HSV1-specific thymidylate kinase activity in infected cells

Several 5-methoxymethyldeoxyuridine (MMdU)-resistant mutants of herpes simplex virus type 1 (HSV1) were classified by measuring their sensitivities to the deoxythymidine kinase (dTK)-dependent antiviral drugs 9-(2-hydroxyethoxymethyl)-guanine (acyclovir, ACV), 1-beta-D-arabinofuranosylthymine (araT), and E-(2)-5-bromovinyldeoxyuridine (BVdU) and to the dTK-independent antiviral drug phosphonoacetate (PAA). Compared to wild-type (WT) virus, all five of the dTK- mutants were highly resistant (greater than or equal to 500-fold) to BVdU and MMdU, moderately resistant to ACV (50- to 100-fold) and araT (10- to 20-fold), but not resistant to PAA. The dTK of the mutant MMdUr-20 (dTK+) appeared to phosphorylate dTMP less well than that of the WT virus, while its affinity for deoxythymidine was not altered. Two other drug-resistant HSV mutants-S1 (isolated against ACV) and B3 (isolated against BVdU)--also showed reduced phosphorylation of dTMP. This suggests that alterations in both dTK and thymidylate kinase activities may determine sensitivity to antiviral drugs.     

Evaluation of sampling methods in research reported in selected clinical nursing journals: implications for nursing practice

This article summarizes standards relating to sampling methodology, identifies deviations from these standards in research studies reported in selected clinical nursing journals, and provides suggestions for improving sampling methods to enhance the applicability of research for nursing practice. A random sample of 30 research reports published in 1986 in five clinical nursing journals was examined. Nearly 97 per cent of the published studies contained at least one major deficiency in sampling methodology. More than two thirds failed to describe the sampling frame, sample size, or number of refusals, withdrawals, and/or cases lost. Thirteen per cent did not report sampling methods. More than half made generalizations that were inappropriate for the sampling method used; 43 per cent did not acknowledge any limitations of their sample. Sample sizes were small, and statistical power to detect significant differences was low. These deficiencies in sampling procedures could detract from the value of the research that nurses are encouraged to use as a basis for practice. This article provides specific recommendations for remedying these deficiencies to help ensure the scientific merit of the research published for nursing practice.     

Bright circularly polarized soft X-ray high harmonics for X-ray magnetic circular dichroism

We demonstrate, to our knowledge, the first bright circularly polarized high-harmonic beams in the soft X-ray region of the electromagnetic spectrum, and use them to implement X-ray magnetic circular dichroism measurements in a tabletop-scale setup. Using counterrotating circularly polarized laser fields at 1.3 and 0.79 µm, we generate circularly polarized harmonics with photon energies exceeding 160 eV. The harmonic spectra emerge as a sequence of closely spaced pairs of left and right circularly polarized peaks, with energies determined by conservation of energy and spin angular momentum. We explain the single-atom and macroscopic physics by identifying the dominant electron quantum trajectories and optimal phase-matching conditions. The first advanced phase-matched propagation simulations for circularly polarized harmonics reveal the influence of the finite phase-matching temporal window on the spectrum, as well as the unique polarization-shaped attosecond pulse train. Finally, we use, to our knowledge, the first tabletop X-ray magnetic circular dichroism measurements at the N_4,5 absorption edges of Gd to validate the high degree of circularity, brightness, and stability of this light source. These results demonstrate the feasibility of manipulating the polarization, spectrum, and temporal shape of high harmonics in the soft X-ray region by manipulating the driving laser waveform.High-harmonic generation (HHG) results from an extreme nonlinear quantum response of atoms to intense laser fields. When implemented in a phase-matched geometry, bright, coherent HHG beams can extend to photon energies beyond 1.6 keV (1, 2). For many years, however, bright HHG was limited to linear polarization, precluding many applications in probing and characterizing magnetic materials and nanostructures, as well as chiral phenomena in general. Although X-ray optics can in principle be used to convert extreme UV (EUV) and X-ray light from linear to circular polarization, in practice such optics are challenging to fabricate and have poor throughput and limited bandwidth (3). A more appealing option is the direct generation of elliptically polarized (4–6) and circularly polarized (7–9) high harmonics. In recent work we showed that by using a combination of 0.8 and 0.4 µm counterrotating driving fields, bright (i.e., phase-matched) EUV HHG with circular polarization can be generated at wavelengths λ > 18 nm and used for EUV magnetic dichroism measurements (10–13).Here we make, to our knowledge, the first experimental demonstration of circularly polarized harmonics in the soft X-ray region to wavelengths λ < 8 nm, and use them to implement soft X-ray magnetic circular dichroism (XMCD) measurements using a tabletop-scale setup. By using counterrotating driving lasers at 0.79 µm (1.57 eV) and 1.3 µm (0.95 eV), we generate bright circularly polarized soft X-ray HHG beams with photon energies greater than 160 eV (14) and with flux comparable to the HHG flux obtained using linearly polarized 800-nm driving lasers (15). Moreover we implement, to our knowledge, the first advanced simulations of the coherent buildup of circularly polarized high harmonics to show how the macroscopic phase-matching physics and ellipticity of the driving lasers influence the HHG spectra, number of bright attosecond bursts, and the degree of circular polarization.This work presents several new capabilities and findings. First, circularly polarized HHG provides a unique route for generating bright narrowband (λ/Δλ > 400) harmonic peaks in the soft X-ray region, to complement the soft X-ray supercontinua that are produced with linearly polarized mid-IR lasers (2, 15, 16). This capability is significant because it provides an elegant and efficient route for shaping soft X-ray light by manipulating the driving laser light, and is very useful for applications in high-resolution coherent imaging (17–21) and photoelectron spectroscopies. Second, we show that the macroscopic phase-matching physics of circularly polarized soft X-ray HHG driven by mid-IR lasers has similarities to linearly polarized HHG, where the number of bright attosecond bursts is limited by the finite phase-matching temporal window. Third, we implement the first tabletop XMCD measurements at the N_4,5 absorption edges of Gd. The Gd/Fe multilayer sample is a candidate material for next-generation all-optical magnetic storage devices (22), but has been inaccessible to HHG XMCD until now. This capability also opens up the possibility of probing spin dynamics in rare-earth elements using HHG, which has been successfully used for 3d transition metals to uncover the fastest spin dynamics using EUV HHG (23, 24). Finally, and most importantly, these results demonstrate the universal nature of circularly polarized HHG that can be generated across the EUV and soft X-ray spectral regions using a broad range of driving laser wavelengths.     

Successful treatment of invasive aspergillosis in a heart transplant patient

Infection continues to be a major cause of mortality in cardiac transplant patients, and even though there has been significant progress in diagnosing and treating many infectious disease problems, invasive aspergillosis in the transplant patient represents a serious and usually fatal complication. Even with successful early diagnosis, the use of free amphotericin B (a polyene antibiotic) has failed to cure disseminated infection. We report the case of a 46-year-old transplant patient who failed to respond to treatment with free amphotericin B after a 7-day period of treatment for biopsy-proven pulmonary aspergillosis. However, a subsequent substitution of a liposomal form of amphotericin B was used, and the patient responded well to a total dose of 1.5 mg. After 7 months, the patient continues free of infection. This experience suggests that the introduction of liposomal amphotericin B may give new hope for treating an otherwise lethal infection.     

Sequence analyses of herpesviral enzymes suggest an ancient origin for human sexual behavior.

Comparison of the amino acid sequences of the deoxythymidine kinases of herpes simplex (HSV) and of marmoset herpes viruses (MHV) suggests a divergence time of 8 to 10 million years ago for HSV-1 and -2. Like MHV, HSV-1 and -2 cause local infections in their natural hosts, and direct contact between two individuals during the brief period of infectivity is needed for transmission. Because B virus, a nearer relative of HSV, depends on both oral and genital routes of transmission, we postulate that ancestral HSV (aHSV) was similar, and that for HSV-1 and -2 to diverge, genital and oral sites had to become microbiologically somewhat isolated from each other, while oral--oral and genital--genital contact had to be facilitated to maintain both aHSV strains. We propose that acquisition of continual sexual attractiveness by the ancestral human female and the adoption of close face-to-face mating, two hallmarks of human sexual behavior, provided the conditions for the divergence.     

Vascular endothelial growth factor-165 gene therapy promotes cardiomyogenesis in reperfused myocardial infarction

BACKGROUND: Vascular endothelial growth factor (VEGF)-165 promotes cardiomyogenesis in chronic myocardial ischemia and nonreperfused myocardial infarction (MI). It is unknown whether this effect is present in reperfused MI. We sought to investigate the effect of VEGF-165 gene therapy on cardiomyogenesis after reperfused MI. METHODS AND RESULTS: Twenty-four Yucatan minipigs underwent thoracotomy and a vascular clamp was placed in the left circumflex artery. Reperfusion was reestablished after 90 minutes, and VEGF-165 gene therapy or placebo was administered. A replication-deficient recombinant human adenovirus serotype 5 was used for gene transfer (Ad5-VEGF165). The same viral vector devoid of VEGF gene (Ad5-beta-galactosidase) was used as placebo. Two administration routes were tested, intramyocardial (IM) injection and circumflex intracoronary (IC) infusion. The pigs were assigned to one of the following groups: IM Ad5-VEGF165 (n = 6), IM Ad5-betaGal (n = 6), IC Ad5-VEGF165 (n = 6), and IC Ad5-betaGal (n = 6). All pigs received 5-bromo-2'-deoxyuridine (BrdU) 250 mg IV twice a week to label cells undergoing DNA replication. The hearts were explanted at 4 weeks. BrdU-labeled cardiomyocytes in the peri-infarct area were counted by a pathologist blinded to group assignment. The number of BrdU-labeled cardiomyocytes per million cells was 4-fold higher in the group receiving IM VEGF-165 (64 +/- 11.4) vs. IM placebo (16 +/- 10.6), P = 0.034. No difference in infarct size or ventricular function was observed between the groups. CONCLUSIONS: IM VEGF-165 gene therapy promotes cardiomyogenesis in reperfused MI. However, no benefit in infarct size or cardiac function was observed at 4 weeks. The origin of these cells remains unknown and needs to be determined.     

Pneumococcal vaccine response in cirrhosis and liver transplantation

Cirrhosis is a major risk factor for severe pneumococcal infection, and patients evaluated for liver transplantation routinely receive pneumococcal vaccine. This study followed serologic antibody levels of 45 adults evaluated for transplantation and 13 age-matched control subjects. All received 23-valent pneumococcal polysaccharide vaccine (PPS). Serum anti-PPS levels and antibodies specific for capsular types 3 and 23 were measured by ELISA before and 1 and 6 months after vaccination. Antibody levels for the 25 patients who received transplants also were measured immediately before and 3 months after transplantation. Control subjects had higher IgG responses to the whole vaccine, whereas patients appeared to produce more IgM and IgA. IgA, and possibly IgM levels, also declined faster in patients than in control subjects. All anti-PPS levels were at or below prevaccination baselines by 3 months after transplantation. These data suggest that vaccination with PPS may not be effective for patients during and after liver transplantation.     

Monocytes expressing CX3CR1 orchestrate the development of vincristine-induced pain

A major dose-limiting side effect associated with cancer-treating antineoplastic drugs is the development of neuropathic pain, which is not readily relieved by available analgesics. A better understanding of the mechanisms that underlie pain generation has potential to provide targets for prophylactic management of chemotherapy pain. Here, we delineate a pathway for pain that is induced by the chemotherapeutic drug vincristine sulfate (VCR). In a murine model of chemotherapy-induced allodynia, VCR treatment induced upregulation of endothelial cell adhesion properties, resulting in the infiltration of circulating CX3CR1⁺ monocytes into the sciatic nerve. At the endothelial-nerve interface, CX3CR1⁺ monocytes were activated by the chemokine CX3CL1 (also known as fractalkine [FKN]), which promoted production of reactive oxygen species that in turn activated the receptor TRPA1 in sensory neurons and evoked the pain response. Furthermore, mice lacking CX3CR1 exhibited a delay in the development of allodynia following VCR administration. Together, our data suggest that CX3CR1 antagonists and inhibition of FKN proteolytic shedding, possibly by targeting ADAM10/17 and/or cathepsin S, have potential as peripheral approaches for the prophylactic treatment of chemotherapy-induced pain.     

New insights into mechanism of Eustachian tube ventilation based on cine computed tomography images

There is debate concerning the mechanism of Eustachian tube (ET) ventilation. While a mechanism of complete opening has been advocated previously, sequential contraction of the levator veli palatini and medial pterygoid muscles followed by the tensor veli palatini and lateral pterygoid muscles may produce a transient sequential opening mechanism, allowing an air bolus to traverse the ET. This may explain confusion surrounding sonotubometry reports that not every swallow leads to sound passage in normal subjects. We hypothesize that the ET may not need to open completely when ventilating the middle ear; rather, a discrete air bolus can pass through it. Five normal and five disordered subjects underwent low-radiation dose cine computed tomography (CT) scans of the ET. Sixteen contiguous 2.5?mm slice locations were chosen through a 4?cm area in the nasopharynx that were parallel to and encompassed the entire ET. Twelve images were acquired at each slice over 4.8?s during swallowing and other tasks. Serial images were analyzed. An air bolus was observed passing through the ET in the normal subjects, but not the subject with ET dysfunction. Medial and lateral pterygoid contractions were also observed. A new hypothetical mechanism of transient sequential ET ventilation is presented. This is not a definitive conclusion, as the number of scans taken and maneuvers used was limited. Improved understanding of ET ventilation may facilitate management of middle ear disease as treatment evolves from ventilatory tube placement to ET manipulation.