Flecainide attenuates rate adaptation of ventricular repolarization in guinea-pig heart

Objectives: Flecainide is class Ic antiarrhythmic agent that was found to increase the risk of sudden cardiac death. Arrhythmic responses to flecainide could be precipitated by exercise, suggesting a role played by inappropriate rate adaptation of ventricular repolarization. This study therefore examined flecainide effect on adaptation of the QT interval and ventricular action potential duration (APD) to abrupt reductions of the cardiac cycle length. Design: ECG and ventricular epicardial and endocardial monophasic APD were recorded in isolated, perfused guinea-pig heart preparations upon a sustained cardiac acceleration (rapid pacing for 30 s), and following a single perturbation of the cycle length evoked by extrasystolic stimulation. Results: Sustained increase in heart rate was associated with progressive bi-exponential shortening of the QT interval and APD. Flecainide prolonged ventricular repolarization, delayed its rate adaptation, and decreased the amplitude of QT interval and APD shortening upon rapid cardiac pacing. During extrasystolic stimulation, flecainide attenuated APD shortening in premature ventricular beats, with effect being greater upon using a longer basic drive cycle length (S₁-S₁=550 ms versus S₁-S₁=300 ms). Conclusions: Flecainide-induced arrhythmia may be partly accounted for by attenuated adaptation of ventricular repolarization to sudden changes in cardiac cycle length provoked by transient tachycardia or ectopic beats.     

Nanocrystalline TiO2 doped by small amount of pre-synthesized colloidal CdS nanoparticles for photocatalytic degradation of 1,2,4-trichlorobenzene

The chemical stability and hydrophobic nature of chloroarenes make them a persistent environmental hazard. Modeling of 1,2,4-trichlorobenzene (1,2,4-TCB) degradation in alcohol-water solution under UV irradiation was carried out with the aim of probing how the 1,2,4-TCB might behave in the environment. The photocatalytic activity of both bare TiO₂ and TiO₂ doped by colloidal CdS nanoparticles synthesized by the sol-gel method has been investigated in the processes of 1,2,4-TCB photodegradation in the aqueous protic solvent. Non-sensitized TiO₂ cannot be regarded as catalyst for the 1,2,4-TCB photodecomposition. On the contrary, the CdS/TiO₂ composite accelerated the 1,2,4-TCB photodegradation process. The concentration of CdS/TiO₂ was shown to effect on the 1,2,4-TCB photolysis mechanisms, which resulted in the quantitative ratios of the 1,2,4-TCB photolysis products.     

Photoinactivation of Mycobacteria in vitro and in a new murine model of localized Mycobacterium bovis BCG-induced granulomatous infection

Treatment of tuberculosis is currently hindered by prolonged antibiotic regimens and the emergence of significant drug resistance. Alternatives and adjuncts to standard antimycobacterial agents are needed. We propose that a direct attack utilizing photosensitizers and light-based treatments may be effective in curtailing Mycobacterium tuberculosis in discrete anatomical sites in the most infectious phase of pulmonary tuberculosis. To demonstrate experimental proof of principle, we have applied established photodynamic therapy (PDT) technology to in vitro cultures and an in vivo mouse model using Mycobacterium bovis BCG. We report here in vitro and in vivo PDT efficacy studies and the use of a three-dimensional collagen gel as a delivery vehicle for BCG, subcutaneously inserted, to induce specifically localized granuloma-like lesions in mice. When a benzoporphyrin derivative was utilized as the photosensitive agent, exposure to light killed extracellular and intracellular BCG in significant numbers. Collagen scaffolds containing BCG inserted in situ in BALB/c mice for 3 months mimicked granulomatous lesions and demonstrated a marked cellular infiltration upon histological examination, with evidence of caseating necrosis and fibrous capsule formation. When 10(5) BCG were present in the in vivo-induced granulomas, a significant reduction in viable mycobacterial cells was demonstrated in PDT-treated granulomas compared to those of controls. We conclude that PDT has potential in the treatment of localized mycobacterial infections, such as pulmonary granulomas and cavities.     

Vaccination with photodynamic therapy-treated macrophages induces highly suppressive T-regulatory cells

Background/purpose: The present study explores whether photodynamic therapy (PDT)‐induced apoptosis can increase the number of tolerogenic regulatory T cells (Treg) and limit collateral tissue damage. Methods: BALB/c mice were vaccinated subcutaneously three times with PDT‐induced apoptotic or thaw‐frozen, necrotic non‐infected autologous macrophages (MΦ). Two weeks after the last vaccination, mice were infected intradermally with 10⁶ promastigotes of Leishmania major. Results: Mice that received PDT‐induced apoptotic MΦ had fewer parasites and higher numbers of Treg than mice vaccinated with thaw‐frozen necrotic MΦ or phosphate‐buffered saline (PBS). Interleukin (IL)‐4 and IL‐6 were significantly suppressed, while IL‐10 was increased in mice that received the PDT‐induced apoptotic MΦ. The role of Treg in this process was confirmed through Treg transfer from vaccinated to naïve mice. Mice receiving CD4⁺CD25⁺ cells from mice vaccinated with PDT‐induced apoptotic MΦ showed smaller lesions 3 weeks after infection and lower parasitic burdens than mice that received Tregs from mice of thaw‐frozen necrotic MΦ or PBS groups. These changes were mediated by the depletion of CD3⁺CD8⁺ and NKT cells and increased levels of IL‐12p70 and interferon‐γ, IL‐10, and TGF‐β in the cutaneous leishmaniasis lesions. Conclusion: Vaccination with apoptotic MΦ‐induced tolerogenic Treg cells that limited collateral tissue damage and diminished parasitic burden.     

Expression of lysostaphin in HeLa cells protects from host cell killing by intracellular Staphylococcus aureus

The Staphylococcus aureus-specific cell wall endopeptidase lysostaphin was used as a model for an intracellular acting bactericidal antibiotic. HeLa cells were transfected with an expression vector directing the heterologous expression of lysostaphin in the cytoplasm. Expression, subcellular localization and enzymatic activity of lysostaphin were investigated by immunoblotting, fluorescent microscopy and agar diffusion assays. Both transiently and stably transfected HeLa cells showed a strong expression of active lysostaphin. After infection with S. aureus, the intracellular number of S. aureus and the host cell viability were determined. This staphylolytic activity resulted in a strong reduction of intracellular S. aureus in a time- and dose-dependent manner. Furthermore, host cells expressing lysostaphin became protected from S. aureus-induced cell death. Our data demonstrate the potential of intracellularly acting cell-wall active drugs or antibiotics that kill S. aureus without causing harm to the infected host cells.     

Contractile responses to selective phosphodiesterase inhibitors following chronic β-adrenoreceptor activation

Contractile responses to phosphodiesterase (PDE) inhibitors are attenuated in heart failure, an effect limiting the clinical value of these agents. In this study, we sought to determine whether abnormalities in the β-adrenoreceptor (β-AR)–cyclic adenosine monophosphate (cAMP) signal transduction are sufficient to account for downregulation of PDE inhibitor-induced inotropic responses following chronic sympathetic activation. Sustained β-AR activation produced by administration of isoproterenol (ISO) (50 μg kg⁻¹ day⁻¹ i.p. for 1 month) to rats resulted in cardiac hypertrophy, but did not affect baseline cardiac systolic function, as assessed in vivo by echocardiography and ex vivo under controlled loading conditions and heart rate (left ventricular systolic pressure–volume and stress–strain relations). Moreover, chronic ISO administration did not alter the baseline myocardial norepinephrine release or inotropic responses to incremental concentrations of Ca²⁺ in isolated, perfused heart preparations. However, left ventricular contractile responses to ISO, the PDE III inhibitor amrinone, and the PDE IV inhibitor rolipram were attenuated following chronic β-AR activation. Myocardial cAMP concentrations after stimulation with amrinone and rolipram were similar in ISO-treated and control rats. However, in ISO-treated rats, a marked decrease in contractile responsiveness to the cell-permeable, PDE-resistant cAMP analogue, 8-bromoadenosine cAMP, was noted. In conclusion, these data suggest that in cardiac disease, sustained β-AR activation, without producing ventricular systolic dysfunction or enhanced myocardial norepinephrine release, is sufficient to account for the downregulation of contractile responses to PDE inhibitors. This effect appears to be largely mediated through abnormalities in signal transduction between cAMP and Ca²⁺-induced Ca²⁺ release.     

Dependence of cross-bridge kinetics on myosin light chain isoforms in rabbit and rat skeletal muscle fibres

Cross-bridge kinetics underlying stretch-induced force transients was studied in fibres with different myosin light chain (MLC) isoforms from skeletal muscles of rabbit and rat. The force transients were induced by stepwise stretches (< 0.3% of fibre length) applied on maximally Ca²⁺-activated skinned fibres. Fast fibre types IIB, IID (or IIX) and IIA and the slow fibre type I containing the myosin heavy chain isoforms MHC-IIb, MHC-IId (or MHC-IIx), MHC-IIa and MHC-I, respectively, were investigated. The MLC isoform content varied within fibre types. Fast fibre types contained the fast regulatory MLC isoform MLC2f and different proportions of the fast alkali MLC isoforms MLC1f and MLC3f. Type I fibres contained the slow regulatory MLC isoform MLC2s and the slow alkali MLC isoform MLC1s. Slow MLC isoforms were also present in several type IIA fibres. The kinetics of force transients differed by a factor of about 30 between fibre types (order from fastest to slowest kinetics: IIB > IID > IIA ≫ I). The kinetics of the force transients was not dependent on the relative content of MLC1f and MLC3f. Type IIA fibres containing fast and slow MLC isoforms were about 1.2 times slower than type IIA fibres containing only fast MLC isoforms. We conclude that while the cross-bridge kinetics is mainly determined by the MHC isoforms present, it is affected by fast and slow MLC isoforms but not by the relative content of MLC1f and MLC3f. Thus, the physiological role of fast and slow MLC isoforms in type IIA fibres is a fine-tuning of the cross-bridge kinetics.     

Monte Carlo study of photon fields from a flattening filter-free clinical accelerator

In conventional clinical linear accelerators, the flattening filter scatters and absorbs a large fraction of primary photons. Increasing the beam-on time, which also increases the out-of-field exposure to patients, compensates for the reduction in photon fluence. In recent years, intensity modulated radiation therapy has been introduced, yielding better dose distributions than conventional three-dimensional conformal therapy. The drawback of this method is the further increase in beam-on time. An accelerator with the flattening filter removed, which would increase photon fluence greatly, could deliver considerably higher dose rates. The objective of the present study is to investigate the dosimetric properties of 6 and 18 MV photon beams from an accelerator without a flattening filter. The dosimetric data were generated using the Monte Carlo programs BEAMnrc and DOSXYZnrc. The accelerator model was based on the Varian Clinac 2100 design. We compared depth doses, dose rates, lateral profiles, doses outside collimation, total and collimator scatter factors for an accelerator with and without a flatteneing filter. The study showed that removing the filter increased the dose rate on the central axis by a factor of 2.31 (6 MV) and 5.45 (18 MV) at a given target current. Because the flattening filter is a major source of head scatter photons, its removal from the beam line could reduce the out-of-field dose.