Uso do protocolo no serviço de psicologia e avaliação psicodinâmica em candidatos à cirurgia bariátrica

A clínica da Obesidade Mórbida e a Cirurgia Bariátrica exige estudos e acompanhamentos do paciente. Os benefícios e riscos do emagrecimento por tratamento cirúrgico devem servir como ponto de alerta aos profissionais da saúde. O uso do questionário no serviço de psicologia é norteado pela escuta psicológica. Objetivos: 1) Descrever o perfil sócio-demográfico candidatos à cirurgia bariátrica. 2) Analisar a percepção dos pacientes sobre características de personalidade associadas à obesidade e transtornos alimentares. 3) Descrever os conteúdos psicodinâmicos da narrativa do sujeito e avaliar o sistema tensional inconsciente de dois pacientes por meio do Teste das Relações Objetais de Phillipson (TRO). Método: O delineamento metodológico com análise de dados pelo método epidemiológico e estudo de caso clínico, orientação psicanalítica. Na primeira etapa foram consultados 300 questionários do serviço de psicologia e na segunda dois pacientes com ganho de peso após 24 meses. São pacientes que procuraram tratamento em clínica especializada, em uma metrópole do sudeste brasileiro, sob consentimento pós-informado. Os questionários foram preenchidos por 227 mulheres e 73 homens; com média de idade igual a 36 anos; escolaridade ensino médio e superior, 53%; maioria casados; IMC entre grave e super mórbido (94,3%). Técnicas cirúrgicas indicadas Capella Bypass e Fobi-Capella (67%). Resultados: características psicológicas referidas pelos pacientes, a ansiedade apontou em 93,7% das respostas, seguidas por impulsividade, depressão, tolerância à frustração, baixa auto-estima, resolvedor de problemas dos outros (mais de 50%). No histórico familiar da obesidade está em mais de 70% depressão e uso do álcool em 30%; realização de psicoterapia (30%) e medicamentos para depressão e ansiedade (10%). Na segunda etapa, foi realizado o diagnóstico psicodinâmico, por meio do Teste das Relações Objetais de Phillipson com duas pacientes, cuja análise indicou necessidade de psicoterapia psicanalítica, pois tinham fixações na posição esquizoparanóide e apresentavam dificuldade em lidar com perdas e baixa motivação para mudança e insigth. Conclusões: Com a aplicação do questionário e o registro das observações empíricas, este questionário de entrevista semidirigida preenche condições de melhor acessar e avaliar os conteúdos revelados pelos pacientes. As contradições entre as respostas e o discurso, no contato individual com o psicólogo, apontam a necessidade de investimento no preparo do paciente para a cirurgia e mais acentuadamente o acompanhamento psicológico no primeiro ano do pós-operatório. Há um pensamento mágico a ser trabalhado durante a aplicação do questionário sobre as crenças frente à cirurgia e o emagrecimento e assim convocar o paciente a ocupar o lugar do sujeito implicado em seu processo pré e pós-operatório. O TRO contribuiu na compreensão do diagnóstico psicodinâmico de pacientes com ganho de peso após cirurgia e reforçou a necessidade de maior investimento no pré-operatório.(AU)

Uso do protocolo no serviço de psicologia e avaliação psicodinâmica em candidatos à cirurgia bariátrica

A clínica da Obesidade Mórbida e a Cirurgia Bariátrica exige estudos e acompanhamentos do paciente. Os benefícios e riscos do emagrecimento por tratamento cirúrgico devem servir como ponto de alerta aos profissionais da saúde. O uso do questionário no serviço de psicologia é norteado pela escuta psicológica. Objetivos: 1) Descrever o perfil sócio-demográfico candidatos à cirurgia bariátrica. 2) Analisar a percepção dos pacientes sobre características de personalidade associadas à obesidade e transtornos alimentares. 3) Descrever os conteúdos psicodinâmicos da narrativa do sujeito e avaliar o sistema tensional inconsciente de dois pacientes por meio do Teste das Relações Objetais de Phillipson (TRO). Método: O delineamento metodológico com análise de dados pelo método epidemiológico e estudo de caso clínico, orientação psicanalítica. Na primeira etapa foram consultados 300 questionários do serviço de psicologia e na segunda dois pacientes com ganho de peso após 24 meses. São pacientes que procuraram tratamento em clínica especializada, em uma metrópole do sudeste brasileiro, sob consentimento pós-informado. Os questionários foram preenchidos por 227 mulheres e 73 homens; com média de idade igual a 36 anos; escolaridade ensino médio e superior, 53%; maioria casados; IMC entre grave e super mórbido (94,3%). Técnicas cirúrgicas indicadas Capella Bypass e Fobi-Capella (67%). Resultados: características psicológicas referidas pelos pacientes, a ansiedade apontou em 93,7% das respostas, seguidas por impulsividade, depressão, tolerância à frustração, baixa auto-estima, resolvedor de problemas dos outros (mais de 50%). No histórico familiar da obesidade está em mais de 70% depressão e uso do álcool em 30%; realização de psicoterapia (30%) e medicamentos para depressão e ansiedade (10%). Na segunda etapa, foi realizado o diagnóstico psicodinâmico, por meio do Teste das Relações Objetais de Phillipson com duas pacientes, cuja análise indicou necessidade de psicoterapia psicanalítica, pois tinham fixações na posição esquizoparanóide e apresentavam dificuldade em lidar com perdas e baixa motivação para mudança e insigth. Conclusões: Com a aplicação do questionário e o registro das observações empíricas, este questionário de entrevista semidirigida preenche condições de melhor acessar e avaliar os conteúdos revelados pelos pacientes. As contradições entre as respostas e o discurso, no contato individual com o psicólogo, apontam a necessidade de investimento no preparo do paciente para a cirurgia e mais acentuadamente o acompanhamento psicológico no primeiro ano do pós-operatório. Há um pensamento mágico a ser trabalhado durante a aplicação do questionário sobre as crenças frente à cirurgia e o emagrecimento e assim convocar o paciente a ocupar o lugar do sujeito implicado em seu processo pré e pós-operatório. O TRO contribuiu na compreensão do diagnóstico psicodinâmico de pacientes com ganho de peso após cirurgia e reforçou a necessidade de maior investimento no pré-operatório.(AU)

Phospholipase D activity is required for suppression of yeast phosphatidylinositol transfer protein defects

Yeast phosphatidylinositol transfer protein (Sec14p) function is essential for production of Golgi-derived secretory vesicles, and this requirement is bypassed by mutations in at least seven genes. Analyses of such ‘bypass Sec14p’ mutants suggest that Sec14p acts to maintain an essential Golgi membrane diacylglycerol (DAG) pool that somehow acts to promote Golgi secretory function. SPO14 encodes the sole yeast phosphatidylinositol-4,5-bisphosphate-activated phospholipase D (PLD). PLD function, while essential for meiosis, is dispensable for vegetative growth. Herein, we report specific physiological circumstances under which an unanticipated requirement for PLD activity in yeast vegetative Golgi secretory function is revealed. This PLD involvement is essential in ‘bypass Sec14p’ mutants where normally Sec14p-dependent Golgi secretory reactions are occurring in a Sec14p-independent manner. PLD catalytic activity is necessary but not sufficient for ‘bypass Sec14p’, and yeast operating under ‘bypass Sec14p’ conditions are ethanol-sensitive. These data suggest that PLD supports ‘bypass Sec14p’ by generating a phosphatidic acid pool that is somehow utilized in supporting yeast Golgi secretory function.

Loss of heterozygosity induced by a chromosomal double-strand break

The repair of chromosomal double-strand breaks (DSBs) is necessary for genomic integrity in all organisms. Genetic consequences of misrepair include chromosomal loss, deletion, and duplication resulting in loss of heterozygosity (LOH), a common finding in human solid tumors. Although work with radiation-sensitive cell lines suggests that mammalian cells primarily rejoin DSBs by nonhomologous mechanisms, alternative mechanisms that are implicated in chromosomal LOH, such as allelic recombination, may also occur. We have examined chromosomal DSB repair between homologs in a gene targeted mammalian cell line at the retinoblastoma (Rb) locus. We have found that allelic recombinational repair occurs in mammalian cells and is increased at least two orders of magnitude by the induction of a chromosomal DSB. One consequence of allelic recombination is LOH at the Rb locus. Some of the repair events also resulted in other types of genetic instability, including deletions and duplications. We speculate that mammalian cells may have developed efficient nonhomologous DSB repair processes to bypass allelic recombination and the potential for reduction to homozygosity.

A novel bacterial tyrosine kinase essential for cell division and differentiation

Protein kinases play central roles in the regulation of eukaryotic and prokaryotic cell growth, division, and differentiation. The Caulobacter crescentus divL gene encodes a novel bacterial tyrosine kinase essential for cell viability and division. Although the DivL protein is homologous to the ubiquitous bacterial histidine protein kinases (HPKs), it differs from previously studied members of this protein kinase family in that it contains a tyrosine residue (Tyr-550) in the conserved H-box instead of a histidine residue, which is the expected site of autophosphorylation. DivL is autophosphorylated on Tyr-550 in vitro, and this tyrosine residue is essential for cell viability and regulation of the cell division cycle. Purified DivL also catalyzes phosphorylation of CtrA and activates transcription in vitro of the cell cycle-regulated fliF promoter. Suppressor mutations in ctrA bypass the conditional cell division phenotype of cold-sensitive divL mutants, providing genetic evidence that DivL function in cell cycle and developmental regulation is mediated, at least in part, by the global response regulator CtrA. DivL is the only reported HPK homologue whose function has been shown to require autophosphorylation on a tyrosine, and, thus, it represents a new class of kinases within this superfamily of protein kinases.

An alternative pathway for rod signals in the rodent retina: Rod photoreceptors, cone bipolar cells, and the localization of glutamate receptors

In the mammalian retina, extensive processing of spatiotemporal and chromatic information occurs. One key principle in signal transfer through the retina is parallel processing. Two of these parallel pathways are the ON- and OFF-channels transmitting light and dark signals. This dual system is created in the outer plexiform layer, the first relay station in retinal signal transfer. Photoreceptors release glutamate onto ON- and OFF-type bipolar cells, which are functionally distinguished by their postsynaptic expression of different types of glutamate receptors, namely ionotropic and metabotropic glutamate receptors. In the current concept, rod photoreceptors connect only to rod bipolar cells (ON-type) and cone photoreceptors connect only to cone bipolar cells (ON- and OFF-type). We have studied the distribution of (RS)-α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor subunits at the synapses in the outer plexiform layer of the rodent retina by immunoelectron microscopy and serial section reconstruction. We report a non-classical synaptic contact and an alternative pathway for rod signals in the retina. Rod photoreceptors made synaptic contact with putative OFF-cone bipolar cells that expressed the AMPA glutamate receptor subunits GluR1 and GluR2 on their dendrites. Thus, in the retina of mouse and rat, an alternative pathway for rod signals exists, where rod photoreceptors bypass the rod bipolar cell and directly excite OFF-cone bipolar cells through an ionotropic sign-conserving AMPA glutamate receptor.

The β subunit sliding DNA clamp is responsible for unassisted mutagenic translesion replication by DNA polymerase III holoenzyme

The replication of damaged nucleotides that have escaped DNA repair leads to the formation of mutations caused by misincorporation opposite the lesion. In Escherichia coli, this process is under tight regulation of the SOS stress response and is carried out by DNA polymerase III in a process that involves also the RecA, UmuD′ and UmuC proteins. We have shown that DNA polymerase III holoenzyme is able to replicate, unassisted, through a synthetic abasic site in a gapped duplex plasmid. Here, we show that DNA polymerase III*, a subassembly of DNA polymerase III holoenzyme lacking the β subunit, is blocked very effectively by the synthetic abasic site in the same DNA substrate. Addition of the β subunit caused a dramatic increase of at least 28-fold in the ability of the polymerase to perform translesion replication, reaching 52% bypass in 5 min. When the ssDNA region in the gapped plasmid was extended from 22 nucleotides to 350 nucleotides, translesion replication still depended on the β subunit, but it was reduced by 80%. DNA sequence analysis of translesion replication products revealed mostly −1 frameshifts. This mutation type is changed to base substitution by the addition of UmuD′, UmuC, and RecA, as demonstrated in a reconstituted SOS translesion replication reaction. These results indicate that the β subunit sliding DNA clamp is the major determinant in the ability of DNA polymerase III holoenzyme to perform unassisted translesion replication and that this unassisted bypass produces primarily frameshifts.

Cyclin E, a redundant cyclin in breast cancer

Cyclin E is an important regulator of cell cycle progression that together with cyclin-dependent kinase (cdk) 2 is crucial for the G1/S transition during the mammalian cell cycle. Previously, we showed that severe overexpression of cyclin E protein in tumor cells and tissues results in the appearance of lower molecular weight isoforms of cyclin E, which together with cdk2 can form a kinase complex active throughout the cell cycle. In this study, we report that one of the substrates of this constitutively active cyclin E/cdk2 complex is retinoblastoma susceptibility gene product (pRb) in populations of breast cancer cells and tissues that also overexpress p16. In these tumor cells and tissues, we show that the expression of p16 and pRb is not mutually exclusive. Overexpression of p16 in these cells results in sequestering of cdk4 and cdk6, rendering cyclin D1/cdk complexes inactive. However, pRb appears to be phosphorylated throughout the cell cycle following an initial lag, revealing a time course similar to phosphorylation of glutathione S-transferase retinoblastoma by cyclin E immunoprecipitates prepared from these synchronized cells. Hence, cyclin E kinase complexes can function redundantly and replace the loss of cyclin D-dependent kinase complexes that functionally inactivate pRb. In addition, the constitutively overexpressed cyclin E is also the predominant cyclin found in p107/E2F complexes throughout the tumor, but not the normal, cell cycle. These observations suggest that overexpression of cyclin E in tumor cells, which also overexpress p16, can bypass the cyclin D/cdk4-cdk6/p16/pRb feedback loop, providing yet another mechanism by which tumors can gain a growth advantage.

Pressure-mediated oligonucleotide transfection of rat and human cardiovascular tissues

The application of gene therapy to human disease is currently restricted by the relatively low efficiency and potential hazards of methods of oligonucleotide or gene delivery. Antisense or transcription factor decoy oligonucleotides have been shown to be effective at altering gene expression in cell culture expreriments, but their in vivo application is limited by the efficiency of cellular delivery, the intracellular stability of the compounds, and their duration of activity. We report herein the development of a highly efficient method for naked oligodeoxynucleotide (ODN) transfection into cardiovascular tissues by using controlled, nondistending pressure without the use of viral vectors, lipid formulations, or exposure to other adjunctive, potentially hazardous substances. In this study, we have documented the ability of ex vivo, pressure-mediated transfection to achieve nuclear localization of fluorescent (FITC)-labeled ODN in approximately 90% and 50% of cells in intact human saphenous vein and rat myocardium, respectively. We have further documented that pressure-mediated delivery of antisense ODN can functionally inhibit target gene expression in both of these tissues in a sequence-specific manner at the mRNA and protein levels. This oligonucleotide transfection system may represent a safe means of achieving the intraoperative genetic engineering of failure-resistant human bypass grafts and may provide an avenue for the genetic manipultation of cardiac allograft rejection, allograft vasculopathy, or other transplant diseases.

Rapid restoration of visual pigment and function with oral retinoid in a mouse model of childhood blindness

Mutations in the retinal pigment epithelium gene encoding RPE65 are a cause of the incurable early-onset recessive human retinal degenerations known as Leber congenital amaurosis. Rpe65-deficient mice, a model of Leber congenital amaurosis, have no rod photopigment and severely impaired rod physiology. We analyzed retinoid flow in this model and then intervened by using oral 9-cis-retinal, attempting to bypass the biochemical block caused by the genetic abnormality. Within 48 h, there was formation of rod photopigment and dramatic improvement in rod physiology, thus demonstrating that mechanism-based pharmacological intervention has the potential to restore vision in otherwise incurable genetic retinal degenerations.

Response of human DNA polymerase ι to DNA lesions

Lesion bypass is an important mechanism to overcome replication blockage by DNA damage. Translesion synthesis requires a DNA polymerase (Pol). Human Pol ι encoded by the RAD30B gene is a recently identified DNA polymerase that shares sequence similarity to Pol η. To investigate whether human Pol ι plays a role in lesion bypass we examined the response of this polymerase to several types of DNA damage in vitro. Surprisingly, 8-oxoguanine significantly blocked human Pol ι. Nevertheless, translesion DNA synthesis opposite 8-oxoguanine was observed with increasing concentrations of purified human Pol ι, resulting in predominant C and less frequent A incorporation opposite the lesion. Opposite a template abasic site human Pol ι efficiently incorporated a G, less frequently a T and even less frequently an A. Opposite an AAF-adducted guanine, human Pol ι was able to incorporate predominantly a C. In both cases, however, further DNA synthesis was not observed. Purified human Pol ι responded to a template TT (6–4) photoproduct by inserting predominantly an A opposite the 3′ T of the lesion before aborting DNA synthesis. In contrast, human Pol ι was largely unresponsive to a template TT cis-syn cyclobutane dimer. These results suggest a role for human Pol ι in DNA lesion bypass.

RECODE: a database of frameshifting, bypassing and codon redefinition utilized for gene expression

The RECODE database is a compilation of ‘programmed’ translational recoding events taken from the scientific literature and personal communications. The database deals with programmed ribosomal frameshifting, codon redefinition and translational bypass occurring in a variety of organisms. The entries for each event include the sequences of the corresponding genes, their encoded proteins for both the normal and alternate decoding, the types of the recoding events involved, trans-factors and cis-elements that influence recoding. The database is freely available at http://recode.genetics.utah.edu/.

Evidence for an Intrinsic Toxicity of Phosphatidylcholine to Sec14p-dependent Protein Transport from the Yeast Golgi Complex

Yeast phosphatidylinositol-transfer protein (Sec14p) is essential for Golgi secretory function and cell viability. This requirement of Sec14p is relieved by genetic inactivation of the cytidine diphosphate-choline pathway for phosphatidycholine (PtdCho) biosynthesis. Standard phenotypic analyses indicate that inactivation of the phosphatidylethanolamine (PtdEtn) pathway for PtdCho biosynthesis, however, does not rescue the growth and secretory defects associated with Sec14p deficiency. We now report inhibition of choline uptake from the media reveals an efficient “bypass Sec14p” phenotype associated with PtdEtn-methylation pathway defects. We further show that the bypass Sec14p phenotype associated with PtdEtn-methylation pathway defects resembles other bypass Sec14p mutations in its dependence on phospholipase D activity. Finally, we find that increased dosage of enzymes that catalyze phospholipase D-independent turnover of PtdCho, via mechanisms that do not result in a direct production of phosphatidic acid or diacylglycerol, effect a partial rescue of sec14-1ts-associated growth defects. Taken together, these data support the idea that PtdCho is intrinsically toxic to yeast Golgi secretory function.

Human testis expresses a specific poly(A)-binding protein

In testis mRNA stability and translation initiation are extensively under the control of poly(A)-binding proteins (PABP). Here we have cloned a new human testis-specific PABP (PABP3) of 631 amino acids (70.1 kDa) with 92.5% identical residues to the ubiquitous PABP1. A northern blot of multiple human tissues hybridised with PABP3- and PABP1-specific oligonucleotide probes revealed two PABP3 mRNAs (2.1 and 2.5 kb) detected only in testis, whereas PABP1 mRNA (3.2 kb) was present in all tested tissues. In human adult testis, PABP3 mRNA expression was restricted to round spermatids, whereas PABP1 was expressed in these cells as well as in pachytene spermatocytes. PABP3-specific antibodies identified a protein of 70 kDa in human testis extracts. This protein binds poly(A) with a slightly lower affinity as compared to PABP1. The human PABP3 gene is intronless with a transcription start site 61 nt upstream from the initiation codon. A sequence of 256 bp upstream from the transcription start site drives the promoter activity of PABP3 and its tissue-specific expression. The expression of PABP3 might be a way to bypass PABP1 translational repression and to produce the amount of PABP needed for active mRNA translation in spermatids.