Establishment of an
in vitro model of bacteria adhesion to respiratory epithelial cells- Journal of Third Military Medical University Vol. 40, Issue 11, Pages: 973-978(2018)
- 作者机构：
- 作者简介：
- 基金信息：the General Program of National Natural Science Foundation of China(81373451)
DOI：10.16016/j.1000-5404.201802030
CLC：Received：05 February 2018，
Published：2018-06
- 稿件说明：
移动端阅览
Ruoxi WANG, Qian WANG, Fengjun SUN, et al. Establishment of an
in vitro model of bacteria adhesion to respiratory epithelial cells[J]. Journal of Third Military Medical University, 2018, 40(11): 973-978.Ruoxi WANG, Qian WANG, Fengjun SUN, et al. Establishment of an
in vitro model of bacteria adhesion to respiratory epithelial cells[J]. Journal of Third Military Medical University, 2018, 40(11): 973-978. DOI： 10.16016/j.1000-5404.201802030.
摘要
Abstract
Objective
2
To establish an
in vitro
model to study the adhesion of bacteria to respiratory epithelial cells based on the construction of flow-chamber.
Methods
2
After 1×10
5
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% CO
2
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 2×10
8
10
8
5×10
7
and 2.5×10
7
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
2
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
2
Under appropriate flow rate and flow duration
our model is an alternative to the conventional plate for studying the adhesion of bact
eria to the airway epithelial cells
with the advantages of being more close to the
in vivo
environment
high accuracy and real-time observation.
关键词
Keywords
references
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