The KCl‐cotransporter KCC2 linked to epilepsy

In the mature brain, the neurotransmitter GABA can cause a postsynaptic hyperpolarization via activation of chloride permeant GABA_A receptor channels. This hyperpolarizing response critically depends on chloride extrusion via the KCl‐cotransporter KCC2 1 . Its knockdown in mice impairs synaptic inhibition by changing the electrochemical potential for chloride and thus increases neuronal excitability 2 3 . Two independent groups provide first evidence now, published in EMBO reports, that rare variants of KCC2 confer an increased risk of epilepsy in men 4 5 .     

Alteration in mitochondrial Ca<Superscript>2+</Superscript> uptake disrupts insulin signaling in hypertrophic cardiomyocytes

Background

Cardiac hypertrophy is characterized by alterations in both cardiac bioenergetics and insulin sensitivity. Insulin promotes glucose uptake by cardiomyocytes and its use as a substrate for glycolysis and mitochondrial oxidation in order to maintain the high cardiac energy demands. Insulin stimulates Ca²⁺ release from the endoplasmic reticulum, however, how this translates to changes in mitochondrial metabolism in either healthy or hypertrophic cardiomyocytes is not fully understood.

Results

In the present study we investigated insulin-dependent mitochondrial Ca²⁺ signaling in normal and norepinephrine or insulin like growth factor-1-induced hypertrophic cardiomyocytes. Using mitochondrion-selective Ca²⁺-fluorescent probes we showed that insulin increases mitochondrial Ca²⁺ levels. This signal was inhibited by the pharmacological blockade of either the inositol 1,4,5-triphosphate receptor or the mitochondrial Ca²⁺ uniporter, as well as by siRNA-dependent mitochondrial Ca²⁺ uniporter knockdown. Norepinephrine-stimulated cardiomyocytes showed a significant decrease in endoplasmic reticulum-mitochondrial contacts compared to either control or insulin like growth factor-1-stimulated cells. This resulted in a reduction in mitochondrial Ca²⁺ uptake, Akt activation, glucose uptake and oxygen consumption in response to insulin. Blocking mitochondrial Ca²⁺ uptake was sufficient to mimic the effect of norepinephrine-induced cardiomyocyte hypertrophy on insulin signaling.

Conclusions

Mitochondrial Ca²⁺ uptake is a key event in insulin signaling and metabolism in cardiomyocytes.

     

Chronic High-Fat Diet Impairs Collecting Lymphatic Vessel Function in Mice

Lymphatic vessels play an essential role in intestinal lipid uptake, and impairment of lymphatic vessel function leads to enhanced adipose tissue accumulation in patients with lymphedema and in genetic mouse models of lymphatic dysfunction. However, the effects of obesity on lymphatic function have been poorly studied. We investigated if and how adipose tissue accumulation influences lymphatic function. Using a lymphatic specific tracer, we performed in vivo near-infrared (NIR) imaging to assess the function of collecting lymphatic vessels in mice fed normal chow or high-fat diet (HFD). Histological and whole mount analyses were performed to investigate the morphological changes in initial and the collecting lymphatic vessels. HFD was associated with impaired collecting lymphatic vessel function, as evidenced by reduced frequency of contractions and diminished response to mechanostimulation. Moreover, we found a significant negative correlation between collecting lymphatic vessel function and body weight. Whole mount analyses showed an enlargement of contractile collecting lymphatic vessels of the hind limb. In K14-VEGF-C mice, HFD resulted in a reduced spreading of the tracer within dermal lymphatic vessels. These findings indicate that adipose tissue expansion due to HFD leads to a functional impairment of the lymphatic vasculature, predominantly in collecting lymphatic vessels.     

Type III restriction endonucleases are heterotrimeric: comprising one helicase–nuclease subunit and a dimeric methyltransferase that binds only one specific DNA

Fundamental aspects of the biochemistry of Type III restriction endonucleases remain unresolved despite being characterized by numerous research groups in the past decades. One such feature is the subunit stoichiometry of these hetero-oligomeric enzyme complexes, which has important implications for the reaction mechanism. In this study, we present a series of results obtained by native mass spectrometry and size exclusion chromatography with multi-angle light scattering consistent with a 1:2 ratio of Res to Mod subunits in the EcoP15I, EcoPI and PstII complexes as the main holoenzyme species and a 1:1 stoichiometry of specific DNA (sDNA) binding by EcoP15I and EcoPI. Our data are also consistent with a model where ATP hydrolysis activated by recognition site binding leads to release of the enzyme from the site, dissociation from the substrate via a free DNA end and cleavage of the DNA. These results are discussed critically in the light of the published literature, aiming to resolve controversies and discuss consequences in terms of the reaction mechanism.     

Accuracy of patient-reported height loss and risk factors for height loss among postmenopausal women

Background

Since loss of height may indicate vertebral fracture, the accuracy of the information on height is relevant for clinical practice. We undertook this study to compare reported and measured loss of height among post-menopausal women in a primary care setting. We also analyzed the determinants of this height loss.

Methods

In an observational study conducted between December 2007 and May 2008, we asked 1779 randomly selected general practitioners to recruit the first five female patients who were more than 60 years of age, regardless of the reason for the consultation. Using a questionnaire, physicians collected data on demographic and clinical variables, history of osteoporosis and current anti-osteoporotic treatment. We used three assessments of height: tallest height in early adulthood recalled by the patient, estimated current height reported by the patient at the visit and current measured height. We defined loss of height as the difference between the patient’s tallest recalled height and her current measured height.

Results

A total of 8610 patients were included in the analysis; the mean age was 70.9 (standard deviation [SD] 7.2) years. The mean loss of height was 4.5 cm. The mean current reported height was 2.1 (SD 2.5) cm lower than the tallest recalled height and 2.4 (SD 2.6) cm lower than the measured current height. The best predictors of a loss of height of 3 cm or more were age (odds ratio [OR] 1.09, 95% confidence interval [CI] 1.08–1.10), previous vertebral fracture (OR 1.49, 95% CI 1.16–1.91), previous nonvertebral fracture (OR 1.26, 95% CI 1.06–1.51), thoracic kyphosis (OR 2.07, 95% CI 1.69–2.55), scoliosis (OR 1.35, 95% CI 1.12–1.63), back pain (OR 1.22, 95% CI 1.07–1.39) and osteoporosis (OR 1.39, 95% CI 1.20–1.60).

Interpretation

Our study showed that the patients’ estimated current height was not correct, with a mean difference of −2.5 cm from the current measured height. The mean height loss was 4.5 cm. Previous vertebral fracture and thoracic kyphosis were strong determinants of the height loss.Loss of height is common with advancing age.^1,2 Causes include changes in curvature of the spine, narrowing of intervertebral discs and vertebral fractures. Height loss is associated with back pain and thoracic hyperkyphosis.^3,4 Two-thirds of adults have back pain at any time. Controversies exist about the need for radiographs of the spine: Does the benefit of detecting treatable disorders of the spine such as vertebral fracture outweigh the harm of unnecessary radiographs? Loss of height is usually recorded as one of the clinical signs to help identify postmenopausal women with vertebral fractures.⁵ The use of this parameter to decide whether radiography is needed depends on the threshold for height loss associated with a strong risk of vertebral fracture. The thresholds useful in clinical practice to detect prevalent vertebral fracture range from 3 cm to 6 cm,^6–9 with the risk of prevalent fracture increasing with the magnitude of the height loss. Thus, the accuracy of the information on height is relevant for clinical practice.We conducted this study to compare reported and measured loss of height in a large population of women more than 60 years old in a primary care setting and to analyze the determinants of this height loss.     

Amino-Acid Co-Variation in HIV-1 Gag Subtype C: HLA-Mediated Selection Pressure and Compensatory Dynamics

Background

Despite high potential for HIV-1 genetic variation, the emergence of some mutations is constrained by fitness costs, and may be associated with compensatory amino acid (AA) co-variation. To characterize the interplay between Cytotoxic T Lymphocyte (CTL)-mediated pressure and HIV-1 evolutionary pathways, we investigated AA co-variation in Gag sequences obtained from 449 South African individuals chronically infected with HIV-1 subtype C.

Methodology/Principal Findings

Individuals with CTL responses biased toward Gag presented lower viral loads than individuals with under-represented Gag-specific CTL responses. Using methods that account for founder effects and HLA linkage disequilibrium, we identified 35 AA sites under Human Leukocyte Antigen (HLA)-restricted CTL selection pressure and 534 AA-to-AA interactions. Analysis of two-dimensional distances between co-varying residues revealed local stabilization mechanisms since 40% of associations involved neighboring residues. Key features of our co-variation analysis included sites with a high number of co-varying partners, such as HLA-associated sites, which had on average 55% more connections than other co-varying sites.

Conclusions/Significance

Clusters of co-varying AA around HLA-associated sites (especially at typically conserved sites) suggested that cooperative interactions act to preserve the local structural stability and protein function when CTL escape mutations occur. These results expose HLA-imprinted HIV-1 polymorphisms and their interlinked mutational paths in Gag that are likely due to opposite selective pressures from host CTL-mediated responses and viral fitness constraints.     

Revision and affinities ofGalium (Rubiaceae) in Madagascar

The only representatives ofRubiaceae-Rubieae in Madagascar are five species ofGalium. G. thunbergianum andG. chloroionanthum are essentially afromontane species, ± widely distributed in both the African mainland and Madagascar. The new speciesG. andringitrense andG. ankaratrense are endemic to Madagascar but show close affinities to two afromontane—afroalpine groups from the African mainland, theG. simense—G. ruwenzoriense andG. glaciale complex respectively.G. polyacanthum (new combination) exhibits certain primitive morphological traits and appears to be an old Madagascan endemic without close allies. New chromosome counts for populations from the African mainland demonstrate thatG. chloroionanthum andG. simense are 4x (x=11). Distribution patterns and origins of theGalium species in Madagascar and their relationships with African taxa are discussed.