E. *The P value of correlation between regional GMV and age less than.05; Abbreviations: MNI, Montreal Neurological Institute. doi:10.1371/journal.pone.0056663.tbe associated with Bcl-2 expression, differences in Bcl-2 expression levels among the Bcl-2 rs956572 allelic variants may influence the age-related rates of GM volume decline in these regions. Based on our findings, the Bcl-2 rs956572 polymorphism has the most prominent effect on age-related GM volume reductions in the cerebellum. Significant interconnections of the cerebellum with the hippocampus and the occipital and temporal regions of the cerebral cortex have been implicated in the integration of sensory information, visuospatial organization, visual memory, procedural learning, and the control of behavior and motivation [52?6]. Because the cerebellum may have extensive outgoing 
connections to these regions, Bcl-2 rs956572 polymorphism may indirectly modulate GM volume reduction in the lingual gyrus, the middle temporal gyrus, and the parahippocampal gyrus through direct impacts on the cerebellum. In our study, the age-related reduction in GM volume in the frontal and parietal lobes were not associated with Bcl-2 genotype. Although Bcl-2 expression is widespread in all brain regions, the effect of Bcl-2 expression on the trajectory of maturation or degeneration during brain aging may vary considerably in the cortex [50]. Analysis of post-mortem brain samples from patients with Alzheimer disease showed that the level of Bcl-2 expression were significantly higher in the cerebellum than in the frontal lobe [57]. Therefore, the effect of the Bcl-2 genotype on age- or neuropsychiatric disease-related changes in regional GM volumes warrants further investigation. The need for statistically sufficient sample sizes in imaging studies of genetic variation has become increasingly recognized. The relatively large and, by international standards, homogenous sample of participants that were reviewed in our study lend credibility to our findings, based on previously proposed recommendations regarding cohort sizes [58]. However, the cross-sectional nature of our study design may represent a limitation to our findings. Prospective studies have demonstrated HIV-RT inhibitor 1 greater sensitivity for clarifying the GM volume changes in specific brain regions during the aging process [59]. In addition, it is possible that, rather than having a direct effect of GM volume, the Bcl-2 rs956572 polymorphism may be in linkage disequilibrium with the truly associated allele. Such linkage likely varies among different populations, which would confound the generalization of findings based on a homogenous Chinese cohort, such as ours. Furthermore, the addition of a clinical control group with a psychiatric disorder, such as bipolar disorder, to future study designs may yield added knowledge of the dual role of Bcl-2 in aging and disease states. In conclusion, our findings of the effects of Bcl-2 rs956572 polymorphism on age-related KS-176 site morphologic changes in the brain indicate that Bcl-2 G homozygosity confers a protective effect against age-related GM volume reduction in several brain regions, particularly in the cerebellum. Although the underlying molecular mechanisms remain unclear, our findings support the hypothesis that Bcl-2-related genetic factors play a critical role in the effects of aging in the brain.AcknowledgmentsWe thank Ms Ashley for English editing.Author ContributionsConceived and designed the experiments: M.E. *The P value of correlation between regional GMV and age less than.05; Abbreviations: MNI, Montreal Neurological Institute. doi:10.1371/journal.pone.0056663.tbe associated with Bcl-2 expression, differences in Bcl-2 expression levels among the Bcl-2 rs956572 allelic variants may influence the age-related rates of GM volume decline in these regions. Based on our findings, the Bcl-2 rs956572 polymorphism has the most prominent effect on age-related GM volume reductions in the cerebellum. Significant interconnections of the cerebellum with the hippocampus and the occipital and temporal regions of the cerebral cortex have been implicated in the integration of sensory information, visuospatial organization, visual memory, procedural learning, and the control of behavior and motivation [52?6]. Because the cerebellum may have extensive outgoing connections to these regions, Bcl-2 rs956572 polymorphism may indirectly modulate GM volume reduction in the lingual gyrus, the middle temporal gyrus, and the parahippocampal gyrus through direct impacts on the cerebellum. In our study, the age-related reduction in GM volume in the frontal and parietal lobes were not associated with Bcl-2 genotype. Although Bcl-2 expression is widespread in all brain regions, the effect of Bcl-2 expression on the trajectory of maturation or degeneration during brain aging may vary considerably in the cortex [50]. Analysis of post-mortem brain samples from patients with Alzheimer disease showed that the level of Bcl-2 expression were significantly higher in the cerebellum than in the frontal lobe [57]. Therefore, the effect of the Bcl-2 genotype on age- or neuropsychiatric disease-related changes in regional GM volumes warrants further investigation. The need for statistically sufficient sample sizes in imaging studies of genetic variation has become increasingly recognized. The relatively large and, by international standards, homogenous sample of participants that were reviewed in our study lend credibility to our findings, based on previously proposed recommendations regarding cohort sizes [58]. However, the cross-sectional nature of our study design may represent a limitation to our findings. Prospective studies have demonstrated greater sensitivity for clarifying the GM volume changes in specific brain regions during the aging process [59]. In addition, it is possible that, rather than having a direct effect of GM volume, the Bcl-2 rs956572 polymorphism may be in linkage disequilibrium with the truly associated allele. Such 
linkage likely varies among different populations, which would confound the generalization of findings based on a homogenous Chinese cohort, such as ours. Furthermore, the addition of a clinical control group with a psychiatric disorder, such as bipolar disorder, to future study designs may yield added knowledge of the dual role of Bcl-2 in aging and disease states. In conclusion, our findings of the effects of Bcl-2 rs956572 polymorphism on age-related morphologic changes in the brain indicate that Bcl-2 G homozygosity confers a protective effect against age-related GM volume reduction in several brain regions, particularly in the cerebellum. Although the underlying molecular mechanisms remain unclear, our findings support the hypothesis that Bcl-2-related genetic factors play a critical role in the effects of aging in the brain.AcknowledgmentsWe thank Ms Ashley for English editing.Author ContributionsConceived and designed the experiments: M.
