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Micronucleus test using cultured new born rat astrocytes
Y Miyakoshi1, Y Suzuki, M Ooida
1Department of Public Health and Environmental Medicine, Jikei University School of Medicine, Tokyo, Japan.
Abstract:
Micronuclei is induced in cytoplasm as a consequence of the formation of chromosomal fragments or remaining chromosomes during cell division by the cause of clastogens or spindle poisons, and is used as an indicator of genotoxicity screening tests. There are few short-term genotoxicity screening tests using brain cells. We attempted to establish a new in vitro micronucleus test (MN test) system by use of central nervous system cells. Primary cultured astrocytes were prepared from newborn male Sprague-Dawley (SD) rats. In growth curve of astrocytes, doubling time was determined to be 31 h. In time study, the highest frequency of micronuclei was observed at 48 h, 72 h and 6 h-exposure-66 h-recovery by vincristine (VCR), mitomycin C (MMC) without metabolic activation system and cyclophosphamide (CPM) with metabolic activation system, respectively. Dose-response relationships between micronucleus frequency and concentrations of MMC, VCR and CPM were observed, respectively. It is suggested that the in vitro MN test using new born rat-astrocytes could be used as a screening test of environmental and occupational genotoxic chemicals in the central nervous system cells.
Insights
This study introduces a new in vitro micronucleus test using rat astrocytes to screen for genotoxic chemicals. This method assesses potential damage to central nervous system cells from environmental and occupational exposures.
Area of Science:
- Toxicology
- Cell Biology
- Neuroscience
Background:
- Micronuclei formation indicates genotoxicity from chromosomal damage during cell division.
- Existing short-term genotoxicity tests have limited application to brain cells.
- There is a need for in vitro tests assessing central nervous system (CNS) cell exposure.
Purpose of the Study:
- To establish a novel in vitro micronucleus (MN) test system using primary cultured astrocytes from Sprague-Dawley rats.
- To evaluate the efficacy of this system for detecting genotoxic chemicals targeting CNS cells.
Main Methods:
- Primary astrocytes were cultured from newborn male Sprague-Dawley rats.
- Astrocyte growth kinetics were analyzed, determining a doubling time of 31 hours.
- Time-course and dose-response studies were conducted using genotoxic agents like vincristine, mitomycin C, and cyclophosphamide.
Main Results:
- The highest micronuclei frequency was observed at specific exposure-recovery times for each tested chemical.
- A clear dose-response relationship was established between chemical concentration and micronuclei frequency.
- The in vitro MN test demonstrated sensitivity to genotoxic agents without requiring a metabolic activation system for some chemicals.
Conclusions:
- The developed in vitro micronucleus test using rat astrocytes is a viable screening tool.
- This system can be used to identify genotoxic chemicals impacting central nervous system cells.
- It offers a promising method for assessing environmental and occupational genotoxic risks to the CNS.