Hybrid Closed Loop Overcomes the Impact of Missed or Suboptimal Meal Boluses on Glucose Control in Children with Type 1 Diabetes Compared to Sensor-Augmented Pump Therapy.

Background: It is unclear whether hybrid closed-loop (HCL) therapy attenuates the metabolic impact of missed or suboptimal meal insulin bolus compared with sensor-augmented pump (SAP) therapy in children with type 1 diabetes in free-living conditions. Methods: This is an ancillary study from a multicenter randomized controlled trial that compared 24/7 HCL with evening and night (E/N) HCL for 36 weeks in children between 6 and 12 years old. In the present study, the 60 children from the E/N arm underwent a SAP phase, an E/N HCL for 18 weeks, then a 24/7 phase for 18 weeks, extended for 36 more weeks. The last 28-30 days of each of the four phases were analyzed according to meal bolus management (cumulated 6817 days). The primary endpoint was the percentage of time that the sensor glucose was in the target range (TIR, 70-180 mg/dL) according to the number of missed boluses per day. Findings: TIR was 54% ± 10% with SAP, 63% ± 7% with E/N HCL, and steadily 67% ± 7% with 24/7 HCL. From the SAP phase to 72 weeks of HCL, the percentage of days with at least one missed meal bolus increased from 12% to 22%. Estimated marginal (EM) mean TIR when no bolus was missed was 54% (95% confidence intervals [CI] 53-56) in SAP and it was 13% higher (95% CI 11-15) in the 24/7 HCL phase. EM mean TIR with 1 and ≥2 missed boluses/day was 49.5% (95% CI 46-52) and 45% (95% CI 39-51) in SAP, and it was 15% (95% CI 14-16) and 17% higher (95% CI 6-28), respectively, in the 24/7 HCL phase (P < 0.05 for all comparisons vs. SAP). Interpretation: HCL persistently improves glycemic control compared with SAP, even in case of meal bolus omission. ClinicalTrials.gov (NCT03739099).

Sublethal effects of metal toxicity and the measure of plant fitness in ecotoxicological experiments.

Anthropogenic pollution is a major driver of global environmental change. To be properly addressed, the study of the impact of pollutants must consider both lethal effects and sublethal effects on individual fitness. However, measuring fitness remains challenging. In plants, the total number of seeds produced, i.e. the seed set, is traditionally considered, but is not readily accessible. Instead, performance traits related to survival, e.g., vegetative biomass and reproductive success, can be measured, but their correlation with seed set has rarely been investigated. To develop accurate estimates of seed set, relationships among 15 vegetative and reproductive traits were analyzed. For this purpose, Noccaea caerulescens (Brassicaceae), a model plant to study local adaptation to metal-contaminated environments, was used. To investigate putative variation in trait relationships, sampling included several accessions cultivated in contrasting experimental conditions. To test their applicability, selected estimates were used in the first generation of a Laboratory Natural Selection (LNS) experiment exposing experimentally plants to zinc soil pollution. Principal component analyses revealed statistical independence between vegetative and reproductive traits. Traits showing the strongest positive correlation with seed set were the number of non-aborted silicles, and the product of this number and mean silicle length. They thus appeared the most appropriate to document sublethal or fitness effects of environmental contaminants in plant ecotoxicological studies. The relevance of both estimates was confirmed by using them to assess the fitness of parental plants of the first generation of an LNS experiment: the same families consistently displayed the highest or the lowest performance values in two independent experimental metal-exposed populations. Thus, both these fitness estimates could be used to determine the expected number of offspring and the composition of successive generations in further LNS experiments investigating the impact of multi-generational exposure of a plant species to environmental pollution.

Outcomes of hybrid closed-loop insulin delivery activated 24/7 versus evening and night in free-living prepubertal children with type 1 diabetes: A multicentre, randomized clinical trial.

AIM: To assess the safety and efficacy of hybrid closed-loop (HCL) insulin delivery 24/7 versus only evening and night (E/N), and on extended 24/7 use, in free-living children with type 1 diabetes. MATERIALS AND METHODS: Prepubertal children (n = 122; 49 females/73 males; age, 8.6 ± 1.6 years; diabetes duration, 5.2 ± 2.3 years; insulin pump use, 4.6 ± 2.5 years; HbA1c 7.7% ± 0.7%/61 ± 5 mmol/mol) from four centres were randomized for 24/7 versus E/N activation of the Tandem Control-IQ system for 18 weeks. Afterwards, all children used the activated system 24/7 for 18 more weeks. The primary outcome was the percentage of time spent in the 70-180 mg/dL glucose range (TIR). RESULTS: HCL was active 94.1% and 51.1% of the time in the 24/7 and E/N modes, respectively. TIR from baseline increased more in the 24/7 versus the E/N mode (52.9% ± 9.5% to 67.3% ± 5.6% [+14.4%, 95% CI 12.4%-16.7%] vs. 55.1% ± 10.8% to 64.7% ± 7.0% [+9.6%, 95% CI 7.4%-11.6%]; P = .001). Mean percentage time below range was similarly reduced, from 4.2% and 4.6% to 2.7%, and the mean percentage time above range decreased more in the 24/7 mode (41.9% to 30.0% [-11.9%, 95% CI 9.7%-14.6%] vs. 39.8% to 32.6% [-7.2%, 95% CI 5.0%-9.9%]; P = .007). TIR increased through the whole range of baseline levels and always more with 24/7 use. The results were maintained during the extension phase in those initially on 24/7 use and improved in those with initial E/N use up to those with 24/7 use. Neither ketoacidosis nor severe hypoglycaemia occurred. CONCLUSIONS: The current study shows the safety and efficacy of the Tandem Control-IQ system in free-living children with type 1 diabetes for both E/N and 24/7 use; 24/7 use shows better outcomes, sustained for up to 36 weeks with no safety issues.

Effects of advanced carbohydrate counting on glucose control and quality of life in children with type 1 diabetes.

OBJECTIVE: The effect of advanced carbohydrate counting (ACC) on metabolic and quality of life (QOL) outcomes is uncertain in children with type 1 diabetes. Our aim was to determine whether ACC would improve HbA1c and QOL scores as compared with standard nutrition in this population. METHODS: We randomized 87 patients using pump and rapid-acting analogs in a 1 year randomized multicenter study (age 9.6 ± 3.5 years, diabetes duration 4.6 ± 2.7 years, HbA1c 7.8 ± 0.5% [62 ± 5 mmol/mol]). The ACC group received CC education and the control group received traditional dietary education. HbA1c was measured every 3 months. At 0 and 1 year, general, diabetes-specific, and diet-related QOL were respectively assessed by the KIDSCREEN and WHO-5 questionnaires, the diabetes-specific module of the DISABKIDS, and the diet restriction items of the DSQOLS. RESULTS: Mean HbA1c was lower in the ACC than the control group at 3 months (P < .05) and tended to be lower at 6 months (P = .10), 9 months (P = .10), but not at 12 months. The mean of individual average HbA1c during the one-year study period (from M3 to M12) was 7.63 ± 0.43 in the ACC vs 7.85 ± 0.47% in the control group (60 ± 5 vs 62 ± 5 mmol/mol)(P < .05). ACC was associated with significantly higher scores at 1 year on the KIDSCREEN children's psychological scale and the KIDSCREEN parents' physical scale, the DISABKIDS children's treatment scale, and the children's and parents' dietary restriction scales of the DSQOLS (indicating better QOL or lower perceived diet restriction). CONCLUSIONS: ACC may be associated with small improvements in metabolic control and QOL scores in children.

Can zinc pollution promote adaptive evolution in plants? Insights from a one-generation selection experiment.

Human activities generate environmental stresses that can lead plant populations to become extinct. Population survival would require the evolution of adaptive responses that increase tolerance to these stresses. Thus, in pseudometallophyte species that have colonized anthropogenic metalliferous habitats, the evolution of increased metal tolerance is expected in metallicolous populations. However, the mechanisms by which metal tolerance evolves remain unclear. In this study, parent populations were created from non-metallicolous families of Noccaea caerulescens. They were cultivated for one generation in mesocosms and under various levels of zinc (Zn) contamination to assess whether Zn in soil represents a selective pressure. Individual plant fitness estimates were used to create descendant populations, which were cultivated in controlled conditions with moderate Zn contamination to test for adaptive evolution in functional traits. The number of families showing high fitness estimates in mesocosms was progressively reduced with increasing Zn levels in soil, suggesting increasing selection for metal tolerance. In the next generation, adaptive evolution was suggested for some physiological and ecological traits in descendants of the most exposed populations, together with a significant decrease of Zn hyperaccumulation. Our results confirm experimentally that Zn alone can be a significant evolutionary pressure promoting adaptive divergence among populations.

Eosinophilic granulomatosis with polyangiitis in children: Data from the French RespiRare® cohort.

OBJECTIVES: To describe the characteristics of pediatric cases of eosinophilic granulomatosis with polyangiitis (EGPA), a systemic necrotizing vasculitis rarely diagnosed in children, retrieved from the French Reference Center for rare pediatric lung diseases and compared with adult cases included in the French Vasculitis Study Group cohort. METHODS: We collected information on pediatric EGPA disease presentation, management, and outcome. Cases met the Lanham criteria and/or American College of Rheumatology classification criteria. RESULTS: Fourteen cases of pediatric EGPA were included, from 1980 to 2012, with a median follow-up of 58.5 months. Median age at diagnosis was 12.3 years. All cases had respiratory involvement. The organ systems most frequently involved were the upper airway (85%), skin (71%), digestive tract (64%), and heart (57%). Neurological and renal involvement were rare. Four of the fourteen children were positive for ANCA (30.7%). During follow-up, three children required intensive care and one child died. The relapse rate was 64%. In comparison with an adult cohort, we found more ENT, heart, and digestive-tract involvement, and fewer neurological manifestations. In children, the delay between asthma onset and diagnosis was shorter, and biopsies showed fewer features of vasculitis. CONCLUSION: This French cohort is the biggest pediatric EGPA series described to date, with a long follow-up period. The findings confirm that pediatric EGPA has specific clinical, radiological, and histological characteristics that differ from adult EGPA. Development of systemic symptoms, and consequently diagnosis, occur with a shorter delay in children, mainly during the eosinophilic phase and leading to a specific presentation.

Transformation by Polyomavirus Middle T Antigen Involves a Unique Bimodal Interaction with the Hippo Effector YAP.

UNLABELLED: Murine polyomavirus has repeatedly provided insights into tumorigenesis, revealing key control mechanisms such as tyrosine phosphorylation and phosphoinositide 3-kinase (PI3K) signaling. We recently demonstrated that polyomavirus small T antigen (ST) binds YAP, a major effector of Hippo signaling, to regulate differentiation. Here we characterize YAP as a target of middle T antigen (MT) important for transformation. Through a surface including residues R103 and D182, wild-type MT binds to the YAP WW domains. Mutation of either R103 or D182 of MT abrogates YAP binding without affecting binding to other signaling molecules or the strength of PI3K or Ras signaling. Either genetic abrogation of YAP binding to MT or silencing of YAP via short hairpin RNA (shRNA) reduced MT transformation, suggesting that YAP makes a positive contribution to the transformed phenotype. MT targets YAP both by activating signaling pathways that affect it and by binding to it. MT signaling, whether from wild-type MT or the YAP-binding MT mutant, promoted YAP phosphorylation at S127 and S381/397 (YAP2/YAP1). Consistent with the known functions of these phosphorylated serines, MT signaling leads to the loss of YAP from the nucleus and degradation. Binding of YAP to MT brings it together with protein phosphatase 2A (PP2A), leading to the dephosphorylation of YAP in the MT complex. It also leads to the enrichment of YAP in membranes. Taken together, these results indicate that YAP promotes MT transformation via mechanisms that may depart from YAP's canonical oncogenic transcriptional activation functions. IMPORTANCE: The highly conserved Hippo/YAP pathway is important for tissue development and homeostasis. Increasingly, changes in this pathway are being associated with cancer. Middle T antigen (MT) is the primary polyomavirus oncogene responsible for tumor formation. In this study, we show that MT signaling promotes YAP phosphorylation, loss from the nucleus, and increased turnover. Notably, MT genetics demonstrate that YAP binding to MT is important for transformation. Because MT also binds PP2A, YAP bound to MT is dephosphorylated, stabilized, and localized to membranes. Taken together, these results indicate that YAP promotes MT transformation via mechanisms that depart from YAP's canonical oncogenic transcriptional activation functions.

NBR1 enables autophagy-dependent focal adhesion turnover.

Autophagy is a catabolic pathway involving the sequestration of cellular contents into a double-membrane vesicle, the autophagosome. Although recent studies have demonstrated that autophagy supports cell migration, the underlying mechanisms remain unknown. Using live-cell imaging, we uncover that autophagy promotes optimal migratory rate and facilitates the dynamic assembly and disassembly of cell-matrix focal adhesions (FAs), which is essential for efficient motility. Additionally, our studies reveal that autophagosomes associate with FAs primarily during disassembly, suggesting autophagy locally facilitates the destabilization of cell-matrix contact sites. Furthermore, we identify the selective autophagy cargo receptor neighbor of BRCA1 (NBR1) as a key mediator of autophagy-dependent FA remodeling. NBR1 depletion impairs FA turnover and decreases targeting of autophagosomes to FAs, whereas ectopic expression of autophagy-competent, but not autophagy-defective, NBR1 enhances FA disassembly and reduces FA lifetime during migration. Our findings provide mechanistic insight into how autophagy promotes migration by revealing a requirement for NBR1-mediated selective autophagy in enabling FA disassembly in motile cells.

Polyomavirus small T antigen interacts with yes-associated protein to regulate cell survival and differentiation.

Murine polyomavirus small t antigen (PyST) regulates cell cycle, cell survival, apoptosis, and differentiation and cooperates with middle T antigen (MT) to transform primary cells in vitro and in vivo. Like all polyomavirus T antigens, PyST functions largely via its interactions with host cell proteins. Here, we show that PyST binds both Yes-associated protein 1 (YAP1) and YAP2, integral parts of the Hippo signaling pathway, which is a subject of increasing interest in human cancer. The transcription factor TEAD, which is a known target of YAP, is also found in PyST complexes. PyST enhanced YAP association with protein phosphatase 2A (PP2A), leading to decreased YAP phosphorylation. PyST increased YAP levels by decreasing its degradation. This effect was mediated by a reduction in YAP association with ß-transducin repeat protein (ßTRCP), which is known to regulate YAP turnover in a phosphorylation-dependent manner. Genetic analysis has identified PyST mutants defective in YAP binding. These mutants demonstrated that YAP binding is important for PyST to block myoblast differentiation and to synergize with the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX) to promote cell death in 3T3-L1 preadipocytes placed under differentiation conditions. In addition to YAP binding, both of these phenotypes require PyST binding to PP2A. Importance: The Hippo/YAP pathway is a highly conserved cascade important for tissue development and homeostasis. Defects in this pathway are increasingly being associated with cancer. Polyomavirus small t antigen is a viral oncogene that cooperates with middle T antigen in transformation. On its own, small t antigen controls cell survival and differentiation. By binding YAP, small t antigen brings it together with protein phosphatase 2A. This work shows how this association of small t antigen with YAP is important for its effects on cell phenotype. It also suggests that PyST can be used to characterize cellular processes that are regulated by YAP.

Protein phosphatase 2A isoforms utilizing Aß scaffolds regulate differentiation through control of Akt protein.

Protein phosphatase 2A (PP2A) regulates almost all cell signaling pathways. It consists of a scaffolding A subunit to which a catalytic C subunit and one of many regulatory B subunits bind. Of the more than 80 PP2A isoforms, 10% use Aß as a scaffold. This study demonstrates the isoform-specific function of the A scaffold subunits. Polyomaviruses have shown the importance of phosphotyrosine, PI3K, and p53 in transformation. Comparisons of polyoma and SV40 small T antigens implicate Aß in the control of differentiation. Knockdown of Aß enhanced differentiation. Akt signaling regulated differentiation; its activation or inhibition promoted or blocked it, respectively. Aß bound Akt. Enhancement of PP2A Aß/Akt interaction by polyoma small T antigen increased turnover of Akt Ser-473 phosphorylation. Conversely, knockdown of Aß promoted Akt activity and reduced turnover of phosphate at Ser-473 of Akt. These data provide new insight into the regulation of Akt, a protein of extreme importance in cancer. Furthermore, our results suggest that the role for Aß in differentiation and perhaps tumor suppression may lie partly in its ability to negatively regulate Akt.

Investigation of the geographical scale of adaptive phenological variation and its underlying genetics in Arabidopsis thaliana.

Despite the increasing number of genomic tools, identifying the genetics underlying adaptive complex traits remains challenging in the model species Arabidopsis thaliana. This is due, at least in part, to the lack of data on the geographical scale of adaptive phenotypic variation. The aims of this study were (i) to tease apart the historical roles of adaptive and nonselective processes in shaping phenological variation in A. thaliana in France and (ii) to gain insights into the spatial scale of adaptive variation by identifying the putative selective agents responsible for this selection. Forty-nine natural stands from four climatically contrasted French regions were characterized (i) phenologically for six traits, (ii) genetically using 135 SNP markers and (iii) ecologically for 42 variables. Up to 63% of phenological variation could be explained by neutral genetic diversity. The remaining phenological variation displayed stronger associations with ecological variation within regions than among regions, suggesting the importance of local selective agents in shaping adaptive phenological variation. Although climatic conditions have often been suggested as the main selective agents acting on phenology in A. thaliana, both edaphic conditions and interspecific competition appear to be strong selective agents in some regions. In a first attempt to identify the genetics of phenological variation at different geographical scales, we phenotyped worldwide accessions and local polymorphic populations from the French RegMap in a genome-wide association (GWA) mapping study. The genomic regions associated with phenological variation depended upon the geographical scale considered, stressing the need to account for the scale of adaptive phenotypic variation when choosing accession panels for GWAS.

Genome-wide association mapping of flowering time in Arabidopsis thaliana in nature: genetics for underlying components and reaction norms across two successive years.

Often used as a proxy for the transition to reproduction, flowering time (FT) is an integrative trait of two successive biological processes, i.e. bolting time (BT) and the interval between bolting and flowering time (INT). In this study, we aimed to identify candidate genes associated with these composite traits in Arabidopsis thaliana using a field experiment. Genome-wide association (GWA) mapping was performed on BT, INT and FT based on a sample of 179 worldwide natural accessions genotyped for 216,509 SNPs. The high resolution conferred by GWA mapping indicates that FT is an integrative trait at the genetic level, with distinct genetics for BT and INT. BT is shaped largely by genes involved in the circadian clock whereas INT is shaped by genes involved in both the hormone pathways and cold acclimation. Finally, the florigen TSF appears to be the main integrator of environmental and internal signals in ecologically realistic conditions. Based on FT scored in a previous field experiment, we also studied the genetics underlying reaction norms across two years. Only four genes were common to both years, emphasizing the need to repeat field experiments. The gene regulation model appeared as the main genetic model for genotype × year interactions.

Adaptation to climate across the Arabidopsis thaliana genome.

Understanding the genetic bases and modes of adaptation to current climatic conditions is essential to accurately predict responses to future environmental change. We conducted a genome-wide scan to identify climate-adaptive genetic loci and pathways in the plant Arabidopsis thaliana. Amino acid-changing variants were significantly enriched among the loci strongly correlated with climate, suggesting that our scan effectively detects adaptive alleles. Moreover, from our results, we successfully predicted relative fitness among a set of geographically diverse A. thaliana accessions when grown together in a common environment. Our results provide a set of candidates for dissecting the molecular bases of climate adaptations, as well as insights about the prevalence of selective sweeps, which has implications for predicting the rate of adaptation.

Linkage and association mapping of Arabidopsis thaliana flowering time in nature.

Flowering time is a key life-history trait in the plant life cycle. Most studies to unravel the genetics of flowering time in Arabidopsis thaliana have been performed under greenhouse conditions. Here, we describe a study about the genetics of flowering time that differs from previous studies in two important ways: first, we measure flowering time in a more complex and ecologically realistic environment; and, second, we combine the advantages of genome-wide association (GWA) and traditional linkage (QTL) mapping. Our experiments involved phenotyping nearly 20,000 plants over 2 winters under field conditions, including 184 worldwide natural accessions genotyped for 216,509 SNPs and 4,366 RILs derived from 13 independent crosses chosen to maximize genetic and phenotypic diversity. Based on a photothermal time model, the flowering time variation scored in our field experiment was poorly correlated with the flowering time variation previously obtained under greenhouse conditions, reinforcing previous demonstrations of the importance of genotype by environment interactions in A. thaliana and the need to study adaptive variation under natural conditions. The use of 4,366 RILs provides great power for dissecting the genetic architecture of flowering time in A. thaliana under our specific field conditions. We describe more than 60 additive QTLs, all with relatively small to medium effects and organized in 5 major clusters. We show that QTL mapping increases our power to distinguish true from false associations in GWA mapping. QTL mapping also permits the identification of false negatives, that is, causative SNPs that are lost when applying GWA methods that control for population structure. Major genes underpinning flowering time in the greenhouse were not associated with flowering time in this study. Instead, we found a prevalence of genes involved in the regulation of the plant circadian clock. Furthermore, we identified new genomic regions lacking obvious candidate genes.

Genome-wide association study of 107 phenotypes in Arabidopsis thaliana inbred lines.

Although pioneered by human geneticists as a potential solution to the challenging problem of finding the genetic basis of common human diseases, genome-wide association (GWA) studies have, owing to advances in genotyping and sequencing technology, become an obvious general approach for studying the genetics of natural variation and traits of agricultural importance. They are particularly useful when inbred lines are available, because once these lines have been genotyped they can be phenotyped multiple times, making it possible (as well as extremely cost effective) to study many different traits in many different environments, while replicating the phenotypic measurements to reduce environmental noise. Here we demonstrate the power of this approach by carrying out a GWA study of 107 phenotypes in Arabidopsis thaliana, a widely distributed, predominantly self-fertilizing model plant known to harbour considerable genetic variation for many adaptively important traits. Our results are dramatically different from those of human GWA studies, in that we identify many common alleles of major effect, but they are also, in many cases, harder to interpret because confounding by complex genetics and population structure make it difficult to distinguish true associations from false. However, a-priori candidates are significantly over-represented among these associations as well, making many of them excellent candidates for follow-up experiments. Our study demonstrates the feasibility of GWA studies in A. thaliana and suggests that the approach will be appropriate for many other organisms.