Estimation of Gestational Age via Image Analysis of Anterior Lens Capsule Vascularity in Preterm Infants: A Pilot Study

Introduction: Anterior lens capsule vascularity (ALCV) is resorbed in the developing fetus from 27 to 35 weeks gestation. In this pilot study, we evaluated the feasibility and validity of combining smartphone ophthalmoscope videos of ALCV and image analysis for gestational age estimation.Methods: ALCV videos were captured longitudinally in preterm neonates from delivery using a PanOptic® Ophthalmoscope with an iExaminer® adapter (Welch-Allyn). ALCV video frames were manually selected and quantified using semi-automatic image analysis. A predictive model based on ALCV features was compared to gold-standard ultrasound gestational age estimates.Results: A total of 64 image-capture sessions were carried out in 24 neonates. Ultrasound-estimated gestational age and ALCV-predicted gestational age estimates indicate that the two methods are similar (r = 0.78, p < 0.0001). ALCV estimates of gestational age were within 0.11 ± 1.3 weeks of ultrasound estimates. In the final model, gestational age was predicted within ± 1 week for 54% and within ± 2 weeks for 86% of the measures.Conclusions: This novel application of smartphone ophthalmoscopy and ALCV image analysis may provide a safe, accurate and non-invasive technology to estimate postnatal gestational age, especially in low income countries where gestational age may not be known at birth

Deferoxamine produces nitric oxide under ferricyanide oxidation, blood incubation, and UV-irradiation.

Deferoxamine (DFO), an iron chelator, is used therapeutically for the removal of excess iron in multiple clinical conditions such as beta thalassemia and intracerebral hemorrhage. DFO is also used as an iron chelator and hypoxia-mimetic agent in in vivo and in vitro basic research. Here we unexpectedly discover DFO to be a nitric oxide (NO) precursor in experiments where it was intended to act as an iron chelator. Production of NO from aqueous solutions of DFO was directly observed by ozone-based chemiluminescence using a ferricyanide-based assay and was confirmed by electron paramagnetic resonance (EPR). DFO also produced NO following exposure to ultraviolet light, and its incubation with sheep adult and fetal blood resulted in considerable formation of iron nitrosyl hemoglobin, as confirmed by both visible spectroscopy and EPR. These results suggest that experiments using DFO can be confounded by concomitant production of NO, and offer new insight into some of DFOs unexplained clinical side effects such as hypotension

Chronic High-Altitude Hypoxia Alters Iron and Nitric Oxide Homeostasis in Fetal and Maternal Sheep Blood and Aorta

The mammalian fetus thrives at oxygen tensions much lower than those of adults. Gestation at high altitude superimposes hypoxic stresses on the fetus resulting in increased erythropoiesis. We hypothesized that chronic hypoxia at high altitude alters the homeostasis of iron and bioactive nitric oxide metabolites (NOx) in gestation. To test for this, electron paramagnetic resonance was used to provide unique measurements of iron, metalloproteins, and free radicals in the blood and aorta of fetal and maternal sheep from either high or low altitudes (3801 or 300 m). Using ozone-based chemiluminescence with selectivity for various NOx species, we determined the NOx levels in these samples immediately after collection. These experiments demonstrated a systemic redistribution of iron in high altitude fetuses as manifested by a decrease in both chelatable and total iron in the aorta and an increase in non-transferrin bound iron and total iron in plasma. Likewise, high altitude altered the redox status diversely in fetal blood and aorta. This study also found significant increases in blood and aortic tissue NOx in fetuses and mothers at high altitude. In addition, gradients in NOx concentrations observed between fetus and mother, umbilical artery and vein, and plasma and RBCs demonstrated complex dynamic homeostasis of NOx among these circulatory compartments, such as placental generation and efflux as well as fetal consumption of iron-nitrosyls in RBCs, probably HbNO. In conclusion, these results may suggest the utilization of iron from non-hematopoietic tissues iron for erythropoiesis in the fetus and increased NO bioavailability in response to chronic hypoxic stress at high altitude during gestation

Artifacts Introduced by Sample Handling in Chemiluminescence Assays of Nitric Oxide Metabolites

We recently developed a combination of four chemiluminescence-based assays for selective detection of different nitric oxide (NO) metabolites, including nitrite, S-nitrosothiols (SNOs), heme-nitrosyl (heme-NO), and dinitrosyl iron complexes (DNICs). However, these NO species (NOx) may be under dynamic equilibria during sample handling, which affects the final determination made from the readout of assays. Using fetal and maternal sheep from low and high altitudes (300 and 3801 m, respectively) as models of different NOx levels and compositions, we tested the hypothesis that sample handling introduces artifacts in chemiluminescence assays of NOx. Here, we demonstrate the following: (1) room temperature placement is associated with an increase and decrease in NOx in plasma and whole blood samples, respectively; (2) snap freezing and thawing lead to the interconversion of different NOx in plasma; (3) snap freezing and homogenization in liquid nitrogen eliminate a significant fraction of NOx in the aorta of stressed animals; (4) A “stop solution” commonly used to preserve nitrite and SNOs leads to the interconversion of different NOx in blood, while deproteinization results in a significant increase in detectable NOx; (5) some reagents widely used in sample pretreatments, such as mercury chloride, acid sulfanilamide, N-ethylmaleimide, ferricyanide, and anticoagulant ethylenediaminetetraacetic acid, have unintended effects that destabilize SNO, DNICs, and/or heme-NO; (6) blood, including the residual blood clot left in the washed purge vessel, quenches the signal of nitrite when using ascorbic acid and acetic acid as the purge vessel reagent; and (7) new limitations to the four chemiluminescence-based assays. This study points out the need for re-evaluation of previous chemiluminescence measurements of NOx, and calls for special attention to be paid to sample handling, as it can introduce significant artifacts into NOx assays

Iron nitrosyl complexes are formed from nitrite in the human placenta.

Placental nitric oxide (NO) is critical for maintaining perfusion in the maternal-fetal-placental circulation during normal pregnancy. NO and its many metabolites are also increased in pregnancies complicated by maternal inflammation such as preeclampsia, fetal growth restriction, gestational diabetes, and bacterial infection. However, it is unclear how increased levels of NO or its metabolites affect placental function or how the placenta deals with excessive levels of NO or its metabolites. Since there is uncertainty over the direction of change in plasma levels of NO metabolites in preeclampsia, we measured the levels of these metabolites at the placental tissue level. We found that NO metabolites are increased in placentas from patients with preeclampsia compared to healthy controls. We also discovered by ozone-based chemiluminescence and electron paramagnetic resonance that nitrite is efficiently converted into iron nitrosyl complexes (FeNOs) within the human placenta and also observed the existence of endogenous FeNOs within placentas from sheep and rats. We show these nitrite-derived FeNOs are relatively short-lived, predominantly protein-bound, heme-FeNOs. The efficient formation of FeNOs from nitrite in the human placenta hints toward the importance of both nitrite and FeNOs in placental physiology or pathology. As iron nitrosylation is an important posttranslational modification that affects the activity of multiple iron-containing proteins such as those in the electron transport chain, or those involved in epigenetic regulation, we conclude that FeNOs merit increased study in pregnancy complications

Gestational Hypoxia and Programing of Lung Metabolism

Gestational hypoxia is a risk factor in the development of pulmonary hypertension in the newborn and other sequela, however, the mechanisms associated with the disease remain poorly understood. This review highlights disruption of metabolism by antenatal high altitude hypoxia and the impact this has on pulmonary hypertension in the newborn with discussion of model organisms and human populations. There is particular emphasis on modifications in glucose and lipid metabolism along with alterations in mitochondrial function. Additional focus is placed on increases in oxidative stress and the progression of pulmonary vascular disease in the newborn and on the need for further exploration using a combination of contemporary and classical approaches

A novel, noninvasive, predictive epilepsy biomarker with clinical potential.

A significant proportion of temporal lobe epilepsy (TLE), a common, intractable brain disorder, arises in children with febrile status epilepticus (FSE). Preventative therapy development is hampered by our inability to identify early the FSE individuals who will develop TLE. In a naturalistic rat model of FSE, we used high-magnetic-field MRI and long-term video EEG to seek clinically relevant noninvasive markers of epileptogenesis and found that reduced amygdala T2 relaxation times in high-magnetic-field MRI hours after FSE predicted experimental TLE. Reduced T2 values likely represented paramagnetic susceptibility effects derived from increased unsaturated venous hemoglobin, suggesting augmented oxygen utilization after FSE termination. Indeed, T2 correlated with energy-demanding intracellular translocation of the injury-sensor high-mobility group box 1 (HMGB1), a trigger of inflammatory cascades implicated in epileptogenesis. Use of deoxyhemoglobin-sensitive MRI sequences enabled visualization of the predictive changes on lower-field, clinically relevant scanners. This novel MRI signature delineates the onset and suggests mechanisms of epileptogenesis that follow experimental FSE