Analysis of the extreme diversity of salivary alpha-amylase isoforms generated by physiological proteolysis using liquid chromatography-tandem mass spectrometry

Saliva is a crucial biofluid for oral health and is also of increasing importance as a non-invasive source of disease biomarkers. Salivary alpha-amylase is an abundant protein in saliva, and changes in amylase expression have been previously associated with a variety of diseases and conditions. Salivary alpha-amylase is subject to a high diversity of post-translational modifications, including physiological proteolysis in the oral cavity. Here we developed methodology for rapid sample preparation and non-targeted LC-ESI-MS/MS analysis of saliva from healthy subjects and observed an extreme diversity of alpha-amylase proteolytic isoforms. Our results emphasize the importance of consideration of post-translational events such as proteolysis in proteomic studies, biomarker discovery and validation, particularly in saliva. (C) 2012 Elsevier B.V. All rights reserved.

Collection and determination of nucleotide metabolites in neonatal and adult saliva by high performance liquid chromatography with tandem mass spectrometry

Saliva contains a number of biochemical components which may be useful for diagnosis/monitoring of metabolic disorders, and as markers of cancer or heart disease. Saliva collection is attractive as a non-invasive sampling method for infants and elderly patients. We present a method suitable for saliva collection from neonates. We have applied this technique for the determination of salivary nucleotide metabolites. Saliva was collected from 10 healthy neonates using washed cotton swabs, and directly from 10 adults. Two methods for saliva extraction from oral swabs were evaluated. The analytes were then separated using high performance liquid chromatography (HPLC) with tandem mass spectrometry (MS/MS). The limits of detection for 14 purine/pyrimidine metabolites were variable, ranging from 0.01 to 1.0 mu M. Recovery of hydrophobic purine/pyrimidine metabolites from cotton tips was consistently high using water/acetonitrile extraction (92.7-111%) compared with water extraction alone. The concentrations of these metabolites were significantly higher in neonatal saliva than in adults. Preliminary ranges for nucleotide metabolites in neonatal and adult saliva are reported. Hypoxanthine and xanthine were grossly raised in neonates (49.3 +/- 25.4; 30.9 +/- 19.5 mu M respectively) compared to adults (4.3 +/- 3.3; 4.6 +/- 4.5 mu M); nucleosides were also markedly raised in neonates. This study focuses on three essential details: contamination of oral swabs during manufacturing and how to overcome this; weighing swabs to accurately measure small saliva volumes; and methods for extracting saliva metabolites of interest from cotton swabs. A method is described for determining nucleotide metabolites using HPLC with photo-diode array or MS/MS. The advantages of utilising saliva are highlighted. Nucleotide metabolites were not simply in equilibrium with plasma, but may be actively secreted into saliva, and this process is more active in neonates than adults. (C) 2013 Elsevier B.V. All rights reserved.

Identification of salivary N-glycoproteins and measurement of glycosylation site occupancy by boronate glycoprotein enrichment and liquid chromatography/electrospray ionization tandem mass spectrometry

RATIONALE Diseases including cancer and congenital disorders of glycosylation have been associated with changes in the site-specific extent of protein glycosylation. Saliva can be non-invasively sampled and is rich in glycoproteins, giving it the potential to be a useful biofluid for the discovery and detection of disease biomarkers associated with changes in glycosylation. METHODS Saliva was collected from healthy individuals and glycoproteins were enriched using phenylboronic acid based glycoprotein enrichment resin. Proteins were deglycosylated with peptide-N-glycosidase F and digested with AspN or trypsin. Desalted peptides and deglycosylated peptides were separated by reversed-phase liquid chromatography and detected with on-line electrospray ionization quadrupole-time-of-flight mass spectrometry using a 5600 TripleTof instrument. Site-specific glycosylation occupancy was semi-quantitatively determined from the abundance of deglycosylated and nonglycosylated versions of each given peptide. RESULTS Glycoprotein enrichment identified 67 independent glycosylation sites from 24 unique proteins, a 3.9-fold increase in the number of glycosylation sites identified. Enrichment of glycoproteins rather than glycopeptides allowed detection of both deglycosylated and nonglycosylated versions of each peptide, and thereby robust measurement of site-specific occupancy at 21 asparagines. Healthy individuals showed limited biological variability in occupancy, with partially modified sites having characteristics consistent with inefficient glycosylation by oligosaccharyltransferase. Inclusion of negative controls without enzymatic deglycosylation controlled for spontaneous chemical deamidation, and identified asparagines previously incorrectly annotated as glycosylated. CONCLUSIONS We developed a sample preparation and mass spectrometry detection strategy for rapid and efficient measurement of site-specific glycosylation occupancy on diverse salivary glycoproteins suitable for biomarker discovery and detection of changes in glycosylation occupancy in human disease.

Work and family : the perception of balance among female teachers in northern Malaysia

Women’s participation in paid employment has become a common scenario even in non-western developing countries. For example in Malaysia, the trend is growing although the traditional gender role remains strong in Malaysian society. Even though working, women are still expected to assume major responsibilities at home. Thus, as opposed to men, women in this society face the challenge to satisfactorily balance work and family. This study was carried out to explore how Malaysian women perceive the meaning of a balanced work-family life. Sampling women teachers, the interview findings revealed that work-family balance was mainly perceived in terms of an individual’s ‘ability to fulfill role obligation’ appropriately in both the work and family domains. A few participants also viewed balance in the context of role satisfaction and role interference. Overall, the results support the assumption in the literature that perceptions of work-family experience are not universal, rather, the construct of work-family balance is culture-specific.

Commercial friendships between gay salesmen and straight female customers

Perhaps it is now sacrosanct in marketing to contemplate that many service encounters, especially those in retail settings, are social encounters in which bonds between and among customers and employees are critical drivers of consumption (Beatty et al., 1996; Rosenbaum, 2006). Indeed, within retail settings, it is often possible for salespeople and customers to form so-called “commercial friendships” (Price and Arnould, 1999). These friendships result in both salespeople and their customers having social interactions that are close to those experienced in personal friendships (Swan et al., 2001), and which are extremely satisfying for all parties. Outside of marketing, the social science literature (Grigoriou, 2004; Rumens, 2008; Russell, DelPriore, Butterfield, and Hill, 2013) and popular press (de la Cruz and Dolby, 2007; Hopcke and Rafaty, 1999; Tilmann-Healy, 2001, Whitney, 1990) is replete with knowledge regarding the “absolutely fabulous” friendships (Hopcke and Rafaty, 1999) that often form between gay men and straight women. In fact, Western culture regularly highlights the compatibility of gay men and straight women in film, television, and writing, to the extent that they have now influenced popular thinking on the topic, so that gay men and straight females are viewed as sharing common plights and interests (Rumens, 2008). Yet, thus far, marketing researchers have looked askance at the effect of friendships between gay male employees and heterosexual female customers in consumption settings, such as retail stores and boutiques. Indeed, with the exception of Peretz’s (1995) participant observation regarding how young and outwardly gay salesmen use their ambiguous gender to sell women’s clothing, in a Paris-based luxury boutique, any theoretical explorations regarding retail-based commercial friendships between gay salesmen and female customers are non-existent—until now. This research addresses this apparent chasm in the literature by putting forth an original framework that shows how the emotional closeness between gay salesmen and female customers, due to the absence of sexual interest and inter-female competition, results in an intense emotional closeness, that facilitates pleasurable retail transactions, customer satisfaction, loyalty, and positive word-of-mouth. In doing so, this work extends the commercial friendship paradigm by considering retail-based, commercial friendships between an under-researched marketplace dyad; gay men and straight females. It is worth noting here that some straight women may find the idea of commercial friendships with gay salesmen as undesirable, due to the very notion of having relationships with retail organizations or employees (Noble and Phillips, 2004), or a personal disdain for homosexuality.

A Labour of Sex? Male and Female Prostitution

Perspectives in Human Sexuality is an overview of key debates, research findings and theories in the area of sex and sexuality. This textbook written especially for undergraduate students offers a detailed and comprehensive introduction to sex and sexuality from an Australian and New Zealand perspective. It examines controversial issues such as sex and age, sex work and gay, lesbian and queer sex in a fair and balanced manner.