Quantitative Differentiation of Both Free 3′-OH and 5′-OH DNA Ends Between Heat-induced Apoptosis and Necrosis

Hayashi, R.; Ito, Yuko; Matsumoto, Kenji; Fujino, Yuko; Otsuki, Yoshinori

doi:10.1177/002215549804600909

Cited by 44 publications

(29 citation statements)

References 28 publications

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“…Alternatively, PCD activity may have been reduced by the stress-mediated upregulation of heat shock proteins that are known inhibitors of PCD. [6][7][8] A shift from PCD to necrosis occurs in several vertebrate systems in response to temperature, 29,32 other stressors and disease. [33][34][35] A similar pattern in the anemone, a lower invertebrate, suggests that the mechanisms controlling the shift between different forms of cell death in response to stress may be highly conserved.…”

Section: Discussionmentioning

confidence: 99%

Heat stress induces different forms of cell death in sea anemones and their endosymbiotic algae depending on temperature and duration

et al. 2004

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Bleaching of reef building corals and other symbiotic cnidarians due to the loss of their dinoflagellate algal symbionts ( ¼ zooxanthellae), and/or their photosynthetic pigments, is a common sign of environmental stress. Mass bleaching events are becoming an increasingly important cause of mortality and reef degradation on a global scale, linked by many to global climate change. However, the cellular mechanisms of stress-induced bleaching remain largely unresolved. In this study, the frequency of apoptosislike and necrosis-like cell death was determined in the symbiotic sea anemone Aiptasia sp. using criteria that had previously been validated for this symbiosis as indicators of programmed cell death (PCD) and necrosis. Results indicate that PCD and necrosis occur simultaneously in both host tissues and zooxanthellae subject to environmentally relevant doses of heat stress. Frequency of PCD in the anemone endoderm increased within minutes of treatment. Peak rates of apoptosis-like cell death in the host were coincident with the timing of loss of zooxanthellae during bleaching. The proportion of apoptosis-like host cells subsequently declined while cell necrosis increased. In the zooxanthellae, both apoptosis-like and necrosis-like activity increased throughout the duration of the experiment (6 days), dependent on temperature dose. A stress-mediated PCD pathway is an important part of the thermal stress response in the sea anemone symbiosis and this study suggests that PCD may play different roles in different components of the symbiosis during bleaching.

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Section: Discussionmentioning

confidence: 99%

Heat stress induces different forms of cell death in sea anemones and their endosymbiotic algae depending on temperature and duration

et al. 2004

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“…Furthermore, DNA fragmentation also occurs in necrosis. 30 For this reason, we paid attention to perform TUNEL staining always under constant conditions of reaction time, reagent concentrations and room temperature.…”

Section: Discussionmentioning

confidence: 99%

Post-traumatic moderate systemic hypothermia reduces TUNEL positive cells following spinal cord injury in rat

et al. 2004

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Study design: A standardized animal model of contusive spinal cord injury (SCI) with incomplete paraplegia was used to test the hypothesis that moderate systemic hypothermia reduces neural cell death. Terminal deoxynucleotidyl transferase [TdT]-mediated deoxyuridine triphosphate [dUTP] nick-end labeling (TUNEL) staining was used as a marker of apoptosis or cell damage. Objective: To determine whether or not moderate hypothermia could have a neuroprotective effect in neural cell death following spinal cord injury in rats. Setting: Kagawa Medical University, Japan. Methods: Male Sprague-Dawley (SD) rats (n ¼ 39) weighing on average 300 g (280-320 g) were used to prepare SCI models. After receiving contusive injury at T11/12, rats were killed at 24 h, 72 h, or 7 days after injury. The spinal cord was removed en bloc and of examined at five segments: 5 and 10 mm rostral to the center of injury, center of injury, and 5 and 10 mm caudal to the center of injury. Rats that received hypothermia (321C/4 h) were killed at the same time points as those that received normothermia (371C/3 h). The specimens were stained with hematoxylin and eosin, and subjected to in situ nick-end labeling (TUNEL), a specific method for visualizing cell death in the spinal cord. Results: At 24 h postinjury, TUNEL positive cells (TPC) decreased significantly 10 mm rostral to center of injury in hypothermic animals compared to the normothermia group. At 72 h post-SCI, TPC also decreased significantly at 5 mm rostral, and 5 and 10 mm caudal to the lesion center compared to normothermic animals. At 7 days postinjury, a significant decrease of TPC was observed at the 5 mm rostral and 5 mm caudal sites compared to normothermic animals. Conclusion: These results indicate that systemic hypothermia has a neuroprotective effect following SCI by attenuating post-traumatic TPC.

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“…Recent studies using the TUNEL method have cast doubt on its specificity of detecting apoptosis because the TUNEL method detects not only apoptotic cells but also necrotic and mitotic cells (Grasl-Kraupp et al 1995;Sit et al 1997;Wolvekamp et al 1998;Otsuki 2000Otsuki ,2004. The TUNEL method by TEM (TUNEL/TEM) was developed for retrospective studies using archival epoxyresin-embedded materials and enables the morphological discrimination of apoptotic cells among many cell types including necrotic or mitotic cells based on ultrastructural characteristics (Inoki et al 1997;Hayashi et al 1998;Otsuki 2000Otsuki ,2004. To date, several studies of psoriasis have shown that almost all proliferative keratinocytes are TUNEL positive, although psoriasis is a hyperproliferative disease (Laporte et al 2000).…”

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Method of Specific Detection of Apoptosis Using Formamide-induced DNA Denaturation Assay

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Shibata

Kusakabe

et al. 2006

J Histochem Cytochem.

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S U M M A R Y We compared the reliability between apoptosis detection methods, namely, the terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) method and formamide-induced DNA denaturation assay using a monoclonal antibody (MAb) to single-stranded DNA (ssDNA) (formamide-MAb assay). Reaction targets in these methods are different: the TUNEL method recognizes free 39-OH DNA ends, whereas the formamide-MAb assay detects ssDNA itself (25-30 bp). We found that the formamideMAb assay immunohistochemically detected apoptotic cells, whereas the TUNEL method detected apoptotic cells as well as mitotic and necrotic cells. The TUNEL method recognized not only 39-OH DNA ends cleaved by DNase during apoptosis but also constitutive physiological nicking that occurs in DNA duplication and histone posttranslational modifications during mitosis and random DNA breaks during necrotic execution. By electron microscopy, the mean labeling density (the number of 39-OH DNA ends/nuclear area) obtained by the TUNEL method was determined to be consistently higher than that (the number of ssDNAs/ nuclear area) obtained by the formamide-MAb assay. On the basis of these findings, we conclude that the formamide-MAb assay was more specific than the TUNEL method for the detection of apoptotic cells using electron microscopy; however, the labeling intensity of the formamide-MAb assay was slightly weaker than that of the TUNEL method. (J Histochem Cytochem 54:683-692, 2006)

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Quantitative Differentiation of Both Free 3′-OH and 5′-OH DNA Ends Between Heat-induced Apoptosis and Necrosis

Cited by 44 publications

References 28 publications

Heat stress induces different forms of cell death in sea anemones and their endosymbiotic algae depending on temperature and duration

Heat stress induces different forms of cell death in sea anemones and their endosymbiotic algae depending on temperature and duration

Post-traumatic moderate systemic hypothermia reduces TUNEL positive cells following spinal cord injury in rat

Method of Specific Detection of Apoptosis Using Formamide-induced DNA Denaturation Assay

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