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[Effect of CCM3 gene defect on lead-induced cell genotoxicity in mouse embryonic fibroblasts]
Xiaolin Su1, Xiumei Xing, Guanchao Lai
1Department of Toxicology, School of Public Health, Sun Yat-sen University of Medical Sciences, Guangzhou 510080, China.
Objective:
To investigate the effect of CCM3 gene defection on lead induced cell genotoxicity in mouse embryonic fibroblasts.
Methods:
C57 female mice were mated with CCM3 gene heterozygous male mice. E13.5 embryos were taken to isolate primary mouse embryonic fibroblasts. After genotyping, wild type and heterozygous cells were treated with different doses of lead acetate. Cell viability, genotoxicity and protein expression were detected by MTS assay, CB micronucleus method and Western blot, respectively.
Results:
Mouse embryonic fibroblasts with lead acetate treatment for 24 h, wild-type cells 100.00 µmol/L lead acetate-treated group (69.16±1.36) and the control group (100.00±2.33) compared to cells decreased by 30%, CCM3 heterozygous type cell 100.00 µmol/L lead acetate-treated group (87.16±5.50) and the control group (100.00±2.06) compared to cells decreased by 13%, the difference was statistically significant (F values were 98.59, 82.63, P<0.001). Lead acetate treatment after 48 h, wild-type cells 100.00 µmol/L lead acetate-treated group (51.99±5.62) and the control group (100.00±3.11) compared to cells decreased by 50%, heterozygous type cells 100.00 µmol/L lead acetate treatment group (66.33±4.06) and the control group (100.00±5.72) compared to cells decreased by 35%, the differences were statistically significant (F values were 82.63, 36.86, P < 0.001). The results of CBMN test showed that with increased dose, micronucleus cell rate of two genotypes showed an increasing trend, in the wild-type cells, the micronucleus cell rate (/1 000) for the control group, 29.6±2.2, 6.25 µmol/L dose group 47.3±6.6, 25 µmol/L dose group 55.5±9.1, 100.00 µmol/L dose group 66.8±3.5; heterozygous cells micronucleus cell rate (/1 000) for the control group, 35.3±5.6, 6.25 µmol/L dose of 50.0±8.3, 25.00 µmol/L dose group 57.0±8.5, 100.00 µmol/L dose group 58.8±2.1. Micronucleus cell rates (/1 000) were significant differences, in 100.00 µmol/L dose groups of two genotypes. Western blot results showed that wild-type cells CCM3 expression 100.00 µmol/L lead acetate-treated group (0.70±0.03) was 1.32 times higher than the control group (0.53±0.07), heterozygous cells CCM3 expression 100.00 µmol/L lead acetate-treated group (0.48±0.02) was 1.77 times higher than control group that of 0.27±0.04, there was statistically significant difference (F values were 14.77, 25.74, P < 0.001); wild-type cells γ-H2AX expression 100.00 µmol/L lead acetate-treated group (0.69±0.03) was 1.06 times higher than the control group (0.65±0.07), heterozygous cells γ-H2AX expression 100.00 µmol/L lead acetate-treated group (0.99±0.04) was 1.55 times higher than the control group CCM3 expression levels (0.64±0.06), there was statistically significant difference (wild-type cells: F = 7.08, P = 0.012, heterozygous type cell: F = 13.49, P = 0.002).
Conclusion:
CCM3 gene may play a role in lead-induced genetic toxicity of mouse embryonic fibroblasts, CCM3 gene-lead interactions effects on mouse embryonic fibroblasts cell toxicity.
Insights
CCM3 gene deficiency increases lead-induced genotoxicity in mouse embryonic fibroblasts. This suggests CCM3 plays a protective role against lead
Area of Science:
- Genetics
- Toxicology
- Cell Biology
Background:
- Lead exposure is a significant environmental health concern, known to induce cellular damage.
- Genetic factors can influence an individual's susceptibility to environmental toxins.
- The role of the CCM3 gene in response to heavy metal toxicity is not well understood.
Purpose of the Study:
- To investigate the impact of CCM3 gene deficiency on lead-induced genotoxicity.
- To determine if CCM3 gene status affects cell viability and DNA damage in response to lead.
- To analyze CCM3 and DNA damage marker protein expression under lead exposure.
Main Methods:
- Primary mouse embryonic fibroblasts were isolated from wild-type and CCM3 heterozygous embryos.
- Cells were exposed to varying concentrations of lead acetate.
- Cell viability (MTS assay), genotoxicity (CBMN assay), and protein expression (Western blot) were assessed.
Main Results:
- Lead acetate significantly reduced cell viability in both genotypes, with a more pronounced effect in wild-type cells after 48 hours.
- The frequency of micronuclei, an indicator of genotoxicity, increased with lead dose in both cell types, with higher rates in wild-type cells at the highest dose.
- CCM3 expression was upregulated by lead exposure in both genotypes, while γ-H2AX (a DNA damage marker) expression increased significantly, particularly in heterozygous cells.
Conclusions:
- CCM3 gene deficiency exacerbates lead-induced genotoxicity in mouse embryonic fibroblasts.
- CCM3 may play a protective role against lead-induced cellular damage.
- The interaction between the CCM3 gene and lead exposure influences cellular toxicity.
