Monocrotaline presence in the Crotalaria (Fabaceae) plant genus and its influence on arthropods in agroecosystems

Abstract Crotalaria (Fabaceae) occurs abundantly in tropical and subtropical regions and has about 600 known species. These plants are widely used in agriculture, mainly as cover plants and green manures, in addition to their use in the management of phytonematodes. A striking feature of these species is the production of pyrrolizidine alkaloids (PAs), secondary allelochemicals involved in plant defense against herbivores. In Crotalaria species, monocrotaline is the predominant PA, which has many biological activities reported, including cytotoxicity, tumorigenicity, hepatotoxicity and neurotoxicity, with a wide range of ecological interactions. Thus, studies have sought to elucidate the effects of this compound to promote an increase in flora and fauna (mainly insects and nematodes) associated with agroecosystems, favoring the natural biological control. This review summarizes information about the monocrotaline, showing such effects in these environments, both above and below ground, and their potential use in pest management programs.

Resumo Crotalaria (Linnaeus, 1753) (Fabaceae) ocorre abundantemente em regiões tropicais e subtropicais e tem cerca de 600 espécies conhecidas. Estas plantas são amplamente utilizadas na agricultura, principalmente como cobertura e adubos verdes, além da sua utilização no manejo de fitonematoides. Uma característica marcante destas espécies é a produção de alcalóides pirrolizidinicos (APs), aleloquímicos secundários envolvidos na defesa das plantas contra os herbívoros. Nas espécies de Crotalaria, a monocrotalina é a AP predominante, que tem muitas atividades biológicas relatadas, incluindo citotoxicidade, tumorigenicidade, hepatotoxicidade e neurotoxicidade, além de uma vasta gama de interações ecológicas. Assim, estudos têm procurado elucidar os efeitos desse composto para promover um incremento na flora e fauna (principalmente insetos e nematoides) associados aos agroecossistemas, favorecendo o controle biológico natural. Esta revisão compila informações sobre a monocrotalina, mostrando tais efeitos nesses ambientes, tanto acima como abaixo do solo e a sua potencial utilização em programas de manejo de pragas.

Monocrotaline presence in the Crotalaria (Fabaceae) plant genus and its influence on arthropods in agroecosystems / Presença de monocrotalina em plantas do gênero Crotalaria (Fabaceae) e a sua influência sobre artrópodes nos agroecossistemas

Abstract Crotalaria (Fabaceae) occurs abundantly in tropical and subtropical regions and has about 600 known species. These plants are widely used in agriculture, mainly as cover plants and green manures, in addition to their use in the management of phytonematodes. A striking feature of these species is the production of pyrrolizidine alkaloids (PAs), secondary allelochemicals involved in plant defense against herbivores. In Crotalaria species, monocrotaline is the predominant PA, which has many biological activities reported, including cytotoxicity, tumorigenicity, hepatotoxicity and neurotoxicity, with a wide range of ecological interactions. Thus, studies have sought to elucidate the effects of this compound to promote an increase in flora and fauna (mainly insects and nematodes) associated with agroecosystems, favoring the natural biological control. This review summarizes information about the monocrotaline, showing such effects in these environments, both above and below ground, and their potential use in pest management programs.

Resumo Crotalaria (Linnaeus, 1753) (Fabaceae) ocorre abundantemente em regiões tropicais e subtropicais e tem cerca de 600 espécies conhecidas. Estas plantas são amplamente utilizadas na agricultura, principalmente como cobertura e adubos verdes, além da sua utilização no manejo de fitonematoides. Uma característica marcante destas espécies é a produção de alcalóides pirrolizidinicos (APs), aleloquímicos secundários envolvidos na defesa das plantas contra os herbívoros. Nas espécies de Crotalaria, a monocrotalina é a AP predominante, que tem muitas atividades biológicas relatadas, incluindo citotoxicidade, tumorigenicidade, hepatotoxicidade e neurotoxicidade, além de uma vasta gama de interações ecológicas. Assim, estudos têm procurado elucidar os efeitos desse composto para promover um incremento na flora e fauna (principalmente insetos e nematoides) associados aos agroecossistemas, favorecendo o controle biológico natural. Esta revisão compila informações sobre a monocrotalina, mostrando tais efeitos nesses ambientes, tanto acima como abaixo do solo e a sua potencial utilização em programas de manejo de pragas.

Monocrotaline presence in the Crotalaria (Fabaceae) plant genus and its influence on arthropods in agroecosystems.

Crotalaria (Fabaceae) occurs abundantly in tropical and subtropical regions and has about 600 known species. These plants are widely used in agriculture, mainly as cover plants and green manures, in addition to their use in the management of phytonematodes. A striking feature of these species is the production of pyrrolizidine alkaloids (PAs), secondary allelochemicals involved in plant defense against herbivores. In Crotalaria species, monocrotaline is the predominant PA, which has many biological activities reported, including cytotoxicity, tumorigenicity, hepatotoxicity and neurotoxicity, with a wide range of ecological interactions. Thus, studies have sought to elucidate the effects of this compound to promote an increase in flora and fauna (mainly insects and nematodes) associated with agroecosystems, favoring the natural biological control. This review summarizes information about the monocrotaline, showing such effects in these environments, both above and below ground, and their potential use in pest management programs.

CT volume of enhancement of disease (VED) can predict the early response to treatment and overall survival in patients with advanced HCC treated with sorafenib.

OBJECTIVES: To analyse the predictive value of the volume of enhancement of disease (VED), based on the CT arterial enhancement coefficient (ΔArt%), in the evaluation of the sorafenib response in patients with advanced hepatocellular carcinoma (HCC). METHODS: Patients with sorafenib-treated advanced HCC, who underwent a multiphase contrast-enhanced CT before (T0) and after 60-70 days of starting therapy (T1), were included. The same target lesions utilised for the response evaluation according to modified Response Evaluation Criteria in Solid Tumors criteria were retrospectively used for the ΔArt% calculation ([(HUarterial phase - HUunenhanced phase) / HUunenhanced phase] × 100). ΔArt% was weighted for the lesion volume to obtain the VED. We compared VEDT0 and VEDT1 values in patients with clinical benefit (CB) or progressive disease (PD). The impact of VED, ancillary imaging findings, and blood chemistries on survival probability was evaluated. RESULTS: Thirty-two patients (25 men, mean age 65.8 years) analysed between 2012 and 2016 were selected. At T1, 8 patients had CB and 24 had PD. VEDT0 was > 70% in 8/8 CB patients compared with 12/24 PD patients (p = 0.011). Patients with VEDT0 > 70% showed a significantly higher median survival than those with lower VEDT0 (451.5 days vs. 209.5 days, p = 0.032). Patients with VEDT0 > 70% and alpha-fetoproteinT0 ≤ 400 ng/ml had significantly longer survival than all other three combinations. In multivariate analysis, VEDT0 > 70% emerged as the only factor independently associated with survival (p = 0.037). CONCLUSION: In patients with advanced HCC treated with sorafenib, VED is a novel radiologic parameter obtained by contrast-enhanced CT, which could be helpful in selecting patients who are more likely to respond to sorafenib, and with a longer survival. KEY POINTS: • To achieve the best results of treatment with sorafenib in advanced HCC, a strict selection of patients is needed. • New radiologic parameters predictive of the response to sorafenib would be essential. • Volume of enhancement of disease (VED) is a novel radiologic parameter obtained by contrast-enhanced CT, which could be helpful in selecting patients who are more likely to respond to therapy, and with a longer survival.

Role of SLC22A1 polymorphic variants in drug disposition, therapeutic responses, and drug-drug interactions.

The SCL22A1 gene encodes the broad selectivity transporter hOCT1. hOCT1 is expressed in most epithelial barriers thereby contributing to drug pharmacokinetics. It is also expressed in different drug target cells, including immune system cells and others. Thus, this membrane protein might also contribute to drug pharmacodynamics. Up to 1000 hOCT1 polymorphisms have been identified so far, although only a small fraction of those have been mechanistically studied. A paradigm in the field of drug transporter pharmacogenetics is the impact of hOCT1 gene variability on metformin clinical parameters, affecting area under the concentration-time curve, Cmax and responsiveness. However, hOCT1 also mediates the translocation of a variety of drugs used as anticancer, antiviral, anti-inflammatory, antiemetic agents as well as drugs used in the treatment of neurological diseases among. This review focuses exclusively on those drugs for which some pharmacogenetic data are available, and aims at highlighting the need for further clinical research in this area.

Human organic cation transporter 1 (hOCT1) as a mediator of bendamustine uptake and cytotoxicity in chronic lymphocytic leukemia (CLL) cells.

Bendamustine is used in the treatment of chronic lymphocytic leukemia (CLL). Routes for bendamustine entry into target cells are unknown. This study aimed at identifying transporter proteins implicated in bendamustine uptake. Our results showed that hOCT1 is a bendamustine transporter, as bendamustine could cis-inhibit the uptake of a canonical hOCT1 substrate, with a Ki in the micromolar range, consistent with the EC50 values of the cytotoxicity triggered by this drug in HEK293 cells expressing hOCT1. hOCT1 polymorphic variants determining impaired bendamustine-transporter interaction, consistently reduced bendamustine cytotoxicity in HEK293 cells stably expressing them. Exome genotyping of the SLC22A1 gene, encoding hOCT1, was undertaken in a cohort of 241 CLL patients. Ex vivo cytotoxicity to bendamustine was measured in a subset of cases and shown to correlate with SLC22A1 polymorphic variants. In conclusion, hOCT1 is a suitable bendamustine transporter, thereby contributing to its cytotoxic effect depending upon the hOCT1 genetic variants expressed.

Nucleoside transporters and human organic cation transporter 1 determine the cellular handling of DNA-methyltransferase inhibitors.

BACKGROUND AND PURPOSE: Inhibitors of DNA methyltransferases (DNMTs), such as azacytidine, decitabine and zebularine, are used for the epigenetic treatment of cancer. Their action may depend upon their translocation across the plasma membrane. The aim of this study was to identify transporter proteins contributing to DNMT inhibitor action. EXPERIMENTAL APPROACH: Drug interactions with selected hCNT and hENT proteins were studied in transiently transfected HeLa and MDCK cells. Interaction with human organic cation transporters (hOCTs) was assessed in transiently transfected HeLa cells and Xenopus laevis oocytes. KEY RESULTS: Zebularine uptake was mediated by hCNT1, hCNT3 and hENT2. Decitabine interacted with but was not translocated by any nucleoside transporter (NT) type. hCNT expression at the apical domain of MDCK cells promoted net vectorial flux of zebularine. Neither hOCT1 nor hOCT2 transported decitabine, but both were involved in the efflux of zebularine, suggesting these proteins act as efflux transporters. hOCT1 polymorphic variants, known to alter function, decreased zebularine efflux. CONCLUSIONS AND IMPLICATIONS: This study highlights the influence of human NTs and hOCTs on the pharmacokinetics and pharmacodynamics of selected DNMT inhibitors. As hOCTs may also behave as efflux transporters, they could contribute either to chemoresistance or to chemosensitivity, depending upon the nature of the drug or combination of drugs being used in cancer therapy.

Hemodynamic performance of stented and stentless aortic bioprostheses.

BACKGROUND: This study compares the hemodynamic performance of stented and stentless bioprostheses used for aortic valve replacement in patients with aortic stenosis and small aortic root. METHODS: Between 1995 and 1998, 37 patients with a 21-mm aortic annulus (group 1) underwent aortic valve replacement with either a 21-mm Edwards Perimount or a 23-mm St. Jude Toronto bioprosthesis whereas 47 patients with a 23-mm aortic annulus (group 2) received either a 23-mm Medtronic Mosaic or a 25-mm Edwards Prima bioprosthesis. In each group mean and peak gradients, effective orifice area index, and left ventricular mass index were compared during follow-up. RESULTS: Group 1 patients showed a significant reduction of mean (p < 0.001) and peak gradients (p = 0.001) during follow-up, more evident for St. Jude Toronto versus Edwards Perimount (p = 0.02 and p = 0.05, respectively). Group 2 patients showed a significant reduction of mean and peak gradients (p < 0.001), more evident for Edwards Prima versus Medtronic Mosaic (p < 0.001 and p = 0.07, respectively). Effective orifice area index significantly increased only in group 1 (p = 0.005). Left ventricular mass index significantly decreased in all patients regardless of the type of valve (p < 0.001). Patients with Edwards Prima showed a trend to a higher regression of left ventricular mass index versus Medtronic Mosaic recipients (p = 0.07). CONCLUSIONS: After aortic valve replacement, stented and stentless bioprostheses exhibited similar results with a more evident hemodynamic improvement during follow-up in the stentless valves. Stented bioprostheses of new generation, however, may parallel the hemodynamic performance of stentless valves and appear to be a valid alternative for aortic valve replacement in elderly patients with a small aortic annulus.

Dissection of atrial septum after mitral valve replacement.

We report a patient who presented with paraprosthetic leak complicated by dissection of the interatrial septum after mitral valve replacement. A review of the literature provides confirmation that only 3 cases have been previously reported of this potential, albeit extremely rare, complication of prosthetic mitral valve replacement. Prosthesis oversizing and improper mitral annular handling appeared to be the predisposing factors of this complication.

Hemodynamic assessment of the Medtronic Mosaic bioprosthesis in the aortic position.

BACKGROUND AND AIM OF THE STUDY: The Medtronic Mosaic bioprosthesis (MMB) is a newly developed tissue valve which incorporates several features such as a low-profile semi-flexible stent, zero-pressure tissue fixation, and anti-mineralization pretreatment with alpha-amino oleic acid aimed to improve hemodynamics and prevent structural deterioration. METHODS: Between November 1995 to June 1999, 62 patients underwent aortic valve replacement (AVR) with the MMB; 41 of these who reached the one-year follow up interval and who had isolated AVR without any concomitant procedure with size 23 mm and 25 mm MMB were evaluated by serial echocardiography after three, six and 12 months. RESULTS: For size 23 mm valves, mean and peak gradients were 12+/-3 and 21+/-6 mmHg at 3 months, 12+/-3 and 20+/-5 mmHg at 6 months, and 12+/-4 and 20+/-6 mmHg at 12 months. For size 25 mm valves, mean and peak gradients were 13+/-2 and 22+/-4 mmHg at 3 months, 12+/-3 and 21+/-5 mmHg at 6 months, and 12+/-4 and 22+/-6 mmHg at 12 months. In patients with 23 mm MMB, left ventricular mass index (LVMi) was reduced from 181+/-34 g/m2 before surgery to 158+/-32, 150+/-28 and 140+/-25 g/m2 at 3, 6 and 12 months after AVR (p <0.001); in patients with 25 mm MMB, LVMi was reduced from 182+/-28 g/m2 before surgery to 165+/-25, 156+/-24 and 146+/-19 g/m2 at 3, 6 and 12 months after AVR (p <0.001). CONCLUSION: Our results indicate that MMB is associated with low mean and peak gradients and significant reduction in LVMi throughout the postoperative period. Thus, the MMB appears to be an excellent cardiac valve substitute in elderly subjects who require AVR.

Performance of 21-mm size perimount aortic bioprosthesis in the elderly.

BACKGROUND: Aortic valve replacement in elderly patients with a small aortic annulus may pose difficult problems in terms of prosthesis selection. We have evaluated the hemodynamic performance of the 21-mm Carpentier-Edwards Perimount bioprosthesis implanted in elderly patients. METHODS: From July 1996 to June 1998, 19 patients (17 women and 2 men, mean age 76+/-4 years and mean body surface area 1.73+/-0.13 m2), had aortic valve replacement with a 21-mm Carpentier-Edwards Perimount bioprosthesis. The hemodynamic performance of the valve was evaluated in 16 patients, who completed at least a 6-month follow-up interval, with transthoracic color-Doppler echocardiography with particular reference to peak and mean transprosthetic gradients, effective orifice area index, and regression of left ventricular mass index. RESULTS: There were no late deaths and no major postoperative complications. At a mean follow-up of 12+/-7 months, compared to discharge, all patients showed clinical improvement with a significant reduction of peak gradient (from 23+/-4 to 21+/-6 mm Hg, p = 0.04) and left ventricular mass index (from 181+/-23 to 153+/-20 g/m2; p<0.001), whereas mean gradient (from 13+/-3 to 13+/-4 mm Hg, p = not significant) and effective orifice area index (from 1.12+/-0.34 to 1.13+/-0.28 cm2/m2, p = not significant) remained substantially unchanged. CONCLUSIONS: The use of a 21-mm Carpentier-Edwards Perimount bioprosthesis is associated with low transprosthetic gradients and significant reduction in left ventricular hypertrophy after aortic valve replacement. The results of our study suggest that a 21-m Carpentier-Edwards Perimount bioprosthesis should be considered a valid option in elderly patients with aortic valve disease and a small aortic annulus.

Prospective evaluation of frequency and nature of transcranial high-intensity Doppler signals in prosthetic valve recipients.

BACKGROUND AND AIM OF THE STUDY: In asymptomatic prosthetic valve recipients, high-intensity transient signals (HITS) observed with transcranial Doppler (TCD) are a phenomenon of obscure clinical relevance which nature has not yet been elucidated convincingly. METHODS: Eighty-three patients without carotid disease, history of cerebrovascular accidents, and with negative preoperative TCD undergoing either valve replacement (mitral, n = 11; aortic, n = 56; mitral + aortic, n = 6; 40 mechanical prostheses, 29 biological prostheses, 10 homografts) or mitral repair (n = 10) were evaluated prospectively by means of TCD at discharge, three months and one year after surgery, to analyze the presence, incidence and characteristics of HITS. Furthermore, in 12 patients positive for HITS, TCD was repeated during a 30-min period of 100% O2 inhalation. RESULTS: Twenty-five patients (30%) were positive for HITS at all postoperative controls, although no neurological symptoms were observed. Mechanical prostheses showed a significantly higher incidence of HITS (85%) than biological prostheses (10%, p <0.001), repaired mitral valves (0%, p <0.001) and homografts (0%, p <0.001). At multivariate analysis the presence of a mechanical prosthesis was the only significant predictor of detection of HITS after valve replacement. During O2 inhalation, a significant decrease in the number of HITS per hour (55 +/- 79 versus 22 +/- 31, p = 0.002) occurred, which returned to initial values when room-air breathing was resumed. CONCLUSIONS: Prosthetic valve replacement, particularly when mechanical devices are used, is associated with the generation of HITS which persist throughout the follow up period, but remain clinically silent. The decrease of HITS during O2 inhalation strongly supports the hypothesis of the gaseous nature of such signals and confirms the validity of this method in helping to differentiate gaseous microemboli from solid microemboli in prosthetic valve recipients.

Recurrent myxoma of the left ventricle. Case report and review of the literature.

A 29-year-old female was found to have a left ventricular mass while in the 14th week of gestation. Seven years earlier she had undergone removal of a left ventricular myxoma. At re-operation, after elective interruption of pregnancy, a recurrent left ventricular myxoma was successfully excised. According to a review of the literature recurrence of an isolated, localized left ventricular myxoma has not been previously reported.

Modulation of hepatitis C virus NS3 protease and helicase activities through the interaction with NS4A.

The hepatitis C virus nonstructural 3 protein (NS3) possesses a serine protease activity in the N-terminal one-third, whereas RNA-stimulated NTPase and helicase activities reside in the C-terminal portion. The serine protease activity is required for proteolytic processing at the NS3-NS4A, NS4A-NS4B, NS4B-NS5A, and NS5A-NS5B polyprotein cleavage sites. NS3 forms a complex with NS4A, a 54-residue polypeptide that was shown to act as an essential cofactor of the NS3 protease. We have expressed in Escherichia coli the NS3-NS4A precursor; cleavage at the junction between NS3 and NS4A occurs during expression in the bacteria cells, resulting in the formation of a soluble noncovalent complex with a sub-nanomolar dissociation constant. We have assessed the minimal ionic strength and detergent and glycerol concentrations required for maximal proteolytic activity and stability of the purified NS3-NS4A complex. Using a peptide substrate derived from the NS5A-NS5B junction, the catalytic efficiency (kcat/Km) of NS3-NS4A-associated protease under optimized conditions was 55 000 s-1 M-1, very similar to that measured with a recombinant complex purified from eukaryotic cells. Dissociation of the NS3-NS4A complex was found to be fully reversible. No helicase activity was exhibited by the purified NS3-NS4A complex, but NS3 was fully active as a helicase upon dissociation of NS4A. On the other hand, both basal and poly(U)-induced NTPase activity and ssRNA binding activity associated with the NS3-NS4A complex were very similar to those exhibited by NS3 alone. Therefore, NS4A appears to uncouple the ATPase/ssRNA binding and RNA unwinding activities associated with NS3.

Selection of functional variants of the NS3-NS4A protease of hepatitis C virus by using chimeric sindbis viruses.

The NS3-NS4A serine protease of hepatitis C virus (HCV) mediates four specific cleavages of the viral polyprotein and its activity is considered essential for the biogenesis of the HCV replication machinery. Despite extensive biochemical and structural characterization, the analysis of natural variants of this enzyme has been limited by the lack of an efficient replication system for HCV in cultured cells. We have recently described the generation of chimeric HCV-Sindbis viruses whose propagation depends on the NS3-NS4A catalytic activity. NS3-NS4A gene sequences were fused to the gene coding for the Sindbis virus structural polyprotein in such a way that processing of the chimeric polyprotein, nucleocapsid assembly, and production of infectious viruses required NS3-NS4A-mediated proteolysis (G. Filocamo, L. Pacini, and G. Migliaccio, J. Virol. 71:1417-1427, 1997). Here we report the use of these chimeric viruses to select and characterize active variants of the NS3-NS4A protease. Our original chimeric viruses displayed a temperature-sensitive phenotype and formed lysis plaques much smaller than those formed by wild-type (wt) Sindbis virus. By serially passaging these chimeric viruses on BHK cells, we have selected virus variants which formed lysis plaques larger than those produced by their progenitors and produced NS3-NS4A proteins different in size and/or sequence from those of the original viruses. Characterization of the selected protease variants revealed that all of the mutated proteases still efficiently processed the chimeric polyprotein in infected cells and also cleaved an HCV substrate in vitro. One of the selected proteases was expressed in a bacterial system and showed a catalytic efficiency comparable to that of the wt recombinant protease.

The Edwards Prima stentless valve: hemodynamic performance at one year.

BACKGROUND: The Edwards Prima stentless valve (EPSV) is a porcine aortic root cylinder with resected coronary ostia, fixed in glutaraldehyde at low pressure. The purpose of this study was to evaluate the hemodynamic performance of the EPSV 1 year after aortic valve replacement. METHODS: From December 1994 to February 1996, 29 patients underwent aortic valve replacement with EPSV used in the subcoronary position (group 1, n = 23) or as a root replacement (group 2, n = 6). Hemodynamic performance of EPSV was assessed by two-dimensional Doppler echocardiography at 1 week, 6 months, and 1 year by calculating peak transprosthetic velocity, peak and mean gradients, effective orifice area, degree of aortic regurgitation, and regression of left ventricular hypertrophy. RESULTS: There were no operative deaths. One patient in group 2 died after 3 months at reoperation for endocarditis. In group 1 early mean gradient (25+/-5 mm Hg for 23 mm and 19+/-5 mm Hg for 25 mm) decreased at 6 months and 1 year in the 23-mm size (17+/-7 mm Hg and 15+/-4 mm Hg, p < 0.01) and at 1 year in the 25-mm size (14+/-4 mm Hg, p = 0.03) without modifications of the effective orifice area in both sizes. A significant reduction in left ventricular hypertrophy occurred at 6 months and 1 year in both sizes. In group 2 lower early gradients were recorded with subsequent improvement at follow-up; reduction in left ventricular hypertrophy occurred as well. CONCLUSIONS: The EPSV used in the subcoronary position has shown high early gradients, which partially regressed at 6 months, with further improvement at 1 year. Gradients are attributed to inward folding of the Dacron cloth at the right coronary ostium, being more evident in patients with aortic stenosis without dilatation of the aortic root and coronary ostia close to the annulus. In such patients a better early hemodynamic result can be obtained by using the EPSV as a root replacement.

Multiple enzymatic activities associated with recombinant NS3 protein of hepatitis C virus.

The hepatitis C virus (HCV) nonstructural 3 protein (NS3) contains at least two domains associated with multiple enzymatic activities; a serine protease activity resides in the N-terminal one-third of the protein, whereas RNA helicase activity and RNA-stimulated nucleoside triphosphatase activity are associated with the C-terminal portion. To study the possible mutual influence of these enzymatic activities, a full-length NS3 polypeptide of 67 kDa was expressed as a nonfusion protein in Escherichia coli, purified to homogeneity, and shown to retain all three enzymatic activities. The protease activity of the full-length NS3 was strongly dependent on the activation by a synthetic peptide spanning the central hydrophobic core of the NS4A cofactor. Once complexed with the NS4A-derived peptide, the full-length NS3 protein and the isolated N-terminal protease domain cleaved synthetic peptide substrates with comparable efficiency. We show that, as in the case of the isolated protease domain, the protease activity of full-length NS3 undergoes inhibition by the N-terminal cleavage products of substrate peptides corresponding to the NS4A-NS4B and NS5A-NS5B. We have also characterized and quantified the NS3 ATPase, RNA helicase, and RNA-binding activities under optimized reaction conditions. Compared with the isolated N-terminal and C-terminal domains, recombinant full-length NS3 did not show significant differences in the three enzymatic activities analyzed in independent in vitro assays. We have further explored the possible interdependence of the NS3 N-terminal and C-terminal domains by analyzing the effect of polynucleotides on the modulation of all NS3 enzymatic functions. Our results demonstrated that the observed inhibition of the NS3 proteolytic activity by single-stranded RNA is mediated by direct interaction with the protease domain rather than with the helicase RNA-binding domain.

[Transmyocardial revascularization with a holmium laser: preliminary results]. / Rivascolarizzazione transmiocardica con laser ad olmio: risultati preliminari.

BACKGROUND: Transmyocardial laser revascularization (TMLR) aims to improve perfusion of the ventricular wall via laser-created transmural channels. We present the results of TMLR with a holmium laser as sole therapy in patients with angina refractory to medical treatment and extensive coronary artery disease unsuitable for angioplasty or coronary artery by-pass grafting. METHODS: From November 1995 to February 1997, twenty-two patients underwent isolated TMLR with a holmium laser. Five patients (23%) were female; the mean age was 67 +/- 7 years (range 53 to 74 years). Previous myocardial revascularization procedures had been performed in 17 patients (77%). Mean preoperative angina class was 3.4 +/- 0.5 and unstable angina was present in 7 patients (32%). RESULTS: There were no hospital deaths. The only postoperative complications were transient supraventricular arrhythmias in 6 patients (27%). Each patient received a mean of 33 +/- 8 channels in 27 +/- 13 minutes. There were two late deaths, 40 days and 4 months after TMLR, due to stroke and myocardial infarction, respectively. Mean follow-up duration was 8 +/- 5 months (range 40 days-15 months). The mean number of hospitalizations due to angina fell from 4.9 +/- 1.5 in the 6 months before TMLR to 1.5 +/- 1.0 in the 6 months following surgery (p < 0.001). At follow-up, mean angina class had significantly improved (1.8 +/- 0.6, p < 0.001), as well as effort tolerance, which increased from a mean of 3.5 +/- 1.4 minutes to 5.1 +/- 1.7 minutes (p = 0.01). 201Tl SPECT at 3 and 6 months did not show any significant changes in the segmental perfusion of the lased and unlased areas. CONCLUSIONS: TMLR with a holmium laser is a simple procedure with low operative mortality and morbidity. Short-term results confirm that clinical improvement is obtained in most patients, although this is not supported by significant changes in myocardial perfusion at short-term follow-up.

Displasia de tricúspide associada à anomalia de Ebstein em cäo: relato de caso / Tricuspid dysplasia associated to Ebstein's anomaly in dog: a case report

A displasia da tricúspide, associada ou näo à anomalia de Ebstein, näo é das cardiopatias mais freqüentes em cäes (Liu; Tilley6, 1976; Eyster et al.4, 1977; Netter9, 1978; Moise8, 1994). A displasia da tricúspide caracteriza-se pelo espessamento focal ou difuso dos folhetos valvulares, subdesenvolvimento das cordoalhas tendíneas e músculos papilares, separaçäo incompleta dos componentes valvares da parede ventricular, além da agenesia do tecido valvular. Simultaneamente ou näo a esta valvulopatia congênita, pode ocorrer a inserçäo mais baixa do aparelho valvar na parede do ventrículo direito, condiçäo esta conhecida como anomalia de Ebstein (Liu, Tilley6, 1976; Eyster et al.4, 1977; Moise8, 1994). No presente relato, os autores descrevem um caso de displasia da tricúspide associada à malformaçäo de Ebstein, em cäo Weimaraner, macho, de 3 meses de idade, cujo diagnóstico clínico foi estribado no exame físico, eletrocardiográfico, radiográfico e ecocardiográfico. O animal apresentou uma sobrevida de 15 meses após a realizaçäo do diagnóstico, vindo a óbito de forma súbita e natural, e a existência das alteraçöes congênitas foram confirmadas no exame necroscópico