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Background: Formalin fixation, duration of tissue storage and tissue enrichment techniques can affect DNA methylation yield but these effects have not been quantitatively measured. The aim is to investigate the relative impact of these conditions on DNA methylation in rectal cancer., Methods: 10 rectal cancers with matched undissected fresh frozen tissues, laser capture microdissected (LCM) formalin-fixed paraffin-embedded (FFPE) tissues, manual macrodissected FFPE tissues, adjacent normal mucosa and stromal tissues were analysed for APC and LINE-1 methylation using bisulphite pyrosequencing., Results: FFPE cancer tissues, which had been stored for at least 4 years showed similar APC and LINE-1 methylation changes to matched fresh frozen cancer tissues. Laser capture microdissection did not increase the degree of methylation detected compared to manual macrodissection. Analysis of stromal tissues showed that they had undergone significant methylation changes compared to adjacent macroscopically normal mucosa, but not to the same extent as cancer tissues., Conclusion: Reliable DNA methylation results can be obtained from FFPE rectal cancer tissues, which have been in long-term storage. Because only minor differences in methylation between macrodissected and LCM cancer tissues were found, our results do not support the routine use of LCM to enrich for cancer cells for DNA methylation studies., (© 2013.)

In many rodent laboratories, blood samples are collected from rats using the tail vein nick procedure and analyzed to quantify blood corticosterone levels as an indicator of stress. The standard method of corticosterone quantification often requires the collection of a relatively large volume of blood, followed by the extraction of the blood plasma. An alternative blood sampling method requires the collection of only a drop of blood on paper (the 'drop' method), minimizing handling of the animals, and does not require plasma extraction. The authors aimed to validate the drop method of blood sampling for use in corticosterone quantification. They induced stress in rats by cerebral ischemia, collected blood samples at various intervals using both the drop method and the plasma extraction method and then quantified corticosterone by radioimmunoassay. Corticosterone levels of the ischemic rats were compared with those of sham-operated rats and those of ischemic rats that had been given metyrapone, a glucocorticoid synthesis inhibitor, prior to vessel occlusion. Blood corticosterone levels in the samples obtained from the same animal using the two different methods were highly correlated for all rats. The authors further provide a regression model that can be used to predict plasma corticosterone values from those obtained from the drop blood samples. Quantification of corticosterone from only a small drop of blood has many practical and ethical advantages and should be considered as an alternative to standard methods.

Protein engineering techniques can facilitate the direct de-convolution of specific domains, regions, and particular amino acids that contribute to protein function. Many tools are available to aid this enterprise and herein we describe one such tool, a technique we term "Molecular Scanning" (MS). MS is analogous to previously described alanine scanning in that it samples potentially functional sequence space, but differs in that it uses Error-Prone polymerase chain reaction to randomly introduce all amino acids across the sequence space, as opposed to simply introducing alanine at each desired position. We commonly use MS in conjunction with ribosome-display, selecting for specific character traits (e.g., improved affinity) which allows us to sample functionally relevant diversity on a reasonably large scale. This approach is amenable to a variety of different mutational techniques and display technologies as dictated by user requirements or needs. In this chapter we present a general outline of the process as we have previously successfully applied it.

This paper provides field evidence on the effects of diet on educational outcomes, exploiting a campaign lead in the UK in 2004, which introduced drastic changes in the meals offered in the schools of one borough - Greenwich - shifting from low-budget processed meals towards healthier options. We evaluate the effect of the campaign on educational outcomes in primary schools using a difference in differences approach; comparing educational outcomes in primary schools (Key Stage 2 outcomes more specifically) before and after the reform, using the neighbouring Local Education Authorities as a control group. We find evidence that educational outcomes did improve significantly in English and Science. We also find that authorized absences - which are most likely linked to illness and health - fell by 14%., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Purpose: To evaluate the mammographic features of breast cancer that favor lesion detection with single reading and computer-aided detection (CAD) or with double reading., Materials and Methods: The Computer Aided Detection Evaluation Trial II study was approved by the ethics committee, and all participants provided written informed consent. A total of 31,057 women were recruited from three screening centers between September 2006 and August 2007. They were randomly allocated to the double reading group, the single reading with CAD group, or the double reading and single reading with CAD group at a ratio of 1:1:28, respectively. In this study, cancers in the women whose mammograms were read with both single reading with CAD and double reading were retrospectively reviewed. The original mammograms were obtained for each case and reviewed by two of three experienced breast radiologists in consensus. The method of detection was noted. The size and predominant mammographic feature of the cancer were recorded, as was the breast density. CAD marking data were reviewed to determine if the cancer had been correctly marked., Results: A total of 227 cancers were detected in 28,204 women. A total of 170 cases were recalled with both reading regimens. Lesion types were masses (66%), microcalcifications (25%), parenchymal deformities (6%), and asymmetric densities (3%). The ability of the reading regimens to correctly prompt the reader to recall cases varied significantly by lesion type (P < .001). More parenchymal deformities were recalled with double reading, whereas more asymmetric densities were recalled with single reading with CAD. There was no difference in the ability of either reading regimen to prompt the reader to correctly recall masses or microcalcifications. CAD correctly prompted 100% of microcalcifications, 87% of mass lesions, 80% of asymmetric densities, and 50% of parenchymal deformities. CAD correctly marked 93% of spiculated masses compared with 80% of ill-defined masses (P = .054). There was a significant trend for cancers detected with double reading to occur only in women with a denser mammographic background pattern (P = .02). Size had no effect on lesion detection., Conclusion: Readers using either single reading with CAD or double reading need to be aware of the strengths and weaknesses of reading regimens to avoid missing the more challenging cancer cases.

The mechanisms underlying Golgi targeting and vesiculation are unknown, although the responsible phosphatidylinositol 4-phosphate (PtdIns(4)P) ligand and four-phosphate-adaptor protein (FAPP) modules have been defined. The micelle-bound structure of the FAPP1 pleckstrin homology domain reveals how its prominent wedge independently tubulates Golgi membranes by leaflet penetration. Mutations compromising the exposed hydrophobicity of full-length FAPP2 abolish lipid monolayer binding and compression. The trafficking process begins with an electrostatic approach, phosphoinositide sampling and perpendicular penetration. Extensive protein contacts with PtdIns(4)P and neighbouring phospholipids reshape the bilayer and initiate tubulation through a conserved wedge with features shared by diverse protein modules.

Objective: Basal phenotype has been found to be an independent poor prognostic factor for breast cancer. The aim of this study was to assess the mammographic appearance of screening-detected breast carcinoma with the basal phenotype., Materials and Methods: A series of 1,944 consecutively enrolled patients with operable invasive breast cancer underwent immunohistochemical analysis with cytokeratin 5/6 and cytokeratin 14 markers to identify tumors exhibiting basal phenotype characteristics. Among those patients, 356 women with breast cancer were common to a prospectively collected database of screening-detected cases of breast cancer. The predominant mammographic appearance and any associated features were reported by experienced image readers blinded to phenotype status. A chi-square test was used to assess difference between the mammographic appearances of a group of tumors with the basal phenotype and those of a group with the nonbasal phenotype., Results: Forty-one (12%) of the screening-detected tumors had basal phenotypic expression, and these were compared with 309 (88%) nonbasal tumors. Basal-phenotype tumors were significantly more likely to manifest as an ill-defined mass (basal phenotype, 25 [61%] of 41 tumors; nonbasal phenotype, 75 [24%] of 309 tumors; p < 0.001) or with comedo calcification (basal phenotype, nine [22%] of 41 tumors; nonbasal phenotype, 30 [10%] of 309 tumors; p = 0.019). Nonbasal-phenotype tumors were more likely to manifest as a spiculated mass (nonbasal phenotype, 150 [49%] of 309 tumors; basal phenotype, eight [20%] of 41 tumors; p < 0.001). The low rate of spiculation in basal tumors was independent of histologic grade., Conclusion: Screening-detected breast tumors with a basal phenotype have a mammographic appearance different from that of nonbasal tumors. This finding may explain the good prognostic value of mammographic spiculation reported in previous studies.

Objective: The aim of our study was to assess the effect of mammographic parenchymal pattern on patient survival, mammographic features, and pathologic features of breast cancer in a screened population., Materials and Methods: We classified the parenchymal pattern (according to BI-RADS) of 759 screened women who presented with a screening-detected (n = 455) or interval (n = 304) invasive breast cancer. Pathologic details (tumor size, histologic grade, lymph node stage, vascular invasion, and histologic type) and mammographic appearances were recorded. Breast cancer-specific survival was ascertained, with a median follow-up of 9.0 years., Results: An excess of interval cancers was seen in women with dense breasts (p < 0.0001). Screening-detected (but not interval) tumors were significantly smaller in fatty breasts (p = 0.014). Tumor grade, lymph node stage, vascular invasion, and histologic type did not vary significantly with mammographic parenchymal pattern in screening-detected or interval cancers. Screening-detected cancers in fatty breasts were more likely to appear as indistinct (p = 0.003) or spiculated (p = 0.002) masses in contrast to cancers in dense breasts, which more commonly appeared as architectural distortions (p < 0.0001). No significant breast cancer-specific survival difference was seen by mammographic parenchymal pattern for screening-detected cancers (p = 0.75), interval cancers (p = 0.82), or both groups combined (p = 0.12)., Conclusion: The prognosis of screened women presenting with breast cancer is unrelated to dense mammographic parenchymal pattern despite an excess of interval cancers and larger screening-detected tumors in this group. These data support the mammographic screening of women with dense parenchymal patterns.

Interferon-alpha (IFN-alpha) is used as a front-line treatment for cancer and other diseases. Reports of depression as a consequence of IFN-alpha therapy scatter the literature, generating interest in the CNS disruptions elicited by this cytokine. In the present work, we investigated the short- and long-term effects of a single systemic injection of vehicle, 10, or 1000 units of IFN-alpha on temperature, body weight, food intake, sickness behaviours, locomotor activity, and brain stimulation reward (BSR) thresholds elicited from the ventral tegmental area in female Long-Evans rats. Pioneered for studying motivational processes, BSR has been exploited as a tool for tracking hedonic status in animal models of depression. In this study, the main findings were that IFN-alpha did not induce anhedonia as defined by no increase in frequency thresholds. However, the analyses of sickness behaviours unveiled a significant increase in piloerection in all sham control animals that received an IFN-alpha injection while the BSR animal scores remained relatively unchanged between pre- and post-injection days. This pattern was also evident in the overall total sickness behaviour scores. Our data suggest that a single exposure to IFN-alpha treatment in female rats elicits long-term somatic effects, without altering hedonic status.

Objective: The aim of this study was to review the prognostic significance of pathologic and radiologic factors for screening-detected invasive breast cancers of any size., Materials and Methods: The patient group was a consecutive series of 470 screening-detected invasive breast cancers that were diagnosed between 1988 and 1998. Data regarding tumor type, grade, maximum invasive diameter, lymph node status, and the presence or absence of vascular invasion were recorded, as were the mammographic features of the lesion. Survival was ascertained from hospital records and a cancer registry. Differences in survival were assessed using Kaplan-Meier survival curves with a log-rank test for difference. The significance of any correlations was assessed using the chi-square test and the chi-square test for trend. Multivariate analysis used a Cox proportional hazards model., Results: At univariate analysis, large invasive size, the presence of definite vascular invasion, high histologic grade, and nodal involvement were associated with poorer breast-cancer-specific survival. Mammographic spiculation (the presence of either a spiculated mass or distortion) was associated with more prolonged breast-cancer-specific survival. The presence or absence of mammographic comedo calcification did not influence breast-cancer-specific survival. In a Cox multivariate analysis that included those factors significant in univariate analysis, size, grade, nodal stage, and mammographic spiculation maintained their prognostic significance., Conclusion: Mammographic spiculation is an independent, good prognostic factor for screening-detected invasive breast cancer. The mechanism of how mammographic spiculation confers a beneficial prognostic effect is not clear.