An inducible CiliaGFP mouse model for in vivo visualization and analysis of cilia in live tissue

BACKGROUND: Cilia are found on nearly every cell type in the mammalian body, and have been historically classified as either motile or immotile. Motile cilia are important for fluid and cellular movement; however, the roles of non-motile or primary cilia in most tissues remain unknown. Several genetic syndromes, called the ciliopathies, are associated with defects in cilia structure or function and have a wide range of clinical presentations. Much of what we know about the formation and maintenance of cilia comes from model systems like C. elegans and Chalmydomonas. Studies of mammalian cilia in live tissues have been hampered by difficulty visualizing them. RESULTS: To facilitate analyses of mammalian cilia function we generated an inducible Cilia(GFP) mouse by targeting mouse cDNA encoding a cilia-localized protein somatostatin receptor 3 fused to GFP (Sstr3::GFP) into the ROSA26 locus. In this system, Sstr3::GFP is expressed from the ubiquitous ROSA26 promoter after Cre mediated deletion of an upstream Neo cassette flanked by lox P sites. Fluorescent cilia labeling was observed in a variety of live tissues and after fixation. Both cell-type specific and temporally regulated cilia labeling were obtained using multiple Cre lines. The analysis of renal cilia in anesthetized live mice demonstrates that cilia commonly lay nearly parallel to the apical surface of the tubule. In contrast, in more deeply anesthetized mice the cilia display a synchronized, repetitive oscillation that ceases upon death, suggesting a relationship to heart beat, blood pressure or glomerular filtration. CONCLUSIONS: The ability to visualize cilia in live samples within the Cilia(GFP) mouse will greatly aid studies of ciliary function. This mouse will be useful for in vivo genetic and pharmacological screens to assess pathways regulating cilia motility, signaling, assembly, trafficking, resorption and length control and to study cilia regulated physiology in relation to ciliopathy phenotypes

Association between sex steroid hormones and subsequent hyperglycemia during pregnancy

ObjectiveSex steroid hormones may play a role in insulin resistance and glucose dysregulation. However, evidence regarding associations between early-pregnancy sex steroid hormones and hyperglycemia during pregnancy is limited. The primary objective of this study was to assess the relationships between first trimester sex steroid hormones and the subsequent development of hyperglycemia during pregnancy; with secondary evaluation of sex steroid hormones levels in mid-late pregnancy, concurrent with and subsequent to diagnosis of gestational diabetes.MethodsRetrospective analysis of a prospective pregnancy cohort study was conducted. Medically low-risk participants with no known major endocrine disorders were recruited in the first trimester of pregnancy (n=319). Sex steroid hormones in each trimester, including total testosterone, free testosterone, estrone, estradiol, and estriol, were assessed using high-performance liquid chromatography and tandem mass spectrometry. Glucose levels of the 1-hour oral glucose tolerance test and gestational diabetes diagnosis were abstracted from medical records. Multivariable linear regression models were fitted to assess the associations of individual first trimester sex steroids and glucose levels.ResultsIn adjusted models, first trimester total testosterone (β=5.24, 95% CI: 0.01, 10.46, p=0.05) and free testosterone (β=5.98, 95% CI: 0.97, 10.98, p=0.02) were positively associated with subsequent glucose concentrations and gestational diabetes diagnosis (total testosterone: OR=3.63, 95% CI: 1.50, 8.78; free testosterone: OR=3.69; 95% CI: 1.56, 8.73). First trimester estrone was also positively associated with gestational diabetes (OR=3.66, 95% CI: 1.56, 8.55). In mid-late pregnancy, pregnant people with gestational diabetes had lower total testosterone levels (β=-0.19, 95% CI: -0.36, -0.02) after adjustment for first trimester total testosterone.ConclusionEarly-pregnancy sex steroid hormones, including total testosterone, free testosterone, and estrone, were positively associated with glucose levels and gestational diabetes in mid-late pregnancy. These hormones may serve as early predictors of gestational diabetes in combination with other risk factors

Identification of unique neoantigen qualities in long-term survivors of pancreatic cancer

Pancreatic ductal adenocarcinoma is a lethal cancer with fewer than 7% of patients surviving past 5 years. T-cell immunity has been linked to the exceptional outcome of the few long-term survivors1,2, yet the relevant antigens remain unknown. Here we use genetic, immunohistochemical and transcriptional immunoprofiling, computational biophysics, and functional assays to identify T-cell antigens in long-term survivors of pancreatic cancer. Using whole-exome sequencing and in silico neoantigen prediction, we found that tumours with both the highest neoantigen number and the most abundant CD8+ T-cell infiltrates, but neither alone, stratified patients with the longest survival. Investigating the specific neoantigen qualities promoting T-cell activation in long-term survivors, we discovered that these individuals were enriched in neoantigen qualities defined by a fitness model, and neoantigens in the tumour antigen MUC16 (also known as CA125). A neoantigen quality fitness model conferring greater immunogenicity to neoantigens with differential presentation and homology to infectious disease-derived peptides identified long-term survivors in two independent datasets, whereas a neoantigen quantity model ascribing greater immunogenicity to increasing neoantigen number alone did not. We detected intratumoural and lasting circulating T-cell reactivity to both high-quality and MUC16 neoantigens in long-term survivors of pancreatic cancer, including clones with specificity to both high-quality neoantigens and predicted cross-reactive microbial epitopes, consistent with neoantigen molecular mimicry. Notably, we observed selective loss of high-quality and MUC16 neoantigenic clones on metastatic progression, suggesting neoantigen immunoediting. Our results identify neoantigens with unique qualities as T-cell targets in pancreatic ductal adenocarcinoma. More broadly, we identify neoantigen quality as a biomarker for immunogenic tumours that may guide the application of immunotherapies

The Influence of Pain Severity and Interference on Satisfaction with Pain Management among Middle-Aged and Older Adults

Background. Health outcomes are often contingent on how effective the individual is able to manage existent illness-related symptoms. This is all the more relevant among chronic pain patients. Objective. This study aimed to identify indicators of pain treatment satisfaction among middle-aged and older adults (N=150) receiving outpatient treatment from a comprehensive cancer center. Methods. Patients were surveyed on questions assessing pain treatment satisfaction, pain severity, and additional social characteristics. Results. Descriptive data showed that middle-aged adults reported more pain locations, greater pain severity, and less satisfaction with pain treatment. A multivariate model was specified, showing older adults being more satisfied with their pain treatment. For the middle-aged adults, treatment satisfaction was generally lower with greater pain severity. This counters that for the older adults, where treatment satisfaction remained consistent despite increased levels of pain severity. Conclusion. These findings address an important issue regarding how pain is experienced across the life course. This suggests that general assumptions cannot be made about the health outcomes of older adults. Beyond the descriptive definitions of pain, there remains the need to develop models that account for determinants that may account for the pain experience among a diverse adult population

Food parenting practices for 5 to 12 year old children: a concept map analysis of parenting and nutrition experts input

Background: Parents are an important influence on children's dietary intake and eating behaviors. However, the lack of a conceptual framework and inconsistent assessment of food parenting practices limits our understanding of which food parenting practices are most influential on children. The aim of this study was to develop a food parenting practice conceptual framework using systematic approaches of literature reviews and expert input. Method: A previously completed systematic review of food parenting practice instruments and a qualitative study of parents informed the development of a food parenting practice item bank consisting of 3632 food parenting practice items. The original item bank was further reduced to 110 key food parenting concepts using binning and winnowing techniques. A panel of 32 experts in parenting and nutrition were invited to sort the food parenting practice concepts into categories that reflected their perceptions of a food parenting practice conceptual framework. Multi-dimensional scaling produced a point map of the sorted concepts and hierarchical cluster analysis identified potential solutions. Subjective modifications were used to identify two potential solutions, with additional feedback from the expert panel requested. Results: The experts came from 8 countries and 25 participated in the sorting and 23 provided additional feedback. A parsimonious and a comprehensive concept map were developed based on the clustering of the food parenting practice constructs. The parsimonious concept map contained 7 constructs, while the comprehensive concept map contained 17 constructs and was informed by a previously published content map for food parenting practices. Most of the experts (52%) preferred the comprehensive concept map, while 35% preferred to present both solutions. Conclusion: The comprehensive food parenting practice conceptual map will provide the basis for developing a calibrated Item Response Modeling (IRM) item bank that can be used with computerized adaptive testing. Such an item bank will allow for more consistency in measuring food parenting practices across studies to better assess the impact of food parenting practices on child outcomes and the effect of interventions that target parents as agents of change