| Objective:With live attenuated mumps vaccine (LAMV) as an oncolytic virus, we work on its inhibitory effect on tumor cell lines, analyze its anti-tumor mechanism, and explore new approaches to cancer treatment.Methods:In this paper, live attenuated mumps vaccine (Hangzhou Center for Disease Control), mouse gastric cancer cell line (MFC) and human Wilm’s tumor cell line (G401) were materials mainly used. Three groups were involved in this experiment, namely, live attenuated mumps vaccine virus stock, mumps virus stock*10-1group (ie,10%of the stock in viral titer) and the control group.1. Using Vero cells to amplify the virus, collecting live attenuated mumps vaccine. Determination viral titer using plaque forming units PFU method.2. Tumor cells MFC and G401were cultured in vitro to detect in inhibition effect of LAMV using CCK-8kit.3. Tumor cells MFC and G401were cultured in vitro to test the apoptosis rates by means of FITC Annexin V/PI kit and flow cytometers in3consecutive days after LAMV was inoculated.4. The whole genome DNA was extracted after3days’ infection of LAMV. DNA electrophoresis of MFC, G401tumor cells was done to check to DNA Ladders from apoptotic bodies.5. The whole proteins were extracted after3days’ infection of LAMV. Western Blot was performed to detect autophagy-related proteins P62, LC3(LC3-I, LC3-II) trends.6. Preliminary experiments carried out in vivo by injecting MFC cells to BALB/c mice forelimbs. Tumor-bearing animal model was established. Then injection of high-dose and low-dose mumps virus and PBS control volume, to explore mumps virus inhibition effects on solid tumors growing in mice.Results:1. With Vero cells as hosts of mumps virus, we successfully harvested a great deal of live attenuated mumps vaccine virus. We determined the mumps titer with PFU. Purification can further improve the virus titer.2. LAMV’s inhibition rate on MFC and G401reached58.31%and49.52%, very effectively compared to measles and varicella vaccine virus, indicating that mumps virus has a strong ability to induce cell death.3. MFC and G401tumor cells infected with LAMV over time, the morphological changes included the increase of intercellular gaps, elongation of cells, accumulation of brown particles in cells with nuclei disappearance. Cell membranes dissolved with fragments and ultimately death.4. Flow cytometers showed the apoptosis rates of three groups were3.98%,3.44%and2.16%(24h);21.2%,2.72%and2.39%(48h);51.0%,4.63%and2.12%(72h) after MFC infected with LAMV. In addition, DNA electrophoresis detected DNA Ladder on MFC with mumps virus. Thus, induction of MFC apoptosis leads to MFC cell death following treatment with LAMV.5. Western Blot showed decline in P62, and elevation in LC3and LC3-Ⅱ/LC3-Ⅰ ratio after G401infected with LAMV, indicating that induction of autophagy leads to G401cell death following treatment with LAMV.6. We successively constructed the tumor-bearing mice model of gastric cancer. Initial experiments found that LAMV can inhibit tumor growth.Conclusion:1. Live attenuated mumps vaccine can effectively inhibit the proliferation of tumor cells in vitro (MFC, G401), and promote tumor cell deformation, sparse until death.2. Live attenuated mumps vaccine induce tumor cell death with different mechanisms. Apoptosis is activated in MFC, while autophagy in G401, leading to programmed cell death.3. We constructed the tumor-bearing animal model with MFC. LAMV can inhibit tumor growth in vivo.Thus, live attenuated mumps vaccine as oncolytic virus, provides a promising therapy for tumor treatment. |
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