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Method for establishing EBV virus infected artificial respiratory tract epithelium model

A technology of artificial respiratory tract and virus infection, which is applied in the field of establishment of epithelial model of artificial respiratory tract infected by EBV virus, and can solve the problem of low efficiency of epithelial cells

Active Publication Date: 2020-12-08
山东银丰生命科学研究院 +1
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  • Abstract
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Problems solved by technology

[0006] Aiming at the above-mentioned prior art, aiming at the low efficiency of the existing EBV virus infecting epithelial cells and the limiting factors that cannot be effectively replicated, the present invention provides a method for establishing an EBV virus-infected artificial respiratory epithelial model-derived from stem cells differentiated from the nasal cavity basal layer In vitro virus infection model constructed from pseudostratified respiratory epithelium

Method used

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  • Method for establishing EBV virus infected artificial respiratory tract epithelium model
  • Method for establishing EBV virus infected artificial respiratory tract epithelium model
  • Method for establishing EBV virus infected artificial respiratory tract epithelium model

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Embodiment 1E

[0025] The construction of embodiment 1 EBV virus infection artificial respiratory tract epithelial model

[0026] The steps are as follows (the technical route is as follows figure 1 shown):

[0027] (1) Rinse the nasopharyngeal mucosal tissues surgically resected in patients with nasopharyngeal cysts in PBS buffer (4°C) (put them in immediately after surgical resection), remove surface mucus, and peel off submucosal tissues; use at 4°C Dispase II (dispase II) (purchased from Sigma, used at a concentration of 10 mg / ml) was digested for 8 to 12 hours; the digestion was terminated with DMEM culture solution containing 10% FBS, and single cells were slowly mechanically broken into single cells with a sample gun, and the supernatant was removed by centrifugation. Cleared to obtain cell pellets;

[0028] (2) The cell pellet obtained after the above centrifugation was suspended with FG medium, added fetal calf serum (the amount added was 0.02% of the FG medium, volume percentage)...

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Abstract

The invention discloses a method for establishing an EBV (Epstein-Barr Virus) infected artificial respiratory tract epithelium model. The method comprises the following steps: (1) digesting nasopharyngeal mucosa tissues by using Dispase II, performing beating into single cells, and performing centrifuging to remove supernate, so as to obtain cell precipitate; (2) suspending and culturing the cellprecipitate by using an epithelial cell culture medium; (3) performing digesting into single cells, planting the single cells in a small chamber above the 24-hole support membrane, and performing culturing until the support membrane is overgrown; (4) completely removing the culture solution above the support membrane, and continuously culturing for 2-3 weeks; and (5) when large-area cilium swinging is observed, adding Akata cells to carry out cell contact mediated EBV infection, and when successful infection, obtaining the artificial respiratory tract epithelium model infected with the EBV. According to the establishing method, nasopharyngeal mucosa tissue is taken as a material part, nasopharyngeal epithelial cells can be differentiated into completely polarized pseudo-multilayer respiratory tract epithelial tissue through gas-liquid interface culture, and the nasopharyngeal epithelial tissue is completely simulated by basal layer cells, secretory cuppy cells and swinging cilium cells.

Description

technical field [0001] The invention relates to a method for establishing an artificial respiratory epithelial model infected by EBV virus. Background technique [0002] The full name of EBV virus is Epstein-Barr virus, which is a virus strain established by Epstein and Barr in 1964 for the first time using Burkitt African children's lymphoma cells successfully through in vitro suspension culture. EBV is universally infective in the population, and worldwide, serology shows that more than 90% of people have been infected by EBV. EBV is mainly transmitted and transmitted through saliva and blood. Infection mostly occurs in early childhood, usually without or only with weak clinical symptoms. EBV infection can induce a variety of malignant tumors, including Burkitt's lymphoma, Hodgkin's lymphoma, nasopharyngeal carcinoma, and gastric carcinoma. At present, there is no effective drug for treating EBV infection, and the carcinogenicity caused by EBV infection cannot be effect...

Claims

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Application Information

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IPC IPC(8): C12N5/10C12N5/071C12N15/869
CPCC12N5/0625C12N15/86C12N2509/00C12N2710/16243Y02A50/30
Inventor 于凤刚魏本杰
Owner 山东银丰生命科学研究院
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