Structure of Excitatory Synapses and GABAA Receptor Localization at Inhibitory Synapses Are Regulated by Neuroplastin-65

Formation, maintenance, and activity of excitatory and inhibitory synapses are essential for neuronal network function. Cell adhesion molecules (CAMs) are crucially involved in these processes. The CAM neuroplastin-65 (Np65) highly expressed during periods of synapse formation and stabilization is present at the pre- and postsynaptic membranes. Np65 can translocate into synapses in response to electrical stimulation and it interacts with subtypes of GABA_A receptors in inhibitory synapses. Here, we report that in the murine hippocampus and in hippocampal primary culture, neurons of the CA1 region and the dentate gyrus (DG) express high Np65 levels, whereas expression in CA3 neurons is lower. In neuroplastin-deficient (Np^−/−) mice the number of excitatory synapses in CA1 and DG, but not CA3 regions is reduced. Notably this picture is mirrored in mature Np^−/− hippocampal cultures or in mature CA1 and DG wild-type (Np^+/+) neurons treated with a function-blocking recombinant Np65-Fc extracellular fragment. Although the number of GABAergic synapses was unchanged in Np^−/− neurons or in mature Np65-Fc-treated Np^+/+ neurons, the ratio of excitatory to inhibitory synapses was significantly lower in Np^−/− cultures. Furthermore, GABA_A receptor composition was altered at inhibitory synapses in Np^−/− neurons as the α1 to α2 GABA_A receptor subunit ratio was increased. Changes of excitatory and inhibitory synaptic function in Np^−/− neurons were confirmed evaluating the presynaptic release function and using patch clamp recording. These data demonstrate that Np65 is an important regulator of the number and function of synapses in the hippocampus.     

Sediment Nitrification, Denitrification, and Nitrous Oxide Production in a Deep Arctic Lake

We used a combination of ¹⁵N tracer methods and a C₂H₂ blockage technique to determine the role of sediment nitrification and denitrification in a deep oligotrophic arctic lake. Inorganic nitrogen concentrations ranged between 40 and 600 nmol · cm⁻³, increasing with depth below the sediment-water interface. Nitrate concentrations were at least 10 times lower, and nitrate was only detectable within the top 0 to 6 cm of sediment. Eh and pH profiles showed an oxidized surface zone underlain by more reduced conditions. The lake water never became anoxic. Sediment Eh values ranged from −7 to 484 mV, decreasing with depth, whereas pH ranged from 6.0 to 7.3, usually increasing with depth. The average nitrification rate (49 ng of N · cm⁻³ · day⁻¹) was similar to the average denitrification rate (44 ng of N · cm⁻³ · day⁻¹). In situ N₂O production from nitrification and denitrification ranged from 0 to 25 ng of N · cm⁻³ · day⁻¹. Denitrification appears to depend on the supply of nitrate by nitrification, such that the two processes are coupled functionally in this sediment system. However, the low rates result in only a small nitrogen loss.     

Aftereffects of low and high temperature pretreatment on leaf resistance, transpiration, and leaf temperature in xanthium

Leaf resistance for water vapor (total diffusion resistance minus boundary layer resistance), transpiration, and leaf temperature were measured in attached leaves of greenhouse-grown Xanthium strumarium L. plants that had been pretreated for 72 hours with high (40 C day, 35 C night), or low (10 C day, 5 C night) air temperatures. Measurements were made in a wind tunnel at light intensity of 1.15 cal cm⁻² min⁻¹, air temperatures between 5 and 45 C, and wind speed of 65 cm sec⁻¹. Leaf resistances in low temperature pretreated plants were higher (8 to 27 sec cm⁻¹) than in controls or high temperature pretreated plants (0.5 to 3 sec cm⁻¹) at leaf temperatures between 5 and 25 C. Thus, the pretreatment influenced stomatal aperture.     

Quantification of Left Ventricular Torsion and Diastolic Recoil Using Cardiovascular Magnetic Resonance Myocardial Feature Tracking

Objectives

Cardiovascular magnetic resonance feature tracking (CMR-FT) offers quantification of myocardial deformation from routine cine images. However, data using CMR-FT to quantify left ventricular (LV) torsion and diastolic recoil are not yet available. We therefore sought to evaluate the feasibility and reproducibility of CMR-FT to quantify LV torsion and peak recoil rate using an optimal anatomical approach.

Methods

Short-axis cine stacks were acquired at rest and during dobutamine stimulation (10 and 20 µg·kg⁻¹·min⁻¹) in 10 healthy volunteers. Rotational displacement was analysed for all slices. A complete 3D-LV rotational model was developed using linear interpolation between adjacent slices. Torsion was defined as the difference between apical and basal rotation, divided by slice distance. Depending on the distance between the most apical (defined as 0% LV distance) and basal (defined as 100% LV distance) slices, four different models for the calculation of torsion were examined: Model-1 (25–75%), Model-2 (0–100%), Model-3 (25–100%) and Model-4 (0–75%). Analysis included subendocardial, subepicardial and global torsion and recoil rate (mean of subendocardial and subepicardial values).

Results

Quantification of torsion and recoil rate was feasible in all subjects. There was no significant difference between the different models at rest. However, only Model-1 (25–75%) discriminated between rest and stress (Global Torsion: 2.7±1.5°cm⁻¹, 3.6±2.0°cm⁻¹, 5.1±2.2°cm⁻¹, p<0.01; Global Recoil Rate: −30.1±11.1°cm⁻¹s⁻¹,−46.9±15.0°cm⁻¹s⁻¹,−68.9±32.3°cm⁻¹s⁻¹, p<0.01; for rest, 10 and 20 µg·kg⁻¹·min⁻¹ of dobutamine, respectively). Reproducibility was sufficient for all parameters as determined by Bland-Altman analysis, intraclass correlation coefficients and coefficient of variation.

Conclusions

CMR-FT based derivation of myocardial torsion and recoil rate is feasible and reproducible at rest and with dobutamine stress. Using an optimal anatomical approach measuring rotation at 25% and 75% apical and basal LV locations allows effective quantification of torsion and recoil dynamics. Application of these new measures of deformation by CMR-FT should next be explored in disease states.     

Persistent Organic Pollutants and Early Menopause in U.S. Women

Objective

Endocrine-disrupting chemicals (EDCs) adversely affect human health. Our objective was to determine the association of EDC exposure with earlier age of menopause.

Methods

Cross-sectional survey using National Health and Nutrition Examination Survey (NHANES) data from 1999 to 2008 (n = 31,575 females). Eligible participants included: menopausal women >30 years of age; not currently pregnant, breastfeeding, using hormonal contraception; no history of bilateral oophorectomy or hysterectomy. Exposures, defined by serum lipid and urine creatinine-adjusted measures of EDCs, data were analyzed: > 90^th percentile of the EDC distribution among all women, log-transformed EDC level, and decile of EDC level. Multi linear regression models considered complex survey design characteristics and adjusted for age, race/ethnicity, smoking, body mass index. EDCs were stratified into long (>1 year), short, and unknown half-lives; principle analyses were performed on those with long half-lives as well as phthalates, known reproductive toxicants. Secondary analysis determined whether the odds of being menopausal increased with EDC exposure among women aged 45–55 years.

Findings

This analysis examined 111 EDCs and focused on known reproductive toxicants or chemicals with half-lives >1 year. Women with high levels of β-hexachlorocyclohexane, mirex, p,p’-DDE, 1,2,3,4,6,7,8-heptachlorodibenzofuran, mono-(2-ethyl-5-hydroxyhexyl) and mono-(2-ethyl-5-oxohexyl) phthalate, polychlorinated biphenyl congeners −70, −99, −105, −118, −138, −153, −156, −170, and −183 had mean ages of menopause 1.9 to 3.8 years earlier than women with lower levels of these chemicals. EDC-exposed women were up to 6 times more likely to be menopausal than non-exposed women.

Conclusions

This study of a representative sample of US women documents an association between EDCs and earlier age at menopause. We identified 15 EDCs that warrant closer evaluation because of their persistence and potential detrimental effects on ovarian function. Earlier menopause can alter the quantity and quality of a woman’s life and has profound implications for fertility, human reproduction, and our global society.     

Impairment of Visual Function and Retinal ER Stress Activation in Wfs1-Deficient Mice

Wolfram syndrome is an early onset genetic disease (1/180,000) featuring diabetes mellitus and optic neuropathy, associated to mutations in the WFS1 gene. Wfs1−/− mouse model shows pancreatic beta cell atrophy, but its visual performance has not been investigated, prompting us to study its visual function and histopathology of the retina and optic nerve. Electroretinogram and visual evoked potentials (VEPs) were performed in Wfs1^−/− and Wfs1^+/+ mice at 3, 6, 9 and 12 months of age. Fundi were pictured with Micron III apparatus. Retinal ganglion cell (RGC) abundance was determined from Brn3a immunolabeling of retinal sections. RGC axonal loss was quantified by electron microscopy in transversal optic nerve sections. Endoplasmic reticulum stress was assessed using immunoglobulin binding protein (BiP), protein disulfide isomerase (PDI) and inositol-requiring enzyme 1 alpha (Ire1α) markers. Electroretinograms amplitudes were slightly reduced and latencies increased with time in Wfs1^−/− mice. Similarly, VEPs showed decreased N+P amplitudes and increased N-wave latency. Analysis of unfolded protein response signaling revealed an activation of endoplasmic reticulum stress in Wfs1 ^−/− mutant mouse retinas. Altogether, progressive VEPs alterations with minimal neuronal cell loss suggest functional alteration of the action potential in the Wfs1 ^−/− optic pathways.     

Bax-Induced Apoptosis in Leber's Congenital Amaurosis: A Dual Role in Rod and Cone Degeneration

Pathogenesis in the Rpe65^−/− mouse model of Leber''s congenital amaurosis (LCA) is characterized by a slow and progressive degeneration of the rod photoreceptors. On the opposite, cones degenerate rapidly at early ages. Retinal degeneration in Rpe65 ^−/− mice, showing a null mutation in the gene encoding the retinal pigment epithelium 65-kDa protein (Rpe65), was previously reported to depend on continuous activation of a residual transduction cascade by unliganded opsin. However, the mechanisms of apoptotic signals triggered by abnormal phototransduction remain elusive. We previously reported that activation of a Bcl-2-dependent pathway was associated with apoptosis of rod photoreceptors in Rpe65^−/− mice during the course of the disease. In this study we first assessed whether activation of Bcl-2-mediated apoptotic pathway was dependent on constitutive activation of the visual cascade through opsin apoprotein. We then challenged the direct role of pro-apoptotic Bax protein in triggering apoptosis of rod and cone photoreceptors.Quantitative PCR analysis showed that increased expression of pro-apoptotic Bax and decreased level of anti-apoptotic Bcl-2 were restored in Rpe65^−/−/Gnat1^−/− mice lacking the Gnat1 gene encoding rod transducin. Moreover, photoreceptor apoptosis was prevented as assessed by TUNEL assay. These data indicate that abnormal activity of opsin apoprotein induces retinal cell apoptosis through the Bcl-2-mediated pathway. Following immunohistological and real-time PCR analyses, we further observed that decreased expression of rod genes in Rpe65-deficient mice was rescued in Rpe65^−/−/Bax^−/− mice. Histological and TUNEL studies confirmed that rod cell demise and apoptosis in diseased Rpe65^−/− mice were dependent on Bax-induced pathway. Surprisingly, early loss of cones was not prevented in Rpe65^−/−/Bax^−/− mice, indicating that pro-apoptotic Bax was not involved in the pathogenesis of cone cell death in Rpe65-deficient mice.This is the first report, to our knowledge, that a single genetic mutation can trigger two independent apoptotic pathways in rod and cone photoreceptors in Rpe65-dependent LCA disease. These results highlight the necessity to investigate and understand the specific death signaling pathways committed in rods and cones to develop effective therapeutic approaches to treat RP diseases.     

Air quality improvement and cognitive decline in community-dwelling older women in the United States: A longitudinal cohort study

BackgroundLate-life exposure to ambient air pollution is a modifiable risk factor for dementia, but epidemiological studies have shown inconsistent evidence for cognitive decline. Air quality (AQ) improvement has been associated with improved cardiopulmonary health and decreased mortality, but to the best of our knowledge, no studies have examined the association with cognitive function. We examined whether AQ improvement was associated with slower rate of cognitive decline in older women aged 74 to 92 years.Methods and findingsWe studied a cohort of 2,232 women residing in the 48 contiguous US states that were recruited from more than 40 study sites located in 24 states and Washington, DC from the Women’s Health Initiative (WHI) Memory Study (WHIMS)-Epidemiology of Cognitive Health Outcomes (WHIMS-ECHO) study. They were predominantly non-Hispanic White women and were dementia free at baseline in 2008 to 2012. Measures of annual (2008 to 2018) cognitive function included the modified Telephone Interview for Cognitive Status (TICSm) and the telephone-based California Verbal Learning Test (CVLT). We used regionalized universal kriging models to estimate annual concentrations (1996 to 2012) of fine particulate matter (PM_2.5) and nitrogen dioxide (NO₂) at residential locations. Estimates were aggregated to the 3-year average immediately preceding (recent exposure) and 10 years prior to (remote exposure) WHIMS-ECHO enrollment. Individual-level improved AQ was calculated as the reduction from remote to recent exposures. Linear mixed effect models were used to examine the associations between improved AQ and the rates of cognitive declines in TICSm and CVLT trajectories, adjusting for sociodemographic (age; geographic region; race/ethnicity; education; income; and employment), lifestyle (physical activity; smoking; and alcohol), and clinical characteristics (prior hormone use; hormone therapy assignment; depression; cardiovascular disease (CVD); hypercholesterolemia; hypertension; diabetes; and body mass index [BMI]). For both PM_2.5 and NO₂, AQ improved significantly over the 10 years before WHIMS-ECHO enrollment. During a median of 6.2 (interquartile range [IQR] = 5.0) years of follow-up, declines in both general cognitive status (β = −0.42/year, 95% CI: −0.44, −0.40) and episodic memory (β = −0.59/year, 95% CI: −0.64, −0.54) were observed. Greater AQ improvement was associated with slower decline in TICSm (β_{PM2.5improvement} = 0.026 per year for improved PM_2.5 by each IQR = 1.79 μg/m³ reduction, 95% CI: 0.001, 0.05; β_{NO2improvement} = 0.034 per year for improved NO₂ by each IQR = 3.92 parts per billion [ppb] reduction, 95% CI: 0.01, 0.06) and CVLT (β_{PM2.5 improvement} = 0.070 per year for improved PM_2.5 by each IQR = 1.79 μg/m³ reduction, 95% CI: 0.02, 0.12; β_{NO2improvement} = 0.060 per year for improved NO₂ by each IQR = 3.97 ppb reduction, 95% CI: 0.005, 0.12) after adjusting for covariates. The respective associations with TICSm and CVLT were equivalent to the slower decline rate found with 0.9 to 1.2 and1.4 to 1.6 years of younger age and did not significantly differ by age, region, education, Apolipoprotein E (ApoE) e4 genotypes, or cardiovascular risk factors. The main limitations of this study include measurement error in exposure estimates, potential unmeasured confounding, and limited generalizability.ConclusionsIn this study, we found that greater improvement in long-term AQ in late life was associated with slower cognitive declines in older women. This novel observation strengthens the epidemiologic evidence of an association between air pollution and cognitive aging.

Diana Younan and colleagues investigate whether air quality improvement is associated with rate of cognitive decline in community-dwelling older women in the United States.     

Regional Distribution and Evolution of Gray Matter Damage in Different Populations of Multiple Sclerosis Patients

Background

Both gray-matter (GM) atrophy and lesions occur from the earliest stages of Multiple Sclerosis (MS) and are one of the major determinants of long-term clinical outcomes. Nevertheless, the relationship between focal and diffuse GM damage has not been clarified yet. Here we investigate the regional distribution and temporal evolution of cortical thinning and how it is influenced by the local appearance of new GM lesions at different stages of the disease in different populations of MS patients.

Methods

We studied twenty MS patients with clinically isolated syndrome (CIS), 27 with early relapsing-remitting MS (RRMS, disease duration <5 years), 29 with late RRMS (disease duration ≥ 5 years) and 20 with secondary-progressive MS (SPMS). The distribution and evolution of regional cortical thickness and GM lesions were assessed during 5-year follow-up.

Results

The results showed that new lesions appeared more frequently in hippocampus and parahippocampal gyri (9.1%), insula (8.9%), cingulate cortex (8.3%), superior frontal gyrus (8.1%), and cerebellum (6.5%). The aforementioned regions showed the greatest reduction in thickness/volume, although (several) differences were observed across subgroups. The correlation between the appearance of new cortical lesions and cortical thinning was stronger in CIS (r² = 50.0, p<0.001) and in early RRMS (r² = 52.3, p<0.001), compared to late RRMS (r² = 25.5, p<0.001) and SPMS (r² = 6.3, p = 0.133).

Conclusions

We conclude that GM atrophy and lesions appear to be different signatures of cortical disease in MS having in common overlapping spatio-temporal distribution patterns. However, the correlation between focal and diffuse damage is only moderate and more evident in the early phase of the disease.     

Seasonal Mesophotic Coral Bleaching of Stylophora pistillata in the Northern Red Sea

Coral bleaching occurs when environmental stress induces breakdown of the coral-algae symbiosis and the host initiates algae expulsion. Two types of coral bleaching had been thoroughly discussed in the scientific literature; the first is primarily associated with mass coral bleaching events; the second is a seasonal loss of algae and/or pigments. Here, we describe a phenomenon that has been witnessed for repeated summers in the mesophotic zone (40–63 m) in the northern Red Sea: seasonal bleaching and recovery of several hermatypic coral species. In this study, we followed the recurring bleaching process of the common coral Stylophora pistillata. Bleaching occurred from April to September with a 66% decline in chlorophyll a concentration, while recovery began in October. Using aquarium and transplantation experiments, we explored environmental factors such as temperature, photon flux density and heterotrophic food availability. Our experiments and observations did not yield one single factor, alone, responsible for the seasonal bleaching. The dinoflagellate symbionts (of the genus Symbiodinium) in shallow (5 m) Stylophora pistillata were found to have a net photosynthetic rate of 56.98–92.19 µmol O₂ cm⁻² day⁻¹. However, those from mesophotic depth (60 m) during months when they are not bleached are net consumers of oxygen having a net photosynthetic rate between −12.86 - (−10.24) µmol O₂ cm⁻² day⁻¹. But during months when these mesophotic corals are partially-bleached, they yielded higher net production, between −2.83–0.76 µmol O₂ cm⁻² day⁻¹. This study opens research questions as to why mesophotic zooxanthellae are more successfully meeting the corals metabolic requirements when Chl a concentration decreases by over 60% during summer and early fall.     

Mouse models of telomere dysfunction phenocopy skeletal changes found in human age-related osteoporosis

A major medical challenge in the elderly is osteoporosis and the high risk of fracture. Telomere dysfunction is a cause of cellular senescence and telomere shortening, which occurs with age in cells from most human tissues, including bone. Telomere defects contribute to the pathogenesis of two progeroid disorders characterized by premature osteoporosis, Werner syndrome and dyskeratosis congenital. It is hypothesized that telomere shortening contributes to bone aging. We evaluated the skeletal phenotypes of mice with disrupted telomere maintenance mechanisms as models for human bone aging, including mutants in Werner helicase (Wrn^−/−), telomerase (Terc^−/−) and Wrn^−/−Terc^−/− double mutants. Compared with young wild-type (WT) mice, micro-computerized tomography analysis revealed that young Terc^−/− and Wrn^−/−Terc^−/− mice have decreased trabecular bone volume, trabecular number and trabecular thickness, as well as increased trabecular spacing. In cortical bone, young Terc^−/− and Wrn^−/−Terc^−/− mice have increased cortical thinning, and increased porosity relative to age-matched WT mice. These trabecular and cortical changes were accelerated with age in Terc^−/− and Wrn^−/−Terc^−/− mice compared with older WT mice. Histological quantification of osteoblasts in aged mice showed a similar number of osteoblasts in all genotypes; however, significant decreases in osteoid, mineralization surface, mineral apposition rate and bone formation rate in older Terc^−/− and Wrn^−/−Terc^−/− bone suggest that osteoblast dysfunction is a prominent feature of precocious aging in these mice. Except in the Wrn^−/− single mutant, osteoclast number did not increase in any genotype. Significant alterations in mechanical parameters (structure model index, degree of anistrophy and moment of inertia) of the Terc^−/− and Wrn^−/−Terc^−/− femurs compared with WT mice were also observed. Young Wrn^−/−Terc^−/− mice had a statistically significant increase in bone-marrow fat content compared with young WT mice, which remained elevated in aged double mutants. Taken together, our results suggest that Terc^−/− and Wrn^−/−Terc^−/− mutants recapitulate the human bone aging phenotype and are useful models for studying age-related osteoporosis.KEY WORDS: Aging, Bone histomorphometry, Osteoporosis     

Reduced Hippocampal Volume in Healthy Young ApoE4 Carriers: An MRI Study

The E4 allele of the ApoE gene has consistently been shown to be related to an increased risk of Alzheimer''s disease (AD). The E4 allele is also associated with functional and structural grey matter (GM) changes in healthy young, middle-aged and older subjects. Here, we assess volumes of deep grey matter structures of 22 healthy younger ApoE4 carriers and 22 non-carriers (20–38 years). Volumes of the nucleus accumbens, amygdala, caudate nucleus, hippocampus, pallidum, putamen, thalamus and brain stem were calculated by FMRIB''s Integrated Registration and Segmentation Tool (FIRST) algorithm. A significant drop in volume was found in the right hippocampus of ApoE4 carriers (ApoE4+) relative to non-carriers (ApoE4−), while there was a borderline significant decrease in the volume of the left hippocampus of ApoE4 carriers. The volumes of no other structures were found to be significantly affected by genotype. Atrophy has been found to be a sensitive marker of neurodegenerative changes, and our results show that within a healthy young population, the presence of the ApoE4+ carrier gene leads to volume reduction in a structure that is vitally important for memory formation. Our results suggest that the hippocampus may be particularly vulnerable to further degeneration in ApoE4 carriers as they enter middle and old age. Although volume reductions were noted bilaterally in the hippocampus, atrophy was more pronounced in the right hippocampus. This finding relates to previous work which has noted a compensatory increase in right hemisphere activity in ApoE4 carriers in response to preclinical declines in memory function. Possession of the ApoE4 allele may lead to greater predilection for right hemisphere atrophy even in healthy young subjects in their twenties.     

Investigation on Crude and High-Temperature Heated Coffee Oil by ATR-FTIR Spectroscopy along with Antioxidant and Antimicrobial Properties

The coffee oil has a promising potential to be used in food industry, but an efficient use, especially in products that required high-temperature heating, depends on its chemical composition and the changes induced by processing. Since there is little information on this topic, the aim of our study was to investigate the crude green and roasted coffee oil (GCO, RCO) and heated (HGCO, HRCO) for 1 h at 200°C, by Fourier Transform Infrared (FTIR) spectroscopy and in terms of antioxidant and antimicrobial properties. The results of FTIR spectroscopy revealed that no statistically significant differences (one-way ANOVA, p>0.05) in the oxidative status of GCO and RCO were found. The coffee oils heating induced significant spectral changes in the regions 3100–3600 cm^–1, 2800–3050 cm^–1 and 1680–1780 cm^–1 proved by the differences in the absorbance ratios A 3009 cm⁻¹/A 2922 cm⁻¹, A 3009 cm⁻¹/A 2853 cm⁻¹, A 3009 cm⁻¹/A 1744 cm⁻¹, A 1744 cm⁻¹/A 2922 cm⁻¹. These alterations were related to the reduction of the unsaturation degree due to primary and secondary oxidation processes of the lipid fraction. The radical scavenging ability of oils investigated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay revealed that the IC50 value of GCO was significantly lower than of RCO (p<0.05). The IC50 values of crude coffee oils were lower than those of heated samples. The antioxidant activity of oils was attributed to both antioxidant compounds with free-radical scavenging capacity and to lipids oxidation products generated by heating. In the first 6 h of incubation, the inhibitory activity of crude oils against E. coli and E. faecalis was not significantly different to the control (p>0.05). Also, HGCO and HRCO showed significantly different inhibitory potential related to the control (p<0.05). The heating induced statistically significant decreases in the effectiveness of coffee oils against the tested bacteria. GCO proved to be the most effective among investigated coffee oils against the tested bacteria.     

Dynamics of Microbial Communities on Marine Snow Aggregates: Colonization, Growth, Detachment, and Grazing Mortality of Attached Bacteria

We studied the dynamics of microbial communities attached to model aggregates (4-mm-diameter agar spheres) and the component processes of colonization, detachment, growth, and grazing mortality. Agar spheres incubated in raw seawater were rapidly colonized by bacteria, followed by flagellates and ciliates. Colonization can be described as a diffusion process, and encounter volume rates were estimated at about 0.01 and 0.1 cm³ h⁻¹ for bacteria and flagellates, respectively. After initial colonization, the abundances of flagellates and ciliates remained approximately constant at 10³ to 10⁴ and ~10² cells sphere⁻¹, respectively, whereas bacterial populations increased at a declining rate to >10⁷ cells sphere⁻¹. Attached microorganisms initially detached at high specific rates of ~10⁻² min⁻¹, but the bacteria gradually became irreversibly attached to the spheres. Bacterial growth (0 to 2 day⁻¹) was density dependent and declined hyperbolically when cell density exceeded a threshold. Bacterivorous flagellates grazed on the sphere surface at an average saturated rate of 15 bacteria flagellate⁻¹ h⁻¹. At low bacterial densities, the flagellate surface clearance rate was ~5 × 10⁻⁷ cm² min⁻¹, but it declined hyperbolically with increasing bacterial density. Using the experimentally estimated process rates and integrating the component processes in a simple model reproduces the main features of the observed microbial population dynamics. Differences between observed and predicted population dynamics suggest, however, that other factors, e.g., antagonistic interactions between bacteria, are of importance in shaping marine snow microbial communities.     

Integration of New Genes into Cellular Networks,and Their Structural Maturation

It has been recently discovered that new genes can originate de novo from noncoding DNA, and several biological traits including expression or sequence composition form a continuum from noncoding sequences to conserved genes. In this article, using yeast genes I test whether the integration of new genes into cellular networks and their structural maturation shows such a continuum by analyzing their changes with gene age. I show that 1) The number of regulatory, protein–protein, and genetic interactions increases continuously with gene age, although with very different rates. New regulatory interactions emerge rapidly within a few million years, while the number of protein–protein and genetic interactions increases slowly, with a rate of 2–2.25 × 10⁻⁸/year and 4.8 × 10⁻⁸/year, respectively. 2) Gene essentiality evolves relatively quickly: the youngest essential genes appear in proto-genes ∼14 MY old. 3) In contrast to interactions, the secondary structure of proteins and their robustness to mutations indicate that new genes face a bottleneck in their evolution: proto-genes are characterized by high β-strand content, high aggregation propensity, and low robustness against mutations, while conserved genes are characterized by lower strand content and higher stability, most likely due to the higher probability of gene loss among young genes and accumulation of neutral mutations.     

Cystathionine γ-Lyase-deficient Mice Require Dietary Cysteine to Protect against Acute Lethal Myopathy and Oxidative Injury

Cysteine is considered a nonessential amino acid in mammals as it is synthesized from methionine via trans-sulfuration. However, premature infants or patients with hepatic failure may require dietary cysteine due to a lack of cystathionine γ-lyase (CTH), a key trans-sulfuration enzyme. Here, we generated CTH-deficient (Cth^−/−) mice as an animal model of cystathioninemia/cystathioninuria. Cth^−/− mice developed normally in general but displayed hypercystathioninemia/hyperhomocysteinemia though not hypermethioninemia. When fed a low cyst(e)ine diet, Cth^−/− mice showed acute skeletal muscle atrophy (myopathy) accompanied by enhanced gene expression of asparagine synthetase and reduced contents of glutathione in livers and skeletal muscles, and intracellular accumulation of LC3 and p62 in skeletal myofibers; they finally died of severe paralysis of the extremities. Cth^−/− hepatocytes required cystine in a culture medium and showed greater sensitivity to oxidative stress. Cth^−/− mice exhibited systemic vulnerability to oxidative injury, which became more prominent when they were fed the low cyst(e)ine diet. These results reveal novel roles of trans-sulfuration previously unrecognized in mice lacking another trans-sulfuration enzyme cystathionine β-synthase (Cbs^−/−). Because Cbs^−/− mice display hyperhomocysteinemia and hypermethioninemia, our results raise questions against the homocysteine-based etiology of CBS deficiency and the current newborn screening for homocysteinemia using Guthrie''s method, which detects hypermethioninemia.     

Increased Corneal Epithelial Turnover Contributes to Abnormal Homeostasis in the Pax6+/? Mouse Model of Aniridia

We aimed to test previous predictions that limbal epithelial stem cells (LESCs) are quantitatively deficient or qualitatively defective in Pax6^+/− mice and decline with age in wild-type (WT) mice. Consistent with previous studies, corneal epithelial stripe patterns coarsened with age in WT mosaics. Mosaic patterns were also coarser in Pax6^+/− mosaics than WT at 15 weeks but not at 3 weeks, which excludes a developmental explanation and strengthens the prediction that Pax6^+/− mice have a LESC-deficiency. To investigate how Pax6 genotype and age affected corneal homeostasis, we compared corneal epithelial cell turnover and label-retaining cells (LRCs; putative LESCs) in Pax6^+/− and WT mice at 15 and 30 weeks. Limbal BrdU-LRC numbers were not reduced in the older WT mice, so this analysis failed to support the predicted age-related decline in slow-cycling LESC numbers in WT corneas. Similarly, limbal BrdU-LRC numbers were not reduced in Pax6^+/− heterozygotes but BrdU-LRCs were also present in Pax6^+/− corneas. It seems likely that Pax6^+/− LRCs are not exclusively stem cells and some may be terminally differentiated CD31-positive blood vessel cells, which invade the Pax6^+/− cornea. It was not, therefore, possible to use this approach to test the prediction that Pax6^+/− corneas had fewer LESCs than WT. However, short-term BrdU labelling showed that basal to suprabasal movement (leading to cell loss) occurred more rapidly in Pax6^+/− than WT mice. This implies that epithelial cell loss is higher in Pax6^+/− mice. If increased corneal epithelial cell loss exceeds the cell production capacity it could cause corneal homeostasis to become unstable, resulting in progressive corneal deterioration. Although it remains unclear whether Pax6^+/− mice have LESC-deficiency, we suggest that features of corneal deterioration, that are often taken as evidence of LESC-deficiency, might occur in the absence of stem cell deficiency if corneal homeostasis is destabilised by excessive cell loss.     

Fatty acid and very low density lipoprotein metabolism in obese African American and Caucasian women with type 2 diabetes

Type 2 diabetes mellitus (T2DM) is associated with increased plasma triglyceride (TG) concentrations, but African Americans (AA) have lower plasma TG than Caucasians (CC). We evaluated the hypothesis that obese AA women have lower plasma TG than obese CC women do because of differences in lipid kinetics. Eleven AA and 11 CC obese women with T2DM, matched on body mass index (BMI) (AA = 37 ± 1, CC = 37 ± 1 kg/m²), age, duration of diabetes, percentage body fat, and insulin sensitivity (S_I, determined by an intravenous glucose tolerance test), were studied. Plasma TG concentration (AA = 1.14 ± 0.11, CC = 1.88 ± 0.18 mmol/l), FFA rate of appearance (R_a) into plasma (AA = 419 ± 27, CC = 503 ± 31 µmol·min⁻¹), and total VLDL-TG secretion rate (AA = 18 ± 2, CC = 29 ± 4 µmol·min⁻¹) were lower in AA than CC women (all P < 0.05). In contrast, plasma total apolipoprotein (apo)B-100 concentration (AA = 1,542 ± 179, CC = 1,620 ± 118 nmol/l) and VLDL-apoB-100 secretion rate (AA = 1.3 ± 0.1, CC = 1.3 ± 0.1 nmol·min⁻¹) were similar in both groups, so the molar ratio of VLDL-TG secretion rate to VLDL-apoB-100 secretion rate was lower in AA women than in CC women. VLDL-TG concentration was lower in AA women due to lower total VLDL-TG secretion rate. However, the VLDL-apoB-100 secretion rate was the same in both groups, demonstrating that AA women secrete smaller VLDL particles containing less TG than do CC women.     

Mice Doubly-Deficient in Lysosomal Hexosaminidase A and Neuraminidase 4 Show Epileptic Crises and Rapid Neuronal Loss

Tay-Sachs disease is a severe lysosomal disorder caused by mutations in the HexA gene coding for the α-subunit of lysosomal β-hexosaminidase A, which converts G_M2 to G_M3 ganglioside. Hexa^−/− mice, depleted of β-hexosaminidase A, remain asymptomatic to 1 year of age, because they catabolise G_M2 ganglioside via a lysosomal sialidase into glycolipid G_A2, which is further processed by β-hexosaminidase B to lactosyl-ceramide, thereby bypassing the β-hexosaminidase A defect. Since this bypass is not effective in humans, infantile Tay-Sachs disease is fatal in the first years of life. Previously, we identified a novel ganglioside metabolizing sialidase, Neu4, abundantly expressed in mouse brain neurons. Now we demonstrate that mice with targeted disruption of both Neu4 and Hexa genes (Neu4 ^−/−;Hexa ^−/−) show epileptic seizures with 40% penetrance correlating with polyspike discharges on the cortical electrodes of the electroencephalogram. Single knockout Hexa ^−/− or Neu4 ^−/− siblings do not show such symptoms. Further, double-knockout but not single-knockout mice have multiple degenerating neurons in the cortex and hippocampus and multiple layers of cortical neurons accumulating G_M2 ganglioside. Together, our data suggest that the Neu4 block exacerbates the disease in Hexa^−/− mice, indicating that Neu4 is a modifier gene in the mouse model of Tay-Sachs disease, reducing the disease severity through the metabolic bypass. However, while disease severity in the double mutant is increased, it is not profound suggesting that Neu4 is not the only sialidase contributing to the metabolic bypass in Hexa ^−/− mice.