Overexpression of ERG in the prostate epithelium, due to chromosomal translocations, contributes to prostate tumorigenesis. Here, genomic analysis of ERG siRNA-treated prostate cells harboring the endogenous TMPRSS2-ERG fusion revealed an inverse relationship between ERG and Annexin A2 (ANXA2) expression at both the RNA and protein level. ANXA2, a Ca2+-dependent and phospholipid-binding protein, is involved in various cellular functions, including maintenance of epithelial cell polarity. Mechanistic studies defined the prostate-specific transcription start site of ANXA2 and showed that the recruitment of ERG to the ANXA2 promoter is required for transcriptional repression by ERG. Knockdown of ERG enhanced the apical localization of ANXA2, the bundling of actin filaments at cell-cell junctions and formation of a polarized epithelial phenotype. ERG overexpression disrupted ANXA2 mediated cell polarity and promoted epithelial mesenchymal transition (EMT) by inhibiting CDC42 and RHOA, and by activating cofilin. Immunohistochemistry (IHC), demonstrated a reciprocal relationship of ANXA2 and ERG expression in a large fraction of primary prostate cancer clinical specimens. ANXA2 was absent or markedly reduced in ERG(+) tumors, which were mostly well-differentiated. ERG(-) tumors, meanwhile, expressed moderate to high levels of ANXA2, and were either poorly-differentiated or displayed subsets of poorly-differentiated cells. Taken together, the transcriptional repression of ANXA2 by ERG in prostate epithelial cells plays a critical role in abrogating differentiation, promoting EMT, and in the reciprocal correlation of ERG and ANXA2 expression observed in human prostate cancer. Implications: ANXA2 is a new component of the ERG network with potential to enhance biological stratification and therapeutic targeting of ERG stratified prostate cancers.
Phosphoserine-containing peptides have been shown to exert antioxidative stress effects, by lowering lipid peroxidation, increasing intracellular glutathione, and increasing the expression of antioxidant enzymes in human intestinal epithelial cells. However, the role of phosphoserine residues in antioxidative stress activity, and their mechanism of action, remains unknown.
Erythroblast transformation-specific-related gene (ERG) fusions, the most common and validated prostate cancer (CaP) genome alteration, result in alterations in the expression of the ERG oncoprotein. Significantly lower frequencies of ERG have been reported in tumors of African American (AA) in comparison to Caucasian American (CA) men. Building on our preliminary observations, this study has focused on the increased association of the ERG-negative status with higher-grade prostate tumors in AA men. Representative whole-mount prostate sections from a matched cohort of 63 AA and 63 CA men with Gleason scores of 4+3 and those with Gleason scores of 8-10 were analyzed for ERG oncoprotein by immunohistochemistry. The striking finding of this study was that ERG expression was 3 times more likely to be present in the higher-grade index tumors of CA men compared to AA men (31 of 63 vs. 10 of 63 patients, respectively; P<0.0001). Although the mechanisms underlying these differences have not been elucidated, the present study along with our previous observations underscores that ERG typing may enhance the understanding of ethnic differences and future targeted therapy of CaP.
Sepsis is a systemic inflammatory response syndrome during infection. Therapeutic agents are essential to protect the host from sepsis. Ovocalyxin-36 (OCX-36) is a chicken eggshell membrane protein and shares protein sequence and gene organization homology with bactericidal permeability-increasing protein (BPI), lipopolysaccharide-binding protein (LBP) and palate, lung and nasal epithelium clone (PLUNC) proteins that play a major role in innate immune protection. We recently reported that OCX-36 binds to both lipopolysaccharide (LPS) and lipoteichoic acid (LTA) (Cordeiro et al., 2013, PLoS ONE 8, e84112), which is an important activity to neutralize endotoxins and non-endotoxin pyrogens during an inflammatory response. Here we investigated the immune modulating effects of OCX-36 and enzymatically digested OCX-36 (dOCX-36) in vitro and in a mouse model of endotoxemia. OCX-36 alone dose-dependently induced both TNF-? and nitric oxide (NO) production by RAW 264.7 macrophage cells, and this immunostimulatory effect was reduced by enzymatic digestion. In the presence of LPS, dOCX-36 was more effective than intact OCX-36 at reducing LPS-induced secretion of TNF-? from RAW 264.7 cells, but did not reduce NO production. In contrast, OCX-36 increased LPS-induced NO production, both in the presence and absence of FBS, PCR array analysis confirmed that OCX-36 and dOCX-36 differentially regulated genes involved in innate immunity, and dOCX-36 down-regulated the expression of genes involved in LPS signaling and inflammatory responses. In vivo, dOCX-36 was more effective at reducing LPS-induced inflammatory symptoms and inhibiting the local production of pro-inflammatory mediators in the small intestine. These results suggest that OCX-36 and OCX-36 derived peptides may differentially modulate innate immune responses, and support our hypothesis that OCX-36 derived peptides have potential therapeutic applications in sepsis.
Gene fusion between TMPRSS2 promoter and the ERG proto-oncogene is a major genomic alteration found in over half of prostate cancers (CaP), which leads to aberrant androgen dependent ERG expression. Despite extensive analysis for the biological functions of ERG in CaP, there is no systematic evaluation of the ERG responsive proteome (ERP). ERP has the potential to define new biomarkers and therapeutic targets for prostate tumors stratified by ERG expression.
ERG oncogene fusions (predominantly TMPRSS2-ERG) represent the most common (50-70% frequency) and validated prostate cancer (CaP) genome alteration in the Western countries. A common TMPRSS2-ERG fusion type leads to the androgen dependent tumor cell specific expression of the TMPRSS2-ERG fusion transcript and amino terminally truncated ERG oncoprotein. CaP prevalence and aggressiveness, as well as genomic alterations vary in different geographic locations in the world. Recent studies from our group highlighted significantly lower frequency of ERG alterations in prostate index tumors of African American men (~30%) in comparison to Caucasian Americans (~60%). Further, much lower frequencies (10 -25%) of ERG alterations have been reported in studies from China and Japan. There is no study on ERG alterations in CaP patients from India, representing a significant portion of the world male population. This study focuses on the frequency of ERG oncoprotein expression in CaP patients from India.
We evaluated the antiinflammatory activity of soy-derived di- and tripeptides in a dextran sodium sulfate (DSS)-induced pig model of intestinal inflammation. In the DSS-positive control (POS) and DSS-positive with soy peptide treatment (SOY) groups (n = 6/group), DSS was administered to piglets via i.g. catheter for 5 d, followed by a 5-d administration of saline or soy-derived peptides, respectively. A negative control (NEG) group received saline in lieu of the DSS and soy peptides. The severity of inflammation was assessed by clinical signs, morphological and histological measurements, gut permeability, and neutrophil infiltration. Local production of TNF and IL6 were measured by ELISA, colonic and ileal inflammatory gene expression were assessed by real-time RT-PCR, and CD4+CD25+ lymphocyte populations were analyzed by flow cytometry. Crypt elongation and muscle thickness, d-mannitol gut permeation, colonic expression of the inflammatory mediators IFNG, IL1B, TNF, RORC, and IL17A as well as the FOXP3 T-regulatory transcription factor, and myeloperoxidase activity were lower (P < 0.05) in the SOY pigs than in POS pigs. Messenger RNA levels of ileal IFNG, TNF, IL12B, and IL17A were lower (P < 0.05) and FOXP3 expression was greater (P < 0.05) in SOY piglets than in the POS group. In the mesenteric lymph nodes, CD4+CD25+ T cells were higher (P < 0.05) in both the POS and SOY groups than in NEG controls. Soy-derived peptides exert antiinflammatory activity in vivo, suggesting their usefulness for the treatment of inflammatory disorders.
Oxidative stress is involved in the initiation and propagation of chronic intestinal pathologies. Bioactive peptides such as egg yolk-derived phosvitin phosphopeptides (PPP3) have been previously shown to reduce in vitro oxidative stress by up-regulating glutathione synthesis and antioxidant enzyme activities. Peptide and gene expression profile analysis of the PPP3 peptides can provide insight into structures involved in signal transduction mechanisms in the antioxidative stress response. The objectives of this research were to identify the PPP3 amino acid sequences before and after simulated gastrointestinal digestion and to assess the genes influenced by PPP3. Peptide sequences were analyzed using ESI Q-TOF-MS/MS, and the expression profile of 84 human oxidative stress and antioxidant defense genes were analyzed. Undigested PPP3 was composed of three main peptides: GTEPDAKTSSSSSSASSTATSSSSSSASSPNRKKPMDE (phosvitin-PV residues 4-41), NSKSSSSSSKSSSSSSRSRSSSKSSSSSSSSSSSSSSKSSSSR (PV residues 155-197), and EDDSSSSSSSSVLSKIWGRHEIYQ (PV residues 244-257) and their fragments. There was limited degradation of PPP3 after gastrointestinal digestion as deduced from the fragment sizes of digested PPP3, which ranged from 5 to 32 amino acids. These fragments were rich in contiguous serines and, in some cases, monoesterified with phosphate. Both undigested and digested PPP3 significantly reduced IL-8 secretion in H(2)O(2)-induced Caco-2 cells, indicating that antioxidative stress bioactivity is retained upon digestion. After PPP3 pretreatment, antioxidant genes associated with oxygen and reactive oxygen species (ROS) metabolism and cellular responses to chemical stimulus, oxidative stress, and ROS are up-regulated in the presence and absence of oxidative stress, thereby contributing to the prevention of intestinal oxidative stress and the promotion of gut health.
Long-term oxidative stress in the gastrointestinal tract can lead to the development of chronic intestinal disorders. Many food-derived antioxidants are effective in vitro, but the variable reports of in vivo efficacy and the pro-oxidant nature of some antioxidants necessitate alternative strategies for the reduction of in vivo oxidative stress. Compounds that up-regulate the production of endogenous antioxidants such as glutathione (GSH) and antioxidant enzymes provide novel approaches for the restoration of redox homeostatis. Egg yolk peptides (EYP) prepared from Alcalase and protease N digestion of delipidated egg yolk proteins were found to exhibit antioxidative stress properties. The effect of EYP supplementation was examined in a hydrogen peroxide-induced human colon cell line and in an animal model of intestinal oxidative stress. EYP significantly reduced the pro-inflammatory cytokine, IL-8, in Caco-2 cells. In piglets given intraperitoneal infusions of hydrogen peroxide, EYP treatment increased GSH and gamma-glutamylcysteine synthetase mRNA expression and activity, significantly increased antioxidant enzyme activities, in particular catalase and glutathione S-transferase activities, and reduced protein and lipid oxidation in the duodenum, jejunum, ileum, and colon. Furthermore, EYP boosted the systemic antioxidant status in blood by increasing the GSH concentration in red blood cells. These results suggest that EYP supplementation is a novel strategy for the reduction of intestinal oxidative stress.
Neural-cadherin is a member of the cadherin gene family encoding the N-cadherin protein that mediates cell adhesion. N-cadherin is a marker of Sertoli cells and is also expressed in germ cells of varying stages of maturation. The purpose of this study was to determine the presence and distribution of this protein by immunohistochemistry in 105 germ cell tumors of both single and mixed histological types and 12 gonadal stromal tumors. Twenty-four germ cell tumors consisted of one cell type and the remaining were mixed. Of the 23 seminomas in either pure or mixed tumors, 74% were positive. Two spermatocytic seminomas were positive. Of the 83 cases with yolk sac tumor, 99% were positive for N-cadherin. The teratomas were positive in 73% in neuroectodermal and / or glandular components. In contrast, 87% of embryonal carcinomas did not express N-cadherin. Only 17% of the syncytiotrophoblastic cells were positive for N-cadherin. In conclusion, N-cadherin expression is very helpful in the identification of yolk sac tumors. In addition to glypican-3 and Sal-like protein 4, N-cadherin can be beneficial for the diagnosis and classification of this subtype of testicular germ cell tumor. Nine of the 12 gonadal stromal tumors were positive to a variable extent.
To systematically evaluate the ETS-related gene (ERG) alterations in the multifocal tumor context using whole-mount prostatectomy specimens from African and Caucasian American patients matched for age, pathologic grade and stage. Oncogenic activation of the ERG is the most common early genomic alteration in patients with prostate cancer (CaP) in Western countries. However, ERG alterations have not been systematically examined in African American patients with a known greater risk of CaP incidence and mortality.
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