Molecular phenotyping of laboratory mouse strains using 500 multiple reaction monitoring mass spectrometry plasma assays.

Mouse is the predominant experimental model for the study of human disease due, in part, to phylogenetic relationship, ease of breeding, and the availability of molecular tools for genetic manipulation. Advances in genome-editing methodologies, such as CRISPR-Cas9, enable the rapid production of new transgenic mouse strains, necessitating complementary high-throughput and systematic phenotyping technologies. In contrast to traditional protein phenotyping techniques, multiple reaction monitoring (MRM) mass spectrometry can be highly multiplexed without forgoing specificity or quantitative precision. Here we present MRM assays for the quantitation of 500 proteins and subsequently determine reference concentration values for plasma proteins across five laboratory mouse strains that are typically used in biomedical research, revealing inter-strain and intra-strain phenotypic differences. These 500 MRM assays will have a broad range of research applications including high-throughput phenotypic validation of novel transgenic mice, identification of candidate biomarkers, and general research applications requiring multiplexed and precise protein quantification.

Multiplexed MRM-based assays for the quantitation of proteins in mouse plasma and heart tissue.

The mouse is the most commonly used laboratory animal, with more than 14 million mice being used for research each year in North America alone. The number and diversity of mouse models is increasing rapidly through genetic engineering strategies, but detailed characterization of these models is still challenging because most phenotypic information is derived from time-consuming histological and biochemical analyses. To expand the biochemists' toolkit, we generated a set of targeted proteomic assays for mouse plasma and heart tissue, utilizing bottom-up LC/MRM-MS with isotope-labeled peptides as internal standards. Protein quantitation was performed using reverse standard curves, with LC-MS platform and curve performance evaluated by quality control standards. The assays comprising the final panel (101 peptides for 81 proteins in plasma; 227 peptides for 159 proteins in heart tissue) have been rigorously developed under a fit-for-purpose approach and utilize stable-isotope labeled peptides for every analyte to provide high-quality, precise relative quantitation. In addition, the peptides have been tested to be interference-free and the assay is highly multiplexed, with reproducibly determined protein concentrations spanning >4 orders of magnitude. The developed assays have been used in a small pilot study to demonstrate their application to molecular phenotyping or biomarker discovery/verification studies.

A preliminary mapping of individual, relational, and social factors that impede disclosure of childhood sexual abuse.

Uncovering the pathways to disclosures of child sexual abuse (CSA) and the factors influencing the willingness of victims to talk about the abuse is paramount to the development of powerful practice and policy initiatives. Framed as a long interview method utilizing a grounded theory approach to analyze data, the objective of the current study was to provide a preliminary mapping of the barriers to CSA disclosures through an ecological systemic lens, from a sample of 67 male and female CSA adult survivors, all of whom had recently received counselling services. The current project led to the identification of three broad categories of barriers that were each comprised of several subthemes, namely: Barriers from Within (internalized victim-blaming, mechanisms to protect oneself, and immature development at time of abuse); Barriers in Relation to Others (violence and dysfunction in the family, power dynamics, awareness of the impact of telling, and fragile social network); and Barriers in Relation to the Social World (labelling, taboo of sexuality, lack of services available, and culture or time period). This study points to the importance of using a broad ecological framework to understand the factors that inhibit disclosure of CSA, as barriers to disclosure do not constrain solely the victims. Results are discussed in light of their implications for research, prevention and intervention programs, and social policies and media campaigns, as the burden is on the larger community to create a climate of safety and transparency that makes the telling of CSA possible.

Unsuccessful trial of labor in women with and without previous cesarean delivery.

OBJECTIVE: To compare maternal and neonatal outcomes after unsuccessful labor in women with and those without prior cesarean delivery. METHODS: This was a retrospective cohort study of all women in labor delivered by cesarean section (CS) from November 2004 through December 2006. The study population was dichotomized by previous CS and compared for various maternal and neonatal outcomes. Student t-test, χ² and Fisher exact tests were used for analysis. RESULTS: There was a significantly higher rate of symptomatic uterine rupture [3/100 (3%) vs. 0/449 (0%), p = 0.006], asymptomatic uterine scar dehiscence [6/100 (6%) vs. 0/449 (0%), p = 0.0001], and bladder injury [2/100 (2%) vs. 0/100 (0%), p = 0.001], among women with prior cesarean delivery compared to those without. The rate of respiratory distress syndrome [(6/100) (6%) vs. 10/449 (2.2%), p = 0.05] and meconium aspiration [4/100 (4%) vs. 2/449 (0.4%), p = 0.01] was also significantly higher among neonates of women with prior cesarean delivery. However, the rate of endomyometritis [3/100 (3%) vs. 50/449 (11.1%), p = 0.009] and febrile morbidity [17/100 (17%) vs. 144/449 (32.1%), p  = 0.003] was significantly lower among women with prior cesarean delivery compared to those without prior cesarean birth. CONCLUSIONS: Compared to laboring women without previous cesarean delivery, women with previous cesarean delivery have increased maternal and neonatal morbidity. Febrile morbidity was, however, lower among women with previous cesarean delivery. These differential findings should further inform our perinatal counseling of women contemplating trial of labor after a previous cesarean delivery.

Computed tomographic confirmation of distally placed Essure* microinsert.

OBJECTIVE: To highlight the use of CT scan to confirm proper placement of the Essure microinsert tubal occlusion device. DESIGN: Case report. SETTING: Community-based, outpatient setting. PATIENT(S): A 37-year-old woman (G2P2002) who desired permanent sterilization. INTERVENTION(S): The patient underwent hysteroscopic placement of Essure microinserts, which was complicated by inability to confirm intraluminal placement of the left implant by hysteroscopic visualization. A CT scan performed that day confirmed intraluminal placement and ruled out intraperitoneal placement of the insert. MAIN OUTCOME MEASURE(S): Confirmation of correct insert placement. RESULT(S): A CT scan confirmed the placement of the Essure insert within the fallopian tube lumen and excluded perforation into the peritoneal cavity. CONCLUSION(S): Computed tomography is a reliable way to determine whether Essure microinserts are properly placed when this cannot be determined by hysteroscopic visualization. Follow-up hysterosalpingography continues to be required to confirm tissue ingrowth at 3 months' time.

Abetalipoproteinemia complicating the puerperium.

BACKGROUND: Abetalipoproteinemia is a rare, autosomal recessive disease, in which the absence of beta-lipoprotein results in the malabsorption of fat-soluble vitamins. There are few reported complications from abetalipoproteinemia during pregnancy. We present a case of untreated abetalipoproteinemia complicating the puerperium. CASE: A 23-year-old, gravida 3, para 0020 woman presented to an outside facility in labor, and her delivery was complicated by postpartum hemorrhage and a large vulvar hematoma. She was coagulopathic and transferred for suspected disseminated intravascular coagulation. Her preexisting medical history was not appreciated by the transferring facility. CONCLUSION: Abetalipoproteinemia in pregnancy is rare. Untreated disease conveys multi-system organ dysfunction and has ramifications in labor and delivery. Clinicians must elicit a comprehensive medical history to properly manage complications in the puerperium.

The role of type and token frequency in using past tense morphemes correctly.

Type and token frequency have been thought to be important in the acquisition of past tense morphology, particularly in differentiating regular and irregular forms. In this study we tested the role of frequency in two ways: (1) in bilingual children, who typically use and hear either language less often than monolingual children and (2) cross-linguistically: French and English have different patterns of frequency of regular/irregular verbs. Ten French-English bilingual children, 10 French monolingual and 10 English monolingual children between 4 and 6 years watched a cartoon and re-told the story. The results demonstrated that the bilingual children were less accurate than the monolingual children. Their accuracy in both French and English regular and irregular verbs corresponded to frequency in the input language. These results are consistent with the hypothesis that children learn past tense morphemes by analogy with other words in their vocabularies. We propose a developmental sequence based on conservative generalization across a growing set of verbs.