Cancer‐associated somatic DICER1 hotspot mutations cause defective miRNA processing and reverse‐strand expression bias to predominantly mature 3p strands through loss of 5p strand cleavage

Our group recently described recurrent somatic mutations of the miRNA processing gene DICER1 in non‐epithelial ovarian cancer. Mutations appeared to be clustered around each of four critical metal‐binding residues in the RNase IIIB domain of DICER1. This domain is responsible for cleavage of the 3′ end of the 5p miRNA strand of a pre‐mRNA hairpin. To investigate the effects of these cancer‐associated 'hotspot' mutations, we engineered mouse DICER1‐deficient ES cells to express wild‐type and an allelic series of the mutant DICER1 variants. Global miRNA and mRNA profiles from cells carrying the metal‐binding site mutations were compared to each other and to wild‐type DICER1. The miRNA and mRNA profiles generated through the expression of the hotspot mutations were virtually identical, and the DICER1 hotspot mutation‐carrying cells were distinct from both wild‐type and DICER1‐deficient cells. Further, miRNA profiles showed that mutant DICER1 results in a dramatic loss in processing of mature 5p miRNA strands but were still able to create 3p strand miRNAs. Messenger RNA (mRNA) profile changes were consistent with the loss of 5p strand miRNAs and showed enriched expression for predicted targets of the lost 5p‐derived miRNAs. We therefore conclude that cancer‐associated somatic hotspot mutations of DICER1, affecting any one of four metal‐binding residues in the RNase IIIB domain, are functionally equivalent with respect to miRNA processing and are hypomorphic alleles, yielding a global loss in processing of mature 5p strand miRNA. We further propose that this resulting 3p strand bias in mature miRNA expression likely underpins the oncogenic potential of these hotspot mutations. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Influence of oxygen tension on the viscoelastic behavior of red blood cells in sickle cell disease

Although the rheological behavior of sickle cell suspensions and of hemoglobin S solutions is known to be strongly dependent on oxygen tension (PO2), little data exist concerning the influence of PO2 on the viscoelasticity of individual HbSS RBC. We have used micropipette aspiration techniques to test the deformation response of both HbSS and control HbAA RBC over a wide range of PO2 at 23 degrees C. Sickled, spiculed HbSS cells were present for PO2 approximately less than 35 mm Hg; for a number of these cells, the deformation response was essentially elastic and an effective membrane rigidity (EMR) was calculated. EMR increased with decreasing PO2 and was approximately 5 to 50 times higher than the equivalent rigidity of oxygenated HbSS RBC. In addition, the rate of membrane deformation was very slow for sickled cells; the half-time for the deformation process increased as PO2 was lowered and was about two orders of magnitude longer than the equivalent time for normal RBC. Other sickled cells exhibited plastic deformation when subjected to comparable deforming forces and experienced irreversible membrane deformation and budding. At all PO2 levels tested, some HbSS RBC remained as discocytes; these cells had normal membrane elasticity and membrane viscosity. Furthermore, changes in PO2 did not affect the membrane properties of HbAA RBC. Thus, gross abnormalities in the deformation response of HbSS RBC were only detected after morphological sickling had occurred. These abnormalities most likely arose from changes in the cytoplasmic HbS viscoelasticity and, if present in vivo, would be expected to impair the flow of HbSS cells in the microcirculation.     

A C-Peptide-Based Model of Pancreatic Insulin Secretion in Extremely Preterm Neonates in Intensive Care

Background:

Model-based glycemic control relies on sufficiency of underlying models to describe underlying patient physiology. In particular, very preterm infant glucose-insulin metabolism can differ significantly from adults, and is relatively unstudied. In this study, C-peptide concentrations are used to develop insulin-secretion models for the purposes of glycemic control in neonatal intensive care.

Methods:

Plasma C-peptide, insulin, and blood glucose concentrations (BGC) were retrospectively analyzed from a cohort of 41 hyperglycemic very preterm (median age 27.2 [26.2-28.7] weeks) and very low birth-weight infants (median birth weight 839 [735-1000] g). A 2-compartment model of C-peptide kinetics was used to estimate insulin secretion. Insulin secretion was examined with respect to nutritional intake, exogenous and plasma insulin concentration, and BGC.

Results:

Insulin secretion was found to be highly variable between patients and over time, and could not be modeled with respect to age, weight, or protein or dextrose intake. In 13 of 54 samples exogenous insulin was being administered, and insulin secretion was lower. However, low data numbers make this result inconclusive. Insulin secretion was found to increase with BG, with a stronger association in female infants than males (R² = .51 vs R² = .13, and R² = .26 for the combined cohort).

Conclusions:

A sex-based insulin secretion model was created and incorporated into a model-based glycemic control framework. Nutritional intake did not predict insulin secretion, indicating that insulin secretion is a complex function of a number of metabolic factors.     

Do current therapeutic anti-Aβ antibodies for Alzheimer’s disease engage the target?

Reducing amyloid-β peptide (Aβ) burden at the pre-symptomatic stages of Alzheimer’s disease (AD) is currently the advocated clinical strategy for treating this disease. The most developed method for targeting Aβ is the use of monoclonal antibodies including bapineuzumab, solanezumab and crenezumab. We have synthesized these antibodies and used surface plasmon resonance (SPR) and mass spectrometry to characterize and compare the ability of these antibodies to target Aβ in transgenic mouse tissue as well as human AD tissue. SPR analysis showed that the antibodies were able to bind Aβ with high affinity. All of the antibodies were able to bind Aβ in mouse tissue. However, significant differences were observed in human brain tissue. While bapineuzumab was able to capture a variety of N-terminally truncated Aβ species, the Aβ detected using solanezumab was barely above detection limits while crenezumab did not detect any Aβ. None of the antibodies were able to detect any Aβ species in human blood. Immunoprecipitation experiments using plasma from AD subjects showed that both solanezumab and crenezumab have extensive cross-reactivity with non-Aβ related proteins. Bapineuzumab demonstrated target engagement with brain Aβ, consistent with published clinical data. Solanezumab and crenezumab did not, most likely as a result of a lack of specificity due to cross-reactivity with other proteins containing epitope overlap. This lack of target engagement raises questions as to whether solanezumab and crenezumab are suitable drug candidates for the preventative clinical trials for AD.     

Interpersonal distress is associated with sleep and arousal in insomnia and good sleepers

Objective

The interpersonal environment is strongly linked to sleep. However, little is known about interpersonal distress and its association with sleep. We examined the associations among interpersonal distress, objective and subjective sleep in people with and without insomnia.

Methods

Participants in this cross-sectional observational study included men and women with insomnia (n = 28) and good sleeper controls (n = 38). Interpersonal distress was measured with the Inventory of Interpersonal Problems. Sleep parameters included insomnia severity, self-reported presleep arousal, and sleep quality; and polysomnographically-assessed sleep latency (SL), total sleep time (TST), wake after sleep onset (WASO), percent delta (stage 3 + 4 NREM), percent REM, and EEG beta power. Hierarchical linear regression was used to assess the relationship between distress from interpersonal problems and sleep and the extent to which relationships differed among insomnia patients and controls.

Results

More interpersonal distress was associated with more self-reported arousal and higher percentage of REM. More interpersonal distress was associated with greater insomnia severity and more cognitive presleep arousal for individuals with insomnia, but not for controls. Contrary to expectations, interpersonal distress was associated with shorter sleep latency in the insomnia group. Results were attenuated, but still significant, after adjusting for depression symptoms.

Conclusion

Distress from interpersonal problems is associated with greater self-reported arousal and higher percent REM. Individuals with insomnia who report more distress from interpersonal problems have greater insomnia severity and cognitive presleep arousal, perhaps due to rumination. These findings extend our knowledge of the association between interpersonal stressors and sleep. Assessment and consideration of interpersonal distress could provide a novel target for insomnia treatment.     

Improvement and Recovery from Eating Disorders: A Patient Perspective

This study presents a patient perspective on improvement and recovery from eating disorders. Forty-eight women recruited from patient organizations and a university eating disorder unit participated in the study. Participants completed standard questionnaires and a qualitative interview. "A wish to change," professional treatment, nonprofessional care, and important persons in the women's lives were identified as important recovery factors. Recovery included improved acceptance of oneself, interpersonal relations, problem solving, and body satisfaction, which was not entirely dependent on symptom absence. The patient perspective concurred with clinical reasoning about recovery.     

Environmental Factors Predictive of No-Show Visits in Radiology: Observations of Three Million Outpatient Imaging Visits Over 16 Years

PurposeTo evaluate the impact of environmental and socioeconomic factors on outpatient cancellations and “no-show visits” (NSVs) in radiology.Materials and MethodsWe conducted a retrospective analysis by collecting environmental factor data related to outpatient radiology visits occurring between 2000 and 2015 at our multihospital academic institution. Appointment attendance records were joined with daily weather observations from the National Oceanic and Atmospheric Administration and estimated median income from the US Census American Community Survey. A multivariate logistic regression model was built to examine relationships between NSV rate and median income, commute distance, maximum daily temperature, and daily snowfall.ResultsThere were 270,574 (8.0%) cancellations and 87,407 (2.6%) NSVs among 3,379,947 scheduled outpatient radiology appointments and 575,206 unique patients from 2000 to 2015. Overall cancellation rates decreased from 14% to 8%, and NSV rates decreased from 6% to 1% as median income increased from $20,000 to $120,000 per year. In a multivariate model, the odds of NSV decreased 10.7% per $10,000 increase in median income (95% confidence interval [CI]: 10.3%-11.1%) and 2.0% per 10°F increase in maximum daily temperature (95% CI: 1.3%-1.6%). The odds of NSV increased 1.4% per 10-mile increase in commute distance (95% CI: 1.3%-1.6%) and 4.5% per 1-inch increase in daily snowfall (95% CI: 3.6%-5.3%). Commute distance was more strongly associated with NSV for those in the two lower tertiles of income than the highest tertile (P < .001).ConclusionEnvironmental factors are strongly associated with patients’ attendance at scheduled outpatient radiology examinations. Modeling of appointment failure risk based on environmental features can help increase the attendance of outpatient radiology appointments.