Musa spp. cultivars as a neutralising source against some toxic activities of Bothrops and Crotalus genus snake venoms.

Accidents involving snakes from Bothrops spp. and Crotalus spp. constitute the most important cause of envenomation in Brazil and Argentina. Musa spp. (banana) have been reported to be used in popular medicine against snakebite by the members of the Canudos Settlement, located in Goiás. In this way, the aim of this work was to evaluate the antivenom effect of the Ouro (AA), Prata (AAB), Prata-anã (AAB) and Figo (ABB) cultivars against in vitro (phospholipase, coagulation and proteolytic) and in vivo (lethality and toxicity) activities caused by the venoms and toxicity (Artemia salina nauplii and Danio rerio embryos) of Musa spp. as well as the annotation of chemical compounds possibly related to these activities. From the in vitro antiophidic tests with the sap, we observed 100% inhibition of the phospholipase and coagulant activities with the cultivars Prata-anã and Figo against the venoms of B. alternatus and C. d. collineatus, B. diporus and B. pauloensis, respectively, and neutralisation of the lethality against the B. diporus venom. It was observed that the cultivars of Musa spp. did not show toxicity against Artemia salina nauplii and Danio rerio embryos. The sap analysis via HPLC-MS/MS allowed the annotation of the 13 compounds: abscisic acid, shikimic acid, citric acid, quinic acid, afzelechin, Glp-hexose, glucose, sucrose, isorhamnetin-3-O-galactoside-6-raminoside, kaempferol-3-glucoside-3-raminoside, myricetin-3-O-rutinoside, procyanidin B1 and rutin. Therefore, it can be seen that Musa spp. is a potential therapeutic agent that can act to neutralise the effects caused by snakebites.

Ovarian Real-World International Consortium (ORWIC): A multicentre, real-world analysis of epithelial ovarian cancer treatment and outcomes.

Introduction: Much drug development and published analysis for epithelial ovarian cancer (EOC) focuses on early-line treatment. Full sequences of treatment from diagnosis to death and the impact of later lines of therapy are rarely studied. We describe the establishment of an international network of cancer centers configured to compare real-world treatment pathways in UK, Portugal, Germany, South Korea, France and Romania (the Ovarian Real-World International Consortium; ORWIC). Methods: 3344 patients diagnosed with EOC (2012-2018) were analysed using a common data model and hub and spoke programming approach applied to existing electronic medical records. Consistent definition of line of therapy between sites and an efficient approach to analysis within the limitations of local information governance was achieved. Results: Median age of participants was 53-67 years old and 5-29% were ECOG >1. Between 62% and 84% of patients were diagnosed with late-stage disease (FIGO III-IV). Sites treating younger and fitter patients had higher rates of debulking surgery for those diagnosed at late stage than sites with older, more frail patients. At least 21% of patients treated with systemic anti-cancer therapy (SACT) had recurrent disease following second-line therapy (2L); up to 11 lines of SACT treatment were recorded for some patients. Platinum-based SACT was consistently used across sites at 1L, but choices at 2L varied, with hormone therapies commonly used in the UK and Portugal. The use (and type) of maintenance therapy following 1L also varied. Beyond 2L, there was little consensus between sites on treatment choice: trial compounds and unspecified combinations of other agents were common. Discussion: Specific treatment sequences are reported up to 4L and the establishment of this network facilitates future analysis of comparative outcomes per line of treatment with the aim of optimizing available options for patients with recurrent EOC. In particular, this real-world network can be used to assess the growing use of PARP inhibitors. The real-world optimization of advanced line treatment will be especially important for patients not usually eligible for involvement with clinical trials. The resources to enable this analysis to be implemented elsewhere are supplied and the network will seek to grow in coverage of further sites.

Comparative evaluation of alternative batteries of genetic markers to complement autosomal STRs in kinship investigations: autosomal indels vs. X-chromosome STRs.

Kinship investigations such as paternity are currently solved using sets of (commercially available) highly polymorphic autosomal short tandem repeats (STRs), which lead to powerful likelihood ratios (LR). Still, some difficult cases arise whenever the kinship is much more remote or if the alternative hypotheses are not correctly formulated due to the lack of information (for e.g. there is an unknown relationship between the alleged and the true fathers). In these situations, beyond the routinely used marker set, laboratories usually enlarge the number and/or the type of markers analysed. Among these, autosomal indels and X-chromosome STRs have gained popularity. The aim of this study was to compare the results obtained after complementing an initial set of autosomal STRs with indels or with X-chromosome-specific STRs in simulated paternity cases where the alleged father is a close relative of the real one. Results show that in paternity cases where a low number of incompatibilities are observed, the best strategy is to increase the number of autosomal STRs under analysis. Nevertheless, if these are not available, our study globally shows that in father-daughter duos, a set of 12 X-STRs is more advantageous than 38 highly diverse autosomal biallelic markers. Additionally, the usefulness of X-STRs was also evaluated in cases where only a close relative of the alleged parent (father or mother) is available for testing. For those situations where these markers have the power to exclude, strong LR values are obtained. In the remaining cases, LRs are usually weak and sometimes the results are more likely under the wrong kinship hypothesis.

Cytotoxic activity in basal and tumoral cell lines of the C0K3N3 protein from the snake venom Crotalus durissus collilineatus, variety crotamine negative.

Snake venoms are natural sources of bioactive substances with therapeutic potential. In this work, we evaluated the cytotoxicity of the Crotalus durissus collilineatus, negative crotamine variety and the isolated fraction C0K3N3 in BALB C/3T3 and K562 cell lines. The results indicate that the C0K3N3 protein is more cytotoxic against the K562 tumor cell line than in the 3T3 baseline.

Benzoic Acid Derivatives as Prodrugs for the Treatment of Tuberculosis.

One interesting approach to fight tuberculosis is the use of prodrugs that often have shown improved biological activities over drugs with poor absorption or difficulty to cross membranes. Previous studies demonstrate that weak acids such as benzoic acid, present antimycobacterial activity. Moreover, esters of those acids revealed to be a viable alternative since they may diffuse more easily through the cell membranes. Previously we showed that mycobacteria can easily activate benzoic acid esters by conversion to the corresponding acid. Since Zhang postulated that the activity of the acids can be dependent on their pKa, we set up to synthesize a library of benzoates with different electron withdrawing groups (4-chloro, 2,6-dichloro, 3,5-dichloro, 4-nitro, and 3,5 dinitro), to modulate pKa of the liberated acid and different alkoxy substituents (propyl, hexyl, and phenyl) to modulate their lipophilicity, and tested the activity of the esters and the corresponding free acids against mycobacteria. We also studied the activation of the esters by mycobacterial enzymes and the stability of the compounds in buffer and plasma. We concluded that all the benzoates in our study can be activated by mycobacterial enzymes and that the phenyl and hexyl esters presented higher activity than the corresponding free acids, with the nitrobenzoates, and especially the dinitrobenzoates, showing very interesting antitubercular activity that deserve further exploration. Our results did not show a correlation between the activity and the pKa of the acids.

Acute toxicity, antinociceptive, and anti-inflammatory activities of the orally administered crotamine in mice.

Crotamine is a polypeptide toxin isolated from rattlesnake venom. Although several studies have been developed identifying many biological effects of isolated crotamine, none of them evaluated its acute toxicity, antinociceptive, and anti-inflammatory activities through oral administration. All in vivo experiments from this study were performed in mice. The up-and-down procedure and hippocratic screening were carried out to evaluate possible pharmacological and toxic effects. Antinociceptive and anti-inflammatory activities of this toxin were evaluated using acetic acid-induced abdominal writhing, formalin-induced pain assays, croton oil-induced ear edema, and carrageenan-induced pleurisy. Crotamine did not cause lethality or signs of intoxication up to the maximum dose tested (10.88 mg/kg). The number of contortions was reduced significantly by 34, 57, and 74% at the oral doses of 0.08, 0.16, and 0.32 mg/kg, respectively. At the dose of 0.16 mg/kg, crotamine decreases pain time-reactivity at neurogenic phase by 45% and at inflammatory phase by 60%. Also, crotamine elicited antiedematogenic activity through the attenuation of the croton oil-induced ear edema by 77%. In the carrageenan-induced pleurisy, the leukocyte, neutrophil, and mononuclear cell migration to the lesion site were reduced by 52%, 46%, and 59%, respectively. Altogether, crotamine demonstrated in vivo antinociceptive and anti-inflammatory effect through acute oral administration, generating an anti-migratory mechanism of action at non-toxic doses.

 Health-related Quality of Life and Depression in Older Patients With Pressure Ulcers.

UNLABELLED:  Objectives. To evaluate health-related quality of life (HRQoL) and depression in older patients with pressure ulcers who were living at home in the community. METHODS: A cross-sectional analytical study conducted in southern Minas Gerais, Brazil. Forty-two outpatients 60 years and older, showing no cognitive deficit who were living in the community, participated in the study. They were divided into two groups: the study group (21 patients with pressure ulcers), and control group (21 patients without pressure ulcers). The instruments Mini-Mental State Examination (MMSE), Medical Outcomes Study 36-Item Short Form Health Survey questionnaire (SF-36), and 15-item Geriatric Depression Scale (GDS-15) were used, respectively, to identify cognitive impairment, and assess HRQoL and depression. RESULTS: Among the 36 pressure ulcers detected on examination, Stage II ulcers were the most common finding (50%), and the sacral region was the most common location (44.5%). Patients with pressure ulcers had significantly lower HRQoL scores than controls in all SF-36 domains, with the lowest scores being reported for physical functioning, role physical and role emotional (P <0.0001). Seventeen (80.9%) of the patients in the study group were identified as having depression (P = 0.002). CONCLUSION: A high rate of depression (GDS-15) was found in elderly patients with pressure ulcers, who also reported lower HRQoL scores in all SF-36 domains compared with controls.

A hyaluronidase from Potamotrygon motoro (freshwater stingrays) venom: isolation and characterization.

Freshwater stingrays (Potamotrygon motoro) are known to cause human accidents through a sting located in its tail. In the State of Goiás, this accident happens especially during the fishing season of the Araguaia River. The P. motoro venom extracted from the sting presented hyaluronidase activity. The enzyme was purified by gel filtration on Sephacryl S-100 and ion-exchange chromatography on SP-Sepharose. A typical procedure provided 376.4-fold purification with a 2.94% yield. The molecular weight of the purified enzyme was 79 kDa as estimated by gel filtration on Sephacryl S-100. The K(m) and V(max) values for hyaluronidase, using hyaluronic acid as substrate, were 4.91 microg/ml and 2.02 U/min, respectively. The pH optimum for the enzyme was pH 4.2 and maximum activity was obtained at 40 degrees C. The hyaluronidase from P. motoro was shown to be heat instable, being stabilized by bovine albumin and DTT, and inhibited by Fe(2+), Mn(2+), Cu(2+) and heparin.

Risk Factors for Pressure Ulcers in Hospitalized Elderly without Significant Cognitive Impairment.

PURPOSE: To evaluate risk factors for pressure ulcers (PU) in hospitalized elderly without significant cognitive impairment. METHODS: From July 2005 to February 2006, 40 hospitalized elderly patients without cognitive deficit were evaluated in a university hospital in Pouso Alegre, Brazil. Twenty patients with a PU formed the study group and 20 without a PU formed the control group. The Mini Mental State Examination (MMSE) was used to assess cognitive status. Pressure ulcers were classified using the National Pressure Ulcer Advisory Panel (NPUAP) guidelines, followed by evaluation of risk factors for PU using the Braden scale. The chi-squared test was applied and for the Braden scale the Mann-Whitney test was used. RESULTS: In the study group, 14 (70%) of the subjects were women and 6 (30%) were men. The average age was 71.5 years. The average score for the MMSE was 19.7. The average time of hospitalization was 23.1 days for the study group and 13 days for the control. In the Braden scale, the risk factors such as humidity, activity, mobility, friction, and shear force were significant (P < 0.05). CONCLUSION: The data from the present study demonstrate that hospitalized elderly patients have an increased risk for the development of PU. Humidity, activity, mobility, friction, and shear are important risk factors during the hospitalization period.

Effects of Crotalus durissus collilineatus venom in the isolated rat kidney.

Ophidian accidents caused by the subspecies Crotalus durissus are responsible for high morbity and mortality rates. Acute renal failure is a common complication observed in these accidents. The aim of the present study was to investigate the renal effects promoted by the venom of C. d. collilineatus and its fractions, crotoxin and phospholipase A2. C. d. collilineatus (Cdc; 30 microg mL(-1)), crotoxin (CTX; 10 microg mL(-1)) and phospholipase A2 (PLA2; 10 microg mL(-1)) were tested in isolated rat kidney. The first 30 min of each experiment were used as an internal control and Cdc or its fractions, CTX and PLA2 were added to the system after this period. All experiments lasted 120 min. The venom of Cdc decreased perfusion pressure (PP; control120 = 110.3 +/- 3.69 mmHg; Cdc120 = 96.7+/-8.1 mmHg), renal vascular resistance (RVR; control120 = 6.42+/-0.78 mmHg mL g(-1) min(-1); Cdc120 = 4.8+/-0.56 mmHg/mL g(-1) min(-1)), urinary flow (UF; control120 = 0.19+/-0.03 mL g(-1) min(-1); Cdc120 = 0.12 +/- 0.01 mL g(-1) min(-1)), and glomerular filtration rate (GFR; control120 = 0.79 +/- 0.07 mL g(-1) min(-1); Cdc120 = 0.53 +/- 0.09 mL g(-1) min(-1)), but had no effect on the percent of sodium tubular transport (%TNa+), percent of chloride tubular transport (%TK+) and percent of potassium tubular transport (%TCl-). CTX and PLA2 reduced the GFR, while UF, PP and RVR remained stable during the full 120 min of perfusion. Crotoxin administration also diminished the %TK+ (control120 = 69.94 +/- 6.49; CTX120 = 33.28 +/- 4.78) and %TCl- (control120 = 79.53 +/- 2.67; CTX120 = 64.62 +/- 6.93). PLA2 reduced the %TK+, but exerted no effect on the %TNa+ or on that of TCl-. In conclusion, the C. d. collilineatus venom altered the renal functional parameters evaluated. We suggest that crotoxin and phospholipase A2 were involved in this process, since the renal effects observed would be due to the synergistic action of the components of the venom.

Identification of antinociceptive fraction of snake venom from Crotalus durissus collilineatus crotamine-negative and its acute toxicity evaluation.

The crude venom of the snake Crotalus durissus collilineatus (CDC) promotes neurological signs and symptoms in accidents involving humans and animals and the victims reports analgesia at the bite site, without tissue destruction. Studies shows that CDC has analgesic activity, among others. The crude venom is considered unsuitable for therapeutic purposes, with encouragement to the fractionation and purification of the same. Thus, the aim with CDC venom is: to perform fractionation by preparative HPLC; to test the antinociceptive activity of fractions and acute toxicity of active fractions. The CDC was fractionated on preparative HPLC-PDA (Oliveira et al., 2015) and the fractions were tested for their antinociceptive activity for writhing test by acetic acid (0.6%) in mice. For one of the fractions, which showed high analgesic effect both p.o. and i.p. routes, it evaluated the acute toxicity by the up and down method (OECD, 2001). In the fractionation by HPLC-PDA, CDC yielded 10 peaks (P1P10). SDS-PAGE showed that there was a good separation of components of the venom. All peaks were evaluated for their ability to reduce writhing, and the only one that apparently showed antinociceptive effect was Fr5 fraction (40 µg/kg). The Fr5 was able to reduce by 47% the number of contortions (i.p.) and 87% (p.o.), compared to control. The Fr5 fraction showed no morbidity and no mortality in the acute toxicity test (dose of 1000 µg/kg, p.o.); so it was not possible to estimate the LD50. According to the results, it can be stated that the venom and Fr5 of Crotalus durissus collilineatus snake of crotamine-negative type, may exhibit antinociceptive activity by suppressing nociception induced by acetic acid, suggesting it is related to effects on peripheral sites spinal and presents low acute toxicity values in experimental animals.

Reevaluating the role of 1,10-phenanthroline in oxidative reactions involving ferrous ions and DNA damage.

It is widely believed that the iron chelator 1,10-phenanthroline (phen) is able to fully block the Fenton reaction by forming a complex (Fe(phen)3(2+), also known as ferroin) that cannot react with H2O2. We observed that phen cannot fully prevent 2-deoxyribose (5 mM) degradation induced by Fenton reagents (30 microM Fe(II) plus 100-500 microM H2O2); protection varied from 55% to 66% when the phen/Fe(II) ratio was 3:1 to 20:1. Inhibition of 2-deoxyribose damage was nearly unchanged if phen was pre-incubated with Fe(II). Moreover, preformed Fe(phen)3(2+) complex added to the solution containing H2O2 was able to induce 2-deoxyribose degradation and methane sulfinic acid formation from the oxidation of 5% DMSO. The partially protective effect of phen was unchanged with the use of either phosphate or HEPES as buffers (5 mM, pH 7.2), or in unbuffered media (pH 5.1). Both DMSO oxidation and 2-deoxyribose degradation correlated with the increase in Fe(phen)3(2+) concentration. Strand breaks in plasmid pTARGETtrade mark DNA induced by Fenton reagents (1 microM Fe(II) plus 25 microM H2O2) in HEPES buffer could only be partially prevented by phen, even when the chelator was 16 times more concentrated than Fe(II). In these experiments, Fe(phen)3(2+) and DNA were pre-incubated from 1 to 10 min before addition of H2O2. Moreover, a high level of DNA strand breakage was observed when iron and phen are added to the reaction immediately before H2O2. On the other hand, phen fully prevented 2-deoxyribose degradation induced by the autoxidation of 30 microM Fe(II) in phosphate-buffered (3 to 30 mM) media. Our data provide evidence that the Fe(phen)3(2+) complex induces in vitro oxidative damage in the presence of H2O2 (possibly by means of Fe(phen)3(2+) dissociation into Fe(phen)2(2+)), but they show that the complex cannot undergo autoxidation.

Identification of crotamine in the venom of Crotalus durissus collilineatus by three different methods.

The use of mice for the identification of crotamine has been the motive of discussions of bioethical character and technical (efficiency), so that a reassessment of the use of animals in experiments is global trend the search for alternative tests. The objective of this study was to standardize a method for HPLC-PDA to identify the presence of crotamine in the venom of rattlesnakes, aiming to propose an alternative methodology to reduce or replace the use of animals. The Cdc was evaluated as to the presence of crotamine by 3 methods: traditional test lethality in mice (Mus musculus) swiss albino male, 18-22 g (i.p.), polyacrylamide gel electrophoresis (SDS-PAGE) and HPLC-PDA. The venoms of 50 specimens of Crotalus durissus collilineatus, held in the serpentarium CEPB/PUC Goiás, were obtained by manual massage of the gland, making the collection individually. To identify the band corresponding to crotamine, the venoms of specimens, analysis was performed on SDS-PAGE and references. Procedure in mice with 20% of the samples tested positive for crotamine, 24% negative and 56% uncertain outcome. With the SDS-PAGE was identified crotamine in 26% of samples, 26% negative and 48% continued with uncertain outcome. By HPLC method showed the presence myotoxin in 86% of samples, with 14% negative. The tests conducted in this study indicated that methodology which utilizes animals for identifying the presence of crotamine the venom of C. durissus can safely be replaced by the test SDS-PAGE and HPLC, since the methods are reproducible, and do not undergo any interference biological animal and mainly contribute to reducing the number of animals used for laboratory tests.

[Financial incentives for employees as part of a policy to upgrade care in a public hospital in Volta Redonda, Rio de Janeiro State, Brazil]. / O pagamento de incentivo financeiro para os funcionários como parte da política de qualificação da assistência de um hospital público, Volta Redonda, Rio de Janeiro, Brasil.

The authors present and analyze six years of regular and steady application of an institutional evaluation policy based on financial incentives in a public hospital in Volta Redonda, Rio de Janeiro State, Brazil, as a part of the hospital's administrative modernization policy. This type of policy is considered implicitly capable of developing a sequence of strategic wagers: (1) release of financial resources for payment of bonuses; (2) creation of an operationally feasible and sensitive evaluation instrument; (3) creation of adequate management mechanisms to improve evaluation policy; (4) employee adherence to the hospital upgrading policy based on the bonus system; and (5) maintenance of the effects of evaluation policy over time. The article discusses the "degree of success" of each of these wagers in an attempt to portray possible gains throughout the process, while also identifying inherent difficulties in such a policy.

Antibacterial activity of the skin secretion of Phyllomedusa azurea (Anura: Hylidae) from the Central Brazil Cerrado / Actividad antibacterial de secreciones de la piel de Phyllomedusa azurea (Anura: Hylidae) en Cerrado, Brasil centro

Abstract Phyllomedusa azurea is a frog species well distributed geographically in South America, including Brazilian biomes as Pantanal and Cerrado. Compared with other anurans from the Phyllomedusinae family, there are few reports on the bioactive potential of skin-derived molecules from this species. In this perspective, the aim of the present study was to evaluate the in vitro antibacterial activity of skin secretion of P. azurea by detection of Minimum Inhibitory Concentration (MIC) of the growth of bacterial indicator strains and to determine if occurs a changing in the bacterial cell envelope permeability. The MIC determination was carried out by the microdilution plate method. The absorbance was measured and analyzed statistically using the t-test to compare two groups (0.05 % of significance). The impact of the crude extract on cell envelope permeability of Staphylococcus aureus ATCC 25923 was conducted by the crystal violet assay, and the absorbance was measured spectrophotometry followed by the calculation of the crystal violet uptake percentage. The specific MIC for S. aureus ATCC 25923 and Escherichia coli ATCC 25922 was 31.25 µg/mL, while for Bacillus subtilis ATCC 6633 was 125 µg/mL and Pseudomonas aeruginosa ATCC 27853 was 250 µg/mL. The treatment with crescent concentrations of frog skin secretion increased the crystal violet uptake by S. aureus ATCC 25923 cells, suggesting an action on the cell plasma membrane. The results demonstrated that the skin secretion of P. azurea presents antibacterial activity and merit further investigations to characterize the bioactive molecules.(AU)

Resumen P. azurea es una especie de rana bien distribuida geográficamente en América del Sur, que incluye biomas brasileños como Pantanal y Cerrado. En comparación con otros anuros de Phyllomedusinae, existen pocos informes sobre el potencial bioactivo de las moléculas derivadas de la piel de esta especie. En esta perspectiva, el objetivo del presente estudio fue evaluar la actividad antibacteriana in vitro de la secreción de la piel de P. azurea mediante la detección de la Concentración Inhibitoria Mínima (CIM) del crecimiento de cepas indicadoras bacterianas y determinar si ocurre un cambio en la permeabilidad de la envoltura celular bacteriana. La determinación de MIC se llevó a cabo mediante el método de la placa de microdilución. La absorbancia se midió y se analizó estadísticamente mediante la prueba t para comparar dos grupos (0.05 de significancia). El impacto del extracto crudo sobre la permeabilidad de la envoltura celular de Staphylococcus aureus ATCC 25923 se realizó mediante el ensayo de cristal violeta, y se midió la absorbancia mediante espectrofotometría seguida del cálculo del porcentaje de absorción de violeta cristal. La CIM específica para S. aureus ATCC 25923 y Escherichia coli ATCC 25922 fue 31.25 μg / ml, mientras que para Bacillus subtilis ATCC 6633 de 125 μg / ml y Pseudomonas aeruginosa ATCC 27853 de 250 μg / ml. El tratamiento con concentraciones crecientes de secreción de piel de rana aumentó la absorción de violeta cristal por las células de S. aureus ATCC 25923, sugiriendo una acción sobre la membrana plasmática de la célula. Los resultados demostraron que la secreción de la piel de P. azurea presenta actividad antibacteriana y amerita más investigaciones para caracterizar las moléculas bioactivas.(AU)

Crotalus durissus collilineatus Venom Induces TNF- α and IL-10 Production in Human Peripheral Blood Mononuclear Cells.

Snake venom has been the subject of numerous studies in an attempt to find properties and biological effects that may be beneficial to man. In this study we evaluated in vitro the effects of Crotalus durissus terrificus (Cdt) and Crotalus durissus collilineatus (Cdc) venom in human peripheral blood mononuclear cells (PBMCs). At 24 h, a significant decrease of viable cells was observed in cells stimulated with the Cdc venom at 0.0005 mg/mL and 0.005 mg/mL compared to the negative control. At 48 h, a significant decrease of viable cells was observed only in cells stimulated with Cdc venom at 0.005 mg/mL. A significant increase of TNF- α and IL-10 was detected 48 hours after culture of PBMC with Cdc, but not with Cdt venom. The expression of CD69 and PD1 (programmed death-1), activation and regulatory cell markers, on CD8+ and CD8- T cells did not change in the presence of Cdt and Cdc venom. Our results suggest the presence of proinflammatory and anti-inflammatory components in the Cdc venom. Further analysis should be done to identify those Cdc venom components as it has been done for the Cdt venom by other authors, indicating that modulatory components are found in the venom of different species of Crotalus snakes.

Evaluation of effectiveness and outcome of PKU screening and management in the State of Sergipe, Brazil.

OBJECTIVES: Phenylketonuria (PKU) was the first inherited metabolic disease known to cause mental retardation for which a newborn screening program (NBS) was developed. The objective of this study was to evaluate the effectiveness of PKU NBS and the management of cases in the northeastern Brazilian state of Sergipe (SE). MATERIALS AND METHODS: We reviewed the phenylalanine concentrations in filter-paper collected from the heel (PKUneo) of 43,449 newborns; blood concentrations obtained by venipuncture in the subjects with abnormal PKUneo; the children's age at several phases of the program, the incidence of the disease from January 2007 to June 2008; and metabolic control of the patients. RESULTS: The coverage of NBS/SE was 78.93%. The children's age was 10 ± 7 days at PKUneo collection. Twelve children were recalled based on the PKUneo cutoff value at 28 ± 13 days. From these, the concentrations of phenylalanine collected by venipuncture were normal in five children. The incidence of hyperphenylalaninemia was 1/43,449, and of PKU was 1/8,690 (5 cases). One suspected subject died. Another death occurred in the cohort, in a confirmed PKU case. PKU treatment began within 51 ± 12 days of life. In the four patients under dietary phenylalanine restriction, metabolic control was often difficult. CONCLUSIONS: PKU NBS/SE has satisfactory coverage and adequate cutoff for recalling patients and diagnosis, but the onset of treatment is delayed, and follow-up metabolic control is frequently inadequate.

Assessing paternities with inconclusive STR results: The suitability of bi-allelic markers.

In paternity testing the genetic profiles of the individuals are used to compare the relative likelihoods of the alleged father and the child being related as father/offspring against, usually, being unrelated. In the great majority of the cases, analyses with the widely used sets of short tandem repeat markers (STRs) provide powerful statistical evidence favouring one of the alternative hypotheses. Nevertheless, there are situations where the final statistical result is ambiguous, mostly because the alleged father shows incompatible genotypes at a few loci along with a very high paternity index in the remaining systems. In these cases, the possibility that the alleged father is actually a close relative of the real one (son, father or brother) can reasonably be raised. In such cases, when the statistical evidence obtained is considered as insufficient, the common practice is to extend the set of analysed markers. In this context, many authors have suggested that bi-allelic markers, such as single nucleotide (SNP) or insertion/deletion (Indel) polymorphisms, are markers of choice, as they are incomparably less prone to mutation than STRs. In this work we address the soundness of this claim and the consequences of this strategy, analyzing the a priori odds both for (a) expected number of Mendelian incompatibilities, and (b) expected values for the final likelihood ratios. Moreover, one hundred real pairs of second degree relatives, typed for two sets of markers: 15 STRs plus 38 Indels, were used to simulate paternity testing. Our data show that, for the number of markers commonly considered, the results from an extended battery of SNPs or Indels should be interpreted with caution when relatives are possibly involved.

Instability of mRNA expression signatures of drug transporters in chronic myeloid leukemia patients resistant to imatinib.

Despite the success of imatinib mesylate (IM) in the treatment of chronic myeloid leukemia (CML), approximately 30% of patients are resistant to therapy, mostly due to unknown causes. To profile the expression signatures of drug transporters throughout IM therapy and correlate them with resistance, we quantified mRNA expression levels of the SLC22A12, ABCB1, ABCC1, ABCG2 and MVP genes in consecutive samples from peripheral blood or bone marrow of CML patients who were either responsive or resistant to IM. Additionally we identified and quantified BCR-ABL1 transcript variants and analyzed 1236T>C ABCB1 and 480G>C SLC22A1 polymorphisms. A relationship between the type of BCR-ABL1 transcript or ABCB1 or SLC22A1 genotype and response to treatment was not discovered. However, the studied genes had higher expression levels in follow-up compared to the diagnostic samples, demonstrating a possible induction in expression. IM-sensitive patients presented significantly higher values of SLC22A1 expression, suggesting higher drug influx. Most importantly, while responding patients demonstrated stable expression signatures in consecutive samples, there was considerable variation in IM-resistant patients, indicating that single point sampling expression signatures are not reliable in predicting clinical outcomes or prognostic features in these patients. Studies that assessed consecutive samples from CML patients in order to evaluate the variation in expression levels of transporter genes are limited yet our study emphasizes the importance of such approaches.

Development of imatinib and dasatinib resistance: dynamics of expression of drug transporters ABCB1, ABCC1, ABCG2, MVP, and SLC22A1.

About 20% of patients with chronic myeloid leukemia (CML) do not respond to treatment with imatinib either initially or because of acquired resistance. To study the development of CML drug resistance, an in vitro experimental system comprising cell lines with different resistance levels was established by exposing K562 cells to increasing concentrations of imatinib and dasatinib anticancer agents. The mRNA levels of BCR- ABL1 and of genes involved in drug transport or redistribution (ABCB1, ABCC1, ABCC3, ABCG2, MVP, and SLC22A1) were measured and the ABL1 kinase domain sequenced. Results excluded BCR- ABL1 overexpression and mutations as relevant resistance mechanisms. Most studied transporters were overexpressed in the majority of resistant cell lines. Their expression pattern was dynamic: varying with resistance level and chronic drug exposure. Studied efflux transporters may have an important role at the initial stages of resistance, but after prolonged exposure and for higher doses of drugs other mechanisms might take place.