultured with T cells. The interferon-γand interleukin-4 productions were measured by enzyme-linked immunosorbent assay. RESULTS AND CONCLUSION:(1) The expression of CD80/CD86 on the mature dendritic cells of asthmatic group was significantly higher than that in normal control group (al P<0.05). (2) After siRNA was transferred into mature dendritic cells, the expression level of CD80/CD86 mRNA and protein in siRNA group was significantly lower than other groups (al P<0.05). (3) After siRNA transfection, the level of interferon-γfrom the supernatant of mature dendritic cells and T cells co-culture system was significantly increased in the siRNA group compared with other groups (al P<0.05), while interleukin-4 production in the siRNA group was significantly decreased (al P<0.05). These findings suggest that high expression of CD80/CD86 on mature dendritic cells of asthmatic mice is observed, specific siRNA can effectively inhibit the expression of CD80/CD86, thus increasing interferon-γproduction and decreasing interleukin-4 production, which contributes to regulate the Th1/Th2 imbalance.
