Acute efficacy of contiguous versus temporally discontiguous point-by-point radiofrequency pulmonary vein isolation in patients with paroxysmal atrial fibrillation: a randomized study.

PURPOSE: Durable pulmonary vein (PV) isolation (PVI) determines the clinical success of catheter ablation for atrial fibrillation. In this randomized study, we investigated whether the temporally discontiguous deployment of ablation lesions adversely affected the acute efficacy of PVI. METHODS: Thirty-six consecutive patients with drug-refractory paroxysmal atrial fibrillation (aged 59 ± 11, 58% males) were randomized 1:1 to either discontiguous (D-PVI) or contiguous (C-PVI) encircling radiofrequency (RF) lesions around ipsilateral PVs. A contact force-sensing catheter was used targeting a final interlesion distance < 6 mm and the ablation index of 400-450 (anterior wall) and 300-350 (posterior wall). The study endpoint was defined as failure of first-pass PVI or acute PV reconnection during a waiting time (> 30 min) followed by adenosine challenge. RESULTS: The total RF time, number of RF lesions, and mean interlesion distance were comparable in both groups. Total endpoint rates were 1/36 (3%) in the D-PVI vs 4/36 (11%) in the C-PVI groups; P = 0.34 for superiority, P = 0.008 for non-inferiority. Adenosine-induced reconnection of right PVs was the only endpoint in the D-PVI group. In the C-PVI group, first-pass PVI failed in 2 right PVs and spontaneous reconnection occurred in 2 other circles (left and right PVs). CONCLUSION: Temporally discontiguous deployment of RF lesions is not associated with lower procedural PVI efficacy when strict criteria for interlesion distance and ablation index are applied. The development of local edema around each ablation site does not prevent effective RF lesion formation at adjacent positions. TRIAL REGISTRATION: clinicaltrials.gov (NCT03332862).

Corrosion Processes on Weathering Steel Bridges Influenced by Deposition of De-Icing Salts.

The safety and durability of bridges designed from weathering steels are conditioned by the development of a sufficiently protective layer of corrosion products. Air pollution, microclimate around the bridge, time of wetness, structural solution of the bridge, and the position and orientation of the surface within the bridge structure all influence the development of protective layers on the surface of the weathering steel. In this article, attention is focused mainly on the microclimatic effects resulting from the road traffic under the bridge. The influence of chloride deposition on the development of corrosion products is evaluated using experimental in situ testing. Two neighboring bridges made of weathering steel and crossing different types of obstacles were selected for this experiment. Relations and dependences between the measured parameters (deposition rate of chlorides, corrosion rates, thickness of corrosion products and the amount of chlorides in corrosion products) are evaluated and discussed.

Immunity to the Dual Threat of Silica Exposure and Mycobacterium tuberculosis.

Exposure to silica and the consequent development of silicosis are well-known health problems in countries with mining and other dust producing industries. Apart from its direct fibrotic effect on lung tissue, chronic and immunomodulatory character of silica causes susceptibility to tuberculosis (TB) leading to a significantly higher TB incidence in silica-exposed populations. The presence of silica particles in the lung and silicosis may facilitate initiation of tuberculous infection and progression to active TB, and exacerbate the course and outcome of TB, including prognosis and survival. However, the exact mechanisms of the involvement of silica in the pathological processes during mycobacterial infection are not yet fully understood. In this review, we focus on the host's immunological response to both silica and Mycobacterium tuberculosis, on agents of innate and adaptive immunity, and particularly on silica-induced immunological modifications in co-exposure that influence disease pathogenesis. We review what is known about the impact of silica and Mycobacterium tuberculosis or their co-exposure on the host's immune system, especially an impact that goes beyond an exclusive focus on macrophages as the first line of the defense. In both silicosis and TB, acquired immunity plays a major role in the restriction and/or elimination of pathogenic agents. Further research is needed to determine the effects of silica in adaptive immunity and in the pathogenesis of TB.

Cell cycle profiling by image and flow cytometry: The optimised protocol for the detection of replicational activity using 5-Bromo-2'-deoxyuridine, low concentration of hydrochloric acid and exonuclease III.

The approach for the detection of replicational activity in cells using 5-bromo-2'-deoxyuridine, a low concentration of hydrochloric acid and exonuclease III is presented in the study. The described method was optimised with the aim to provide a fast and robust tool for the detection of DNA synthesis with minimal impact on the cellular structures using image and flow cytometry. The approach is based on the introduction of breaks into the DNA by the low concentration of hydrochloric acid followed by the subsequent enzymatic extension of these breaks using exonuclease III. Our data showed that the method has only a minimal effect on the tested protein localisations and is applicable both for formaldehyde- and ethanol-fixed cells. The approach partially also preserves the fluorescence of the fluorescent proteins in the HeLa cells expressing Fluorescent Ubiquitin Cell Cycle Indicator. In the case of the short labelling pulses that disabled the use of 5-ethynyl-2'-deoxyuridine because of the low specific signal, the described method provided a bright signal enabling reliable recognition of replicating cells. The optimized protocol was also successfully tested for the detection of trifluridine, the nucleoside used as an antiviral drug and in combination with tipiracil also for the treatment of some types of cancer.

Trilobolide-steroid hybrids: Synthesis, cytotoxic and antimycobacterial activity.

Sesquiterpene lactone trilobolide is a sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) inhibitor, thus depleting the Ins(1,4,5)P3-sensitive intracellular calcium stores. Here, we describe a synthesis of a series of 6 trilobolide-steroids conjugates (estradiol, pregnene, dehydroepiandrosterone, and testosterone). We found that the newly synthesized Tb-based compounds possess different remarkable biological activities. Cancer cell cytotoxicity and preferential selectivity is represented in our study by a Tb-pregnene derivative. The most cytotoxic clickates of estradiol and pregnene were studied by FACS where impact on cell cycle and RNA synthesis was observed; live-cell microscopy revealed the impact on cell organelle morphology particularly endoplasmic reticulum, mitochondria and nucleus. Further, we have studied the estrogenic and androgenic properties of the clickate molecules using cell-based luciferase assays. Finally, antimycobacterial tests revealed that testosterone and estradiol derivatives potentiated the antimycobacterial activity up to IC50 of 10.6µM.

Pathophysiologically relevant in vitro tumor models for drug screening.

The alarming rate of failure of clinical trials is a major hurdle in cancer therapy that partly results from the inadequate use of in vitro tumor models for the screening of promising hits and leads in preclinical studies. 2D cultures of cancer cell lines that are primarily used for drug screening do not adequately recapitulate tumor microenvironment (TME) complexities compared with 3D cancer cell cultures and tumor-derived primary cell cultures. In this review, we focus on the potential use of in vitro tumor models that reproduce in vivo tumor complexities for effective drug selection in the preclinical stages of drug development.

Synthesis and biological activity evaluation of hydrazone derivatives based on a Tröger's base skeleton.

We report the design and synthesis of novel anticancer agents based on bis-hydrazones separated by a rigid Tröger's base skeleton. This novel approach combines a biologically active moiety (hydrazone) with this scaffold (Tröger's base) to construct DNA intercalators. Evaluation of the anticancer activity of these agents using seven cancer cell lines and two healthy cell lines found that several derivatives had potent anticancer activity and excellent selectivity indexes toward cancer cells. The antimicrobial activities were tested on a set of thirteen bacterial stains, but the prepared compounds were not active. Complexation studies using biologically important metal ions demonstrated that these compounds are able to bind Cu(2+), Fe(3+), Co(2+), Ni(2+) and Zn(2+). DNA intercalation studies showed that the compounds themselves do not interact with DNA, but their metallocomplexes do interact, most likely via intercalation into DNA.

C-MYC and C-FOS expression changes and cellular aspects of the photodynamic reaction with photosensitizers TMPyP and ClAlPcS2.

Photodynamic therapy (PDT) is based on the tumor-selective accumulation of photosensitizer followed by irradiation with light of an appropriate wavelength. After irradiation and in the presence of oxygen, photosensitizer induces cellular damage. The aim of this study was to evaluate effects of two photosensitizers TMPyP and ClAlPcS2 on cell lines to obtain better insight into their mechanisms of action. We determined cell viability, reactive oxygen species (ROS) generation and changes in expression levels of two important early response genes, C-MYC and C-FOS, on tumor MCF7 (human breast adenocarcinoma) and G361 (human melanoma) cell lines and non-tumor BJ cell line (human fibroblast) after photodynamic reaction with TMPyP and ClAlPcS2 as photosensitizers. In addition TMPyP and ClAlPcS2 cellular uptake and clearance and antioxidant capacity of the mentioned cell lines were investigated. We found appropriate therapeutic doses and confirmed that both tested photosensitizers are photodynamically efficient in treatment used cells in vitro. TMPyP is more efficient; it had higher ROS production and toxicity after irradiation by intermediate therapeutic doses than ClAlPcS2. We revealed that both TMPyP and ClAlPcS2-PDT increased C-FOS expression on tumor cell lines (G361 and MCF7), but not on non-tumor BJ cell line. Conversely, both TMPyP and ClAlPcS2-PDT decreased C-MYC expression on non-tumor BJ cell line but not on tumor cell lines. As first we tested these photosensitizers in such extent and we believe that it can help to better understand mechanisms of PDT and increase its efficiency and applicability.

Structural basis for inhibition of mycobacterial and human adenosine kinase by 7-substituted 7-(Het)aryl-7-deazaadenine ribonucleosides.

Adenosine kinase (ADK) from Mycobacterium tuberculosis (Mtb) was selected as a target for design of antimycobacterial nucleosides. Screening of 7-(het)aryl-7-deazaadenine ribonucleosides with Mtb and human (h) ADKs and testing with wild-type and drug-resistant Mtb strains identified specific inhibitors of Mtb ADK with micromolar antimycobacterial activity and low cytotoxicity. X-ray structures of complexes of Mtb and hADKs with 7-ethynyl-7-deazaadenosine showed differences in inhibitor interactions in the adenosine binding sites. 1D (1)H STD NMR experiments revealed that these inhibitors are readily accommodated into the ATP and adenosine binding sites of Mtb ADK, whereas they bind preferentially into the adenosine site of hADK. Occupation of the Mtb ADK ATP site with inhibitors and formation of catalytically less competent semiopen conformation of MtbADK after inhibitor binding in the adenosine site explain the lack of phosphorylation of 7-substituted-7-deazaadenosines. Semiempirical quantum mechanical analysis confirmed different affinity of nucleosides for the Mtb ADK adenosine and ATP sites.

Synthesis, cytostatic, antimicrobial, and anti-HCV activity of 6-substituted 7-(het)aryl-7-deazapurine ribonucleosides.

A series of 80 7-(het)aryl- and 7-ethynyl-7-deazapurine ribonucleosides bearing a methoxy, methylsulfanyl, methylamino, dimethylamino, methyl, or oxo group at position 6, or 2,6-disubstituted derivatives bearing a methyl or amino group at position 2, were prepared, and the biological activity of the compounds was studied and compared with that of the parent 7-(het)aryl-7-deazaadenosine series. Several of the compounds, in particular 6-substituted 7-deazapurine derivatives bearing a furyl or ethynyl group at position 7, were significantly cytotoxic at low nanomolar concentrations whereas most were much less potent or inactive. Promising activity was observed with some compounds against Mycobacterium bovis and also against hepatitis C virus in a replicon assay.

Combination of prednisolone and low dosed dexamethasone exhibits greater in vitro antileukemic activity than equiactive dose of prednisolone and overcomes prednisolone drug resistance in acute childhood lymphoblastic leukemia.

INTRODUCTION: Glucocorticoids, particularly prednisone/ prednisolone and dexamethasone, play a prominent role in the treatment of pediatric patients with acute lymphoblastic leukemia due to their ability to induce apoptosis in susceptible cells. Current therapeutic protocols use prednisone for both the prophase and the induction phase of the therapy because the greater antileukemic activity of dexamethasone is compromised by its high frequency of serious adverse reactions. AIM: To compare, for the first time, the in vitro antileukemic activity of prednisolone alone to that of a combination of prednisolone and dexamethasone using dexamethasone at a very low and presumably safe dosage (1/50 w/w). METHODS: Lymphoblasts were isolated from bone marrow and/or blood samples from children with newly diagnosed acute lymphoblastic leukemia. The cytotoxic activity of prednisolone, dexamethasone and the prednisolone/dexamethasone combination against isolated leukemia cells was analyzed using the MTT cytotoxicity assay. RESULTS: We observed differences in the in vitro antileukemic activity of prednisolone and dexamethasone in 21% of the tested patients. 3% of the children were prednisolone sensitive but dexamethasone resistant, while 18% were prednisolone resistant and dexamethasone sensitive. 32% were sensitive to both glucocorticoids and 18% were resistant to both. Cells from patients with good in vivo responses to prednisone monotherapy were more responsive to prednisolone in vitro than were cells from patients with poor prednisone responses (P<0.07). Importantly, we demonstrated that the use of even a minimal dose (1/50 w/w) of dexamethasone with prednisolone dramatically increases the in vitro anti-leukemic activity of prednisolone (P<0.0006). CONCLUSION: The high inter-individual variability of acute lymphoblastic leukemia responses to glucocorticoids suggest that either patients should be selected for prednisone or dexamethasone treatment on the basis of predictive biomarkers or that prednisone should be used directly in combination with a very low and safe dose of dexamethasone to potentiate its antileukemic activity. The latter option is likely to be cheaper and more efficient, and therefore warrants further clinical investigation to assess its efficacy and safety in treating childhood acute lymphoblastic leukemia.

Synthesis and significant cytostatic activity of 7-hetaryl-7-deazaadenosines.

A series of 7-aryl- and 7-hetaryl-7-deazaadenosines was prepared by the cross-coupling reactions of unprotected or protected 7-iodo-7-deazaadenosines with (het)arylboronic acids, stannanes, or zinc halides. Nucleosides bearing 5-membered heterocycles at the position 7 exerted potent in vitro antiproliferative effects against a broad panel of hematological and solid tumor cell lines. Cell cycle analysis indicated profound inhibition of RNA synthesis and induction of apoptosis in treated cells. Intracellular conversion to triphosphates has been detected with active compounds. The triphosphate metabolites showed only a weak inhibitory effect on human RNA polymerase II, suggesting potentially other mechanisms for the inhibition of RNA synthesis and quick onset of apoptosis. Initial in vivo evaluation demonstrated an effect of 7-(2-thienyl)-7-deazaadenine ribonucleoside on the survival rate in syngeneic P388D1 mouse leukemia model.