A prospective cohort study to identify and evaluate endotypes of venous thromboembolism: Rationale and design of the Genotyping and Molecular Phenotyping in Venous ThromboEmbolism project (GMP-VTE).

Several clinical, genetic and acquired risk factors for venous thromboembolism (VTE) have been identified. However, the molecular pathophysiology and mechanisms of disease progression remain poorly understood. This is reflected by uncertainties regarding the primary and secondary prevention of VTE and the optimal duration of antithrombotic therapy. A growing body of literature points to clinically relevant differences between VTE phenotypes (e.g. deep vein thrombosis (DVT) versus pulmonary embolism (PE), unprovoked versus provoked VTE). Extensive links to cardiovascular, inflammatory and immune-related morbidities are testament to the complexity of the disease. The GMP-VTE project is a prospective, multi-center cohort study on individuals with objectively confirmed VTE. Sequential data sampling was performed at the time of the acute event and during serial follow-up investigations. Various data levels (e.g. clinical, genetic, proteomic and platelet data) are available for multi-dimensional data analyses by means of advanced statistical, bioinformatic and machine learning methods. The GMP-VTE project comprises n = 663 individuals with acute VTE (mean age: 60.3 ±â€¯15.9 years; female sex: 42.8%). In detail, 28.4% individuals (n = 188) had acute isolated DVT, whereas 71.6% subjects (n = 475) had PE with or without concomitant DVT. In the study sample, 28.9% (n = 129) of individuals with PE and 30.1% (n = 55) of individuals with isolated DVT had a recurrent VTE event at the time of study enrolment. The systems-oriented approach for the comprehensive dataset of the GMP-VTE project may generate new biological insights into the pathophysiology of VTE and refine our current understanding and management of VTE.

'Simply to be let in': opening the doors to lower-income older adults and their companion animals.

Inspired by poet J.L. Borges's intimations on acceptance, this commentary highlights the eviction of an older woman and her kitten from an affordable independent living facility as representing exclusionary practices and policies that compromise the ability for some lower-income older adults to age-in-place. Even as efforts to promote aging-in-place have prioritized housing as a key need, and public health evidence suggests benefits from animal companionship later in life, there is a shortage of social and other types of affordable housing in Canada and elsewhere that allows older adult tenants to reside independently with companion animals. Within the private housing market, however, companion animals may be leveraged as a marketing tactic, with 'pets' being welcomed into rental arrangements. In light of this means-patterned discrepancy, this commentary troubles the persistent undervaluing of human-animal relationships that exists at policy and practice levels. Furthermore, banning companion animals from affordable housing subsumes an accepted yet insidious practise of regulating the lives of older adults who have not achieved idealized conceptions of responsible aging, including home ownership. We draw these two concerns together by advocating for adequate provision of affordable housing opportunities where lower-income older adults may be granted the choice to establish a home that includes a companion animal as they age-in-place.

Detection of MET mRNA in gastric cancer in situ. Comparison with immunohistochemistry and sandwich immunoassays.

Determination of predictive biomarkers by immunohistochemistry (IHC) relies on antibodies with high selectivity. RNA in situ hybridization (RNA ISH) may be used to confirm IHC and may potentially replace it if suitable antibodies are not available or are insufficiently selective to discriminate closely related protein isoforms. We validated RNA ISH as specificity control for IHC and as a potential alternative method for selecting patients for treatment with MET inhibitors. MET, the HGF receptor, is encoded by the MET proto-oncogene that may be activated by mutation or amplification. MET expression and activity were tested in a panel of control cell lines. MET could be detected in formalin fixed paraffin, embedded (FFPE) samples by IHC and RNA ISH, and this was confirmed by sandwich immunoassays of fresh frozen samples. Gastric cancer cell lines with high MET expression and phosphorylation of tyrosine-1349 respond to the MET inhibitor, BAY-853474. High expression and phosphorylation of MET is a predictive biomarker for response to MET inhibitors. We then analyzed MET expression and activity in a matched set of FFPE vs. fresh frozen tumor samples consisting of 20 cases of gastric cancer. Two of 20 clinical samples investigated exhibited high MET expression with RNA ISH and IHC. Both cases were shown by sandwich immunoassays to exhibits strong functional activity. Expression levels and functional activity in these two cases were in a range that predicted response to treatment. Our findings indicate that owing to its high selectivity, RNA ISH can be used to confirm findings obtained by IHC and potentially may replace IHC for certain targets if no suitable antibodies are available. RNA ISH is a valid platform for testing predictive biomarkers for patient selection.

Cardiovascular autonomic neuropathy contributes to left ventricular diastolic dysfunction in subjects with Type 2 diabetes and impaired glucose tolerance undergoing coronary angiography.

AIMS: Left ventricular diastolic dysfunction is considered a precursor of diabetic cardiomyopathy, while diabetic cardiovascular autonomic neuropathy is associated with an increased risk of mortality. This study aimed to evaluate the association between left ventricular diastolic dysfunction and cardiovascular autonomic neuropathy, both diagnosed according to the current guidelines. METHODS: We evaluated 145 patients referred for an elective coronary angiography, 52 of whom had Type 2 diabetes and 48 had impaired glucose tolerance, while 45 subjects had normal glucose tolerance. Cardiovascular autonomic neuropathy was diagnosed using autonomic function tests, while left ventricular diastolic dysfunction was verified by tissue Doppler imaging echocardiography. RESULTS: Cardiovascular autonomic neuropathy was diagnosed in 15 (28.8%) patients with Type 2 diabetes and in six (12.5%) individuals with impaired glucose tolerance. The rates of left ventricular diastolic dysfunction were 81 and 33% in patients with and without cardiovascular autonomic neuropathy, respectively (P < 0.001). In the cardiovascular autonomic neuropathy group (n = 21), early diastolic relaxation velocity (Em) was significantly reduced (5.4 ± 0.9 vs. 7.3 ± 2.1 cm/s; P < 0.001) and the E/Em ratio was significantly higher (13.6 ± 4.6 vs. 10.3 ± 3.4 cm/s, P < 0.001) as compared with the group without cardiovascular autonomic neuropathy (n = 79). These findings remained significant after adjustment for age, sex, coronary artery disease, hypertension and HbA(1c) . A severe form of left ventricular diastolic dysfunction was observed in 33 and 15% of patients with and without cardiovascular autonomic neuropathy, respectively (P = 0.001). CONCLUSION: Cardiovascular autonomic neuropathy is associated with a higher prevalence and a more severe form of left ventricular diastolic dysfunction in patients with diabetes or impaired glucose tolerance undergoing coronary angiography. Because both cardiovascular autonomic neuropathy and left ventricular diastolic dysfunction are associated with increased cardiovascular morbidity and mortality, screening for patients with left ventricular diastolic dysfunction and cardiovascular autonomic neuropathy with diabetes or impaired glucose tolerance may identify those at high risk.

Preimplantation genetic diagnosis and reproductive autonomy.

This paper examines how reproductive autonomy of women and couples is affected by the availability of preimplantation genetic diagnosis (PGD). Libertarians have argued that PGD enhances reproductive autonomy by increasing options available to prospective parents. We acknowledge that PGD presents prospective parents with more options, but note that the libertarian view of reproductive autonomy overlooks important aspects of the social practice of PGD by focusing too narrowly on the absence of explicit constraints as the main guarantee of reproductive autonomy. We endorse instead a feminist relational account of autonomy in which exercising autonomy involves paying attention to the relevant contextual features that configure and shape prospective parents' choices and understanding of their reproductive options. On this view, PGD has a negative potential to constrain reproductive autonomy. We recommend that this negative potential be addressed by making sure the availability of PGD is accompanied by increased attention to the relevant educational needs of prospective parents and the general public, as well as the availability of adequate social supports for people with disabilities, their care providers and families.

Acid/base transport in a model of the proximal tubule brush border: impact of carbonic anhydrase.

A mathematical model of the brush border of the proximal tubule (T. A. Krahn, P. S. Aronson, and A. M. Weinstein. Bull. Math. Biol, 56: 459-490, 1994) has been extended by the inclusion of CO2 and H2CO3 as diffusible species and by the inclusion of finite rate constants for the hydration of CO2. This permits the simulation of carbonic anhydrase (CA) activity and its inhibition. We confirm the result of our previous study, which is that, in the presence of CA, the unstirred layer has only a modest effect on the observed formic acid permeability. CA inhibition results in disequilibrium pH gradients, and the effect of these gradients on formic acid permeability depends on the presence of other membrane transport proteins. We also examined the impact of CA activity on the flux of total CO2 through the brush border. Under physiological conditions, CA inhibition depressed NaHCO3 reabsorption through the brush border by interfering with the HCO3(-)-facilitated diffusion of CO2. However, the determination of brush-border CO2 permeability, using an imposed CO2 gradient, was relatively uninfluenced by CA activity. Finally, we inserted a kinetic representation of the Na+/H+ exchanger into the brush-border model. Even when luminal and cytosolic diffusion coefficients were increased 1,000-fold, there was no effect on brush-border Na+ flux. This suggests that variations in the unstirred layer cannot be responsible for the flow dependence of Na+ reabsorption.

Weak acid permeability of a villous membrane: formic acid transport across rat proximal tubule.

Chloride/formate exchange, in parallel with Na+/H+ exchange and nonionic diffusion of H2CO2, has been proposed as a mechanism of electroneutral transcellular Cl- reabsorption by the proximal tubule. However, the measured brush border H2CO2 permeability of the rat proximal tubule is at least an order of magnitude too low to support sufficient H2CO2 recycling. To investigate the possibility that an unstirred layer within the brush border might depress the measured H2CO2 permeability, we constructed a mathematical model of a villous membrane. Axial fluxes along villous and intervillous spaces were specified by Nernst-Planck diffusion equations. Model parameters were set to achieve agreement with ion and water fluxes measured in the rat proximal tubule. The equations were solved numerically to generate steady-state concentration profiles in the villous and intervillous spaces. An apparent brush border H2CO2 permeability was determined by perturbing luminal [H2CO2] and calculating the change in H2CO2 flux. Overall, the ratio of apparent brush border H2CO2 permeability to cell membrane H2CO2 permeability was greater than 90%. Contributing to the small decrease in apparent permeability are finite diffusion coefficients, folding of the membrane, and acidification of the luminal solution. An approximate analysis of this system shows the critical parameters of brush border formate transport to be the actual membrane H2CO2 permeability, and the diffusion coefficients of HCO2- and HCO3-. Nevertheless, decreasing the diffusion coefficients by one order of magnitude failed to depress apparent brush border H2CO2 permeability by more than an additional 25%. We conclude that although permeability is systematically underestimated across a villous membrane, unstirred layer effects in the brush border are still too small to resolve the discrepancy between the measured value of H2CO2 permeability and the value needed to allow recycling.

Long-term follow-up of patients with avascular necrosis after treatment of slipped capital femoral epiphysis.

A retrospective review of 264 patients treated for slipped capital femoral epiphysis (SCFE) identified 36 who developed avascular necrosis (AVN). Twenty-two patients (24 hips) were evaluated at an average follow-up of 31 years. Nine have undergone reconstructive surgery: four during adolescence and five during adulthood. The remaining 13 patients (15 hips) have had no further operations, but all show degenerative changes on current roentgenograms. The natural history appears to be that of gradual degenerative changes for which reconstructive surgery most often can be delayed until adulthood.

Effect of isoprenaline on active Na transport in sheep cardiac Purkinje fibres.

The effect of isoprenaline (ISO; 5.10(-8) to 10(-6) M) on active Na transport was studied in depolarized sheep cardiac Purkinje fibres. Membrane current (I) and intracellular Na activity were measured simultaneously during enhanced Na pumping in voltage clamped preparations. ISO stimulated enhanced active Na transport but did not affect membrane current or intracellular Na concentration (ciNa) in the steady state under the chosen experimental conditions. The stimulatory effect of ISO was mediated by beta 1-adrenoceptors via a cAMP dependent pathway. The effect depended on the extracellular K concentration (coK) and was inhibited by external Ba ions. Complementary experiments on isolated sheep Purkinje cells revealed no ISO induced alteration of the Na pump current. The mechanism of the ISO induced stimulation of enhanced Na pumping in sheep Purkinje fibres probably involves an augmented K efflux. A direct effect on the pump molecule seems unlikely.

The dependence of sodium pump current on internal Na concentration and membrane potential in cardioballs from sheep Purkinje fibres.

The effect of various intracellular Na concentrations (CiNa) and membrane potentials on the Na pump current (Ip) was studied in isolated, cultured sheep cardiac Purkinje cells ('cardioballs'). Ip was identified as cardiac steroid sensitive current. The dependence of Ip on CiNa was investigated at a membrane potential of -40 mV by means of whole-cell recording from cardioballs internally perfused with media containing various Na concentrations. Internal perfusion with a Na free solution abolished Ip. The amplitude of Ip as a function of CiNa displayed saturation kinetics. Half maximal activation of Ip occurred at a CiNa of about 9 mM. The maximal Ip density was estimated to be 1.1 microA/cm2. The potential dependence of Ip was studied by conventional whole-cell recording under various ionic conditions. Generally Ip displayed little voltage dependence at membrane potentials positive to -20 mV. Ip declined at more negative potentials. The pump cycle probably includes only one voltage sensitive step. The potential dependence of Ip was more pronounced at lower CiNa or lower concentrations of the external pump activator Cs+. The findings are in line with the idea that increasingly steeper ionic gradients against which the cations are pumped strengthen the voltage dependence of Ip in the potential range studied. Other factors probably affecting the pump current-voltage (Ip-V) relation are discussed. The results suggest that Ip varies during electrical activity.

Activation of the Na pump current by external K and Cs ions in cardioballs from sheep Purkinje fibres.

The Na pump current (Ip) was measured at 30-33 degrees C as a function of the extracellular K+ or Cs+ concentrations ([K]o; [Cs]o) by means of whole cell recording from cardioballs isolated from sheep Purkinje fibres. The results show that the magnitude of Ip is an s-shaped, saturating function of [K]o or [Cs]o, respectively. Half maximal Ip activation occurs at 1.2 mM Ko or 3.2 mM Cso (clamp potential: -20 mV). These values are appreciably lower than earlier measurements on sheep Purkinje fibres indicated. The Hill coefficients for Ip activation by external K+ or Cs+ were calculated to be 1.94 and 1.73, respectively. The numbers suggest that Ip activation requires at least two external activator cations.