BACKGROUND: The Multi-Omics for Mothers and Infants consortium aims to improve birth outcomes. Preterm birth is a major obstetrical complication globally and causes significant infant and childhood morbidity and mortality. OBJECTIVE: We analyzed placental samples (basal plate, placenta or chorionic villi, and the chorionic plate) collected by the 5 Multi-Omics for Mothers and Infants sites, namely The Alliance for Maternal and Newborn Health Improvement Bangladesh, The Alliance for Maternal and Newborn Health Improvement Pakistan, The Alliance for Maternal and Newborn Health Improvement Tanzania, The Global Alliance to Prevent Prematurity and Stillbirth Bangladesh, and The Global Alliance to Prevent Prematurity and Stillbirth Zambia. The goal was to analyze the morphology and gene expression of samples collected from preterm and uncomplicated term births. STUDY DESIGN: The teams provided biopsies from 166 singleton preterm (<37 weeks' gestation) and 175 term (>= 37 weeks' gestation) deliveries. The samples were fixed in formalin and paraffin embedded. Tissue sections from these samples were stained with hematoxylin and eosin and subjected to morphologic analyses. Other placental biopsies (n=35 preterm, 21 term) were flash frozen, which enabled RNA purification for bulk transcriptomics. RESULTS: The morphologic analyses revealed a surprisingly high rate of inflammation that involved the basal plate, placenta or chorionic villi, and the chorionic plate. The rate of inflammation in chorionic villus samples, likely attributable to chronic villitis, ranged from 25% (Pakistan site) to 60% (Zambia site) of cases. Leukocyte infiltration in this location vs in the basal plate or chorionic plate correlated with preterm birth. Our transcriptomic analyses identified 267 genes that were differentially expressed between placentas from preterm vs those from term births (123 upregulated, 144 downregulated). Mapping the differentially expressed genes onto single-cell RNA sequencing data from human placentas suggested that all the component cell types, either singly or in subsets, contributed to the observed dysregulation. Consistent with the histopathologic findings, gene ontology analyses highlighted the presence of leukocyte infiltration or activation and inflammatory responses in both the fetal and maternal compartments. CONCLUSION: The relationship between placental inflammation and preterm birth is appreciated in developed countries. In this study, we showed that this link also exists in developing geographies. In addition, among the participating sites, we found geographic- and population- based differences in placental inflammation and preterm birth, suggesting the importance of local factors.

BACKGROUND:Fluoride exposure during pregnancy has been associated with various effects on offspring, including changes in behavior and IQ. To provide clues to possible mechanisms by which fluoride may affect human fetal development, we completed proteomic analyses of cord blood serum collected from second-trimester pregnant women residing in northern California, USA. OBJECTIVE:To identify changes in cord blood proteins associated with maternal serum fluoride concentration in pregnant women. METHODS:The proteomes of 19 archived second-trimester cord blood samples from women living in northern California, USA, and having varied serum fluoride concentrations, were analyzed by quantitative mass spectrometry. The 327 proteins that were quantified were characterized by their abundance relative to maternal serum fluoride concentration, and subjected to pathway analyses using PANTHER and Ingenuity Pathway Analysis processes. RESULTS:Pathway analyses showed significant increases in process related to reactive oxygen species and cellular oxidant detoxification, associated with increasing maternal serum fluoride concentrations. Pathways showing significant decreases included complement cascade, suggesting alterations in alterations in process associated with inflammation. CONCLUSION:Maternal fluoride exposure, as measured by serum fluoride concentrations in a small, but representative sample of women from northern California, USA, showed significant changes in the second trimester cord blood proteome relative to maternal serum fluoride concentration.

BackgroundThe Multi-Omics for Mothers and Infants (MOMI) consortium aims to improve birth outcomes. Preterm birth is a major obstetric complication globally causing significant infant and childhood morbidity and mortality.ObjectivesWe analyzed placental samples (basal plate, placenta/chorionic villi and/or the chorionic plate) collected by the 5 MOMI sites: The Alliance for Maternal and Newborn Health Improvement (AMANHI) Bangladesh, AMANHI Pakistan, AMANHI Tanzania, The Global Alliance to Prevent Prematurity and Stillbirth (GAPPS) Bangladesh and GAPPS Zambia. The goal was to analyze the morphology and gene expression of samples collected from preterm and uncomplicated term births.Study designThe teams provided biopsies from 166 singleton preterm (<37 weeks) and 175 term (≥37 weeks) deliveries. They were formalin-fixed and paraffin embedded. Tissue sections from these samples were stained with hematoxylin and eosin and subjected to morphological analyses. Other placental biopsies (n = 35 preterm, 21 term) were flash frozen, which enabled RNA purification for bulk transcriptomics.ResultsThe morphological analyses revealed a surprisingly high rate of inflammation involving the basal plate, placenta/chorionic villi and/or the chorionic plate. The rate in chorionic villus samples, likely attributable to chronic villitis, ranged from 25% (Pakistan site) to 60% (Zambia site) of cases. Leukocyte infiltration in this location vs. the basal plate or chorionic plate correlated with preterm birth. Our transcriptomic analyses identified 267 genes as differentially expressed (DE) between placentas from preterm vs. term births (123 upregulated, 144 downregulated). Mapping the DE genes onto single cell RNA-seq data from human placentas suggested that all the component cell types, either singly or in subsets, contributed to the observed dysregulation. Consistent with the histopathological findings, GO (Gene Ontology) analyses highlighted leukocyte infiltration/activation and inflammatory responses in both the fetal and maternal compartments.ConclusionThe relationship between placental inflammation and preterm birth is appreciated in developed countries. Here, we show that this link also exists in developing geographies. Also, among the participating sites, we found geographic- and/or population-based differences in placental inflammation and preterm birth, suggesting the importance of local factors.

ABSTRACT Human placental architecture is complex. Its surface epithelium, specialized for transport, forms by fusion of cytotrophoblast progenitors into multinucleated syncytiotrophoblasts. Near the uterine surface, these progenitors assume a different fate, becoming cancer-like cells that invade its lining and blood vessels. The latter process physically connects the placenta to the mother and shunts uterine blood to the syncytiotrophoblasts. Isolation of trophoblast subtypes is technically challenging. Upon removal, syncytiotrophoblasts disintegrate and invasive cytotrophoblasts are admixed with uterine cells. We used laser capture to circumvent these obstacles. This enabled isolation of syncytiotrophoblasts and two subpopulations of invasive cytotrophoblasts from cell columns and the endovascular compartment of spiral arteries. Transcriptional profiling revealed numerous genes, the placental or trophoblast expression of which was not known, including neurotensin and C4ORF36. Using mass spectrometry, discovery of differentially expressed mRNAs was extended to the protein level. We also found that invasive cytotrophoblasts expressed cannabinoid receptor 1. Unexpectedly, screening agonists and antagonists showed that signals from this receptor promote invasion. Together, these results revealed previously unseen gene expression patterns that translate to the protein level. Our data also suggested that endogenous and exogenous cannabinoids can affect human placental development.

Purpose This paper investigates the effect of Walmart presence on professional service firms within the finance and insurance sector. This paper also discusses the finance and insurance industry’s place within the Walmart ripple effect—changed retail sector leads to changed professional support services resulting in altered levels of social capital within the community. Design/Methodology The study uses county-level data from the state of Nebraska to examine the relationship between cumulative Walmart exposure and the number of firms, number of employees, and payroll levels within the finance and insurance (F&I) sector. These same variables are also investigated within two specific F&I industries—insurance and banking. Findings The study documents a negative association between cumulative Walmart presence and the number of employees in the finance and insurance industry and a positive relationship between Walmart exposure and F&I payroll levels. The insurance industry results are consistent with the F&I sector result relating to payroll, but show a significant positive relationship between cumulative Walmart exposure and number of insurance agencies. Empirical results from the commercial banking industry are consistent with both the F&I sector and insurance industries with a positive relationship between Walmart presence over time and payroll levels. Originality/value This study contributes to the Walmart effect literature by considering the impact of Walmart presence beyond the retail sector. The paper investigates the effect of Walmart on the finance and insurance profession, a sector that 1) competes directly with Walmart Supercenters for money services, 2) provides support services for retail businesses impacted by Walmart services, and 3) potentially suffers from centralized benefits provided to employees by Walmart headquarters rather than local insurers. The paper concludes that the mixed impact on professional firms results in social capital that could be both strengthened and challenged by Walmart presence.

We are interested in the effects of dividend reduction decisions on the firms that make them. We compare financial ratios of distressed dividend-reducing firms to distressed firms that did not reduce dividends, of nondistressed dividend-reducing firms to nondistressed firms that did not reduce dividends, and ratios of distressed dividend- reducing firms to nondistressed dividend-reducing firms. Results indicate benefits of dividend reductions to both distressed and nondistressed firms. We find a cost to nondistressed firms, i.e., a drop in the market value of the firm in the year following the dividend reduction.

We present a strategy to discover recombinant monoclonal antibodies (mAbs) to specific cancers and demonstrate this approach using basal subtype breast cancers. A phage antibody library was depleted of antibodies to common cell surface molecules by incubation with luminal breast cancer cell lines, and then selected on a single basal-like breast cancer cell line (MDA-MB-231) for binding associated receptor-mediated endocytosis. Additional profiling against two luminal and four basal-like cell lines revealed 61 unique basal-specific mAbs from a pool of 1440 phage antibodies. The unique mAbs were further screened on nine basal and seven luminal cell lines to identify those with the greatest affinity, specificity, and internalizing capability for basal-like breast cancer cells. Among the internalizing basal-specific mAbs were those recognizing four transmembrane receptors (EphA2, CD44, CD73 and EGFR), identified by immunoprecipitation-mass spectrometry and yeast-displayed antigen screening. Basal-like breast cancer expression of these four receptors was confirmed using a bioinformatic approach, and expression microarray data on 683 intrinsically subtyped primary breast tumors. This overall approach, which sequentially employs phage display antibody library selection, antigen identification and bioinformatic confirmation of antigen expression by cancer subtypes, offers efficient production of high-affinity mAbs with diagnostic and therapeutic utility against specific cancer subtypes.

We used isobaric mass tagging (iTRAQ) and lectin affinity capture mass spectrometry (MS)-based workflows for global analyses of parotid saliva (PS) and whole saliva (WS) samples obtained from patients diagnosed with primary Sjögren's Syndrome (pSS) who were enrolled in the Sjögren's International Collaborative Clinical Alliance (SICCA) as compared with two control groups. The iTRAQ analyses revealed up- and down-regulation of numerous proteins that could be involved in the disease process (e.g., histones) or attempts to mitigate the ensuing damage (e.g., bactericidal/permeability increasing fold containing family (BPIF) members). An immunoblot approach applied to independent sample sets confirmed the pSS associated up-regulation of β2-microglobulin (in PS) and down-regulation of carbonic anhydrase VI (in WS) and BPIFB2 (in PS). Beyond the proteome, we profiled the N-glycosites of pSS and control samples. They were enriched for glycopeptides using lectins Aleuria aurantia and wheat germ agglutinin, which recognize fucose and sialic acid/N-acetyl glucosamine, respectively. MS analyses showed that pSS is associated with increased N-glycosylation of numerous salivary glycoproteins in PS and WS. The observed alterations of the salivary proteome and N-glycome could be used as pSS biomarkers enabling easier and earlier detection of this syndrome while lending potential new insights into the disease process.

Numerous affinity purification-mass spectrometry (AP-MS) and yeast two-hybrid screens have each defined thousands of pairwise protein-protein interactions (PPIs), most of which are between functionally unrelated proteins. The accuracy of these networks, however, is under debate. Here, we present an AP-MS survey of the bacterium Desulfovibrio vulgaris together with a critical reanalysis of nine published bacterial yeast two-hybrid and AP-MS screens. We have identified 459 high confidence PPIs from D. vulgaris and 391 from Escherichia coli. Compared with the nine published interactomes, our two networks are smaller, are much less highly connected, and have significantly lower false discovery rates. In addition, our interactomes are much more enriched in protein pairs that are encoded in the same operon, have similar functions, and are reproducibly detected in other physical interaction assays than the pairs reported in prior studies. Our work establishes more stringent benchmarks for the properties of protein interactomes and suggests that bona fide PPIs much more frequently involve protein partners that are annotated with similar functions or that can be validated in independent assays than earlier studies suggested.

Identifying protein-protein interactions (PPIs) at an acceptable false discovery rate (FDR) is challenging. Previously we identified several hundred PPIs from affinity purification - mass spectrometry (AP-MS) data for the bacteria Escherichia coli and Desulfovibrio vulgaris. These two interactomes have lower FDRs than any of the nine interactomes proposed previously for bacteria and are more enriched in PPIs validated by other data than the nine earlier interactomes. To more thoroughly determine the accuracy of ours or other interactomes and to discover further PPIs de novo, here we present a quantitative tagless method that employs iTRAQ MS to measure the copurification of endogenous proteins through orthogonal chromatography steps. 5273 fractions from a four-step fractionation of a D. vulgaris protein extract were assayed, resulting in the detection of 1242 proteins. Protein partners from our D. vulgaris and E. coli AP-MS interactomes copurify as frequently as pairs belonging to three benchmark data sets of well-characterized PPIs. In contrast, the protein pairs from the nine other bacterial interactomes copurify two- to 20-fold less often. We also identify 200 high confidence D. vulgaris PPIs based on tagless copurification and colocalization in the genome. These PPIs are as strongly validated by other data as our AP-MS interactomes and overlap with our AP-MS interactome for D.vulgaris within 3% of expectation, once FDRs and false negative rates are taken into account. Finally, we reanalyzed data from two quantitative tagless screens of human cell extracts. We estimate that the novel PPIs reported in these studies have an FDR of at least 85% and find that less than 7% of the novel PPIs identified in each screen overlap. Our results establish that a quantitative tagless method can be used to validate and identify PPIs, but that such data must be analyzed carefully to minimize the FDR.

There is an increasing need in biology and clinical medicine to robustly and reliably measure tens to hundreds of peptides and proteins in clinical and biological samples with high sensitivity, specificity, reproducibility, and repeatability. Previously, we demonstrated that LC-MRM-MS with isotope dilution has suitable performance for quantitative measurements of small numbers of relatively abundant proteins in human plasma and that the resulting assays can be transferred across laboratories while maintaining high reproducibility and quantitative precision. Here, we significantly extend that earlier work, demonstrating that 11 laboratories using 14 LC-MS systems can develop, determine analytical figures of merit, and apply highly multiplexed MRM-MS assays targeting 125 peptides derived from 27 cancer-relevant proteins and seven control proteins to precisely and reproducibly measure the analytes in human plasma. To ensure consistent generation of high quality data, we incorporated a system suitability protocol (SSP) into our experimental design. The SSP enabled real-time monitoring of LC-MRM-MS performance during assay development and implementation, facilitating early detection and correction of chromatographic and instrumental problems. Low to sub-nanogram/ml sensitivity for proteins in plasma was achieved by one-step immunoaffinity depletion of 14 abundant plasma proteins prior to analysis. Median intra- and interlaboratory reproducibility was <20%, sufficient for most biological studies and candidate protein biomarker verification. Digestion recovery of peptides was assessed and quantitative accuracy improved using heavy-isotope-labeled versions of the proteins as internal standards. Using the highly multiplexed assay, participating laboratories were able to precisely and reproducibly determine the levels of a series of analytes in blinded samples used to simulate an interlaboratory clinical study of patient samples. Our study further establishes that LC-MRM-MS using stable isotope dilution, with appropriate attention to analytical validation and appropriate quality control measures, enables sensitive, specific, reproducible, and quantitative measurements of proteins and peptides in complex biological matrices such as plasma.

Background: The carbohydrate portions of salivary glycoproteins play important roles, including mediating bacterial and leukocyte adhesion. Salivary glycosylation is complex. Many of its glycoproteins present ABO and Lewis blood group determinants. An individual's genetic complement and secretor status govern the expression of blood group antigens. We queried the extent to which salivary glycosylation varies according to blood group and secretor status. First, we screened submandibular/sublingual and parotid salivas collected as ductal secretions for reactivity with a panel of 16 lectins. We selected three lectins that reacted with the largest number of glycoproteins and one that recognized uncommon lactosamine-containing structures. Ductal salivas representing a secretor with complex blood group expression and a nonsecretor with a simple pattern were separated by SDS-PAGE. Gel slices were trypsin digested and the glycopeptides were individually separated on each of the four lectins. The bound fractions were deN-glycosylated. LC-MS/MS identified the original glycosylation sites, the peptide sequences, and the parent proteins.Results: The results revealed novel salivary N-glycosites and glycoproteins not previously reported. As compared to the secretor, nonsecretor saliva had higher levels of N-glycosylation albeit with simpler structures.Conclusions: Together, the results suggested a molecular basis for inter-individual variations in salivary protein glycosylation with functional implications for oral health.

Endometriosis, ectopic growth of the uterine lining (endometrium), which affects 6-11% of reproductive age women, is associated with pelvic pain and infertility. We investigated the peritoneal fluid (PF), urine and omental fat (OF) proteomes of women with endometriosis vs. individuals with no surgically visualized endometriosis. All participants were enrolled in the NICHD-funded ENDO Study. A two-step proteomic study was performed. The first, a broad survey, employed a semi-quantitative gel LC-mass spectrometry (MS) workflow: SDS PAGE fractionation, trypsin digestion and LC MS/MS. The results showed sample integrity but failed to detect any differences between women with and without endometriosis. The second step was a quantitative analysis of OF samples. We employed another sample set (n = 30) from women +/- disease and isobaric mass-tag (iTRAQ) chemistry to label peptides and 2D LC MS/MS for protein identification and quantification. Three proteins matrix metalloproteinase-9, neutrophil elastase, and FAM49B-were significantly lower in abundance in samples from women with endometriosis. Interestingly, neutrophil elastase and FAM49B levels were associated with higher levels of a subset of endocrine disrupting chemicals (EDCs) that were previously measured in the same samples. The results of these experiments showed the feasibility of associating endometriosis with changes in the OF protein repertoire and EDC levels.Biological significanceEndometriosis, pathological growth of the uterine lining, is associated with significant morbidities, including pain and infertility. However, the causes of this common condition are poorly understood. This study determined whether endometriosis was associated with changes in the protein composition of peritoneal fluid, urine and/or omental fat. A protein of unknown function (FAM49B) and two proteinases (metalloproteinase-9, neutrophil elastase) were down regulated in OF samples from women with versus without endometriosis. These findings suggested proteinase imbalances at sites that were distant from the endometriotic lesions. Additionally, FAM49B and neutrophil elastase levels were associated with higher levels of a subset of environmental chemicals that were quantified in the same samples, suggesting other possible associations. Thus, this work generated hypotheses that will be tested in further studies. (C) 2014 Elsevier B.V. All rights reserved.

ABSTRACT: In March of 1989 the oil tanker Exxon Valdez ran aground and ruptured spilling thousands of barrels of oil into Prince William Sound. That oil spill created a large contingent liability for Exxon. The final adjudication of lawsuits relative to the oil spill did not happen until 2008. Thus, Exxon had a contingent liability relative to the Exxon Valdez incident for nineteen reporting years. You will read about the spill, Exxon’s attempts to contain it, fines, settlements, court cases, damage awards, and the changing nature of the law regarding punitive damages. You will be asked at key points over the nineteen years of the story to comment on the proper reporting and disclosure of the contingent liability. Making these judgments and discussing them in class will deepen your understanding of contingent liabilities and punitive damage awards, will enhance your critical thinking skills, and if you participate in the discussion, enhance your oral communication skills.

This study focuses on the information technology (IT) outsourcing decision and asks whether reported financial statement data can measure short-term financial effects of the IT outsourcing decision and thus add to the literature on the benefits of outsourcing. In this study we used accounting metrics derived from archival financial data to assess the impact of IT outsourcing on firms performance measures. In the sample of 79 firms from 1986 to 2009, there were 45 firms in the manufacturing sector and 34 firms in the service sector. The comparative study between manufacturing and service sectors will help identify where the higher potential of outsourcing impact lies. Firms performance is measured over a two-year period, one year before and one year after outsourcing decisions were made. For performance measures, we used cost efficiency, productivity, profitability, growth, cash management, and market ratio metrics. Using accounting metrics we show that IT outsourcing has a favorable short-term impact on manufacturing firms cost efficiency, productivity, and cash management. At the same time outsourcing the IT function has little favorable impact on service firms short-term performance measures.

Multiple reaction monitoring (MRM) mass spectrometry coupled with stable isotope dilution (SID) and liquid chromatography (LC) is increasingly used in biological and clinical studies for precise and reproducible quantification of peptides and proteins in complex sample matrices. Robust LC-SID-MRM-MS-based assays that can be replicated across laboratories and ultimately in clinical laboratory settings require standardized protocols to demonstrate that the analysis platforms are performing adequately. We developed a system suitability protocol (SSP), which employs a predigested mixture of six proteins, to facilitate performance evaluation of LC-SID-MRM-MS instrument platforms, configured with nanoflow-LC systems interfaced to triple quadrupole mass spectrometers. The SSP was designed for use with low multiplex analyses as well as high multiplex approaches when software-driven scheduling of data acquisition is required. Performance was assessed by monitoring of a range of chromatographic and mass spectrometric metrics including peak width, chromatographic resolution, peak capacity, and the variability in peak area and analyte retention time (RT) stability. The SSP, which was evaluated in 11 laboratories on a total of 15 different instruments, enabled early diagnoses of LC and MS anomalies that indicated suboptimal LC-MRM-MS performance. The observed range in variation of each of the metrics scrutinized serves to define the criteria for optimized LC-SID-MRM-MS platforms for routine use, with pass/fail criteria for system suitability performance measures defined as peak area coefficient of variation <0.15, peak width coefficient of variation <0.15, standard deviation of RT <0.15 min (9 s), and the RT drift <0.5min (30 s). The deleterious effect of a marginally performing LC-SID-MRM-MS system on the limit of quantification (LOQ) in targeted quantitative assays illustrates the use and need for a SSP to establish robust and reliable system performance. Use of a SSP helps to ensure that analyte quantification measurements can be replicated with good precision within and across multiple laboratories and should facilitate more widespread use of MRM-MS technology by the basic biomedical and clinical laboratory research communities.