Benchmarking nanopore sequencing and rapid genomics feasibility: validation at a quaternary hospital in New Zealand.

Approximately 200 critically ill infants and children in New Zealand are in high-dependency care, many suspected of having genetic conditions, requiring scalable genomic testing. We adopted an acute care genomics protocol from an accredited laboratory and established a clinical pipeline using Oxford Nanopore Technologies PromethION 2 solo system and Fabric GEM™ software. Benchmarking of the pipeline was performed using Global Alliance for Genomics and Health benchmarking tools and Genome in a Bottle samples (HG002-HG007).

Effect of maternal periconceptional undernutrition in sheep on cortisol regulation in offspring from mid-late gestation, through to adulthood.

Introduction: Maternal periconceptional undernutrition (PCUN) alters fetal hypothalamic-pituitary-adrenal axis (HPAA) function and placental glucocorticoid metabolism in sheep. The effects of PCUN on HPAA function in adult life are not known. We investigated the effects of PCUN on fetal adrenal development across gestation and on cortisol regulation in adult offspring.

Whole-slide imaging and a Fiji-based image analysis workflow of immunohistochemistry staining of pancreatic islets.

Quantification of cell populations in tissue sections is frequently examined in studies of human disease. However, traditional manual imaging of sections stained with immunohistochemistry is laborious, time-consuming, and often assesses fields of view rather than the whole tissue section. The analysis is usually manual or utilises expensive proprietary image analysis platforms.

Effect of Bias Gas Flow on Tracheal Cytokine Concentrations in Ventilated Extremely Preterm Infants: A Randomized Controlled Trial.

Background: The objective of this study was to determine whether ventilator bias gas flow affects tracheal aspirate (TA) cytokine concentrations in ventilated extremely preterm infants.

Methods: This is a randomized controlled trial in a tertiary neonatal unit in New Zealand. Preterm infants (<28 weeks' gestation/<1,000 g) requiring intubation in the first 7 days after birth were randomized to bias gas flows of 4 or 10 L/min.

Effects of maternal periconceptional undernutrition in sheep on offspring glucose-insulin axis function into adulthood.

Maternal periconceptional undernutrition (PCUN) affected fetal pancreatic maturation in late gestation lambs and impaired glucose tolerance in 10-month-old sheep. To examine the importance of the timing of maternal undernutrition around conception, a further cohort was born to PCUN ewes [undernourished for 61 d before conception (PreC), 30 d after conception (PostC), or 61 d before until 30 d after conception (PrePostC)], or normally fed ewes (Control) (n = 15-20/group). We compared glucose tolerance, insulin secretion, and sensitivity at 36 months of age.

Effects of preterm birth induced with or without exogenous glucocorticoids on the ovine glucose-insulin axis.

Antenatal exogenous glucocorticoids (ANG) are standard management for women at risk of preterm birth but are reputed to impair glucose tolerance in preterm offspring. We compared lambs born preterm (137 days gestation) following labour induced with exogenous glucocorticoids (G-Prem, glucocorticoid-induced preterm group), or with a progesterone synthesis inhibitor (NG-Prem, non-glucocorticoid-induced preterm group), with term-born lambs (Term; 149 days). We assessed glucose tolerance, insulin secretion and sensitivity at 4 and 10 months n = 11-14/group) and pancreatic and hepatic gene and protein expression at 4 weeks post-term (4 weeks; n = 6/group) and 12 months (12 months; n = 12-13/group).

Brief neonatal nutritional supplementation has sex-specific effects on glucose tolerance and insulin regulating genes in juvenile lambs.

Background: The nutritional plane and composition during fetal life can impact upon growth and epigenetic regulation of genes affecting pancreatic β-cell development and function. However, it is not clear whether β-cell development can be altered by nutritional factors or growth rate after birth. We therefore investigated the effect of neonatal nutritional supplements on growth, glucose tolerance, and pancreatic development in lambs.

Weekly intra-amniotic IGF-1 treatment increases growth of growth-restricted ovine fetuses and up-regulates placental amino acid transporters.

Frequent treatment of the growth-restricted (IUGR) ovine fetus with intra-amniotic IGF-1 increases fetal growth. We aimed to determine whether increased growth was maintained with an extended dosing interval and to examine possible mechanisms. Pregnant ewes were allocated to three groups: Control, and two IUGR groups (induced by placental embolization) treated with weekly intra-amniotic injections of either saline (IUGR) or 360 µg IGF-1 (IGF1).

Differential regulation of igf1 and igf1r mRNA levels in the two hepatic lobes following intrauterine growth restriction and its treatment with intra-amniotic insulin-like growth factor-1 in ovine fetuses.

Intrauterine growth restriction (IUGR) has life-long health implications, yet there is no effective prenatal treatment. Daily intra-amniotic administration of insulin-like growth factor (IGF)-1 to IUGR fetal sheep improves fetal gut maturation but suppresses hepatic igf1 gene expression. Fetal hepatic blood supply is regulated, in part, by shunting of oxygen- and nutrient-rich umbilical venous blood through the ductus venosus, with the left hepatic lobe predominantly supplied by umbilical venous blood and the right hepatic lobe predominantly supplied by the portal circulation.

Studying vertebrate topoisomerase 2 function using a conditional knockdown system in DT40 cells.

DT40 is a B-cell lymphoma-derived avian cell line widely used to study cell autonomous gene function because of the high rates with which DNA constructs are homologously recombined into its genome. Here, we demonstrate that the power of the DT40 system can be extended yet further through the use of RNA interference as an alternative to gene targeting. We have generated and characterized stable DT40 transfectants in which both topo 2 genes have been in situ tagged using gene targeting, and from which the mRNA of both topoisomerase 2 isoforms can be conditionally depleted through the tetracycline-induced expression of short hairpin RNAs.

Depletion of topoisomerase IIalpha leads to shortening of the metaphase interkinetochore distance and abnormal persistence of PICH-coated anaphase threads.

Topoisomerase II (topo II) is a major component of mitotic chromosomes, and its unique decatenating activity has been implicated in many aspects of chromosome dynamics, of which chromosome segregation is the most seriously affected by loss of topo II activity in living cells. There is considerable evidence that topo II plays a role at the centromere including: the centromere-specific accumulation of topo II protein; cytogenetic/molecular mapping of the catalytic activity of topo II to active centromeres; the influence of sumoylated topo II on sister centromere cohesion; and its involvement in the activation of a Mad2-dependent spindle checkpoint. By using a human cell line with a conditional-lethal mutation in the gene encoding DNA topoisomerase IIalpha, we find that depletion of topo IIalpha, while leading to a disorganised metaphase plate, does not have any overt effect on general assembly of kinetochores.