Inflammation and Thrombosis Biomarkers and Incident Frailty in Postmenopausal Women

Materials and Methods 

Results 

Of the baseline characteristics examined (Table 1), lower education, obesity, current smoking, infrequent alcohol consumption, worse health status, more depressive symptoms, diabetes, hypertension, arthritis, and nonsteroidal anti-inflammatory drug and statin use were more common among frailty cases than controls (P ≤.01). Cross-sectional univariate analyses of baseline characteristics in the 900 non-frail control women according to quartile of plasma biomarker levels showed significant associations (P ≤.01) of age with higher D-dimer, factor VIII, and t-PA; BMI with higher CRP and t-PA; hormone therapy use with higher CRP, fibrinogen, and t-PA; hypertension with higher CRP and t-PA; and alcohol consumption with higher fibrinogen.

Table 1. Baseline Characteristic of Women's Health Initiative Observational Study Participants versus Incident Frailty

		Frail Cases		Non-Frail Controls		P Value
		N	%	N	%
	Age group at screening, y
	60-69	485	53.9	485	53.9	1.00
	70-79	415	46.1	415	46.1	.
	Education
	0-8 y	12	1.3	6	0.7	<.001
	Some high school	34	3.8	17	1.9	.
	High school diploma/GED	192	21.5	135	15.1	.
	School after high school	341	38.2	342	38.2	.
	College degree or higher	314	35.2	395	44.1	.
	Ethnicity
	White	865	96.1	865	96.1	1.00
	Black	16	1.8	16	1.8	.
	Hispanic	2	0.2	2	0.2	.
	Asian/Pacific Islander	15	1.7	15	1.7	.
	Unknown	2	0.2	2	0.2	.
	BMI, kg/m2 (full categories)
	Underweight (<18.5)	4	0.4	14	1.6	<.001
	Normal (18.5-24.9)	271	30.2	461	51.5	.
	Overweight (25.0-29.9)	352	39.3	320	35.8	.
	Obesity I (30.0-34.9)	188	21.0	86	9.6	.
	Obesity II (35.0-39.9)	55	6.1	10	1.1	.
	Extreme obesity III (≥40)	26	2.9	4	0.4	.
	Smoking
	Never	472	52.9	478	53.7	.005
	Past	360	40.4	382	42.9	.
	Current	60	6.7	30	3.4	.
	Alcohol
	Non/past drinker	272	30.3	200	22.3	<.001
	<1 drink/wk	314	35.0	251	28.0	.
	1-14 drinks/wk	271	30.2	404	45.1	.
	>14 drinks/wk	41	4.6	40	4.5	.
	Hormone therapy use status
	Never used	378	42.0	400	44.5	.48
	Past user	129	14.3	131	14.6	.
	Current user	393	43.7	368	40.9	.
	Depressive mood
	0	167	18.7	325	36.6	<.001
	1-2	367	41.2	356	40.1	.
	3-4	215	24.1	150	16.9	.
	5+	142	15.9	57	6.4	.
	Activity of daily living disability
	No	869	98.5	878	99.1	.27
	Yes	13	1.5	8	0.9	.
	Treated diabetes (pills or shots)
	No	862	95.8	888	98.8	<.001
	Yes	38	4.2	11	1.2	.
	Hypertensive
	No	448	49.9	564	62.7	<.001
	Yes	450	50.1	336	37.3	.
	History of arthritis
	No	330	36.7	529	58.8	<.001
	Yes	570	63.3	371	41.2	.
	History of hip fracture at age ≥55 y
	No	805	98.9	825	99.8	.03
	Yes	9	1.1	2	0.2	.
	No. of falls in last 12 mo
	None	605	67.3	643	71.6	.02
	1 time	170	18.9	171	19.0	.
	2 times	81	9.0	61	6.8	.
	≥3	43	4.8	23	2.6	.
	Angiotensin-converting enzyme inhibitor use
	No	816	90.7	844	93.8	.01
	Yes	84	9.3	56	6.2	.
	Non-statin antihyperlipidemic medication use
	No	882	98.0	888	98.7	.27
	Yes	18	2.0	12	1.3	.
	Statin use
	No	798	88.7	835	92.8	.003
	Yes	102	11.3	65	7.2	.
	NSAID use
	No	521	57.9	588	65.3	.001
	Yes	379	42.1	312	34.7	.
	Coumarin anticoagulant use
	No	889	98.8	898	99.8	.01
	Yes	11	1.2	2	0.2	.

GED=General Equivalency Diploma; BMI=body mass index; NSAID=nonsteroidal anti-inflammatory drug.

In both minimally adjusted and fully adjusted regression models, women with t-PA or D-dimer levels in the upper quartile were at increased risk of incident frailty relative to women in the lowest respective quartiles; women in the second and third quartiles were at intermediate risk (Table 2). The trend P values associated with D-dimer and t-PA were each P = .04 for the fully adjusted models. Women with IL-6 levels ≥1 pg/mL and those with detectable IL-6 levels less than 1 pg/mL had a higher risk of frailty than women with undetectable IL-6 levels in the minimally adjusted model (P = .03), but the frailty-IL-6 association became nonsignificant on further covariate adjustment (P = .27). Baseline levels of CRP, fibrinogen, and factor VIII were not significantly associated with incident frailty in either minimally or fully adjusted models. Restricting the analysis to subjects with factor VIII levels>30% and fibrinogen levels>100 did not appreciably alter these results.

Table 2. Risk of Frailty by Quartile of Inflammation and Coagulation Biomarkers

	Biomarker Quartile	Model 1 Odds Ratio (95% CI)	Model 2 Odds Ratio (95% CI)
	C-reactive protein Q1 (<1.1 mg/L)	1	1
	C-reactive protein Q2 (1.1-2.3 mg/L)	1.12(0.80-1.56)	1.24(0.84-1.81)
	C-reactive protein Q3 (2.3-4.8 mg/L)	1.13(0.81-1.58)	1.14(0.78-1.65)
	C-reactive protein Q4 (≥4.8 mg/L)	1.15(0.82-1.62)	1.05(0.72-1.54)
	P for trend	0.43	0.95
	D-dimer Q1 (<0.24 ug/mL)	1	1
	D-dimer Q2 (0.24-0.36 ug/mL)	1.14(0.84-1.56)	1.27(0.90-1.79)
	D-dimer Q3 (0.36-0.61 ug/mL)	1.10(0.81-1.49)	1.05(0.75-1.48)
	D-dimer Q4 (≥0.61 ug/mL)	1.37(1.00-1.88)	1.57(1.11-2.22)
	P for trend	0.07	0.04
	Factor VIII Q1 (<64%)	1	1
	Factor VIII Q2 (64%-90%)	1.22(0.90-1.64)	1.17(0.84-1.64)
	Factor VIII Q3 (90%-116%)	1.05(0.76-1.44)	1.05(0.74-1.48)
	Factor VIII Q4 (≥116%)	1.02(0.73-1.42)	1.10(0.76-1.59)
	P for trend	0.72	0.84
	Fibrinogen Q1 (<261 mg/dL)	1	1
	Fibrinogen Q2 (261-305 mg/dL)	0.71(0.52-0.97)	0.70(0.49-1.00)
	Fibrinogen Q3 (305-352 mg/dL)	0.91(0.66-1.25)	0.96(0.67-1.36)
	Fibrinogen Q4 (≥352 mg/dL)	0.95(0.69-1.31)	0.93(0.64-1.34)
	P for trend	0.79	0.80
	Tissue plasminogen activator Q1 (<6 mg/mL)	1	1
	Tissue plasminogen activator Q2 (6-8 mg/mL)	1.14(0.83-1.57)	1.23(0.87-1.75)
	Tissue plasminogen activator Q3 (8-10.3 mg/mL)	1.10(0.80-1.51)	1.20(0.84-1.72)
	Tissue plasminogen activator Q4 (≥10.3 mg/mL)	1.48(1.05-2.07)	1.52(1.05-2.22)
	P for trend	0.03	0.04
	Interleukin-6 (0 pg/mL)	1	1
	Interleukin-6 (0<1 pg/mL)	1.07(0.74-1.54)	0.97(0.64-1.45)
	Interleukin-6 (≥1 pg/mL)	1.41(1.00-1.98)	1.20(0.82-1.76)
	P for trend	0.03	0.27

CI=confidence interval. Model 1 adjusted for hypertension, hormone use, and BMI. Model 2 adjusted for model 1 covariates+education, alcohol consumption, hypertension, arthritis, and smoking.

Additional sensitivity analyses were performed by stratifying the fully adjusted frailty-biomarker models for D-dimer and t-PA according to baseline use of any anti-inflammatory and anticoagulant medication, including angiotensin-converting enzyme inhibitors, statins, non-statin anti-hyperlipidemics, nonsteroidal anti-inflammatory drugs, and warfarin. The frailty odds ratios by quartile of biomarker level were similar to those described above. For example, among the 936 participants (52%) not using anti-inflammatory or anticoagulant medications, the frailty odds ratio for the highest quartile of D-dimer was 1.81 (95% confidence interval, 1.12-2.91) compared with women in the lowest quartile of D-dimer. Similarly, among non-medication users, the frailty odds ratio associated with the highest quartile of tPA was 1.52 (95% confidence interval, 0.93-2.50) compared with women in the lowest quartile of t-PA. There was no significant interaction between either D-dimer or t-PA and anti-inflammatory/anticoagulant medication use on risk of frailty (P values for interaction = .47 and .42, respectively).

Because D-dimer and t-PA measure different aspects of the blood coagulation and fibrinolysis pathways, and circulating levels were uncorrelated with one another among non-frail WHI-OS controls (Spearman's r = −0.04), we also explored whether the combination of high D-dimer and high t-PA levels was associated with a greater risk of frailty compared with each biomarker individually (Table 3). In the fully adjusted regression model, women in the upper quartile for t-PA had only a 1.3-fold increased risk of frailty and women in the upper quartile for D-dimer had only a 1.4-fold increased risk of frailty, compared with women with low levels of both markers. For women in the upper quartile of both t-PA and D-dimer, the risk of incident frailty was even larger (odds ratio, 2.2). Although there was some overlap in the confidence intervals for these risk estimates, the combined effect of t-PA and D-dimer levels on risk frailty was significantly greater than the effect of each biomarker individually (P = .02).

Table 3. Risk of Frailty Comparing Levels of D-dimer and Tissue Plasminogen Activator in the Upper Range of the Distribution

	Biomarker Quartile	Model 1 Odds Ratio (95% CI)	Model 2 Odds Ratio (95% CI)
	Neither D-dimer nor t-PA in the upper quartile	1	1
	Only t-PA in the upper quartile	1.38(1.05-1.82)	1.29(0.94-1.76)
	Only D-dimer in the upper quartile	1.30(0.97-1.74)	1.38(0.99-1.91)
	Both D-dimer and t-PA in the upper quartile	1.90(1.17-3.08)	2.20(1.29-3.75)
	Overall P value(3 df test)	.02	.02

CI=confidence interval; t-PA=tissue plasminogen activator. Model 1 adjusted for hypertension, hormone use, and BMI. Model 2 adjusted for model 1 covariates+education, alcohol consumption, hypertension, arthritis, and smoking.

Discussion 

In a nested case-control study of incident frailty in postmenopausal women, higher levels of the coagulation/fibrinolysis biomarkers D-dimer and t-PA were each associated with increased risk of frailty at 3-year follow-up. The associations between frailty and D-dimer or t-PA were independent of the potential confounding effects of comorbid conditions and lifestyle factors that also are associated with increased coagulation and inflammation biomarkers levels and frailty. To our knowledge, a prospective association between D-dimer and t-PA and incident frailty have not been reported. Moreover, the combined effects of high D-dimer and high t-PA on risk of incident frailty were greater than the risk associated with either biomarker alone. The risks of frailty associated with D-dimer and t-PA were not modified by baseline use of medications with anticoagulant or anti-inflammatory properties. Finally, we observed little evidence for association between coagulation factor VIII, fibrinogen, CRP, or IL-6 levels and development of frailty.

Studies of older adults consistently support associations between markers of thrombosis and inflammation and measures of disability, physical performance, or frailty.4, 5, 6, 7, 8, 9, 10, 11, 12 However, most previous studies assessing the outcome of frailty were cross-sectional analyses, in which the temporal relationships underlying the observed associations are more difficult to tease apart. To our knowledge, there has been one other prospective analysis examining the relationship of biomarkers measured at baseline to development of incident frailty during follow-up in CHS.13 In the report by Barzilay et al,13 CRP and insulin resistance were associated with increased risk of incident frailty in CHS independently of potential confounders, whereas coagulation factor VIII levels had a borderline significant association. The lack of association between CRP and incident frailty in the WHI-OS might reflect differences in gender or other study population characteristics between women from WHI compared with men and women from CHS. D-dimer and t-PA were not evaluated in the analysis of incident frailty by Barzilay et al,13 although higher D-dimer levels were previously reported to be increased with baseline frailty in a cross-sectional analysis from CHS.5 Therefore, additional prospective studies of older adults are required to confirm the role of D-dimer and t-PA as independent risk factors for incident frailty.

D-dimer results from fibrin formation and degradation, and is a marker of activation of the coagulation and fibrinolytic systems. Fibrinolytic activity is largely regulated by tissue-type plasminogen activator (t-PA) released from endothelial cells and by a circulating inhibitor, plasminogen activator inhibitor type 1. t-PA antigen measures free, active t-PA, and t-PA complexed to plasminogen activator inhibitor type 1. Therefore, t-PA and D-dimer each summarize different components of the coagulation/fibrinolytic systems. Both t-PA antigen and D-dimer levels increase with age19, 20 and have been reported to predict future venous thromboembolic21 and arterial thrombotic events22, 23 independently of traditional cardiovascular risk factors. Together with a recent report that frailty predicts increased risk of idiopathic venous thrombotic events,12 the current WHI findings support a possible pathophysiologic connection among aging, activation of the blood coagulation and fibrinolytic systems, and occurrence of frailty. The association between D-dimer and t-PA and development of frailty in postmenopausal women from WHI was present in women without clinical cardiovascular disease, because women with overt cardiovascular disease were excluded from our analyses. Because frailty has been correlated with the extent of underlying atherosclerotic disease,24 studies that include subclinical noninvasive measures of vascular disease are needed to more accurately assess the interrelationships among coagulation/fibrinolysis biomarkers, vascular disease, and frailty.

Frailty has been conceptualized as a pathophysiologic state of global functional decline characterized by increased vulnerability to acute and chronic physical and psychosocial stressors due to impaired homeostatic mechanisms. Acute or chronic stress contributes to activation of the coagulation system and impaired fibrinolysis, including increases in circulating levels of D-dimer and t-PA.25, 26, 27, 28 Therefore, D-dimer and t-PA might hasten the clinical transition to frailty or other aging-related disorders in susceptible older individuals who experience sustained or repeated physical or psychosocial trauma, stress, injury, or “allostatic load.”29 Strengths of the current study include the use of a prospective study design in which we assessed incident cases of frailty that occurred subsequent to measurement of the biomarkers. Although we excluded women with chronic diseases that manifest as frailty and also performed sensitivity analyses according to use of medications with anti-inflammatory or anticoagulant properties, we cannot fully exclude the possibility that residual confounding due to unmeasured factors, such as subclinical atherosclerosis,24 might account for the observed frailty-biomarker associations. Because our study population comprised postmenopausal women of predominantly white race, whether the associations with t-PA and D-dimer with incident frailty are generalizable to men and other ethnic groups also requires additional study.

Other study limitations include the relatively short follow-up duration and lack of objective physical performance measurements of muscle weakness and walking speed, which may be prone to error. Nonetheless, our “modified definition” of frailty has been validated previously through strong association with future mortality, disability, hospitalization, and hip fracture events among older women in the WHI-OS.2 Because of assay and plasma specimen issues that we were not able to fully resolve, a sizeable proportion of our study participants had undetectable IL-6 levels; thus, our ability to assess the risk of frailty with IL-6 was limited, and longitudinal assessment of IL-6 in other frail populations is warranted.

Conclusions 

Plasma levels of D-dimer and t-PA, 2 markers of fibrin turnover and fibrinolysis, were associated prospectively with risk of incident frailty in postmenopausal women. These findings support the role of activation of coagulation and fibrinolytic systems in the pathophysiology and occurrence of frailty in older adults. If these results are confirmed in other study populations, screening of D-dimer and t-PA may be useful for identifying otherwise healthy older individuals at risk for developing frailty or other functional consequences of aging.