Clumsy straight paths: Notes on prison segregation of the lawbreaker with mental suffering

The horror of the reality found in mental hospitals, lack of care, treatment and rights. Simultaneously, the society exposes its prediction, for acts performed as spectacular. Hence it requires the State to make disappear these messengers of the worst, which are devoid of their history, recognition of their human and social complexity. The silent asking for help of this citizen, his case was not heard by the same society. The anguish, suffering and inconvenience direct the individual to perform the "barbaric" act. There is, in this context, lack of assistance to the scream of despair, disregard and disrespect implicit in the model of health care, which becomes, then, the problem of the judiciary. Thus, he should be presented as a "psychopath", irretrievable from madness and for social life, and there is no other way, but his containment and exclusion of social order. To the state remains to discuss terms: Responsibility or Nonimputability? Equality and difference. Still, to live with the dichotomy that the law is influenced by the anti-asylum movement.O horror da realidade encontrada nos manicômios, ausência de cuidados, tratamento e de direitos. Simultaneamente, a sociedade expõe seus agouros, por atos praticados como espetaculares. Daí exige-se do Estado que faça desaparecer com estes mensageiros do pior, os quais são desprovidos de sua historicidade, reconhecimento da sua complexidade humana e social. O pedindo silencioso de socorro deste cidadão, ao seu modo, não foi ouvido por esta mesma sociedade. A angústia, o sofrimento e o transtorno direcionam o indivíduo ao cometimento de ato "bárbaro". Há, neste contexto, ausência de assistência ao grito de desespero, a desconsideração, descaso e desrespeito implícito no modelo de assistência à saúde, que se torna, então, em problema do judiciário. Resta, então, apresentá-lo como um "psicopata", irrecuperável da loucura e do convívio social, não restando outra saída, senão a sua contensão e exclusão da ordem social. Ao Estado, resta discutir termos: Responsabilidade ou Inimputabilidade? Igualdade e diferença. Ainda, conviver com a dicotomia de que a legislação é influenciada pelo Movimento da Luta Antimanicomial

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

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Chemiluminescent determination of leukocyte alkaline phosphatase: An advantageous alternative to the cytochemical assay

The determination of leukocyte alkaline phosphatasd (LAP) is used as an aid to diagnose many diseases in the laboratory. For example, it can be used to distinguish chronic myeloid leukemia (CML) from other myeloproliferative disorders (particularly myelofibrosis and polycythemia) and leukemoid reactions (LR). Traditionally, this test is performed with the use of subjective cytochemical assays that assign a score to the level of LAP. Here we present a nonsubjective, quantitative, sensitive, and inexpensive chemiluminescent technique that determines LAP based on the commercial reagent Immulite (R) (AMPPD). To validate this methodology, intact leukocytes obtained from 32 healthy subjects, nine CML patients, and nine LR patients were submitted to the optimized protocol. By measuring the light emission elicited by four concentrations of neutrophils, we were able to estimate the activity of LAP per cell (the slope of the curve obtained by linear regression). A high linear correlation was found between the chemiluminescent result (slope) and the cytochemical score. The slope for healthy individuals ranged between 0.61 and 8.49 (10(-5) mV.s/cell), with a median of 2.04 (10(-5) mV.s/cell). These results were statistically different from those of CML patients (range = 0.07-1.75, median = 0.79) and LR patients (range = 3.84-47.24, median 9.58; P < 0.05)

Molecular Components of the <i>Neurospora crassa</i> pH Signaling Pathway and Their Regulation by pH and the PAC-3 Transcription Factor

<div><p>Environmental pH induces a stress response triggering a signaling pathway whose components have been identified and characterized in several fungi. <i>Neurospora crassa</i> shares all six components of the <i>Aspergillus nidulans</i> pH signaling pathway, and we investigate here their regulation during an alkaline pH stress response. We show that the <i>N</i>. <i>crassa pal</i> mutant strains, with the exception of Δ<i>pal-9</i>, which is the <i>A</i>. <i>nidulans palI</i> homolog, exhibit low conidiation and are unable to grow at alkaline pH. Moreover, they accumulate the pigment melanin, most likely via regulation of the tyrosinase gene by the pH signaling components. The PAC-3 transcription factor binds to the tyrosinase promoter and negatively regulates its gene expression. PAC-3 also binds to all <i>pal</i> gene promoters, regulating their expression at normal growth pH and/or alkaline pH, which indicates a feedback regulation of PAC-3 in the <i>pal</i> gene expression. In addition, PAC-3 binds to the <i>pac-3</i> promoter only at alkaline pH, most likely influencing the <i>pac-3</i> expression at this pH suggesting that the activation of PAC-3 in <i>N</i>. <i>crassa</i> results from proteolytic processing and gene expression regulation by the pH signaling components. In <i>N</i>. <i>crassa</i>, PAC-3 is proteolytically processed in a single cleavage step predominately at alkaline pH; however, low levels of the processed protein can be observed at normal growth pH. We also demonstrate that PAC-3 preferentially localizes in the nucleus at alkaline pH stress and that the translocation may require the <i>N</i>. <i>crassa</i> importin-α since the PAC-3 nuclear localization signal (NLS) has a strong <i>in vitro</i> affinity with importin-α. The data presented here show that the pH signaling pathway in <i>N</i>. <i>crassa</i> shares all the components with the <i>A</i>. <i>nidulans</i> and <i>S</i>. <i>cerevisiae</i> pathways; however, it exhibits some properties not previously described in either organism.</p></div

Expression of the tyrosinase gene in the wild-type and the Δ<i>pal</i> mutant strains at normal (5.8) and alkaline (7.8) pH.

<p><b>(A)</b> Mycelial samples from the wild-type and Δ<i>pal</i> mutant strains cultured at pH 5.8 for 24 h and shifted to pH 7.8 for 1 h were used to extract total RNA. Gene expression analysis was performed by RT-qPCR on the StepOnePlus^™ Real-Time PCR system (Applied Biosystems) using Power SYBR^® Green and specific primers. The <i>tub-2</i> gene was used as the reference gene. The asterisks indicate significant differences compared to the wild-type strain at the same pH, and circles indicate significant differences between the same mutant strain cultured at a different pH (Student’s <i>t</i>-test, <i>P</i> < 0.01). <b>(B)</b> Representation of the PAC-3 motifs (black circles) in the tyrosinase gene promoter. Dashed boxes indicate the region analyzed by ChIP-PCR. Genomic DNA samples from the Δ<i>pac-3</i> complemented strain subjected to pH stress or not were immunoprecipitated with an anti-mCherry antibody and subjected to PCR using specific primers. The input DNA (I) was used as the positive control, and the reactions without any antibody (no Ab) were used as the negative control. The intensity of the DNA bands in the gel (left side, arrow) was quantified by ImageJ and the results are shown in the right figure. L, 1 kb DNA ladder. *Asterisks indicate significant differences between the no Ab and IP samples at the same pH (Student’s <i>t</i>-test, <i>P</i> < 0.01). All results represent the average of at least three independent experiments. Bars indicate the standard deviation from the biological experiments.</p

Morphological analyses of the <i>pal</i> mutant strains.

<p><b>(A)</b> The strains (10⁷ conidia/mL) were cultured in Erlenmeyer flasks containing solid VM medium plus 2% sucrose at pH 5.8 for 8–10 days. Basal hyphae growth was examined after cultivating the strains on Petri dishes containing solid VM medium plus 2% sucrose at pH 5.8 and 7.8 for 24 h at 30°C. <b>(B)</b> Radial growth of the colonies measured in cm. The results represent at least two independent experiments. *Indicates significant difference between wild-type and mutant strains at the same pH (Student’s <i>t</i>-test, <i>P</i> < 0.01).</p