Journal articles on the topic 'Novela juveni.l'

Olive (Olea europaea L.) is the most characteristic and important oil crop of the Mediterranean region. Traditional olive cultivation is based on few tens cultivars of ancient origin. To improve this crop, novel selections with higher tolerance to biotic and abiotic stress, adaptable to high-density planting systems and resilient to climate change are needed; however, breeding programs are hindered by the long juvenile period of this species and few improved genotypes have been released so far. Genetic transformation could be of great value, in the near future, to develop new varieties or rootstocks in a shorter time; in addition, it has currently become an essential tool for functional genomic studies. The recalcitrance of olive tissues to their in vitro manipulation has been the main bottleneck in the development of genetic transformation procedures in this species; however, some important traits such as fungal resistance, flowering or lipid composition have successfully been manipulated through the genetic transformation of somatic embryos of juvenile or adult origin, providing a proof of the potential role that this technology could have in olive improvement. However, the optimization of these protocols for explants of adult origin is a prerequisite to obtain useful materials for the olive industry. In this review, initially, factors affecting plant regeneration via somatic embryogenesis are discussed. Subsequently, the different transformation approaches explored in olive are reviewed. Finally, transgenic experiments with genes of interest undertaken to manipulate selected traits are discussed.

Laevistrombus canarium, also known as dog conch, is a marine gastropod mollusk widely distributed in the Indo-Pacific region. It is an economically crucial species; however, its population has been declining due to overfishing and overexploitation. In this study, the suitable salinity for juvenile L. canarium was between 20 and 35‰. Diatoms and biological detritus by using flow-water from the fish pool were the most favorable diets for newly metamorphosed and 10 mm juveniles. In the polyculture experiment, L. canarium was cultured with whiteleg shrimp, tilapia, small abalone, purple sea urchin, and collector urchin. Better growth was found in all co-culture groups except with whiteleg shrimp. We also found that the polyculture system with or without substrates significantly affected the growth of juveniles. Additionally, we observed that water temperature was the most crucial factor for growth and survival; a water temperature of less than 10 °C might cause the death of L. canarium. We have proposed a novel polyculture and water-flow method for mass production of L. canarium and evaluated the feasibility and benefits of polyculture with other species. The findings from this work reveal the potentiality of L. canarium in integrated multitrophic aquaculture (IMTA) and its implication for aquaculture and resource restoration.

Abstract Abstract 5102 Juvenile hemochromatosis (JH) is a rare autosomal recessive disorder characterized by the early onset of severe iron overload. A large variety of mutations within the genes encoding hepcidin (HAMP) and hemojuvelin (HJV) have been identified in patients with JH. But in the Chinese population, the prevalence of JH is quite low. No HJV mutation has been reported so far. Methods and results The proband was a 25-year-old young man of Asia decent presented with hypogonadotrophic hypogonadism, diabetes mellitus and heart failure but no family history of iron disorders. His serum iron level was 34μmol/L, with a transferrin concentration of 8.5g/L, serum ferritin concentration was 8140 μg/L. Echocardiography revealed that he had generalized cardiac enlargement, cardiac dysfunction, and severe mitral and tricuspidal insufficiency, pulmonary hypertension. Ultra-sonography showed diffuse hepatomegaly and splenomegaly, seroperitoneum and right hydrothorax. Liver biopsy showed severe diffuse hepatocellular siderosis and cirrhosis,hemosiderin pigmentation. To search for possible variants in the HJV gene, we performed PCR and direct sequencing in proband and his family. A homozygous nonsense mutation in exon 4 of HJV (R329X) was identified in the JH patient and heterozygous mutation of R329X in his father and mother. No mutations were found in other genes associated with adult or juvenile hemochromatosis including HFE, transferrin receptor-2 (TFR2), ferroportin (SLC40A1), hepcidin (HAMP). Conclusions A novel nonsense mutation (R329X) has been identified in the HJV gene for the first time in China. This mutation elevates ferritin levels and leads to JH associated with hypogonadotrophic hypogonadism, diabetes mellitus and severe cardiomyopathy. Disclosures No relevant conflicts of interest to declare.

Exposure to selective serotonin reuptake inhibitors (SSRIs) during development may elicit long-term neuroadaptive changes that could alter the basal regulation of stress-associated physiological and behavioral processes later in life. Currently, the effects of juvenile fluoxetine exposure in rodent models appear to be dependent on the developmental window targeted as well as the duration of drug exposure. The zebrafish (Danio rerio) model is rapidly becoming a useful tool in pharmacological research and can be used to help elucidate some of the long-term effects of fluoxetine exposure prior to sexual maturation on neuroendocrine and behavioral stress markers. In the current study, juvenile zebrafish were chronically exposed to fluoxetine hydrochloride (0 or 100 μg/L) for 14 days (31–44 days post-fertilization (dpf)), then were left untreated until young adulthood. Starting at 90 dpf, basal neuroendocrine stress and behavioral responses of zebrafish were assessed. Cortisol was extracted from the young adult zebrafish body (trunk) and quantified via enzyme-linked immunosorbent assay (ELISA). Anxiety-like behaviors were assessed in response to introduction to the novel tank test. It was expected that juvenile exposure to fluoxetine would (1) reduce basal cortisol levels and (2) elicit anxiolytic effects in the novel tank test in adulthood. However, fluoxetine exposure during the juvenile period was not associated with alterations in basal levels of cortisol nor were there any significant changes in anxiety-like behavior in the young adult zebrafish. Thus, in zebrafish, it does not appear that SSRI exposure during the juvenile period has a long-term adverse or maladaptive impact on the basal expression of cortisol and anxiety-like behavior in adulthood. Further studies are needed to determine if SSRI exposure during this developmental window influences neuroendocrine and behavioral responses to acute stress.

The stocking of predators can have significant consequences on recipient aquatic ecosystems. We investigated some potential ecological impacts of stocking a predatory fish (Lates calcarifer) into a coastal river and a large impoundment in north-eastern Australia. L. calcarifer was mostly found in slower-moving, larger reaches of the river or in the main body of the impoundment where there was abundant suitable habitat. In the tidally influenced freshwater reaches of the coastal river, L. calcarifer predominately consumed aytid and palaemonid shrimp that were associated with local macrophyte beds or littoral grasses. In this area the diets of juvenile stocked and wild L. calcarifer were similar and stocked fish displayed a high degree of site fidelity. Further upstream in the river, away from tidal influence, and in the impoundment, fish were the main prey item. Cannibalism was uncommon and we suggest that, at the current stocking densities, there was little dietary evidence of predatory impacts from L. calcarifer on species of conservation concern. We caution against introducing novel predatory species such as L. calcarifer in or near areas that are outside their natural range and are known to support rare, threatened or endangered species.

SUMMARYAlthough bats are one of the most successful and diverse of mammalian orders, studies that focus upon bat endoparasites are limited. To further knowledge of bat parasitology, pipistrelle bats (Pipistrellus pipistrellus and P. pygmaeus) were acquired from across the Greater Manchester and Lancashire region of England and examined for gastrointestinal helminths using morphological and molecular analyses†. Sixty-eight of 90 adult/juvenile bats (76% prevalence) were infected with at least 1 species of helminth and mean helminth abundance was 48·2 (+/−7·0). All helminths were digenean trematodes and the following species were identified in 51 P. pipistrellus specimens (prevalence in parentheses): Lecithodendrium linstowi (80·4%), L. spathulatum (19·6%), Prosthodendrium sp. (35·3%), Plagiorchis koreanus (29·4%) and Pycnoporus heteroporus (9·8%). Statistical analyses, incorporating multifactorial models, showed that male bats exhibited a significantly more aggregated helminth distribution and lower abundance than females. Positive associations were observed between L. linstowi and L. spathulatum, Prosthodendrium sp. and P. heteroporus and between L. spathulatum and P. koreanus. A revised phylogeny of bat-associated Lecithodendriidae, incorporating novel L. spathulatum and Prosthodendrium sp. 28S rRNA sequences, separated the controversial clade formed by L. linstowi and P. hurkovaae. Further studies are likely to assist the understanding of bat-parasite/pathogen relationships, helminth infracommunity structures and phylogenetics.

Juvenile polyposis syndrome (JPS) is a rare genetic disorder characterized by juvenile polyps of the gastrointestinal tract. We present a new pathogenic mutation of theSMAD4gene and illustrate the need for a multidisciplinary health care approach to facilitate the correct diagnosis. The patient, a 47-year-old Caucasian woman, was diagnosed with anaemia at the age of 12. During the following 30 years, she developed numerous gastrointestinal polyps. The patient underwent several operations, and suffered chronic abdominal pain, malnutrition, and multiple infections. Screening of theSMAD4gene revealed a novel, disease-causing mutation. In 2012, the patient suffered hypoalbuminemia and a large polyp in the small bowel was found. Gamma globulin was given but the patient responded with fever and influenza-like symptoms and refused more treatment. The patient underwent surgery in 2014 and made an uneventful recovery. At follow-up two months later albumin was 38 g/L and IgG was 6.9 g/L. Accurate diagnosis is essential for medical care. For patients with complex symptomatology, often with rare diseases, this is best provided by multidisciplinary teams including representatives from clinical genetics. Patients with aSMAD4mutation should be followed up both for JPS and haemorrhagic hereditary telangiectasia and may develop protein loosing enteropathy and immunodeficiency.

SUMMARYThe efficacy of flocoumafen, a novel anticoagulant rodentide, was evaluated in feeding tests on confined and free-living populations of house mice (Mus musculus L.). In four pen trials, family groups of laboratory-reared wild mice were conditioned to feeding on plain foods and then offered flocoumafen at 0.005% in pinhead oatmeal bait. All 68 mice, comprising juvenile and adult animals, died within 10 days.Ten field trials were carried out, using the same formulated poison bait, against mice infesting farm buildings. Mean treatment success, estimated from live-capture and mortality data, ranged between 87–1 and 100%.The performance of flocoumafen is compared with that of difenacoum, bromadiolone and brodifacoum used at the same concentration in oatmeal bait. Flocoumafen gave an equally effective but quicker kill of mice. It is concluded that flocoumafen is a promising new rodenticide for the control of M. musculus.

Abstract Composite five to seven-day sample s of chlorinated and unchlorinated primary-treated municipal wastewater were collected at the Iona Island treatment plant during a 62-day exposure of juvenile chinook salmon (Oncorhynchus tshawytscha). No differences between chlorinated and unchlorinated samp les were detectable and only 9 chlorinated extractables we re identified. Mass spectrometric analysis of sewage and sludge extracts identified 100 base/neutral components and 60 acidic substances. Some major constituent s we re quantified. Fatty acids, petroleum hydrocarbons, aromatic acids and chemical disinfectants we re predominant. Toxic compounds present included chlorophenols, polynuclear aromatic hydrocarbons (PAH’s) and nonylphenols plus nonylphenolethoxylates. Tetrachlorophenol (TCP) and pentachlorophenol (PCP) reached maximum levels of 7.8 and 13.2 μg · L−l respectively. The PAH’s we re heavily concentrated in sludge samples. Nonylphenol was present in wastewater and sludge but the corresponding ethoxylates occurred only in wastewater. PCB’s were detectable only in sludge. Some novel identifications included two substituted monochiorophenol disinfectants and two generic drugs.

Farmed fish are typically reared at densities much higher than those observed in the wild, but to what extent crowding results in abnormal behaviours that can impact welfare and stress coping styles is subject to debate. Neophobia (i.e. fear of the ‘new’) is thought to be adaptive under natural conditions by limiting risks, but it is potentially maladapted in captivity, where there are no predators or novel foods. We reared juvenile Nile tilapia ( Oreochromis niloticus ) for six weeks at either high (50 g l −1 ) or low density (14 g l −1 ), assessed the extent of skin and eye darkening (two proxies of chronic stress), and exposed them to a novel object in an open test arena, with and without cover, to assess the effects of density on neophobia and stress coping styles. Fish reared at high density were darker, more neophobic, less aggressive, less mobile and less likely to take risks than those reared at low density, and these effects were exacerbated when no cover was available. Thus, the reactive coping style shown by fish at high density was very different from the proactive coping style shown by fish at low density. Our findings provide novel insights into the plasticity of fish behaviour and the effects of aquaculture intensification on one of the world's oldest farmed and most invasive fish, and highlight the importance of considering context. Crowding could have a positive effect on the welfare of tilapia by reducing aggressive behaviour, but it can also make fish chronically stressed and more fearful, which could make them less invasive.

Knowledge of dormancy traits are important in peach breeding. Traditional method selection of seedlings takes a long time because of the juvenile period of plants; therefore, novel application of marker assisted selection methods are needed to accelerate this work. The aims of this study were to test the extent of variability in the PpSOC1 gene among 16 peach cultivars and to establish whether the variability of SOC1 can be used as a functional marker for the timing of endodormancy break based on a 14-year phenology evaluation covering nine consecutive phenology phases, from string stage to ripening. Based on an SSR motif of SOC1, three allele categories were detected: one peach cultivar was heterozygous (203/209), while five of the 15 homozygous cultivars carried a 203 bp allele and the remainder were characterized with 218 bp. There were significant correlations between the PpSOC1 alleles and the various phenology phases, the strongest one being observed at the string stage, marking the end of endodormancy. At this stage, PpSOC1 explained 82.6% of the phenotypic variance; cultivars with the 203 allele reached the string stage 11.7 days earlier than those with 218 bp allele. This finding makes the PpSOC1 screening a valuable method in breeding.

Until recently comic strips were predominantly categorized as either juvenile distraction or some odd adult enthusiasts' hobby. The genre experienced a minor revolution in the 1990s when on the one hand the mass visual media began to explore its rich potential whereas on the other hand the medium's ability to offer "tremendous resources to all writers and artists" (McCloud 212) came under scrntiny, prompting authors like Art Spiegelman to wage an experiment. His biographical Holocaust graphic novel MAUS l and II (1986, 1991) became a bestseller and Pulitzer Prize winner. The paper looks into its 2003 Slovenian edition from the point of view of the undividable entity of drawing and lettering within a panel. It also touches upon certain translation solutions - how closely they correspond to the source text in terms of syntax and transfer of information - but it is not a detailed contrastive analysis as such.

Occurrence and localization of novel antimicrobial and antifeeding compounds in wheat, 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA ) and 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA), and their glucosides, were examined by staining wheat plants ( Triticum aestivum L.) in the juvenile stage of growth by ferric chloride. The methanol extracts of the stained plant tissues gave a characteristic blue color, which was shown by spectroscopic and chromatographic analyses to be exclusively due to benzoxazinones. When ferric chloride was applied to the root in the seedlings, the blue color immediately developed, the staining being strongest at the tip region and becoming lighter towards the basal part. The staining pattern of the radicle in the pre-emerging seed was similar to that in the root, but the coleorhiza was not stained. Little staining was observed in the epidermal layer of the leaf sheath in the shoot but the underlying tissue was stained strongly. The foliage leaf folded in the sheath was also stained, but less intense than the sheath tissue. It is suggested that the DIBOA and DIMBOA are produced within the stained region of the leaf and root. Together with previous findings that the benzoxazinones appear constitutively in wheat during the juvenile stage of growth, their localized occurrence in the tissues exposed to microbial and insect attacks suggests that they act as defense compounds during this vulnerable plant stage.

Background:Higher dosage regimes for Infliximab (IFX) have been described to be effective in partial- or non-responding adults and children with rheumatic disease and appear to be safe (1,2). To optimize IFX treatment in juvenile idiopathic arthritis (JIA) patients, therapeutic drug monitoring (TDM) might be beneficial. To support routine TDM of IFX and dose regimen optimization in JIA patients, more in-depth knowledge of the pharmacokinetic (PK) variability of IFX is needed. Ultimately, as soon as the optimal therapeutic drug ranges will be known, PK model-based simulation can be used to individualize drug dosing recommendations. Individual dosages may be adjusted by taking specific patient characteristics into account that explain inter-patient variability in pharmacokinetics (PK). Inter-patient variability can be quantified and investigated by the population approach.Objectives:Our hypothesis is that optimizing dosage and frequency of IFX administration for individual patients will improve treatment outcome. In this current study, the population PK for IFX are described for JIA patients.Methods:Data including IFX trough concentrations and anti-IFX antibodies of 27 JIA patients on IFX maintenance treatment were retrieved from electronic charts. Three population pharmacokinetic models from literature were validated for our dataset using nonlinear-mixed effects modeling program NONMEM (3,4,5). A novel population pharmacokinetic model was developed based on our study data.Results:A total of 65 obtained blood samples after a median of 32 days after the last IFX infusion (IQR 28-42) were analyzed. The three published models underpredicted the observed trough concentrations. A newly developed one compartment model best described the IFX serum concentration over time data in JIA patients (see Figure 1).Conclusion:Our study shows a novel and the first PK model for IFX in JIA patients. Our main finding was that a one- compartment model best described the IFX serum concentration over time data. Predictive performance of the known models from literature was insufficient for our patient data. Our data also show that different PK models are needed for different age categories (children or adults) and in different diseases.References:[1]Nozaki Y et al. Infliximab dose adjustment can improve the clinical and radiographic outcomes of rheumatoid arthritis patients: REVIVE study results. Biologics. 2018.[2]Tambralli A et al. High doses of infliximab in the management of juvenile idiopathic arthritis. J Rheumatol. 2013.[3]Fasanmade AA et al. Pharmacokinetic properties of infliximab in children and adults with Crohn’s disease: a retrospective analysis of data from 2 phase III clinical trials. Clin Ther. 2011.[4]Xu Z et al. Population pharmacokinetics of infliximab in patients with ankylosing spondylitis. J Clin Pharmacol. 2008.[5]Ternant D et al. Relationship between inflammation and infliximab pharmacokinetics in rheumatoid arthritis. Br J Clin Pharmacol. 2014.Table 1.Patient characteristicsN=27 patientsWeight (kg)52 [40 – 62]BSA1.6 [1.3 – 1.8]Age (years)14 [11 – 17]Male, N (%)6 (22%)CRP (mg/L)0.5 [0.2 – 0.7]WBC count6.5 [5.6 –7.5]Antibodies-to-IFX1 (4%)Dose (mg)300 [200-400]Dose (mg/kg)5.4 [4.9 -7.0]Hemoglobine (mmol/L)7.9 [7.5 -8.2]ESR (mm/h)5 [5-8]Uveitis3 (11%)Disclosure of Interests:None declared.

ABSTRACT During April–October 2019, the West Valley Mosquito and Vector Control District (Ontario, CA) deployed large numbers of In2Care® mosquito traps in a preliminary study to evaluate the trap's potential effectiveness at controlling invasive Aedes aegypti (L.) and Ae. albopictus (Skuse) in 6 cities of San Bernardino County, CA. The trap was used to attract ovipositing females, expose them to the juvenile hormone mimic pyriproxyfen and the entomopathogenic fungus Beauveria bassiana, and autodisseminate pyriproxyfen to other water sources prior to their death from fungal infection. The trap attracted Ae. aegypti and Culex quinquefasciatus, with the latter species predominating at much higher larval densities in the trap reservoirs. Field-collected larvae and pupae from the trap reservoirs showed complete adult emergence inhibition. Furthermore, the trap reservoirs retained high levels of residual larvicidal, pupicidal, and emergence inhibition activity after they were retrieved from the field, as indicated by laboratory bioassays against laboratory colony of Cx. quinquefasciatus. Results of this study support more detailed quantitative local evaluations on trap efficacy to measure the impact of the In2Care mosquito trap on wild invasive Aedes and Culex populations in future mosquito control efforts.

Abstract Juvenile myelomonocytic leukemia (JMML) is a mixed myelodysplastic /myeloproliferative disorder (MDS/MPD) of infancy and early childhood. It is characterized by monocytosis, leukocytosis, elevated fetal hemoglobin, hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF), a low percentage of myeloblasts in the bone marrow, and absence of the Philadelphia chromosome or the BCR/ABL fusion gene. The pathogenesis of JMML has been clearly and definitively linked to dysregulated signal transduction through the RAS signaling pathway. A series of studies conducted over the last decade have shown that mutations or other abnormalities in RAS, NF1, and PTPN11, are potentially responsible for the pathogenesis of JMML in up to 75% of cases. Treatment has been very difficult. There is no effective therapy for JMML. Only allogeneic stem cell transplantation (SCT) can extend survival. However, the relapse rate from allogeneic SCT is inordinately high in JMML (28–55%), with 5-year disease-free survival rates of 25-40%. Rapamycin is a macrolide antibiotic with established clinical applications in organ transplantation. Recent studies have proved that the Mammalian Target of Rapamycin (mTOR) plays an important role in cytokine receptor signaling and induction of apoptosis. Numerous studies have suggested that mTOR functions as a nutritional checkpoint and is connected to energy sensing through AMP-dependent kinase (AMPK) which senses the AMP: ATP ratio in cells. Its function is regulated by the RAS/PI3-kinase pathway. In searching for novel mechanistically-targeted reagents to treat JMML, we conducted an in vitro pilot study with JMML cells. The CFU-GM formation assay was used to test the therapeutic sensitivity of rapamycin to JMML cells. Mononuclear cells (MNCs) from peripheral blood of 9 JMML patients were collected and plated on 0.3% agar medium with rapamycin at a concentration of 1-8nM(0.91-7.28μg/L) and carrier (DMSO). Greater than 50% inhibition of spontaneous CFU-GM growth was observed in all cultures in a dose-dependent fashion, with the exception of one patient sample which had colonies resistant to rapamycin. The effective concentrations in our cultures are equivalent to the safe and tolerable whole blood concentrations achieved in organ transplant patients in clinical settings (5-30μg/L). Our data suggests that rapamycin may be considered as a potentially safe and effective reagent to treat JMML, but that in vitro sensitivity testing might be recommended since one patient sample demonstrated complete resistance to rapamycin in vitro. Further studies are ongoing to explore the mechanism of rapamycin in inhibiting hypersensitivity of JMML cells to GM-CSF.

Infections caused by Fasciola species are widely distributed in cattle and sheep causing significant economic losses, and are emerging as human zoonosis with increasing reports of human cases, especially in children in endemic areas. The current treatment is chemotherapeutic, triclabendazole being the drug of preference since it is active against all parasite stages. Due to the emergence of resistance in several countries, the discovery of new chemical entities with fasciolicidal activity is urgently needed. In our continuous search for new fasciolicide compounds, we identified and characterized six quinoxaline 1,4-di-N-oxide derivatives from our in-house library. We selected them from a screening of novel inhibitors against FhCL1 and FhCL3 proteases, two essential enzymes secreted by juvenile and adult flukes. We report compounds C7, C17, C18, C19, C23, and C24 with an IC50 of less than 10 µM in at least one cathepsin. We studied their binding kinetics in vitro and their enzyme-ligand interactions in silico by molecular docking and molecular dynamic (MD) simulations. These compounds readily kill newly excysted juveniles in vitro and have low cytotoxicity in a Hep-G2 cell line and bovine spermatozoa. Our findings are valuable for the development of new chemotherapeutic approaches against fascioliasis, and other pathologies involving cysteine proteases.

10031 Background: Hematopoietic stem cell transplantation (HSCT) is the only curative therapy for JMML patients (pts). Novel therapies controlling the disorder prior to HSCT are urgently needed. A phase 2, multicenter, open-label study was conducted to evaluate safety and anti-leukemia activity of AZA monotherapy prior to HSCT in pts with newly diagnosed (ND) JMML. Methods: AZA (75 mg/m2 IV) was administered once daily on days 1–7 of each 28-day cycle (C). Primary endpoint was number of pts with clinical complete remission or clinical partial remission (cPR) at C3 day (D) 28 (C3D28). Results: 18 JMML pts (13 PTPN11-, 3 NRAS-, 1 KRAS-, 1 NF1-mutated) were enrolled from 09/2015 to 11/2017. Median (range) white blood cell and platelet (Plt) counts were 19.7 (4.3–59.0) × 109/L and 28 (7–85) × 109/L, respectively. DNA methylation class (MC) was high, intermediate (int), or low in 11, 5, and 2 pts, respectively. 16 pts completed C3 and 5 pts C6. 2 pts discontinued treatment (Tx) pre-C3D28 due to disease progression (PD). 6 pts (33%) had ≥ 1 grade (Gr) 3–4 manageable adverse event (AE) related to AZA. Most common Gr 3–4 AEs related to AZA were neutropenia (2) and anemia (2). 11 pts (61%) were in cPR at C3D28; 7 had PD at C3D28 or prior. All 7 pts of the int/low MC and 4/11 in high MC achieved cPR. 17 pts received HSCT at median of 58 days (37–518) from last AZA dose; 14 were leukemia-free at a median follow-up of 15.7 months (0.1–31.7) after HSCT. 2 pts (high MC) given HSCT relapsed after allograft. 16/18 pts were alive at a median follow-up of 19.8 months (2.6–37.3) from diagnosis. 1 pt discontinuing Tx prior to C3 died from PD; 1 non-responder died from transplant-related causes. Plt response in pts with cPR prompted retrospective comparison of Plt counts at time of HSCT with a historical registry control cohort. Pts with NF1-mutated JMML with higher Plt counts versus other genetic subtypes were excluded. While 7/16 (44%) study pts had Plt counts ≥ 100 × 109/L at HSCT, only 10/58 (17%) historical cohort pts reached this cutoff ( P < 0.01). Conclusions: This study shows that AZA monotherapy was well tolerated in pts with ND JMML. Although the long-term advantage of AZA Tx remains to be fully assessed, responses show it was effective in JMML and provided clinical benefit to pts in this study. Clinical trial information: NCT02447666.

Plants forming a rosette during their juvenile growth phase, such as Arabidopsis thaliana (L.) Heynh., are able to adjust the size, position and orientation of their leaves. These growth responses are under the control of the plants circadian clock and follow a characteristic diurnal rhythm. For instance, increased leaf elongation and hyponasty – defined here as the increase in leaf elevation angle – can be observed when plants are shaded. Shading can either be caused by a decrease in the fluence rate of photosynthetically active radiation (direct shade) or a decrease in the fluence rate of red compared with far-red radiation (neighbour detection). In this paper we report on a phenotyping approach based on laser scanning to measure the diurnal pattern of leaf hyponasty and increase in rosette size. In short days, leaves showed constitutively increased leaf elevation angles compared with long days, but the overall diurnal pattern and the magnitude of up and downward leaf movement was independent of daylength. Shade treatment led to elevated leaf angles during the first day of application, but did not affect the magnitude of up and downward leaf movement in the following day. Using our phenotyping device, individual plants can be non-invasively monitored during several days under different light conditions. Hence, it represents a proper tool to phenotype light- and circadian clock-mediated growth responses in order to better understand the underlying regulatory genetic network.

Abstract Background Juvenile myelomonocytic leukemia(JMML) has usually poor response to chemotherapy, and approximately 50% of patients relapse after hematopoietic stem cell transplantation (HSCT). Recent studies have highlighted the importance of epigenetic aberrations in JMML and proved that some JMML stem cells were associated with hypermethylation. Hence, we desiged the current study to investigate whether low dose Decitabine could improve outcomes of JMML-HSCT.We have reported the preliminary results of low-dose decitabine in the treatment of children with JMML in 2017 ASH as a poster(see blood 2017 130:3232). Then, we will report our latest study. Patients and method 27 patients received HSCT combined with Decitabine between December 2014 and July 2018. Of them,6 patients with NF-1 mutation,11 with PTPN11 mutation, 2 with Kras somatic mutation ,1 with Nras somatic mutation, 3 with multiple mutation (PTPN11+NF-1),2 with monosomy 7,and 3 with uncertain mutation. The median age at diagnosis was 24 months (range: 1-72 months). 26/27 patients received 1~4 course mild chemotherapy(one patient,case 6, received only single course Decitabine therapy)before HSCT.3 patients received HSCT from HLA matched unrelated donors(MUD),and 24 patients received the complementary transplantation(CT), i.e. unrelated cord blood(UCB) was given at day 6 after haploidentical peripheral blood stem cell transplantation(PBSCT) using high dose cyclophosphamide(Cy) post-transplant (PTCy), (see blood 2016 128:1235). Conditioning regimen was composed of Cy, Busulfan (Bu)/ Thiotepa (TT), Fludarabine (Flu) and ATG-F in the MUD-HSCT, and Cy, Bu/TT, Flu and Cytarabine in the CT. Patients received a fixed dose of 8×108/kg mononuclear cells(MNC) in the MUD-HSCT, and a median dose of 45.5×108/kg (range, 26.8~88×108/kg) mobilized peripheral blood MNCs and a median dose of 8.9×107/kg (range, 4.0~12.8×107/kg) UCB nucleated cells in the CT, respectively. GVHD prophylaxis consisted of PTCy, Mycophenolate Mofetil (MMF) and Tacrolimus in the CT, and Thymoglobuline, CsA and MMF in the MUD-HSCT. Decitabine was administrated for 2~4 courses (20mg/m2.d×5 day for each course with 4-week interval) before HSCT to reduce load of leukemia cells and for 2~4 courses (5~10mg/m2.day×5day for each course with the interval of 4~6 weeks) after HSCT to overcome immune-escape of leukemia cells. Results: The median follow-up time was 13months (range, 2-51 months). Full donor cells were engrafted in all patients (donor cell engraftment in case 6 occurred in a salvaged transplant from another haplo-donor after primary failure of first CT).The Overall survival(OS) and Disease-free survival(DFS) was 89.4% and 87.3% respectively. In the CT, haplo-cells and UCB-cells were engrafted in 10 and 14 patients, respectively. The median time to neutrophil more than 0.5x109/L was 31days (range,12~71 days) and 17 days (range, 12~35 days)post-transplant, and to platelet more than 20 x109/L was 22 days (range,9~105 days) and 12 days (range,10~30 days) post-transplant, respectively, in the CT and the MUD- HSCT. All the 3 patients with relapse were haploid-engrated. Two of the three patients with relapse had underwent secondary CT. One of them was Disease-free survival ,and the other died of viral encephalitis(HHV-6) after secondary CT. The cumulative incidence of grades Ⅱ-Ⅳ acute GVHD (aGVHD) was 25.9% (7/27 patients). Case 6 had grade III aGVHD. A case died of grade IV aGVHD(gut) 50 days after the CT. Chronic GVHD(cGVHD) occurred in 5 patients, and no cGVHD more than grade II (NIH criterion) occurred in all patients. The most common complication associated with HSCT was infection. The cumulative incidences of infection plus reactivation of CMV,EBV and HHV-6 were 30% (7/27),3.7%(1/27) and 11.1%(3/27), respectively. Recoverable serious pancytopenia occurred in 3 patients with Decitabine therapy post-HSCT. Conclusion: The combination of hypomethylation agent with HSCT still showed satisfactory results in JMML-HSCT when the follow-up time has been extended for one year. A large-cohort study with extending follow-up time should be developed continuously in the future and the interim results of five-year follow-up time will be reported. Disclosures No relevant conflicts of interest to declare.

Acclimation to low O2 in many organisms involves changes at the level of the transcriptome. Here we used high-throughput RNA sequencing (RNA-Seq) to explore the global transcriptomic response and specific involvement of a suite of hemocyanin (Hc) subunits to low O2 alone and in combination with high CO2, which naturally co-occurs with low O2. Hepatopancreas mRNA of juvenile L. vannamei exposed to air-saturated water, low O2, or low O2/high CO2 for 4 or 24 h was pooled, sequenced (HiSeq 2500) and assembled (Trinity: 52,190 contigs) to create a deep strand-specific reference transcriptome. Annotation of the assembly revealed sequences encoding the previously described small Hc subunit (HcS), and three full-length isoforms of the large subunit (HcL1-3). In addition to this, a previously unidentified full-length Hc subunit was discovered. Phylogenetic analysis demonstrated the subunit to be a β-type Hc subunit (denoted HcB), making this the first report of a β-type hemocyanin subunit in the Penaeoidea. RNAs of individual shrimp were sequenced; regulated genes identified from pairwise comparisons demonstrated a distinct pattern of regulation between prolonged low O2 and low O2/high CO2 treatments by GO term enrichment analysis (Roff-Bentzen, P < 0.0001), showcasing the stabilization of energetically costly translational machinery, mobilization of energy stores, and downregulation of the ubiquitin/proteasomal degradation machinery. Exposure to hypoxia for 24 h resulted in an increase in all of the full-length hemocyanin subunits (HcS, HcL1, HcL2, HcL3, and HcB). The addition of CO2 to hypoxia muted the transcriptomic response of all the Hc subunits to low O2, except for the β-type subunit.

Abstract Introduction: Juvenile myelomonocytic leukemia (JMML) is a rare myeloproliferative neoplasm (MPN) that occurs during childhood and has a poor prognosis. Somatic or germline mutations in canonical RAS pathway genes, i.e., PTPN11, NF1, NRAS, KRAS, and CBL, are reported be detected in approximately 85% patients. Hematopoietic stem cell transplantation (HSCT) is the only curative therapy for JMML. Although spontaneous remission is occasionally observed in others with supportive therapy, some patients show aggressive disease progression despite HSCT. Recent studies have identified several additional genetic events in an array of genes, including SETBP1 and JAK3, but the relationship between genetic alterations and clinical outcomes remains unclear. Patients and Methods: A total of 131 patients (88 boys, 43 girls) with JMML were enrolled in the study. The median age was 15 months (range, 1-160 months). Eighty-two of 131 patients underwent HSCT, and 36 patients died (disease related, n = 27, transplantation-related complications, n = 16, infection, n = 5, unknown, n = 3). We performed comprehensive genetic analyses of the 131 JMML patients using whole-exome sequencing (n = 68, 52%) or targeted deep sequencing (n = 92, 70%), and assessed the impact of genetic alterations on clinical outcomes in 119 patients, excluding 12 patients with Noonan syndrome-related myeloproliferative disorder (NS/MPD). Results: We identified canonical RAS pathway gene mutations in 115 of 131 patients (88%). Although most RAS pathway mutations were mutually exclusive, coexisting secondary RAS pathway mutations were found in nine patients (8%). In addition, 28 patients harbored secondary genetic alterations in other genes, including SETBP1 (n = 10), JAK3 (n = 12), ASXL1 (n = 6), SH3BP1 (n = 1), RRAS2 (n = 2), and SOS1 (n = 3). In total, 34 of 131 patients harbored secondary genetic mutations. In univariate analysis, patients with secondary genetic mutations showed poorer survival rates than patients without these mutations [5-year transplantation-free survival (TFS) (95% CI) = 8.8% (2.3%-21.1%) vs. 24.1% (15.2%-34.1%), p = 0.007; 5-year overall survival (OS) (95% CI) = 49.6% (32.0%-65.0%) vs. 62.3% (50.8%-71.8%), p = 0.135]. On the basis of the dominant canonical RAS pathway mutations classification, patients with PTPN11 and NF1 mutations were significantly associated with the presence of secondary genetic mutations compared to patients with other RAS pathway gene mutations (PTPN11 (20 of 43, 47%), NF1 (5 of 7, 71%), NRAS (2 of 18, 11%), KRAS (4 of 20, 20%), CBL (1 of 17, 6%), p < 0.001). Consistent with previous reports, patients with PTPN11 and NF1 mutations had inferior survival rates than other JMML patients [5-year TFS (95% CI) = 0% vs. 32.7% (21.5%-44.3%), p < 0.001; 5-year OS (95% CI) = 45.3% (31.1%-58.5%) vs. 68.1% (55.2%-78.0%), p = 0.006]. Multivariate survival analysis identified the RAS pathway mutations (i.e., patients with PTPN11 and NF1 mutations vs. others) [TFS: HR (95% CI) = 3.732 (2.382-5.847), p < 0.001; OS: HR (95% CI) = 1.983 (1.117-3.521), p < 0.019] and low platelet count (<33 × 109/L vs. ≥ 33 × 109/L) [TFS: HR (95% CI) = 1.816 (1.160-2.843), p < 0.001] as independent risk factors for TFS and OS. In subgroup analysis of 50 patients with PTPN11 and NF1 mutations, there were no significant survival differences between patients with (n = 25) or without (n = 25) secondary genetic mutations [5-year TFS (95% CI) = 0% vs. 0%, p = 0.753; 5-year OS (95% CI) = 39.6% (20.8%-57.9%) vs. 51.7% (30.9%-69.1%), p = 0.589]. Discussion: Consistent with previous studies, secondary genetic mutations were associated with inferior survival rates, but high correlations were observed in JMML patientswith PTPN11 and NF1 mutations. Our results suggest that comprehensive genetic mutational profiling is essential to estimate prognosis and to stratify JMML patients who require HSCT and/or novel treatment modalities. Disclosures Ogawa: Sumitomo Dainippon Pharma: Research Funding; Takeda Pharmaceuticals: Consultancy, Research Funding; Kan research institute: Consultancy, Research Funding. Kojima:SANOFI: Honoraria, Research Funding.

Krüppel-homolog 1 (Kr-h1) is a zinc finger transcription factor maintaining the status quo in immature insect stages and promoting reproduction in adult insects through the transduction of the Juvenile Hormone (JH) signal. Knockdown studies have shown that precocious silencing of Kr-h1 in the immature stages results in the premature development of adult features. However, the molecular characteristics and reproductive potential of these premature adult insect stages are still poorly understood. Here we report on an adult-like or ‘adultoid’ phenotype of the migratory locust, Locusta migratoria, obtained after a premature metamorphosis induced by the silencing of LmKr-h1 in the penultimate instar. The freshly molted adultoid shows precocious development of adult features, corresponding with increased transcript levels of the adult specifier gene LmE93. Furthermore, accelerated ovarian maturation and vitellogenesis were observed in female adultoids, coinciding with elevated expression of LmCYP15A1 in corpora allata (CA) and LmKr-h1 and vitellogenin genes (LmVg) in fat body, whereas LmE93 and Methoprene-tolerant (LmMet) transcript levels decreased in fat body. In adultoid ovaries, expression of the Halloween genes, Spook (LmSpo) and Phantom (LmPhm), was elevated as well. In addition, the processes of mating and oviposition were severely disturbed in these females. L. migratoria is a well-known, swarm-forming pest insect that can destroy crops and harvests in some of the world’s poorest countries. As such, a better understanding of factors that are capable of significantly reducing the reproductive potential of this pest may be of crucial importance for the development of novel locust control strategies.

Background:Aberrant T cell responses are key in driving autoimmunity and are commonly associated with rheumatoid arthritis (RA). Unravelling pathways of importance in therapeutic partial response and failure is of critical importance, as this will potentially provide new insights into key drivers of immune-mediated pathogenesis.Objectives:To delineate disease-relevant T cell subsets in RA and assess their potential to act as cellular markers amenable to precision medicine approaches, particularly in the context of therapeutic partial or non-response.Methods:FACS-based immunophenotyping and ex-vivo functional response profiles of CD4+CD161+CCR2+CCR5+T cells were performed in peripheral blood mononuclear cells (PBMC) obtained from patients with RA and healthy controls, using previously characterised methodologies. RA patients fulfilled the 2010 ACR/EULAR criteria for RA. All samples were obtained after written consent, with the appropriate ethical approvals in place.Results:RA patients harboured a higher frequency of CCR2+CCR5+cells within the CD4+CD161+T cell compartment compared with healthy controls. In RA patients this T cell subset had a higher proportion of cells that secrete pro-inflammatory cytokines such as IL-17A, GM-CSF, IFN-γ, and TNF. Importantly, the CD4+CD161+CCR2+CCR5+T cell subset was significantly increased in DMARD non-responders compared to both responders and healthy controls. Moreover, in DMARD non-responders, these cells had a propensity to express increased proportions of pro-inflammatory cytokines. Notably, there was also a significant increase in the ratio of effector: regulatory T cell (Teff: Treg) compared to both responders and healthy controls. In addition, the CD4+CD161+CCR2+CCR5+T cell subset was less responsive to suppression by Tregs. In further support of a role for this T cell population in disease pathogenesis, the frequency of CD4+CD161+CCR2+CCR5+T cells significantly correlated with disease activity, as measured by the DAS28 (R2= 0.65; p = 0.003; n=11).Conclusion:Combined, our findings suggest that the CD4+CD161+CCR2+CCR5+T cell subset represents a substantially abnormal T cell subset in RA, exhibiting exaggerated pro-inflammatory responses, numerical abundance relative to Tregs, and resistant to regulation by Tregs. The CD4+CD161+CCR2+CCR5+T cell subset appears to be a marker of therapeutic response status in RA, via its contribution to disease pathology and highlights this subset as a potential therapeutic target in RA.References:[1]McInnes IB, Schett G. The pathogenesis of rheumatoid arthritis.N Engl J Med. 2011;365(23):2205-19.[2]Mexhitaj I, Nyirenda MH, Li R, O’Mahony J, Rezk A, Rozenberg A,et al. Abnormal effector and regulatory T cell subsets in paediatric-onset multiple sclerosis.Brain. 2019;142(3):617-32.[3]Cosmi L, Cimaz R, Maggi L, Santarlasci V, Capone M, Borriello F,et al. Evidence of the transient nature of the Th17 phenotype of CD4+CD161+T cells in the synovial fluid of patients with juvenile idiopathic arthritis.Arthritis Rheum. 2011;63(8):2504-15.Disclosure of Interests:Mukanthu Nyirenda: None declared, Iain McInnes Grant/research support from: Bristol-Myers Squibb, Celgene, Eli Lilly and Company, Janssen, and UCB, Consultant of: AbbVie, Bristol-Myers Squibb, Celgene, Eli Lilly and Company, Gilead, Janssen, Novartis, Pfizer, and UCB, Carl Goodyear: None declared

Abstract Polycythemia vera (PV), is a myeloproliferative disease (MPD) originating in a hematopoietic stem cell resulting in clonal expansion of erythroid progenitors. It is associated with thromboses and malignant transformation. Recently the V617F alteration arising from a mutation in JAK2 has been identified in greater than 90% of cases of PV in adults. PV is rare in children and the frequency of the common JAK2 mutation is significantly lower than in adult patients, indicating that alternative genetic events are involved in the pathogenesis of this disease. We have identified a child diagnosed with PV at the age of 15 months, the youngest described in the literature to date. Initial laboratory values demonstrated a WBC of 33 x109/L, hemoglobin 181 g/L, and platelet count of 579 x109/L. Bone marrow cytogenetics were normal. Erythroid colony forming units demonstrated erythropoietin-independent growth. Peripheral blood, buccal swab and saliva analysis revealed the presence of the common JAK2 V617F mutation, but a B lymphocyte cell line and skin-fibroblast-culture from this patient were negative, indicating that the JAK2 mutation was somatic. Peripheral blood from her parents and older brother demonstrated normal blood counts and wild type JAK2 status. This child was also diagnosed with Neurofibromatosis type 1 (NF1) based on meeting NIH consensus diagnostic criteria diagnostic criteria, having the requisite number of appropriately sized café-au-lait macules and Lisch nodules. NF1 and PV have no previously known association, however NF1 is associated with another MPD, juvenile myelomonocytic leukemia (JMML). Patients with NF1 and JMML demonstrate loss of heterozygosity (LOH) at the NF1 locus while 60% of JMML patients without NF1 alternatively demonstrate somatic mutations in NRAS, KRAS2 or PTPN11. Taken together, these genetic lesions result in hyperactivation of the RAS/MAPK pathway. Low density single nucleotide polymorphism arrays performed on peripheral blood from this patient failed to demonstrate obvious LOH at the NF1 locus. NF1 gene sequencing failed to identify the cause of the NF1 phenotype. Mutations were not identified in the commonly mutated regions of NRAS, KRAS2 or PTPN11. This case reveals the presence of the most commonly acquired somatic JAK2 mutation in a young child with PV and indicates that buccal swabs and saliva are unreliable sources of unaffected tissue for assessing the presence of germline mutations in PV patients. Moreover, it suggests that some patients with clinical NF1 are at risk for developing other MPDs besides JMML. For this patient, a novel unidentified genetic abnormality resulting in the clinical phenotype of NF1 may serve as a predisposing genetic event accounting for the unusually young age of presentation. The investigation of rare children with PV has the potential to provide valuable insight into the molecular interactions underlying the pathogenesis of MPDs.

Background At present, allogeneic-HSCT is still the only proven potentially curative strategy for Juvenile myelomonocytic leukemia(JMML) children, being only able to cure about 50% of patients by either a relative donnor or an unrelated volunteer. Relapse represented the main causes of treatment failure. The data of HLA-haploidentical transplantation is poorly reported so far. But we have reported an encouraging result in the treatment of leukemia and Thalassemia with complementary transplantation(CT), i.e. unrelated umbilical cord blood(UCB) following haploidentical stem cells transplantation(see blood 2016 128:1235). The evidence of aberrant DNA methylation in JMML provided a rationale for the use of the DNA hypomethylating agent 5-azacytidine or Decitabine(DAC). Recent studies have indicated hypomethylating agent maybe reduce the disease burden in the pretransplant window with minimal toxicity and play an immunomodulatory role after transplantation. Therefore, we developed a novel CT strategy combinated with hypomethylation agent for children with JMML. Patients and method 48 patients received CT combined with DAC from December 2014 to April 2019 in two independent HSCT-centers. Of them, 44 patients had molecular genetics mutation(including NF1, PTPN11,Kras,Nras),and 6 patients had cytogenetic abnormalities with monosomy 7. The median age at diagnosis was 22 (1 to 90) ms. (Table.1). 47 of 48 patients received 2~6 courses of DAC therapy (20mg/m2/d ×5 days for each course with 4-weeks interval) and 1~2 courses of mild-moderate cytotoxic chemotherapy (according to the"A-triple-V"regimen from Korea or reduced-intensity FlAG regimen) as "Bridging therapy" in order to reduce the tumor burden prior to HSCT. There was only one patient who was accepted one course of DAC therapy prior to HSCT. In addition, low dose DAC was also administrated 2~6 courses (10mg/m2/day for 3 or 5 days for each course with the interval of 4~6 weeks) for all patients in order to overcome immune-escape of leukemia cells post HSCT. None of the patients underwent splenectomy. Conditioning regimen included DAC ,Cyclophosphamide(Cy) ,Busulfan,Fludarabine and Thiotepa (or replaced by Busulfan). GVHD prophylaxis consisted of Cy on day+3,+4, MMF and Tacrolimus from day+6. Noteworthily, UCB was infused on day+6 (Fig.1). Human leukocyte antigen of UCB-donor/recipient pairs had less than 3/10 loci incompatibility.The median number of infusion mononuclear cells from haploid-donor was 37.8×108/kg, and which of infusion nucleated cells from UCB was 8.8×107/kg(Table.1). Results The median and mean follow-up time was 13.5 and 16.2 (3 to 63)ms ,respectively. Full donor cells engrafted in all patients (donor cell engraftment in one case without any cytotoxic chemotherapy before HSCT, who was occurred in a salvaged HSCT from another haplo-donor and UCB-donor after primary failure of first CT ). 5-years OS, disease-free survival(DFS) and transplant-related mortality were 90.1%,86.9% and 4.4%, respectively(Fig.2). Haplo-cells and UCB-cells engrafted finally in 26 and 22 patients, respectively. The median times to neutrophil more than 0.5×109/L and platelet more than 20×109/L were 29 days and 24 days post-HSCT, respectively (Table.2). Platelets and neutrophi recovered earlier in patients with haplo-engrafted than those with UCB-engrafted (p=0.000)(Table.3). Five patients developed relapse after HSCT, who had PTPN11 mutation. 4 of them were Haplo-cells engraftment. The median time to relapse after HSCT was 5.5 (1.5 to 12 ) months. There ware no significant difference in the OS and DFS between UCB-engrafted group and Haplo-engrafted group(p=0.612 and p=0.227,respectively)(Fig.3).The cumulative incidences of grades Ⅱ-Ⅳ aGVHD was 38% (18 pts).The number of patients with grade III and Ⅳ aGVHD were 7 in total. Chronic GVHD occurred in 7 patients, and no chronic GVHD more than grade II (NIH criterion) occurred in all patients(Table.2).The most common complication associated CT was infection. The reactivation incidences of CMV, EBV and human herpes virus 6 were 27% ,13% and 5.0%, respectively. Recoverable serious pancytopenia occurred in four patients with DAC therapy post-HSCT. Summary CT have significantly improved the therapeutic effect in JMML-HSCT.Hypomethylation agent may play an important role in CT. Neither cord blood-engraftment nor haploid-engraftment demonstrated a clear superiority for CT. Disclosures No relevant conflicts of interest to declare.

Planes minutus (L.) (Crustacea: Brachyura) swims well (forwards only) but has limited endurance; at 28°C crabs swam at the surface for 35–45 min before sinking. The legs of the neuston crab are flattened and rotated into the ‘advanced’ orientation of the legs to the cephalothorax, characteristic of swimming brachyurans. All legs are involved in swimming; pairs 1 and 2 mainly support the crab in the water column, pairs 3 and 4 provide forward propulsion too. In leg-pair 4, the plane of extension and flexion is close to the horizontal and the leg provides lift on the recovery stroke. Plumose hairs are present on the anterior borders of the legs, adding to the propulsive area. Stout spines line the anterior and posterior margins of the legs; they allow the crab to hang upside down from floating substrata, but are also used to hold food items. The chelae of Planes have cutting rather than crushing ‘teeth’. Crabs of the size studied (0·128−0·736 g wet wt) used oxygen (at 30°C) at about twice the rate when swimming (mean 1·014 ml O2 g wet wt−1 h1) as did resting crabs (mean rate 0·495 ml O2 g wet wt−1 h1). No oxygen debt is incurred during swimming. The mean crab density was 1·096 g ml−1 (SD 0·060). Neustonic crabs would not move more than 5 cm away from their floating substrata to catch food. They show a novel ‘revving up’ procedure in which leg pairs 1 to 3 beat while the animal is attached to the substratum by spines on the tips of leg-pair 4. This allows much of the locomotor apparatus to overcome inertia and reach working speed, before an attack is carried out. Crabs ate postlarval and juvenile fish, euphausids, isopods, sea skaters and squid. Gelatinous animals were gener-ally avoided, though salps were captured and the stomach contents eaten. Planes will take prey as big as itself and stores food for later consumption.

Abstract Abstract 61 To identify new therapeutic strategies for AML, we compiled and screened an in-house library of on-patent and off-patent drugs to identify agents cytotoxic to leukemia cells. From this screen, we identified mefloquine, an off-patent drug indicated for the treatment and prophylaxis of malaria. In secondary assays, mefloquine decreased the viability of 9/10 human and murine leukemia cell lines (EC50 3.25–8.0 μM). Moreover, it reduced the viability of 4/5 primary AML samples, but was not cytotoxic to normal hematopoietic cells (EC50>31 μM). Importantly, mefloquine reduced the clonogenic growth of primary AML samples, but not normal hematopoietic cells, and completely inhibited engraftment of primary AML cells into immune deficient mice. Finally, systemic treatment with oral mefloquine (50 mg/kg/day) decreased leukemic burden without evidence of toxicity in 4 mouse models of leukemia, including mice engrafted with primary AML cells. Thus, mefloquine effectively targets leukemic cells, including leukemia stem cells, at concentrations that appear pharmacologically achievable and are not toxic to normal hematopoietic cells. To identify the mechanisms of mefloquine-mediated cell death in AML cells, we performed a binary drug combination screen, hypothesizing that drugs that synergized with mefloquine may share overlapping mechanism of action. From this combination screen of 550 drugs, we identified 18 that reproducibly synergized with mefloquine as measured by the Excess over Bliss additivism score, including 3 members of the artemisinin class of anti-malarials: artemisinin, artesunate and artenimol. Strikingly, 10/18 synergistic compounds, including the artemisinins, were known generators of reactive oxygen species (ROS). Therefore we tested mefloquine's ability to increase ROS in leukemic cells. Mefloquine increased ROS production in leukemia cells in a dose- and time-dependent manner. Co-treatment with ROS scavengers α-tocopherol and N-acetyl-cysteine abrogated mefloquine-induced ROS production and cell death, indicating that ROS production was functionally important for mefloquine-mediated cell death. Moreover, the artemisinins induced ROS as single agents, and synergistically increased ROS when combined with mefloquine. To identify cellular target(s) of mefloquine's anti-leukemic effects, we performed a yeast genome-wide functional screen to identify heterozygous gene deletions that rendered yeast more sensitive to mefloquine. 21/37 genes whose depletion conferred >4-fold sensitivity to mefloquine were associated with function of the yeast vacuole, equivalent to the mammalian lysosome. Consistent with these data, fluorescent confocal microscopy demonstrated that mefloquine and artesunate disrupted lysosomes. Cell death after mefloquine and artesunate treatment was caspase-independent and associated with increased incorporation of monodancylcadaverin in autophagosomes, consistent with the effect of these drugs on the lysosomes. To further explore the anti-leukemic activity of lysosomal disruption, we evaluated the anti-leukemic effects of the known lysosomal disrupter L-leucine-leucine methyl ether (LeuLeuOMe). Similar to mefloquine and artesunate, LeuLeuOMe induced cell death in leukemia cells, increased ROS production, and disrupted the lysosomes. Highlighting the potential clinical utility of lysosomal disrupters for the treatment of leukemia, a patient with relapsed/refractory juvenile myelomonocytic leukemia self-administered artemisinin. The artemisinin cleared the circulating blasts from the circulating blasts and the patient proceeded to allotransplant. Finally, to investigate the basis of leukemic cell hypersensitivity to lysosomal disruption, we assessed lysosomal characteristics of primary AML and normal hematopoietic cells. By gene expression analysis, AML patient samples had higher mRNA levels of the lysosomal cathepsins A, B, C, D, H, L, S and Z, compared to CD34+ normal hematopoietic cells, and cathepsins C, D and Z were significantly over-expressed in the LSC compartment, compared to normal HSCs. In summary, our data demonstrate that lysosomal disruption preferentially targets AML cells and AML stem cells through a mechanism related to increased ROS production. Thus, this work highlights lysosomal disruption as a novel therapeutic strategy for AML. Disclosures: Off Label Use: This study includes a case report of off-label use of the anti-malarial artemisinin in the treatment of a case of juvenile myelomonocytic leukemia.

Abstract Abstract 4676 Introduction Myeloproliferative diseases (MPDs) are a group of haematological disorders characterized by the hyper proliferation of different blood cells in peripheral blood and other hematopoietic organs. The clinical heterogenity of these neoplasms reflects the different gene pathways involved, most of which are only partially known. Given the genetic homology and the physiological similarity to mammals, zebrafish has emerged as an ideal model to study human normal and malignant haematopoiesis. In the last decade several oncogenes involved in the development of hematopoietic neoplasms have been used to model leukemia in zebrafish with the aim to discovery new molecular pathways involved in malignant transformation. Despite the first encouraging results these experimental models failed to fully recapitulate human myeloproliferative disorders. Methods We took advantage of the Gal4/UAS binary system to induce the expression of human oncogenic HRASV12G in the zebrafish hematopoietic compartment. We used a specific transgenic line that drives oncogene expression in zebrafish early hematopoietic progenitors under control of the FLI.1 (Friend Leukemia virus Integration 1) promoter. Results We observed the development of a myelo-erythroid proliferative disease in few days in zebrafish transgenic larva. The pathological phenotype is characterized by the expansion of the hematopoietic tissue, an increased expression of myelo-erythroid specific genes (PU.1, gata1, mpx, c-mpl) associated with a slight increase of staminality markers (lmo2, scl, c-myb, runx.1), and a higher number of l-plastin expressing cells. Moreover blood smear of pathological larva displayed leukemic blasts and the arrest of erythrocyte differentiation whereas kidney marrow of juvenile fish displayed abnormal myelopoiesis characterized by the increase of erythro-myeloid progenitors. We found that the pathological phenotype is associated with a down regulation of the Notch pathway as shown by the decreased gene expression of notch pathways target genes (notch1, notch3, her6). Furthermore we discovered a novel set of genes involved in neoplastic transformation induced by HRASV12 expression through RNA-Seq analysis of pathological larva. Conclusions The expansion of the zebrafish hematopoietic compartment characterized by the hyper-proliferation of the myelo-erythroid progenitors that we found in this model reproduces some of the pathological features of human myeloproliferative disorders. This study showed that forcing oncogene expression in the hemogenic endothelial cells induces the transdifferentiation of the early hemogenic pluripotent stem cells into abnormal myeloerythoid progenitors by repressing the Notch pathways. Transcriptome analysis identified a number of potential effectors of this transformation. Disclosures: No relevant conflicts of interest to declare.

Liu, Ying Bing, Gui-Lan Ye, Xue-Song Liu, Joseph F. Pasternak, and Barbara L. Trommer. GABAA Currents in immature dentate gyrus granule cells. J. Neurophysiol. 80: 2255–2267, 1998. We used whole cell patch clamp and gramicidin perforated patch recordings in hippocampal slices to study γ-aminobutyric acid (GABA) currents in granule cells (GCs) from juvenile rat dentate gyrus (DG). GCs are generated postnatally and asynchronously such that they can be detected at different stages of their maturation in DG within the first month. In contrast, inhibitory interneurons are generated embryonically, and their circuitry is well developed even as their target GCs and GC excitatory connections are still being formed. In this study, two GABA currents evoked in GCs by medial perforant path stimulation are compared. The first, pharmacologically isolated by glutamate receptor blockade, is the product of direct activation of GABA interneurons with monosynaptic input to the recorded GC (monosynaptic GABAA). Monosynaptic GABAA displays slight outward rectification of its current-voltage relation, is 97% eliminated by 10 μM bicuculline and coincides temporally with the excitatory components of GC postsynaptic currents as has been described for GABAA currents in other brain regions. The second is a novel GABA response that is detectable in 10 μM bicuculline and is present on GCs only at the earliest stages of their maturation. Unlike monosynaptic GABAA, this transient GABA is eliminated by glutamate receptor blockade and hence is likely to be generated by interneurons activated via an intervening glutamatergic synapse (polysynaptically). It is predominantly chloride mediated, has a relative bicarbonate/chloride permeability ratio of 26%, and is unchanged by bath-applied saclofen and strychnine or by intracellular calcium chelation. It is 97% antagonized by 100 μM picrotoxin and 99% antagonized by 100 μM bicuculline. This current is thus a relatively bicuculline (BMI)-resistant GABAA current (BMIR-GABAA). Compared with monosynaptic GABAA, BMIR-GABAA has a later peak, slower time course of decay, and marked outward rectification. Its reversal potential is 7–8 mV depolarized to that of monosynaptic GABAA whether recorded in whole cell or with gramicidin perforated patch to preserve native internal chloride concentration. Together these data may suggest that BMIR-GABAA is evoked by an anatomically segregated population of interneurons activating a unique, developmentally regulated GABAA receptor. Further, the transient nature of this current coupled with its temporal characteristics that preclude overlap with the excitatory components of the synaptic response are consistent with a role that is trophic or signaling rather than primarily inhibitory.

The pandemic of COVID-19 has afflicted every individual and has initiated a cascade of directly or indirectly involved events in precipitating mental health issues. The human species is a wanderer and hunter-gatherer by nature, and physical social distancing and nationwide lockdown have confined an individual to physical isolation. The present review article was conceived to address psychosocial and other issues and their aetiology related to the current pandemic of COVID-19. The elderly age group has most suffered the wrath of SARS-CoV-2, and social isolation as a preventive measure may further induce mental health issues. Animal model studies have demonstrated an inappropriate interacting endogenous neurotransmitter milieu of dopamine, serotonin, glutamate, and opioids, induced by social isolation that could probably lead to observable phenomena of deviant psychosocial behavior. Conflicting and manipulated information related to COVID-19 on social media has also been recognized as a global threat. Psychological stress during the current pandemic in frontline health care workers, migrant workers, children, and adolescents is also a serious concern. Mental health issues in the current situation could also be induced by being quarantined, uncertainty in business, jobs, economy, hampered academic activities, increased screen time on social media, and domestic violence incidences. The gravity of mental health issues associated with the pandemic of COVID-19 should be identified at the earliest. Mental health organization dedicated to current and future pandemics should be established along with Government policies addressing psychological issues to prevent and treat mental health issues need to be developed. References World Health Organization (WHO) Coronavirus Disease (COVID-19) Dashboard. 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Abstract Objective The objective of this study was to assess the relationship between adalimumab trough concentrations and treatment response in paediatric patients with JIA. Methods This was a monocentric cohort study of JIA patients treated with adalimumab. Clinical data and samples were collected during routine follow-up. Adalimumab trough concentrations were quantified by a novel liquid chromatography–tandem mass spectrometry assay. Anti-adalimumab antibodies were measured in samples with trough concentrations of ≤5mg/l. Disease activity was evaluated using the clinical Juvenile Arthritis DAS with 71-joint count (cJADAS71). Response to adalimumab was defined according to recent international treat-to-target guidelines. Results A total of 35 adalimumab trough samples were available from 34 paediatric patients with JIA. Although there was no significant difference in adalimumab dose, trough concentrations were significantly lower in patients with secondary failure [median 1.0 mg/l; interquartile range (IQR) 1.0–5.3] compared with patients with primary failure (median 13.97 mg/l; IQR 11.81–16.67) or an adequate response (median 14.94 mg/l; IQR 10.31–16.19) to adalimumab. Conclusion Adalimumab trough concentrations were significantly lower in JIA patients with secondary failure compared with patients with primary failure or an adequate response to adalimumab. Our results suggest that trough concentration measurements could identify JIA patients who require increased adalimumab doses to achieve or maintain therapeutic drug concentrations.

Abstract The forest resources are changing due to fire prevention and depletion of old growth forest. Applications for small-diameter juvenile timber, especially for low-value species, such as ponderosa pine (Pinus ponderosa) are needed. In this study, a novel thin wood strand composite of P. ponderosa, 3.2 mm thick, is introduced as a veneer substitute. Optimization was performed for the processing parameters phenol formaldehyde resin content, platen temperature, and the aspect ratio strand length to thickness (L/t). Mean modulus of elasticity and modulus of rupture values of 10.2 GPa and 79.1 MPa, respectively, were obtained with the optimized formulation (5.5% resin, platen temperature 152°C, and aspect ratio of 430). These values were approximately 2–2.5 times higher than the parent small-diameter ponderosa pine lumber. The results indicate that fast-growing low-density species – such as hybrid poplars, or small-diameter timber with predominantly juvenile wood – could potentially be used to manufacture strand veneers of consistent quality with significantly higher strength and stiffness than parent material.

Abstract Background The brain anatomy in the clade Spiralia can vary from simple, commissural brains (e.g., gastrotrichs, rotifers) to rather complex, partitioned structures (e.g., in cephalopods and annelids). How often and in which lineages complex brains evolved still remains unclear. Nemerteans are a clade of worm-like spiralians, which possess a complex central nervous system (CNS) with a prominent brain, and elaborated chemosensory and neuroglandular cerebral organs, which have been previously suggested as homologs to the annelid mushroom bodies. To understand the developmental and evolutionary origins of the complex brain in nemerteans and spiralians in general, we investigated details of the neuroanatomy and gene expression in the brain and cerebral organs of the juveniles of nemertean Lineus ruber. Results In the juveniles, the CNS is already composed of all major elements present in the adults, including the brain, paired longitudinal lateral nerve cords, and an unpaired dorsal nerve cord, which suggests that further neural development is mostly related with increase in the size but not in complexity. The ultrastructure of the juvenile cerebral organ revealed that it is composed of several distinct cell types present also in the adults. The 12 transcription factors commonly used as brain cell type markers in bilaterians show region-specific expression in the nemertean brain and divide the entire organ into several molecularly distinct areas, partially overlapping with the morphological compartments. Additionally, several of the mushroom body-specific genes are expressed in the developing cerebral organs. Conclusions The dissimilar expression of molecular brain markers between L. ruber and the annelid Platynereis dumerilii indicates that the complex brains present in those two species evolved convergently by independent expansions of non-homologous regions of a simpler brain present in their last common ancestor. Although the same genes are expressed in mushroom bodies and cerebral organs, their spatial expression within organs shows apparent differences between annelids and nemerteans, indicating convergent recruitment of the same genes into patterning of non-homologous organs or hint toward a more complicated evolutionary process, in which conserved and novel cell types contribute to the non-homologous structures.