RESUMEN
BACKGROUND: Extracellular matrix has been shown to improve cell proliferation and reduce intracellular reactive oxygen species levels. However, there is little research on whether extracellular matrix can enhance the antioxidant capacity of umbilical cord stem cells to enhance their application in regenerative medicine and tissue engineering. OBJECTIVE: To investigate the effect of extracellular matrix on umbilical cord stem cell proliferation, antioxidant and osteogenic capacity. METHODS: The umbilical cord stem cells were divided into four groups. In the polystyrene group, the umbilical cord stem cells were cultured with ordinary polystyrene culture plate without other special treatment. In the extracellular matrix group, the umbilical cord stem cells were cultured with extracellular matrix without other special treatment. In the polystyrene + hydrogen peroxide group, the umbilical cord stem cells were cultured with polystyrene plate and pretreated with 200 μmol/L hydrogen peroxide for 2 hours. In the extracellular matrix + hydrogen peroxide group, umbilical cord stem cells were cultured with extracellular matrix and pretreated with 200 μmol/L hydrogen peroxide for 2 hours. The cells were pretreated with 200 μmol/L hydrogen peroxide for 2 hours. Proliferation capacity of umbilical cord stem cells was detected by CCK-8 assay. The cells were cultured for 72 hours after hydrogen peroxide pretreatment for 2 hours. The antioxidant capacity of umbilical cord stem cells was detected by flow cytometry and qRT-PCR. After 2 hours of hydrogen peroxide pretreatment, the cells were induced to differentiate into osteoblasts for 14 days. The osteogenic capacity of umbilical cord stem cells was detected by alizarin red staining and qRT-PCR. RESULTS AND CONCLUSION: The absorbance values of extracellular matrix group and extracellular matrix + hydrogen peroxide group were higher than that of polystyrene group and polystyrene + hydrogen peroxide group, respectively. The levels of reactive oxygen species in the polystyrene + hydrogen peroxide group and the polystyrene group were higher than those in the extracellular matrix group and the extracellular matrix + hydrogen peroxide group, respectively (P < 0.05). The expression levels of antioxidant enzyme-related genes SOD2 and CAT in the extracellular matrix group and extracellular matrix + hydrogen peroxide group were significantly higher than those in the polystyrene group and the polystyrene + hydrogen peroxide group, respectively (P < 0.05). The expression of bone related genes COL-1, RUNX2, OCN, and OSTERIX was highest in the extracellular matrix group, followed by the extracellular matrix + hydrogen peroxide group, and lowest in the polystyrene + hydrogen peroxide group; there was significant difference between the groups (P < 0.05). The results show that extracellular matrix can increase the proliferation capacity, antioxidant capacity and osteogenic differentiation potential of umbilical cord stem cells. It is a method for in vitro amplification and culture of cells with wide application prospects.
RESUMEN
Objective: To establish a method for the determination of bilastine in human plasma, and evaluate the uncertainty by LC-MS/MS. Methods: The uncertainty sources were obtained from the whole process of the determination including repeatability, e-quipment error, weighting, solution preparation, calibration fitting and plasma sample handling. The uncertainty and synthesized un-certainty of each component were calculated, and then the expanded uncertainty was obtained. Results: The expanded uncertainty for low (15 ng·ml-1), medium (400 ng·ml-1) and high(1 200 ng·ml-1) level of bilastine was 1. 45 ng·ml-1, 28. 72 ng·ml-1 and 74. 61 ng·ml-1, respectively (k=2, P=95% ). Conclusion: The uncertainty in the determination of bilastine in human plasma is mainly caused by equipment error, solution preparation, protein precipitation and calibration fitting (especially at low level).