Programmed cell death in the larval salivary glands of Apis mellifera (Hymenoptera, Apidae)

The morphological and histochemical features of degeneration in honeybee (Apis mellifera) salivary glands were investigated in 5th instar larvae and in the pre-pupal period. The distribution and activity patterns of acid phosphatase enzyme were also analysed. As a routine, the larval salivary glands were fixed and processed for light microscopy and transmission electron microscopy. Tissue sections were subsequently stained with haematoxylin-eosin, bromophenol blue, silver, or a variant of the critical electrolyte concentration (CEC) method. Ultrathin sections were contrasted with uranyl acetate and lead citrate. Glands were processed for the histochemical and cytochemical localization of acid phosphatase, as well as biochemical assay to detect its activity pattern. Acid phosphatase activity was histochemically detected in all the salivary glands analysed. The cytochemical results showed acid phosphatase in vesicles, Golgi apparatus and lysosomes during the secretory phase and, additionally, in autophagic structures and luminal secretion during the degenerative phase. These findings were in agreement with the biochemical assay. At the end of the 5th instar, the glandular cells had a vacuolated cytoplasm and pyknotic nuclei, and epithelial cells were shed into the glandular lumen. The transition phase from the 5th instar to the pre-pupal period was characterized by intense vacuolation of the basal cytoplasm and release of parts of the cytoplasm into the lumen by apical blebbing; these blebs contained cytoplasmic RNA, rough endoplasmic reticule and, occasionally, nuclear material. In the pre-pupal phase, the glandular epithelium showed progressive degeneration so that at the end of this phase only nuclei and remnants of the cytoplasm were observed. The nuclei were pyknotic, with peripheral chromatin and blebs. The gland remained in the haemolymph and was recycled during metamorphosis. The programmed cell death in this gland represented a morphological form intermediate between apoptosis and autophagy.

Nuclear alterations associated to programmed cell death in larval salivary glands of Apis mellifera (Hymenoptera : Apidae)

The silk glands of bees are a good model for the study of cell death in insects. With the objective to detect the nuclear features during glandular regression stage, larvae at the last instar and pre-pupae were collected and their silk glands were dissected and processed for ultrastructural analysis and histologically for cytochemical and imunocytochemical analysis. The results showed that the cellular nuclei exhibited characteristics of death by atypical apoptosis as well as autophagic cell death. Among the apoptosis characteristic were: nuclear strangulation with bleb formation in some nuclei, DNA fragmentation in most of the nuclei and nucleolar fragmentation. Centripetal chromatin compaction was observed in many nuclei, forming a perichromatin halo differing from typical apoptotic nuclei. With regards to the characteristics of autophagic-programmed cell death, most relevant was the delay in the collapse of many nuclei. (C) 2006 Elsevier Ltd. All rights reserved.

A new nucleolar body appears in Drosophila saltans salivary gland cells before histolysis, in programmed cell death

The salivary glands of Drosophila saltans ( saltans group, saltans subgroup) analyzed in an advanced stage of programmed cell death showed the appearance of a single, round, nucleolar corpuscle inside the highly altered nucleus of every gland cell, at a time during which the integrity of the original nucleolus was already lost and the original nucleolar material apparently disappeared. In the same nuclei, which already had also lost the characteristic chromosome structure, some delicate chromosome threads were maintained. In many cells, the new nucleolar corpuscle and these chromosome threads are associated. These findings are novel. However, the hypothesis put forward concerning their meaning remains dependent on other studies.

Programmed cell death in salivary glands of Drosophila arizonae and Drosophila mulleri

Programmed cell death (PCD) in insect metamorphosis assumes a great diversity of morphology and controlling processes that are still not well understood. With the objective of obtaining information about the PCD process, salivary glands of Drosophila arizonae and D. mulleri were studied during larval-pupal development. From the results, it can be concluded that the type of the PCD that occurs in these organs is morphologically typical of apoptosis (formation of apoptotic nuclei, followed by fragmentation into apoptotic bodies). Histolysis happens in both species, between 22 and 23 h after pupation. There were no significant differences between the species studied. Apoptosis does not occur simultaneously in all cells. Cytoplasmic acid phosphatase activity gradually increases during development, suggesting the existence of acid phosphatases that are only expressed during the apoptotic stage. Twenty hours after pupation, salivary glands already show biochemical alterations relative to nuclear permeability such as acidification, possibly due to the fusion of lysosomes with the nucleus a few hours before apoptosis. Autophagy seems to act together with apoptosis and has a secondary role in cell death. ©FUNPEC-RP.

Canine transmissible venereal tumors: Aspects related to programmed cell death

Canine transmissible venereal tumor (CTVT) is a neoplasm transmitted by the physical transfer of viable tumor cells by direct contact with injured skin and/or mucous tissue. These cells can transpose across histocompatibility barriers into unrelated hosts. This review focuses on the biology of apoptosis and the interaction of proteins involved in this process, as well as p53, p63 and the antiapoptotic protein Bcl-2. As such, this disease offer unique opportunity to study the biology of transplantable tumours and the interaction of proteins involved in apoptosis process and the prognosis of CTVT.

Involvement of Renal Corpuscle microRNA Expression on Epithelial-to-Mesenchymal Transition in Maternal Low Protein Diet in Adult Programmed Rats

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Programmed cell death-associated gene transcripts in bovine embryos exposed to bovine Herpesvirus type 5

In vitro-produced bovine embryos become infected after exposure to bovine Herpesvirus type 5 (BoHV-5), yet no changes in developmental rates, mitochondrial activity and inhibition of apoptosis are detected in comparison to unexposed embryos. Thus, the aim of the present study was to assess the transcription of mitochondria-mediated apoptosis genes using TaqMan real-time polymerase chain reaction. Transcripts of mcl-1, caspase-2, -3, Apaf-1 and Bax genes were measured after exposure to BoHV-5 in vitro. Mitochondrial dehydrogenase activity was evaluated by MIT test and compared between groups of exposed and unexposed embryos, at day 7 of development. The rate of oocyte maturation was assessed by the extrusion of the first polar body. In summary, BoHV-5 exposed embryos retained their viability, mitochondrial dehydrogenase activity and displayed up-regulation of transcription of survival mcl-1 gene and down-regulation of Bax transcription in relation to mitochondria-mediated pathway which might improve embryo viability. These findings demonstrate that BoHV-5 exposed embryos maintain their viability and mitochondrial dehydrogenase activity with no compromise of embryos produced in vitro. (c) 2013 Elsevier Ltd. All rights reserved.

Estudo do papel da proteína eIF5A na elongação da tradução em S. cerevisiae: análise da relação funcional com o trna de alanina e sua participação na síntese proteica geral da célula

Pós-graduação em Biotecnologia - IQ

Alkaloids extracted from Pterogyne nitens induce apoptosis in malignant breast cell line

In the present study, two alkaloids isolated from Pterogyne nitens, a plant native to Brazil, have been shown to induce apoptosis in human breast cancer cells. These compounds, pterogynine (PGN) and pterogynidine (PGD), were tested for their effect on a human infiltrating ductal carcinoma cell line (ZR-7531). The cell line was treated with each alkaloid at several concentrations. Time-dependence (with or without recuperation time) and concentration-dependence (in the range 0.25-10 mM) were investigated in cytotoxicity and apoptosis assays. The annexin assay indicated an apparently higher percentage of death by necrosis of malignant cells after 24 h exposure to both P. nitens extracts than the Hoechst assay. Thus, our results in the two tests demonstrated that the Hoechst assay can discriminate between late apoptotic cells and necrosis, whereas the flow cytometry-based annexin V assay cannot. We concluded that PGN and PGD have effective antineoplastic activity against human breast cancer cells in vitro, by inducing programmed cell death.

Finite-Element Analysis of Stress on Dental Implant Prosthesis

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anestesia para separação de gêmeos isquiópagos no período neonatal: relato de caso

JUSTIFICATIVA E OBJETIVOS: A separação de gêmeos unidos causa grande interesse devido à complexidade da anestesia e cirurgia, à raridade da patologia e às poucas chances de sobrevida. O objetivo desta descrição é o de contribuir para a literatura existente, relatando os desafios encontrados por nossa equipe no atendimento à cirurgia-anestesia de separação de gêmeos isquiópagos. RELATO do CASO: Pacientes gêmeos, nascidos a termo, de parto cesariano, pesando juntos 5.100 g, classificados como isquiópagus tetrapus. Duas equipes anestésico-cirúrgicas estavam presentes, sendo o procedimento anestésico esquematizado com aparelho de anestesia, cardioscópio, capnógrafo, oxímetro de pulso, termômetro elétrico, estetoscópio esofágico, todos em dobro. Realizou-se indução anestésica com halotano e fentanil, com os gêmeos em posição lateral e com rotação da cabeça em 45º para facilitar a intubação traqueal. Os recém-nascidos foram mantidos em ventilação controlada manualmente, utilizando o sistema de Rees-Baraka. A anestesia foi mantida com halotano, oxigênio e fentanil. Durante o per-operatório, foram encontrados órgãos abdominais duplos, com exceção do cólon, que era único. As bexigas e os ísquios estavam ligados. Ao final da cirurgia as duas crianças apresentavam-se com sinais vitais estáveis. Os gêmeos permaneceram na Unidade de Terapia Intensiva (UTI) Neonatal por quatro semanas e receberam alta em bom estado geral. CONCLUSÕES: Ressalta-se a importância do entrosamento da equipe, do estudo retrospectivo multidisciplinar, da monitorização adequada e acurada observação clínica; todos esses fatores contribuíram para a boa evolução e alta dos gêmeos.

Decrease in the number and apoptosis of alveolar bone osteoclasts in estrogen-treated rats

Bone is a mineralized tissue that is under the influence of several systemic, local and environmental factors. Among systemic factors, estrogen is a hormone well known for its inhibitory function on bone resorption. As alveolar bone of young rats undergoes continuous and intense remodeling to accommodate the growing and erupting tooth, it is a suitable in vivo model for using to study the possible action of estrogen on bone. Thus, in an attempt to investigate the possibility that estrogen may induce the death of osteoclasts, we examined the alveolar bone of estrogen-treated rats.Fifteen, 22-d-old female rats were divided into estrogen, sham and control groups. The estrogen group received estrogen and the sham group received corn oil used as the dilution vehicle. After 8 d, fragments containing alveolar bone were removed and processed for light microscopy and transmission electron microscopy. Sections were stained with hematoxylin and eosin and tartrate-resistant acid phosphatase (TRAP)-an osteoclast marker. Quantitative analysis of the number of TRAP-positive osteoclasts per mm of bone surface was carried out. For detecting apoptosis, sections were analyzed by the Terminal deoxynucleotidyl transferase-mediated dUTP Nick-End Labeling (TUNEL) method; TUNEL/TRAP combined methods were also used.The number of TRAP-positive osteoclasts per mm of bone surface was significantly reduced in the estrogen group compared with the sham and control groups. TRAP-positive osteoclasts exhibiting TUNEL-positive nuclei were observed only in the estrogen group. In addition, in the estrogen group the ultrastructural images revealed shrunken osteoclasts exhibiting nuclei with conspicuous and tortuous masses of condensed chromatin, typical of apoptosis.Our results reinforce the idea that estrogen inhibits bone resorption by promoting a reduction in the number of osteoclasts, thus indicating that this reduction may be, at least in part, a consequence of osteoclast apoptosis.

Structural and ultrastructural aspects of ontogenesis and differentiation of resin secretory cavities in Hymenaea stigonocarpa (Fabaceae-Caesalpinioideae) leaves

Cell lysis in the formation of secretory cavities in plants has been questioned by some authors and considered as result of technical artifacts. To describe the formation of secretory resin cavities in Hymenaea stigonocarpa leaves, leaflet samples at different stages of differentiation were collected, fixed, and processed for light and electron microscopy as per usual methods. The initial cells of secretory resin cavities are protodermal and grow towards the mesophyll ground meristem; these cells then divide producing cell groups that are distinguished by the shape and arrangement of cytoplasm, and density. At the initial stages of differentiation of the secretory cavities, some central cells in these groups show dark cytoplasm and condensed nuclear chromatin. Later, there is cell wall loosening, tonoplast and plasmalemma rupture resulting in cell death. These cells, however, maintain organelle integrity until lysis, when the cell wall degrades and the plasmalemma ruptures, releasing protoplast residues, marked characteristics of programmed cell death. The secretory epithelium remains active until complete leaf expansion when the cavity is filled with resin and the secretory activity ceases. There are no wall residues between central cells in adult cavities. Our results demonstrate lysigeny and the importance of ontogenetic studies in determining the origin of secretory cavities.

The role of the parenchyma sheath and PCD during the development of oil cavities in Pterodon pubescens (Leguminosae-Papilionoideae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)