TUNEL Assay for Apoptotic Cells

Heatwole, Virginia M.

doi:10.1385/1-59259-213-9:141

Cited by 54 publications

(46 citation statements)

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“…Apoptosis in tumor sections was assessed with the use of the terminal nucleotidyl transferase-mediated nick end labeling (TUNEL) assay (22). All components for this procedure were from the ApopTag In situ Apoptosis Detection kit (Chemicon), which was used according to the manufacturer's instructions.…”

Section: Methodsmentioning

confidence: 99%

HIV-1 Protease Inhibitors Nelfinavir and Atazanavir Induce Malignant Glioma Death by Triggering Endoplasmic Reticulum Stress

Pyrko¹,

Kardosh²,

et al. 2007

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HIV type 1 (HIV-1) protease inhibitors (PI) have been shown to have anticancer activity in non-HIV-associated human cancer cells. The underlying mechanism of this effect is unclear. Here, we show that the PIs nelfinavir and atazanavir cause cell death in various malignant glioma cell lines in vitro. The underlying mechanism of this antitumor effect involves the potent stimulation of the endoplasmic reticulum

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Section: Methodsmentioning

confidence: 99%

HIV-1 Protease Inhibitors Nelfinavir and Atazanavir Induce Malignant Glioma Death by Triggering Endoplasmic Reticulum Stress

Pyrko¹,

Kardosh²,

et al. 2007

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“…DNA damage can be detected by TdT (terminal deoxynucleotidyl transferase)-mediated dUTP nick-end labelling (TUNEL) assay, as previously described [36]. Thus, DNA fragmentation was measured using an in situ cell death detection kit, fluorescein, using fluorescein-dUTP as a label according to the manufacturer's instructions.…”

Section: Tunel Assaymentioning

confidence: 99%

“…The TUNEL assay is a well-established method for detection of DNA cleavage, a relatively late apoptotic marker [36]. In addition, this technique has been widely used to determine DNA damage in human sperm [37,38].…”

Section: H2o2-and Progesterone-induced Dna Damagementioning

confidence: 99%

Relationship between Caspase Activity and Apoptotic Markers in Human Sperm in Response to Hydrogen Peroxide and Progesterone

et al. 2009

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Abstract. Apoptosis plays an essential role in normal spermatogenesis, but deregulations of this biological process, which is closely associated with male infertility, have been found. Whereas calcium homeostasis is a key regulator of cell survival, sustained elevation of intracellular calcium plays a role in apoptosis. The aim of this research was to determine the role of two different calcium mobilizing agents, hydrogen peroxide (H2O2) and the physiological agonist progesterone, on the apoptosis process of human ejaculated spermatozoa. Translocation of membrane phosphatidylserine was examined with an annexin V binding assay, DNA damage was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL assay) and caspase-3 activity was assessed using a fluorometric assay. After incubation of spermatozoa for 1 h with either 10 μM H2O2 or 20 μM of progesterone, there was a significant increase in both caspase-3 activity and the percentage of annexin V-positive cells. Similarly, the TUNEL results were significantly higher 1 h after incubation with either 10 μM H2O2 or 20 μM of progesterone. In fact, progesterone-treated cells showed a three-fold increase (from 17.6 to 52.9%) of TUNEL-positive cells compared to untreated cells, while H2O2-treated cells exhibited a two-fold increase (from 17.6 to 37.9%). In sum, our results suggest that spermatozoa treated with calcium mobilizing agents, such as H2O2 and progesterone, seem to undergo an apoptosis process that is dependent on caspase-3 activation. Key words: Apoptosis, Caspases, Phosphatidylserine exposure, Spermatozoa, TUNEL (J. Reprod. Dev. 55: [615][616][617][618][619][620][621] 2009) poptosis is a distinctive form of cell death characterized by a series of morphological and biochemical changes that plays a critical role in many physiological processes such as development, tissue homeostasis, inflammatory responses and disease [1]. It also plays an essential role in gamete maturation and embryogenesis, contributing to appropriate formation of various organs and structures. In fact, many studies in animal models have demonstrated that apoptosis is the underlying mechanism of germ cell death during normal spermatogenesis [2], which has also been reported in humans [3].On the other hand, deregulations of this biological process, which is closely associated with male infertility, have been found, and both men with azoospermia and men with severe oligozoospermia have an increased frequency of apoptotic germ cells in their testicular tissues compared with patients having normal spermatogenesis [2,4,5]. In addition, a certain degree of spontaneous apoptosis and relatively high rates of apoptosis have been detected using an in situ end-labelling (TUNEL) assay in testicular biopsies of infertile patients with testicular insufficiency [6,7]. Similarly, disturbance of cell membrane symmetry with translocation of phospatidylserine residues to the outer layer of the plasma membrane has been observed in germ cells from patients with complete spermi...

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“…4) in X. tropicalis. The TUNEL technology reveals multiple DNA cleavages in cells, a late step specifically encountered in apoptotic cells (Heatwole, 1999;Saraste, 1999;Nagata, 2000). Likewise, Acridine Orange, a DNA intercalater, can be used as a vital marker providing staining of cells undergoing apoptosis (Abrams et al, 1993;Gaumer et al, 2000).…”

Section: Apoptosis In Muscle and Spinal Cord Followed By Tdt-mediatedmentioning

confidence: 99%

Autonomous regulation of muscle fibre fate during metamorphosis in Xenopus tropicalis

Rowe

Coen

Blay

et al. 2002

Developmental Dynamics

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A key event in metamorphosis of anuran amphibians is tail resorption. This composite structure includes epidermal cells, spinal cord, muscle fibres and connective tissue. It is unclear how resorption proceeds and to what extent the signals for the death process are transmitted between cells. We determined the kinetics of metamorphosis, apoptosis, and tail regression in the diploid anuran, Xenopus tropicalis, a species more suited to genetic analysis than the pseudotetraploid, Xenopus laevis. Metamorphosis was found to proceed at a regular and predictable rate in X. tropicalis but not in X. laevis. Caspase 3 activity and mRNA levels were correlated with TdT-mediated dUTP nick endlabeling (TUNEL) signalling and most markedly increased in tail muscle and spinal cord. It has been proposed that muscles die as a result of loss of connectivity with the surrounding matrix. To test this hypothesis, we used direct DNA injection in trunk and tail muscle to overexpress Xenopus Bcl-X L (xR11), an anti-apoptotic gene, along with a marker gene (luciferase or GFP). xR11 significantly inhibited the cell death process in both trunk and tail muscle. This protection was functional even up to stage 64 on completion of tail regression. We conclude that (1) somatic gene transfer can be applied to analyse cell fate in X. tropicalis, and (2) that muscle death can be abrogated despite extracellular matrix loss.

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TUNEL Assay for Apoptotic Cells

Cited by 54 publications

References 0 publications

HIV-1 Protease Inhibitors Nelfinavir and Atazanavir Induce Malignant Glioma Death by Triggering Endoplasmic Reticulum Stress

HIV-1 Protease Inhibitors Nelfinavir and Atazanavir Induce Malignant Glioma Death by Triggering Endoplasmic Reticulum Stress

Relationship between Caspase Activity and Apoptotic Markers in Human Sperm in Response to Hydrogen Peroxide and Progesterone

Autonomous regulation of muscle fibre fate during metamorphosis in Xenopus tropicalis

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