An overview of apoptosis assays detecting DNA fragmentation

Majtnerová, Pavlína; Roušar, Tomáš

doi:10.1007/s11033-018-4258-9

Cited by 223 publications

(163 citation statements)

References 115 publications

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“…As A20 functions as a regulator of cell death in a variety of tissues, we next assessed its function in the regulation of apoptosis in TIGKs. Briefly, A20-competent and -altered TIGKs were challenged with live P. gingivalis (100 MOI) for 24 h and apoptosis was determined through monitoring cleavage of caspase 3 and detecting cytoplasmic histone associated DNA using ELISA (46)(47)(48)(49). P. gingivalis is considered as a weak inducer of apoptosis and our results were consistent with this notion revealing minimum to no apoptosis in A20 competent TIGKs following bacterial infection ( Figures 7A-F).…”

Section: Lack Of A20 Sensitizes Gingival Keratinocytes To Apoptosis Fsupporting

confidence: 84%

A20 Restricts Inflammatory Response and Desensitizes Gingival Keratinocytes to Apoptosis

Mooney

Xia

et al. 2020

Front. Immunol.

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The pathophysiology of periodontal disease involves a perturbed immune system to a dysbiotic microflora leading to unrestrained inflammation, collateral tissue damage, and various systemic complications. Gingival epithelial cells function as an important part of immunity to restrict microbial invasion and orchestrate the subsequent innate responses. A20 (TNFAIP3), an ubiquitin-editing enzyme, is one of the key regulators of inflammation and cell death in numerous tissues including gastrointestinal tract, skin, and lungs. Emerging evidence indicates A20 as an essential molecule in the oral mucosa as well. In this study, we characterized the role of A20 in human telomerase immortalized gingival keratinocytes (TIGKs) through loss and gain of function assays in preclinical models of periodontitis. Depletion of A20 through gene editing in TIGKs significantly increased IL-6 and IL-8 secretion in response to Porphyromonas gingivalis infection while A20 over-expression dampened the cytokine production compared to A20 competent cells through modulating NF-κB signaling pathway. In the subsequent experiments which assessed apoptosis, A20 depleted TIGKs displayed increased levels of cleaved caspase 3 and DNA fragmentation following P. gingivalis infection and TNF/CHX challenge compared to A20 competent cells. Consistently, there was reduced apoptosis in the cells overexpressing A20 compared to the control cells expressing GFP further substantiating the role of A20 in regulating gingival epithelial cell fate in response to exogenous insult. Collectively, our findings reveal first systematic evidence and demonstrate that A20 acts as a regulator of inflammatory response in gingival keratinocytes through its effect on NF-κB signaling and desensitizes cells to bacteria and cytokine induced apoptosis in the oral mucosa. As altered A20 levels can have profound effect on different cellular responses, future studies will determine whether A20-targeted therapies can be exploited to restrain periodontal inflammation and maintain oral mucosa tissue homeostasis.

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Section: Lack Of A20 Sensitizes Gingival Keratinocytes To Apoptosis Fsupporting

confidence: 84%

A20 Restricts Inflammatory Response and Desensitizes Gingival Keratinocytes to Apoptosis

Mooney

Xia

et al. 2020

Front. Immunol.

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“…DNA fragmentation is a step in the cell signaling cascade for induction of apoptotic cell death, downstream of caspase 3 activation and cleavage and activation of the DNA fragmentation factor DFF, and many apoptotic assays are based on detection of DNA fragmentation [ 109 ]. As DNA breaks are coupled to the cell death process, false positive results and classification of cytotoxic compounds as genotoxic can occur if cytotoxicity is not an integrated part of the genotoxicity testing.…”

Section: Cytotoxicity Testing Before Genotoxicity Studiesmentioning

confidence: 99%

Genotoxicity of Nanomaterials: Advanced In Vitro Models and High Throughput Methods for Human Hazard Assessment—A Review

et al. 2020

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Changes in the genetic material can lead to serious human health defects, as mutations in somatic cells may cause cancer and can contribute to other chronic diseases. Genotoxic events can appear at both the DNA, chromosomal or (during mitosis) whole genome level. The study of mechanisms leading to genotoxicity is crucially important, as well as the detection of potentially genotoxic compounds. We consider the current state of the art and describe here the main endpoints applied in standard human in vitro models as well as new advanced 3D models that are closer to the in vivo situation. We performed a literature review of in vitro studies published from 2000–2020 (August) dedicated to the genotoxicity of nanomaterials (NMs) in new models. Methods suitable for detection of genotoxicity of NMs will be presented with a focus on advances in miniaturization, organ-on-a-chip and high throughput methods.

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“…The isolated issues were further sent for histological detection, while H&E staining ( Figure 7G) further con rmed the collected specimen were tumors. TUNEL termed as "Terminal deoxynucleotidyl transferase dUTP nick end labeling", which is a classic marker to detect DNA fragmentation in apoptotic tumor cells [56]. Interestingly, we found that expression of TUNEL was upregulated in HMON@CuS/Gd plus NIR group, while no evident changes were found in other groups, compared with saline group (Figure 7G; Supplementary les, Figure S21).…”

Section: In Vivo Photo-therapeutic Treatments Of Hmon@cus/gd Nanopartmentioning

confidence: 83%

Biodegradable hollow mesoporous organosilica nanotheranostics (HMON) for multi-mode imaging and mild photo-therapeutic-induced mitochondrial damage on gastric cancer

Guo

Chen

et al. 2020

Preprint

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Background: CuS-modified hollow mesoporous organosilica nanoparticles (HMON@CuS) have been preferred as non-invasive treatment for cancer, as near infrared (NIR)-induced photo-thermal effect (PTT) and/or photo-dynamic effect (PDT) could increase cancer cells’ apoptosis. However, the certain role of HMON@CuS-produced-PTT&PDT inducing gastric cancer (GC) cells’ mitochondrial damage, remained unclear. Moreover, theranostic eﬃciency of HMON@CuS might be well improved by applying multi-modal imaging, which could offer an optimal therapeutic region and time window. Herein, new nanotheranostics agents were reported by Gd doped HMON decorated by CuS nanocrystals (called HMON@CuS/Gd).Results: HMON@CuS/Gd exhibited appropriate size distribution, good biocompatibility, L-Glutathione (GSH) responsive degradable properties, high photo-thermal conversion eﬃciency (82.4%) and a simultaneous reactive oxygen species (ROS) generation effect. Meanwhile, HMON@CuS/Gd could efficiently enter GC cells, induce combined mild PTT (43-45 °C) and PDT under mild NIR power density (0.8W/cm2). Surprisingly, it was found that PTT might not be the only factor of cell apoptosis, as ROS induced by PDT also seemed playing an essential role. The NIR-induced ROS could attack mitochondrial transmembrane potentials (MTPs), then promote mitochondrial reactive oxygen species (mitoROS) production. Meanwhile, mitochondrial damage dramatically changed the expression of anti-apoptotic protein (Bcl-2) and pro-apoptotic protein (Bax). Since that, mitochondrial permeability transition pore (mPTP) was opened, followed by inducing more cytochrome c (Cyto C) releasing from mitochondria into cytosol, and finally activated caspase-9/caspase-3-depended cell apoptosis pathway. Our in vivo data also showed that HMON@CuS/Gd exhibited good fluorescence (FL) imaging (wrapping fluorescent agent), enhanced T1 imaging under magnetic resonance imaging (MRI) and infrared thermal (IRT) imaging capacities. Guided by FL/MRI/ IRT trimodal imaging, HMON@CuS/Gd could selectively cause mild photo-therapy at cancer region, efficiently inhibit the growth of GC cells without evident systemic toxicity in vivo. Conclusion: HMON@CuS/Gd could serve as a promising multifunctional nanotheranostic platform and as a cancer photo-therapy agent through inducing mitochondrial dysfunction on GC.

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An overview of apoptosis assays detecting DNA fragmentation

Cited by 223 publications

References 115 publications

A20 Restricts Inflammatory Response and Desensitizes Gingival Keratinocytes to Apoptosis

A20 Restricts Inflammatory Response and Desensitizes Gingival Keratinocytes to Apoptosis

Genotoxicity of Nanomaterials: Advanced In Vitro Models and High Throughput Methods for Human Hazard Assessment—A Review

Biodegradable hollow mesoporous organosilica nanotheranostics (HMON) for multi-mode imaging and mild photo-therapeutic-induced mitochondrial damage on gastric cancer

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