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Background Salmonella infection in livestock and poultry causes salmonellosis, and is mainly treated using antibiotics. However, the misuse use of antibiotics often triggers the emergence of multi-drug-resistant Salmonella strains. Currently, Salmonella phages is safe and effective against Salmonella, serving as the best drug of choice. This study involved 16 Salmonella bacteriophages separated and purified from the sewage and the feces of the broiler farm. A phage, vB_SalP_LDW16, was selected based on the phage host range test. The phage vB_SalP_LDW16 was characterized by the double-layer plate method and transmission electron microscopy. Furthermore, the clinical therapeutic effect of phage vB_SalP_LDW16 was verified by using the pathogenic Salmonella Enteritidis in the SPF chicken model. Results The phage vB_SalP_LDW16 with a wide host range was identified to the family Siphoviridae and the order Caudoviridae, possess a double-stranded DNA and can lyse 88% (22/25) of Salmonella strains stored in the laboratory. Analysis of the biological characteristics, in addition, revealed the optimal multiplicity of infection (MOI) of vB_SalP_LDW16 to be 0.01 and the phage titer to be up to 3 × 1014 PFU/mL. Meanwhile, the phage vB_SalP_LDW16 was found to have some temperature tolerance, while the titer decreases rapidly above 60 ℃, and a wide pH (i.e., 5–12) range as well as relative stability in pH tolerance. The latent period of phage was 10 min, the burst period was 60 min, and the burst size was 110 PFU/cell. Furthermore, gastric juice was also found to highly influence the activity of the phage. The clinical treatment experiments showed that phage vB_SalP_LDW16 was able to significantly reduce the bacterial load in the blood through phage treatment, thereby improving the pathological changes in the intestinal, liver, and heart damage, and promoting the growth and development of the chicken. Conclusions The phage vB_SalP_LDW16 is a highly lytic phage with a wide host range, which can be potentially used for preventing and treating chicken salmonellosis, as an alternative or complementary antibiotic treatment in livestock farming.

Esophageal cancer is the ninth most common malignancy worldwide, ranking sixth in mortality. Platinum-based chemotherapy is commonly used for treating locally advanced esophageal cancer, yet it is ineffective in a large portion of patients. There is a need for reliable molecular markers with direct clinical application for a prospective selection of patients who can benefit from chemotherapy and patients in whom toxicity is likely to outweigh the benefit. The cytotoxic activity of platinum derivatives largely depends on the uptake and accumulation into cells, primarily by organic cation transporters (OCTs). The aim of the study was to investigate the impact of OCT expression on the clinical outcome of patients with esophageal cancer treated with oxaliplatin. Twenty patients with esophageal squamous cell carcinoma (SCC) were prospectively enrolled and surgical specimens used for screening OCT expression level by western blotting and/or immunostaining, and for culture of cancer cells. Sixty-seven patients with SCC who received oxaliplatin and for whom follow-up was available were retrospectively assessed for organic cation/carnitine transporter 2 (OCTN2) expression by real time RT-PCR and immunostaining. OCTN2 staining was also performed in 22 esophageal adenocarcinomas. OCTN2 function in patient-derived cancer cells was evaluated by assessing L-carnitine uptake and sensitivity to oxaliplatin. The impact of OCTN2 on oxaliplatin activity was also assessed in HEK293 cells overexpressing OCTN2. OCTN2 expression was higher in tumor than in normal tissues. In patient-derived cancer cells and HEK293 cells, the expression of OCTN2 sensitized to oxaliplatin. Patients treated with oxaliplatin who had high OCTN2 level in the tumor tissue had a reduced risk of recurrence and a longer survival time than those with low expression of OCTN2 in tumor tissue. In conclusion, OCTN2 is expressed in esophageal cancer and it is likely to contribute to the accumulation and cytotoxic activity of oxaliplatin in patients with esophageal carcinoma treated with oxaliplatin.

Background: Barrett’s esophagus (BE) is the premalignant condition for the development of esophageal adenocarcinoma, and occurs when the stratified epithelium is replaced by an intestinal-type epithelium with goblet cells. Bile acids (BAs) may play a dominant role in esophageal metaplasia, which is characterized by the expression of intestine-specific nuclear transcription factor caudal type homeobox transcription factor 2 (CDX-2). Notch-1 is frequently downregulated in BE, and protects squamous epithelia from becoming malignant and inducing intestinal metaplasia. However, the molecular mechanisms of this pathway are unclear. Methods: We first investigated Notch-1 and CDX-2 in human BE epithelium. Then Wister rats were used to develope animal models to test the role of BAs in the pathogenesis of BE. Effects of deoxycholic acid (DCA) on Notch-1 and CDX-2, as well as Notch signaling pathway blockage with DATP or with si-Hes-1 in vitro were observed through RT–PCR and western blotting techniques in human EAC cells (OE-19) and esophageal epithelial cells (Het-1a). Results: We found that the Notch -1 gene was inhibited in human and rodent BE specimens and BAs induced Barrett’s-like metaplasia. DCA decreases Notch-1 in a time and concentration dependent manner in both EAC cells (OE-19) and esophageal epithelial cells (Het-1a). Notch signaling inhibition increased CDX-2 expression but also blocked the influence of DCA. Conclusions: These data imply that BAs induce BE by inhibiting Notch-1 in esophageal cells. Notch signal pathway inhibition presents a therapeutic strategy for premalignant conditions of the esophagus.

Dongfeng Sun,1 Chengyu Chen,1 Wensi Hu,1 Chenxi Zhong,2 Limin Fan,2 Xiaoming Song,1 Zhibo Gai3 1Department of Thoracic Surgery, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan 250014, People’s Republic of China; 2Department of Thoracic Surgery, Shanghai Chest Hospital, Shanghai JiaoTong University, Shanghai 200030, People’s Republic of China; 3Joint Pharmacology Center, University Hospital Zurich and Liaocheng People’s Hospital, Liaocheng 252000, People’s Republic of China Objective: To investigate the expression of tumor suppressor protein ASK1-interacting protein-1 (AIP1) in human esophageal squamous cell carcinoma (ESCC) and its role in tumor progression, angiogenesis, and prognosis.Methods: A total of 117 biopsy samples were obtained from ESCC patients. None of the patients had distant metastasis before surgery, and did not receive preoperative chemotherapy or radiotherapy. Immunohistochemistry was used to detect the expression of AIP1 protein and vascular endothelial growth factor receptor 2 (VEGFR2) in ESCC specimens collected from 117 patients who underwent esophageal cancer radical surgery. Microvessel density (MVD) was evaluated by immunohistochemical staining of vascular endothelial CD34. The correlation between AIP1 protein and clinicopathological characteristics, tumor angiogenesis, and prognosis was analyzed.Results: The downregulation of AIP1 protein in esophageal carcinoma tissues was detected in 63 cases. This downregulation significantly correlated with lymph node metastasis, clinicopathological staging, and tumor MVD (P

Reactive oxygen species (ROS) are at the center of many physiological and pathological processes. ROS generated due to oxidative stress can potentiate both cancer initiation and progression. Rotenone, which is an inhibitor of the mitochondrial electron transport chain complex I, results in the activation of NOX2 and release of ROS, and has been shown to display anticancer activity through the induction of apoptosis in various cancer cells. The mechanistic link between rotenone-dependent activation of NOX2 and induction of apoptosis is still elusive. In this study, we used the human lung cancer A549 cells to study the molecular mechanism(s) involved between rotenone-dependent activation of NOX2 and impairment of autophagic machinery. We report that acute exposure to rotenone induced mild NOX2-dependnet oxidative stress, which impaired the autophagic flux, resulting in cytosolic accumulation of LC3 and p62/STSQM1. We further show that this induction occurs through the PI3K/Akt/mTORC1 signaling pathway. We furthermore show that chronic exposure to rotenone lead to excessive NOX2-dependent ROS generation, increases autophagy, and decreases p62 level via increased-autophagic flux. Taken together, this study is the first mechanistic elucidation of how rotenone can be used to potently target cancer cells without overhauling the entire cellular machinery. © 2016 IUBMB Life 68(5):388-393, 2016.

Lysosome-associated protein transmembrane-4β (LAPTM4B) is a novel cancer-related gene that is upregulated in many tumors, and which plays important roles in carcinogenesis. It has two alleles, LAPTM4B 1 and LAPTM4B 2. LAPTM4B 1 contains only one copy of a 19-bp sequence in the first exon, whereas LAPTM4B 2 contains two tight tandem segments. Previous studies have shown that LAPTM4B 2 is a risk factor for susceptibility and prognosis of many tumors. The present study investigated the relationship between LAPTM4B polymorphism and non-small cell lung cancer (NSCLC) susceptibility and prognosis. We identified LAPTM4B genotypes with polymerase chain reaction (PCR) in peripheral blood samples. In the adjusted multivariate logistic regression analysis, we found that LAPTM4B 1/2, LAPTM4B 2/2 exhibited 1.48-fold [95% confidence interval (CI), 1.076-2.037] and 2.855-fold (95%CI, 1.722-4.734) increases in the risk of developing NSCLC compared with non-LAPTM4B 2 carriers. Furthermore, our results showed that overall survival time and disease-free survival time of patients with LAPTM4B 2 were significantly shorter than in patients carrying LAPTM4B 1 (P=0.001 and P=0.001, respectively). In addition, multivariate Cox regression analysis revealed that LAPTM4B 2 was also an independent prognostic factor for NSCLC. These results suggest that LAPTM4B polymorphisms may be a prospective marker for evaluating the risk and prognosis of NSCLC.

Lung cancer is the leading cause of cancer-related mortality all over the world. In recent years, pulmonary adenocarcinoma has surpassed squamous cell carcinoma in frequency and is the predominant form of lung cancer in many countries. Epidemiological investigations have shown an inverse relationship between garlic (Allium sativum) consumption and death rate from many cancers. Diallyl trisulfide (DATS) is one of the garlic-derived compounds (also known as: organosulfer compounds, OSC). DATS can induce apoptosis and inhibit the growth of many cancer cell lines. Our study demonstrated that the apoptotic incidents induced by DATS were a mitochondria-dependent caspase cascade through a significant decrease of the anti-apoptotic Bcl-2 that resulted in up-regulation of the ratio of Bax/Bcl-2 and the activity of caspase-3, -8, and -9. Eventually, DATS induced the apoptosis and inhibited the proliferation in a concentration- and time-dependent manner. Furthermore, by establishing an animal model of female BALB/c nude mice with A549 xenografts, we found that oral gavage of DATS significantly retarded growth of A549 xenografts in nude mice without causing weight loss or any other side effects compared with the control group. All the evidence both in vitro and in vivo suggested that DATS could be an ideal anti-cancer drug.

The purposes of the present study were to detect the expression of metastasis-associated protein 1 (MTA1) in patients with esophageal squamous cell cancer (ESCC), and to evaluate the relevance of MTA1 protein expression to the tumor progression, angiogenesis, and prognosis. Both MTA1 protein and intratumoral microvessels were examined by immunohistochemical staining in 131 ESCC patients who successfully underwent subtotal esophagectomy and esophagogastric anastomosis at Qilu Hospital between Jan 2004 and Dec 2005. Intratumoral microvessel density (MVD) was recorded by counting CD-34 positive immunostained endothelial cells. All statistical analyses were performed with SPSS 13.0 statistical software. High expression of MTA1 protein was detected in 57 cases and significantly correlated with tumor invasion depth (P = 0.041), lymph node metastasis (P = 0.021), pathologic stage (P = 0.003), and MVD (P = 0.044). Survival analysis showed that patients with MTA1 protein high expression had significantly poor overall 5-year survival (P = 0.002), and the factor found on multivariate analysis to significantly affect overall survival was only pathologic stage (P = 0.040). Further stratified survival analysis split by pathologic stage demonstrated that MTA1 protein high expression significantly predicted unfavorable prognosis among patients with pathologic stage II disease (P = 0.006). High expression of the MTA1 protein is common in ESCC, and is closely associated with tumor progression, increased tumor angiogenesis, and poor survival. These findings indicate that MTA1 protein has clinical potentials as a useful indicator of progressive phenotype, a promising prognostic predictor to identify patients with poor prognosis, and a potential novel therapeutic target of antiangiogenesis for patients with ESCC.

Reactive oxygen species (ROS) are at the center of many physiological and pathological processes. ROS generated due to oxidative stress can potentiate both cancer initiation and progression. Rotenone, which is an inhibitor of the mitochondrial electron transport chain complex I, results in the activation of NOX2 and release of ROS, and has been shown to display anticancer activity through the induction of apoptosis in various cancer cells. The mechanistic link between rotenone-dependent activation of NOX2 and induction of apoptosis is still elusive. In this study, we used the human lung cancer A549 cells to study the molecular mechanism(s) involved between rotenone-dependent activation of NOX2 and impairment of autophagic machinery. We report that acute exposure to rotenone induced mild NOX2-dependnet oxidative stress, which impaired the autophagic flux, resulting in cytosolic accumulation of LC3 and p62/STSQM1. We further show that this induction occurs through the PI3K/Akt/mTORC1 signaling pathway. We furthermore show that chronic exposure to rotenone lead to excessive NOX2-dependent ROS generation, increases autophagy, and decreases p62 level via increased-autophagic flux. Taken together, this study is the first mechanistic elucidation of how rotenone can be used to potently target cancer cells without overhauling the entire cellular machinery. © 2016 IUBMB Life 68(5):388-393, 2016.

Based on previous findings regarding the angiogenic activities and prognostic roles of metastasis-associated protein 1 (MTA1) in early-stage non-small cell lung cancer, the clinicopathological and prognostic significance of MTA1 protein expression, and its correlation with angiogenesis in lung invasive adenocarcinoma, were further assessed in the present study, according to the 2011 International Association for the Study of Lung Cancer/American Thoracic Society/European Respiratory Society classification. High protein expression levels of MTA1 were commonly observed in patients with lung invasive adenocarcinoma, and were significantly correlated with tumor size (P=0.030), lymph node metastasis (P=0.021) and microvessel density (P=0.015). Survival analysis demonstrated that patients with high protein expression levels of MTA1 exhibited significantly shorter five-year disease-free and overall survival than those patients whose protein expression levels of MTA1 were low (24.5% vs. 48.7%, P=0.001, and 34.7% vs. 59.2%, P=0.005, respectively). In addition, Cox regression multivariate analysis demonstrated that high protein expression levels of MTA1 significantly correlated with unfavorable five-year disease-free survival (P=0.024). These findings indicate that MTA1 protein expression may possess clinical potential as an indicator of progressive phenotype. Therefore, MTA1 is a promising prognostic predictor to identify subgroups of patients with high risk of relapse, and a potentially novel therapeutic target for antiangiogenesis in patients with lung invasive adenocarcinoma.

Background: Golgi phosphoprotein-3 (GOLPH3) is implicated in cancer development and progression. The aim of this study was to evaluate the prognostic significance of GOLPH3 protein and its association with tumor angiogenesis in patients with early-stage NSCLC. Materials and Methods: Immunohistochemistry was performed to determine GOLPH3 protein expression and allow assessment of intratumoral microvessel density (MVD) by counting CD-34 positive immunostained endothelial cells. Correlations of expression with MVD, clinicopathologic features and clinical prognosis were analyzed. Results: A notably higher level of GOLPH3 expression was found in early-stage NSCC tissues at the protein level. However, we do not find any correlation between GOLPH3 expression and clinicopathologic features (p>0.05), although higher MVD was positively associated with GOLPH3 overexpression (p

Studies have shown that genetic activation of TFIIB-related factor 2 (BRF2) represents a unique mechanism of tumorigenesis through the increase in Pol III-mediated transcription. Several studies have shown that BRF2 is overexpressed in several types of cancer and suggest the oncogenic role of BRF2. This study aimed to examine the expression of TFIIB-related factor 2 (BRF2) in patients with esophageal squamous cell cancer (ESCC) and explore the relationship of BRF2 expression with clinicopathologic factors, tumor angiogenesis and prognosis. We found that increased BRF2 protein expression was prevalent in esophageal squamous cell cancer and was significantly associated with deeper tumor invasion (P = 0.039) and microvessel density (P = 0.007). Additionally, expression of BRF2 was found to be an independent prognostic factor in ESCC patients. Furthermore, a significant correlation between high BRF2 expression and shorter overall survival time was found in different subgroups of ESCC patients stratified by the clinical stage, T classification and lymph node metastasis. High expression of BRF2 protein is closely associated with tumor progression and angiogenesis and poor survival of ESCC. BRF2 is a promising biomarker to identify individuals with poor prognostic potential and concludes the possibility of its use as a prognostic marker in patients with ESCC.

Metastasis-associated protein 1 (MTA1) high expression has been detected in a wide variety of human aggressive tumors and plays important roles in the malignant biological behaviors such as invasion, metastasis, and angiogenesis. However, the specific roles and mechanisms of MTA1 protein in regulating the malignant behaviors of non-small-cell lung cancer (NSCLC) cells still remain unclear. To elucidate the detailed functions of MTA1 protein, we down-regulated the MTA1 protein expression in NSCLC cell line by RNA interference (RNAi) in vitro, and found that down-regulation of MTA1 protein significantly inhibited the migration and invasion potentials of 95D cells. Further research revealed that down-regulation of MTA1 protein significantly decreased the activity of matrix metalloproteinase-9, which could be the mechanism responsible for the inhibition of 95D cells migration and invasion. In addition, the tube formation assay demonstrated that the number of complete tubes induced by the conditioned medium of MTA1-siRNA 95D cells was significantly smaller than that of 95D cells. These findings demonstrate that MTA1 protein plays important roles in regulating the migration, invasion, and angiogenesis potentials of 95D cells, suggesting that MTA1 protein down-regulation by RNAi might be a novel therapeutic approach to inhibit the progression of NSCLC.

Lung cancer is the leading cause of cancer-related mortality all over the world. In recent years, pulmonary adenocarcinoma has surpassed squamous cell carcinoma in frequency and is the predominant form of lung cancer in many countries. Epidemiological investigations have shown an inverse relationship between garlic (Allium sativum) consumption and death rate from many cancers. Diallyl trisulfide (DATS) is one of the garlic-derived compounds (also known as: organosulfer compounds, OSC). DATS can induce apoptosis and inhibit the growth of many cancer cell lines. Our study demonstrated that the apoptotic incidents induced by DATS were a mitochondria-dependent caspase cascade through a significant decrease of the anti-apoptotic Bcl-2 that resulted in up-regulation of the ratio of Bax/Bcl-2 and the activity of caspase-3, -8, and -9. Eventually, DATS induced the apoptosis and inhibited the proliferation in a concentration- and time-dependent manner. Furthermore, by establishing an animal model of female BALB/c nude mice with A549 xenografts, we found that oral gavage of DATS significantly retarded growth of A549 xenografts in nude mice without causing weight loss or any other side effects compared with the control group. All the evidence both in vitro and in vivo suggested that DATS could be an ideal anti-cancer drug.

The aims of this work are to detect the expression levels of metastasis-associated protein 1 (MTA1) in patients with early-stage non-small cell lung cancer (NSCLC), and to investigate the relationship of MTA1 protein with clinicopathologic factors, tumor angiogenesis, and prognosis. One hundred and two patients with pathologic stage I NSCLC who successfully underwent curative surgical resection were enrolled in this study. Immunohistochemical staining for MTA1 and CD34 was performed using the streptavidin–peroxidase method, and intratumoral microvessel density (MVD) was recorded by counting CD34-positive immunostained endothelial cells. All statistical analyses were performed with SPSS statistical software to determine the effects of MTA1 protein on clinicopathologic factors, tumor angiogenesis, and prognosis. MTA1 protein overexpression was detected in 41 cases and was significantly associated with MVD (P = 0.008). MTA1 protein overexpression and high MVD were significantly associated with tumor relapse (P = 0.004 and 0.007) and poor 5-year disease-free survival (P = 0.001 and 0.004). Patients with MTA1 protein overexpression and high MVD had significantly poor overall survival (P = 0.005 and 0.043) and disease-specific survival (P = 0.006 and 0.031) at 5 years after operation. Multivariate analysis demonstrated that MTA1 protein overexpression was an independent prognosticator for unfavorable disease-free, overall, and disease-specific survival (P = 0.011, 0.024, and 0.046). MTA1 protein overexpression is common in early-stage NSCLC and is significantly associated with tumor angiogenesis and poor survival. These findings suggest that MTA1 may have clinical potential as a promising predictor to identify individuals with poor prognostic potential and as a possible novel target molecule of antiangiogenic therapy for patients with early-stage NSCLC.

Examination of the genomes of 10 white-pock variants of cowpox virus strain Brighton red (CPV-BR) revealed that 9 of them had lost 32 to 38 kilobase pairs (kbp) from their right-hand ends and that the deleted sequences had been replaced by inverted copies of regions from 21 to 50 kbp long from the left-hand end of the genome. These variants thus possess inverted terminal repeats (ITRs) from 21 to 50 kbp long; all are longer than the ITRs of CPV-BR (10 kbp). The 10th variant is a simple deletion mutant that has lost the sequences between 32 and 12 kbp from the right-hand end of the genome. The limits of the inner ends of the observed deletions (between 32 and 38 kbp from the right-hand end of the CPV-BR genome) appear to be defined by the location of the nearest essential gene on the one hand and the location of the gene that encodes "pock redness" on the other. The genomes of the deletion/duplication white-pock variants appear to have been generated either by single crossover recombinational events between two CPV-BR genomes aligned in opposite directions or by the nonreciprocal transfer of genetic information. The sites where such recombination/transfer occurred were sequenced in four variants. In all of them, the sequences adjacent to such sites show no sequence homology or any other unusual structural feature. The analogous sites at the internal ends of the two ITRs of CPV-BR also were sequenced and also show no unusual features. It is likely that the ITRs of CPV-BR and of its white-pock variants, and probably those of other orthopox-virus genomes, arise as a result of nonhomologous recombination or by random nonreciprocal transfer of genetic information.

Several recombinant cowpox viruses were constructed and used to identify a viral gene that controls the production of hemorrhage in lesions caused by the Brighton Red strain of cowpox virus (CPV-BR). This gene is located in the KpnD fragment of CPV-BR DNA, between 31 and 32 kilobases from the end of the genome. This position corresponds well with that predicted from analyses of the DNA structures of spontaneously generated deletion mutants. The gene responsible for hemorrhage encodes a 38-kDa protein that is one of the most abundant early gene products. The 11-base-pair sequence GAAAATATATT present 84 base pairs upstream of its coding region is also present upstream of three other early genes of vaccinia virus; therefore, this sequence may be involved in the regulation of transcription. There is extensive similarity between the predicted amino acid sequence of the 38-kDa protein and the amino acid sequences of several plasma proteins that are inhibitors of various serine proteases involved in blood coagulation pathways. This suggests that the viral protein may possess a similar biological activity, which may enable it to effect hemorrhage by inhibiting one or more of the serine proteases involved in the host's normal processes of blood coagulation and wound containment.