人呼吸道上皮细胞在 Matrigel 中的三维培养用于评价人类鼻病毒 C 和博卡病毒感染。
Three-dimensional Culture of Human Airway Epithelium in Matrigel for Evaluation of Human Rhinovirus C and Bocavirus Infections.
机构信息
Key Laboratory of Heavy Ion Radiation Biology and Medicine of Chinese Academy of Sciences & Key Laboratory of Space Radiobiology of Gansu Province, Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000, Gansu, China.
Department of Pathogenic Biology, Chengde Medical University, Chengde 067000, Hebei, China.
出版信息
Biomed Environ Sci. 2018 Feb;31(2):136-145. doi: 10.3967/bes2018.016.
OBJECTIVE
Newly identified human rhinovirus C (HRV-C) and human bocavirus (HBoV) cannot propagate in vitro in traditional cell culture models; thus obtaining knowledge about these viruses and developing related vaccines are difficult. Therefore, it is necessary to develop a novel platform for the propagation of these types of viruses.
METHODS
A platform for culturing human airway epithelia in a three-dimensional (3D) pattern using Matrigel as scaffold was developed. The features of 3D culture were identified by immunochemical staining and transmission electron microscopy. Nucleic acid levels of HRV-C and HBoV in 3D cells at designated time points were quantitated by real-time polymerase chain reaction (PCR). Levels of cytokines, whose secretion was induced by the viruses, were measured by ELISA.
RESULTS
Properties of bronchial-like tissues, such as the expression of biomarkers CK5, ZO-1, and PCK, and the development of cilium-like protuberances indicative of the human respiration tract, were observed in 3D-cultured human airway epithelial (HAE) cultures, but not in monolayer-cultured cells. Nucleic acid levels of HRV-C and HBoV and levels of virus-induced cytokines were also measured using the 3D culture system.
CONCLUSION
Our data provide a preliminary indication that the 3D culture model of primary epithelia using a Matrigel scaffold in vitro can be used to propagate HRV-C and HBoV.
目的
新发现的人类鼻病毒 C（HRV-C）和人类博卡病毒（HBoV）不能在传统的细胞培养模型中体外繁殖；因此，了解这些病毒并开发相关疫苗是困难的。因此，有必要开发一种新型平台来繁殖这些类型的病毒。
方法
开发了一种使用 Matrigel 作为支架在三维（3D）模式下培养人呼吸道上皮细胞的平台。通过免疫化学染色和透射电子显微镜鉴定 3D 培养的特征。通过实时聚合酶链反应（PCR）定量测定指定时间点 3D 细胞中 HRV-C 和 HBoV 的核酸水平。通过 ELISA 测量病毒诱导的细胞因子的分泌水平。
结果
在 3D 培养的人呼吸道上皮（HAE）培养物中观察到支气管样组织的特性，如生物标志物 CK5、ZO-1 和 PCK 的表达，以及指示人类呼吸道的纤毛样突起的发育，但在单层培养的细胞中没有。还使用 3D 培养系统测量了 HRV-C 和 HBoV 的核酸水平以及病毒诱导的细胞因子的水平。
结论
我们的数据初步表明，使用 Matrigel 支架体外培养的原代上皮 3D 培养模型可用于繁殖 HRV-C 和 HBoV。
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