Pregnancy influences the selection of appropriate reference genes in mouse tissue: Determination of appropriate reference genes for quantitative reverse transcription PCR studies in tissues from the female mouse reproductive axis.

Selecting stably expressed reference genes which are not affected by physiological or pathophysiological conditions is crucial for reliable quantification in gene expression studies. This study examined the expression stability of a panel of twelve reference genes in tissues from the female mouse reproductive axis and the uterus. Gene expression studies were carried out using reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). cDNA was synthesised from RNA extracted from hypothalami, pituitaries, ovaries and uteri of female mice at ages representing weaning, puberty and adulthood as well as pregnancy (13 ± 1 days post-coitus) (n = a minimum of 3 at each age and at pregnancy). The reference genes examined included 18 s, Actb, Atp5b, B2m, Canx, Cyc1, Eif4a2, Gapdh, Rpl13a, Sdha, Ubc and Ywhaz. The RT-qPCR raw data were imported into the qBASE+ software to analyse the expression stability using GeNorm. These data were also subsequently analysed using other software packages (Delta CT, Normfinder, BestKeeper). A comprehensive ranking was conducted considering all stability rankings generated from the different software analyses. B2m and Eif4a2 deviated from the acceptable range for amplification efficiency and therefore were excluded from the further analyses. The stability of the reference genes is influenced by the software used for the analysis with BestKeeper providing markedly different results than the other analyses. GeNorm analysis of tissues taken at different ages but not including pregnant animals, indicated that the expression of the reference genes is tissue specific with the most stable genes being: in the hypothalamus, Canx and Actb; in the pituitary, Sdha and Cyc1; in the ovary, 18s, Sdha and Ubc; and in the uterus, Ywhaz, Cyc1, Atp5b, 18s and Rpl13a. The optimal number of reference genes to be used was determined to be 2 in the first three tissues while in the uterus, the V-score generated by the GeNorm analysis was higher than 0.15 suggesting that 3 or more genes should be used for normalisation. Inclusion of tissues from pregnant mice changed the reference genes identified as being the most stable: Ubc and Sdha were the most stable genes in the hypothalamus, pituitary and the ovary. The addition of pregnant tissue had no effect on the stability of the genes in uterus (Ywhaz, Cyc1, Atp5b, 18s and Rpl13a). Identification of these stable reference genes will be of use to those interested in studying female fertility and researchers should be alert to the effects of pregnancy on reference gene stability. This study also signifies the importance of re-examining reference gene stability if the experimental conditions are changed, as shown with the introduction of pregnancy as a new factor in this research. [Abstract copyright: Crown Copyright © 2021. Published by Elsevier B.V. All rights reserved.

Introducing open book examinations in clinical education: A case study

During the recent COVID-19 pandemic, in common with educators across the Higher Education sector, the School of Dentistry at the University of Liverpool reimagined the learning and assessment strategy by moving from proctored closed-book assessment to largely unmonitored open-book examinations (OBE). This article discusses understandings from an educator perspective following our implementation of OBE. The educator perspective discussed here indicates that OBE have the potential to be an authentic and acceptable form of assessment, but that some reframing of attitudes towards assessment from all stakeholders and their approaches to assessment is necessary when developing these innovative types of assessment

Use of Accelerometers in a Large Field-Based Study of Children: Protocols, Design Issues, and Effects on Precision

Background: Objective methods can improve accuracy of physical activity measurement in field studies but uncertainties remain about their use. Methods: Children age 11 years from the Avon Longitudinal Study of Parents and Children (ALSPAC), were asked to wear a uni-axial accelerometer (MTI Actigraph) for 7 days. Results: Of 7159 children who attended for assessment, 5595 (78%) provided valid measures. The reliability coefficient for 3 days of recording was .7 and the power to detect a difference of 0.07 SDs (P ≤ .05) was \u3e 90%. Measures tended to be higher on the first day of recording (17 counts/min; 95% CI, 10-24) and if children wore the monitor for fewer days, but these differences were small. The children who provided valid measures of activity were different from those who did not, but the differences were modest. Conclusion: Objective measures of physical activity can be incorporated into large longitudinal studies of children

Relationship Between Sexual Activity, Contraceptive Utilization and Biopsychosocial Characteristics Among Homeless Shelter Adolescents.

Objective: To determine whether biopsychosocial factors are associated with sexual activity and contraceptive utilization among homeless shelter adolescents. Methods: A retrospective study of 440 adolescents at a shelter in Pennsylvania between February 2015 and September 2019 was conducted. The cohort was evaluated to determine what relationship age, gender identity, substance use, and trauma history have with sexual activity and contraceptive utilization. Results: Sexual activity was significantly related to age (mean 15.8+1.4 years in sexually active vs. 14.7+1.6 years in abstinent youth, p\u3c0.001); remote history of self-harm behavior (relative risk ratio (RR) 1.23 [95% CI 1.03-1.46]; p=0.02), history of aggressive behavior (RR 1.21 [95% CI 1.01-1.46]; p=0.04), history of trauma (RR 1.24 [95% CI 1.04-1.48]; p=0.03), and substance use (RR 2.27 [95%CI 1.86-2.77]; p\u3c0.001). There were 55.7% sexually active females vs. 42.50% males reporting contraception use (p=0.01). After adjustment, older age and substance use remained significantly associated with sexual activity (adjusted odds ratio (AOR) 1.58 [95% CI 1.36-1.83]; p\u3c0.001 and AOR 5.18 [95% CI 3.28-8.18]; p\u3c0.001, respectively). Conclusions: Females self-reported sexual activity using contraception more than males. After adjustment, older age and substance use were associated with sexual activity. By better understanding the impact these factors can have on contraceptive utilization, informed policy and practice interventions can be developed and implemented to help increase safe sex practices in spaces where homeless adolescents access healthcare

Risk of Cardiovascular Events and Death—Does Insurance Matter?

BACKGROUND: Many Americans lack health insurance. Despite good evidence that lack of insurance compromises access to care, few prospective studies examine its relationship to health outcomes. OBJECTIVE: To determine the relationship between insurance and cardiovascular outcomes and the relationship between insurance and selected process measures. DESIGN AND PARTICIPANTS: We used data from 15,792 participants in the Atherosclerosis Risk in Communities Study, a prospective cohort study. Participants were enrolled in 1987–1989 and returned for follow-up visits every 3 years, for a total of 4 visits. MAIN OUTCOME MEASURES: We estimated the hazard of myocardial infarction, stroke, and death associated with insurance status using Cox proportional hazard modeling. We used generalized estimating equations to examine the association between insurance status and risk of (1) reporting no routine physical examinations, (2) being unaware of a personal cardiovascular risk condition, and (3) inadequate control of cardiovascular risk conditions. RESULTS: Persons without insurance had higher rates of stroke (adjusted hazard ratio, 95% CI 1.22–2.22) and death (adjusted hazard ratio 1.26, 95% CI 1.03–1.53), but not myocardial infarction, than those who were insured. The uninsured were less likely to report routine physical examinations (adjusted risk ratio 1.13, 95% CI 1.08–1.18); more likely to be unaware of hypertension (adjusted risk ratio 1.12, 95% CI 1.00–1.25) and hyperlipidemia (adjusted risk ratio 1.11, 95% CI 1.03–1.19); and more likely to have poor blood pressure control (adjusted risk ratio 1.23, 95% CI 1.08–1.39). CONCLUSIONS: Lack of health insurance is associated with increased rates of stroke and death and with less awareness and control of cardiovascular risk conditions. Health insurance may improve cardiovascular risk factor awareness, control and outcomes

Bryozoan genera Fenestrulina and Microporella no longer confamilial; multi-gene phylogeny supports separation

Bryozoans are a moderately diverse, mostly marine phylum with a fossil record extending to the early Ordovician. Compared to other phyla, little is known about their phylogenetic relationships at both lower and higher taxonomic levels. Hence, an effort is being made to elucidate the phylogenetic relationships among bryozoans. Here, we present newly sequenced nuclear and mitochondrial genes for 21 cheilostome bryozoans and compile these with existing orthologous molecular data. Using these data, we focus on reconstructing the phylogenetic relationships of Fenestrulina and Microporella, two species-rich genera. They are currently placed in a globally distributed family, Microporellidae, defined by having a semicircular primary orifice and a proximal ascopore, although there are indirect inferences in the morphological literature that suggest they might not be confamilial. Our six-gene phylogenetic analysis reveals that the genera Fenestrulina and Microporella are each monophyletic, with the sister clade to Microporella comprising non-microporellids. These genera thus have a polyphyletic relationship and should not be placed in the same family. Our result supports the reinstatement of the family Fenestrulinidae Jullien, 1888 for Fenestrulina and genera with comparable frontal shield and ooecial morphologies. Our well-supported phylogeny based on independent molecular data lends credit to existing phylogenetic hypotheses based on morphological observations but does not conform to the current classification of these particular bryozoans. This illustrates the general need for a rethink of bryozoan higher-level systematics, ideally based on both morphological and molecular data

Epidemiological Interactions between Urogenital and Intestinal Human Schistosomiasis in the Context of Praziquantel Treatment across Three West African Countries

© 2015 Knowles et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. The attached file is the published version of the article

Ethanol reversal of tolerance to the respiratory depressant effects of morphine

Opioids are the most common drugs associated with unintentional drug overdose. Death results from respiratory depression. Prolonged use of opioids results in the development of tolerance but the degree of tolerance is thought to vary between different effects of the drugs. Many opioid addicts regularly consume alcohol (ethanol), and post-mortem analyses of opioid overdose deaths have revealed an inverse correlation between blood morphine and ethanol levels. In the present study, we determined whether ethanol reduced tolerance to the respiratory depressant effects of opioids. Mice were treated with opioids (morphine, methadone, or buprenorphine) for up to 6 days. Respiration was measured in freely moving animals breathing 5% CO(2) in air in plethysmograph chambers. Antinociception (analgesia) was measured as the latency to remove the tail from a thermal stimulus. Opioid tolerance was assessed by measuring the response to a challenge dose of morphine (10 mg/kg i.p.). Tolerance developed to the respiratory depressant effect of morphine but at a slower rate than tolerance to its antinociceptive effect. A low dose of ethanol (0.3 mg/kg) alone did not depress respiration but in prolonged morphine-treated animals respiratory depression was observed when ethanol was co-administered with the morphine challenge. Ethanol did not alter the brain levels of morphine. In contrast, in methadone- or buprenorphine-treated animals no respiratory depression was observed when ethanol was co-administered along with the morphine challenge. As heroin is converted to morphine in man, selective reversal of morphine tolerance by ethanol may be a contributory factor in heroin overdose deaths