Value of Magnifying Chromoendoscopy and Magnifying Optical Enhancement Technology in Classifying Colorectal Polyps: A Prospective Controlled Study.

BACKGROUND AND AIMS: Magnifying chromoendoscopy (ME-CE) through the observation of pit patterns is a productive way to distinguish between neoplastic and nonneoplastic polyps. Magnifying optical enhancement technology (ME-OE) is an emerging virtual chromoendoscopy imaging technology and appeared to be a promising approach. However, this information is currently not available. This study is aimed at comparing the differential diagnostic value of ME-CE and OE for neoplastic and nonneoplastic polyps. Patients and Methods. Consecutive patients undergoing colonoscopy were randomized (1 : 1) into examination by ME-OE or ME-CE. Histopathological findings were utilized as the reference standard. Accuracy, sensitivity, specificity, and positive and negative predictive values of two endoscopy methods were compared using ME-OE (were classified according to the JNET classification) and ME-CE (were classified according to the Kudo pit pattern classification), respectively, and the time to predict the histological polyp type was compared. And the agreements between the pathological and clinical diagnosis by ME-OE or ME-CE were analyzed. RESULTS: A total of 365 polyps were found in the 220 patients included (ME-OE: 185; ME-CE: 180.202 had nonneoplastic polyps, 163 had neoplastic polyps). The diagnostic accuracy of ME-OE was higher than that of ME-CE (93% vs. 92%, p > 0.05). The average diagnosis time was lower in ME-OE than ME-CE (83 ± 26.4 s vs. 194 ± 17.7 s, p < 0.001). The agreements between the pathological and clinical diagnosis were at least substantial in both groups. CONCLUSION: ME-OE was superlative to ME-CE in predicting the histology of polyps. OE devoted classification would possibly similarly enhance the endoscopist performance. The trial is registered with ChiCT2000032075.

Comparison of white-light endoscopy, optical-enhanced and acetic-acid magnifying endoscopy for detecting gastric intestinal metaplasia: A randomized trial.

BACKGROUND: Gastric intestinal metaplasia (GIM) is a precancerous lesion of the stomach, which severely affects human life and health. Currently, a variety of endoscopic techniques are used to screen/evaluate GIM. Traditional white-light endoscopy (WLE) and acetic-acid chromoendoscopy combined with magnifying endoscopy (ME-AAC) are the interventions of choice due to their diagnostic efficacy for GIM. Optical-enhanced magnifying endoscopy (ME-OE) is a new virtual chromoendoscopy technique to identify GIM, which combines bandwidth-limited light and image enhancement processing technology to enhance the detection of mucosal and vascular details. We hypothesized that ME-OE is superior to WLE and ME-AAC in the evaluation of GIM. AIM: To directly compare the diagnostic value of WLE, ME-AAC, and ME-OE for detection of GIM. METHODS: A total of 156 patients were subjected to consecutive upper gastrointestinal endoscopy examinations using WLE, ME-AAC, and ME-OE. Histopathological findings were utilized as the reference standard. Accuracy, sensitivity, specificity, and positive and negative predictive values of the three endoscopy methods in the diagnosis of GIM were evaluated. Moreover, the time to diagnosis with ME-AAC and ME-OE was analyzed. Two experts and two non-experts evaluated the GIM images diagnosed using ME-OE, and diagnostic accuracy and intra- and inter-observer agreement were analyzed. RESULTS: GIM was detected in 68 of 156 patients (43.6%). The accuracy of ME-OE was highest (91.7%), followed by ME-AAC (86.5%), while that of WLE (51.9%) was lowest. Per-site analysis showed that the overall diagnostic accuracy of ME-OE was higher than that of ME-AAC (P = 0.011) and WLE (P < 0.001). The average diagnosis time was lower in ME-OE than in ME-AAC (64 ± 7 s vs 151 ± 30 s, P < 0.001). Finally, the inter-observer agreement was strong for both experts (k = 0.862) and non-experts (k = 0.800). The internal consistency was strong for experts (k = 0.713, k = 0.724) and moderate for non-experts (k = 0.667, k = 0.598). CONCLUSION: For endoscopists, especially experienced endoscopists, ME-OE is an efficient, convenient, and time-saving endoscopic technique that should be used for the diagnosis of GIM.

Expression of miR-129-5p and miR-433 in the serum of breast cancer patients and their relationship with clinicopathological features.

Expression of miR-129-5p and miR-433 was detected in breast cancer to explore the relationship with clinicopathological features of breast cancer. Seventy-eight patients with breast cancer diagnosed in Zhengzhou Central Hospital Affiliated to Zhengzhou University (Zhengzhou, China) from February 2016 to September 2017 were collected and enrolled into the research group. Additionally, 72 healthy people who underwent physical examination during the same period were selected as the control group. The expression levels of miR-129-5p and miR-433 in peripheral blood of the two groups were detected by fluorescence quantitative PCR (RT-PCR). The relationship between the expression of miR-129-5p, miR-433 and clinicopathological features, clinical stages of breast cancer, and the degree of differentiation were analyzed. Expression of miR-129-5p and miR-433 in the research group was significantly lower than that in the control group (P<0.05). Expression of miR-129-5p in the blood of breast cancer patients was correlated with tumor size, differentiation degree, lymph node metastasis, depth of invasion and clinical stages (P<0.05). Expression level of miR-433 was correlated with the degree of differentiation, lymph node metastasis, depth of invasion and clinical stages (P<0.05). miR-129-5p and miR-433 were positively correlated with differentiation degree (r=0.8507, r=0.7522; P<0.05), and negatively correlated with clinical stages (r=-0.6595, -0.8947; P<0.05). The sensitivity and area under curve (AUC) were higher in joint detection (87.5 and 0.95% respectively), compared with those in single detection. Patients were separated into high and low expression groups according to the median values of miR-129-5p and miR-433. The one-year survival rate of breast cancer patients was analyzed. Patients in the low expression groups had lower survival rates than patients in the high expression groups (P<0.05). In conclusion, the expression of miR-129-5p and miR-433 in peripheral blood of breast cancer patients is lower than that of healthy people, and the expression level is closely related to clinical stages and differentiation degree, which is expected to provide reference value for judging the state of breast cancer patients. The combined detection of miR-129-5p and miR-433 is of great significance in the diagnosis and treatment of breast cancer.

Combining radiation with autophagy inhibition enhances suppression of tumor growth and angiogenesis in esophageal cancer.

Radiotherapy is an effective treatment for esophageal cancer; however, tumor resistance to radiation remains a major biological problem. The present study aimed to investigate whether inhibition of autophagy may decrease overall tumor resistance to radiation. The effects of the autophagy inhibitor 3-methyladenine (3-MA) on radiosensitivity were tested in the EC9706 human esophageal squamous cell carcinoma cell line by colony formation assay. Furthermore, the synergistic cytotoxic effects of 3-MA and radiation were assessed in a tumor xenograft model in nude mice. Mechanistic studies were performed using flow cytometry, immunohistochemistry and western blot analysis. The results of the present study demonstrated that radiation induced an accumulation of autophagosomes and 3-MA effectively inhibited radiation-induced autophagy. Inhibition of autophagy was shown to significantly increase the radiosensitivity of the tumors in vitro and in vivo. The enhancement ratio of sensitization in EC9706 cells was 1.76 when the cells were treated with 10 mM 3-MA, alongside ionizing radiation. In addition, autophagy inhibition increased apoptosis and reduced tumor cell proliferation. The combination of radiation and autophagy inhibition resulted in a significant reduction in tumor volume and vasculature in the murine model. The present study demonstrated in vitro and in vivo that radiation-induced autophagy has a protective effect against cell death, and inhibition of autophagy is able to enhance the radiosensitivity of esophageal squamous cell carcinoma.

Autophagy-related proteins LC3 and Beclin-1 impact the efficacy of chemoradiation on esophageal squamous cell carcinoma.

Autophagy is a bulk protein and organelle degradation process essential for cellular maintenance, cell viability and development. This study investigated the prognostic role of LC3 and Beclin-1, two autophagy-related proteins, in patients with locally advanced esophageal squamous cell carcinoma (ESCC) treated with definitive chemoradiation. The data of 150 patients with stages II-IVa ESCC who had undergone definitive concurrent chemoradiation were studied; LC3 and Beclin-1 protein expression was evaluated by immunohistochemistry and western blot. The correlations between LC3 and Beclin-1 expression, the patients' clinicopathological features, and overall survival were analyzed. Eighty-three patients showed positive LC3 and 84 showed positive Beclin-1 protein expressions, but LC3 and Beclin-1 expression in ESCC was not significantly correlated (P=0.746). LC3 and Beclin-1 expression did not show any association with gender, age, tumor location, and treatment response. The median survival of patients with positive LC3 expression was 23.6 months, while the median survival was 32.0 months in patients with negative LC3 expression (P=0.049). The patients with LC3 and Beclin-1-positive tumors presented much poorer long-term survival. Multivariate analysis showed that clinical stage (P=0.019), treatment response (P=0.002), and LC3 expression level (P=0.021) were independent predictive factors of overall survival in patients with locally advanced ESCC receiving definitive chemoradiation. Patients with LC3 and Beclin-1-negative expression had a better overall survival than those with LC3 and Beclin-1-positive tumors. LC3 expression is an independent prognostic factor for overall survival in ESCC patients treated with definitive chemoradiation.

Product feedback regulation implicated in translational control of the Trypanosoma brucei S-adenosylmethionine decarboxylase regulatory subunit prozyme.

Human African sleeping sickness (HAT) is caused by the parasitic protozoan Trypanosoma brucei. Polyamine biosynthesis is an important drug target in the treatment of HAT. Previously we showed that trypanosomatid S-adenosylmethionine decarboxylase (AdoMetDC), a key enzyme for biosynthesis of the polyamine spermidine, is activated by heterodimer formation with an inactive paralogue termed prozyme. Furthermore, prozyme protein levels were regulated in response to reduced AdoMetDC activity. Herein we show that T. brucei encodes three prozyme transcripts. The 3'UTRs of these transcripts were mapped and chloramphenicol acetyltransferase (CAT) reporter constructs were used to identify a 1.2 kb region that contained a 3'UTR prozyme regulatory element sufficient to upregulate CAT protein levels (but not RNA) upon AdoMetDC inhibition, supporting the hypothesis that prozyme expression is regulated translationally. To gain insight into trans-acting factors, genetic rescue of AdoMetDC RNAi knock-down lines with human AdoMetDC was performed leading to rescue of the cell growth block, and restoration of prozyme protein to wild-type levels. Metabolite analysis showed that prozyme protein levels were inversely proportional to intracellular levels of decarboxylated AdoMet (dcAdoMet). These data suggest that prozyme translation may be regulated by dcAdoMet, a metabolite not previously identified to play a regulatory role.

RNA interference-mediated silencing of ornithine decarboxylase and spermidine synthase genes in Trypanosoma brucei provides insight into regulation of polyamine biosynthesis.

Polyamine biosynthesis is a drug target for the treatment of African sleeping sickness; however, mechanisms regulating the pathway in Trypanosoma brucei are not well understood. Recently, we showed that RNA interference (RNAi)-mediated gene silencing or the inhibition of S-adenosylmethionine decarboxylase (AdoMetDC) led to the upregulation of the AdoMetDC activator, prozyme, and ornithine decarboxylase (ODC) proteins. To determine if this regulatory response is specific to AdoMetDC, we studied the effects of the RNAi-induced silencing of the spermidine synthase (SpdSyn) and ODC genes in bloodstream form T. brucei. The knockdown of either gene product led to the depletion of the polyamine and trypanothione pools and to cell death. Decarboxylated AdoMet levels were elevated, while AdoMet was not affected. There was no significant effect on the protein levels of other polyamine pathway enzymes. The treatment of parasites with the ODC inhibitor alpha-difluoromethylornithine gave similar results to those observed for ODC knockdown. Thus, the cellular response to the loss of AdoMetDC activity is distinctive, suggesting that AdoMetDC activity controls the expression levels of the other spermidine biosynthetic enzymes. RNAi-mediated cell death occurred more rapidly for ODC than for SpdSyn. Further, the ODC RNAi cells were rescued by putrescine, but not spermidine, suggesting that the depletion of both putrescine and spermidine is more detrimental than the depletion of spermidine alone. This finding may contribute to the effectiveness of ODC as a target for the treatment of African sleeping sickness, thus providing important insight into the mechanism of action of a key antitrypanosomal agent.

The central kink region of fowlicidin-2, an alpha-helical host defense peptide, is critically involved in bacterial killing and endotoxin neutralization.

Fowlicidins are a group of newly identified chicken cathelicidin host defense peptides. We have shown that the putatively mature fowlicidin-2 of 31 amino acid residues possesses potent antibacterial and lipopolysaccharide (LPS)- neutralizing activities, but with a noticeable toxicity to mammalian cells. As a first step in exploring the structure-activity relationships of fowlicidin-2, in this study we determined its tertiary structure by nuclear magnetic resonance spectroscopy. Unlike the majority of cathelicidins, which are composed of a predominant alpha-helix with a short hinge sequence near the center, fowlicidin-2 consists of 2 well-defined alpha-helical segments (residues 6-12 and 23-27) connected by a long extensive kink (residues 13-20) induced by proline. To further investigate the functional significance of each of these structural components, several N- and C-terminal deletion analogs of fowlicidin-2 were synthesized and analyzed for their antibacterial, cytotoxic and LPS-neutralizing activities. Our results indicated that neither the N- nor C-terminal alpha-helix alone is sufficient to confer any function. Rather, fowlicidin-2(1-18) and fowlicidin-2(15-31), 2 alpha-helical segments with inclusion of the central cationic kink region, retained substantial capacities to kill bacteria and neutralize the LPS-induced proinflammatory response, relative to the parent peptide. More desirably, these 2 peptide analogs showed substantially reduced toxicity to human erythrocytes and epithelial cells, indicative of improved potential as antibacterial and antisepsis agents. To our knowledge, fowlicidin-2 is the first alpha-helical cathelicidin, with the central kink region shown to be critically important in killing bacteria and neutralizing LPS.

Structure-activity relationships of fowlicidin-1, a cathelicidin antimicrobial peptide in chicken.

Cationic antimicrobial peptides are naturally occurring antibiotics that are actively being explored as a new class of anti-infective agents. We recently identified three cathelicidin antimicrobial peptides from chicken, which have potent and broad-spectrum antibacterial activities in vitro (Xiao Y, Cai Y, Bommineni YR, Fernando SC, Prakash O, Gilliland SE & Zhang G (2006) J Biol Chem281, 2858-2867). Here we report that fowlicidin-1 mainly adopts an alpha-helical conformation with a slight kink induced by glycine close to the center, in addition to a short flexible unstructured region near the N terminus. To gain further insight into the structural requirements for function, a series of truncation and substitution mutants of fowlicidin-1 were synthesized and tested separately for their antibacterial, cytolytic and lipopolysaccharide (LPS)-binding activities. The short C-terminal helical segment after the kink, consisting of a stretch of eight amino acids (residues 16-23), was shown to be critically involved in all three functions, suggesting that this region may be required for the peptide to interact with LPS and lipid membranes and to permeabilize both prokaryotic and eukaryotic cells. We also identified a second segment, comprising three amino acids (residues 5-7) in the N-terminal flexible region, that participates in LPS binding and cytotoxicity but is less important in bacterial killing. The fowlicidin-1 analog, with deletion of the second N-terminal segment (residues 5-7), was found to retain substantial antibacterial potency with a significant reduction in cytotoxicity. Such a peptide analog may have considerable potential for development as an anti-infective agent.

Identification and functional characterization of three chicken cathelicidins with potent antimicrobial activity.

Cathelicidins comprise a family of antimicrobial peptides sharing a highly conserved cathelin domain. Here we report that the entire chicken genome encodes three cathelicidins, namely fowlicidin-1 to -3, which are densely clustered within a 7.5-kb distance at the proximal end of chromosome 2p. Each fowlicidin gene adopts a fourexon, three-intron structure, typical for a mammalian cathelicidin. Phylogenetic analysis revealed that fowlicidins and a group of distantly related mammalian cathelicidins known as neutrophilic granule proteins are likely to originate from a common ancestral gene prior to the separation of birds from mammals, whereas other classic mammalian cathelicidins may have been duplicated from the primordial gene for neutrophilic granule proteins after mammals and birds are diverged. Similar to ovine cathelicidin SMAP-29, putatively mature fowlicidins displayed potent and salt-independent activities against a range of Gram-negative and Gram-positive bacteria, including antibiotic-resistant strains, with minimum inhibitory concentrations in the range of 0.4-2.0 microm for most strains. Fowlicidin-1 and -2 also showed cytotoxicity, with 50% killing of mammalian erythrocytes or epithelial cells in the range of 6-40 microm. In addition, two fowlicidins demonstrated a strong positive cooperativity in binding lipopolysaccharide (LPS), resulting in nearly complete blockage of LPS-mediated proinflammatory gene expression in RAW264.7 cells. Taken together, fowlicidin-1 and -2 are clearly among the most potent cathelicidins that have been reported. Their broad spectrum and salt-insensitive antibacterial activities, coupled with their potent LPS-neutralizing activity, make fowlicidins excellent candidates for novel antimicrobial and anti-sepsis agents.

A genome-wide screen identifies a single beta-defensin gene cluster in the chicken: implications for the origin and evolution of mammalian defensins.

BACKGROUND: Defensins comprise a large family of cationic antimicrobial peptides that are characterized by the presence of a conserved cysteine-rich defensin motif. Based on the spacing pattern of cysteines, these defensins are broadly divided into five groups, namely plant, invertebrate, alpha-, beta-, and theta-defensins, with the last three groups being mostly found in mammalian species. However, the evolutionary relationships among these five groups of defensins remain controversial. RESULTS: Following a comprehensive screen, here we report that the chicken genome encodes a total of 13 different beta-defensins but with no other groups of defensins being discovered. These chicken beta-defensin genes, designated as Gallinacin 1-13, are clustered densely within a 86-Kb distance on the chromosome 3q3.5-q3.7. The deduced peptides vary from 63 to 104 amino acid residues in length sharing the characteristic defensin motif. Based on the tissue expression pattern, 13 beta-defensin genes can be divided into two subgroups with Gallinacin 1-7 being predominantly expressed in bone marrow and the respiratory tract and the remaining genes being restricted to liver and the urogenital tract. Comparative analysis of the defensin clusters among chicken, mouse, and human suggested that vertebrate defensins have evolved from a single beta-defensin-like gene, which has undergone rapid duplication, diversification, and translocation in various vertebrate lineages during evolution. CONCLUSIONS: We conclude that the chicken genome encodes only beta-defensin sequences and that all mammalian defensins are evolved from a common beta-defensin-like ancestor. The alpha-defensins arose from beta-defensins by gene duplication, which may have occurred after the divergence of mammals from other vertebrates, and theta-defensins have arisen from alpha-defensins specific to the primate lineage. Further analysis of these defensins in different vertebrate lineages will shed light on the mechanisms of host defense and evolution of innate immunity.