A molecular switch for neuroprotective astrocyte reactivity.

The intrinsic mechanisms that regulate neurotoxic versus neuroprotective astrocyte phenotypes and their effects on central nervous system degeneration and repair remain poorly understood. Here we show that injured white matter astrocytes differentiate into two distinct C3-positive and C3-negative reactive populations, previously simplified as neurotoxic (A1) and neuroprotective (A2)1,2, which can be further subdivided into unique subpopulations defined by proliferation and differential gene expression signatures. We find the balance of neurotoxic versus neuroprotective astrocytes is regulated by discrete pools of compartmented cyclic adenosine monophosphate derived from soluble adenylyl cyclase and show that proliferating neuroprotective astrocytes inhibit microglial activation and downstream neurotoxic astrocyte differentiation to promote retinal ganglion cell survival. Finally, we report a new, therapeutically tractable viral vector to specifically target optic nerve head astrocytes and show that raising nuclear or depleting cytoplasmic cyclic AMP in reactive astrocytes inhibits deleterious microglial or macrophage cell activation and promotes retinal ganglion cell survival after optic nerve injury. Thus, soluble adenylyl cyclase and compartmented, nuclear- and cytoplasmic-localized cyclic adenosine monophosphate in reactive astrocytes act as a molecular switch for neuroprotective astrocyte reactivity that can be targeted to inhibit microglial activation and neurotoxic astrocyte differentiation to therapeutic effect. These data expand on and define new reactive astrocyte subtypes and represent a step towards the development of gliotherapeutics for the treatment of glaucoma and other optic neuropathies.

Exploring the effect of the 'Growing Together' parenting education kit on early parenting - study protocol for a cluster randomised controlled trial.

BACKGROUND: Significant gaps exist in education for prospective and new parents, especially for some of the most vulnerable families. Prospective parents would like more information during pregnancy to prepare them for parenting, and need access to trusted and quality information. The Royal Women's Hospital (the Women's) in Melbourne, Australia, a large tertiary referral maternity hospital, developed a parenting education kit known as 'Growing Together'. The kit, designed to guide prospective and new parents from conception until 1 year after birth, includes three components: an A4 sized book, a specifically designed 'App' and a children's story book. We aim to evaluate the impact of the kit on a range of outcomes. METHODS: A two-arm cluster randomised controlled trial will be used. Antenatal clinic days will be randomised to either the intervention or standard care arms. Women in the intervention arm receive the kit at their antenatal booking visit. Women in the standard care arm receive the standard information resources at the Women's. Analyses will be by intention to treat. INCLUSION CRITERIA: primiparous women with adequate English-speaking ability and ≤ 30 weeks' gestation at first pregnancy booking appointment. The primary outcome of the study is the 'experience of motherhood questionnaire' (EMQ), a 20 item validated self-report measure, ranging from 0 to 80, with lower scores indicating better maternal health and wellbeing. To detect a 10% difference in new mothers scoring ≤40 between women who have received the kit (60%) and those who have not (50%), would require 408 per group (total of 816 women) with 95% confidence and 80% power. Allowing for loss to follow up, we aim to recruit 1000 mothers. Secondary outcomes include parents' views and experiences of their care and of the kit during pregnancy and after the birth, parental attachment, knowledge, confidence, wellbeing and health-seeking behaviour; and emotional, developmental and physical health of the infant. Survey data will be collected from mothers at 2, 6 and 12 months postpartum and partners at 6 months. DISCUSSION: This study will provide much needed high-level evidence on the impact of a comprehensive education resource for new parents. TRIAL REGISTRATION: ANZCTRN12615000270516 - Retrospectively registered (23/03/2015); trial started on 16 March 2015.

Global Transcriptional Response to CRISPR/Cas9-AAV6-Based Genome Editing in CD34+ Hematopoietic Stem and Progenitor Cells.

Genome-editing technologies are currently being translated to the clinic. However, cellular effects of the editing machinery have yet to be fully elucidated. Here, we performed global microarray-based gene expression measurements on human CD34+ hematopoietic stem and progenitor cells that underwent editing. We probed effects of the entire editing process as well as each component individually, including electroporation, Cas9 (mRNA or protein) with chemically modified sgRNA, and AAV6 transduction. We identified differentially expressed genes relative to control treatments, which displayed enrichment for particular biological processes. All editing machinery components elicited immune, stress, and apoptotic responses. Cas9 mRNA invoked the greatest amount of transcriptional change, eliciting a distinct viral response and global transcriptional downregulation, particularly of metabolic and cell cycle processes. Electroporation also induced significant transcriptional change, with notable downregulation of metabolic processes. Surprisingly, AAV6 evoked no detectable viral response. We also found Cas9/sgRNA ribonucleoprotein treatment to be well tolerated, in spite of eliciting a DNA damage signature. Overall, this data establishes a benchmark for cellular tolerance of CRISPR/Cas9-AAV6-based genome editing, ensuring that the clinical protocol is as safe and efficient as possible.

Improving influenza vaccination coverage in pregnancy in Melbourne 2010-2011.

BACKGROUND: Seasonal influenza vaccination during pregnancy is effective in preventing serious maternal and infant respiratory illness, but published Australian audits are sparse concerning practice. AIMS: Guided by information gained from new mothers about obstacles to influenza vaccination in July 2010, an educational program for maternity staff and pregnant women was developed with the aim of improving vaccine uptake in 2011. METHODS: One hundred and ninety-nine new mothers in July 2010 and 240 new mothers in July 2011 on postnatal wards of a tertiary obstetric hospital were interviewed about whether influenza vaccine had been offered or given and about reasons why women chose for or against vaccination during pregnancy. RESULTS: Influenza vaccination increased from 30% in 2010 to 40% in 2011 (P = 0.03). Fewer women cited safety concerns for themselves or their babies after the 2011 educational campaign. Comments from 90 women in 2011 give grounds for speculation that the vaccination rate could have been as high as 78% if influenza vaccination had been integrated as part of hospital-based antenatal care. CONCLUSIONS: Influenza vaccine coverage during pregnancy is increasing but obstacles persist, especially absent or inconsistent advice from healthcare workers and lack of ready access during routine antenatal care.

Diagnostic accuracy of maternal serum macrophage inhibitory cytokine-1 and pregnancy-associated plasma protein-A at 6-10 weeks of gestation to predict miscarriage.

OBJECTIVE: To determine whether serum macrophage inhibitory cytokine-1, pregnancy-associated plasma protein-A (PAPP-A), anandamide, or ß-human chorionic gonadotropin (hCG) measured in an asymptomatic population in the middle of the first trimester with a viable fetus predicts subsequent miscarriage. METHODS: We undertook a prospective cohort study at Mercy Hospital for Women between 2004 and 2008. Participants (N=782) were recruited from prenatal clinics, where samples were taken from asymptomatic women at 6 0/7 to 10 6/7 weeks of gestation. We collected samples from only those women for whom we were able to obtain ultrasound evidence of a singleton with fetal cardiac activity. Serum macrophage inhibitory cytokine-1, PAPP-A, anandamide, and ß-hCG concentrations were assayed. RESULTS: Twenty-one (2.7%) miscarried and 761 did not. Among those who miscarried, macrophage inhibitory cytokine-1 and PAPP-A were significantly decreased at 63% (multiples of the median (MOM) 0.63, 25th-75th percentiles 0.33-0.88) and 23% (MOM 0.23, 25th-75th percentiles 0.12-0.48) of levels seen among those with ongoing pregnancies (P<.001 for both comparisons). In contrast, neither serum ß-hCG (MOM 0.99, 25th-75th percentiles 0.46-1.86) nor anandamide (MOM 1.07, 25th-75th percentiles 0.87-1.19) was elevated or decreased among those who miscarried compared with those with ongoing pregnancies. At a fixed 10% false-positive rate (90% specificity), a test combining macrophage inhibitory cytokine-1 and PAPP-A yielded 63% sensitivity and a 6.6 positive likelihood ratio in predicting miscarriage. CONCLUSION: Low serum levels of macrophage inhibitory cytokine-1 and PAPP-A measured from asymptomatic women at 6-10 weeks of gestation with viable pregnancies can predict subsequent miscarriage. These analytes are likely to have an important biological role in early pregnancy and are likely to be useful clinical biomarkers for miscarriage and other early pregnancy complications. LEVEL OF EVIDENCE: II.

Derivational morphophonology: exploring errors in third graders' productions.

PURPOSE: This study describes a post hoc analysis of segmental, stress, and syllabification errors in third graders' productions of derived English words with the stress-changing suffixes -ity and -ic. We investigated whether (a) derived word frequency influences error patterns, (b) stress and syllabification errors always co-occur, and (c) derived word stress pattern affects errors. METHOD: A total of 1,900 productions from 81 third-grade children were transcribed and coded. The targets were 8 high-frequency (HF) and 8 low-frequency (LF) real English words and 8 nonsense (NS) derived words. Participants combined a suffix and a base word to produce a derived word. The vowel-initial suffixes required both stress and syllabic changes in a base word. RESULTS: Children made more segmental and stress errors on NS words than on HF and LF words, but more syllabification errors on LF words than on HF and NS words. More items among the LF words required vowel alternations than among the HF words. When syllabification was not mastered, stress was usually incorrect; however, when syllabification was correct, children often still erred on stress. Derived words that contained 2 trochaic feet were produced with fewer errors than were those that only had 1 trochaic foot. CONCLUSION: Rather than a frequency effect, a lexical effect emerged such that real words were more similar to each other for segmental and stress errors than they were to NS words. Three findings suggest that prosody plays a role in errors: Children made more suprasegmental than segmental errors, they appeared to master syllabification before stress placement, and they produced fewer errors on derived words with 2 trochaic feet than on those with only 1 trochaic foot. Although these results are preliminary, they highlight the role of prosody in morphophonology.

Third graders' metalinguistic skills, reading skills, and stress production in derived English words.

PURPOSE: This study examined relationships between 3rd graders' metalinguistic skills (phonological and morphological awareness), reading skills (decoding and word identification), and accurate stress production in derived words with stress-changing suffixes. METHOD: Seventy-six typically developing 3rd-grade children (M=8;8[years;months]) participated in a battery of tests measuring general oral language ability, phonological and morphological awareness skills, reading skills, and derived word production. RESULTS: Significant positive correlations between stress accuracy in derived words and all other measures were found. Two multiple regressions were run, one with stress accuracy as the outcome variable and the other with decoding as the outcome variable. Metalinguistic and decoding skills independently accounted for a significant proportion of the variance in derived word stress production beyond that accounted for by age and general oral language ability. When decoding was the outcome variable, accurate stress production explained a significant amount of variance (11%) after phonological and morphological awareness were controlled. CONCLUSION: The relationship between accurate stress production and decoding appears to be strong and bidirectional. Possibly, the stress accuracy measure taps into another level of phonological awareness (i.e., morphophonological awareness), which develops later than typical segmental measures of phonological awareness.