Characterisation of the Stromal Microenvironment in Lobular Breast Cancer

SIMPLE SUMMARY: Invasive lobular breast cancer (ILC) accounts for approximately 5–15% of breast cancers, and although response rates to treatments are initially good, an ILC diagnosis is associated with adverse long-term outcomes; better treatments, specifically targeted to this sub-type of breast cancer, are required to improve patient survival. The tumor microenvironment (TME) plays an important role in determining how cancers respond to treatment, and in this study, we carried out an in-depth analysis of the TME in ILC following laser-capture microdissection of the tumor stroma, and analysis of primary cancer-associated fibroblasts (CAFs), which comprise the majority of non-malignant cells within tumors. This identified changes in genes involved in regulation of the extracellular matrix and also growth factor signaling pathways that were differentially regulated in ILC. Further analysis of breast cancer datasets showed that two of these genes which encode a secreted metalloproteinase (PAPPA) and a metalloproteinase inhibitor (TIMP2) were associated with survival outcomes in ILC. ABSTRACT: Invasive lobular carcinoma (ILC) is the second most common histological subtype of breast cancer, and it exhibits a number of clinico-pathological characteristics distinct from the more common invasive ductal carcinoma (IDC). We set out to identify alterations in the tumor microenvironment (TME) of ILC. We used laser-capture microdissection to separate tumor epithelium from stroma in 23 ER+ ILC primary tumors. Gene expression analysis identified 45 genes involved in regulation of the extracellular matrix (ECM) that were enriched in the non-immune stroma of ILC, but not in non-immune stroma from ER+ IDC or normal breast. Of these, 10 were expressed in cancer-associated fibroblasts (CAFs) and were increased in ILC compared to IDC in bulk gene expression datasets, with PAPPA and TIMP2 being associated with better survival in ILC but not IDC. PAPPA, a gene involved in IGF-1 signaling, was the most enriched in the stroma compared to the tumor epithelial compartment in ILC. Analysis of PAPPA- and IGF1-associated genes identified a paracrine signaling pathway, and active PAPP-A was shown to be secreted from primary CAFs. This is the first study to demonstrate molecular differences in the TME between ILC and IDC identifying differences in matrix organization and growth factor signaling pathways

Expression Analysis of the NLRP Gene Family Suggests a Role in Human Preimplantation Development

Background: The NLRP (Nucleotide-binding oligomerization domain, Leucine rich Repeat and Pyrin domain containing) family, also referred to as NALP family, is well known for its roles in apoptosis and inflammation. Several NLRPs have been indicated as being involved in reproduction as well. Methodology: We studied, using the unique human gametes and embryo materials, the expression of the NLRP family in human gametes and preimplantation embryos at different developmental stages, and compared the expression levels between normal and abnormal embryos using real-time PCR. Principal Findings: Among 14 members of the NLRP family, twelve were detected in human oocytes and preimplantation embryos, whereas seven were detected in spermatozoa. Eight NLRPs (NLRP4, 5, 8, 9, 11, 12, 13, and 14) showed a similar expression pattern: their expression levels were high in oocytes and then decreased progressively in embryos, resulting in a very low level in day 5 embryos. However, NLRP2 and NLRP7 showed a different expression pattern: their expression decreased from oocytes to the lowest level by day 3, but increased again by day 5. The expression levels of NLRP5, 9, and 12 were lower in day 1 abnormal embryos but higher in day3 and day5 arrested embryos, when compared with normal embryos at the same stages. NLRP7 was down-regulated in day 1 and day 5 abnormal embryos but over-expressed in day3 arrested embryos

Bifunctional Avidin with Covalently Modifiable Ligand Binding Site

The extensive use of avidin and streptavidin in life sciences originates from the extraordinary tight biotin-binding affinity of these tetrameric proteins. Numerous studies have been performed to modify the biotin-binding affinity of (strept)avidin to improve the existing applications. Even so, (strept)avidin greatly favours its natural ligand, biotin. Here we engineered the biotin-binding pocket of avidin with a single point mutation S16C and thus introduced a chemically active thiol group, which could be covalently coupled with thiol-reactive molecules. This approach was applied to the previously reported bivalent dual chain avidin by modifying one binding site while preserving the other one intact. Maleimide was then coupled to the modified binding site resulting in a decrease in biotin affinity. Furthermore, we showed that this thiol could be covalently coupled to other maleimide derivatives, for instance fluorescent labels, allowing intratetrameric FRET. The bifunctional avidins described here provide improved and novel tools for applications such as the biofunctionalization of surfaces

Heritable symbionts in a world of varying temperature

Heritable microbes represent an important component of the biology, ecology and evolution of many plants, animals and fungi, acting as both parasites and partners. In this review, we examine how heritable symbiont–host interactions may alter host thermal tolerance, and how the dynamics of these interactions may more generally be altered by thermal environment. Obligate symbionts, those required by their host, are considered to represent a thermally sensitive weak point for their host, associated with accumulation of deleterious mutations. As such, these symbionts may represent an important determinant of host thermal envelope and spatial distribution. We then examine the varied relationship between thermal environment and the frequency of facultative symbionts that provide ecologically contingent benefits or act as parasites. We note that some facultative symbionts directly alter host thermotolerance. We outline how thermal environment will alter the benefits/costs of infection more widely, and additionally modulate vertical transmission efficiency. Multiple patterns are observed, with symbionts being cold sensitive in some species and heat sensitive in others, with varying and non-coincident thresholds at which phenotype and transmission are ablated. Nevertheless, it is clear that studies aiming to predict ecological and evolutionary dynamics of symbiont–host interactions need to examine the interaction across a range of thermal environments. Finally, we discuss the importance of thermal sensitivity in predicting the success/failure of symbionts to spread into novel species following natural/engineered introduction

Use of apostrophes by six to nine year old children

This paper investigated six to nine year old children's ability to use contractive and possessive apostrophes, the effects of type and token frequency, and the effect of teaching on this ability. Apostrophes provide an interesting opportunity to examine the basis of frequency effects in rule inference, as type and token frequencies dissociate in apostrophes found in children's early reading materials. Teaching effects are now open to inspection as the National Literacy Strategy Framework for Teaching lays down a specific and detailed curriculum and teaching timetable. Results showed that token but not type frequency influenced children's correct use of apostrophes. There were also effects of teaching

Children's early reading vocabulary: Description and word frequency lists

BACKGROUND: When constructing stimuli for experimental investigations of cognitive processes in early reading development, researchers have to rely on adult or American children's word frequency counts, as no such counts exist for English children. AIM: The present paper introduces a database of children's early reading vocabulary, for use by researchers and teachers. SAMPLE: Texts from 685 books from reading schemes and story books read by 5-7 year-old children were used in the construction of the database. METHOD: All words from the 685 books were typed or scanned into an Oracle database. RESULTS: The resulting up-to-date word frequency list of early print exposure in the UK is available in two forms from a website address given in this paper. This allows access to one list of the words ordered alphabetically and one list of the words ordered by frequency. We also briefly address some fundamental issues underlying early reading vocabulary (e.g., that it is heavily skewed towards low frequencies). Other characteristics of the vocabulary are then discussed. CONCLUSIONS: We hope the word frequency lists will be of use to researchers seeking to control word frequency, and to teachers interested in the vocabulary to which young children are exposed in their reading material

Children's printed word database : Continuities and changes over time in children's early reading vocabulary

In this paper we introduce a comprehensive database of the vocabulary in reading materials used by 5 – 9 year old children in the UK. The database is available on-line http:www.essex.ac.ukpsychologycpwd and allows researchers into early reading development the possibility of rigorous control over critical characteristics of experimental stimuli such as word frequency, regularity and length, frequency of grapheme-phoneme correspondences, orthographic and phonological neighbourhoods etc. The on-line database is also a resource that can be used by practitioners with interests in literacy development and literacy instruction. It can be used to obtain characteristics for a user-generated list of words, or else to generate a list of words according to constraints specified by the user. Here we present an overview of the construction of the database, the materials entered into it, the survey of schools by which we obtained information about the books that were most likely to be used by children in each age group, and the search features available on the database website. We also discuss certain characteristics of the Vocabulary itself and compare these with those reported in an earlier non-representative database reported in Stuart, Dixon, Masterson and Gray (2003). We then present a detailed analysis of the characteristics of Vocabulary in books used in the Reception year, against the background of recent recommendations for change in the early teaching of reading. Finally, we present data showing that the database is indeed already proving a useful resource for both practitioners and researchers

Detection of of <i>NLRPs</i> in human gametes and embryos by PCR.

<p>PCR products were about 70 bp in GV, MI, MII oocytes, day2 (D2), day 3(D3), day 5(D5) embryos and spermatozoa (S). M: marker. <i>NLRP1</i> was detected in GV, MI, MII oocytes and D2 embryos, but not in later stages. <i>NLRP2, 4, 7, 11, 12 and 14</i> were detected in all samples. <i>NLRP3</i> was only detected in GV, MI oocytes and spermatozoa. <i>NLRP5, 8, 9, 13</i> were detected in all oocytes and embryos, but not in spermatozoa. <i>NLPR6</i> was only detected in spermatozoa and <i>NLRP10</i> was not detected in any gametes or embryos (figure not shown).</p

Microarray detection of <i>NLRPs</i> in human oocytes and embryos showed two different expression patterns.

<p>A: <i>NLRP4, 5, 8, 9, 11,12, 13 and14</i> were highly expressed in oocytes and then gradually decreased in embryos with a very low level in day5 (D5) embryos. B: <i>NLRP2</i> and <i>NLRP</i>7 progressively decreased from oocytes to day 3(D3) embryos then showed a sharp increase in D5. Error bars = SEM (standard error of the mean).</p