Abstract
Background and Purpose
Cerebral microbleeds (CMBs) due to cerebral amyloid angiopathy generally occur in lobar regions, while those due to hypertensive vasculopathy are deep. Inflammation may be an underlying mechanism for CMB, with varying associations according to CMB location. Lipoprotein phospholipase-A2 (Lp-PLA2) is a circulating enzyme marker of vascular inflammation associated with risk of ischemic stroke and dementia. We hypothesized that higher Lp-PLA2 levels would be related to higher prevalence of CMBs, with possible regional specificity.
Methods
Framingham Offspring participants aged ≥65 years with available Lp-PLA2 measures and brain MRI were included. Logistic regression models were used to relate Lp-PLA2 activity and mass to presence of CMBs, adjusted for age, sex, medication use (aspirin, anticoagulants, and statins), systolic blood pressure, APOE, current smoking, and diabetes.
Results
819 participants (mean age 73 years; 53% women) were included; 106 (13%) had CMBs; 82 (10%) lobar and 27 (3%) deep. We did not observe significant associations of CMB and LpPLA2 measures in multivariable adjusted analyses. However, there was a significant interaction between APOE genotype and Lp-PLA2 activity in their relation to presence of deep CMBs (p-interaction=0.01). Among persons with APOE ε3/ε3, the OR for deep CMB was 0.95 [0.59–1.53; p=0.83], while among those with at least one ε2 or ε4 allele, OR=3.46 [1.43–8.36; p=0.006].
Conclusions
In our community-based sample of older adults, there was no significant association of Lp-PLA2 with total or lobar CMBs. The association of higher levels of Lp-PLA2 activity with deep CMBs among those with at least one APOE ε2 or ε4 allele merits replication.
Introduction
Cerebral microhemorrhages (CMB) detected on brain MRI are implicated as a risk factor for clinical outcomes such as stroke, dementia and cognitive impairment.1–3 Although CMB in different locations have the same MRI appearance, CMB in lobar regions are generally attributed to cerebral amyloid angiopathy (CAA) and those in deep regions to hypertensive vasculopathy.4, 5 Recent studies suggest an association of inflammation to both CAA and hypertension affecting the cerebral vessels.6 Inflammation has been related to CMB7 and diseases with high prevalence of CMB, such as stroke and dementia.8 We selected a specific marker of vascular inflammation, Lipoprotein phospholipase A2 (Lp-PLA2) and studied its relation to CMB, testing the hypothesis that higher Lp-PLA2 levels are associated with a greater prevalence of CMBs, and that the association would be stronger for lobar CMBs.
Materials and Methods
Framingham Offspring participants (baseline characteristics; Table 1) who attended the 7th Offspring examination (1999–2001), had blood drawn at this examination for Lp-PLA2 mass and activity assay, and subsequently underwent brain MRI with gradient-echo sequences were included. Only participants above age 65 years were included since the prevalence of CMB is known to be low in younger persons.9 The Institutional Review Board of Boston University Medical Center approved the study protocol and informed consent was obtained from all subjects. Inclusion and exclusion criteria, details of brain MRI protocol, CMB detection methods and reproducibility measures, and lipoprotein phospholipase A2 measurements are provided on the online supplement (see http://stroke.ahajournals.org). CMBs were defined using standard criteria recently published.10
Table 1.
Characteristics of study sample (N=819)
| No CMB (n=713) | CMB | |||
|---|---|---|---|---|
| All CMB (n=106) | Lobar (n=82) | Deep (n=27) | ||
| Clinical Characteristics | ||||
| Men, n (%) | 331 (46.4) | 58 (54.7) | 47 (57.3) | 14 (51.9) |
| Age at Exam 7, yrs, mean [SD] | 67 (6) | 69 (6) | 69 (5) | 71 (5) |
| Age at MRI, yrs, mean [SD] | 73 (6) | 75 (5) | 75 (5) | 76 (6) |
| Time interval between Exam 7 and MRI, yrs, mean [SD] | 5.9 (2.3) | 6.0 (2.2) | 6.1 (2.1) | 5.7 (2.2) |
| Systolic blood pressure, mm Hg, mean [SD] | 129 (18) | 134 (19) | 132 (16) | 138 (18) |
| HDL cholesterol (mg/dL) | 54 (17) | 50 (17) | 49 (17) | 52 (17) |
| LDL cholesterol (mg/dL) | 120 (31) | 120 (36) | 120 (37) | 126 (37) |
| Diabetes mellitus, n (%) | 82 (11.5) | 13 (12.3) | 9 (11.0) | 4 (14.8) |
| Current smokers, n (%), | 53 (7.4) | 9 (8.5) | 6 (7.3) | 1 (3.7) |
| Hypertension, n (%) | 351 (49.2) | 66 (62.3) | 48 (58.5) | 22 (81.5) |
| Hypertension treatment, n (%) | 255 (35.8) | 50 (47.2) | 37 (45.1) | 16 (59.3) |
| Aspirin use, n (%) | 274 (38.5) | 49 (46.2) | 37 (45.1) | 15 (55.6) |
| Anticoagulant use, n (%) | 10 (1.4) | 3 (2.8) | 3 (3.7) | 2 (7.4) |
| Statin use, n (%) | 142 (19.9) | 34 (32.1) | 28 (34.2) | 9 (33.3) |
| APOE Status, n (%) | ||||
| ε3/ε3 | 440 (62.7) | 67 (64.4) | 51 (63.8) | 21 (77.8) |
| ε2/ε2- ε2/ε3- ε2/ε4- ε3/ε4- ε4/ε4 | 262 (37.3) | 37 (35.6) | 29 (36.3) | 6 (22.2) |
| Lipoprotein Phospholipase A2 measures (LpPLA2) | ||||
| LpPLA2 activity nmol/mL/min, mean [SD] | 143 (34) | 145 (36) | 147 (35) | 152 (48) |
| LpPLA2 mass ng/mL, mean [SD] | 293 (91) | 292 (86) | 295 (83) | 315 (113) |
Abbreviations: HDL, high density lipoprotein; LDL, low density lipoprotein.
Multiple logistic regression analyses were used to relate Lp-PLA2 activity and mass to presence of CMBs, overall and stratified by CMB location. All models were adjusted for age, sex and time interval between exam 7 and brain MRI acquisition. Secondary models were additionally adjusted for systolic blood pressure, diabetes, current smoking, statin use, aspirin use, anticoagulants, and APOE status.
We tested for interaction with the APOE genotype, and with antihypertensive medication use in the association between LpPLA2 measures and presence or absence of CMB (online supplement see http://stroke.ahajournals.org).
Results
CMB prevalence was 13% (106/819) with most participants having single CMB, and CMB located more frequently in lobar regions (online supplement see http://stroke.ahajournals.org).
We did not observe a significant association of LpPLA2 measures and CMB (online supplement Table 2) and the relation did not change according to CMB location, or additional adjustment for LDL and HDL cholesterol levels.
We observed a significant interaction between APOE genotype and Lp-PLA2 activity in their relation to presence of deep CMBs (p-interaction=0.01). Among persons with APOE ε3/ε3, the multivariable-adjusted OR for deep CMB was 0.95 [95% CI 0.59–1.53; p=0.83], while among those with at least one ε2 or ε4 allele, the multivariable adjusted OR was 3.46 [95% CI 1.43–8.36; p=0.006] (Table 2). No significant interaction was observed with antihypertensive medication use.
Table 2.
Interaction between APOE genotype and Lp-PLA2 measures in relation to presence of CMB
| APOE Group | N | LpPLA2 Activity | LpPLA2 Mass | |||||
|---|---|---|---|---|---|---|---|---|
| OR (95% CI) | P-value | P-interaction† | OR (95% CI) | P-value | P-interaction† | |||
| Any CMB | ||||||||
| Model 1* | 33 | 67 | 0.91 (0.69–1.20) | 0.50 | 0.31 | 0.90 (0.69–1.18) | 0.46 | 0.42 |
| 22,23,24,34,44 | 37 | 1.14 (0.82–1.58) | 0.44 | 1.07 (0.77–1.49) | 0.67 | |||
| Model 2** | 33 | 67 | 1.00 (0.75–1.33) | 0.98 | 0.42 | 0.95 (0.72–1.26) | 0.73 | 0.47 |
| 22,23,24, 34,44 | 37 | 1.09 (0.78–1.52) | 0.60 | 1.00 (0.70–1.41) | 0.98 | |||
| Lobar | ||||||||
| Model 1* | 33 | 51 | 1.01 (0.74–1.37) | 0.97 | 0.72 | 0.94 (0.70–1.28) | 0.71 | 0.47 |
| 22,23,24, 34,44 | 29 | 1.10 (0.76–1.58) | 0.62 | 1.11 (0.77–1.59) | 0.59 | |||
| Model 2** | 33 | 51 | 1.14 (0.82–1.59) | 0.44 | 0.85 | 1.01 (0.74–1.37) | 0.95 | 0.53 |
| 22,23,24, 34,44 | 29 | 1.06 (0.73–1.53) | 0.76 | 1.04 (0.71–1.52) | 0.86 | |||
| Deep | ||||||||
| Model 1* | 33 | 21 | 0.89 (0.57–1.40) | 0.62 | 0.01 | 1.03 (0.67–1.59) | 0.88 | 0.20 |
| 22,23,24, 34,44 | 6 | 2.76 (1.33–5.73) | 0.007 | 1.70 (0.92–3.12) | 0.09 | |||
| Model 2a*** | 33 | 21 | 0.95 (0.59–1.53) | 0.83 | 0.01 | 1.08 (0.70–1.66) | 0.74 | 0.22 |
| 22,23,24, 34,44 | 6 | 3.46 (1.43–8.36) | 0.006 | 1.78 (0.92–3.45) | 0.09 | |||
N=number of CMB. Odds Ratio (OR) of CMB per standard deviation change in LpPLA2 activity (1 SD=31 for women, 33 for men) and mass (1 SD=84 for women, 96 for men).
Interaction between LpPLA2 and APOE group.
Model 1 adjusts for age at MRI, sex, and time interval between exam 7 and MRI.
Model 2 additionally adjusts for aspirin use, anticoagulant use, statin use, diabetes status, smoking status, systolic blood pressure.
Model 2a adjusts for age at MRI, sex, time between exam 7 and MRI, aspirin use, anticoagulant use, statin use, systolic blood pressure.
Discussion
The present study explores for the first time the relation of LpPLA2 with cerebral microhemorrhages detected on brain MRI. No significant association of LpPLA2 measures with CMB presence was seen in multivariable adjusted models. There was a significant interaction with APOE status such that in participants who were carriers of at least one APOE ε2 or ε4 alleles, higher LpPA2 activity was related to the presence of deep CMB. However, as this was an interaction of only Lp-PLA2 activity (not mass) in the small subgroup of deep CMBs further study is required.
LpPLA2 is a recognized marker of vascular inflammation and cardiovascular risk, which has been related to ischemic small vessel disease detected on brain MRI.11 Our results did not support a role for LpPLA2 measures in hemorrhage prone small vessel disease represented by CMB when taken all together or with lobar CMB alone, but suggested a possible relation of LpPLA2 activity and deep CMB in persons with an APOE ε2 or ε4 allele. Although we cannot draw firm conclusions based on our data given the small sample size of participants with deep CMB, if replicated in other studies, this finding would suggest that LpPLA2 may not be a marker for CAA related CMB, but a more-specific systemic marker of vascular inflammation related to hypertensive vasculopathy, consistent with LpPLA2 associations with cardiovascular risk factors.12 It would also support a role for APOE as modifier of the relation of inflammation due to vascular risk factors and risk of CMB, similar to what has been reported in relation to clinical ICH.13, 14 APOE may also modify CMB risk via alternate mechanisms such as endothelial dysfunction implicated in hypertensive vasculopathy, and Lp-PLA2 has been related to endothelial dysfunction.15
The present study is limited by its cross-sectional nature, assessing MRI and LpPLA2 measures at only a single time-points (online supplement see http://stroke.ahajournals.org). The predominant white, European descent of Framingham Heart Study participants limits generalization to other groups, and the small sample size of participants with the deep CMB does not allow excluding that chance accounts for the findings.
In conclusion, we did not observe a significant association of Lp-PLA2 with total or lobar CMBs. The interaction of APOE status and the relation of LpPLA2 activity to CMB, with higher LpPLA2 activity in participants with CMB in deep cerebral regions needs to be interpreted with caution and further studies are required. If it is confirmed by others, LpPLA2 measures may potentially contribute to development of risk models for hypertensive hemorrhage, support mechanistic effects of LpPLA2 and hypertension on cerebrovascular integrity; both elevated LpPLA2 and hypertension are potentially modifiable conditions.
Supplementary Material
Acknowledgments
We thank Drs Steven Greenberg and Anand Viswanathan for their collaboration in the training of the main reader of MRI data (JRR) in CMB detection using current definitions.
Sources of funding:
This work (design and conduct of the study, collection and management of the data) was supported by the Framingham Heart Study’s National Heart, Lung, and Blood Institute contract (N01-HC-25195) and by grants from the National Institute of Neurological Disorders and Stroke (R01 NS17950), the National Institute on Aging (R01 AG16495; AG08122; AG033193; AG031287; K23AG038444) and NIH grant (1RO1 HL64753; R01 HL076784; 1 R01 AG028321).
Dr. Romero was also supported by grant award from GlaxoSmithKline protocol WEUSRTP3833.
Lp-PLA2 activity measurements were provided by GlaxoSmithKline and mass measurements by diaDexus at no cost to the FHS.
Footnotes
Conflict of Interest Disclosures:
Dr Irizarry is a stock and options holding employee of GlaxoSmithKline. Dr Romero received funding grant award from GlaxoSmithKline protocol WEUSRTP3833. The current analysis was partially supported by GlaxoSmithKline.
Statistical analyses: by Sarah R. Preis and Alexa S. Beiser, academic
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