The gastrointestinal tract is a specialized organ in which dynamic interactions between host cells and the complex environment occur in addition to food digestion. Together with the chemical barrier of the mucosal layer and the cellular immune system, the epithelial cell layer performs a pivotal role as the first physical barrier against external factors and maintains a symbiotic relationship with commensal bacteria. The tight junction proteins, including occludin, claudins, and zonula occludens, are crucial for the maintenance of epithelial barrier integrity. To allow the transport of essential molecules and restrict harmful substances, the intracellular signaling transduction system and a number of extracellular stimuli such as cytokines, small GTPases, and post-translational modifications dynamically modulate the tight junction protein complexes. An imbalance in these regulations leads to compromised barrier integrity and is linked with pathological conditions. Despite the obscurity of the causal relationship, the loss of barrier integrity is considered to contribute to inflammatory bowel disease, obesity, and metabolic disorders. The elucidation of the role of diseases in barrier integrity and the underlying regulatory mechanisms have improved our understanding of the intestinal barrier to allow the development of novel and potent therapeutic approaches.

The gut microbiota has an important role in the gut barrier, inflammation and metabolic functions. Studies have identified a close association between the intestinal barrier and metabolic diseases, including obesity and type 2 diabetes (T2D). Recently, Akkermansia muciniphila has been reported as a beneficial bacterium that reduces gut barrier disruption and insulin resistance. Here we evaluated the role of A. muciniphila-derived extracellular vesicles (AmEVs) in the regulation of gut permeability. We found that there are more AmEVs in the fecal samples of healthy controls compared with those of patients with T2D. In addition, AmEV administration enhanced tight junction function, reduced body weight gain and improved glucose tolerance in high-fat diet (HFD)-induced diabetic mice. To test the direct effect of AmEVs on human epithelial cells, cultured Caco-2 cells were treated with these vesicles. AmEVs decreased the gut permeability of lipopolysaccharide-treated Caco-2 cells, whereas Escherichia coli-derived EVs had no significant effect. Interestingly, the expression of occludin was increased by AmEV treatment. Overall, these results imply that AmEVs may act as a functional moiety for controlling gut permeability and that the regulation of intestinal barrier integrity can improve metabolic functions in HFD-fed mice.

Ulcerative colitis is a multi-factorial disease involving a dysregulated immune response. Disruptions to the intestinal epithelial barrier and translocation of bacteria, resulting in inflammation, are common in colitis. The mechanisms underlying epithelial barrier dysfunction or regulation of tight junction proteins during disease progression of colitis have not been clearly elucidated. Increase in phospholipase D (PLD) activity is associated with disease severity in colitis animal models. However, the role of PLD2 in the maintenance of intestinal barrier integrity remains elusive. We have generated intestinalspecific Pld2 knockout mice (Pld2 IEC-KO) to investigate the mechanism of intestinal epithelial PLD2 in colitis. We show that the knockout of Pld2 confers protection against dextran sodium sulphate (DSS)-induced colitis in mice. Treatment with DSS induced the expression of PLD2 and downregulated occludin in colon epithelial cells. PLD2 was shown to mediate phosphorylation of occludin and induce its proteasomal degradation in a c-Src kinase-dependent pathway. Additionally, we have shown that treatment with an inhibitor of PLD2 can rescue mice from DSS-induced colitis. To our knowledge, this is the first report showing that PLD2 is pivotal in the regulation of the integrity of epithelial tight junctions and occludin turn over, thereby implicating it in the pathogenesis of colitis.Ulcerative colitis (UC) is a multi-factorial disease with genetic, immunological, environmental, and diet-related factors contributing to its etiology 1, 2 . Incidence of UC is increasing worldwide, and current research is focused on elucidating the origin and mechanism of disease initiation and perpetuation. A chronically dysregulated response, mediated by the host's immune cells against normal gut microbiota, results in severe inflammation and is the hallmark of UC 3 . Genome-wide association studies have revealed several genes associated with UC. These studies have highlighted the role of immune cell-and intestinal barrier-associated genes in the development of UC; however, convincing evidence, enabling the understanding of the complex aetiology of UC, is still lacking 4 .Maintenance of proper barrier integrity is an essential function of the epithelial layer. Disruptions in the intestinal epithelial monolayer lead to bacterial translocation across the membrane and subsequent inflammation 5 . Patients with UC show an increased intestinal permeability 6, 7 . Tight junction proteins are a major class of proteins responsible for junctional sealing and selective transport of molecules across the epithelial barrier 4,8 . Altered or compromised expression of epithelial tight junction proteins is a key factor in the pathogenesis of the disease 9, 10 . However, the mechanisms underlying epithelial barrier dysfunction or regulation of epithelial tight junction proteins during disease progression remain unexplored.

A ngiogenesis involves complex endothelial cell (EC) behaviors, such as proliferation, survival, migration, and tube formation. The most important stimulus promoting angiogenesis is tissue hypoxia. Hypoxia mediates several processes in ECs that are required for each step of angiogenesis.1 Hypoxia-induced angiogenesis is closely related to pathological situation. The dysregulation of these EC behaviors and thus abnormal angiogenesis are critically associated with hypoxia-induced pathological angiogenesis that occurs during the course of several diseases, such as cancer and vascular retinopathy.2,3 Therefore, identification of specific molecules involved in hypoxia-induced angiogenesis will facilitate studies to clarify the various molecular mechanisms involved in pathological angiogenesis and may aid the discovery of novel angiogenic drug targets.© 2014 American Heart Association, Inc. Objective-Aberrant regulation of the proliferation, survival, and migration of endothelial cells (ECs) is closely related to the abnormal angiogenesis that occurs in hypoxia-induced pathological situations, such as cancer and vascular retinopathy. Hypoxic conditions and the subsequent upregulation of hypoxia-inducible factor-1α and target genes are important for the angiogenic functions of ECs. Phospholipase D2 (PLD2) is a crucial signaling mediator that stimulates the production of the second messenger phosphatidic acid. PLD2 is involved in various cellular functions; however, its specific roles in ECs under hypoxia and in vivo angiogenesis remain unclear. In the present study, we investigated the potential roles of PLD2 in ECs under hypoxia and in hypoxia-induced pathological angiogenesis in vivo. Approach and Results-Pld2 knockout ECs exhibited decreased hypoxia-induced cellular responses in survival, migration, and thus vessel sprouting. Analysis of hypoxia-induced gene expression revealed that PLD2 deficiency disrupted the upregulation of hypoxia-inducible factor-1α target genes, including VEGF, PFKFB3, HMOX-1, and NTRK2. Consistent with this, PLD2 contributed to hypoxia-induced hypoxia-inducible factor-1α expression at the translational level. The roles of PLD2 in hypoxia-induced in vivo pathological angiogenesis were assessed using oxygen-induced retinopathy and tumor implantation models in endothelial-specific Pld2 knockout mice. Pld2 endothelial-specific knockout retinae showed decreased neovascular tuft formation, despite a larger avascular region. Tumor growth and tumor blood vessel formation were also reduced in Pld2 endothelial-specific knockout mice. Cellular responses to hypoxic stress are mediated by multiple mechanisms. Hypoxia-inducible factor-1 (HIF-1) is the most prominent factor that mediates cellular responses to hypoxia by inducing the expression of several target genes. Conclusions-Our 1Because HIF-1 activation is preferentially modulated by changes in the amount of the HIF-1α subunit, several factors and mechanisms have been suggested. Among these, the regulation of HIF-1α degradation through post-translat...

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DNA-dependent protein kinase (DNA-PK), a member of phosphatidylinositol-kinase family, is a key protein in mammalian DNA double-strand break (DSB) repair that helps to maintain genomic integrity. DNA-PK also plays a central role in immune cell development and protects telomerase during cellular aging. Epigenetic deregulation due to endogenous and exogenous factors may affect the normal function of DNA-PK, which in turn could impair DNA repair and contribute to genomic instability. Recent studies implicate a role for epigenetics in the regulation of DNA-PK expression in normal and cancer cells, which may impact cancer progression and metastasis as well as provide opportunities for treatment and use of DNA-PK as a novel cancer biomarker. In addition, several small molecules and biological agents have been recently identified that can inhibit DNA-PK function or expression, and thus hold promise for cancer treatments. This review discusses the impact of epigenetic alterations and the expression of DNA-PK in relation to the DNA repair mechanisms with a focus on its differential levels in normal and cancer cells.

Shedding of cancer cells from the primary site or undetectable bone marrow region into the circulatory system, resulting in clinically overt metastasis or dissemination, is the hallmark of unfavorable invasive cancers. The shed cells remain in circulation until they extravasate to form a secondary metastatic lesion or undergo anoikis. The circulating tumor cells (CTCs) found as single cells or clusters carry a plethora of information, are acknowledged as potential biomarkers for predicting cancer prognosis and cancer progression, and are supposed to play key roles in determining tailored therapies for advanced diseases. With the advent of novel technologies that allow the precise isolation of CTCs, more and more clinical trials are focusing on the prognostic and predictive potential of CTCs. In this review, we summarize the role of CTCs as a predictive marker for cancer incidence, relapse, and response to therapy.

PurposeThe incidence of <i>BRAF</i> V600E mutation in non-small cell lung carcinoma (NSCLC) is lower than 2%, which poses difficulties in finding legitimate patients for targeted therapy. We investigated the predictive factors pertaining to <i>BRAF</i> V600E and the effectiveness of the VE1 antibody as a screening method for patient selection.Materials and MethodsThe study was designed into two steps. In a first group, <i>BRAF</i>-mutated NSCLCs were identified from sequencing data to determine the features of <i>BRAF</i> V600E mutation. The results of the first group helped the collection of adenocarcinomas with a papillary or micropapillary pattern but without <i>EGFR</i> or <i>ALK</i> alterations as a second group so that the frequency of <i>BRAF</i> V600E mutation could be calculated. The sensitivity and specificity of the VE1 were compared with <i>BRAF</i> V600E status. ResultsAmong 39 <i>BRAF</i>-mutated NSCLCs in the first group, 20 (51%) were V600E. <i>BRAF</i> V600E mutation was more common in female patients and showed no significant correlation with smoking status. Nineteen cases were adenocarcinomas without <i>EGFR</i> and <i>ALK</i> alterations. The most common patterns of invasion were papillary and micropapillary along with central fibrosis. The sensitivity and specificity of the VE1 were 90.0% and 92.3%, respectively. In the second group, 6.7% of cases were VE1-positive, indicating that the prevalence was significantly higher than that reported in previous studies (0.3-1.8%). Conclusion<i>BRAF</i> V600E-mutated NSCLCs could be enriched with the application of clinicopathologic parameters, which are not perfect. Therefore, additional VE1 immunohistochemistry may be useful as a screening method.