ANO2 Genetic Variants and Anti-VEGF Treatment Response in Neovascular AMD: A Pharmacogenetic Substudy of VIEW 1 and VIEW 2.

Purpose: This analysis investigated potential associations between gene variants and clinical end points in the VIEW 1 and 2 randomized clinical trials of intravitreal aflibercept and ranibizumab in neovascular age-related macular degeneration (AMD). Methods: A genome-wide association analysis was conducted in a subgroup of patients from VIEW 1 and 2 consenting to the optional pharmacogenetic analysis. Results: Data were pooled from 780 samples from patients representative of the overall VIEW 1 and 2 populations. After Bonferroni correction for multiplicity and statistical adjustment for baseline risk factors, no significant associations were found between previously identified prognostic AMD gene variants and treatment response according to key prespecified VIEW 1 and 2 end points. Genome-wide, there were no significant genetic associations in patients experiencing gains of ≥15 Early Treatment of Diabetic Retinopathy Study letters after 1 or 2 years of treatment. A cluster of variants in ANO2 (encoding anoctamin 2, a calcium-activated chloride channel expressed on photoreceptor cells) on chromosome 12 reached the level of significance for loss of ≥5 letters after 1 year of treatment (P < 5 × 10-8), with the ANO2 rs2110166 SNP demonstrating highly significant association (P = 1.99 × 10-8). Carriers of the ANO2 rs2110166 TT genotype showed a robust increase in visual acuity versus baseline compared with a small decrease in those with the TC genotype. Conclusions: None of the potential prognostic candidate genes were associated with the clinical end points for treated patients. Preliminary analyses suggest an association of ANO2 with retinal function, with a potential impact on vision of approximately one line over at least the first year. Further investigation of the function of ANO2 in retinal pathophysiology is merited.

Potential microemboli formation risk and its management during the heated saline-enhanced radiofrequency needle-tip catheter ablation.

BACKGROUND: The potential risk of embolic events during ablation in the left ventricle (LV) with a heated saline-enhanced radiofrequency (SERF) needle-tip ablation catheter has not been characterized. OBJECTIVE: This study aimed to investigate the formation of microemboli or other untoward events during SERF ablation. METHODS: Ninety-three radiofrequency (RF) ablation procedures were performed in the LV of 14 pigs by using a SERF catheter (35 W, 70 seconds, and 60°C; normal or degassed saline [NS or DS] irrigation with a flow rate of 10 mL/min) vs a standard irrigated-tip radiofrequency (S-RF) catheter (30 or 50 W, 30 seconds, and 17 mL/min). Microbubble formation was graded on the basis of intracardiac echocardiography. Microbubbles, microembolic signals, and microparticles were monitored using our established model. RESULTS: There was no significant difference in microbubble volume among SERF-NS, SERF-DS, and S-RF 30 W with "grade 1" intracardiac echocardiography microbubbles (median and 25th-75th percentiles 0.201 [0.011-3.13], 0.455 [0.06-2.66], and 0.004 µL [0.00-0.16 µL], respectively). There was no significant difference in microembolic signals among SERF-NS, SERF-DS, and S-RF 30 W with grade 1 bubbles (n = 8.0 ± 5.8, n = 7.6 ± 4.2, and n = 6.1 ± 6.1, respectively). Both SERF-NS and SERF-DS created larger lesions than did both S-RF 30 W and S-RF 50 W deliveries (mean 1241.5 ± 658.6, 1497.7 ± 893.4, 75.0 ± 24.8, and 184.0 ± 93.8 mm3; P < .001). There was no significant difference in microparticle incidence among groups (P = .675). No evidence of embolic events was found in the brain and other organs at the histology assessment. CONCLUSION: In the setting of SERF ablation, significantly large LV lesions can be created without any increment in embolic microbubble or particle events. Grade 1 microbubble is related to the efficacy and safety.

Viral Infection Leads to a Unique Suite of Allelopathic Chemical Signals in Three Diatom Host-Virus Pairs.

Ecophysiological stress and the grazing of diatoms are known to elicit the production of chemical defense compounds called oxylipins, which are toxic to a wide range of marine organisms. Here we show that (1) the viral infection and lysis of diatoms resulted in oxylipin production; (2) the suite of compounds produced depended on the diatom host and the infecting virus; and (3) the virus-mediated oxylipidome was distinct, in both magnitude and diversity, from oxylipins produced due to stress associated with the growth phase. We used high-resolution accurate-mass mass spectrometry to observe changes in the dissolved lipidome of diatom cells infected with viruses over 3 to 4 days, compared to diatom cells in exponential, stationary, and decline phases of growth. Three host virus pairs were used as model systems: Chaetoceros tenuissimus infected with CtenDNAV; C. tenuissimus infected with CtenRNAV; and Chaetoceros socialis infected with CsfrRNAV. Several of the compounds that were significantly overproduced during viral infection are known to decrease the reproductive success of copepods and interfere with microzooplankton grazing. Specifically, oxylipins associated with allelopathy towards zooplankton from the 6-, 9-, 11-, and 15-lipogenase (LOX) pathways were significantly more abundant during viral lysis. 9-hydroperoxy hexadecatetraenoic acid was identified as the strongest biomarker for the infection of Chaetoceros diatoms. C. tenuissimus produced longer, more oxidized oxylipins when lysed by CtenRNAV compared to CtenDNAV. However, CtenDNAV caused a more statistically significant response in the lipidome, producing more oxylipins from known diatom LOX pathways than CtenRNAV. A smaller set of compounds was significantly more abundant in stationary and declining C. tenuissimus and C. socialis controls. Two allelopathic oxylipins in the 15-LOX pathway and essential fatty acids, arachidonic acid (ARA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) were more abundant in the stationary phase than during the lysis of C. socialis. The host-virus pair comparisons underscore the species-level differences in oxylipin production and the value of screening more host-virus systems. We propose that the viral infection of diatoms elicits chemical defense via oxylipins which deters grazing with downstream trophic and biogeochemical effects.

Comparison of Glutathione Nanoparticles, CoEnzyme Q10, and Fish Oil for Prevention of Oxygen-Induced Retinopathy in Neonatal Rats.

Notch ligands and receptors are important for cell specification and angiogenesis, but their role in oxygen-induced retinopathy (OIR) is not well studied. Delta-like ligand (DLL)-4/Notch inhibits angiogenesis, while Jagged-1/Notch promotes angiogenesis. We tested the hypothesis that early supplementation with antioxidants and/or fish oil curtails severe OIR by inducing DLL-4/Notch and reducing Jagged-1/Notch. Newborn rats were exposed to brief intermittent hypoxia (IH) during hyperoxia, during which they received daily oral supplements of (1) fish oil, (2) coenzyme Q10 (CoQ10) in olive oil (OO), (3) glutathione nanoparticles (nGSH), (4) fish oil + CoQ10, or (5) OO (controls) from birth (P0) to P14. At P14, the pups were placed in room air (RA) until P21, with no further treatment. Oxidative stress, apoptosis, ocular histopathology, and Notch signaling were assessed. Neonatal IH resulted in severe retinal damage consistent with retinopathy of prematurity (ROP). Retinal damage was associated with induced oxidative stress and Jagged-1/Notch signaling, as well as reduced DLL-4/Notch signaling. All treatments reversed these outcomes, but nGSH produced the most beneficial outcomes. Severe OIR promoted the induction of Jagged-1/Notch and curtailed DLL-4/Notch, which was an effect that could be reversed with nGSH supplementation. These findings may indicate a potential alternate pathway for ROP treatment and/or prevention.

Early versus late caffeine and/or non-steroidal anti-inflammatory drugs (NSAIDS) for prevention of intermittent hypoxia-induced neuroinflammation in the neonatal rat.

Preterm infants often experience frequent intermittent hypoxia (IH) episodes which are associated with neuroinflammation. We tested the hypotheses that early caffeine and/or non-steroidal inflammatory drugs (NSAIDs) confer superior therapeutic benefits for protection against IH-induced neuroinflammation than late treatment. Newborn rats were exposed to IH or hyperoxia (50% O2) from birth (P0) to P14. For early treatment, the pups were administered: 1) daily caffeine (Caff) citrate (Cafcit, 20 mg/kg IP loading on P0, followed by 5 mg/kg from P1-P14); 2) ketorolac (Keto) topical ocular solution in both eyes from P0 to P14; 3) ibuprofen (Ibu, Neoprofen, 10 mg/kg loading dose on P0 followed by 5 mg/kg/day on P1 and P2); 4) Caff+Keto co-treatment; 5) Caff+Ibu co-treatment; or 6) equivalent volume saline (Sal). On P14, animals were placed in room air (RA) with no further treatment until P21. For late treatment, pups were exposed from P0 to P14, then placed in RA during which they received similar treatments from P15-P21 (Sal, Caff, and/or Keto), or P15-P17 (Ibu). RA controls were similarly treated. At P21, whole brains were assessed for histopathology, apoptosis, myelination, and biomarkers of inflammation. IH caused significant brain injury and hemorrhage, inflammation, reduced myelination, and apoptosis. Early treatment with Caff alone or in combination with NSAIDs conferred better neuroprotection against IH-induced damage than late treatment. Early postnatal treatment during a critical time of brain development, may be preferable for the prevention of IH-induced brain injury in preterm infants.

Novel Role for Cardiolipin as a Target of Therapy to Mitigate Myocardial Injury Caused by Venoarterial Extracorporeal Membrane Oxygenation.

BACKGROUND: Cardiolipin is a mitochondrial-specific phospholipid that maintains integrity of the electron transport chain (ETC) and plays a central role in myocardial ischemia/reperfusion injury. Tafazzin is an enzyme that is required for cardiolipin maturation. Venoarterial extracorporeal membrane oxygenation (VA-ECMO) use to provide hemodynamic support for acute myocardial infarction has grown exponentially, is associated with poor outcomes, and is under active clinical investigation, yet the mechanistic effect of VA-ECMO on myocardial damage in acute myocardial infarction remains poorly understood. We hypothesized that VA-ECMO acutely depletes myocardial cardiolipin and exacerbates myocardial injury in acute myocardial infarction. METHODS: We examined cardiolipin and tafazzin levels in human subjects with heart failure and healthy swine exposed to VA-ECMO and used a swine model of closed-chest myocardial ischemia/reperfusion injury to evaluate the effect of VA-ECMO on cardiolipin expression, myocardial injury, and mitochondrial function. RESULTS: Cardiolipin and tafazzin levels are significantly reduced in the left ventricles of individuals requiring VA-ECMO compared with individuals without VA-ECMO before heart transplantation. Six hours of exposure to VA-ECMO also decreased left ventricular levels of cardiolipin and tafazzin in healthy swine compared with sham controls. To explore whether cardiolipin depletion by VA-ECMO increases infarct size, we performed left anterior descending artery occlusion for a total of 120 minutes followed by 180 minutes of reperfusion in adult swine in the presence and absence of MTP-131, an amphipathic molecule that interacts with cardiolipin to stabilize the inner mitochondrial membrane. Compared with reperfusion alone, VA-ECMO activation beginning after 90 minutes of left anterior descending artery occlusion increased infarct size (36±8% versus 48±7%; P<0.001). VA-ECMO also decreased cardiolipin and tafazzin levels, disrupted mitochondrial integrity, reduced electron transport chain function, and promoted oxidative stress. Compared with reperfusion alone or VA-ECMO before reperfusion, delivery of MTP-131 before VA-ECMO activation reduced infarct size (22±8%; P=0.03 versus reperfusion alone and P<0.001 versus VA-ECMO alone). MTP-131 restored cardiolipin and tafazzin levels, stabilized mitochondrial function, and reduced oxidative stress in the left ventricle. CONCLUSIONS: We identified a novel mechanism by which VA-ECMO promotes myocardial injury and further identify cardiolipin as an important target of therapy to reduce infarct size and to preserve mitochondrial function in the setting of VA-ECMO for acute myocardial infarction.

Real-World Management of Macular Edema Secondary to Retinal Vein Occlusion with Intravitreal Aflibercept: 24-month Results from the AURIGA Observational Study.

INTRODUCTION: AURIGA is the largest real-world study to date to evaluate intravitreal aflibercept (IVT-AFL) treatment of diabetic macular edema or macular edema secondary to retinal vein occlusion (RVO) in routine clinical practice. Here, we report the 24-month outcomes in the RVO cohort from France, Germany, Italy, and Taiwan. METHODS: AURIGA (NCT03161912) was a prospective observational study. Eligible patients with RVO were enrolled for whom the decision to treat with IVT-AFL had already been made by the attending physician. Patients were treated with IVT-AFL for up to 24 months at physician discretion according to local practice. The primary endpoint was mean change in visual acuity (VA; Early Treatment Diabetic Retinopathy Study [ETDRS] letters) from baseline to month (M) 12. All statistical analyses were descriptive. RESULTS: In 554 treatment-naïve and 65 previously treated patients with RVO, the respective mean (95% confidence interval) change in VA from baseline was + 12.5 (10.8, 14.3) and + 7.9 (3.3, 12.6) letters by M12 and + 11.4 (9.4, 13.3) and + 4.4 (- 0.6, 9.5) letters by M24 (baseline mean ± standard deviation: 51.0 ± 21.9 and 51.9 ± 20.4 letters); 44.0% of treatment-naïve and 27.9% of previously treated patients reported ≥ 15-letter gains by M24. By M24, the mean change in central retinal thickness from baseline was - 247 (- 267, - 227) µm in treatment-naïve patients and - 147 (- 192, - 102) µm in previously treated patients. From baseline to M6, M12, and M24, treatment-naïve patients received a total of 4.0 ± 1.3, 5.5 ± 2.5, and 6.9 ± 4.2 injections, respectively, and previously treated patients received 3.8 ± 1.5, 5.0 ± 2.2, and 6.3 ± 3.7 injections, respectively. The safety profile of IVT-AFL was consistent with that of previous studies. CONCLUSIONS: In AURIGA, patients with RVO experienced clinically relevant functional and anatomic improvements following IVT-AFL treatment in routine clinical practice. These improvements were largely maintained in treatment-naïve patients over the 24-month study despite the decreasing treatment frequency, suggesting long-term durability of IVT-AFL treatment outcomes. Infographic available for this article. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03161912 (May 19, 2017). INFOGRAPHIC.

Ubiquitous, B12-dependent virioplankton utilizing ribonucleotide-triphosphate reductase demonstrate interseasonal dynamics and associate with a diverse range of bacterial hosts in the pelagic ocean.

Through infection and lysis of their coexisting bacterial hosts, viruses impact the biogeochemical cycles sustaining globally significant pelagic oceanic ecosystems. Currently, little is known of the ecological interactions between lytic viruses and their bacterial hosts underlying these biogeochemical impacts at ecosystem scales. This study focused on populations of lytic viruses carrying the B12-dependent Class II monomeric ribonucleotide reductase (RNR) gene, ribonucleotide-triphosphate reductase (Class II RTPR), documenting seasonal changes in pelagic virioplankton and bacterioplankton using amplicon sequences of Class II RTPR and the 16S rRNA gene, respectively. Amplicon sequence libraries were analyzed using compositional data analysis tools that account for the compositional nature of these data. Both virio- and bacterioplankton communities responded to environmental changes typically seen across seasonal cycles as well as shorter term upwelling-downwelling events. Defining Class II RTPR-carrying viral populations according to major phylogenetic clades proved a more robust means of exploring virioplankton ecology than operational taxonomic units defined by percent sequence homology. Virioplankton Class II RTPR populations showed positive associations with a broad phylogenetic diversity of bacterioplankton including dominant taxa within pelagic oceanic ecosystems such as Prochlorococcus and SAR11. Temporal changes in Class II RTPR virioplankton, occurring as both free viruses and within infected cells, indicated possible viral-host pairs undergoing sustained infection and lysis cycles throughout the seasonal study. Phylogenetic relationships inferred from Class II RTPR sequences mirrored ecological patterns in virio- and bacterioplankton populations demonstrating possible genome to phenome associations for an essential viral replication gene.

Factors Influencing Neonatal Gut Microbiome and Health with a Focus on Necrotizing Enterocolitis.

Maturational changes in the gut start in utero and rapidly progress after birth, with some functions becoming fully developed several months or years post birth including the acquisition of a full gut microbiome, which is made up of trillions of bacteria of thousands of species. Many factors influence the normal development of the neonatal and infantile microbiome, resulting in dysbiosis, which is associated with various interventions used for neonatal morbidities and survival. Extremely low gestational age neonates (<28 weeks' gestation) frequently experience recurring arterial oxygen desaturations, or apneas, during the first few weeks of life. Apnea, or the cessation of breathing lasting 15-20 s or more, occurs due to immature respiratory control and is commonly associated with intermittent hypoxia (IH). Chronic IH induces oxygen radical diseases of the neonate, including necrotizing enterocolitis (NEC), the most common and devastating gastrointestinal disease in preterm infants. NEC is associated with an immature intestinal structure and function and involves dysbiosis of the gut microbiome, inflammation, and necrosis of the intestinal mucosal layer. This review describes the factors that influence the neonatal gut microbiome and dysbiosis, which predispose preterm infants to NEC. Current and future management and therapies, including the avoidance of dysbiosis, the use of a human milk diet, probiotics, prebiotics, synbiotics, restricted antibiotics, and fecal transplantation, for the prevention of NEC and the promotion of a healthy gut microbiome are also reviewed. Interventions directed at boosting endogenous and/or exogenous antioxidant supplementation may not only help with prevention, but may also lessen the severity or shorten the course of the disease.

Comparison of hyperoxia or normoxia resolution of intermittent hypoxia and intermittent hyperoxia episodes on liver histopathology, IGF-1, IGFBP-3, and GHBP in neonatal rats.

OBJECTIVE: Extremely low gestational age neonates requiring oxygen therapy for chronic lung disease experience repeated fluctuations in arterial oxygen saturation, or intermittent hypoxia (IH), during the first few weeks of life. These events are associated with a high risk for reduced growth, hypertension, and insulin resistance in later life. This study tested the hypothesis that IH, or intermittent hyperoxia have similar negative effects on the liver; somatic growth; and liver insulin-like growth factor (IGF)-I, IGF binding protein (BP)-3, and growth hormone binding protein (GHBP), regardless of resolution in normoxia or hyperoxia between episodes. DESIGN: Newborn rats on the first day of life (P0) were exposed to two IH paradigms: 1) hyperoxia (50% O2) with brief hypoxia (12% O2); or 2) normoxia (21% O2) with hypoxia (12% O2); intermittent hypoxia (50% O2/21% O2); hyperoxia only (50% O2); or room air (RA, 21% O2). Pups were euthanized on P14 or placed in RA until P21. Controls remained in RA from P0-P21. Growth, liver histopathology, apoptosis, IGFI, IGFBP-3, and GHBP were assessed. RESULTS: Pathological findings of the liver hepatocytes, including cellular swelling, steatosis, apoptosis, necrosis and focal sinusoid congestion were seen in the IH and intermittent hyperoxia groups, and were particularly severe in the 21-12% O2 group during exposure (P14) with no significant improvements during recovery/reoxygenation (P21). These effects were associated with induction of HIF1α, and reductions in liver IGFI, IGFBP-3, and GHBP. CONCLUSIONS: Exposure to IH or intermittent hyperoxia during the first few weeks of life regardless of resolution in RA or hyperoxia is detrimental to the immature liver. These findings may suggest that interventions to prevent frequent fluctuations in oxygen saturation during early neonatal life remain a high priority.

PD-L1 Downregulation and DNA Methylation Inhibition for Molecular Therapy against Cancer Stem Cells in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a heterogeneous cancer characterized by various cellular subtypes. This study investigates the potential of a combination strategy using immunotherapy and epigenetic reprogramming against HCC. We used a transgenic HCC mouse C57BL/6J-TG(ALB1HBV)44BRI/J to assess the dynamics of the programmed death receptor and its ligand (PD-1/PD-L1) and DNA methylation markers. In parallel, PD-L1 RNA silencing was performed in various human HCC cell lines, while combination therapy was performed in a co-culture system using long-term exposure of 5-Azacytidine (5-AZA) and an anti-PD-L1. Data from the mouse model showed that the expressions of Pdcd1, Pdcd1l1, and DNA methyltransferase 1 (Dnmt1) were significantly higher in HCC as compared to the wild-type mice (p < 0.01), supported by the high presence of PD-L1 methylated DNA. In HCC cell lines, PD-L1 silencing was accompanied by DNMT1 reduction, mostly noted in aggressive HCC cell lines, followed by the dysregulation of the cancer stem cell marker EpCAM. In combination therapy, the growth of HCC cells and lymphocytes was limited by the PD-L1 antibody, further reduced in the presence of 5-AZA by up to 20% (p < 0.001). The data demonstrated that combination therapy might be an option as a potential treatment for heterogeneous HCC.

A Randomised Control Trial Investigating the Efficacy of the MapMe Intervention on Parental Ability to Correctly Categorise Overweight in Their Child and the Impact on Child BMI Z-Score Change at 1 Year.

Research suggests parental ability to recognise when their child has overweight is limited. It is hypothesised that recognition of child overweight/obesity is fundamental to its prevention, acting as a potential barrier to parental action to improve their child's health-related behaviours and/or help seeking. The purpose of this study was to investigate the efficacy of an intervention (MapMe) to improve parental ability to correctly categorise their child as having overweight one-month post-intervention, and reduce child body mass index (BMI) z-score 12 months post-intervention. MapMe consists of body image scales of known child BMI and information on the consequences of childhood overweight, associated health-related behaviours and sources of support. We conducted a three-arm (paper-based MapMe, web-based MapMe and control) randomised control trial in fifteen English local authority areas with parents/guardians of 4-5- and 10-11-year-old children. Parental categorisation of child weight status was assessed using the question 'How would you describe your child's weight at the moment?' Response options were: underweight, healthy weight, overweight, and very overweight. Child weight status and BMI z-scores were calculated using objectively measured height and weight data and UK90 clinical thresholds. There was no difference in the percentage of parents correctly categorising their child as having overweight/very overweight (n = 264: 41% control, 48% web-based, and 43% paper-based, p = 0.646). BMI z-scores were significantly reduced for the intervention group at 12 months post-intervention compared to controls (n = 338, mean difference in BMI z-score change -0.11 (95% CI -0.202 to -0.020, p = 0.017). MapMe was associated with a decrease in BMI z-score 12 months post-intervention, although there was no direct evidence of improved parental ability to correctly categorise child overweight status. Further work is needed to replicate these findings in a larger sample of children, investigate mechanisms of action, and determine the use of MapMe as a public health initiative.

Transvalvular Unloading Mitigates Ventricular Injury Due to Venoarterial Extracorporeal Membrane Oxygenation in Acute Myocardial Infarction.

Whether extracorporeal membrane oxygenation (ECMO) with Impella, known as EC-Pella, limits cardiac damage in acute myocardial infarction remains unknown. The authors now report that the combination of transvalvular unloading and ECMO (EC-Pella) initiated before reperfusion reduced infarct size compared with ECMO alone before reperfusion in a preclinical model of acute myocardial infarction. EC-Pella also reduced left ventricular pressure-volume area when transvalvular unloading was applied before, not after, activation of ECMO. The authors further observed that EC-Pella increased cardioprotective signaling but failed to rescue mitochondrial dysfunction compared with ECMO alone. These findings suggest that ECMO can increase infarct size in acute myocardial infarction and that EC-Pella can mitigate this effect but also suggest that left ventricular unloading and myocardial salvage may be uncoupled in the presence of ECMO in acute myocardial infarction. These observations implicate mechanisms beyond hemodynamic load as part of the injury cascade associated with ECMO in acute myocardial infarction.

Comparison of Bevacizumab and Aflibercept for Suppression of Angiogenesis in Human Retinal Microvascular Endothelial Cells.

Bevacizumab (Avastin) is a vascular endothelial growth factor (VEGF) inhibitor that is widely used for aggressive posterior retinopathy of prematurity (APROP). Its use is associated with multiple adverse effects. Aflibercept (Eylea) is a VEGFR-1 analogue that is approved for ocular use, but its efficacy for APROP is less studied. We tested the hypothesis that Eylea is as effective as Avastin for suppression of intermittent hypoxia (IH)-induced angiogenesis. Human retinal microvascular endothelial cells (HRECs) were treated with Avastin and low- or high-dose Eylea and exposed to normoxia, hyperoxia (50% O2), or neonatal IH for 24, 48, or 72 h. Cells were assessed for migration and tube formation capacities, as well as biomarkers of angiogenesis and oxidative stress. Both doses of Eylea suppressed migration and tube formation in all oxygen environments, although the effect was not as robust as Avastin. Furthermore, the lower dose of Eylea appeared to be more effective than the higher dose. Eylea induced soluble VEGFR-1 (sVEGFR-1) coincident with high IGF-I levels and decreased Notch/Jagged-1, demonstrating a functional association. Given the role of VEGFR-1 and Notch as guidance cues for vascular sprouting, these data suggest that Eylea may promote normal vascular patterning in a dose-dependent manner.

Altered growth and death in dilution-based viral predation assays.

Viral lysis of phytoplankton is one of the most common forms of death on Earth. Building on an assay used extensively to assess rates of phytoplankton loss to predation by grazers, lysis rates are increasingly quantified through dilution-based techniques. In this approach, dilution of viruses and hosts are expected to reduce infection rates and thus increase host net growth rates (i.e., accumulation rates). The difference between diluted and undiluted host growth rates is interpreted as a measurable proxy for the rate of viral lytic death. These assays are usually conducted in volumes ≥ 1 L. To increase throughput, we implemented a miniaturized, high-throughput, high-replication, flow cytometric microplate dilution assay to measure viral lysis in environmental samples sourced from a suburban pond and the North Atlantic Ocean. The most notable outcome we observed was a decline in phytoplankton densities that was exacerbated by dilution, instead of the increased growth rates expected from lowered virus-phytoplankton encounters. We sought to explain this counterintuitive outcome using theoretical, environmental, and experimental analyses. Our study shows that, while die-offs could be partly explained by a 'plate effect' due to small incubation volumes and cells adhering to walls, the declines in phytoplankton densities are not volume-dependent. Rather, they are driven by many density- and physiology-dependent effects of dilution on predation pressure, nutrient limitation, and growth, all of which violate the original assumptions of dilution assays. As these effects are volume-independent, these processes likely occur in all dilution assays that our analyses show to be remarkably sensitive to dilution-altered phytoplankton growth and insensitive to actual predation pressure. Incorporating altered growth as well as predation, we present a logical framework that categorizes locations by the relative dominance of these mechanisms, with general applicability to dilution-based assays.

Biomarkers of lung alveolarization and microvascular maturation in response to intermittent hypoxia and/or early antioxidant/fish oil supplementation in neonatal rats.

OBJECTIVE: Extremely preterm infants experience frequent intermittent hypoxia (IH) episodes during oxygen therapy which causes significant damage to the lungs and curtails important signaling pathways that regulate normal lung alveolarization and microvascular maturation. We tested the hypothesis that early supplementation with fish oil and/or antioxidants in rats exposed to neonatal IH improves expression of lung biomarkers of alveolarization and microvascular maturation, and reduces IH-induced lung injury. STUDY DESIGN/METHODS: From birth (P0) to P14, rat pups were exposed to room air (RA) or neonatal IH during which they received daily oral supplementation with either: (1) olive oil (OO) (control); (2) Coenzyme Q10 (CoQ10) in OO; (3) fish oil; (4) glutathione nanoparticles (nGSH); or (5) fish oil +CoQ10. At P14 pups were placed in RA until P21 with no further treatment. RA controls were similarly treated. Lung growth and alveolarization, histopathology, apoptosis, oxidative stress and biomarkers of alveolarization and microvascular maturation were determined. RESULTS: Neonatal IH was associated with reduced lung weights and severe histopathological outcomes. These effects were curtailed with fish oil and nGSH. nGSH was also protective against apoptosis, while CoQ10 prevented IH-induced ROS production. Of all treatments, nGSH and CoQ10 + fish oil-induced vascular endothelial growth factor165 and CD31 (Platelet endothelial cell adhesion molecule-1), which are associated with angiogenesis. CoQ10 + fish oil improved alveolarization in RA and IH despite evidence of hemorrhage. CONCLUSIONS: The benefits of nGSH and CoQ10 + fish oil suggest an antioxidant effect which may be required to curtail IH-induced lung injury. Further clinical assessment of the effectiveness of nGSH is warranted.

Differences Between Self-Reported and Objectively Measured Hearing Loss at Age 61-63 Years: The Newcastle Thousand Families Birth Cohort.

PURPOSE: Hearing loss is most prevalent among older adults, yet underestimated by patients, clinicians, and research communities. This study aimed to assess the accuracy of self-reported hearing difficulties among a group of adults aged 61-63 years, against audiometric measures. METHOD: The analysis used a sample (N = 346) of the Newcastle Thousand Families Study birth cohort. Data from audiological examinations and self-reported hearing difficulties were used to compare subjective and objective hearing. Hearing aid use was also assessed. RESULTS: Over 40% of the participants had some level of hearing loss (n = 155, 44.8%), and 31% (n = 133) of these reported having hearing problems during subjective assessment. Only 18 (10%) of those with objectively measured hearing loss reported using hearing aids. CONCLUSIONS: There was an inconsistency between measured and self-perceived hearing loss among adults aged 61-63 years in this cohort. The small number of hearing aid users in the cohort is a concern, in that people in this age group appear not to be getting the help they need. The data also add to the research evidence that people at this age underestimate their hearing loss.

Aflibercept Suppression of Angiopoietin-2 in a Rabbit Retinal Vascular Hyperpermeability Model.

Purpose: Anti-vascular endothelial growth factor (anti-VEGF) therapies, which attenuate the capacity of VEGF to bind to VEGF receptors, are standard-of-care options for various retinal disorders that are characterized by pathologic retinal angiogenesis and vascular permeability. Multiple receptors and ligands have also been reported as being involved in these pathways, including angiopoietin-1 (ANG1) and angiopoietin-2 (ANG2). Methods: Electrochemiluminescence immunoassays were used to detect human VEGF (hVEGF), as well as rabbit ANG2 and basic fibroblast growth factor protein levels in vitreous samples derived from a study evaluating the efficacy of the anti-VEGF agents ranibizumab, aflibercept, and brolucizumab in an hVEGF165-induced rabbit retinal vascular hyperpermeability model. Results: hVEGF was completely suppressed in rabbit vitreous after anti-VEGF treatment for 28 days. ANG2 protein in vitreous and ANGPT2 mRNA in retina tissue were similarly suppressed, although the anti-VEGF agents do not directly bind to ANG2. Aflibercept demonstrated the greatest inhibitory effect in ANG2 levels in vitreous, which correlated with strong, durable suppression of intraocular hVEGF levels. Conclusions: This study explored the effects of anti-VEGF therapies beyond direct binding of VEGF by evaluating protein levels and the expression of target genes involved in angiogenesis and associated molecular mechanisms in the rabbit retina and choroid. Translational Relevance: In vivo data suggest that anti-VEGF agents currently used for the treatment of retinal diseases could provide beneficial effects beyond direct binding of VEGF, including suppression of ANG2 protein and ANGPT2 mRNA.

Virus infection of phytoplankton increases average molar mass and reduces hygroscopicity of aerosolized organic matter.

Viral infection of phytoplankton is a pervasive mechanism of cell death and bloom termination, which leads to the production of dissolved and colloidal organic matter that can be aerosolized into the atmosphere. Earth-observing satellites can track the growth and death of phytoplankton blooms on weekly time scales but the impact of viral infection on the cloud forming potential of associated aerosols is largely unknown. Here, we determine the influence of viral-derived organic matter, purified viruses, and marine hydrogels on the cloud condensation nuclei activity of their aerosolized solutions, compared to organic exudates from healthy phytoplankton. Dissolved organic material derived from exponentially growing and infected cells of well-characterized eukaryotic phytoplankton host-virus systems, including viruses from diatoms, coccolithophores and chlorophytes, was concentrated, desalted, and nebulized to form aerosol particles composed of primarily of organic matter. Aerosols from infected phytoplankton cultures resulted in an increase in critical activation diameter and average molar mass in three out of five combinations evaluated, along with a decrease in organic kappa (hygroscopicity) compared to healthy cultures and seawater controls. The infected samples also displayed evidence of increased surface tension depression at realistic cloud water vapor supersaturations. Amending the samples with xanthan gum to simulate marine hydrogels increased variability in organic kappa and surface tension in aerosols with high organic to salt ratios. Our findings suggest that the pulses of increased dissolved organic matter associated with viral infection in surface waters may increase the molar mass of dissolved organic compounds relative to surface waters occupied by healthy phytoplankton or low phytoplankton biomass.