Ligation of multiple protein domains using orthogonal inteins with non-native splice junctions.

Protein splicing is a self-catalyzed process in which an internal protein domain (the intein) is excised from its flanking sequences, linking them together with a canonical peptide bond. Trans-inteins are separated in two different precursor polypeptide chains that must assemble to catalytically self-excise and ligate the corresponding flanking exteins to join even when expressed separately either in vitro or in vivo. They are very interesting to construct full proteins from separate domains because their common small size favors chemical synthesis approaches. Therefore, trans-inteins have multiple applications such as protein modification and purification, structural characterization of protein domains or production of intein-based biosensors, among others. For many of these applications, when using more than one trans-intein, orthogonality between them is a critical issue to ensure the proper ligation of the exteins. Here, we confirm the orthogonality (lack of cross-reactivity) of four different trans- or split inteins, gp41-1, gp41-8, IMPDH-1 and NrdJ-1 both in vivo and in vitro, and built different constructs that allow for the sequential fusion of up to four protein fragments into one final spliced product. We have characterized the splicing efficiency of these constructs. All harbor non-native extein residues at the splice junction between the trans-intein and the neighboring exteins, except for the essential Ser + 1. Our results show that it is possible to ligate four different protein domains using inteins gp41-1, IMPDH-1 and NrdJ-1 with non-native extein residues to obtain a final four-domain spliced product with a not negligible yield that keeps its native sequence.

Ir(III) Half-Sandwich Photosensitizers with a π-Expansive Ligand for Efficient Anticancer Photodynamic Therapy.

One approach to reduce the side effects of chemotherapy in cancer treatment is photodynamic therapy (PDT), which allows spatiotemporal control of the cytotoxicity. We have used the strategy of coordinating π-expansive ligands to increase the excited state lifetimes of Ir(III) half-sandwich complexes in order to facilitate the generation of 1O2. We have obtained derivatives of formulas [Cp*Ir(C∧N)Cl] and [Cp*Ir(C∧N)L]BF4 with different degrees of π-expansion in the C∧N ligands. Complexes with the more π-expansive ligand are very effective photosensitizers with phototoxic indexes PI > 2000. Furthermore, PI values of 63 were achieved with red light. Time-dependent density functional theory (TD-DFT) calculations nicely explain the effect of the π-expansion. The complexes produce reactive oxygen species (ROS) at the cellular level, causing mitochondrial membrane depolarization, cleavage of DNA, nicotinamide adenine dinucleotide (NADH) oxidation, as well as lysosomal damage. Consequently, cell death by apoptosis and secondary necrosis is activated. Thus, we describe the first class of half-sandwich iridium cyclometalated complexes active in PDT.

Purification and Biological Properties of Raniseptins-3 and -6, Two Antimicrobial Peptides from Boana raniceps (Cope, 1862) Skin Secretion.

The number of multidrug-resistant pathogenic microorganisms has been growing in recent years, most of which is due to the inappropriate use of the commercial antibiotics that are currently available. The dissemination of antimicrobial resistance represents a serious global public health problem. Thus, it is necessary to search for and develop new drugs that can act as antimicrobial agents. Antimicrobial peptides are a promising alternative for the development of new therapeutic drugs. Anurans' skin glands are a rich source of broad-spectrum antimicrobial compounds and hylids, a large and diverse family of tree frogs, are known as an important source of antimicrobial peptides. In the present study, two novel antimicrobial peptides, named Raniseptins-3 and -6, were isolated from Boana raniceps skin secretion and their structural and biological properties were evaluated. Raniseptins-3 and -6 are cationic, rich in hydrophobic residues, and adopt an α-helix conformation in the presence of SDS (35 mM). Both peptides are active against Gram-negative bacteria and Gram-positive pathogens, with low hemolytic activity at therapeutic concentrations. No activity was observed for yeasts, but the peptides are highly cytotoxic against B16F10 murine melanoma cells and NIH3T3 mouse fibroblast cells. None of the tested compounds showed improvement trends in the MTT and LDH parameters of MHV-3 infected cells at the concentrations tested.

Impact of Bifidobacterium animalis subsp. lactis HN019 probiotic in the prevention of periodontitis associated with immunosuppression.

BACKGROUND: This study evaluated the effects of the probiotic Bifidobacterium animalis subsp. lactis HN019 (HN019) in the development of experimental periodontitis (EP) in rats submitted to chemotherapy (5-fluorouracil [5FU]). METHODS: Eighty male rats were divided into the following groups: control (C); treated with 5FU (60 mg/kg at day 30 and 40 mg/kg at day 32); treated with probiotic (HN019) (daily, for 44 days, starting at day 0); treatment with 5FU and probiotic (5FU-HN019); only EP (EP) (ligature placed on lower first molars at day 30, maintained for 14 days); EP and treatment with 5FU (EP-5FU); EP and treatment with probiotic (EP-HN019); and EP and treatment with 5FU and probiotic (EP-5FU-HN019). Euthanasia occurred at day 44. Morphometric, histomorphometric, microtomographic, immunohistochemical, immunoenzymatic, and gene expressions analyses were performed. The data obtained were statistically analyzed (p < 0.05). RESULTS: The EP-5FU-HN019 group showed less bone and connective tissue loss when compared with EP-5FU group, while EP-HN019 and EP-5FU-HN019 groups had greater bone volume than EP and EP-5FU groups, respectively (p < 0.05). A decrease in immunostaining for tartrate-resistant acid phosphatase and RANKL, an increase for osteoprotegerin and lower interleukin-1ß levels were observed in EP-5FU-HN019 group, when compared with EP-5FU group (p < 0.0001). Probiotic therapy led to an increase in the proportions of B. lactis in the feces (p = 0.0018), but not in the biofilm, and reduced the expression of Fusobacterium nucleatum and Prevotella intermedia in the biofilm (p < 0.0001). CONCLUSION: B. lactis HN019 reduced the severity of EP in rats submitted to chemotherapy, modulating immunoinflammatory parameters in periodontal tissues and reducing periodontopathogens expression on biofilm in rats submitted to chemotherapy.

Dinuclear Iron Complexes of Iminopyridine-Based Ligands as Selective Cytotoxins for Tumor Cells and Inhibitors of Cancer Cell Migration.

A family of dinuclear iron (II) compounds with iminopyridine-based ligands displays selective cytotoxic activity against cancer cell lines. All compounds have IC50 values 2-6 fold lower than that of cisplatin, and 30-90 fold lower than that of carboplatin for the tumor cell lines assayed. Comparing the IC50 values between tumor and non-tumor cell lines, the selectivity indexes range from 3.2 to 34, compound 10, [Fe2(4)2(CH3CN)4](BF4)4, showing the highest selectivity. Those compounds carrying substituents on the iminopyridine ring show the same cytotoxicity as those without substituents. However, the electronic effects of the substituents on position 6 may be important for the cytotoxicity of the complexes, and consequently for their selectivity. All compounds act over DNA, promoting cuts on both strands in the presence of reactive oxygen species. Since compound 10 presented the highest selectivity, its cytotoxic effect was further characterized. It induces apoptosis, affects cell cycle phase distribution in a cell-dependent manner, and its cytotoxic effect is linked to reactive oxygen species generation. In addition, it decreases tumor cell migration, showing potential antimetastatic effects. These properties make compound 10 a good lead antitumor agent among all compounds studied here.

Brazilian Theraphosidae: a toxicological point of view.

The Theraphosidae family includes the largest number of species of the Mygalomorphae infraorder, with hundreds of species currently catalogued. However, there is a huge lack on physiologic and even ecologic information available, especially in Brazil, which is the most biodiverse country in the world. Over the years, spiders have been presented as a source of multiple biologically active compounds with basic roles, such as primary defense against pathogenic microorganisms or modulation of metabolic pathways and as specialized hunters. Spider venoms also evolved in order to enable the capture of prey by interaction with a diversity of molecular targets of interest, raising their pharmaceutical potential for the development of new drugs. Among the activities found in compounds isolated from venoms and hemocytes of Brazilian Theraphosidae there are antimicrobial, antifungal, antiparasitic and antitumoral, as well as properties related to proteinase action and neuromuscular blockage modulated by ionic voltage-gated channel interaction. These characteristics are present in different species from multiple genera, which is strong evidence of the important role in spider survival. The present review aims to compile the main results of studies from the last decades on Brazilian Theraphosidae with special focus on results obtained with the crude venom or compounds isolated from both venom and hemocytes, and their physiological and chemical characterization.

A Nuclear-Directed Ribonuclease Variant Targets Cancer Stem Cells and Inhibits Migration and Invasion of Breast Cancer Cells.

Despite the significant advances in cancer research made in recent years, this disease remains one of the leading causes of death worldwide. In part, this is due to the fact that after therapy, a subpopulation of self-renewing tumor cells can survive and promote cancer relapse, resistance to therapies and metastasis. Targeting these cancer stem cells (CSCs) is therefore essential to improve the clinical outcome of cancer patients. In this sense, multi-targeted drugs may be promising agents targeting CSC-associated multifocal effects. We have previously constructed different human pancreatic ribonuclease (RNase) variants that are cytotoxic for tumor cells due to a non-classical nuclear localization signal introduced in their sequence. These cytotoxic RNases affect the expression of multiple genes involved in deregulated metabolic and signaling pathways in cancer cells and are highly cytotoxic for multidrug-resistant tumor cell lines. Here, we show that these cytotoxic nuclear-directed RNases are highly selective for tumor cell lines grown in 3D, inhibit CSCs' development and diminish the self-renewal capacity of the CSCs population. Moreover, these human RNase variants reduce the migration and invasiveness of highly invasive breast cancer cells and downregulate N-cadherin expression.

The Selectivity for Tumor Cells of Nuclear-Directed Cytotoxic RNases Is Mediated by the Nuclear/Cytoplasmic Distribution of p27KIP1.

Although single targeted anti-cancer drugs are envisaged as safer treatments because they do not affect normal cells, cancer is a very complex disease to be eradicated with a single targeted drug. Alternatively, multi-targeted drugs may be more effective and the tumor cells may be less prone to develop drug resistance although these drugs may be less specific for cancer cells. We have previously developed a new strategy to endow human pancreatic ribonuclease with antitumor action by introducing in its sequence a non-classical nuclear localization signal. These engineered proteins cleave multiple species of nuclear RNA promoting apoptosis of tumor cells. Interestingly, these enzymes, on ovarian cancer cells, affect the expression of multiple genes implicated in metabolic and signaling pathways that are critic for the development of cancer. Since most of these targeted pathways are not highly relevant for non-proliferating cells, we envisioned the possibility that nuclear directed-ribonucleases were specific for tumor cells. Here, we show that these enzymes are much more cytotoxic for tumor cells in vitro. Although the mechanism of selectivity of NLSPE5 is not fully understood, herein we show that p27KIP1 displays an important role on the higher resistance of non-tumor cells to these ribonucleases.

Strengths and Challenges of Secretory Ribonucleases as AntiTumor Agents.

Approaches to develop effective drugs to kill cancer cells are mainly focused either on the improvement of the currently used chemotherapeutics or on the development of targeted therapies aimed at the selective destruction of cancer cells by steering specific molecules and/or enhancing the immune response. The former strategy is limited by its genotoxicity and severe side effects, while the second one is not always effective due to tumor cell heterogeneity and variability of targets in cancer cells. Between these two strategies, several approaches target different types of RNA in tumor cells. RNA degradation alters gene expression at different levels inducing cell death. However, unlike DNA targeting, it is a pleotropic but a non-genotoxic process. Among the ways to destroy RNA, we find the use of ribonucleases with antitumor properties. In the last few years, there has been a significant progress in the understanding of the mechanism by which these enzymes kill cancer cells and in the development of more effective variants. All the approaches seek to maintain the requirements of the ribonucleases to be specifically cytotoxic for tumor cells. These requirements start with the competence of the enzymes to interact with the cell membrane, a process that is critical for their internalization and selectivity for tumor cells and continue with the downstream effects mainly relying on changes in the RNA molecular profile, which are not only due to the ribonucleolytic activity of these enzymes. Although the great improvements achieved in the antitumor activity by designing new ribonuclease variants, some drawbacks still need to be addressed. In the present review, we will focus on the known mechanisms used by ribonucleases to kill cancer cells and on recent strategies to solve the shortcomings that they show as antitumor agents, mainly their pharmacokinetics.

Brazilian Theraphosidae: a toxicological point of view

The Theraphosidae family includes the largest number of species of the Mygalomorphae infraorder, with hundreds of species currently catalogued. However, there is a huge lack on physiologic and even ecologic information available, especially in Brazil, which is the most biodiverse country in the world. Over the years, spiders have been presented as a source of multiple biologically active compounds with basic roles, such as primary defense against pathogenic microorganisms or modulation of metabolic pathways and as specialized hunters. Spider venoms also evolved in order to enable the capture of prey by interaction with a diversity of molecular targets of interest, raising their pharmaceutical potential for the development of new drugs. Among the activities found in compounds isolated from venoms and hemocytes of Brazilian Theraphosidae there are antimicrobial, antifungal, antiparasitic and antitumoral, as well as properties related to proteinase action and neuromuscular blockage modulated by ionic voltage-gated channel interaction. These characteristics are present in different species from multiple genera, which is strong evidence of the important role in spider survival. The present review aims to compile the main results of studies from the last decades on Brazilian Theraphosidae with special focus on results obtained with the crude venom or compounds isolated from both venom and hemocytes, and their physiological and chemical characterization.(AU)

Brazilian Theraphosidae: a toxicological point of view

Abstract The Theraphosidae family includes the largest number of species of the Mygalomorphae infraorder, with hundreds of species currently catalogued. However, there is a huge lack on physiologic and even ecologic information available, especially in Brazil, which is the most biodiverse country in the world. Over the years, spiders have been presented as a source of multiple biologically active compounds with basic roles, such as primary defense against pathogenic microorganisms or modulation of metabolic pathways and as specialized hunters. Spider venoms also evolved in order to enable the capture of prey by interaction with a diversity of molecular targets of interest, raising their pharmaceutical potential for the development of new drugs. Among the activities found in compounds isolated from venoms and hemocytes of Brazilian Theraphosidae there are antimicrobial, antifungal, antiparasitic and antitumoral, as well as properties related to proteinase action and neuromuscular blockage modulated by ionic voltage-gated channel interaction. These characteristics are present in different species from multiple genera, which is strong evidence of the important role in spider survival. The present review aims to compile the main results of studies from the last decades on Brazilian Theraphosidae with special focus on results obtained with the crude venom or compounds isolated from both venom and hemocytes, and their physiological and chemical characterization.

Construction of Highly Stable Cytotoxic Nuclear-Directed Ribonucleases.

Ribonucleases are proteins whose use is promising in anticancer therapy. We have previously constructed different human pancreatic ribonuclease variants that are selectively cytotoxic for tumor cells by introducing a nuclear localization signal into their sequence. However, these modifications produced an important decrease in their stability compromising their behavior in vivo. Here, we show that we can significantly increase the thermal stability of these cytotoxic proteins by introducing additional disulfide bonds by site-directed mutagenesis. One of these variants increases its thermal stability by around 17 °C, without affecting its catalytic activity while maintaining the cytotoxic activity against tumor cells. We also show that the most stable variant is significantly more resistant to proteolysis when incubated with proteinase K or with human sera, suggesting that its half-live could be increased in vivo once administered.

Apoptin, A Versatile Protein with Selective Antitumor Activity.

BACKGROUND: Research in the field of antitumor chemotherapeutics pursues a key issue, drug selectivity for cancer cells. In the last 20 years, a group of proteins has attracted scientific interest as cancer chemotherapeutics due to their ability to specifically kill cancer cells while leaving normal cells undamaged. One of these proteins is apoptin. METHODS: In this study, the recent available literature regarding cell death mechanisms induced by apoptin has been reviewed. Delivering this drug to tumor cells is a challenge because it spontaneously forms soluble non-covalent aggregates. This led us to include in this review the different approaches for obtaining the maximum efficiency of apoptin entry to cancer cells. RESULTS: This review provides an up-to-date summary of the mechanisms by which apoptin induces selective apoptosis in tumor cells while leaving normal cells undamaged. It highlights the relationship between the apoptosis mechanism induced by this protein and its functional motifs. Apoptin has been described as an intrinsically disordered protein, which explains its ability to interact with multiple partners and affect multiple pathways inside the cell. Characterization of the different partners and pathways induced by apoptin has begun to shed light on the molecular basis of apoptin's tumor-selective cytotoxicity. CONCLUSION: The findings confirm the interest in apoptin as a potentially safe antitumor drug. Research still needed to be conducted to find an effective way to deliver apoptin for use in clinics.

Desarrollo de un sistema georreferenciado para la gestión, movilidad y monitoreo de atención primaria de la salud comunitaria / Development of a georeferenced system for the management, mobility and monitoring of primary care in community health

RESUMEN Los agentes de salud comunitaria en terreno practican acciones de fuerte impronta territorial, tanto en relación con prevención y asistencia, como a partir de un acercamiento de los servicios de salud a las familias. Estas prácticas en terreno, construyen un permanente mapeo colectivo basado en el conocimiento que los agentes poseen del territorio de actuación. A partir de un intenso trabajo de campo iniciado en el año 2012, en conjunto con organizaciones sociales locales e instituciones de salud de Brasil y Argentina, se inició el proyecto binacional App+Salud, que tiene como objetivo el desarrollo de un sistema de monitoreo y georreferencia para la gestión de la salud comunitaria, en formato de aplicación, para celular o tablet y acceso web, que considere la condiciones básicas, de ambiente y movilidad histórica de los usuarios, y que vincule metodologías de actualización cartográfica a partir de cartografía social con plataformas de mapeos colaborativos libres. Como resultado, se ha obtenido el desarrollo del sistema App+Salud, en nivel beta, para lo cual se profundizó en un trabajo de intercambio de saberes interdisciplinares.

ABSTRACT Community health agents carry out significant actions on the ground to provide prevention and care and bring health services to families. These practices in the field constitute a constant collective mapping process using the knowledge of the territory that the agents possess. Based on intensive fieldwork starting in 2012 in conjunction with local social organizations and health institutions in Brazil and Argentina, the binational project App+Health (App+Salud in Spanish and App+Saúde in Portugués) was initiated. The project's objective was to develop a monitoring and georeferencing system for community health management in the form of an application for cell phones or tablets with internet access, which would take into account the basic conditions of the environment and movement history of users and connect map updating methodologies using social cartography with free collaborative mapping platforms. As a result, the beta version of App+Health was developed, through a heightened process of exchange of interdisciplinary knowledge.

A family of manganese complexes containing heterocyclic-based ligands with cytotoxic properties.

We describe the synthesis of three new manganese (II) complexes containing the bidentate ligands 2-(1-methyl-3-pyrazolyl)pyridine (pypz-Me) and ethyl 2-(3-(pyridine-2-yl)-1H-pyrazol-1-yl)acetate (pypz-CH2COOEt), with formula [MnX2(pypz-Me)2] (X = Cl-1, CF3SO3-2) and [Mn(CF3SO3)2(pypz-CH2COOEt)2] 3. Complexes 1-3 have been characterized through analytical, spectroscopic and electrochemical techniques, as well as by monocrystal X-ray diffraction analysis. The complexes show a six-coordinated Mn(II) ion though different stereoisomers have been isolated for the three compounds. Complexes 1-3, together with the previously described compounds [MnCl2(pypz-H)2] 4, [Mn(CF3SO3)2(pypz-H)2] 5, [Mn(NO3)2(pypz-H)2] 6, [MnCl2(H2O)2(pypz-H)2] 7 (pypz-H = 2-(3-pyrazolyl)pyridine) and ([Mn(CF3SO3)2((-)-L)2] 8, ((-)-L = (-)-pinene[5,6]bipyridine), were tested in vitro for cytotoxic activity against NCI-H460 and OVCAR-8 cancer cell lines. The geometry of a specific compound does not seem to influence its activity in a significant extent. However, among the tested compounds those that display hydrophobic substituents on the pyrazole ring and triflate ions as labile ligands show the best antiproliferative properties. Specifically, compound 8 containing the pinene-bipyridine ligand presents an antiproliferative activity similar to that of cisplatin and higher than that of carboplatin, and displays selectivity for tumour cells. Its antiproliferative effect is due to the generation of ROS species that allow the compound to interact with DNA.

Transcriptional profiling of NCI/ADR-RES cells unveils a complex network of signaling pathways and molecular mechanisms of drug resistance.

BACKGROUND: Ovarian cancer has the highest mortality rate among all the gynecological cancers. This is mostly due to the resistance of ovarian cancer to current chemotherapy regimens. Therefore, it is of crucial importance to identify the molecular mechanisms associated with chemoresistance. METHODS: NCI/ADR-RES is a multidrug-resistant cell line that is a model for the study of drug resistance in ovarian cancer. We carried out a microarray-derived transcriptional profiling analysis of NCI/ADR-RES to identify differentially expressed genes relative to its parental OVCAR-8. RESULTS: Gene-expression profiling has allowed the identification of genes and pathways that may be important for the development of drug resistance in ovarian cancer. The NCI/ADR-RES cell line has differential expression of genes involved in drug extrusion, inactivation, and efficacy, as well as genes involved in the architectural and functional reorganization of the extracellular matrix. These genes are controlled through different signaling pathways, including MAPK-Akt, Wnt, and Notch. CONCLUSION: Our findings highlight the importance of using orthogonal therapies that target completely independent pathways to overcome mechanisms of resistance to both classical chemotherapeutic agents and molecularly targeted drugs.

Entorno y cuidado: circunstancias que influyen en la calidad de la atención médica percibida por los colombianos / Environment and care: circumstances that influence the quality of healthcare perceived by the people in Colombia / Ambiente e cuidado: circunstâncias que influenciam a qualidade da atenção médica percebida pelos colombianos

Introducción. El análisis de las situaciones de salud, en el acto del cuidado de la enfermería, requiere de conocimientos, pues es el marco conceptual y la guía de esta labor. Aunque el cuidado integral de la salud, según Watson, depende del compromiso moral del enfermero de preservar la dignidad humana; son frecuentes los informes que muestran una mala percepción de la prestación de servicios de salud. En los informes, se indica que el 36% de los encuestados tienen una mala o muy mala percepción de la calidad de la atención médica. El objetivo es reflexionar sobre las circunstancias que influyen en la calidad de la atención médica percibida por los colombianos. Temas de reflexión. Se abordará el tema desde los patrones de conocimiento de la enfermería: el empírico, el estético, el ético, el sociocultural y el personal. Discusión. Las condiciones que influyen en la percepción de la calidad de la atención médica se agrupan en dos conceptos del metaparadigma de la enfermería. El primero es el entorno, en el que existen barreras de oferta y demanda, como la tecnificación, la infraestructura y los recursos del sistema de salud. El segundo es el cuidado, en el que la condición laboral tiene incidencia, ya que los profesionales de enfermería sienten culpabilidad, agotamiento y estrés, y comprometen la buena percepción del cuidado médico y el sentido de trascendencia profesional. Conclusiones. El acto del cuidado de la enfermería está rodeado de situaciones poco favorables, dadas las barreras en el sistema de salud colombiano, que comprometen la labor del enfermero. Por lo tanto, es necesario reflexionar y desarrollar conceptos relacionados con los patrones de conocimiento de la disciplina, que inciden en la formación de los nuevos profesionales. Esto, con el fin de cumplir a cabalidad el deber ser y el hacer de la enfermería en Colombia. [Duque-Castro JA, Ortiz-Urbano J, Rengifo-Arias DM. Entorno y cuidado: circunstancias que influyen en la calidad de la atención percibida por los colombianos. MedUNAB. 2018;21(2):60-68. doi:10.29375/01237047.2799].

Introduction. The analysis of health situations, in terms of nursing care, requires knowledge, since it is the conceptual framework and guide for this work. Although comprehensive healthcare, according to Watson, depends on the nurse's moral commitment to preserve human dignity, there are frequent reports that show a poor perception of the provision of healthcare services. The reports indicate that 36% of those surveyed had a poor or very poor perception of the quality of healthcare. The objective is to reflect on the circumstances that influence the quality of healthcare perceived by the people in Colombia. Topics of reflection. The topic will be addressed through the patterns of knowledge of nursing: empirical, aesthetic, ethical, sociocultural and personal. Discussion. The conditions that influence the perception of healthcare quality are grouped into two nursing metaparadigm concepts. The first is the environment, in which there are supply and demand barriers, such as technification, infrastructure and resources of the healthcare system. The second is the care, on which the employment status has an influence, since nursing professionals feel guilt, exhaustion and stress, and compromise the good perception of healthcare and the sense of professional importance. Conclusions. The act of caring in nursing is surrounded by unfavorable situations, given the barriers in the Colombian healthcare system that compromise the work of nurses. Therefore, it is necessary to reflect and develop concepts related to the patterns of knowledge of this discipline, which have an effect on the training of new professionals. This is in order to fully comply with the duty and practice of nursing in Colombia. [Duque-Castro JA, Ortiz-Urbano J, Rengifo-Arias DM. Environment and care: circumstances that influence the quality of medical care perceived by the people in Colombia. MedUNAB. 2018;21(2):60-68. doi:10.29375/01237047.2799].

Introdução. A análise das situações de saúde, no ato do cuidado da enfermagem, requer conhecimento, pois é o referencial conceitual e o guia desta atividade. Embora os cuidados integrais de saúde, segundo Watson, dependem do compromisso moral do enfermeiro para preservar a dignidade humana; há relatos frequentes que mostram uma má percepção da prestação de serviços de saúde. Nos relatórios, indica-se que 36% dos entrevistados têm uma percepção ruim ou muito ruim da qualidade do atendimento médico. O objetivo é refletir sobre as circunstâncias que influenciam a qualidade da atenção médica percebida pelos colombianos. Temas para reflexão. O tema será abordado a partir dos padrões de conhecimento da enfermagem: o empírico, o estético, o ético, o sociocultural e o pessoal. Discussão. As condições que influenciam a percepção da qualidade da atenção médica são agrupadas em dois conceitos do metaparadigma da enfermagem. O primeiro é o ambiente, no qual existem barreiras de oferta e demanda, como a tecnificação, a infraestrutura e os recursos do sistema de saúde. O segundo é o cuidado, no qual a condição de trabalho tem impacto, uma vez que os profissionais de enfermagem sentem-se culpados, exaustos e estressados, comprometendo a boa percepção do cuidado médico e o sentido de transcendência profissional. Conclusões. O ato do cuidado de enfermagem está cercado de situações desfavoráveis, dadas as barreiras no sistema de saúde colombiano, que comprometem o trabalho do enfermeiro. Portanto, é necessário refletir e desenvolver conceitos relacionados aos padrões de conhecimento da disciplina, que afetam a formação dos novos profissionais. Isso, a fim de cumprir integralmente o dever de ser e exercer a enfermagem na Colômbia. [Duque-Castro JA, Ortiz-Urbano J, Rengifo-Arias DM. Ambiente e cuidado: circunstâncias que influenciam a qualidade da atenção médica percebida pelos colombianos. MedUNAB. 2018;21(2):60-68. doi:10.29375/01237047.2799].

A truncated apoptin protein variant selectively kills cancer cells.

Apoptin is a nonstructural protein encoded by one of the three open reading frames of the chicken anemia virus genome. It has attracted a great deal of interest due to its ability to induce apoptosis in multiple transformed and malignant mammalian cell lines without affecting primary and non-transformed cells. However, the use of Apoptin as an anticancer drug is restricted by its strong tendency to aggregate. A number of methods to overcome this problem have been proposed, including transduction techniques to deliver the Apoptin gene into tumor cells, but all such methods have certain drawbacks. Here we describe that a truncated variant of Apoptin, lacking residues 1 to 43, is a soluble, non-aggregating protein that maintains most of the biological properties of wild-type Apoptin when transfected into cells. We show that the cytotoxic effect of this variant is also present when it is added exogenously to cancer cells, but not to normal cells. In addition to the interest this protein has attracted as a promising therapeutic strategy, it is also an excellent model to study the structural properties of Apoptin and how they relate to its mechanism of action.

Activating transcription factor 3 is crucial for antitumor activity and to strengthen the antiviral properties of Onconase.

Onconase is a ribonuclease that presents both antitumor and antiviral properties linked to its ribonucleolytic activity and represents a new class of RNA-damaging drugs. It has reached clinical trials for the treatment of several cancers and human papilloma virus warts. Onconase targets different RNAs in the cell cytosol but Onconase-treated cells present features that are different from a simple arrest of protein synthesis. We have used microarray-derived transcriptional profiling to identify Onconase-regulated genes in two ovarian cancer cell lines (NCI/ADR-RES and OVCAR-8). RT-qPCR analyses have confirmed the microarray findings. We have identified a network of up-regulated genes implicated in different signaling pathways that may explain the cytotoxic effects exerted by Onconase. Among these genes, activating transcription factor 3 (ATF3) plays a central role in the key events triggered by Onconase in treated cancer cells that finally lead to apoptosis. This mechanism, mediated by ATF3, is cell-type independent. Up-regulation of ATF3 may also explain the antiviral properties of this ribonuclease because this factor is involved in halting viral genome replication, keeping virus latency or preventing viral oncogenesis. Finally, Onconase-regulated genes are different from those affected by nuclear-directed ribonucleases.

Insights into the mechanism of Apoptin's exquisitely selective anti-tumor action from atomic level characterization of its conformation and dynamics.

Apoptin is a 121 residue protein which forms large, soluble aggregates and possesses an exceptionally selectively cytotoxic action on cancer cells. In the accompanying paper, we described the design, production and initial characterization of an Apoptin truncated variant called H6-ApopΔProΔLeu. Whereas both the variant and wild type protein possess similar selective cytotoxicity against cancer cells following transfection, only the variant is cytotoxic when added externally. Remarkably, as observed by gel filtration chromatography and dynamic light scattering, H6-ApopΔProΔLeu lacks the tendency of wild type Apoptin to form large aggregates, which greatly facilitated the study of its biological properties. Here, we characterize the conformation and dynamics of H6-ApopΔProΔLeu. Using a battery of 2D, 3D and (4,2)D NMR spectra, the essentially complete 1H, 13C and 15N resonance assignments of H6-ApopΔProΔLeu were obtained. The analysis of these data shows that the variant is an intrinsically disordered protein, which lacks a preferred conformation. This conclusion is corroborated by a lack of protection against proteolytic cleavage and hydrogen/deuterium exchange. Moreover, the CD spectra are dominated by random coil contributions. Finally, 1H-15N NOE ratios are low, which indicates flexibility on the ps-ns time scale. Interestingly, H6-ApopΔProΔLeu's intrinsically disordered ensemble is not significantly altered by the redox state of its Cys residues or by Thr phosphorylation, which has been proposed to play a key role in Apoptin's selective cytotoxicity. These results serve to better comprehend Apoptin's remarkably selective anticancer action and provide a framework for the future design of improved Apoptin variants.