We applied novel voxel-wise VBM and TBSS analyses to assess the relationship of APOEε4 genetic variation on brain structure in healthy elderly subjects. We found decreased medial temporal gray matter volume and global cognitive performance in carriers of the ε4 allele. APOE is the only established genetic risk factor for late-onset AD [8
], and carriers of the ε4 allele, despite being nondemented, express small measurable endophenotypes of brain structure and function that are similar to individuals with AD.
We found that those cognitively intact, nondemented subjects with an ε4 allele had significantly decreased global cognitive performance compared to APOEε4 negative participants. Interestingly, this decrease was primarily due to ε4-carriers performing significantly worse on Logical Memory I and Letter Number Sequencing and a trend for decreased performance in Delayed Logical Memory. Poorer performance on Logical Memory I has been reported to be an indicator of risk for AD in healthy subjects [64
] and both Logical Memory and Letter Number Sequencing have been noted as markers of early dysfunction in AD [65
]. While presence of an ε4 allele along with these deficits may mark risk for AD, decline in memory performance and hippocampal volume are also commonly observed in normal aging [67
Some have argued that morphological changes in ε4 carriers, such as decreased hippocampal volume, represent brain changes associated with preclinical AD [7
]. Modest APOEε4-related brain volume difference was observed in regions known to be affected in AD, namely the medial temporal cortex, including the hippocampus and amygdala. Decreased hippocampal volume is an early feature of AD and most likely occurs several years before the onset of symptoms [68
], perhaps even in childhood for those carrying an APOEε4 allele [69
]. AD subjects with an APOEε4 allele have lower hippocampal [10
] and amygdala [12
] gray matter volumes. Past VBM studies reported reduced entorinal and subiculum cortical thickness in nondemented ε4 carriers [70
] and decreased hippocampal gray matter density in ε4 homozygotes (but not heterozygotes) relative to noncarriers [23
This study demonstrates ε4-related decreased hippocampal and amygdala volume in nondemented elderly men and women who have been carefully screened for evidence of clinical change using sensitive diagnostic methods (CDR). ε4-carriers also had a trend for lower GM regional volumes in the left parahippocampal gyrus, right inferior parietal cortex, left superior frontal cortex, left angular gyrus, and right precuneus. Decreased gray matter volume in the parahippocampal gyrus, posterior temporal (supramarginal gyrus and angular gyrus), inferior parietal cortex have been reported in early AD subjects compared to healthy controls [53
]. However, our current, cross-sectional findings cannot be interpreted to be either a direct manifestation of the disease process or predisposition to later development. Longitudinal follow-up will determine how these morphological differences represent gray matter changes that may lead to a disease state.
We identified a trend for regional volume increases in ε4 carriers compared to non-carriers, primarily in the frontal cortex. A number of studies have noted increased frontal lobe volume [10
] as well as increased executive function in APOEε4 carriers with AD [76
], although the same has not been reported in nondemented subjects. Regionally increased brain volume in nondemented ε4 carriers may represent preservation of brain health, or alternate protective genetic or environmental mechanisms at work. Indeed, our sample of elderly ε4-carriers are nondemented despite the high risk for AD imparted by the ε4-allele. ε4 carriers typically manifest AD at an earlier age than our present cohort. Alternatively, it is possible these results represent a false positive. Given that our hippocampal cluster extended into subgyral space, the SVC did not reveal a significant increase in this region, and lacking a mechanistic model for increased volume, this possibility cannot be ruled out in the present study. It is possible that our individuals selected out for being nondemented are a residual group of ε4-carriers not fully representative of the population of ε4-carriers. Nevertheless, the present data suggest that despite the impact of the ε4 allele on cognitive and imaging measures in healthy elderly, there are mediating protective mechanisms preventing disease onset in this particular group.
As a post-hoc analysis we extracted the cluster of significantly decreased volume in carriers of the ε4 allele and tested if various factors related to lifestyle, physical activity or cardiorespiratory fitness were related to APOEε4 related brain volume decreases. None of these factors significantly covaried with change in brain morphology associated with the APOE gene. There may be other factors influencing brain volume changes that we have not controlled for such as other genes or medication, and future studies looking at epistasis between interacting gene variants of interest may shed light on more complex genetic mechanisms at work. Several studies have linked APOE risk effects to lifestyle [77
], and while our study consisted of a variety of lifestyle measures, these measures and our sample size may have limited sensitivity for identifying this type of geneenvironment interaction.
We found that APOEε4 genetic variation may be associated with disrupted white matter integrity in the parahippocampal gyrus of healthy elderly subjects. A large number of studies have found decreased white matter strength in posterior temporal regions, including parahippocampal tracts, in subjects with AD [78
]. AD-related hippocampal atrophy has been correlated with abnormalities in the cingulum bundle fiber tract [84
], which mainly includes fibers connecting the parahippocampal gyrus and hippocampus proper to the posterior cingulate cortex [86
]. Moreover several DTI studies in nondemented subjects have reported a significant relationship between the APOEε4 allele and decreased white matter integrity in temporo-parietal regions [25
]. A recent DTI study focused on the parahippocampal gyrus and found decreased FA in healthy carriers of the ε4 allele, similar to our own results [26
]. Another region of interest study found decreased FA in the posterior corpus callosum of individuals carrying a copy of the APOEε4 allele [25
]. Taken together, these results and ours suggest that genetic variation in APOEε4 may influence disease-related white matter integrity in posterior white matter tracts, especially those branching from the medial temporal cortex.
TBSS was created to alleviate alignment-related problems in DTI data, and thus restricts the analysis to a group-estimated skeleton strictly of white matter. Ideally this would increase the power of the analysis to detect the small fractional anisotropy changes that might occur in a cognitively healthy group. Although it is possible that restriction in white matter to the FA skeleton could increase the chance of a Type II error, a recent study using TBSS in AD subjects reported disease-related FA decreases at a statistical threshold similar to our own in the parahippocampal gyrus [88
]. Usage of the FMRIB58_FA standard template might also introduce bias through the registration process, due to the age discrepancy between the template subjects and the age of our subjects. It is also possible that TB-SS could be vulnerable to detecting the vascular burden of the white matter, and that this, in turn, is affecting the APOEε4 results in the white matter. While TBSS confines analyses to a narrow skeleton within the white matter tracts of greatest integrity, the presence of white matter lesions may have influenced the results. We excluded individuals with a history of clinical stroke, diabetes, and cardiovascular disease, minimizing the potential influence of vascular-related changes.
Overall we sought to use automated morphological analysis techniques to characterize the genetic variation in APOE associated with both gray matter volume and white matter viability in carefully screened, non-demented elderly subjects. Using voxel-based analyses in several imaging modalities, we validate previous studies identifying decreased hippocampal and amygdala volume in APOEε4 carriers and decreased FA in the parahippocampal gyrus in one group of subjects. We also identify selective decreases in memory-related cognitive performance in otherwise healthy elderly individuals carrying the ε4 allele. Our finding of a trend for increased frontal gray matter associated with the risk allele sets the groundwork for future studies to identify potential protective mechanisms at work in individuals at risk for AD. More complex approaches to characterizing genetic epistasis using these comprehensive, unbiased image analyses in a larger group, along with relevant environmental, lifestyle, or other genetic measures may inform risk for the disease as well as the mechanisms involved in the disease pathology.