Mechanisms of exercise pressor reflex dysfunction in heart failure

Doctor of PhilosophyDepartment of KinesiologySteven W CoppHeart failure patients with reduced ejection fraction (HF-rEF) have augmented sympathetic nervous system activity (SNA) at rest, and during exercise which is linked to decreased exercise tolerance and increased mortality. A reflex arising from within contracting skeletal muscles, termed the exercise pressor reflex, contributes importantly to that exaggerated SNA in HF-rEF. This reflex is activated when the sensory endings of thin fiber muscle afferents are stimulated by mechanical and/or metabolic signals associated with skeletal muscle contraction. Activation of this reflex in healthy subjects acts to increase and redistribute cardiac output and increase blood flow to the contracting skeletal muscles. Conversely, activation of this reflex in HF-rEF patients contributes to an exaggerated increase in SNA and augmented peripheral and coronary vasoconstriction and increases a heart failure patient’s risk of myocardial ischemia and fibrillation. Thus, in health the exercise pressor reflex functions to support the metabolic demands of exercise, whereas in HF-rEF the reflex becomes dysfunctional and impairs exercise tolerance. In this sequence of experiments presented in my dissertation, we used a rat model of myocardial infarction-induced HF-rEF (produced by coronary artery ligation). To isolate the exercise pressor reflex, we electrically stimulated the sciatic nerve to elicit hindlimb skeletal muscle contractions in decerebrated, unanesthetized rats. We also passively stretched the hindlimb skeletal muscle to isolate the activation of muscle afferents that are mechanically sensitive which allows us to study the mechanoreflex component of the exercise pressor reflex isolated from the metaboreflex component of the exercise pressor reflex. The contraction and stretch maneuvers produced an exaggerated increase in renal SNA (RSNA) and mean arterial pressure (MAP) in rats with HF-rEF compared to sham-operated control rats (SHAM rats). In the first study (chapter 2), we found that hindlimb arterial injection of a thromboxane A2 receptor (TxA₂-R) antagonist reduced the RSNA and MAP response to muscle stretch in rats with HF-rEF, but not SHAM rats. In the second study (chapter 3), we found that hindlimb arterial injection of a TxA₂-R antagonist reduced the RSNA and MAP response to muscle contraction in rats with HF-rEF, but not SHAM rats. In the third study (chapter 4), we found that preventing translocation of protein kinase C subtype epsilon (PKCε), an important second messenger which is translocated to the cell membrane when Gq proteins, such as TxA₂-Rs, are activated, reduced the RSNA and MAP response to muscle contraction and muscle stretch in rats with HF-rEF, but not SHAM rats. In the final study, we found that hindlimb arterial injection of an acid-sensing ion channel subtype 1a (ASIC1a) antagonist reduced the RSNA and MAP response to muscle contraction and RSNA response to muscle stretch in rats with HF-rEF, but not SHAM rats. The culmination of these studies indicates that TxA₂-R and ASIC1a on the sensory endings of thin fiber muscle afferents as well as second messenger signaling involving PKCε within the sensory ending of these afferents, contributes importantly to evoking an exaggerated increase in SNA during exercise in HF-rEF consequent to mechanoreflex and exercise pressor reflex activation

Activated factor IX, factor XI and tissue factor identify patients with permanent atrial fibrillation treated with warfarin who are at risk of ischemic stroke

Introduction: Previously, we have demonstrated that significant proportions of patients with various cardiovascular diseases have active tissue factor and active factor XIa in their plasma. In the current study, we evaluated active tissue factor and active factors (F)XI and FIX in plasma from patients with atrial fibrillation. Material and methods: In 110 consecutive patients with permanent atrial fibrillation receiving warfarin, we determined active tissue factor, together with plasma FIXa and FXIa, using clotting assays by measuring the response to inhibitory monoclonal antibodies. Results: Sixteen (14.5%) patients had detectable active tissue factor and active FXIa, including 11 subjects with both factors, while FIXa was observed in 28 (25.7%) patients. The three positive groups did not differ from the patients without these factors with regard to demographic and clinical characteristics. Von Willebrand factor was higher in the active tissue factor-positive group (p < 0.0001) and FXIa-positive group (p = 0.0037). Individuals positive for active tissue factor and FXIa had higher plasma interleukin-6 levels (p = 0.0014 and 0.0322, respectively). The presence of active tissue factor, FXIa and FIXa in anticoagulated patients with permanent atrial fibrillation correlated with elevated von Willebrand factor and interleukin-6. During a 3-year follow-up, ischemic stroke (n = 12, 10.9%) occurred more commonly among atrial fibrillation patients who had circulating TF (p = 0.002) or FXIa (p = 0.013). Conclusions: These data suggest that circulating active coagulation factors, in particular TF and FXIa, can be detected despite oral anticoagulation in a significant proportion of patients with atrial fibrillation, and could represent novel markers of persistent prothrombotic alterations predisposing to ischemic stroke

Delayed thrombin generation is associated with minor bleedings in venous thromboembolism patients on rivaroxaban : usefulness of calibrated automated thrombography

leeding is the most feared and difficult to predict adverse event of anticoagulation. We sought to investigate whether calibrated automated thrombography (CAT) parameters are associated with minor bleeding (MB) in anticoagulated patients following venous thromboembolism (VTE). Enrolled were 132 patients on rivaroxaban, 145 on vitamin K antagonists (VKA) and 31 controls who stopped anticoagulation. Prior to the next dose of the anticoagulant, we measured CAT parameters, along with rivaroxaban concentration and INR. During a median follow-up of 10 months, we recorded minor and major bleedings. On rivaroxaban, 27 (20.5%) patients with MB had longer time to start thrombin generation, lower peak thrombin generation and lower endogenous thrombin potential compared with subjects without MB (all p < 0.001). All CAT parameters, except for peak thrombin generation (p = 0.049), were similar in VKA patients with (n = 25, 17.2%) vs. without MBs. By logistic regression, time to start thrombin generation (p = 0.007) and unprovoked VTE (p = 0.041) independently predicted MBs on rivaroxaban. Major bleedings were more frequent in patients with MBs (17.3% vs. 1.8%, p < 0.001). Abnormal CAT parameters characterize VTE patients prone to MBs on rivaroxaban, but not on VKA. Time to start thrombin generation measured about 24 h since the last rivaroxaban dose might help predict MBs

Altered fibrin clot properties in advanced lung cancer : impact of chemotherapy

Background: Faster formation of dense and poorly lyzable fibrin networks have been reported in patients at risk of thromboembolism, including cancer patients. We sought to investigate whether chemotherapy affects plasma fibrin clot properties and their determinants in lung cancer patients. Methods: In this observational study we enrolled 83 consecutive patients with advanced inoperable lung cancer. Plasma fibrin clot permeability (Ks), turbidimetric analysis of clot formation, clot lysis time (CLT), microparticle-associated tissue factor (MP-TF) activity, and thrombin generation parameters were investigated at enrolment and 3–4 months after standard chemotherapy. Results: Lung cancer patients after 4 (range, 4–5) cycles of chemotherapy had 35.6% higher D dimer, 22.1% lower MP-TF activity, and unaltered fibrinogen compared with baseline. Chemotherapy resulted also in 7.5% increased Ks, 8.6% prolonged lag phase, and 5.4% shortened CLT, while thrombin generation was unchanged. Chemotherapy-related differences in clot structure were confirmed by scanning electron microscopy images. Fibrin clot properties after chemotherapy did not differ among histological types of lung cancer, cancer stages or chemotherapy regimens. Interestingly, never smoking (n=13, 16%) was associated with looser post-treatment fibrin structure as reflected by 12.3% higher Ks. Multiple linear regression showed that more advanced cancer stage, higher peak thrombin generation, and higher white blood cell count determined post-treatment change in Ks, while active smoking was associated with change in CLT. Conclusions: Three-month chemotherapy in lung cancer patients improves clot properties despite unaffected thrombin generation, suggesting that anticancer treatment might quickly produce antithrombotic actions

Thrombin generation profiles in deep venous thrombosis

BACKGROUND: Reliable markers and methods to predict risk for thrombosis are essential to clinical management. OBJECTIVE: Using an integrated approach that defines an individual’s comprehensive coagulation phenotype might prove valuable in identifying individuals at risk for experiencing a thrombotic event. METHODS: Using a numerical simulation model, we generated tissue factor (TF) initiated thrombin curves using coagulation factor levels from the Leiden Thrombophilia Study population and evaluated thrombotic risk, by sex, age, smoking, alcohol consumption, body mass index (BMI) and oral contraceptive (OC) use. We quantitated the initiation, propagation and termination phases of each individuals’ comprehensive TF-initiated thrombin generation curve by the parameters: time to 10nM thrombin, maximum time, level and rate (MaxR) of thrombin generated and total thrombin. RESULTS: The greatest risk association was obtained using MaxR; with a 2.6 fold increased risk at MaxR exceeding the 90(th) percentile. The odds ratio (OR) for MaxR was 3.9 in men, 2.1 in women, and 2.9 in women on OCs. The association of risk with thrombin generation did not differ by age (OR:2.8≤45 years>OR:2.5), BMI (OR:2.9≤26 kg/m(2)>OR:2.3) or alcohol use. In both numerical simulations and empirical synthetic plasma, OC use created extreme shifts in thrombin generation in both control women and women with a prior thrombosis, with a larger shift in thrombin generation in control women. This suggests an interaction of OC use with underlying prothrombotic abnormalities. CONCLUSIONS: Thrombin generation based upon the individual’s blood composition is associated with the risk for thrombosis and may be useful as a predictive marker for evaluating thrombosis on an individual basis

The Inhibitory Effect of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) on the Monophenolase and Diphenolase Activities of Mushroom Tyrosinase

In the present work, we investigated the effect of non-steroidal anti-inflammatory drugs (NSAIDs) on the monophenolase and diphenolase activity of mushroom tyrosinase. The results showed that diflunisal and indomethacin inhibited both monophenolase and diphenolase activity. For monophenolase activity, the lag time was extended in the presence of diflunisal. In the presence of indomethacin, the lag time did not change. IC50 values of monophenolase activity were estimated to be 0.112 mM (diflunisal) and 1.78 mM (indomethacin). Kinetic studies of monophenolase activity revealed that both diflunisal and indomethacin were non-competitive inhibitors. For diphenolase activity, IC50 values were estimated to be 0.197 mM (diflunisal) and 0.509 mM (indomethacin). Diflunisal and indomethacin were also found to be non-competitive diphenolase inhibitors

Analysis of the potential of cancer cell lines to release tissue factor-containing microvesicles: correlation with tissue factor and PAR2 expression

BackgroundDespite the association of cancer-derived circulating tissue factor (TF)-containing microvesicles and hypercoagulable state, correlations with the incidence of thrombosis remain unclear.MethodsIn this study the upregulation of TF release upon activation of various cancer cell lines, and the correlation with TF and PAR2 expression and/or activity was examined. Microvesicle release was induced by PAR2 activation in seventeen cell lines and released microvesicle density, microvesicle-associated TF activity, and phoshpatidylserine-mediated activity were measured. The time-course for TF release was monitored over 90 min in each cell line. In addition, TF mRNA expression, cellular TF protein and cell-surface TF activities were quantified. Moreover, the relative expression of PAR2 mRNA and cellular protein were analysed. Any correlations between the above parameters were examined by determining the Pearson’s correlation coefficients.ResultsTF release as microvesicles peaked between 30–60 min post-activation in the majority of cell lines tested. The magnitude of the maximal TF release positively correlated with TF mRNA (c = 0.717; p