|Table of Contents|

Establishment of an in vitro model of bacteria adhesion to respiratory epithelial cells

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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

Issue:
2018年第11期
Page:
973-978
Research Field:
基础医学
Publishing date:

Info

Title:

Establishment of an in vitro model of bacteria adhesion to respiratory epithelial cells

Author(s):

WANG Ruoxi WANG Qian SUN Fengjun LIU Yao ZHANG Lei LI Xiaoyu XIA Peiyuan

Department of Pharmacy, First Affiliated Hospital, Army Medical University (Third Military Medical University), Chongqing, 400038, China

Keywords:

flow chamber bacterium epithelial cells adhesion

PACS:
R322.3; R329-33; R378
DOI:
-
Abstract:

Objective    To establish an in vitro model to study the adhesion of bacteria to respiratory epithelial cells based on the construction of flow-chamber. Methods    After 1×105 HBE cells were seeded in the flow chamber precoated with 2 mg/mL bovine collagen, the chamber was cultured in RPMI 1640 medium containing 20% serum and incubated at 37 ℃ in a 5% CO2 incubator for 24 h. Once the bottom was covered with HBE cells, the chamber could be used for further experiments. When the number of cells in the flow chamber was taken as the evaluation index, the effects of the flow rate, the flow duration and the mobile phase composition on the cellular model were investigated by orthogonal design, and the optimal experimental conditions were screened. Then under the obtain optimal conditions, after the bacteria were adhered to the cells at 2×108, 108, 5×107, and 2.5×107 CFU/mL inoculated concentrations respectively, the amount of bacterial adhesion was taken as an index, and the results were compared with the conventional plate culture for bacterial adhering to cells, so as to determine whether the flow chamber is a viable model of bacteria adhesion to cells. The amount of bacteria adhering to cells was characterized by SYTO9 fluorescently labeled bacteria. Results    The influencing factors in optimal experimental conditions in order of their impacts were as follows: flow rate>flow duration>mobile phase composition. The ANOVA study showed that the impacts of flow rate and flow duration were significant (P<0.05), and while those of mobile phase composition were not. After fluorescent staining, the model realized the real-time fluorescence observation of the bacteria adhesion to the respiratory epithelial cells. According to the quantitative results of bacterial adhesion, with the increase of multiplicity of infection, both the flow chamber and the conventional culture plate detected the elevated amount of adhering bacteria, in a linear manner. However, with the same multiplicity of infection, the conventional culture plate model detected significantly larger amount of adhering bacteria when compared to the flow chamber (P<0.05). Conclusion   Under appropriate flow rate and flow duration, our model is an alternative to the conventional plate for studying the adhesion of bacteria to the airway epithelial cells, with the advantages of being more close to the in vivo environment, high accuracy and real-time observation

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Last Update: 2018-06-14