24 July 2019 : Laboratory Research
LaSota Strain Expressing The Rabies Virus Glycoprotein (rL-RVG) Suppresses Gastric Cancer by Inhibiting the Alpha 7 Nicotinic Acetylcholine Receptor (α7 nAChR)/Phosphoinositide 3-Kinase (PI3K)/AKT PathwayXuefeng Bu1A, Chaoyun Yin2B, Xuanfeng Zhang12C, Anwei Zhang12D, Xiaomei Shao2E, Yao Zhang23F, Yulan Yan3G*
Med Sci Monit 2019; 25:5482-5492
BACKGROUND: The recombinant avirulent Newcastle disease virus (NDV) LaSota strain expressing the rabies virus glycoprotein (rL-RVG) can induce much greater apoptosis than can NDV in gastric carcinoma cells, but the mechanisms involved remains unclear.
MATERIAL AND METHODS: The 2 gastric carcinoma cell lines were divided into the rL-RVG group, the NDV group, and the PBS group. MTT assay was used to detect and analyze cell viability. siRNA for α7-nAChR, α7-nAChR antagonist, or α7-nAChR agonist, AKT antagonist, and p-AKT agonist were used for pretreatment. The protein expressions of RVG, NDV, α7-nAChR, cleaved caspase-3, p-AKT, PI3K, Bcl-2, and Bax proteins were detected by Western blot assay. Immunofluorescence was used to detect expressions of α7-nAChR proteins. Light microscopy, flow cytometry, and TUNEL assay were used to assess apoptosis.
RESULTS: The results showed that 2 virus concentrations over 10³ dilution caused greater cell proliferation inhibition. rL-RVG treatment increased the expression of α7-nAChR, cleaved caspase-3, and Bax protein but decreased the expression of p-AKT, PI3K, and Bcl-2 protein. When the groups were pretreated with α7-nAChR antagonist, the α7-nAChR, cleaved caspase-3, and Bax protein expression increased, but the expression of p-AKT, PI3K, and Bcl-2 protein was clearly decreased. However, the results in the α7-nAChR agonist group were the opposite. When treated with the AKT antagonist, the result was the same as in the rL-RVG treatment group. The result in the AKT agonist group was the opposite of that in the AKT antagonist group. Compared with the NDV group, the results of light microscopy, FCM, and TUNEL assay showed that α7-nAChR antagonist significantly affected the apoptosis of gastric cancer cells in the rL-RVG group.
CONCLUSIONS: rL-RVG leads to much greater apoptosis through the α7-nAChR/PI3K/AKT pathway.
Keywords: alpha7 Nicotinic Acetylcholine Receptor, Apoptosis, Newcastle disease virus, Oncogene Protein v-akt, Rabies virus, Stomach Neoplasms, Caspase 3, Cell Proliferation, Cell Survival, Glycoproteins, Peptide Fragments, phosphatidylinositol 3-kinase, Phosphatidylinositol 3-Kinases, Proto-Oncogene Proteins c-akt, Proto-Oncogene Proteins c-bcl-2, Signal Transduction, Viral Proteins, bcl-2-Associated X Protein
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