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Abstract Objectives: This study aims to explore the effect of silencing Beclin-1 gene on autophagy and apoptosis of Benign Prostatic Hyperplasia (BPH) (BPH-1) cells under the condition of Androgen Deprivation (AD) and Autophagy Inhibition (AI). Methods: Control group (BPH-1 group), empty carrier group (sh-RNA-BPH-1 group) and Beclin-1 silenced group (sh-Beclin1-BPH-1 group) were set. The Beclin-1 gene silencing efficiency was detected by RT-PCR and Western blot. Autophagic flux was monitored by GFP-LC3 cleavage assay and cell apoptosis was analyzed by flow cytometry. The protein expression levels of LC3, Caspase-3, PARP-1, Bcl-2, and Bax were detected by Western blot. Results: The transfection of sh-Beclin-1 obviously down-regulated the expression of Beclin-1 at both mRNA and protein levels. Under the conditions of AD and AI, silencing of Beclin-1 restrained the autophagy of BPH-1 cells, as evidenced by a decreased number of autophagosomes and down-regulation of LC3-II protein (p < 0.001). The results of flow cytometry showed that the apoptotic rate of sh-Beclin-1 group was elevated significantly compared to the other two groups (p < 0.01). Western blot results showed that silencing of Beclin-1 promoted 89 kd fragmentation of PARP-1 (p < 0.001) and Caspase-3 activation (p < 0.01). Moreover, silencing of Beclin-1 resulted in declined Bcl-2 and augmented Bax protein expression in BPH-1 cells (p < 0.01), which ultimately led to a decreased Bcl-2/Bax ratio. Conclusions: The results indicated that the silencing of Beclin-1 gene hampered autophagy while activating apoptosis in BPH-1 cells. Thus, Beclin-1 may participate in an antagonistic relationship between autophagy and apoptosis in BPH.
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@#Objective To explore the relationship between Beclin 1 level and lymph node metastasis in patients with non-small cell lung cancer. Method A total of 204 surgical specimens of patients with non-small cell lung cancer from September 2011 to September 2016 were collected in our hospital. There were 116 males and 88 females . Beclin 1 levels were detected by Western blotting. There were 116 males and 88 females at average age of 55.3±11.2 years. The patients were divided into three groups including a group N0 (no lymph node metastasis), a group N1(intralobar and interlobar lymph node metastases, and no mediastinal lymph node metastasis), and a group N2 (mediastinal lymph node metastasis). The differences of Beclin 1 levels in tumor tissues and lymph nodes of patients with N0, N1 and N2 were statistically analyzed. Results Among 204 patients of lung cancer, 36 patients were squamous cell carcinoma and 168 patients were adenocarcinoma. The levels of Beclin 1 in tumor tissues of N0, N1 and N2 groups decreased gradually with a statistical difference (P<0.05). In the three groups, the levels of Beclin 1 in the lung hilum and intrapulmonary lymph nodes (N1 Beclin 1) of N1 and N2 groups were less than that of N0 group with a statistical difference (P<0.01). In the three groups, the level of Beclin 1 in the mediastinal lymph nodes (N2 Beclin 1) of N2 group was less than that of the N0 and N1 groups with a statistical difference (P<0.01). In the N1 group, the level of N1 Beclin 1 was less than that of N2 group (P<0.01). In the N2 group, though the level of N1 Beclin 1 was less than N2 Beclin 1, there was no statistical difference (P>0.05). Conclusion Beclin 1 level can be used as a reference index to judge the benign and malignant lung masses, and lymph node Beclin 1 level can be used as an important reference index to help determine whether there is lymph node metastasis in lung cancer.
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Objective To analyze the expressions of astrocyte elevated gene-1 (AEG-1 )and Beclin-1 (BECN-1 )in papillary thyroid carcinoma (PTC)and their clinical significances.Methods Thirty patients with PTC who were performed total thyroidectomy in the General Hospital Affiliated to Tianjin Medical University from October 201 5 to May 201 6 were collected.All patients were diagnosed with unilateral PTC.The tumor tissues were taken from the patients′ipsilateral thyroids,and 30 cases of normal tissues were taken from the patients′contralateral normal thyroids.The expressions of AEG-1 and BECN-1 in PTC tissues were detected by reverse transcription-poly-merase chain reaction (RT-PCR).The relationships between the expression status and clinicopathologic features were analyzed.Results The positive expression levels [M(QR )]of AEG-1 in PTC and the normal tissues were 0.626 1 (0.741 4)and 0.049 8 (0.01 1 8)respectively,with a significant difference (Z=-4.488,P=0.000). The positive expression levels of BECN-1 in PTC and the normal tissues were 0.067 9 (0.1 98 1 )and 0.785 7 (0.361 7)respectively,with a significant difference (Z=-5.441 ,P=0.000).In PTC,the expressions of AEG-1 and BECN-1 were related to TNMstaging (Z=-3.980,P=0.000;Z=-2.265,P=0.023)and lymphatic metas-tasis (Z=-3.1 90,P=0.001;Z=-2.640,P=0.008),but they were not related to age (Z=-1 .203,P=0.229;Z=-1 .1 62,P=0.245),gender (Z=-1 .222,P=0.222;Z=-0.453,P=0.651 )and tumor size (Z=-1 .496,P=0.1 35;Z=-1 .1 66,P=0.244).The expression of AEG-1 was negatively correlated with that of BECN-1 (r=-0.343,P=0.007).Conclusion The expression of AEG-1 in PTC is higher than that in normal tissues and the expression of BECN-1 in PTC is lower than that in normal tissues,and the expressions of AEG-1 and BECN-1 are related to TNM staging and lymphatic metastasis,which are expected to become prognostic indicators.
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Objective: To investigate the effect of autophagy-related gene Beclin 1 on growth of xenografts of human lung adenocarcinoma A549 cells in BALB/c nude mice. Methods: The recombined plasmids pRNAT-U6.2/Lenti-si423 and pLenex-Beclin 1 were transiently transfected by lipofectin regeant into A549 cells, respectively. The expression levels of Beclin 1 mRNA and protein in A549 cells were detected by real-time fluorogenic quantitative-PCR (RFQ-PCR) and Western blotting, respectively. The proliferation rate of A549 cells transfected with recombinant vector pRNAT-U6.2/Lenti-si423 or pLenex-Beclin 1 was determined by MTT assay. The A549 cells expressing Beclin 1 or with Beclin 1 silencing were subcutaneously injected into right axillary region of nude mice. The growth rate and size of xenograft tumor were observed, and the expression levels of Beclin 1 mRNA and protein in xenograft tumor were determined by RFQ-PCR and immunohistochemistry, respectively. Results: The expression levels of Beclin 1 mRNA and protein were increased in A549 cells transfected with recombinant vector pLenex-Beclin 1, and the cell proliferation in vitro was inhibited. The expression levels of Beclin 1 mRNA and protein were decreased in A549 cells transfected with recombinant vector pRNAT-U6.2/Lenti-si423. As compared with Beclin 1-silencing group, the growth rate of subcutaneous xenograft was slowing down with lower weight and smaller volume and the relative expression levels of Beclin 1 mRNA and protein were both higher in Beclin 1-overexpression group. Conclusion: Autophagy-related gene Beclin 1 can inhibit the growth of xenograft of human lung adenocarcinoma A549 cells in nude mice, and it may become a new target for tumor therapy. Copyright© 2011 by TUMOR.