Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis

Koopman, Gerrit; Reutelingsperger, CP; Kuijten, G. A. M.; Keehnen, RM; Pals, Steven T.; Oers, MH van

doi:10.1182/blood.v84.5.1415.bloodjournal8451415

Cited by 1,862 publications

(94 citation statements)

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“…4D) and displayed AV/PI positive staining consistent with apoptosis (Fig. 4D,F,G) 49 . Morphological analysis further indicated that neutrophils cultured for 30 hours presented as a mixture of viable cells and apoptotic cells with condensed and rounded nuclei 50 (Fig.…”

Section: Resultsmentioning

confidence: 72%

Characterization of IL-10-producing neutrophils in cattle infected with Ostertagia ostertagi

et al. 2019

Sci Rep

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IL-10 is a master regulator of immune responses, but its cellular source and function in cattle during the initial phase of immune priming have not been well established. Despite a massive B cell response in the abomasal draining lymph nodes in Ostertagia ostertagi (OO)-infected cattle, protective immunity is slow to develop, and partial protection requires years of repeated exposure. In addressing this problem, our initial hypothesis was that B cells produce IL-10 that downregulates the host protective immune response. However, our results showed that neutrophils made up the majority of IL-10-producing cells in circulation and in secondary lymphoid tissues, particularly the spleen (80%). Conversely, IL-10-producing B cells were rare. In addition, approximately 10% to 20% of the neutrophils in the blood and spleen expressed MHC II and were IL-10 negative, suggesting that neutrophils could also participate in antigen presentation. In vitro investigation of bovine neutrophils revealed that exposure thereof to OO extract increased IL-10 and MHC II expression in these cells in a dose-dependent manner, consistent with IL-10+/MHC II+ neutrophils detected in cattle shortly after experimental OO infection. Co-culture of untreated neutrophils with anti-CD3 antibody (Ab)-stimulated CD4+ T cells led to enhanced T cell activation; also, IL-10 depletion with neutralizing Ab enhanced the stimulatory function of neutrophils. OO extract depressed neutrophil stimulation of CD4+ T cells in the presence of IL-10-neutralizing Ab, suggesting that OO utilizes both IL-10-dependent and independent mechanisms to manipulate the bovine immune response. Finally, contact and viability were required for T cell-stimulatory neutrophil function. This report, to the best of our knowledge, is the first to demonstrate that neutrophil-derived IL-10 is directly involved in T cell regulation in cattle. Our data suggest that neutrophils and neutrophil-derived IL-10 are co-opted by nematode parasites and other pathogens to attenuate host immune responses and facilitate pathogen survival.

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Section: Resultsmentioning

confidence: 72%

Characterization of IL-10-producing neutrophils in cattle infected with Ostertagia ostertagi

et al. 2019

Sci Rep

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“…If our hypotheses are correct, we would expect perforin to colocalize with exposed PS 28 on the CTL plasma membrane. To test this, we performed confocal microscopy on activated CTLs that were pre-treated with annexin V-Alexa Fluor 568, as a marker for PS exposed on the cell surface 29 . Importantly, this pre-labelling with annexin V ensured detection only of PS already exposed on the CTL membrane before perforin addition, thus avoiding any contribution of PS flip/flop upon perforin association with the plasma membrane 30 .…”

Section: Resultsmentioning

confidence: 99%

Lipid order and charge protect killer T cells from accidental death

et al. 2019

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Killer T cells (cytotoxic T lymphocytes, CTLs) maintain immune homoeostasis by eliminating virus-infected and cancerous cells. CTLs achieve this by forming an immunological synapse with their targets and secreting a pore-forming protein (perforin) and pro-apoptotic serine proteases (granzymes) into the synaptic cleft. Although the CTL and the target cell are both exposed to perforin within the synapse, only the target cell membrane is disrupted, while the CTL is invariably spared. How CTLs escape unscathed remains a mystery. Here, we report that CTLs achieve this via two protective properties of their plasma membrane within the synapse: high lipid order repels perforin and, in addition, exposed phosphatidylserine sequesters and inactivates perforin. The resulting resistance of CTLs to perforin explains their ability to kill target cells in rapid succession and to survive these encounters. Furthermore, these mechanisms imply an unsuspected role for plasma membrane organization in protecting cells from immune attack.

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“…Organisms evolve the cell suicide mechanism termed apoptosis to eliminate the redundant, damaged, or infected cells. 26 The Annexin V-FITC assay perceives such early apoptotic cells through flow cytometry assay in a cell population. In order to determine whether the cumin nanoemulsion induces cell death by apoptosis, Annexin V-FITC Assay was performed using cumin nanoemulsion at a concentration of 1.5 µL/mL.…”

Section: Flow Cytometrymentioning

confidence: 99%

<p>Ultrasonic Nanoemulsification of <em>Cuminum cyminum</em> Essential Oil and Its Applications in Medicine</p>

Nirmala¹,

Durai²,

Rao³

et al. 2020

IJN

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Background and Study: Cumin seed oil (extracted from Cuminum cyminum) has many applications but conclusive evidence of its therapeutic uses has not been presented. This study has explored the anticancer and antibacterial properties of the seed oil. Methods: The cumin nanoemulsion was prepared with Tween 80 non-ionic surfactant employing ultra-sonication technology. The anticancer activity of the nanoscale-based emulsion was evaluated through cell viability (MTT), antiproliferation evaluation through clonogenic assay, and apoptosis through Annexin V-FITC assay. Agar well diffusion was used to study the antimicrobial activity, and this was supported by membrane integrity analysis. Results: A thorough study of process parameters, aimed at obtaining the optimal surface concentration and emulsification time, was completed. GC-MS data indicated cumaldehyde as a major component. The resultant droplet diameter after a sonication time of 5 min was 10.4 ± 0.5 nm. MTT assay revealed the IC 50 value at 1.5 µL/mL and the early induction of apoptosis was evident. Tongue carcinoma cell line treated with cumin nanoemulsion presented a diminished colony formation. The nanoemulsion exhibited significant antibacterial activity against S. aureus. A significant cytoplasmic leakage was observed on treatment with cumin nanoemulsion. The consequences of the analysis projected cumin as a potential component for cancer therapy. Conclusion: This study provides definitive evidence for cumin essential oil nanoemulsion as a legitimate plant-based medicine that can bypass the drawbacks of the present aggressive treatment of cancer, can overcome the antimicrobial resistance, and can also meet all prerequisites.

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Annexin V for flow cytometric detection of phosphatidylserine expression on B cells undergoing apoptosis

Cited by 1,862 publications

References 0 publications

Characterization of IL-10-producing neutrophils in cattle infected with Ostertagia ostertagi

Characterization of IL-10-producing neutrophils in cattle infected with Ostertagia ostertagi

Lipid order and charge protect killer T cells from accidental death

<p>Ultrasonic Nanoemulsification of <em>Cuminum cyminum</em> Essential Oil and Its Applications in Medicine</p>

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