Metformin promotes autophagy activity and constrains HSV-1 replication in neuroblastoma cells

Problem considered: Metformin is an antihyperglycemic drug with effective clinical functions in type 2 diabetes patients. Recent evidence indicates that metformin can change natural cellular signaling to induce autophagy. Furthermore, infection could rise autophagy activity, but herpes simplex virus 1 (HSV-1) constructs, particularly virulence factors, constrain autophagy promotion, we used HSV-1 because of its ability to escape from autophagy by inhibiting beclin-1 formation, reversing PKR signaling and also induce mTOR activity. This study aimed to compare the extent of autophagic activity in metformin-treated cells infected with HSV-1 and determine the effect of metformin on HSV-1 replication. Methods: Neuroblastoma cells were cultured and then divided into the two separate groups of infected with HSV-1 and expose with metformin. Autophagy was evaluated via microtubule-associated protein 1 light chain 3 (LC3-II) monitoring by affix staining antibodies to this protein using flow cytometry. To evaluate viral load, HSV-1 was inoculated for 4 h post metformin treatment, and to evaluate viral load in the post-infection pathway, cells were inoculated with HSV-1. Then, after a 2-hour viral attachment time, metformin was administered to the cells. Viral nucleic acid was purified 48 h post-infection, and virus titer was calculated by real-time polymerase chain reaction (PCR). Results: Our real-time PCR results indicated that HSV-1 viral load in pre-infection pathway reduced from 1.3 x 10(6) in positive control to 8 x 10(5) in cells exposed to metformin. Additionally, the TCID50/ml at 12 and 24 h after infection was 10 times lower than in HSV-1 positive control. However, after 48 h, HSV-1 titer in the metformin-treated group was equal with non-treated cells. For the post-infection pathway, real-time PCR results exhibited that HSV-1 titer was 2.3 x 10(6) in virus control cells compared to 1.8 x 10(6) in treated cells. That is, TCID50/ml in the medium of metformin-treated cells was 10 times lower in comparison with non-treated cells after 72 h. Conclusion: The study of the effect of metformin on neuroblastoma cells indicates high autophagic flux and lesser HSV-1 propagation in treated cells.