Characterization of LRRTM and NGR gene families: Expression and functions

Some leucine-rich repeat (LRR) -containing membrane proteins are known regulators of neuronal growth and synapse formation. In this work I characterize two gene families encoding neuronal LRR membrane proteins, namely the LRRTM (leucine-rich repeat, transmembrane neuronal) and NGR (Nogo-66 receptor) families. I studied LRRTM and NGR family member's mRNA tissue distribution by RT-PCR and by in situ hybridization. Subcellular localization of LRRTM1 protein was studied in neurons and in non-neuronal cells. I discovered that LRRTM and NGR family mRNAs are predominantly expressed in the nervous system, and that each gene possesses a specific expression pattern. I also established that LRRTM and NGR family mRNAs are expressed by neurons, and not by glial cells. Within neurons, LRRTM1 protein is not transported to the plasma membrane; rather it localizes to endoplasmic reticulum. Nogo-A (RTN4), MAG, and OMgp are myelin-associated proteins that bind to NgR1 to limit axonal regeneration after central nervous system injury. To better understand the functions of NgR2 and NgR3, and to explore the possible redundancy in the signaling of myelin inhibitors of neurite growth, I mapped the interactions between NgR family and the known and candidate NgR1 ligands. I identified high-affinity interactions between RTN2-66, RTN3-66 and NgR1. I also demonstrate that Rtn3 mRNA is expressed in the same glial cell population of mouse spinal cord white matter as Nogo-A mRNA, and thus it could have a role in myelin inhibition of axonal growth. To understand how NgR1 interacts with multiple structurally divergent ligands, I aimed first to map in more detail the nature of Nogo-A:NgR1 interactions, and then to systematically map the binding sites of multiple myelin ligands in NgR1 by using a library of NgR1 expression constructs encoding proteins with one or multiple surface residues mutated to alanine. My analysis of the Nogo-A:NgR1 -interactions revealed a novel interaction site between the proteins, suggesting a trivalent Nogo-A:NgR1-interaction. Our analysis also defined a central binding region on the concave side of NgR1's LRR domain that is required for the binding of all known ligands, and a surrounding region critical for binding MAG and OMgp. To better understand the biological role of LRRTMs, I generated Lrrtm1 and Lrrtm3 knock out mice. I show here that reporter genes expressed from the targeted loci can be used for maping the neuronal connections of Lrrtm1 and Lrrtm3 expressing neurons in finer detail. With regard to LRRTM1's role in humans, we found a strong association between a 70 kb-spanning haplotype in the proposed promoter region of LRRTM1 gene and two possibly related phenotypes: left-handedness and schizophrenia. Interestingly, the responsible haplotype was linked to phenotypic variability only when paternally inherited. In summary, I identified two families of neuronal receptor-like proteins, and mapped their expression and certain protein-protein interactions. The identification of a central binding region in NgR1 shared by multiple ligands may facilitate the design and development of small molecule therapeutics blocking binding of all NgR1 ligands. Additionally, the genetic association data suggests that allelic variation upstream of LRRTM1 may play a role in the development of left-right brain asymmetry in humans. Lrrtm1 and Lrrtm3 knock out mice developed as a part of this study will likely be useful for schizophrenia and Alzheimer s disease research.

Monimuotoisuus ja esteettömyys : Näkövammaisten asiantuntijoiden työelämäkokemuksia

The purpose of this study is to analyse education, employment, and work-life experiences of visually impaired persons in expert jobs. The empirical data consists of 30 thematic interviews (24 visually impaired persons, 1 family-member of a visually impaired person, 5 persons working with diversity issues), of supplementary articles, and of statistics on the socio-economic status of the visually impaired. The interviewees experiences of education and employment have been analysed by a qualitative method. The analysis has been deepened by reflecting it against the recent discussion on the concept of diversity. The author s methodological choice as a disability researcher has been to treat the interviewees as co-researchers rather than objects of research. Accessibility in its different forms is a prerequisite of diversity in the workplace, and this study examines what kind of accessibility is required by visually impaired professionals. Access to working life depends on the attitudes prejudices and expectations that society has towards a minority group. Social accessibility is connected with internal relationships in the workplace, and achieving social accessibility is a bilateral process. Information technology has revolutionised the visually impaired people s possibilities of accessing information and performing expert tasks. Accessible environment, good mobility skills, and transportation services enable visually impaired employees to get to their workplaces and to navigate there with ease. Integration has raised the level of education and widened the selection of career options for the visually impaired. However, even visually impaired people with academic degrees often need employment support services. Visually impaired professionals are mainly employed in the public and third sector. Achieving diversity in the labour market is a multiactor process. Social support services are needed, as well as courage and readiness from employers to hire people with disabilities. The organisations of the visually impaired play an important role in affecting the attitudes and providing peer support. Visually impaired employees need good professional skills, blindness skills, and social courage, and they need to be comfortable with their disability. In the workplace, diversity may actualise as diverse ways of working: the work is done by using technical aids or other means of compensating for the lack of eyesight. When an employee must find compensatory solutions for disability-related limitations at work, this will also develop his/her problem-solving abilities. Key words: visually impaired, diversity, accessibility, working life

Eron aika : Tyttärien kertomuksia ikääntyneen vanhemman kuolemasta

There is a relative absence of sociological and cultural research on how people deal with the death of a family member in the contemporary western societies. Research on this topic has been dominated by the experts of psychology, psychiatry and therapy, who mention the social context only in passing, if at all. This gives an impression that the white westerners bereavement experience is a purely psychological phenomenon, an inner journey, which follows a natural, universal path. Yet, as Tony Walter (1999) states, ignoring the influence of culture not only impoverishes the understanding of those work with bereaved people, but it also impoverishes sociology and cultural studies by excluding from their domain a key social phenomenon. This study explores the cultural dimension of grief through narratives told by fifteen of recently bereaved Finnish women. Focussing on one sex only, the study rests on the assumption of the gendered nature of bereavement experience. However, the aim of the study is not to pinpoint the gender differences in grief and mourning, but to shed light on women s ways of dealing with the loss of a loved one in a social context. Furthermore, the study focuses on a certain kind of loss: the death of an elderly parent. Due to the growth in the life expectancy rate, this has presumably become the most typical type of bereavement in contemporary, ageing societies. Most of population will face the death of a parent as they reach the middle years of the life course. The data of this study is gathered with interviews, in which the interviewees were invited to tell a narrative of their bereavement. Narrative constitutes a central concept in this study. It refers to a particular form of talk, which is organised around consequential events. But there are also other, deeper layers that have been added to this concept. Several scholars see narratives as the most important way in which we make sense of experience. Personal narratives provide rich material for mapping the interconnections between individual and culture. As a form of thought, narrative marries singular circumstances with shared expectations and understandings that are learned through participation in a specific culture (Garro & Mattingly 2000). This study attempts to capture the cultural dimension of narrative with the concept of script , which originates in cognitive science (Schank & Abelson 1977) and has recently been adopted to narratology (Herman 2002). Script refers to a data structure that informs how events usually unfold in certain situations. Scripts are used in interpreting events and representing them verbally to others. They are based on dominant forms of knowledge that vary according to time and place. The questions that were posed in this study are the following. What kind of experiences bereaved daughters narrate? What kind of cultural scripts they employ as they attempt to make sense of these experiences? How these scripts are used in their narratives? It became apparent that for the most of the daughters interviewed in this study the single most important part of the bereavement narrative was to form an account of how and why the parent died. They produced lengthy and detailed descriptions of the last stage of a parent s life in contrast with the rest of the interview. These stories took their start from a turn in the parent s physical condition, from which the dying process could in retrospect be seen to have started, and which often took place several years before the death. In addition, daughters also talked about their grief reactions and how they have adjusted to a life without the deceased parent. The ways in which the last stage of life was told reflect not only the characteristic features of late modernity but also processes of marginalisation and exclusion. Revivalist script and medical script, identified by Clive Seale as the dominant, competing models for dying well in the late modern societies, were not widely utilised in the narratives. They could only be applied in situations in which the parent had died from cancer and at somewhat younger age than the average. Death that took place in deep old age was told in a different way. The lack of positive models for narrating this kind of death was acknowledged in the study. This can be seen as a symptom of the societal devaluing of the deaths of older people and it affects also daughters accounts of their grief. Several daughters told about situations in which their loss, although subjectively experienced, was nonetheless denied by other people.

mRNA-binding protein HuR in breast carcinogenesis

Breast cancer is the most common cancer among women. Although its prognosis has improved nowadays, methods to predict the progression of the disease or to treat it are not comprehensive. This thesis work was initiated to elucidate in breast carcinogenesis the role of HuR, a ubiquitously expressed mRNA-binding protein that regulates gene expression posttranscriptionally. HuR is predominantly nuclear, but it shuttles between the nucleus and the cytoplasm, and this nucleocytoplasmic translocation is important for its function as a RNA-stabilizing and translational regulator. HuR has been associated with diverse cellular processes, for example carcinogenesis. The specific aims of my thesis work were to study the prognostic value of HuR in breast cancer and to clarify the mechanisms by which HuR contributes to breast carcinogenesis. My ultimate goal is, by better understanding the role of HuR in breast carcinogenesis, to aid in the discovery of novel targets for cancer therapies. HuR expression and localization was studied in paraffin-embedded preinvasive (atypical ductal hyperplasia, ADH, and ductal carcinoma in situ, DCIS) specimens as well in sporadic and familial breast cancer specimens. Our results show that cytoplasmic HuR expression was already elevated in ADH and remained elevated in DCIS as well as in cancer specimens. Clinicopathological analysis showed that cytoplasmic HuR expression associated with the more aggressive form of the disease in DCIS, and in cancer specimens it proved an independent marker for poor prognosis. Importantly, cytoplasmic HuR expression was significantly associated with poor outcome in the subgroups of small (2 cm) and axillary lymph node-negative breast cancers. HuR proved to be the first mRNA stability protein the expression of which is associated in breast cancer with poor outcome. To explore the mechanisms of HuR in breast carcinogenesis, lentiviral constructs were developed to inhibit and to overexpress the HuR expression in a breast epithelial cell line (184B5Me). Our results suggest that HuR mediates breast carcinogenesis by participating in processes important in cell transformation, in programmed cell death, and in cell invasion. Global gene expression analysis shows that HuR regulates genes participating in diverse cellular processes, and affects several pathways important in cancer development. In addition, we identified two novel target transcripts (connective tissue growth factor, CTGF, and Ras oncogene family member 31, RAB31) for HuR. In conclusion, because cytoplasmic HuR expression in breast cancer can predict the outcome of the disease it could serve in clinics as a prognostic marker. HuR accumulates in the cytoplasm even at its non-invasive stage (ADH and DCIS) of the carcinogenic process and supports functions essential in cell alteration. These data suggest that HuR contributes to carcinogenesis of the breast epithelium.

Effects of gender, and SLCO1B1 and CYP7A1 polymorphisms on plasma bile acids in humans and the pharmacokinetics of therapeutic ursodeoxycholic acid

Bile acids are important steroid-derived molecules essential for fat absorption in the small intestine. They are produced in the liver and secreted into the bile. Bile acids are transported by bile flow to the small intestine, where they aid the digestion of lipids. Most bile acids are reabsorbed in the small intestine and return to the liver through the portal vein. The whole recycling process is referred to as the enterohepatic circulation, during which only a small amount of bile acids are removed from the body via faeces. The enterohepatic circulation of bile acids involves the delicate coordination of a number of bile acid transporters expressed in the liver and the small intestine. Organic anion transporting polypeptide 1B1 (OATP1B1), encoded by the solute carrier organic anion transporter family, member 1B1 (SLCO1B1) gene, mediates the sodium independent hepatocellular uptake of bile acids. Two common SNPs in the SLCO1B1 gene are well known to affect the transport activity of OATP1B1. Moreover, bile acid synthesis is an important elimination route for cholesterol. Cholesterol 7α-hydroxylase (CYP7A1) is the rate-limiting enzyme of bile acid production. The aim of this thesis was to investigate the effects of SLCO1B1 polymorphism on the fasting plasma levels of individual endogenous bile acids and a bile acid synthesis marker, and the pharmacokinetics of exogenously administered ursodeoxycholic acid (UDCA). Furthermore, the effects of CYP7A1 genetic polymorphism and gender on the fasting plasma concentrations of individual endogenous bile acids and the bile acid synthesis marker were evaluated. Firstly, a high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for the determination of bile acids was developed (Study I). A retrospective study examined the effects of SLCO1B1 genetic polymorphism on the fasting plasma concentrations of individual bile acids and a bile acid synthesis marker in 65 healthy subjects (Study II). In another retrospective study with 143 healthy individuals, the effects of CYP7A1 genetic polymorphism and gender as well as SLCO1B1 polymorphism on the fasting plasma levels of individual bile acids and the bile acid synthesis marker were investigated (Study III). The effects of SLCO1B1 polymorphism on the pharmacokinetics of exogenously administered UDCA were evaluated in a prospective genotype panel study including 27 healthy volunteers (Study IV). A robust, sensitive and simple HPLC-MS/MS method was developed for the simultaneous determination of 16 individual bile acids in human plasma. The method validation parameters for all the analytes met the requirements of the FDA (Food and Drug Administration) bioanalytical guidelines. This HPLC-MS/MS method was applied in Studies II-IV. In Study II, the fasting plasma concentrations of several bile acids and the bile acid synthesis marker seemed to be affected by SLCO1B1 genetic polymorphism, but these findings were not replicated in Study III with a larger sample size. Moreover, SLCO1B1 polymorphism had no effect on the pharmacokinetic parameters of exogenously administered UDCA. Furthermore, no consistent association was observed between CYP7A1 genetic polymorphism and the fasting plasma concentrations of individual bile acids or the bile acid synthesis marker. In contrast, gender had a major effect on the fasting plasma concentrations of several bile acids and also total bile acids. In conclusion, gender, but not SLCO1B1 or CYP7A1 polymorphisms, has a major effect on the fasting plasma concentrations of individual bile acids. Moreover, the common genetic polymorphism of CYP7A1 is unlikely to influence the activity of CYP7A1 under normal physiological conditions. OATP1B1 does not play an important role in the in vivo disposition of exogenously administered UDCA.

Roles of Salmonella enterica serovar Typhimurium encoded Peptidase N during systemic infection of Ifnγ(-/-) mice.

Pathogen encoded peptidases are known to be important during infection; however, their roles in modulating host responses in immunocompromised individuals are not well studied. The roles of S. typhimurium (WT) encoded Peptidase N (PepN), a major aminopeptidase and sole M1 family member, was studied in mice lacking Interferon-γ (IFNγ), a cytokine important for immunity. S. typhimurium lacking pepN (ΔpepN) displays enhanced colony forming units (CFU) compared to WT in peripheral organs during systemic infection in C57BL/6 mice. However, Ifnγ(-/-) mice show higher CFU compared to C57BL/6 mice, resulting in lower fold differences between WT and ΔpepN. Concomitantly, reintroduction of pepN in ΔpepN (ΔpepN/pepN) reduces CFU, demonstrating pepN-dependence. Interestingly, expression of a catalytically inactive PepN (ΔpepN/E298A) also lowers CFU, demonstrating that the decrease in CFU is independent of the catalytic activity of PepN. In addition, three distinct differences are observed between infection of C57BL/6 and Ifnγ(-/-) mice: First, serum amounts of TNFα and IL1β post infection are significantly lower in Ifnγ(-/-) mice. Second, histological analysis of C57BL/6 mice reveals that damage in spleen and liver upon infection with WT or ΔpepN is greater compared to ΔpepN/pepN or ΔpepN/E298A. On the other hand, Ifnγ(-/-) mice are highly susceptible to organ damage by all strains of S. typhimurium used in this study. Finally, greater survival of C57BL/6, but not Ifnγ(-/-) mice, is observed upon infection with ΔpepN/pepN or ΔpepN/E298A. Overall, the roles of the host encoded IFNγ during infection with S. typhimurium strains with varying degrees of virulence are highlighted.

Catalytic activity of Peptidase N is required for adaptation of Escherichia coli to nutritional downshift and high temperature stress

Peptidase N (PepN), the sole M1 family member in Escherichia coli, displays broad substrate specificity and modulates stress responses: it lowers resistance to sodium salicylate (NaSal)-induced stress but is required during nutritional downshift and high temperature (NDHT) stress. The expression of PepN does not significantly change during different growth phases in LB or NaSal-induced stress; however, PepN amounts are lower during NDHT stress. To gain mechanistic insights on the roles of catalytic activity of PepN in modulating these two stress responses, alanine mutants of PepN replacing E264 (GAMEN motif) and E298 (HEXXH motif) were generated. There are no major structural changes between purified wild type (WT) and mutant proteins, which are catalytically inactive. Importantly, growth profiles of Delta pepN upon expression of WT or mutant proteins demonstrated the importance of catalytic activity during NDHT but not NaSal-induced stress. Further fluorescamine reactivity studies demonstrated that the catalytic activity of PepN is required to generate higher intracellular amounts of free N-terminal amino acids; consequently, the lower growth of Delta pepN during NDHT stress increases with high amounts of casamino acids. Together, this study sheds insights on the expression and functional roles of the catalytic activity of PepN during adaptation to NDHT stress. (C) 2012 Elsevier GmbH. All rights reserved.

Differential cellular recognition pattern to M. tuberculosis targets defined by IFN-gamma and IL-17 production in blood from TB plus patients from Honduras as compared to health care workers: TB and immune responses in patients from Honduras

Background: A better understanding of the quality of cellular immune responses directed against molecularly defined targets will guide the development of TB diagnostics and identification of molecularly defined, clinically relevant M.tb vaccine candidates. Methods: Recombinant proteins (n = 8) and peptide pools (n = 14) from M. tuberculosis (M.tb) targets were used to compare cellular immune responses defined by IFN-gamma and IL-17 production using a Whole Blood Assay (WBA) in a cohort of 148 individuals, i.e. patients with TB + (n = 38), TB- individuals with other pulmonary diseases (n = 81) and individuals exposed to TB without evidence of clinical TB (health care workers, n = 29). Results: M.tb antigens Rv2958c (glycosyltransferase), Rv2962c (mycolyltransferase), Rv1886c (Ag85B), Rv3804c (Ag85A), and the PPE family member Rv3347c were frequently recognized, defined by IFN-gamma production, in blood from healthy individuals exposed to M.tb (health care workers). A different recognition pattern was found for IL-17 production in blood from M.tb exposed individuals responding to TB10.4 (Rv0288), Ag85B (Rv1886c) and the PPE family members Rv0978c and Rv1917c. Conclusions: The pattern of immune target recognition is different in regard to IFN-gamma and IL-17 production to defined molecular M.tb targets in PBMCs from individuals frequently exposed to M.tb. The data represent the first mapping of cellular immune responses against M.tb targets in TB patients from Honduras.

SIRT6 Promotes COX-2 Expression and Acts as an Oncogene in Skin Cancer

SIRT6 is a SIR2 family member that regulates multiple molecular pathways involved in metabolism, genomic stability, and aging. It has been proposed previously that SIRT6 is a tumor suppressor in cancer. Here, we challenge this concept by presenting evidence that skin-specific deletion of SIRT6 in the mouse inhibits skin tumorigenesis. SIRT6 promoted expression of COX-2 by repressing AMPK signaling, thereby increasing cell proliferation and survival in the skin epidermis. SIRT6 expression in skin keratinocytes was increased by exposure to UVB light through activation of the AKT pathway. Clinically, we found that SIRT6 was upregulated in human skin squamous cell carcinoma. Taken together, our results provide evidence that SIRT6 functions as an oncogene in the epidermis and suggest greater complexity to its role in epithelial carcinogenesis. (C) 2014 AACR.

Lynx1 and the β2V287L mutation affect the stoichiometry of the α4β2 nicotinic acetylcholine receptor

GPI-anchored neurotoxin-like receptor binding proteins, such as lynx modulators, are topologically positioned to exert pharmacological effects by binding to the extracellular portion of nAChRs. These actions are generally thought to proceed when both lynx and the nAChRs are on the plasma membrane. Here, we demonstrate that lynx1 also exerts effects on α4β2 nAChRs within the endoplasmic reticulum. Lynx affects assembly of nascent α4 and β2 subunits, and alters the stoichiometry of the population that reaches the plasma membrane. Additionally, these data suggest that lynx1 alters nAChR stoichiometry primarily through this intracellular interaction, rather than via effects on plasma membrane nAChRs. To our knowledge, these data represent the first test of the hypothesis that a lynx family member, or indeed any GPI-anchored protein, could act within the cell to alter assembly of multi-subunit protein.

Representações sociais do câncer para o familiar do paciente oncológico em tratamento quimioterápico

Considerando-se que a família vivencia e partilha junto com o doente todos os sentimentos, emoções e angústias que envolve o diagnóstico e o tratamento do câncer, o presente estudo teve como objetivos descrever as dimensões das representações sociais acerca do câncer para familiares do cliente oncológico em tratamento quimioterápico ambulatorial em uma unidade de referência para o seu tratamento; analisar a representação social do câncer elaborada por familiares do cliente oncológico em tratamento quimioterápico ambulatorial; e discutir as contribuições do enfermeiro junto à família do cliente oncológico em tratamento quimioterápico ambulatorial a partir da construção representacional do câncer para os sujeitos do estudo. De caráter qualitativo, o caminho metodológico foi construído com base na Teoria das Representações Sociais. Os sujeitos foram 30 familiares que estavam acompanhando o doente durante o tratamento quimioterápico. Os dados foram coletados a partir da realização de entrevistas semi-estruturadas e analisados através da análise de conteúdo de Bardin (1979), sistematizada por Oliveira (2008), com o auxílio do software QRS Nvivo 2.0. Da análise dos dados emergiram seis categorias que compõem o campo representacional e se expressa através das dimensões representacionais concretizadas nas seguintes categorias: sentimentos compartilhados por familiares de clientes oncológicos em tratamento quimioterápico, que mostra que, ao se depararem com a doença e sua dura realidade, os familiares são acometidos por diversos tipos de sentimentos; imagens, metáforas e conceitos no existir da família que enfrenta a doença, onde os familiares revelaram que o câncer é percebido, entre outras coisas, como um monstro que invade a vida das pessoas e dela passa a tomar conta e a dominá-la; preconceitos e estigmas na vivência do câncer, que revelou que ainda hoje existem representações e estigmas presentes na sociedade e em suas construções culturais acerca do câncer; diferentes práticas desenvolvidas no contexto da doença e do processo de adoecimento pelo câncer, que evidenciou as diferentes práticas presentes no discurso dos sujeitos, quais sejam, a de religiosidade no contexto do câncer, a de enfrentamento da doença, a de comunicação-ocultamento e de atitudes da família ao estar no mundo frente ao câncer; o processo de ancoragem e o conhecimento adquirido após a experiência do câncer, onde surgiram os conhecimentos que os sujeitos adquiriram acerca do câncer e alguns elementos do processo de ancoragem do câncer; as vivências do enfermeiro que trabalha em oncologia e suas contribuições junto à família que alerta os enfermeiros para a necessidade de intervenções efetivas direcionadas à assistência integral do indivíduo, levando em consideração a importância da família. Conclui-se que ao se descobrir acompanhando um familiar que tem câncer, a família passa a viver um outro mundo, no qual a possibilidade de morte se mostra de forma inevitável e iminente. Diante disso, a família passa a valorizar não apenas o cuidado dispensado ao doente, mas também anseia por uma atenção profissional que contemple seu existir e seu modo de viver.

Assistência paliativa em oncologia na perspectiva do familiar: contribuições da enfermagem

O estudo emergiu da minha experiência profissional como enfermeira de um serviço especializado em cuidados paliativos em oncologia. A abordagem paliativa exige da equipe maior interação, não apenas na realização de procedimentos terapêuticos necessários naquele momento, mas, sobretudo, em orientar adequadamente, quanto aos cuidados realizados com pacientes que se encontram em estado de doença avançada, bem como promover a adesão terapêutica do familiar, através de ações interdisciplinares. Estudo de natureza qualitativa com referencial teórico metodológico na fenomenologia sociológica de Alfred Schutz, que tem como enfoque o significado da ação. Nesse sentido, as vivências e as ações dos familiares constituem fontes de significados subjetivos das experiências adquiridas, ao cuidar de um familiar em tratamento paliativo em oncologia. A partir dessas reflexões, o objeto de estudo é a experiência vivida pela família em cuidar de pessoa em tratamento paliativo em oncologia e tem como objetivo compreender a perspectiva de cuidar do familiar de pessoa em tratamento paliativo em oncologia. O cenário onde foi realizado o estudo é o Instituto Nacional de Câncer/MS, situado no município do Rio de Janeiro, na Unidade de Cuidados Paliativos/HCIV. Os sujeitos participantes foram, 20 familiares de pacientes em tratamento paliativo em oncologia. A apreensão das falas, deu-se mediante entrevista fenomenológica, guiada por meio da questão orientadora: como você está vivendo a experiência de cuidar de seu familiar em tratamento paliativo em oncologia? O estudo permitiu compreender que o familiar de pessoa em tratamento paliativo em oncologia se mostrou como aquele que ajuda, apóia, estando junto, não abandonando, dando apoio e representando a família, na perspectiva de enfrentar suas próprias dificuldades diante do tratamento paliativo em oncologia e tentar entender a abordagem paliativa que tem como foco reduzir a dor, superando o rótulo de terminal. Portanto, oferecer uma forma de compreender o cuidado desse familiar, situado no mundo institucional, possibilitando apreender o vivido e os significados que alicerçam esta prática conduz a um comportamento, ações e relações, ou seja, um modo de pensar que oriente o individuo no seu cotidiano para ser e estar com o outro nessa fase da vida. Nesse sentido, a escuta e a comunicação se constituem como os pilares do cuidar dos familiares em cuidados paliativos em oncologia. A inserção da família durante todo o processo é fundamental para os cuidados realizados com a pessoa, evidenciando para o enfermeiro a importância de atender às expectativas de ambos, agindo como facilitador na implementação do cuidar. Assim sendo, as ações de enfermagem podem contribuir e auxiliar a família a descobrir suas próprias soluções para as situações que envolvem o suporte às necessidades apresentadas.

A vida após a alta

Esta tese discute a vida após a alta hospitalar através de uma etnografia da experiência de um grupo de famílias, ao cuidarem de um membro dependente de cuidados de saúde. Foi realizada entre agosto de 2001 e julho de 2005 em Juiz de Fora-MG. O foco da investigação foi a convivência cotidiana da família com um membro requerendo cuidados de saúde especializados após ter recebido assistência de alta complexidade. O objetivo foi compreender a questão: como a família cuida, em casa, de um familiar que necessita de cuidados de saúde após a alta hospitalar? Adotamos a etnografia orientada por Geertz (1989), que nos permitiu, através da análise interpretativa das teias de significados apreendidas pela observaçãoconvivência com os sujeitos, uma compreensão de como o fenômeno (cuidado) se evidencia e se transforma em experiência nas relações que se estabelecem dentro e fora da família. A identificação das famílias-sujeito iniciou com a observação das internações na Unidade de Terapia Intensiva (UTI) do Hospital Universitário da Universidade Federal de Juiz de Fora, no segundo semestre de 2003, seguida da observação nas enfermarias e, posteriormente, nos seus domicílios. De 137 pacientes internados nesta UTI naquele semestre, 59 foram a óbito, 12 foram transferidos para outro hospital da cidade, por demandarem tecnologias não oferecidas pela instituição e 66 tiveram alta hospitalar na condição de melhorado. Destes, observamos 12 casos, residentes na cidade de Juiz de Fora, que compartilham de uma mesma cultura assistencial e que foram submetidas a um mesmo padrão de organização e de fluxo de atendimento no sistema de saúde local. Destas, uma família se destacou como principal sujeito, pela repetição de eventos significativos à questão principal da pesquisa e utilizamos dados de outras cinco dentre as observadas. Os dados foram arquivados em um banco de dados qualitativos LOGOS. A prioridade nos cuidados com o corpo; a dependência dos serviços de saúde especializados do SUS e as alterações na organização e no funcionamento da família, com redefinições de papéis, para se adaptar à realidade de convivência com um membro doente, estão dentre os achados. Dois fenômenos que se relacionam com o desenvolvimento da experiência de cuidar pelas famílias se destacaram: a individualização na família, que parece influenciar a forma de abordagem clínica (individualizada) pelos profissionais de saúde, e uma concepção de família como sujeito coletivo Bourdieu (1998) como possibilidade para o planejamento de ações coletivas. O sofrimento, observado pela contínua convivência dos sujeitos com sentimentos de angústia nas trajetórias de busca de cuidados no Sistema, nas instituições de saúde, evidenciou a desassistência a que esse grupo de cidadãos está exposto no modelo assistencial vigente. A lida das famílias com uma diversidade de cuidados, incluindo a prática de cuidados técnicos desencadeou uma rede extrafamiliar de aproximações, para o enfrentamento das necessidades. Apesar da dependência de tecnologias, de saberes técnicos e das restrições no acesso a esses, evidenciou-se um tipo de autonomia pelos sujeitos na prática de cuidados no espaço intrafamiliar e no entorno micro-sociológico de convivência. No Sistema de Saúde a preferência primeira das famílias para busca de ajuda é o hospital, depois, as unidades de referência secundária e, por último, as Equipes de Saúde da Família (ESF). A procura pelas ESF é por que estas representam parte obrigatória no fluxo inicial dos usuários do SUS local, garantem a aquisição de medicamentos, oferecem serviços de natureza cartorial, como atestados e pareceres para juiz e, ainda, encaminhamentos e solicitação de exames.

Benefício de prestação continuada: desafios e questões atuais

A presente Tese busca refletir sobre as características e o significado do Benefício de Prestação Continuada (BPC) como componente da política de Assistência Social. Este benefício constitui o principal direito da Assistência Social, pois é o único garantido constitucionalmente e garante o pagamento de um salário mínimo mensal a idosos com 65 anos ou mais e pessoas com deficiência, cuja renda mensal familiar seja inferior a do salário mínimo por pessoa. A Tese procura analisar, portanto, as mudanças propostas, os resultados alcançados e os desafios que se colocam para o BPC no município do Rio de Janeiro, em face das novas perspectivas apresentadas para este benefício a partir de 2005 pelo Sistema Único de Assistência Social (SUAS) e, posteriormente, pelo Decreto 6.214/2007. Para a efetivação dessa proposta foi realizada uma pesquisa que procurou compreender a situação e as características atuais do desenvolvimento deste Benefício, no município em apreço, no âmbito da política de Assistência Social e do Instituto Nacional de Seguro Social (INSS). O estudo desenvolvido pela Tese revelou, entre outros aspectos, que apesar do intenso movimento, em termos de proposições, que se processa na esfera federal da política de Assistência Social com vistas a encaminhar as mudanças previstas para o BPC, este esforço não tem o respaldo político necessário que dê impulso a essas propostas e crie condições efetivas para elas se materializarem, principalmente na esfera municipal, que é onde a política, de fato, se concretiza. Assim, no período analisado, o ano de 2010, mantinha-se a concepção do BPC como um fator externo à política municipal de Assistência Social do Rio de Janeiro. Na realidade, essa situação reflete o histórico distanciamento da Assistência Social em relação a este benefício, que apesar de ter sido fundamental para a institucionalização desta política como uma das áreas componentes da Seguridade Social, continua, na atualidade, a não ter uma real identidade com a Assistência Social.

Influence of specific HSP70 domains on fibril formation of the yeast prion protein Ure2.

Ure2p is the protein determinant of the Saccharomyces cerevisiae prion state [URE3]. Constitutive overexpression of the HSP70 family member SSA1 cures cells of [URE3]. Here, we show that Ssa1p increases the lag time of Ure2p fibril formation in vitro in the presence or absence of nucleotide. The presence of the HSP40 co-chaperone Ydj1p has an additive effect on the inhibition of Ure2p fibril formation, whereas the Ydj1p H34Q mutant shows reduced inhibition alone and in combination with Ssa1p. In order to investigate the structural basis of these effects, we constructed and tested an Ssa1p mutant lacking the ATPase domain, as well as a series of C-terminal truncation mutants. The results indicate that Ssa1p can bind to Ure2p and delay fibril formation even in the absence of the ATPase domain, but interaction of Ure2p with the substrate-binding domain is strongly influenced by the C-terminal lid region. Dynamic light scattering, quartz crystal microbalance assays, pull-down assays and kinetic analysis indicate that Ssa1p interacts with both native Ure2p and fibril seeds, and reduces the rate of Ure2p fibril elongation in a concentration-dependent manner. These results provide new insights into the structural and mechanistic basis for inhibition of Ure2p fibril formation by Ssa1p and Ydj1p.