The location of the tetraethylammonium (TEA) binding site in the outer vestibule of K
+ channels, and the mechanism by which external TEA slows C-type inactivation, have been considered ...well-understood. The prevailing model has been that TEA is coordinated by four amino acid side chains at the position equivalent to
Shaker T449, and that TEA prevents a constriction that underlies inactivation via a foot-in-the-door mechanism at this same position. However, a growing body of evidence has suggested that this picture may not be entirely correct. In this study, we reexamined these two issues, using both the Kv2.1 and
Shaker potassium channels. In contrast to results previously obtained with
Shaker, substitution of the tyrosine at Kv2.1 position 380 (equivalent to
Shaker 449) with a threonine or cysteine had a relatively minor effect on TEA potency. In both Kv2.1 and
Shaker, modification of cysteines at position 380/449 by 2-(trimethylammonium)ethyl methanethiosulfonate (MTSET) proceeded at identical rates in the absence and presence of TEA. Additional experiments in
Shaker demonstrated that TEA bound well to C-type inactivated channels, but did not interfere with MTSET modification of C449 in inactivated channels. Together, these findings rule out the possibility that TEA binding involves an intimate interaction with the four side chains at the position equivalent to
Shaker 449. Moreover, these results argue against the model whereby TEA slows inactivation via a foot-in-the-door mechanism at position 449, and also argue against the hypothesis that the position 449 side chains move toward the center of the conduction pathway during inactivation. Occupancy by TEA completely prevented MTSET modification of a cysteine in the outer-vestibule turret (Kv2.1 position 356/
Shaker position 425), which has been shown to interfere with both TEA binding and the interaction of K
+ with an external binding site. Together, these data suggest that TEA is stabilized in a more external position in the outer vestibule, and does not bind via direct coordination with any specific outer-vestibule residues.
The aim of this study was to evaluate the relation between myocardial perfusion and ST-segment changes in patients with acute myocardial infarction treated with successful direct angioplasty. ...Thirty-seven patients, successfully treated with direct angioplasty, underwent myocardial contrast echocardiography before and after angioplasty. The sum of ST-segment elevation divided by the number of the leads involved (ST-segment elevation index) was calculated at 1, 5, 10, 20, and 30 minutes after restoration of a Thrombolysis In Myocardial Infarction trial grade 3 flow. After recanalization, myocardial reperfusion within the risk area was observed in 26 patients, whereas a no-reflow phenomenon occurred in 11. In patients with myocardial reperfusion, the ST-segment elevation index progressively declined, whereas in patients with no reflow, no significant change was observed. Reduction of ≥50% in the ST-segment elevation index occurred in 20 of the 26 patients with reflow and in 1 of the 11 with no reflow (p = 0.0002). An additional increase of ≥30% in the ST-segment elevation index occurred in 3 patients with reflow and in 7 with no reflow (p = 0.003). Sensitivity, specificity, positive and negative predictive values, and accuracy of the reduction in the ST-segment elevation index for predicting microvascular reflow were 77%, 91%, 95%, 62%, and 81%, respectively. The corresponding values of the increase in ST-segment elevation index for predicting no reflow were 64%, 88%, 70%, 85%, and 81%, respectively. In conclusion, after successful angioplasty, different patterns of myocardial perfusion are associated with different ST-segment changes. Analysis of ST-segment changes predicts the degree of myocardial reperfusion.
Objectives. This prospective observational study was conducted to examine the apparent impact of a systematic direct percutaneous transluminal coronary angioplasty (PTCA) strategy on mortality in a ...series of 66 consecutive patients with acute myocardial infarction (AMI) complicated by cardiogenic shock, and to analyze the predictors of outcome after successful direct PTCA.
Background. Previous studies have reported encouraging results with PTCA in patients with AMI complicated by cardiogenic shock, but a biased case selection for PTCA may have heavily influenced the observed outcomes.
Methods. All patients admitted with AMI were considered eligible for direct PTCA, including those with the most profound shock, and no upper age limit was used. The treatment protocol also included stenting of the infarct-related artery for a poor or suboptimal angiographic result after conventional PTCA.
Results. Between January 1995 and March 1997, 364 consecutive patients underwent direct PTCA, and in 66 patients AMI was complicated by cardiogenic shock. In patients with cardiogenic shock, direct PTCA had a success rate of 94%; an optimal angiographic result was achieved in 85%; primary stenting of the infarct-related artery was accomplished in 47%; and the in-hospital mortality rate was 26%. Univariate analysis showed that patient age, chronic coronary occlusion and completeness of revascularization were significantly related to in-hospital mortality. The mean follow-up period was 16 ± 8 months. Survival rate at 6 months was 71%. Comparison of event-free survival in patients with a stented or nonstented infarct-related artery suggests an initial and long-term benefit of primary stenting.
Conclusions. Systematic direct PTCA, including stent-supported PTCA, can establish a Thrombolysis in Myocardial Infarction (TIMI) grade 3 flow in the great majority of patients presenting with AMI and early cardiogenic shock. High performance criteria, including new devices such as coronary stents, should be considered in randomized trials where mechanical revascularization therapy is being tested.
Objectives The aim of this study was to assess the effects of serelaxin on short-term changes in markers of organ damage and congestion and relate them to 180-day mortality in patients with acute ...heart failure. Background Hospitalization for acute heart failure is associated with high post-discharge mortality, and this may be related to organ damage. Methods The Pre-RELAX-AHF (Relaxin in Acute Heart Failure) phase II study and RELAX-AHF phase III study were international, multicenter, double-blind, placebo-controlled trials in which patients hospitalized for acute heart failure were randomized within 16 h to intravenous placebo or serelaxin. Each patient was followed daily to day 5 or discharge and at days 5, 14, and 60 after enrollment. Vital status was assessed through 180 days. In RELAX-AHF, laboratory evaluations were performed daily to day 5 and at day 14. Plasma levels of biomarkers were measured at baseline and days 2, 5, and 14. All-cause mortality was assessed as a safety endpoint in both studies. Results Serelaxin reduced 180-day mortality, with similar effects in the phase II and phase III studies (combined studies: N = 1,395; hazard ratio: 0.62; 95% confidence interval: 0.43 to 0.88; p = 0.0076). In RELAX-AHF, changes in markers of cardiac (high-sensitivity cardiac troponin T), renal (creatinine and cystatin-C), and hepatic (aspartate transaminase and alanine transaminase) damage and of decongestion (N-terminal pro–brain natriuretic peptide) at day 2 and worsening heart failure during admission were associated with 180-day mortality. Serelaxin administration improved these markers, consistent with the prevention of organ damage and faster decongestion. Conclusions Early administration of serelaxin was associated with a reduction of 180-day mortality, and this occurred with fewer signs of organ damage and more rapid relief of congestion during the first days after admission.
The objectives of this paper are to evaluate the efficacy and tolerability of topiramate, given at the dose of 100 mg/day, in the prophylactic treatment of migraine. The hypothesis that migraine is ...the result of a condition of neuronal hyperexcitability and the quest for drugs that are able to limit the number of crises justifies the attempt to utilise the new antiepileptic drugs in the prophylaxis of this pathology, which is so important due to its high prevalence and due to the high disability it causes. The study was randomised double-blind versus placebo, lasting 16 weeks, and was preceded by a run-in period of 4 weeks. One hundred and fifteen patients were randomly allocated to treatment with topiramate (TPM) or placebo: 35 patients completed the study in the TPM group and 37 patients in the placebo group. At the end of the double-blind phase of study, in the TPM group, we recorded a significant reduction in the frequency of migraine crises (from 5.26 at baseline to 2.60 in the last 4 weeks), a significant reduction in the quantity of symptomatic drugs taken as compared to the placebo control group (from 6.17+/-1.80 SD to 2.57+/-0.80) and a significant downward trend in the number of days of disability over the 16-week period of therapy. In the TPM group, side effects were transient and well tolerated. TPM has thus proven its efficacy and tolerability in the prophylaxis of migraine.
IFI 16 is a member of the HIN-200 protein family named for their haemopoietic expression, interferon-inducibility and nuclear localisation. These proteins have been characterised as transcriptional ...regulators that modulate the cell cycle. IFI 16 is expressed in some haemopoietic lineages including CD34+ progenitor cells, mature lymphocytes and monocytes, but is absent from granulocytes, erythrocytes and megakaryocytes. We present a wider study of IFI 16 expression in normal human tissues using a monoclonal antibody specifically recognising the C-terminus of IFI 16. As expected, IFI 16 was detected in the nuclei of lymphocytes in the spleen, thymus, lymph node and palatine tonsil, but was also found in epithelial cells in these tissues. Interestingly, IFI 16 protein was also expressed in non-lymphoid tissues including trachea, gastrointestinal tract, skin and testis, but was absent from others including heart and brain. In each tissue, IFI 16 was predominantly expressed in surface epithelial cells and staining was strongest in basal epithelial layers. Therefore, IFI 16 expression is not restricted to cells of the immune system, but is also expressed in epithelial cells. In contrast to the perceived role of HIN-200 proteins as suppressors of cell growth, maximal expression of IFI 16 was in cells with high proliferative potential.
Overexpression of Bcl-2 contributes to resistance of cancer cells to human cytotoxic lymphocytes (CL) by blocking granzyme B (GraB)-induced mitochondrial outer membrane permeabilization (MOMP). Drugs ...that neutralise Bcl-2 (e.g., ABT-737) may therefore be effective adjuvants for immunotherapeutic strategies that use CL to kill cancer cells. Consistent with this we found that ABT-737 effectively restored MOMP in Bcl-2 overexpressing cells treated with GraB or natural killer cells. This effect was observed even if ABT-737 was added up to 16 h after GraB, after which the cells reset their resistant phenotype. Sensitivity to ABT-737 required initial cleavage of Bid by GraB (gctBid) but did not require ongoing GraB activity once Bid had been cleaved. This gctBid remained detectable in cells that were sensitive to ABT-737, but Bax and Bak were only activated if ABT-737 was added to the cells. These studies demonstrate that GraB generates a prolonged pro-apoptotic signal that must remain active for ABT-737 to be effective. The duration of this signal is determined by the longevity of gctBid but not activation of Bax or Bak. This defines a therapeutic window in which ABT-737 and CL synergise to cause maximum death of cancer cells that are resistant to either treatment alone, which will be essential in defining optimum treatment regimens.
The human HIN-200 family member AIM2 was originally identified in a screen for suppressors of melanoma tumorigenicity following introduction of chromosome 6 into the UACC903 human melanoma cell line. ...Although the AIM2 protein contained many of the conserved structural motifs common to other HIN-200 proteins, the biochemical characteristics of AIM2 and the ability of overexpressed AIM2 to phenocopy the effect of introduction of chromosome 6 in the UACC903 cells had not been assessed. Herein we demonstrated that AIM2 was localised within the nucleus of transfected or interferon-treated human cells. In addition, AIM2 could homodimerise via the amino-terminal (PAAD/DAPIN) region and heterodimerise with the related IFI 16 protein. However, overexpressed AIM2 did not significantly affect the growth or survival of UACC903 cells or another human melanoma cell line. These data indicate that AIM2 has many of the biochemical and structural characteristics of HIN-200 proteins, however, its expression is not sufficient to induce a tumor-suppressor-like phenotype.
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