Induction of Autophagy by the Beclin 1 Gene and Its Effect on MDCK Cell Line Necrosis

Authors
F. Taji 1
H. Mohseni Kouchesfehani 1
F. Sheikholeslami 2
K. Baesi 3
A. Abdoli 3
F. Motavalli 3
A. Abdoli 3
1 Department of Animal Biology, Faculty of Biological Science, Kharazmi University, Tehran, Iran
2 WHO Collaborating Center for Reference and Research on Rabies, Pasteur Institute of Iran, Tehran, Iran
3 Department of Hepatitis and AIDS, Pasteur Institute of Iran, Tehran, Iran
Abstract
Objective: Autophagy (self-digestion) is a highly regulated process for the degradation of damaged proteins and intracellular components. Autophagy has multifunctional roles in the protection of cellular homeostasis. Beclin1 is a key regulator molecule in autophagosome formation. Inhibition of autophagy by destruction of the Beclin 1 allele creates sensitivity to metabolic stress. Inhibition of the autophagy under conditions of nutrient deprivation in tumors resistant to apoptosis can lead to necrosis, inflammation and increased tumor growth. This study aims to assess the effect of autophagy induction on the necrosis pathway of MDCK cells.
Methods: We evaluated induction of autophagy by the Beclin1 gene in MDCK cells and assessed the percentage of necrosis cell death by flow cytometry using an Annexin V Staining kit. In order to induce autophagy, the recombinant pcDNA3.1-Beclin 1 was transfected into the MDCK cell line using lipofectamine TM 3000.
Results: Overexpression of the Beclin1 gene in MDCK cells led to induction of autophagy as seen by intracellular autophagosomal indicator LC3-II staining. There were 9.92% positive LC3 structures in transfected cells and 0.15% in untransfected cells. In the transfected and control groups, the rate of necrosis cell death was 1.66% and 0.06%, respectively.
Conclusion: Crosstalk between autophagy and necrosis pathways might affect the fate of the cell life span. Strategies that involve in modulation of autophagy and cell death might lead to therapeutic interventions in diseases. Therefore manipulation of cell death pathways could create new areas in therapeutic uses and interventions.

Keywords

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Pathobiology Reserach
Volume 20, Issue 1
June 2017
Pages 1-15