Background: The Hepatitis E virus (HEV) infection is a serious public health problem, with a mortality rate of approximately 20-25% among pregnant women. DNA vaccination was reported to induce humoral and cellular immune responses in murine models; however, a major problem of the strategy is its limited potency. Methods: In this study, we have investigated whether BECLIN1, as an autophagy-inducing based plasmid, can serve as an immunostimulatory agent and promote the HEV specific protective immunity of naked DNA vaccine encoding truncated ORF-2 (HEV239). Plasmids encoding HEV239 with or without BECLIN1 were used for the BALB/c mice immunization. Results: The results demonstrated a significant increase microtubule-associated protein light chain-3 II (LC3 II) as a marker of autophagy in the HEK293 cell, which were transfected by pVITRO-BECLIN1 plasmid. The immunological effects of adding BECLIN1 to HEV239 ORF-2 DNA vaccines were associated with the induction of the antigen-specific lymphocyte proliferation and the Th1-type cytokine (gamma-interferon (IFN-)), while there were no differences in IL-4 levels between the groups. Examination of humoral immune responses in vaccinated mice represented that immunization with pVITRO-HEV239-BECLIN1 notably increased serum level of IgG2a and total IgG against HEV in comparison to the HEV239 plasmid alone. Conclusions: The strong Th1 immune response induced by the BECLIN1 suggest that induction of autophagy can be an efficient approach to enhance the immunogenicity of DNA vaccine. This promising procedure could be further exploited as a potential therapeutic vaccine candidate in future studies.

Background: Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease, which primarily affects the joints. During RA development, T cells and other immune cells are recruited to the synovial tissue and promote RA. Autophagy is a process in which intracellular organelles and compounds are degraded. Autophagy as a regulator of cell homeostasis can affect immune cells activation and contribute in RA pathogenesis. The aim of this study was to evaluate the autophagy-related genes (Atgs) expression in two groups of RA patients and healthy subjects. Materials and Methods: Peripheral blood was obtained from three groups of donors including 20 patients with early RA, 20 undertreatment RA patients (with methotrexate, hydroxychloroquine and prednisolone therapy) and 20 age-and sex-matched healthy subjects. The expression of two autophagy-related genes was investigated by the real-time PCR technique. Results: The BECLIN-1 expression showed a 3. 41-fold increase in the patients with early RA compared to healthy subjects but in the under treatment patients it was 1. 5 times higher than healthy persons (P<0. 05). The Atg5 gene expression in the early RA patients increased by 2. 4 times more than healthy subjects (P<0. 05). Conclusion: The findings of this study show that in early RA patients, the increased expression of Atgs can promote RA pathogenesis. Also, findings suggest that the decreased autophagy can reduce RA severity.

Background and Aim: Acute lymphoblastic leukemia is the most common cancer in children and juveniles and is also seen in adults with lower frequency. Autophagy is a programmed catabolic process of the cell for destruction of damaged organs and proteins which is carried out by lysozymes. Disruption of autophagy leads to abnormalities in cellular processes associated with cancer. Materials and Methods: In this study, we compared the expression of BECLIN 1 and Atg 10 genes between 50 patients with B-ALL and 18 healthy subjects as our control group, by using RNA extraction and cDNA synthesis, and RT-PCR method. Data were analyzed by statistical tests. Results: The majority of B-ALL patients showed a significant reduction in the BECLIN1 and Atg 10 genes compared to control group (P <0. 05) and the mean expression of the genes (2-Δ Ct ± SD) were 0. 10 ± 0. 49 and 1. 01 ± 0. 27 for ALL patients and control group and 0. 15± 0. 44 and 1. 07± 0. 85 for Atg10 and BECLIN1 respectively. There was no significant correlation between expression levels of BECLIN1and Atg10 in these patients (r =-0. 013, P =0. 926). Conclusion: Considering the reduced expression levels in the essential genes of autophagy in ALL patients in our study and also other studies in this field, disruption of autophagy may be involved in leukomogensis.

Background: The pathogenesis of Gestational Trophoblastic Disease (GTD) is not clearly known.Objective: In this study, immunoexpression of proteins Bcl-2 and BECLIN-1 in trophoblastic lesions and normal trophoblastic tissue was conducted to study the mechanism of apoptotic and autophagic cell death that is expected to complete the study of GTD pathogenesis. Materials and Methods: Bcl-2 and BECLIN-1 immunoexpression were studied on complete hydatidiform mole, partial hydatidiform mole, invasive mole, choriocarcinoma and normal placenta slides. Results: The average total scores of Bcl-2 immunoexpression had a decreasing value, starting from partial hydatidiform mole (3.09), complete hydatidiform mole (2.36), invasive mole (1.18) to choriocarcinoma (0) when compared to normal placenta (6). The results showed no significant difference in BECLIN-1 immunoexpression total score between complete hydatidiform mole, partial hydatidiform mole and invasive mole, namely that the value of the average total score of BECLIN-1 was low (2.27, 2.45 and 2.36), but on the contrary choriocarcinoma showed an increasing strong BECLIN-1 expression with the average total score of 4.57. Conclusion: Bcl-2 expression decreases in line with the excessive proliferation of trophoblast cells in hydatidiform mole and leads to malignancy in invasive mole and choriocarcinoma. The decreased expression of BECLIN-1 that leads to autophagy defects in complete hydatidiform mole, partial hydatidiform mole and invasive mole shows the role of autophagy as tumor suppressor, whereas strong BECLIN-1 expression shows the survival role of autophagy in choriocarcinoma. The change of Bcl-2 activity as antiapoptosis and BECLIN-1 as proautophagy plays a role in pathogenesis of GTD.

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 thecell 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.

Background: There is a growing interest in development of an effective adjuvant system for improving DNA vaccines. Recent findings have confirmed an important role for autophagy in both innate and adaptive immunity. The current study was undertaken to determine the efficacy of autophagy induction with BECLIN-1, as a novel adjuvant system, in mice immunized with human papilloma virus (HPV) DNA vaccine. Methods: To determine whether autophagy induction with BECLIN-1 enhances the efficacy of HPV DNA vaccine, female C57BL/6 mice were challenged with TC-1 tumor cells and were immunized three times at one-week intervals. Two weeks after the final immunization, the mice were sacrificed, and the antitumor effects were assessed by measurement of lymphocyte proliferation, cytotoxicity, cytokine production, and tumor regression. Results: BECLIN-1 in combination with HPV-16 DNA vaccine encoding the E7 antigen induced a higher level of lymphocyte proliferation and cytotoxicity than the DNA vaccine alone. The novel combination increased the production of IFN-γ and highly inhibited tumor progression in comparison with DNA vaccine alone. Conclusion: Administration of BECLIN-1, as an autophagy inducer, with HPV DNA vaccine produces antitumor effects, providing an effective adjuvant for the induction of a strong antitumor immune response.

Purpose: During cancer growth, hypoxia occurs along with autophagy as an adaptive response to overcome cellular stress. Geraniol (GE) is a natural isoprenoid known for its wide anticancer activity and autophagy induction in the cancer cell. To investigate the antihypoxic potential of GE with the incidence of autophagy and apoptotic cell death in A549 CoCl2 treated cells. Methods: A549 cells were incubated for 24 hours with GE and CoCl2 either alone or in combination. We examined the cytotoxicity and cell viability of GE either alone or in combination therapy using MTT and trypan blue assay. GE modulating effect was determined on lipid peroxidation, antioxidant capacity markers, gene expression levels of hypoxia inducible factor-1 (HIF-1), NF-κ, B, vascular endothelial growth factor (VEGF), autophagy factors in different groups, besides apoptotic bodies using acridine orange/ethidium bromide (AO/EB). Results: GE and CoCl2 combination therapy downregulated the expression of HIF-1α,that suppressed A549 cell growth through downregulation of BNIP3 and BECLIN-1 gene expression. This resulted in autophagy and apoptotic cell death, in addition to the downregulation of NF-kB and VEGF expression. Also, GE treatment significantly reduced the oxidative stress markers and restored the antioxidant capacity. Conclusion: GE possesses an antihypoxic effect on A549 CoCl2 treated cells and induces cell death via autophagy along with apoptosis through HIF-1a/BNIP3/BECLIN-1 signaling pathway.

Background and Objective: Despite the protective effect of ischemic preconditioning (IPC) from tissues, its effect on diabetes-induced excessive autophagy is not clear. Autophagy could be inhibited or activated through exercise. The present study aimed to investigate the effect of endurance training and remote IPC on myocardial BECLIN-1 gene expression in diabetic rats.Materials and Method: In this experimental research, 35 Wistar rats were randomly divided into seven groups: normal control (C), diabetic (D), diabetic one leg ischemia (OI), diabetic two legs ischemia (TI), diabetic endurance training (E), diabetes one leg ischemia + endurance training (OIE), diabetes two legs ischemia+ endurance training (TIE). IPC was included three 5-minute cycles of ischemia, followed by five minutes of reperfusion. The endurance training groups performed exercise training for six weeks and five days a week and was included running on a treadmill. The speed for the first week is set at 9 meters per minute for 15 minutes. By the sixth week, the training speed was increased to 18 meters per minute for 30 minutes. BECLIN-1 gene expression was measured by RT-PCR. Data were analyzed by one-way analysis of variance and Tukey post hoc tests.Results: BECLIN-1 gene expression significantly increased in D group compared to C group (p=0.0001). Becli-1 gene expression significantly decreased in OI (p=0.0001), TI (p=0.0001), E (p=0.0001), OIE (p=0.0001) and TIE (p=0.0001) compared to the D.Conclusion: It seems that the effect of ischemia preconditioning in decreasing BECLIN-1 is independent of the ischemia-affected muscle mass. Endurance training with IPC is more effective in decreasing BECLIN-1 in diabetes than any of the exercise and IPC interventions alone.