Variation in Platelet Activation State in Pre-Donation Whole Blood: Effect of Time of Day and ABO Blood Group

Figures & data

Table 1 Donation History, Results of Laboratory Analyses, and Percentage Distribution of Responses to Questionnaire Items by Gender and Age Group (n = 86)

Variables	Overall (n = 86)	Gender		Age Group
		Males (n = 38)	Females (n = 48)	22–50 yr (n = 42)	51–69 yr (n = 44)
Age (years)	50.8 ± 9.7 (22–69)	52.1 ± 8.7	49.5 ± 10.4	42.6 ± 6.5	58.4 ± 4.8^‡
Previous donations
Combined (number)	54 ± 41 (4–256)	61 ± 45	49 ± 37	33 ± 23	74 ± 45^‡
Apheresis PLTs (number)	14 ± 13 (1–43)	15 ± 13	13 ± 13	10 ± 10	18 ± 13^†
Donation interval (week)	10.6 ± 7.3 (2.4–42.8)	12.1 ± 7.3	9.9 ± 7.3	10.4 ± 7.3	10.8 ± 7.4
Hemoglobin (g/dl)	14.1 ± 0.1 (11.7–17.4)	15.0 ± 0.8	13.4 ± 0.8^‡	13.8 ± 1.0	14.4 ± 1.2*
Hematocrit (%)	43 ± 3 (36–5.1)	45 ± 2	41 ± 2^‡	42 ± 3	44 ± 3*
WB-PLT count (10^9/L)	274 ± 40 (185–377)	268 ± 34	279 ± 44	275 ± 38	273 ± 42
MPV (fL)	10.5 ± 0.8 (6.7–12.8)	10.5 ± 0.75	10.5 ± 0.85	10.6 ± 0.83	10.4 ± 0.78
IPF (%)	3.0 ± 1.6 (0.7–8.6)	2.9 ± 1.4	3.1 ± 1.7	3.3 ± 1.9	2.8 ± 1.2
IPC (10^9/L)	8.0 ± 3.9 (2.0–19.6)	7.5 ± 3.5	8.3 ± 4.2	8.7 ± 4.6	7.3 ± 3.0
PDW (%)	12.5 ± 1.5 (9.4–19.6)	12.7 ± 1.8	12.4 ± 1.3	12.6 ± 1.9	12.5 ± 1.1
P-LCR (%)	29.2 ± 5.9 (15.9–48.7)	29.4 ± 6.5	29.1 ± 5.5	29.5 ± 7.0	29.1 ± 4.7
PCT (%)	0.29 ± 0.04 (0.2–0.4)	0.28 ± 0.03	0.29 ± 0.04	0.29 ± 0.04	0.29 ± 0.04
BMI (kg/m^2)	28.1 ± 4.7 (19.6–44.2)	28.2 ± 4.1	28.0 ± 5.1	28.9 ± 5.3	27.3 ± 3.9
Sleep duration (hours)	6.8 ± 0.8 (4.5–9.0)	6.6 ± 0.7	7.0 ± 0.8	6.8 ± 0.8	6.8 ± 0.8
Sleep quality n (%)
Very good/good	73 (85)	32 (84)	41 (85)	35 (83)	38 (86)
Moderate	13 (15)	6 (16)	7 (15)	7 (17)	6 (14)
Recent HB status
Tobacco smoking
Yes	4 (5)	2 (5)	2 (4)	1 (2)	3 (7)
No	82 (95)	36 (95)	46 (96	41 (98)	41 (93)
Physical activity
Yes	38 (44)	17 (45)	21 (44)	13 (31)	25 (57)
No	48 (56)	21 (55)	27 (56)	29 (69)	19 (43)
Alcohol consumption
Yes	9 (11)	5 (13)	4 (8)	2 (5)	7 (16)
No	77 (89)	33 (87)	44 (92)	40 (95)	37 (84)
ABO-Rh distribution
A+/A- (n/n)	40/8	18/3	22/5	17/5	23/3
O+/O- (n/n)	31/7	14/3	17/4	15/5	16/2

Notes: Data are presented as mean ± SD (range) or number (percentage). Column percentages are given. *<0.05; ^†<0.01; ^‡<0.001; Mean values were compared using unpaired t-tests, categorical variables were compared using chi-square or Fisher’s exact test, as appropriate.

Abbreviations: PLT, platelets; WB, whole blood; MPV, mean platelet volume; IPF, immature platelet fraction; IPC, immature platelet count (calculated by multiplying platelet count × IPF); PDW, platelet distribution width; P-LCR, platelet-large cell ratio; PCT, plateletcrit; BMI, body mass index; HB, health behavior during past 24 hours.

Figure 1 Representative flow cytometry plots of activated platelets in pre-donation whole blood samples. (A) Whole blood samples were incubated with anti-CD61 and anti-CD62P (left) or isotype control (middle), washed and analyzed by flow cytometry. Histogram overlay of CD61-positive events is shown to the right. (B) Flow cytometry dot plots showing whole blood samples stimulated with ADP.

Table 2 Correlation of Platelet Activation in Pre-Donation Blood Samples with Collection Time and Other Covariates (n = 84)

Variables	C-Time	P-Count	MPV	IPF	IPC	PDW	P-LCR	MCV	MCH	RDW
%P-selectin+	–0.35^†	–0.03	0.29^†	0.17	0.17	0.21*	0.24*	0.33^†	0.22*	0.28*
C-time		–0.05	–0.19	0.02	–0.01	–0.12	–0.11	–0.16	–0.08	–0.05
P-count			–0.16	–0.37^†	–0.10	–0.30^†	–0.29^†	–0.06	–0.14	–0.13
MPV				0.54^‡	0.60^‡	0.81^‡	0.86^‡	0.15	0.12	0.08
IPF					0.95^‡	0.62^‡	0.63^‡	–0.01	–0.06	0.04
IPC						0.59^‡	0.62^‡	–0.01	–0.07	0.09
PDW							0.96^‡	0.02	0.02	0.00
P-LCR								0.05	0.04	0.00
MCV									0.86^‡	0.53^‡
MCH										0.38^†

Notes: Activated platelets were identified by P-selectin expression above PE-isotype control values. *<0.05; ^†<0.01; ^‡<0.001.

Abbreviations: %P-selectin+, percentage of P-selectin-expressing (ie, activated) platelets; C-time, collection time of pre-donation blood; P-count, platelet count; MCV (fL), mean corpuscular volume; MCH (pg), mean corpuscular hemoglobin; RDW (%), red blood cell distribution width. For other abbreviations, see Table 1.

Table 3 Results of Univariate (Unadjusted) and Backward Multivariable Linear (Adjusted) Models for Association of Platelet Activation in Whole Blood (n = 84) to Collection Time and Other Factors

Variables	Univariate Analyses		Multivariate Analyses
	r-value	p-value	β	Std. Error	Beta	p-value
Collection time	–0.346	0.001	–0.26	0.09	–0.29	<0.001
MPV	0.286	0.008	0.12	0.39	0.06	0.056
PDW	0.209	0.056	–0.23	0.38	–0.22	0.836
P-LCR	0.241	0.027	0.06	0.03	0.23	0.864
MCV	0.332	0.002	0.07	0.05	0.17	0.111
MCH	0.217	0.047	–0.11	0.22	– 0.10	0.997
RDW	0.275	0.011	0.19	0.07	0.28	0.005
ABO group*	0.228	0.036	0.55	0.31	0.17	0.019
			Adjusted coefficient determination [R^2] = 0.298

Notes: Data shown are correlation coefficient (r-value), regression coefficient (β) with standard error and the standardized beta coefficient. *ABO blood group was included as a categorical variable and adjusted for other factors in the model. For abbreviations, see Tables 1 and 2.

Figure 2 Relationship between (A) the percentage of P-selectin-positive platelets and (B) mean platelet volume (MPV, a surrogate marker of platelet activation) with the time of day at which blood samples were collected (collection time). The percentage of P-selectin-positive platelets, as assessed by flow cytometry, correlated inversely (r = –0.35), with the collection time. MPV, as measured via a Sysmex XE-5000 analyzer, was also inversely correlated with the collection time, although it did not reach statistical significance in this study.