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Published on: November 12, 2017
Correlation of DNA fragmentation and chromatin condensation in apoptotic nuclei of the Ser 6 mouse retina
1Department of Ophthalmology, Duke University School of Medicine, Durham, North Carolina 27710, USA.
Abstract:
The form of cell death known as apoptosis was first described in thymocytes. The hallmarks of apoptosis include chromatin condensation, membrane blebbing, formation of apoptotic bodies, and DNA fragmentation. DNA fragmentation can be visualized morphologically by the TdT-mediated dUTP-biotin nick end labeling (TUNEL) method that labels the cut DNA ends. However, at the light microscopic (LM) level, TUNEL-positive nuclei cannot readily be correlated with the other hallmarks of apoptosis. In the retina, chromatin condensation and DNA fragmentation are the major features of developmental cell death as well as photoreceptor degeneration. We performed TUNEL at the electron microscopic (EM) level, which permitted correlation of DNA fragmentation with chromatin condensation. We studied the retinas of transgenic mice (Ser 6) expressing the Pro347Ser mutant rhodopsin gene during developmental cell death (age 7 days) and photoreceptor degeneration (age 21 days). We found that 90% of the nuclei showing chromatin condensation were TUNEL positive as well. Our results demonstrated DNA fragmentation and chromatin condensation in the same cells as they underwent apoptosis in vivo, confirming the notion that these processes are concomitant events, and by implication, that activation of an endogenous endonuclease is an important step in the death process of retinal neurons.
Insights
Apoptosis, a form of cell death, involves DNA fragmentation and chromatin condensation. Electron microscopy confirmed these events occur simultaneously in retinal neurons during development and degeneration.
Area of Science:
- Neuroscience
- Cell Biology
- Genetics
Background:
- Apoptosis, or programmed cell death, is characterized by chromatin condensation and DNA fragmentation.
- The TdT-mediated dUTP-biotin nick end labeling (TUNEL) method detects DNA fragmentation, but correlating it with other apoptotic hallmarks is challenging at the light microscopic level.
- In the retina, chromatin condensation and DNA fragmentation are key features of developmental cell death and photoreceptor degeneration.
Purpose of the Study:
- To correlate DNA fragmentation with chromatin condensation during apoptosis in retinal neurons using electron microscopy.
- To investigate these processes in transgenic mice expressing a mutant rhodopsin gene.
Main Methods:
- Utilized the TUNEL method at the electron microscopic (EM) level.
- Examined retinas of transgenic mice (Pro347Ser mutant rhodopsin) at 7 and 21 days of age.
- Correlated TUNEL-positive nuclei with chromatin condensation.
Main Results:
- Electron microscopy allowed direct correlation of DNA fragmentation with chromatin condensation in apoptotic retinal neurons.
- In transgenic mice, 90% of nuclei exhibiting chromatin condensation were also TUNEL positive.
- Demonstrated that DNA fragmentation and chromatin condensation occur concurrently in vivo.
Conclusions:
- DNA fragmentation and chromatin condensation are concomitant events during apoptosis in retinal neurons.
- These findings support the role of an endogenous endonuclease in the apoptotic death of retinal neurons.
- Provides a high-resolution ultrastructural confirmation of apoptotic processes in the retina.
