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.

The role of apoptosis in the antigen-specific T cell hyporesponsiveness of paracoccidioidomycosis patients

Paracoccidioidomycosis is a deep endemic mycosis associated with an antigen-specific immunodeficiency. To examine the role of apoptosis in this immunodeficiency, peripheral blood mononuclear cells (PBMC) of patients with paracoccidioidomycosis and controls were stimulated with the main antigen of Paracoccidioides brasiliensis (gp43) and an unrelated fungal antigen (from Candida albicans, CMA) and analyzed for annexin V and propidium iodide staining by flow cytometry. Control PBMC proliferated well with both antigens. Patients' PBMC proliferated only with CMA, but presented higher levels of apoptosis with gp43 and CMA than in their own unstimulated cultures. Moreover, gp43-triggered apoptosis in control PBMC was lower than in those of the patients. Thus, patient but not control gp43-stimulated T cells apparently remained anergized and subsequently underwent apoptosis. While CMA-induced apoptosis is likely triggered by activation-induced cell death, this is apparently not the case in gp43-induced apoptosis because of the lack of cell cycling and IL-2 in the gp43-stimulated cultures. However, higher IL-10 levels were found in gp43-stimulated patient PBMC cultures. Addition of a neutralizing anti-IL-10 antibody to the cultures resulted in increased apoptosis levels only in gp43-stimulated patient PBMC cultures. Our results suggest that apoptosis plays a role in the patients' antigen-specific hyporesponsiveness and that IL-10 may have an antiapoptotic role. (C) 2002 Elsevier B.V. (USA).

Cell cycle arrest and apoptosis in TP53 subtypes of bladder carcinoma cell lines treated with cisplatin and gemcitabine

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

Spontaneous germ cell death by apoptosis in epididymis of the adult bat Artibeus lituratus

Many factors can lead cells to apoptosis during the various stages of cell life. This study was undertaken to characterize germ cell death in the epididymis of the adult Artibeus lituratus by histochemical and immunohistochemical techniques using light microscopy and transmission electron microscopy. The results showed that cells with a nuclear phenotype and ultrastructural characteristics of chromatin compaction were common in apoptosis. The Apoptag test confirmed that the suspected cells were apoptotic. It is suggested that immature germ cells, when released from the germinative epithelium, may be directed towards the epididymis instead of being disposed of in the testicle. Furthermore, intact immature cells can leave the testicle in the initial phases of apoptosis and complete this phenomenon in the epididymis.

Cell Death in Salivary Glands of Rhipicephalus (Boophilus) microplus (Canestrini, 1887) (Acari: Ixodidae) Females at Semi-engorged Feeding Stage

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

Cocaine induces cell death and activates the transcription nuclear factor kappa-b in pc12 cells

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

Cytotoxicity of Portland Cement with Different Radiopacifying Agents: A Cell Death Study

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Cell death and quantitative reduction of rests of Malassez according to age

Background and Objective: Rests of Malassez are clusters of epithelial cells that remain in the periodontal ligament throughout life. However, it has been reported that the number of these structures decreases with age, and some epithelial cells undergo apoptosis in rests of Malassez of young and adult rats. Therefore, the purpose of the present study was to investigate the incidence of epithelial cell death and the quantitative changes in the rests of Malassez in rat molars of different ages.Material and Methods: Fragments containing the upper molars of rats aged 29, 45 and 120 d were fixed, decalcified and embedded for analysis by light microscopy. In the sections stained by hematoxylin and eosin, the number of rests of Malassez and the number of nuclei of these epithelial structures were obtained. Moreover, the nuclei exhibiting typical features of cell death were also counted in each rest of Malassez. The terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method for detection of cell death was also carried out.Results: In all groups examined, some rests of Malassez exhibited epithelial cell nuclei with typical features of apoptosis and some of them were also TUNEL positive. From 29 to 120 d of age in rats, the quantitative analysis showed a significant decrease in the total number of rests of Malassez in the cervical, middle and furcation regions of the periodontal ligament. Moreover, a significant decrease of epithelial cell nuclei was concomitant to an increase in the frequency of cell death in the oldest rats.Conclusion: These results suggest that epithelial cell death by apoptosis may be, at least in part, responsible for the reduction in the number of rests of Malassez according to age.

Cell death evaluation in benzo[a]pyrene-transformed human breast epithelial cells after microcell-mediated transfer of chromosomes 11 and 17

The incidence of apoptosis and nuclear instability, including the incidence of catastrophic death, were investigated in benzo[a]pyrene (BP)-transformed human breast epithelial cells (BP1-E cell line) after microcell-mediated transfer of chromosomes 11 and 17. Since the introduction of normal chromosomes 11 and 17 into tumorigenic human breast BP1-E cells reverts some of these cells' characteristics (especially those affected by microsatellite instabilities and loss of heterozygosity) those of parental non-transformed MCF-10F cells, it was expected that the cell death rates would also be affected by this treatment. The transfer of the mentioned chromosomes, especially chromosome 17, to tumorigenic BP1-E cells increased the apoptotic ratios and decreased the nuclear instability ratios, thus showing that the microsatellite instability and loss of heterozygosity induced by BP in these chromosomes of MCF-10F cells affect the control of cell death mechanisms. (C) 2003 Elsevier B.V. All rights reserved.

Modes of cell death in the hypopharyngeal gland of the honey bee (Apis mellifera L)

Different modes of cell death have been revealed in the regressing hypopharyngeal glands of worker honey bees. The hypopharyngeal gland, which is well developed in young nursing bees to produce protein for larval food, was seen to regress naturally in foraging adult worker bees. A range of techniques including histology, cytochemistry, in situ TUNEL, Annexin V and Comet assays indicated that cells within the gland demonstrate progressive symptoms of apoptosis, necrosis and a vacuolar form of programmed cell death. The latter mode of cell death did not display chromatin margination, but was accompanied by an enhanced level of autophagic and hydrolytic activity in which a cytosolic source of acid phosphatase became manifest in the extra-cisternal spaces. Normal and annexin-positive cells were found to occur in the younger nursing bees, whilst necrosis and an aberrant vacuolar type of apoptosis predominated in the older foraging bees. The relevance of these results to the classification of programmed cell death is discussed. (C) 2000 Academic Press.

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.

Fipronil-induced cell death in salivary glands of Rhipicephalus sanguineus (Latreille, 1806) (Acari: Ixodidae) semi-engorged females

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

Markers of cell death in salivary glands of males of the tick Rhipicephalus sanguineus (Latreille, 1806) (Acari, Ixodidae)

The salivary glands of Rhipicephalus sanguineus males at stages: unfed (control), at day seven post-attachment, and at days three and seven post-detachment from the host were examined using methods of enzymatic analysis and cell viability. At these stages of feeding, different staining patterns were observed in the cells of type IV, III, II and I acini, which were affected by degeneration in this sequence. Acid phosphatase reaction was inversely proportional to that of ATPase, while ATPase reaction was proportional to membrane integrity.Salivary gland cells of unfed males exhibited intact nucleus and plasma membrane, suggesting that the acid phosphatase detected may participate in the normal physiology of acini. In males at day seven post-attachment, intact membranes were observed in almost all types of acini, as well as stronger reaction for acid phosphatase, nuclear changes, and decrease in ATPase reaction, changes associated with the degenerative process. At days three and seven post-detachment degeneration progress, being observed loss of membrane integrity, nuclear changes, prominent decrease in ATPase reaction, and an increase in acid phosphatase reaction in the first case and a decreased of it at day seven post-detachment from the host. During cell death, alterations occurred in the following sequence: a) nuclear changes, b) loss of ATPase reaction, c) loss of integrity of the plasma membrane, and d) increase of acid phosphatase. The latter might be associated with the late degradation of cytoplasmic remnants, characterizing the process of cell death in glands of R. sanguineus males as atypical or non-classic apoptosis. (C) 2008 Elsevier B.V. All rights reserved.

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.

Ultrastructural analysis of the effect of mating delay on cell death in the ovaries of virgin honey bee (Apis mellifera L.) queens

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