Implications of Chaoborus pupation and ecdysis in cold water

1. Cold water acted differently to delay and lengthen the pupation period for the larvae of two species of the zooplankton predator Chaoborus (Diptera: Chaoboridae). During Chaoborus pupation, the zooplankton community is released from predation, while the dark-coloured Chaoborus pupae are more susceptible to their own predators.
2. Fourth instar larvae of Chaoborus americanus and C. trivittatus , collected from an oligotrophic lake, were reared individually at 5 °C in the dark. Chaoborus americanus was also reared at 9 and 12 °C under spring photoperiod conditions (L : D, 16 : 8 h). Individuals were observed through pupation to emergence (ecdysis) or death.
3.  Chaoborus americanus pupated at 5, 9 and 12 °C with substantial emergence only at 12 °C. In comparison, C. trivittatus emerged at 5 °C. Light was not a necessary cue for pupation and ecdysis, contrary to previous reports. Cold water delayed the onset and lengthened and increased the variability of the duration of pupation.
4. In Shirley Lake, C. americanus pupated in late June–early July while C. trivittatus pupated first in April and again in June–July.
5.  Chaoborus americanus pupae needed a temperature cue to complete ecdysis. The ecdysis temperature threshold helps to explain the difference in pupation timing, and the geographical distribution, of C. americanus and its relatively inflexible life history contrasted with C. trivittatus . Delayed predator pupation in years with low spring temperature can affect the community dynamics of the prey.     

Size-fractionated Carboxylase Activities During a 32 h Cycle at 30 m Depth in the North-western Mediterranean Sea After an Episodic Wind Event

Using size-fractionation filtration (1 µm), we associatedcarboxylase activities (Rubisco, ß-carboxylases) andchlorophyll measurements with cell enumeration by flow cytometryat a permanent site of the central Ligurian Sea in the north-westernMediterranean Sea (73°25'N–7°51' E). The analyseswere carried out over a day/night cycle (at 30 m depth) followinga strong wind event, during the transition period from springmesotrophic to summer oligotrophic conditions. The highest valuesof Rubisco activity and ß-carboxylase activity perchlorophyll a (Chl a) for >1 µm cells were observedduring the light period of the cycle, reaching 18.9 and 4.3nmol CO₂ (µg Chl a)^–1 h^–1, respectively. Thishigher activity is assumed to be correlated with a dominanceof nanoflagellates in the phytoplankton community. Such phytoplanktonspecies generally had higher ß-carboxylase activity,expressed as a percentage of Rubisco activity (the ßC/Rratio), than diatoms. Using flow cytometry analysis to enumeratethose cells <1 µm in size, we followed the values ofRubisco activity and pigment content expressed per cell, forpicophytoplankton cells. The photoautotrophic activity, measuredas the in vitro Rubisco activity for small picoeukaryote cells,was higher than for cyanobacteria cells with lower apparentcell size. These results suggested an optimum of CO₂ assimilationreached by the pico- and nano-phytoplankton in accordance withthe cell size and growth rates from previous observations inthe literature.     

Barriers to Advance Care Planning in Cancer,Heart Failure and Dementia Patients: A Focus Group Study on General Practitioners' Views and Experiences

Background

The long-term and often lifelong relationship of general practitioners (GPs) with their patients is considered to make them the ideal initiators of advance care planning (ACP). However, in general the incidence of ACP discussions is low and ACP seems to occur more often for cancer patients than for those with dementia or heart failure.

Objective

To identify the barriers, from GPs'' perspective, to initiating ACP and to gain insight into any differences in barriers between the trajectories of patients with cancer, heart failure and dementia.

Method

Five focus groups were held with GPs (n = 36) in Flanders, Belgium. The focus group discussions were transcribed verbatim and analyzed using the method of constant comparative analysis.

Results

Three types of barriers were distinguished: barriers relating to the GP, to the patient and family and to the health care system. In cancer patients, a GP''s lack of knowledge about treatment options and the lack of structural collaboration between the GP and specialist were expressed as barriers. Barriers that occured more often with heart failure and dementia were the lack of GP familiarity with the terminal phase, the lack of key moments to initiate ACP, the patient''s lack of awareness of their diagnosis and prognosis and the fact that patients did not often initiate such discussions themselves. The future lack of decision-making capacity of dementia patients was reported by the GPs as a specific barrier for the initiation of ACP.

Conclusion

The results of our study contribute to a better understanding of the factors hindering GPs in initiating ACP. Multiple barriers need to be overcome, of which many can be addressed through the development of practical guidelines and educational interventions.     

SIRT3, a Mitochondrial NAD+-Dependent Deacetylase,Is Involved in the Regulation of Myoblast Differentiation

Sirtuin 3 (SIRT3), one of the seven mammalian sirtuins, is a mitochondrial NAD⁺-dependent deacetylase known to control key metabolic pathways. SIRT3 deacetylases and activates a large number of mitochondrial enzymes involved in the respiratory chain, in ATP production, and in both the citric acid and urea cycles. We have previously shown that the regulation of myoblast differentiation is tightly linked to mitochondrial activity. Since SIRT3 modulates mitochondrial activity, we decide to address its role during myoblast differentiation. For this purpose, we first investigated the expression of endogenous SIRT3 during C2C12 myoblast differentiation. We further studied the impact of SIRT3 silencing on both the myogenic potential and the mitochondrial activity of C2C12 cells. We showed that SIRT3 protein expression peaked at the onset of myoblast differentiation. The inhibition of SIRT3 expression mediated by the stable integration of SIRT3 short inhibitory RNA (SIRT3shRNA) in C2C12 myoblasts, resulted in: 1) abrogation of terminal differentiation - as evidenced by a marked decrease in the myoblast fusion index and a significant reduction of Myogenin, MyoD, Sirtuin 1 and Troponin T protein expression - restored upon MyoD overexpression; 2) a decrease in peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and citrate synthase protein expression reflecting an alteration of mitochondrial density; and 3) an increased production of reactive oxygen species (ROS) mirrored by the decreased activity of manganese superoxide dismutase (MnSOD). Altogether our data demonstrate that SIRT3 mainly regulates myoblast differentiation via its influence on mitochondrial activity.     

Assessing Cell Cycle Progression of Neural Stem and Progenitor Cells in the Mouse Developing Brain after Genotoxic Stress

Neurons of the cerebral cortex are generated during brain development from different types of neural stem and progenitor cells (NSPC), which form a pseudostratified epithelium lining the lateral ventricles of the embryonic brain. Genotoxic stresses, such as ionizing radiation, have highly deleterious effects on the developing brain related to the high sensitivity of NSPC. Elucidation of the cellular and molecular mechanisms involved depends on the characterization of the DNA damage response of these particular types of cells, which requires an accurate method to determine NSPC progression through the cell cycle in the damaged tissue. Here is shown a method based on successive intraperitoneal injections of EdU and BrdU in pregnant mice and further detection of these two thymidine analogues in coronal sections of the embryonic brain. EdU and BrdU are both incorporated in DNA of replicating cells during S phase and are detected by two different techniques (azide or a specific antibody, respectively), which facilitate their simultaneous detection. EdU and BrdU staining are then determined for each NSPC nucleus in function of its distance from the ventricular margin in a standard region of the dorsal telencephalon. Thus this dual labeling technique allows distinguishing cells that progressed through the cell cycle from those that have activated a cell cycle checkpoint leading to cell cycle arrest in response to DNA damage.An example of experiment is presented, in which EdU was injected before irradiation and BrdU immediately after and analyzes performed within the 4 hr following irradiation. This protocol provides an accurate analysis of the acute DNA damage response of NSPC in function of the phase of the cell cycle at which they have been irradiated. This method is easily transposable to many other systems in order to determine the impact of a particular treatment on cell cycle progression in living tissues.     

Assessing nitrate leaching during the three‐first years of Miscanthus × giganteus from on‐farm measurements and modeling

Miscanthus × giganteus is often regarded as one of the most promising crops to produce sustainable bioenergy. This perennial crop, renowned for its high productivity associated with low input requirements, in particular regarding fertilizers, is thought to have low environmental impacts, but few data are available to confirm this. Our study aimed at assessing nitrate leaching from Miscanthus × giganteus crops in farmers' fields, thus including a wide range of soil and cropping system conditions. We focused on the first years of growth after planting as experimental studies have suggested that Miscanthus × giganteus, once established, results in low nitrate leaching. We combined on‐farm measurements and modeling to estimate drainage, leached nitrogen, and nitrate concentration in drainage water in 38 fields located in Center‐East France during two winters (November 2010 to March 2011, November 2011 to March 2012). Nitrate leaching and nitrate concentration in drainage water were on average very low. Nitrate leaching averaged 6 kg N ha^?1 whereas nitrate concentration averaged 12 mg l^?1. These low values are attributable to the low estimates of drainage water (mean = 166 mm) but also to the low soil mineral nitrogen contents measured at the beginning of winter (mean = 37 kg N ha^?1). Our results were, however, very variable, mainly due to the crop age: nitrate leaching and nitrate concentration were critically higher during the winter following the first growth year of Miscanthus × giganteus, reflecting the low development of the crop. This variability was also explained by the range of soil and cropping conditions explored in the on‐farm design: shallow and/or sandy soils as well as fields where establishment failed had a higher risk of nitrate leaching.     

Decreased DGCR8 Expression and miRNA Dysregulation in Individuals with 22q11.2 Deletion Syndrome

Deletion of the 1.5–3 Mb region of chromosome 22 at locus 11.2 gives rise to the chromosome 22q11.2 deletion syndrome (22q11DS), also known as DiGeorge and Velocardiofacial Syndromes. It is the most common micro-deletion disorder in humans and one of the most common multiple malformation syndromes. The syndrome is characterized by a broad phenotype, whose characterization has expanded considerably within the last decade and includes many associated findings such as craniofacial anomalies (40%), conotruncal defects of the heart (CHD; 70–80%), hypocalcemia (20–60%), and a range of neurocognitive anomalies with high risk of schizophrenia, all with a broad phenotypic variability. These phenotypic features are believed to be the result of a change in the copy number or dosage of the genes located in the deleted region. Despite this relatively clear genetic etiology, very little is known about which genes modulate phenotypic variations in humans or if they are due to combinatorial effects of reduced dosage of multiple genes acting in concert. Here, we report on decreased expression levels of genes within the deletion region of chromosome 22, including DGCR8, in peripheral leukocytes derived from individuals with 22q11DS compared to healthy controls. Furthermore, we found dysregulated miRNA expression in individuals with 22q11DS, including miR-150, miR-194 and miR-185. We postulate this to be related to DGCR8 haploinsufficiency as DGCR8 regulates miRNA biogenesis. Importantly we demonstrate that the level of some miRNAs correlates with brain measures, CHD and thyroid abnormalities, suggesting that the dysregulated miRNAs may contribute to these phenotypes and/or represent relevant blood biomarkers of the disease in individuals with 22q11DS.     

Oxidation of CaMKII determines the cardiotoxic effects of aldosterone

Excessive activation of the β-adrenergic, angiotensin II (Ang II) and aldosterone signaling pathways promotes mortality after myocardial infarction, and antagonists targeting these pathways are core therapies for treating this condition. Catecholamines and Ang II activate the multifunctional Ca(2+)/calmodulin-dependent protein kinase II (CaMKII), the inhibition of which prevents isoproterenol-mediated and Ang II-mediated cardiomyopathy. Here we show that aldosterone exerts direct toxic actions on myocardium by oxidative activation of CaMKII, causing cardiac rupture and increased mortality in mice after myocardial infarction. Aldosterone induces CaMKII oxidation by recruiting NADPH oxidase, and this oxidized and activated CaMKII promotes matrix metalloproteinase 9 (MMP9) expression in cardiomyocytes. Myocardial CaMKII inhibition, overexpression of methionine sulfoxide reductase A (an enzyme that reduces oxidized CaMKII) or NADPH oxidase deficiency prevented aldosterone-enhanced cardiac rupture after myocardial infarction. These findings show that oxidized myocardial CaMKII mediates the cardiotoxic effects of aldosterone on the cardiac matrix and establish CaMKII as a nodal signal for the neurohumoral pathways associated with poor outcomes after myocardial infarction.