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Published online before print November 8, 2007, 10.1097/PSY.0b013e31815a8ba8
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Psychosomatic Medicine 69:910-917 (2007)
© 2007 American Psychosomatic Society

ORIGINAL ARTICLES

Combination of Caregiving Stress and Hormone Replacement Therapy is Associated With Prolonged Platelet Activation to Acute Stress Among Postmenopausal Women

Kirstin Aschbacher, MS, Roland von Känel, MD, Paul J. Mills, PhD, Suzi Hong, PhD, Susan K. Roepke, BA, Brent T. Mausbach, PhD, Thomas L. Patterson, PhD, Michael G. Ziegler, MD, Joel E. Dimsdale, MD, Sonia Ancoli-Israel, PhD and Igor Grant, MD

From the Department of Psychiatry (K.A., R.v.K., P.J.M., S.H., S.K.R., B.T.M., T.L.P., J.D., S.A.-I., I.G.), University of California, San Diego, San Diego, California; Department of General Internal Medicine (R.v.K.), University Hospital Bern, Switzerland; San Diego Veterans Affairs Healthcare System (T.L.P., S.A.-I., I.G.), La Jolla, California, USA; and Department of Medicine (M.G.Z.), University of California, San Diego, San Diego, California.

Address correspondence and reprint requests to Igor Grant, Department of Psychiatry, School of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, California, 92093-0680. E-mail: igrant{at}ucsd.edu


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 NOTES
 REFERENCES
 
Objective: To investigate the combined effects of caregiving and hormone replacement therapy (HRT) on platelet hyperactivity to acute psychological stress. Both HRT and the chronic stress of caregiving have been associated with increased cardiovascular risk, potentially through a mechanism of platelet hyperactivity.

Methods: A total of 78 elderly postmenopausal women (51 caregivers (CG) and 27 noncaregivers (NC)) were assessed for platelet activation in response to a laboratory speech test. Half the sample was taking HRT. Blood was sampled at baseline, post speech, and after 14 minutes of recovery. Platelet activation was assessed through whole blood flow cytometry assays of % aggregates (Agg), and expression of % fibrinogen receptors (FbR) and % P-selectin (P-sel) on platelet surface.

Results: Multivariate repeated-measures analysis of variance revealed that CG taking HRT exhibited significantly prolonged platelet activation in response to acute stress. There was an interaction between HRT and CG on recovery from stress for Agg (F (1,71) = 5.260, p = .025), P-Sel (F(1,71 = 6.426, p = .013), and FbR (F(1,71 = 6.653, p = .012), controlling for age, cardiovascular disease, and aspirin. Among HRT users, regression analysis revealed that CG had delayed recovery of Agg (ß = 0.354, t(34) = 2.154, p = .038) and P-sel (ß = 0.498, t(34)=3.126, p = .004) from stress relative to NC. No caregiving effects on recovery were present among non-HRT users. In addition, these effects were maintained after controlling for health behaviors, medications, and medical conditions.

Conclusion: Chronic dementia caregiving stress in combination with HRT may impair recovery of platelet activation after acute mental stress (i.e., activation levels do not quickly return to resting levels), thereby potentially increasing cardiovascular risk among CG who take HRT.

Key Words: P-selectin • fibrinogen receptor • GP IIb/IIIa • platelet aggregates • dementia • Alzheimer's disease

Abbreviations: HRT = hormone replacement therapy; CG = caregivers; NC = noncaregivers; Agg = % platelet aggregates; P-sel = % platelet P-selectin expression; FbR = % platelet fibrinogen receptor expression; CAM = cellular adhesion molecule; CVD = cardiovascular disease; CAD = coronary artery disease; UCSD = University of California, San Diego; GSI = Global Symptom Inventory; MANOVA = multivariate repeated-measures analysis of variance.


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
METHODS
 RESULTS
 DISCUSSION
 NOTES
 REFERENCES
 
Caring for a spouse with Alzheimer's disease is a significant chronic stressor, which can deleteriously affect cardiovascular health. Dementia caregivers (CG) exhibit impaired immune functioning (1), elevated hemostatic markers of coagulation (2,3), and increased cardiovascular risk (2,4,5), relative to noncaregiving counterparts (6). The considerable toll exacted by caregiving stress is illustrated by one study that found a 63% increase in total mortality among strained CG (7).

Platelet reactivity to acute mental stress, particularly a state of prolonged activation (i.e., delayed recovery from activation) (8), may be one mechanism by which stress is thought to increase the risk of cardiovascular events. Stressful events induce secretion of catecholamines, which, among myriad other platelet agonists, can provoke platelet activation in vivo (9). Given that CG exhibit greater sympathetic arousal (10) and hemostatic reactivity (3), it is plausible that chronically stressed CG may also exhibit elevated platelet reactivity to acute stress.

When platelets become activated, the platelet surface glycoprotein (GP) IIb/IIIa receptor undergoes a conformational change and exposes a binding site for soluble fibrinogen (11,12). Fibrinogen then cross-links platelets by bridging GP IIb/IIIa between adjacent platelets. Platelet stimulation also results in exposure of the granule membrane protein P-selectin on the platelet surface (13). These activation steps accelerate platelet aggregation (13–15), coagulation, and ultimately clot formation when thrombogenic material becomes exposed to the blood after rupture of an atherosclerotic plaque. Importantly, catecholamines may elicit expression of both the fibrinogen receptor and P-selectin on the surface of activated platelets (13,16). P-selectin is viewed as a cellular adhesion molecule (CAM) that facilitates platelet-leukocyte adhesion to and inflammation of the vascular endothelium (17–20). In sum, stress via catecholamine surge can initiate a cascade of platelet-related events, which may facilitate the formation of blood clots, inflammatory processes, and atherothrombotic events downstream.

Platelet agonists may potentially amplify the effects of one another. A body of research has proposed that catecholamines, adenosine diphosphate, and other substances may act in combination as "synergistic agonists," interacting to produce a hyperactive platelet response (9,21). These results suggest the possibility that stress may provoke hyperactive responses when platelets have already been "primed" by other cardiovascular disease (CVD) risk factors. A recent review of cardiovascular reactivity research suggested that the duration of the stress response may be potentially more relevant to cardiovascular outcomes than its intensity (8). Whereas healthy clotting responses are necessary to prevent excessive blood loss and facilitate wound healing, sustained prothrombotic activity may become deleterious.

Use of hormone replacement therapy (HRT) among postmenopausal women may increase CVD risk by elevating coagulation. The Women's Health Initiative, a large randomized trial of HRT among postmenopausal women, was prematurely terminated partly because HRT was found to relate to increased risk of coronary heart disease, stroke, and pulmonary embolism (22–24). These findings have been duplicated in other studies (25,26). Subsequent research suggests that HRT may be related to increased levels of platelet activation (27); however, other studies have reported opposing results (28,29). How can this apparent contradiction be understood? Reviewing a number of human and animal studies on the multifaceted effects of HRT on inflammation, hemostasis, and fibrinolysis, Koh and Yoon (30) proposed that the net effects are more likely to be harmful in the presence of other risk factors for hypercoagulability. For example, they demonstrated that HRT reduced CAM levels in healthy, but not diabetic, postmenopausal women (31,32). Others have reported increased risk of thromboembolism among women taking HRT after the stress of surgical or nonsurgical hospitalization (26). Although stress is a well-recognized psychosocial risk factor for CVD (33), to our knowledge, the potential interactive effects of HRT and chronic stress on platelet activation have not been examined.

Thus, the primary aim of this study was to investigate whether postmenopausal women caring for a spouse with Alzheimer's disease and taking HRT would exhibit hyperactive platelets relative to their peers not taking HRT or to a noncaregiving comparison group. In addition, it was hypothesized that CG might exhibit greater platelet activation to acute psychological stress than noncaregivers (NC) overall, and that HRT would be related to overall platelet hyperactivity.


   METHODS
TOP
 ABSTRACT
 INTRODUCTION
 METHODS
RESULTS
 DISCUSSION
 NOTES
 REFERENCES
 
Participants
These data were obtained between August 2001 and October 2005 as part of a longitudinal study investigating the effects of caregiving and psychological stress on cardiovascular physiology. Seventy-eight elderly women (52–87 years, mean = 69 years) participated in the study. Of these, 51 women were CG of a spouse with Alzheimer's disease, with whom they were living at the time of enrollment, whereas 27 women formed a comparison group of NC. On average, care recipients had received their Alzheimer's diagnosis 7.75 years ago (standard deviation (SD) = 2.49; range 4–15 years), and their average age was 77 years (SD = 6.8; range 59–90 years). The majority of participants were Caucasian (n = 74, 95%). Individuals taking ß blockers were excluded during recruitment in the larger study, given that ß blockade is known to activate many systems of interest in that study (e.g., the sympathetic nervous system) (16). One CG began taking clopidogrel, an antiplatelet medication, shortly after starting the study; none were taking ticlopidine or coumadin; and 17 were taking regular aspirin; thus, these medications were controlled for in the analyses. Additional information on participant medication use is shown in Table 1. Both caregiving and noncaregiving participants were recruited through community support groups, various health fairs, word of mouth/referrals, talks at senior groups, or referrals from the University of California, San Diego (UCSD) Alzheimer Disease Research Center. All volunteers provided written consent to participate in the study, which was approved by the UCSD Institutional Review Board.


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  		TABLE 1. Health Characteristics of Caregivers and Noncaregivers

 

Blood Draw Procedure and Stress Protocol
A laboratory test of psychological stress, previously shown to provoke stress reactivity (34–36), was used to examine in vivo platelet activation. Two trained research nurses administered the stress testing between 8 to 10 AM in the participants' homes. The stress task required participants to deliver a 3-minute speech on an assigned topic regarding an interpersonal dispute to the interviewer. Participants were randomly assigned to one of two topics involving either an accusation of a stolen belt (34) or a conflict with a disreputable automobile repairman over unreasonable costs (37). Blood was drawn using an indwelling 22-gauge venous forearm catheter (as used in previous studies of platelet activity) (38) at three time points: baseline (after 20 minutes of rest in a seated position), immediately post speech, and after 14 minutes of recovery. The first 2 ml of blood was discarded to prevent artificial coagulation activation due to the needle stick.

Measures
Hormone Replacement Therapy
Half of the participants (n = 39) reported taking oral or transdermal HRT (i.e., 22 took estrogen only; 13 took both estrogen and progesterone; two took progesterone/progestin only; one took oral estrogen plus testosterone; and one took an unknown HRT cream) at the time of the stress test. Fifteen of the 27 NC and 24 of the 51 CG reported taking HRT.

Platelet Measures and Assay Procedure
Platelet activation is not a dichotomous "on or off" state but rather a process engaged to differing degrees (11), where the presence of circulating aggregated platelets, combined with the expression of specific markers, suggests a higher degree of activation (15). Flow cytometry (39,40) was utilized to assess three parameters indicative of platelet activation: a) platelet aggregate formation (Agg), i.e., the percentage of platelets that had formed aggregates with other platelets or leukocytes; b) fibrinogen receptor expression (FbR), i.e., percentage of aggregates expressing FbR; and c) P-selectin expression (P-sel), i.e., percentage of aggregates expressing P-sel. Blood was preserved with acid-citrate-dextrose. Blood samples to be used for platelet flow cytometry were kept at room temperature. The antibody staining process was conducted on site within 10 minutes of sample collection by pipetting blood samples into prepared tubes with antibodies, followed by incubation and fixation. A whole blood method was employed to eliminate red blood cell lysis, centrifugation, and washing, further reducing the risk of artificial platelet activation from handling.

Staining procedure was as follows: 5 µl of whole blood was incubated with 30 µl of fluorochrome-conjugated antibodies (Becton Dickenson Immunocytometry Systems, San Jose, California) at saturating concentrations (predetermined) in the dark for 15 minutes at room temperature. The fluorochrome-conjugated antibodies, which bind membrane glycoproteins on the activated platelet surface, included CD61-PerCP (an activation-independent marker of platelets), PAC-1-FITC (an activation-dependent marker indicating the GP IIb/IIIa complex has been converted into a functional fibrinogen-binding receptor) (11), and CD62P-PE (a marker of platelet P-selectin). Samples were fixed with 1 ml 1% formaldehyde in PBS containing 0.1% NaN3 and analyzed within 24 hours of staining using a flow cytometer (Beckman Coulter EPICS Elite, Fullerton, California) equipped with Expo32 Software. Platelets were identified by forward and side scatter and distinguished from red blood cells by the presence of the CD61 antigen. Ten thousand events were collected per tube. Platelet aggregates were identified based on size discrimination using forward scatter and separately gated (11,15). FbR and P-Sel were then quantified as the percentage of the gated population of aggregates that expressed the given activation marker.

Participant Health
The self-reported medical histories of all participants were assessed during in-home interviews on a separate day from the blood draw. Three sets of factors potentially relevant to cardiovascular risk were formed: a) health behavior indices—smoking (never/former/current), alcohol intake (average number of drinks per week), physical activity (average number of days exercise per week), and body mass index; b) preexisting medical conditions—physician's diagnosis of cerebrovascular disease (previous stroke or transient ischemic attack), coronary artery disease (CAD), previous myocardial infarction, angina, coronary artery bypass surgery, other unspecified CAD), diabetes, hypertension, or hypercholesterolemia; and c) medication use—aspirin, other nonsteroidal anti-inflammatory drugs, antidepressants, antihypertensives, statins, or clopidogrel (antiplatelet).

Psychological Distress
The Global Symptom Inventory (GSI) from the 53-item Brief Symptom Inventory (41) was used to compare overall psychological distress across nine symptom dimensions among CG versus NC. Items were scored on a 5-point scale ranging from not at all (0) to extremely (4). The final score represents the sum of all items divided by the overall number of items to form an average score. Psychometric research suggests that the GSI provides a reasonable single index of general psychopathology (42,43). Data on psychological distress were not available for one NC and three CGs.

Data Analyses
Analyses were conducted in three stages: group comparisons and sample characteristics, multivariate repeated-measures analysis of variance (MANOVA) examining platelet reactivity to and recovery from acute stress, and follow-up multiple regression analyses of the simple effects. P-sel was log-transformed and FbR was square root-transformed to approximate normal distributions. Agg did not require transformation. Standardized regression coefficients and partial eta values ({eta}p2) are reported as estimates of effect size.

Before the primary analyses, Pearson correlations of Agg, P-sel, and FbR at all three time points were conducted to determine the extent to which these measures represent a unified construct of overall platelet activation. Each measure of activation correlated highly with itself at all time points (r = .55–0.84, all p < .001). The three measures were moderately correlated with one another in recovery (r = .25–.61, all p < .05). Given these interrelations, we tested our primary hypothesis in two ways: a single omnibus MANOVA with all activation measures entered simultaneously, and follow-up MANOVAs testing each of the three platelet outcomes in separate analyses.

All MANOVAs were conducted as repeated-measures analyses. All utilized "Time" as a within-subjects factor (3 levels [Baseline, Speech, Recovery] to represent change in platelet activation over time due to the acute psychological stress test). Caregiver Status (CG) and HRT (yes/no) were entered as between-subjects factors, controlling for age as a covariate. All interactions were included and were only reported when significant.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
DISCUSSION
 NOTES
 REFERENCES
 
Sample Characteristics and Group Comparisons
The proportion of participants taking HRT did not statistically differ between CG and NC. No significant age differences between HRT and non-HRT users were found. However, CG (mean = 70.33 years, SD) = 8.93) were significantly older than NC (mean = 66.67 years, SD = 5.55) (t(73.90) = –2.23, p = .03), using independent t tests not assuming equal variance. Thus, age was included as a covariate in all analyses. As expected, CG exhibited significantly greater symptoms of psychological distress than NC (t(71.62) = –2.38, p = .02). Scores for the GSI were higher among CG (mean = 0.51, SD = 0.46) compared with NC (mean = 0.32, SD = 0.23). We compared CG and NC on a number of health behavior indicators, preexisting medical conditions, or use of medications, and found CG were significantly more likely to have been diagnosed with CAD (Table 1).

MANOVAs
The study's primary hypothesis that caregiving and HRT would exert interactive effects on stress-induced platelet activation was supported by the significant three-way interaction term, CG x HRT x Time, (Wilk's {lambda}; F(2,72) = 3.327, p = .04, {eta}p2 = 0.085) in the omnibus MANOVA of overall platelet activation (described in Data Analysis). Follow-up specific contrast tests of Time revealed that the CG x HRT interaction was related to significant differences in recovery (F(1,73) = 6.623, p = .01, {eta}p2 = 0.083) but not reactivity. Additional omnibus and recovery contrast tests were also conducted in each separate platelet outcome (Table 2; Figures 1 to 3), revealing a similar pattern of results across all three measures. Significant main effects of HRT and speech reactivity were identified (not reported); however, common statistical practice (44) cautions against interpreting main effects in the presence of a higher order interaction. No significant effects for CG, CG x Time, Age or Age x Time on activation were identified.


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  		TABLE 2. MANOVA Three-Way Interaction Effects on Each Individual Platelet Outcome

 

Figure 116
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  		Figure 1. The figures depict profiles of platelet activation over time in response to an acute speech stressor task. Caregivers (CG) taking hormone replacement therapy (HRT) exhibited significant increases in platelet P-selectin expression and prolonged fibrinogen receptor expression in recovery. Non-caregivers taking HRT exhibited significant declines in the percentage of platelet aggregates at recovery ralative to CG taking HRT.

 


Figure 216
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  		Figure 2. The figures depict profiles of platelet activation over time in response to an acute speech stressor task. Caregivers (CG) taking hormone replacement therapy (HRT) exhibited significant increases in platelet P-selectin expression and prolonged fibrinogen receptor expression in recovery. Non-caregivers taking HRT exhibited significant declines in the percentage of platelet aggregates at recovery ralative to CG taking HRT.

 


Figure 316
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  		Figure 3. The figures depict profiles of platelet activation over time in response to an acute speech stressor task. Caregivers (CG) taking hormone replacement therapy (HRT) exhibited significant increases in platelet P-selectin expression and prolonged fibrinogen receptor expression in recovery. Non-caregivers taking HRT exhibited significant declines in the percentage of platelet aggregates at recovery ralative to CG taking HRT.

 
Health Behaviors, Medications, and Preexisting Medical Conditions
Relevant covariates for inclusion in subsequent analyses were determined by the following regression analyses. The effects of each of the three covariate sets (health behaviors, preexisting medical conditions, and medication use) (see Participant Health for details) on residualized recovery scores were tested in the total sample, at the same time controlling for age, CG, HRT, and CG x HRT. Only aspirin was significantly related to any measure of platelet activation. Additionally, we created a composite variable, "cardiovascular disease" (CVD—yes/no; marking the presence of any CVD condition), which was significantly related to activation. Clopidogrel (n = 1) and statins (n = 2) were dropped because of the small numbers of participants taking them, and the fact that their removal did not change the multivariate results.

Thus, when including age, aspirin, and CVD, CVD was associated with significantly elevated levels of FbR expression at recovery (ß = 0.492, t(71) = 2.142, p = .04), but not P-sel or Agg. In contrast, aspirin usage was associated with lower recovery levels of P-sel (ß = –0.235, t(71) = –2.167, p = .03) and Agg (ß = –0.251, t(71) = –2.255, p = .03), but not FbR. Age was not a significant main effect in any of the analyses. Because some literature suggests that the effects of HRT may be moderated by age, we also added an age x HRT interaction term. The CG x HRT interactions with all measures of activation remained significant, although the Age x HRT interaction was not.

To establish whether the differences apparently attributable to caregiver differences were in fact due to the higher prevalence of CAD among CG, the repeated-measures analyses were repeated in the subsample of 64 participants who did not have CAD (38 CG and 26 NC). The HRT distribution did not differ: {chi}2 (1) = 1.502, p = .22. The three-way interaction remained significant for speech recovery on all outcomes: Agg (F(1,59) = 5.993, p = .02, {eta}p2 = 0.092), P-Sel (F(1,59) = 4.375, p = .04, {eta}p2 = 0.069), and FbR (F(1,59) = 4.624, p = .04, {eta}p2 = 0.073).

Follow-Up Analyses of Simple Effects
Follow-up regression analyses clarified the caregiving by HRT interaction by looking at both sets of simple effects. Analyses of caregiving effects on platelet activation recovery among HRT versus non-HRT users are reported in Table 3. The overall pattern reveals that being a caregiver was associated with greater platelet activation, only among half of the sample taking HRT. The alternate simple effects analyses of HRT effects among CG versus NC are reported in Table 4. These results show that among CG, HRT use significantly increased P-Sel and FbR expression, whereas among NC, HRT use was associated with marginally significantly decreased Agg.


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  		TABLE 3. Simple Effects of Caregiving on Platelet Recovery Among HRT Users versus Non-HRT Users

 

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  		TABLE 4. Simple Effects of HRT on Platelet Recovery Among Caregivers Versus Noncaregivers

 

Effects of HRT on Circulating Levels of Platelet Activation
The main multivariate HRT effects are difficult to interpret given reactivity differences between the second and third time points; however, basal levels of platelet activation were unassociated with the CG x HRT interaction. Therefore, regression analyses of the full model investigated the independent effects of HRT on each platelet activation measure at baseline, adjusting for age and CG status. HRT usage was significantly associated with increased basal levels of Agg (ß = 0.469, t(71) = 2.468, p = .02) and FbR (ß = 0.381, t(71) = 2.002, p = .049), but not P-sel. Neither CG status nor the CG x HRT interaction was significantly related to basal levels, independent of HRT.


   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
NOTES
 REFERENCES
 
Women face difficult decisions in weighing the cost to benefit profile of HRT, given the contradictions of recent scientific evidence. Our findings suggest that HRT seems to have particularly heightened effects on platelet activation among postmenopausal women coping with the chronic stress of caring for a spouse with Alzheimer's disease. In response to acute psychological stress of an interpersonal nature, postmenopausal female CG taking HRT exhibited higher levels of platelet Agg, FbR, and P-sel expression after 14 minutes of recovery, when compared with CG not taking HRT as well as NC with or without HRT. These results persisted when controlling for age, medication use, indices of health behaviors, and preexisting diagnoses of CVD. Consistent with past literature, aspirin was associated with improved recovery (45,46), whereas preexisting CVD was associated with prolonged activation (47,48). Moreover, there was an overall tendency, such that, among CG taking HRT, activation levels increased in the recovery period, whereas among NC taking HRT, activation levels decreased. Theoretically, prolonged platelet hyperactivity (i.e., slower recovery after an acute stressor) could potentially contribute to the development of atherosclerosis or cardiovascular events by promoting increased thrombus formation, vascular inflammation, and interactions with the endothelial wall.

These findings present the possibility that chronic stress may "prime" platelets to mount prolonged activation responses to everyday hassles; e.g., epinephrine, secreted under stress, may amplify platelet activation by other agonists (9). These findings confirm the results of previous studies demonstrating that CG exhibit heightened levels of psychological distress and burden (49,50); moreover, stress among CG is linked to increased procoagulant reactivity (2,3) and overall mortality (7). However, little is known about platelet reactivity to acute stress among CG. Moreover, these analyses speak to the possible cardiovascular sequelae of superimposing acute stress over chronic stress. The findings suggest that the deleterious effects of chronic stress may manifest as an inability of physiological systems to recover quickly from everyday stressful events. Evidence suggests that the risk of acute coronary syndromes increases in the 2 hours after an emotional trigger (51). These events may be more likely to occur, when sympathetic tone or the functional/anatomical integrity of the vasculature is disturbed (52), potentially due to chronic stress (51,53).

Our findings suggest that HRT was significantly associated with elevated resting levels of in vivo platelet activity, whereas the combined effect of caregiving and HRT was associated with a prolonged activation response to the acute psychological challenge. Among the entire sample, HRT use was associated with increased resting levels of platelet Agg and expression of the FbR on them, but not with P-sel expression. The activation-dependent conformational change of the GP IIb/IIIa receptor allows platelets to cross-link via fibrinogen, yielding platelet Agg, thereby enabling thrombus formation (12). In contrast, among CG only, HRT use was associated not only with an excessive number of Agg and fibrinogen binding sites, but also with prolonged P-sel expression during recovery. Notably, although CG taking HRT exhibited the worst P-sel recovery, none of the groups achieved full P-sel recovery (i.e., return to basal levels) 14 minutes after cessation of the stressor. One previous study found that platelet-leukocyte Agg remained elevated 30 minutes after stress, even among apparently healthy male controls (48). These findings may speak to the sensitivity of P-sel and platelet-leukocyte interactions to stress. P-sel is believed to have broad functions with respect to platelet interaction with leukocytes and the vascular endothelium (17), making it a crucial mediator in the development of atherosclerotic lesions (54). Merten et al. (55) proposed that P-sel has an additive role in platelet aggregation in response to pulsatile sheer stress, distinct from that of the FbR. Although speculative, such research suggests the concomitant engagement of fibrinogen binding site and P-sel expression in response to acute stress, such as that noted among CG taking HRT, may potentially lead to larger and more stable Agg (13).

What underlying mechanisms might account for the observed findings of delayed recovery from platelet activation among CG taking HRT? A body of literature suggests that epinephrine may act as a "synergistic agonist," amplifying the effects of other weak platelet agonists (9,21,56,57). These synergistic effects may also exhibit a biphasic profile (58), suggesting a physiological rationale for distinguishing between the magnitude of the initial reactivity response versus prolonged recovery. Moreover, previous research showed that elevated sympathetic reactivity was associated with caregiver psychological distress (59). It is therefore possible that stress-induced sympathetic arousal could trigger an exaggerated response in platelets preactivated by HRT. By acting on estrogen receptors, HRT is believed to modify a number of factors that may contribute to increased coagulation, including tissue factor pathway inhibitor and 5-HT2a receptors expressed on platelets (60,61). Moreover, platelets are also known to express estrogen receptors, although relatively little is known about their function, particularly with regard to chronic stress and HRT (62–64). In addition, HRT and stress are both known to modulate the hypothalamic-pituitary-adrenal axis and the immune system (65), constituting additional possible pathways of interaction.

One limitation of the study is that it is cross-sectional and quasi-experimental in nature (i.e., participants were not randomized to HRT). However, given the controversies on adverse cardiovascular effects of HRT, it would be ethically difficult to randomly assign elderly individuals with an increased prevalence of preexisting CVD to either HRT or placebo in a double-blind study. We found that the combined effects of caregiving and HRT persisted in the subsample of participants who had not been diagnosed with CAD, as well as in the total sample when statistically controlling for age, CVD, and aspirin use; however, it remains conceivable that increased subclinical endothelial damage among CG could account for the worsened platelet response to HRT. The main focus of our study was to investigate the effect of caregiving stress rather than HRT effects on platelet activation. Because of sample size, we were not in a position to explore whether various types of HRT regimens (e.g., estrogen only versus estrogen and progestin) had different associations with platelet activation. Nonetheless, although further research is needed, the stability of the primary findings across multiple analyses, general use of HRT, and platelet outcomes is promising. In particular, future prospective studies might explore whether CG who take HRT or have impaired platelet recovery exhibit faster disease progression, as indicated by subclinical markers of endothelial injury (e.g., increased carotid arterial intima-media thickness) (66), or heightened risk of cardiovascular events (e.g., fatal and nonfatal acute myocardial infarction).

In conclusion, these results present intriguing findings regarding the potential interaction between the chronic stress of dementia caregiving and HRT use among elderly women in terms of cardiovascular health. If confirmed, these results suggest that health providers may want to take chronic stress into account when determining cardiovascular risk factors and evaluating the risk/benefit ratio in prescribing HRT. The study suggests a theoretical model of how the general diathesis-stress model (67) might apply to risk assessment for adverse drug reactions; i.e., pharmaco-risk superimposed against a background of chronic stress may accelerate an underlying disease process, such as atherosclerosis. Because the benefit of risk reduction strategies is proportionate to the level of risk, CG at high risk of CVD should derive extra benefit from preventive measures such as low-dose aspirin and close monitoring by their health providers. This study emphasizes how cardiovascular risk, as it relates to platelet hyperactivity in our study, may occur through multiplicative interaction of risk factors.

The authors are grateful to Susan Calleran, MA, Carolyn Swenerton, RN, and Sharyn Wilenski, RN.


   NOTES
TOP
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 NOTES
REFERENCES
 
Received for publication April 24, 2007; revision received July 10, 2007.

Primary support from award AG15301 (National Institute of Aging (NIA)). Additional support from NIA grants 05131, 08415, 05131, & 23989, and award 00827 from the National Center for Research Resources. The NIA was not involved in the design or conduct of the study; collection, management, analysis, or interpretation of the data; or preparation, review, or approval of the manuscript. The authors report no conflicts of interest.

DOI:10.1097/PSY.0b013e31815a8ba8


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 METHODS
 RESULTS
 DISCUSSION
 NOTES
 REFERENCES
 

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