RESUMO
OBJECTIVE: Three different kinds of transfection reagents were used to mediate the transfection of gadolinium-diethylenetriamine penta-acetic acid (Gd-DTPA) into human umbilical-cord-derived mesenchymal stem cells (hUCMSCs). The efficacy of different transfection reagents and the feasibility of NMR tracer in vitro of magnetized stem cells were estimated. METHODS: After purification by tissue explants adherent method, the biological characteristics of hUCMSCs in vitro were identified by subculture and amplification. Calcium phosphate, Effectene and liposome2000 were used to transfect Gd-DTPA-labeled hUCMSCs respectively, and cell counting was used to mediate the transfection of Gd-DTPA into hUCMSCs, which were then induced to lipoblast and osteoblast in vitro. The determination of the transfection activities of the transfection reagents was conducted by measuring the magnetic resonance imaging (MRI) signal intensity of the Gd-DTPA-labeled cells and the concentration of gadolinium ion in the cells. Furthermore, the relationship between the signal intensity of Gd-DTPA-labeled hUCMSCsMRI, cell subculture and generations was studied. RESULTS: Primary cells were obtained by tissue explants adherent for two weeks. The cells displayed a long spindle form and grew in swirl. After two passage generations, the cellular morphology became more homogeneous. The result detected by the flow cytometer showed that CD29C, D44, CD90, and CD105 were highly expressed, while no CD45, CD40, and HLA-DR expression was detected in the third generation cells. Directional induction in vitro caused the differentiation into lipoblast and osteoblast. After transfected by calcium phosphate, Effectene and liposome 2000, the signal intensity of stem cells was 2281.2±118.8, 2031.9±59.7 and 1887.4±40.8 measured by MRI. Differences between these three groups were statistically significant (P<0.05). The concentrations of gadolinium ion in three groups of stem cells were 0.178±0.009mg/L, 0.158±0.003mg/L and 0.120±0.002mg/L respectively, examined by inductively coupled plasma atomic emission spectrometry. No significant differences were found among these three groups (P<0.05). The proliferation and differentiation abilities of the Gd-DTPA-labeled stem cells were not affected. A minimum 5×10(4) Gd-DTPA-labeled stem cells could be traced with MRI in vitro and presented in high signal. The trace duration time in vitro was about 12days. CONCLUSIONS: Tissue explants adherent method can be availably applied to purify hUCMSCs. The Effectene method was proved to have the best transfection effect. The proliferation ability and differentiation potency of Gd-DTPA-labeled hUCMSCs were not affected, and the NMR of labeled stem cells in vitro was proved to be feasible.
Assuntos
Rastreamento de Células/métodos , Sangue Fetal/citologia , Gadolínio DTPA , Imageamento por Ressonância Magnética/métodos , Espectroscopia de Ressonância Magnética/métodos , Células-Tronco Mesenquimais/citologia , Células Cultivadas , Meios de Contraste , Transplante de Células-Tronco de Sangue do Cordão Umbilical/métodos , Estudos de Viabilidade , Gadolínio DTPA/farmacocinética , Humanos , Células-Tronco Mesenquimais/metabolismo , Coloração e Rotulagem/métodos , Transfecção/métodosRESUMO
BACKGROUND: Functional single nucleotide polymorphisms of x-ray repair cross-complementing protein 1 (XRCC1) have been suspected to contribute to uterine cervical cancer risk for a long time; however, most previous case-control studies were small sized and biased. Additionally, recent studies suggested that XRCC1 polymorphisms could be a biomarker of response to platinum-based chemotherapy. METHODS: A comprehensive search was conducted to retrieve eligible studies and odds ratios (ORs) and 95% confidence intervals (95% CIs) were calculated to measure association strength. RESULTS: A total of 13 studies were identified and analyzed. We found that the Arg194Trp polymorphism (Trp vs. Arg, OR=1.342, 95% CI: 1.176) was associated with increased risk of cervical cancer, while no significant association was found with Arg280His (His vs. Arg, OR=1.059, 95% CI: 0.863, 1.299) or Arg399Gln (Gln vs. Arg, OR=1.144, 95% CI: 0.938, 1.394). As for response to platinum- based chemotherapy, the variant XRCC1 399Gln allele (Gln vs. Arg, OR=0.345, 95% CI: 0.163, 0.729) was linked with a poor response; however, the Arg194Trp polymorphism (TrpArg vs. ArgArg, OR=6.421, 95% CI: 1.573, 26.205) predicted a good response. CONCLUSION: The Arg194Trp polymorphism of XRCC1 increases risk of cervical cancer; the variant 399Gln allele predicts poor response to platinum-based chemotherapy, while the Arg194Trp polymorphism indicates a good response.
