Thrombus resolution and hemodynamic recovery using ultrasound-accelerated thrombolysis in acute pulmonary embolism.

PURPOSE: To evaluate retrospectively the safety profile and clinical success of ultrasound-accelerated thrombolysis for acute pulmonary embolism (PE) with a standard lytic infusion protocol. MATERIALS AND METHODS: A retrospective study was performed at a single center treating patients with acute PE between October 2009 and April 2012. On diagnosis of submassive or massive PE by pulmonary computed tomography angiography or ventilation/perfusion scan, all patients received anticoagulation and treatment using the EkoSonic endovascular system (EKOS Corporation, Bothell, Washington). The ultrasound-accelerated thrombolytic infusion catheters were placed into the affected pulmonary arteries to facilitate administration of recombinant tissue plasminogen activator at 0.5-1.0mg/h/catheter. RESULTS: Treatment of 60 patients (35 men, 25 women; age 61 y±16; 53 bilateral PE; 48 submassive PE) resulted in complete thrombus clearance (≥90%) in 57% and near-complete (50%-90%) clearance in 41% of patients after infusion of 35.1 mg±11.1 of recombinant tissue plasminogen activator over 19.6 hours±6.0. Measurements before and after treatment showed a decrease in pulmonary artery pressure (47 mm Hg±15 to 38 mm Hg±12 [systolic], P<.001) and Miller score (25±3 to 17±6, P<.001). There were 57 patients who survived to discharge. All three patients who died in the hospital presented with massive PE. On 90-day follow-up, 56 patients (93%) were alive. CONCLUSIONS: The current study demonstrates effectiveness and safety of ultrasound-accelerated thrombolysis in patients with acute PE with a large thrombus burden.

Context-independent, temperature-dependent helical propensities for amino acid residues.

Assigned from data sets measured in water at 2, 25, and 60 degrees C containing (13)C=O NMR chemical shifts and [theta](222) ellipticities, helical propensities are reported for the 20 genetically coded amino acids, as well as for norvaline and norleucine. These have been introduced by chemical synthesis at central sites within length-optimized, spaced, solubilized Ala(19) hosts. The resulting polyalanine-derived, quantitative propensity sets express for each residue its temperature-dependent but context-independent tendency to forego a coil state and join a preexisting helical conformation. At 2 degrees C their rank ordering is: P << G < H < C, T, N < S < Y, F, W < V, D < K < Q < I < R, M < L < E < A; at 60 degrees C the rank becomes: H, P < G < C < R, K < T, Y, F < N, V < S < Q < W, D < I, M < E < A < L. The DeltaDeltaG values, kcal/mol, relative to alanine, for the cluster T, N, S, Y, F, W, V, D, Q, imply that at 2 degrees C all are strong breakers: DeltaDeltaG(mean) = +0.63 +/- 0.11, but at 60 degrees C their breaking tendencies are dramatically attenuated and converge toward the mean: DeltaDeltaG(mean) = +0.25 +/- 0.07. Accurate modeling of helix-rich proteins found in thermophiles, mesophiles, and organisms that flourish near 0 degrees C thus requires appropriately matched propensity sets. Comparisons are offered between the temperature-dependent propensity assignments of this study and those previously assigned by the Scheraga group; the special problems that attend propensity assignments for charged residues are illustrated by lysine guest data; and comparisons of errors in helicity assignments from shifts and ellipticity data show that the former provide superior precision and accuracy.

Field-recorded data on habitat, density, growth and movement of Nephrops norvegicus.

The availability of growth data in N. norvegicus is important for management purposes due to a lack of aging criteria and the commercial importance of fisheries in this species. Growth varies as a function of stock density, hence comparisons of growth rates between stocks at known density is particularly valuable. Growth is also related to starting size in males, making raw data on size-specific growth rates more valuable. Internally injected passive tags allowed us to track the growth of male and female individuals over one or two years. The spatial position of tagged recaptures was recorded to measure site fidelity of tagged releases. A total of 3300 pots were fished and their spatial positions were recorded to enable Catch Per Unit Effort calculations. Similarly, spatially geo-referenced v-notching and notched recovery enables spatially gridded densities to be calculated. Finally, acoustic mapping was carried out both on and off the fishing ground and was ground-truthed with sedimentology from grabs at 22 stations. These data are useful for fisheries and macroecological studies.

A Prospective, Single-Arm, Multicenter Trial of Ultrasound-Facilitated, Catheter-Directed, Low-Dose Fibrinolysis for Acute Massive and Submassive Pulmonary Embolism: The SEATTLE II Study.

OBJECTIVES: This study conducted a prospective, single-arm, multicenter trial to evaluate the safety and efficacy of ultrasound-facilitated, catheter-directed, low-dose fibrinolysis, using the EkoSonic Endovascular System (EKOS, Bothell, Washington). BACKGROUND: Systemic fibrinolysis for acute pulmonary embolism (PE) reduces cardiovascular collapse but causes hemorrhagic stroke at a rate exceeding 2%. METHODS: Eligible patients had a proximal PE and a right ventricular (RV)-to-left ventricular (LV) diameter ratio ≥0.9 on chest computed tomography (CT). We included 150 patients with acute massive (n = 31) or submassive (n = 119) PE. We used 24 mg of tissue-plasminogen activator (t-PA) administered either as 1 mg/h for 24 h with a unilateral catheter or 1 mg/h/catheter for 12 h with bilateral catheters. The primary safety outcome was major bleeding within 72 h of procedure initiation. The primary efficacy outcome was the change in the chest CT-measured RV/LV diameter ratio within 48 h of procedure initiation. RESULTS: Mean RV/LV diameter ratio decreased from baseline to 48 h post-procedure (1.55 vs. 1.13; mean difference, -0.42; p < 0.0001). Mean pulmonary artery systolic pressure (51.4 mm Hg vs. 36.9 mm Hg; p < 0.0001) and modified Miller Index score (22.5 vs. 15.8; p < 0.0001) also decreased post-procedure. One GUSTO (Global Utilization of Streptokinase and Tissue Plasminogen Activator for Occluded Coronary Arteries)-defined severe bleed (groin hematoma with transient hypotension) and 16 GUSTO-defined moderate bleeding events occurred in 15 patients (10%). No patient experienced intracranial hemorrhage. CONCLUSIONS: Ultrasound-facilitated, catheter-directed, low-dose fibrinolysis decreased RV dilation, reduced pulmonary hypertension, decreased anatomic thrombus burden, and minimized intracranial hemorrhage in patients with acute massive and submassive PE. (A Prospective, Single-arm, Multi-center Trial of EkoSonic® Endovascular System and Activase for Treatment of Acute Pulmonary Embolism (PE) [SEATTLE II]; NCT01513759).

Minimally invasive parathyroidectomy for recurrent or persistent hyperparathyroidism using carbon track localization.

BACKGROUND: The present study documents the use of carbon tracking to localize parathyroid adenomas in three patients with persistent or recurrent parathyroid disease. METHODS: Three patients requiring second or third operations for hyperparathyroidism were operated upon after the parathyroid lesion had been localized preoperatively using a suspension of carbon particles in water. The enlarged parathyroid glands were identified by using one or more of the following: computed axial tomography, magnetic resonance imaging, ultrasound or Sestamibi nuclear scan. The lesion was then localized with ultrasound or computed axial tomography and a carbon track was inserted from the lesion to the skin, leaving a small skin tattoo as a marker for the surgeon. Each patient underwent a minimally invasive parathyroid operation. RESULTS: For each of the three patients the recurrent or persistent parathyroid adenoma was successfully identified and removed via a small incision. CONCLUSION: Minimally invasive techniques for primary hyperparathyroidism are changing our approach to parathyroid surgery. However, a minimally invasive technique is less applicable when the disease is persistent or recurrent. Although, when the recurrent or persistent parathyroid adenoma can be identified, localization and carbon tracking have proved useful in allowing the surgeon to remove the lesion via a minimally invasive technique.

Chaotropic Anions Strongly Stabilize Short, N-Capped Uncharged Peptide Helicies: A New Look at the Perchlorate Effect.

Regiospecific binding of perchlorate ions to the N-terminus of short-chained template-substituted polyalanine sequences in water dramatically increases helicity.

Large Circular Dichroism Ellipticities for N-Templated Helical Polypeptides Are Inconsistent with Currently Accepted Helicity Algorithms.

An unprecedented, high degree of helicity as judged by CD spectroscopy is observed in N-templated model peptides of the type AcHel-(Ala4 Lys)n Ala2 -NH2 (AcHel-Ala peptide pictured; AcHel is an N-terminal helix-inducing template for polypeptides). These results raise concern over the current methods for determining 100 % helicity.

Helix-coil energetics for helix formers and breakers reflect context and temperature: mutants of helically robust, guest-sensitive homopeptide hosts.

The natural amino acids are primarily helix breakers at the low assignment temperatures characteristic of many studies, but recent genomic analyses of thermophilic proteins suggest that at high temperatures, some breakers may become strong helix formers. Moreover, the breaker/former inventory has not been previously characterized at the physiologically relevant temperature of 37 degrees C. The versatility of 13C==O NMR chemical shifts as helicity reporters allows construction of two mutant peptide series, tailored to expand the range of temperature assignments for helical propensities and derived from the core hosts tL-Ala9XxxAla9-tL and tL-AlaNva4XxxNva4Ala9-tL, Nva=norvaline. For three limiting guests Xxx, the helix former Nva and the breakers Gly and Pro, we report wXxx[T] assignments at seven temperatures from 2 to 80 degrees C, validating our reasoning and paving the way for assignment of a definitive wXxx[T] data-base.

Temperature- and length-dependent energetics of formation for polyalanine helices in water: assignment of w(Ala)(n,T) and temperature-dependent CD ellipticity standards.

Length-dependent helical propensities w(Ala)(n,T) at T = 10, 25, and 60 degrees C are assigned from t/c values and NMR 13C chemical shifts for series 1 peptides TrpLys(m)Inp2(t)Leu-Ala(n)(t)LeuInp2Lys(m)NH2, n = 15, 19, and 25, m = 5, in water. Van't Hoff analysis of w(Ala)(n,T) show that alpha-helix formation is primarily enthalpy-driven. For series 2 peptides Ac-Trp Lys5Inp2(t)Leu-(beta)AspHel-Ala(n)-beta-(t)LeuInp2Lys5NH2, n = 12 and 22, which contain exceptionally helical Ala(n) cores, protection factor-derived fractional helicities FH are assigned in the range 10-30 degrees C in water and used to calibrate temperature-dependent CD ellipticities [theta](lambda,H,n,T). These are applied to CD data for series 1 peptides, 12 < or = n < or = 45, to confirm the w(Ala)(n,T) assignments at T = 25 and 60 degrees C. The [theta](lambda,H,n,T) are temperature dependent within the wavelength region, 222 +/- 12 nm, and yield a temperature correction for calculation of FH from experimental values of [theta](222,n,T,Exp).

Energetic characterization of short helical polyalanine peptides in water: analysis of 13C=O chemical shift data.

Measured at 2 degrees C in water, NMR chemical shifts of (13)C=O labeled central alanine residues of peptides W-Lys(5)-(t)L(3)-Ala(n)-(t)L(3)-Lys(5)NH(2), n = 9, 11, 13, 15, 19 and W-Lys(5)-(t)L(3)-a-Ala(n)-A-Inp-(t)L(2)-Lys(5)NH(2) (a = D-Ala; (t)L = tert-leucine; Inp = 4-carboxypiperidine) are used to assign jt(L) and ct(L), the N- and C-terminal (t)L capping parameters and length-dependent values for w(Ala)(n), the alanine helical propensity for Ala(n) peptides. These parameters allow Lifson-Roig characterization of the stabilities of Ala(n)() helices in water. To facilitate chemical shift characterization, different (13)C/(12)C ratios are incorporated into specific Ala sites to code up to six residue sites per peptide. Large left/right chemical shift anisotropies are intrinsic to helical polyalanines, and a correcting L-R-based model is introduced. Capping parameters jt(L) = ct(L) lie in the range of 0.3 to 0.5; the (t)L residues are thus moderately helix-destabilizing. For helical conformations of lengths shorter than eight residues, assigned values for w(Ala) approach 1.0 but increase monotonically with length to a value of 1.59 for w(Ala)(19).

Water-solubilized, cap-stabilized, helical polyalanines: calibration standards for NMR and CD analyses.

NMR and CD studies are reported for two length series of solubilized, spaced, highly helical polyalanines that are N-capped by the optimal helix stabilizer (beta)Asp-Hel and C-capped by beta-aminoalanine beta and that are studied in water at 2 degrees C, pH 1-8. NMR analysis yields a structural characterization of the peptide Ac(beta)AspHelAla(8)betaNH(2) and selected members of one (beta)AspHelAla(n)beta series. At pH > 4.5 the (beta)AspHel cap provides a preorganized triad of carboxylate anion and two amide residues that is complementary to the helical polyalanine N-terminus. The C-terminal beta-aminoalanine assumes a helix-stabilizing conformation consistent with literature precedents. H(N)CO NMR experiments applied to capped, uniformly (13)C- and (15)N-labeled Ala(8) and Ala(12) peptides define Ala(n) hydrogen bonding signatures as alpha-helical without detectable 3(10) character. Relative NH-->ND exchange rates yield site protection factors PF(i) that define uniquely high fractional helicities FH for the peptide Ala(n) regions. These Ala(n) calibration series, studied in water and lacking helix-stabilizing tertiary structure, yield the first (13)C NMR chemical shifts, (3)J(HNH)(alpha) coupling constants, and CD ellipticities [theta(Molar)](lambda,n) characteristic of a fully helical alanine within an Ala(n) context. CD data are used to assign parameters X and [theta](lambda,infinity), required for rigorous calculation of FH values from CD ellipticities.

Consistent helicities from CD and template t/c data for N-templated polyalanines: progress toward resolution of the alanine helicity problem.

The helicity reporting parameters t/c and [theta](222) have been measured at 2, 25, and 60 degrees C in water for the solubilized polyalanine series Ac-Hel-A(n)-(t)LInp(2)K(4)W-NH(2) of length 4 < or = n < or = 14 that bears the helix-initiating and monitoring N-cap Ac-Hel and the spaced solubilizer (t)LInp(2)K(4)W-NH(2) as a C-cap. Correlation of t/c with length shows that the helical propensity for n < or = 6 is ca. 1.0, consistent with our early reports, but that a dramatic increase in temperature dependence and helical propensity occurs for n > or = 8. A model based on hydrogen-bonding cooperativity is proposed to explain this finding, and both t/c and [theta](222) are modeled successfully by length-dependent alanine propensities at 2 degrees C of 1.03 for n = 6, 1.15, for 7 < or = n < or = 9 and 1.26 for n > or = 10. The implications of these results for the energetics of helix formation by alanine-rich peptide sequences are discussed.

Solubilized, spaced polyalanines: a context-free system for determining amino acid alpha-helix propensities.

The logical design principles behind a system of properly water-solubilized, spaced polyalanines are presented. Peptides conforming to these design principles are shown to be unaggregated, and their helical properties as measured by the circular dichroism (CD) residue ellipticity at 222 nm, [theta](222), are shown to be dependent upon the lengths of their alanine regions. It is further demonstrated that CD contributions of the alanine cores are independent of the CD contributions attributable to other features of the peptides. The CD response of these polyalanines to variations in temperature and salt or denaturant concentration is described. CD data for a series of peptides with Ala(n) cores varying in length from 12 to 45 residues are presented that allow calculation of the helical propensity, w(Ala), in a purely alanine context. Mathematical modeling of these unprecedented data reveals the insufficiency of currently accepted literature helicity modeling parameters. A modification to the standard Lifson-Roig algorithm is introduced based on hydrogen-bonding cooperativity.

Dual wavelength parametric test of two-state models for circular dichroism spectra of helical polypeptides: anomalous dichroic properties of alanine-rich peptides.

A two-state helix-coil model underlies all calculations of fractional helicities FH from CD spectra of helical polypeptides. The presence of an isodichroic point near 203 nm is widely assumed to validate this model, but is shown here to provide inadequate validation for alanine-rich peptides. A parametric correlation with constant slope B between CD ellipticities at a pair of wavelengths is introduced as a more rigorous two-state test. Correlations of temperature-dependent [theta](222) vs [theta](208) values are reported for a variety of peptides. Constant slopes B are observed for literature CD data obtained from fragments of helical proteins and dimeric helical coiled coils, but parametric correlations of CD data for alanine-rich peptides consistently exhibit anomalous concave upward curvature, characterized by local slopes that are linearly temperature dependent. Low-temperature CD studies together with parametric correlations at a series of wavelengths demonstrate that the curvature anomaly is maximal at 222 nm and localized in the 215-230 nm wavelength region. Precedented structural variation of the phi, psi dihedral angles of the alpha-helix is suggested as a possible explanation. For the important case of alanine-rich peptides, experiments are proposed that may yield temperature corrections for [theta](222) and permit reliable calculations of FH from [theta](222) values.