Since the emergence of SARS-CoV-2 in China in December 2019 and the subsequent coronavirus
disease 2019 (COVID-19), and as a consequence of the rapid spread worldwide, the World
Health Organization (WHO) declared the outbreak a serious public health emergency
of international concern.[1]
COVID-19 is now considered a multisystem disease with deregulation of multiple physiological
pathways, including hemostasis.[2] Indeed, several lines of evidence established a link between COVID-19 severity and
thrombotic events, including thrombocytopenia, platelet hyperreactivity, and severe
bleeding.[3]
[4]
[5]
[6]
[7]
[8]
To face the pandemic situation, development of vaccines seems to be crucial to prevent
COVID-19 infection. In the past few months, numerous vaccines have been developed,
among them the Oxford–AstraZeneca ChAdOx1 nCoV-19 (Oxford-AstraZeneca) vaccine (AZD1222).
This vaccine is a chimpanzee adenoviral-vectored vaccine with full length SARS-CoV-2
spike insert, developed at the University of Oxford. Randomized trials have allowed
the assessment of the Oxford-AstraZeneca vaccine safety and efficacy.[9]
[10] In the original protocol trial, volunteers aged 18 years and older received two
standard doses of 5 × 1010 viral particles per dose administered 28 days apart. The protocol amendments allowed
other trial participants to receive a booster more than 28 days after their first
dose (up to 12 weeks).[9]
[10]
[11] Overall, vaccine efficacy after a single standard dose from day 22 to day 90 after
vaccination was 76.0%. More than 14 days after the second dose, vaccine efficacy was
66.7%. Efficacy was higher in participants who received a longer prime-boost interval
than in those with a short interval (<6 weeks). There were no hospital admissions
for COVID-19 in the Oxford-AstraZeneca vaccine group after the initial 21-day exclusion
period. There were two deaths in the Oxford-AstraZeneca vaccine group, but these were
considered unrelated to vaccination.[9]
[10]
[11]
Recently, several cases of thrombosis, thrombocytopenia, and severe bleeding have
been reported following administration of the Oxford-AstraZeneca vaccine.[12] Denmark was the first country to temporarily suspend the use of the Oxford-AstraZeneca
vaccine as a precautionary move after reports of thrombosis in some people, including
one person who developed pulmonary embolism and died 10 days after receiving at least
one dose. Several other European countries soon followed suit.[12] In total, 37 thrombosis cases have been reported out of more than 17 million people
vaccinated in the European Union and Britain. Five of the cases were deep vein thrombosis,
and 22 were pulmonary embolisms.[13]
Although platelets from COVID-19 patients have shown hyperreactivity,[4]
[5]
[6] no scientific data have been able to demonstrate platelet dysfunction in people
injected with the Oxford-AstraZeneca vaccine.
The WHO and the European Medicines Agency (EMA) stated that the association of the
vaccine with the increased risk of blood clots is not justifiable and advised to continue
vaccinations.[13]
Therefore, this study aimed to assess platelet function by measuring the aggregation
of platelets from participants who had been injected with the Oxford-AstraZeneca vaccine
compared with platelets from a cohort of healthy donors.
Participants vaccinated with a single dose (n = 35) and with two doses (n = 42) and patients with severe COVID-19 (n = 10) who were admitted to the Cheikh Zaïd Hospital of Abulcasis University (Rabat,
Morocco) were included in this study. Of note, the second dose was injected 28 days
after the first injection following the original protocol for administration of the
Oxford-AstraZeneca vaccine. Sex- and age-matched healthy blood donors were used as
controls. Recruitment was approved by the Ethics Committee of Cheikh Zaïd Hospital
(CEFCZ/PR/2020/PR04) and complies with the Declaration of Helsinki. All participants
gave their written informed consent. Patients who were receiving medications that
interfere with platelet function within 2 weeks before the experiment started were
excluded from the study. Vaccinated participants who previously had COVID-19 were
also excluded from this study. [Table 1] provides basic demographic and clinical data for patients.
Table 1
Demographic and clinical data of study participants
|
Index
|
Healthy donors
|
AZ one dose
|
AZ two doses
|
p-Value
|
|
Healthy vs. one dose
|
Healthy vs. two doses
|
One dose vs. two doses
|
|
No. of patients
|
10
|
35
|
42
|
|
|
|
|
Female/male
|
4/6
|
16/19
|
23/19
|
–
|
–
|
–
|
|
Age, y
|
51.57 ± 14.98
|
56.15 ± 8.12
|
54.15 ± 13.10
|
0.62
|
0.69
|
0.86
|
|
Weight, kg
|
83.26 ± 23.14
|
76.44 ± 10.10
|
75.68 ± 9.38
|
0.37
|
0.35
|
>0.99
|
|
Platelet number, ×109/L
|
217 ± 81.28
|
230 ± 88.93
|
208 ± 101.14
|
0.83
|
0.18
|
0.22
|
|
Lymphocyte number, × 109/L
|
1.11 ± 0.82
|
1.18 ± 0.66
|
1.24 ± 0.89
|
0.34
|
0.21
|
0.91
|
|
ALT, U/L
|
18.32 ± 6.03
|
15.32 ± 6.80
|
18.16 ± 4.57
|
0.49
|
>0.99
|
0.49
|
|
AST, U/L
|
16.49 ± 4.24
|
15.91 ± 3.42
|
15.44 ± 4.96
|
0.96
|
0.92
|
>0.99
|
|
LDH, U/L
|
362.86 ± 115.71
|
404.66 ± 149.92
|
329.50 ± 178.25
|
0.38
|
0.24
|
0.07
|
|
C-reactive protein, mg/L
|
9.13 ± 4.08
|
14.59 ± 6.52
|
16.19 ± 7.89
|
0.05
|
0.04
|
0.90
|
|
D-dimers, mg/L
|
0.42 ± 0.32
|
0.96 ± 0.21
|
0.89 ± 0.58
|
0.01
|
0.01
|
0.76
|
|
Fibrinogen, mg/dL
|
314.36 ± 91.52
|
330.18 ± 80.07
|
326.10 ± 96.15
|
0.44
|
0.46
|
0.88
|
Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; AZ,
AstraZeneca; LDH, lactate dehydrogenase.
Note: Data are presented as mean ± standard deviation. Statistical analysis: unpaired
Student's t-test was used to calculate p-values. Bold numbers indicate statistical significance at p ≤ 0.05.
Washed platelets were prepared as previously described.[14] Briefly, platelet-rich plasma (PRP) was obtained by centrifugation of acid citrate
dextrose (ratio of 1:5) anticoagulated blood at 200 g for 15 minutes. Platelets were
then pelleted from PRP, to which 1 mg/mL of Prostaglandin E1 (PGE1) was added, washed with HBSS-Hank's sodium citrate buffer (138-mM NaCl, 5-mM KCl,
0.34-mM Na2HPO4, 0.4-mM KH2PO4, 4.2-mM Na2HCO3, 5.6-mM glucose, 10-mM HEPES, 12.9-mM sodium citrate, pH 7.4), also containing PGE1 (0.5 mg/mL), and finally resuspended in HBSS-Hank's buffer containing 2-mM MgCl2 and 2-mM CaCl2. Platelets were adjusted to 250 × 106/mL and allowed to rest at 37°C for 30 minutes before platelet aggregation tests.
Platelet aggregation was monitored and recorded as previously described using an 8-channel
optical aggregometer (SD Medical Innovation; Frouard, France).[15]
Our results show, as demonstrated in several published studies,[4]
[5]
[6] that platelets from COVID-19 patients are hyperreactive ([Fig. 1A]). In addition, our findings document that vaccination by Oxford-AstraZeneca vaccine
(one or two doses) was not associated with enhanced platelet aggregation.
Fig. 1 Oxford-AstraZeneca vaccine does not affect platelet aggregation. Platelet aggregation
was evaluated in healthy donors (n = 10), patients with severe coronavirus disease 2019 (COVID-19) (n = 10), participants who received a single dose of AstraZeneca vaccine (n = 35), and participants who received two doses of AstraZeneca vaccine (n = 42). Quantification (%) of maximal platelet aggregation after stimulation with
(A) 0.05 or (B) 2 U/mL of α-thrombin. Data are represented as mean ± SD. Statistical analysis: data
were normally distributed (Shapiro − Wilk test). One-way ANOVA (analysis of variance)
with subsequent Sidak multiple comparisons test. ****p < 0.0001.
Indeed, in response to a suboptimal α-thrombin concentration (0.05 U/mL), no significant
difference in platelet aggregation was recorded ([Fig. 1A]). At a higher concentration of α-thrombin (2 U/mL), platelets from vaccinated people
aggregate at a maximum rate >90% ([Fig. 1B]).
Overall, our findings demonstrate that platelets from vaccinated participants are
not hyperreactive. To the best of our knowledge, this is the first report that implies
a normal platelet function of vaccinated participants. These findings do not support
any link between thrombosis and vaccination with the Oxford-AstraZeneca vaccine.
