Trends among platelet function, arterial calcium, and vascular function measures

Figures & data

Figure 1. Overview of study design (adapted from Grech et al. 2021).

Whole blood was drawn from OMNI-2, Gen3, and NOS participants during Exam 3 (2016–2019). Whole blood and isolated PRP were used to take platelet activation measures using 5 different assays. Arterial tonometry measures were also gathered from these cohorts during Exam 3. Calcium measures were gathered from Gen3/OMNI-2 participants during Exam 2 (2008–2011), and CT vessel diameters were gathered from Gen3 participants during Exam 1 (2002–2005). A linear mixed effect model, which adjusted for age, sex, aspirin, and various CVD risk factors was used to assess the associations between the three main variable categories.Image sources:Arterial tonometry image: by Mikael Häggström, MD. Public Domain (CC0 1.0).Aortic and arterial calcium image: This Photo by Unknown Author is licensed under CC BY-SA.

Table I. Demographic information.

Trait	Tonometry (2016–2019)	Calcium (2008–2011)	Diameter (2002–2005)
	Mean (SD)/N(%)	Mean (SD)/N(%)	Mean (SD)/N(%)
Sex (female)	1807 (53.88)	762 (45.71)	763 (46.10)
Age (years)	54.40 (9.27)	58.84 (5.94)	58.88 (5.98)
Lipid Medications	836 (24.93)	546 (32.77)	544 (32.89)
Anti-hypertensive medications	934 (27.86)	577 (34.63)	572 (34.58)
Aspirin use	716 (21.35)	460 (27.59)	456 (27.55)
P2Y12 inhibitor use	32 (0.95)	19 (1.14)	22 (1.33)
BMI (kg/m2)	28.49 (5.77)	28.93 (5.79)	28.93 (5.80)
Total Cholesterol (mg/dL)	189.39 (36.60)	190.70 (36.70)	190.73 (36.89)
HDL (mg/dL)	59.58 (19.47)	59.31 (20.41)	59.37 (20.52)
Fasting glucose (mg/dL)	100.10 (21.18)	102.65 (23.12)	102.58 (22.99)
Smoking	217 (6.47)	97 (5.82)	99 (5.99)
Type 2 diabetes	296 (8.83)	186 (11.16)	185 (11.19)
AAo (N = 1649)			31.94 (3.60)
DAo (N = 1649)			23.39 (2.43)
AbAo (N = 1655)			17.33 (1.88)
CAC (N = 1667)		467 (28.01)
TAC (N = 1415)		483 (34.13)
AAS (N = 1421)		658 (46.31)
Zc (N = 3354) (dyne × sec/cm^Citation5	210.20 (74.56)
CPP(N = 3354) (mmHg)	60.04 (16.34)
niCFPWV (N = 3349) (ms/m)	−130.65 (26.41)

The following table is a summary of the relevant demographic, tonometry, diameter, and calcium measures gathered from Gen3/NOS/OMNI-2 cohorts. Platelet and tonometry measures were gathered from Gen3/NOS/OMNI-2 cohorts during Exam 3 (2016–2019), vessel calcium measures were gathered from Gen3/OMNI-2 cohorts during Exam 2 (2008–2011), and vessel diameter measures were gathered from Gen3 cohort during Exam 1 (2002–2005). N = 3,429.

Abbreviations: AAo: antero-posterior diameter of the ascending thoracic aorta obtained at the level of the right pulmonary artery; DAo: Antero-posterior diameter of the descending thoracic aorta obtained at the level of the right pulmonary artery; AbAo: antero-posterior diameter of the abdominal aorta 5 cm above the aorto-common iliac bifurcation; CAC: coronary artery calcium score; TAC: thoracic aortic calcium score; AAS: abdominal aorta calcium score; Zc: characteristic impedance of the aorta; CPP: central pulse pressure; niCFPWV: negative inverse carotid femoral pulse wave velocity.

Figure 2. Heatmap of Z statistics associating platelet activation, arterial calcium, tonometry, and diameter measures.

Red indicates increased platelet reactivity with an increase in the cardiovascular traits; blue indicates reduced platelet reactivity with a decrease in the cardiovascular traits. * (p ≤ .05); ** (p ≤ .01). We used linear mixed effects models, which included CVD risk factors, to calculate the betas (β) and standard errors (SE) for this display. Calcium values had 1 added and were ln-transformed. Following this transformation, the β and SE values were used to calculate z-statistics (β/SE). The following z-statistic values were multiplied by -1 to accurately depict the platelet activation directionality: Agg20, Agg40, ecMax, EC50, disaggregation traits (4 total), and lag time traits (2 total).Abbreviations: AAo: antero-posterior diameter of the ascending thoracic aorta obtained at the level of the right pulmonary artery; AAS: abdominal aorta calcium score; AbAo: antero-posterior diameter of the abdominal aorta 5 cm above the aorto-common iliac bifurcation; CAC: coronary artery calcium score; CPP: central pulse pressure; DAo: Antero-posterior diameter of the descending thoracic aorta obtained at the level of the right pulmonary artery; niCFPWV: negative inverse carotid femoral pulse wave velocity; TAC: thoracic aortic calcium score; Zc: characteristic impedance of the aorta.

Table II. Aortic and arterial calcium results (risk factor-adjusted model).

Agonist	Concentration	Assay Type	Platelet Trait	Calcium Trait	Beta	SE	Z-Statistic	P-Value	N
AA	0.03–1.0 mM	Optimul	ecMAX	AAS	−0.147	0.052	−2.824	4.75E–03	1162
ADP	1.82 μM	LTA	FinAgg	CAC	0.143	0.053	2.672	7.54E–03	1586
ADP	20 µM	Flow Cytometry (WB)	PAC1% (WB)	TAC	−0.18	0.071	−2.537	1.12E–02	1153
Ristocetin	1.5 mg/mL	LTA	FinAgg	TAC	0.168	0.066	2.528	1.15E–02	1351
ADP	0.95 μM	LTA	PrimSlope	CAC	0.143	0.057	2.498	1.25E–02	1502
Ristocetin	1.5 mg/mL	LTA	MaxAgg	TAC	0.162	0.066	2.457	1.40E–02	1351
ADP	1.82 μM	LTA	MaxAgg	CAC	0.133	0.056	2.395	1.66E–02	1586
Ristocetin	1.5 mg/mL	LTA	AUC	TAC	0.158	0.067	2.364	1.81E–02	1350
ADP	1.82 μM	LTA	AUC	CAC	0.126	0.054	2.330	1.98E–02	1586
Collagen	190 µg/mL	LTA	PrimSlope	AAS	−0.119	0.052	−2.314	2.07E–02	1370
Ristocetin	0.14–4.0 mg/ml	Optimul	ecMAX	AAS	−0.122	0.053	−2.289	2.21E–02	1234
ADP	1.82 μM	LTA	PrimSlope	CAC	0.126	0.058	2.178	2.94E–02	1586
U46619	0.005–40 µM	Optimul	ecMAX	CAC	−0.121	0.056	−2.174	2.97E–02	1442
Collagen	0.01–40 µg/ml	Optimul	ecMAX	AAS	0.09	0.043	2.098	3.59E–02	1256
Collagen	190 µg/mL	LTA	AUC	TAC	0.113	0.054	2.097	3.60E–02	1364
U46619	0.005–40 µM	Optimul	eMax	TAC	0.146	0.07	2.097	3.60E–02	1243
Ristocetin	1.5 mg/mL	LTA	PrimSlope	TAC	0.134	0.065	2.069	3.85E–02	1350
ADP	20 µM	Flow Cytometry (WB)	PAC1_Psel_DoublePos% (WB)	TAC	−0.147	0.071	−2.067	3.88E–02	1153
TRAP-6	0.67 mM	LTA	PrimAgg	TAC	0.135	0.066	2.046	4.07E–02	1360
AA	1.6 mM	LTA	FinAgg	CAC	0.087	0.043	2.038	4.16E–02	1587
ADP	0.95 μM	LTA	MaxAgg	CAC	0.118	0.058	2.029	4.25E–02	1502
TRAP-6	0.67 mM	LTA	PrimSlope	TAC	0.134	0.068	1.976	4.81E–02	1360

Trending results (p < .05) of multiple testing analysis of aortic and arterial calcium traits and platelet traits. Positive beta/z-statistic values indicate increased platelet reactivity with an increase in the cardiovascular traits; negative beta/z-statistic values indicate reduced platelet reactivity with a decrease in the cardiovascular traits. We used linear mixed effects models, which included CVD risk factors, to calculate the betas (β) and standard errors (SE) for this display. Calcium values had 1 added and were ln-transformed. Following this transformation, the β and SE values were used to calculate z-statistics (β/SE). The following z-statistic values were multiplied by −1 to accurately depict the platelet activation directionality: Agg20, Agg40, ecMax, EC50, disaggregation traits (4 total), and lag time traits (2 total).

Abbreviations: CAC: coronary artery calcium score; AAS: abdominal aorta calcium score; TAC: thoracic aorta calcium score.

Table IV. Arterial tonometry analysis results (risk factor-adjusted model).

Agonist	Concentration	Assay Type	Platelet Trait	Tonometry Trait	Beta	SE	Z-Statistic	P-Value	N
ADP	0.95 μM	LTA	SecSlp	niCFPWV	−0.0019	0.00064	−3.040	2.36E–03	3049
Collagen	61.0 μg/mL	Multiplate	AUC	Zc	0.00072	0.00024	3.010	2.59E–03	3336
ADP	0.95 μM	LTA	PrimAgg	CPP	0.0034	0.0011	2.990	2.81E–03	3059
ADP	0.95 μM	LTA	SecAgg	niCFPWV	−0.0017	0.00064	−2.680	7.39E–03	3049
ADP	0.95 μM	LTA	PrimSlope	CPP	0.0029	0.0011	2.630	8.46E–03	3059
Epinephrine	0.0004–10 µM	Optimul	ecMax	niCFPWV	−0.0018	0.00071	−2.548	1.08E–02	2987
Ristocetin	1.5 mg/mL	LTA	PrimSlope	CPP	−0.0029	0.0012	−2.490	1.26E–02	3224
Collagen	190 µg/mL	LTA	PrimSlope	Zc	−0.00054	0.00022	−2.460	1.40E–02	3264
Epinephrine	0.0004–10 µM	Optimul	aucMean	Zc	0.00060	0.00025	2.370	1.78E–02	2996
Epinephrine	0.0004–10 µM	Optimul	eMax	Zc	0.00058	0.00026	2.260	2.41E–02	3014
Epinephrine	0.0004–10 µM	Optimul	Slope	niCFPWV	−0.0021	0.00094	−2.200	2.80E–02	2986
Ristocetin	1.5 mg/mL	LTA	FinAgg	niCFPWV	0.0020	0.00090	2.200	2.80E–02	3212
Ristocetin	1.5 mg/mL	LTA	MaxAgg	niCFPWV	0.0019	0.00090	2.130	3.34E–02	3212
Epinephrine	100 μM	LTA	SecAgg	niCFPWV	0.0015	0.00070	2.090	3.67E–02	3250
Epinephrine	100 μM	LTA	SecSlp	niCFPWV	0.0014	0.00070	2.030	4.19E–02	3250
AA	0.03–1.0 mM	Optimul	Agg40	Zc	0.00046	0.00023	2.027	4.27E–02	2658
ADP	0.95 μM	LTA	PrimAgg	Zc	0.00051	0.00025	2.020	4.36E–02	3059
Collagen	61.0 μg/mL	Multiplate	Agg	Zc	0.00048	0.00024	1.980	4.76E–02	3336
Collagen	61.0 μg/mL	Multiplate	AUC	niCFPWV	−0.0017	0.00085	−1.960	4.96E–02	3324
Epinephrine	0.0004–10 µM	Optimul	Agg20	Zc	0.00052	0.00027	1.962	4.98E–02	2943

Trending results (p < .05) of multiple testing analysis of arterial tonometry and platelet traits. Positive beta/z-statistic values indicate increased platelet reactivity with an increase in the cardiovascular traits; negative beta/z-statistic values indicate reduced platelet reactivity with a decrease in the cardiovascular traits. We used linear mixed effects models, which included CVD risk factors, to calculate the betas (β) and standard errors (SE) for this display. The β and SE values were used to calculate z-statistics (β/SE). The following z-statistic values were multiplied by −1 to accurately depict the platelet activation directionality: Agg20, Agg40, ecMax, EC50, disaggregation traits (4 total), and lag time traits (2 total).

Abbreviations: niCFPWV: negative inverse carotid femoral pulse wave velocity; CPP: central pulse pressure; Zc: characteristic impedance of the aorta.

Table III. Vessel diameter results (risk factor-adjusted model).

Agonist	Concentration	Assay Type	Platelet Trait	Diameter Trait	Beta	SE	Z-Statistic	P-Value	N
Epinephrine	0.0004–10 µM	Optimul	eMax	DAo	−0.048	0.015	−3.251	1.15E–03	1468
ADP	0.005–40 µM	Optimul	eMax	DAo	−0.046	0.016	−2.949	3.18E–03	1473
Epinephrine	0.0004–10 µM	Optimul	eMax	AbAo	−0.053	0.019	−2.820	4.81E–03	1472
TRAP-6	1.5 μM-1.8 mM	Optimul	eMax	DAo	−0.045	0.016	−2.735	6.24E–03	1284
Ristocetin	1.5 mg/mL	LTA	PrimAgg	AAo	−0.019	0.008	−2.311	2.08E–02	1557
AA	0.03–1.0 mM	Optimul	Slope	DAo	−0.037	0.016	−2.285	2.23E–02	1329
TRAP-6	1.5 μM-1.8 mM	Optimul	AUC	DAo	−0.038	0.017	−2.275	2.29E–02	1284
AA	0.03–1.0 mM	Optimul	ecMAX	DAo	−0.029	0.013	−2.266	2.35E–02	1330
Collagen (shear)	N/A	T-TAS	AUC	DAo	0.059	0.026	2.246	2.47E–02	399
Ristocetin	0.14–4.0 mg/ml	Optimul	EC50	AAo	−0.019	0.009	−2.243	2.49E–02	1414
Ristocetin	0.14–4.0 mg/ml	Optimul	Agg40	AAo	−0.019	0.009	−2.199	2.79E–02	1404
ADP	0.005–40 µM	Optimul	Agg40	DAo	−0.03	0.014	−2.176	2.96E–02	1457
TRAP-6	1.5 μM-1.8 mM	Optimul	eMax	AbAo	−0.045	0.021	−2.139	3.24E–02	1286
ADP	5.71 μM	LTA	AUC	AbAo	−0.035	0.017	−2.104	3.54E–02	1566
U46619	0.005–40 µM	Optimul	AUC	DAo	−0.032	0.015	−2.093	3.63E–02	1430
U46619	0.005–40 µM	Optimul	Agg40	DAo	−0.032	0.015	−2.085	3.71E–02	1411
Ristocetin	1.5 mg/mL	LTA	Disagg	AbAo	0.03	0.015	2.051	4.02E–02	1563
Ristocetin	0.14–4 mg/ml	Optimul	eMax	DAo	−0.032	0.016	−2.044	4.10E–02	1446
ADP	5.71 μM	LTA	MaxAgg	AbAo	−0.033	0.016	−2.020	4.34E–02	1568
U46619	0.005–40 µM	Optimul	EC50	DAo	−0.03	0.015	−2.008	4.46E–02	1417
Ristocetin	1.5 mg/mL	LTA	PrimSlope	AAo	−0.017	0.008	−2.004	4.51E–02	1557
Epinephrine	100 μM	LTA	PrimSlope	AAo	−0.015	0.007	−1.985	4.71E–02	1570
TRAP-6	1.5 μM-1.8 mM	Optimul	aucMean	AbAo	−0.042	0.021	−1.971	4.87E–02	1286
U46619	0.005–40 µM	Optimul	ecMAX	DAo	−0.026	0.013	−1.970	4.88E–02	1430
ADP	0.005–40 µM	Optimul	aucMean	DAo	−0.028	0.014	−1.967	4.92E–02	1471
Epinephrine	0.0004–10 µM	Optimul	aucMean	DAo	−0.028	0.014	−1.963	4.96E–02	1462
ADP	0.005–40 µM	Optimul	EC50	DAo	−0.027	0.014	−1.963	4.97E–02	1461

Trending results (p < .05) of multiple testing analysis of aortic and arterial diameter traits and platelet traits. Positive beta/z-statistic values indicate increased platelet reactivity with an increase in the cardiovascular traits; negative beta/z-statistic values indicate reduced platelet reactivity with a decrease in the cardiovascular traits. We used linear mixed effects models, which included CVD risk factors, to calculate the betas (β) and standard errors (SE) for this display. The β and SE values were used to calculate z-statistics (β/SE). The following z-statistic values were multiplied by −1 to accurately depict the platelet activation directionality: Agg20, Agg40, ecMax, EC50, disaggregation traits (4 total), and lag time traits (2 total).

Abbreviations: AAo: antero-posterior diameter of the ascending thoracic aorta obtained at the level of the right pulmonary artery; AbAo, antero-posterior diameter of the abdominal aorta 5 cm above the aorto-common iliac bifurcation; DAo, Antero-posterior diameter of the descending thoracic aorta obtained at the level of the right.

Amable PR, Carias RB, Teixeira MV, da Cruz Pacheco I, Correa Do Amaral RJ, Granjeiro JM, Borojevic R. Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors. Stem Cell Res Ther. 2013;4(3):67. doi:10.1186/scrt218.

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Data availability statement

Framingham Heart Study variables are deposited in the NIH dbGaP repository under the accession # phs000007.v32.p13 and available for application to access by qualified researchers https://www.ncbi.nlm.nih.gov/gap/.