ARTICLE

https://doi.org/10.1038/s41467-021-25010-x

OPEN

Malignant cerebral infarction after ChAdOx1
nCov-19 vaccination: a catastrophic variant
of vaccine-induced immune thrombotic
thrombocytopenia
1✉

M. De Michele
O. G. Schiavo1, E. Sbardella1, I. Berto7, L. Petraglia1,10, N. Caracciolo7,10, M. Chiara8, S. Truglia9 & D. Toni7

1, F. Pulcinelli4, B. Cerbelli5, E. Merenda6,

2, A. Chistolini

3, E. Nicolini

, M. Iacobucci

;
,
:
)
(
0
9
8
7
6
5
4
3
2
1

Vaccine-induced thrombotic thrombocytopenia with cerebral venous thrombosis is a syn-
drome recently described in young adults within two weeks from the first dose of the
ChAdOx1 nCoV-19 vaccine. Here we report two cases of malignant middle cerebral artery
(MCA) infarct and thrombocytopenia 9-10 days following ChAdOx1 nCoV-19 vaccination.
The two cases arrived in our facility around the same time but from different geographical
areas, potentially excluding epidemiological links; meanwhile, no abnormality was found in
the respective vaccine batches. Patient 1 was a 57-year-old woman who underwent
decompressive craniectomy despite two prior, successful mechanical thrombectomies.
Patient 2 was a 55-year-old woman who developed a fatal bilateral malignant MCA infarct.
Both patients manifested pulmonary and portal vein thrombosis and high level of antibodies
to platelet factor 4-polyanion complexes. None of the patients had ever received heparin in
the past before stroke onset. Our observations of rare arterial thrombosis may contribute to
assessment of possible adverse effects associated with COVID-19 vaccination.

1 Emergency Department, Stroke Unit, Sapienza University of Rome, Rome, Italy. 2 Neuroradiology Unit, Department of Human Neurosciences, Sapienza
University of Rome, Rome, Italy. 3 Hematology, Department of Translational and Precision Medicine, Sapienza University of Rome, Rome, Italy. 4 Department
of Experimental Medicine, Sapienza University of Rome, Rome, Italy. 5 Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of
Rome, Rome, Italy. 6 Department of Radiological, Oncological and Pathological Sciences, Sapienza University of Rome, Rome, Italy. 7 Department of Human
Neurosciences, Sapienza University of Rome, Rome, Italy. 8 Neurosurgery, Department of Human Neurosciences, Sapienza University of Rome, Rome, Italy.
9 Lupus Clinic, Rheumatology Unit, Dipartimento Di Scienze Cliniche Internistiche Anestesiologiche e Cardiovascolari, Sapienza University of Rome,
Rome, Italy. 10These authors contributed equally: L. Petraglia, N. Caracciolo.

email: M.DeMichele@policlinicoumberto1.it

✉

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

1

 
 
 
 
 
 
ARTICLE

NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

Global vaccination against severe acute respiratory syndrome-

coronarovirus-2 (Sars-cov-2) has become a priority for
mitigating COVID-19 pandemic. Chimpanzee Adenovirus
encoding the SARS-CoV-2 Spike glycoprotein (ChAdOx1) is the
Oxford-AstraZeneca vaccine (re-named Vaxzevria) authorized by
the European Medicines Agency on 29 January 2021 (ref. 1). Three
recently published papers2–4 reported case series of unusual and
mostly devastating thrombosis,
in particular cerebral venous
thrombosis, associated with thrombocytopenia and a high level of
antibodies against platelet
factor 4 (PF4)-heparin, developing
between 5 and 15 days from the first dose of ChAdOx1 nCoV-19
vaccine.

A vaccine-induced immune thrombotic thrombocytopenia
(VITT) has been suggested to name this clinical syndrome, in
order to distinguish it from the heparin-induced thrombocy-
topenia (HIT)2. Afterward, four VITT cases presenting with
ischemic stroke after vaccination with ChAdOx1 nCoV-19 were
reported5,6. A similar syndrome was also observed in some
cases of young adult women (median age was 37, range 18–59
years of age) who had received the Ad26.COV2.S vaccine
(Johnson & Johnson/Janssen)7–9. Pathophysiological mechan-
isms of post-vaccination PF4–polyanion antibodies induction
are still unknown.

Here we describe a possible variant of the VITT characterized
by a prevalent catastrophic involvement of the arterial brain
district. Both patients developed thrombocytopenia with anti-
infarct and venous
PF4/polyanion antibodies,
pulmonary and portal vein thrombosis, within 10 days after
ChAdOx1 nCoV-19 vaccination. Physicians should be aware
about this rare but severe presentation of VITT, in order to
promptly start the appropriate treatment.

large cerebral

Results
Patient characteristics. Patient 1 was a 57-year-old healthy woman
with a past medical history of mild hypothyroidism, in follow-up care
after breast cancer treated surgically in 2012. Last mammography and
breast ultrasound performed 4 months earlier were normal. She
presented to our Emergency Department (ED) 9 days after the first
dose of ChAdOx1 nCoV-19 vaccine and 1 h after the onset of sudden
left hemiplegia, right gaze deviation, dysarthria, and left neglect,
caused by right middle cerebral artery (MCA) occlusion. Since blood
tests showed severe isolated thrombocytopenia (44,000 mm3), the
patient did not receive intravenous thrombolysis and underwent
successful mechanical thrombectomy after platelet transfusion. Two
hours later she underwent a second successful endovascular treat-
ment after evidence of a repeated MCA occlusion at brain magnetic
resonance (MR) and worsening of the neurological symptoms. In
both procedures thrombus was collected and analyzed. Unfortu-
nately, 12 h apart the patient developed a malignant infarct (that is an
ischemia involving the whole territory of the MCA which causes
space-occupying cerebral edema leading to rapid neurological
deterioration)10, due to re-occlusion of the right MCA. Decom-
pressive craniectomy was performed on day 3 from stroke onset
(Fig. 1). Intravenous betamethasone 4 mg b.i.d. was prescribed. Total
body computed tomography (CT) scan showed extensive pulmonary
artery and portal vein thrombosis, with no evidence of underlying
malignancy. A trans-thoracic echocardiogram (TTE) revealed a
structurally normal heart. Transcranial
color Doppler ultra-
sonography with bubble test was negative for right-to-left shunt.
Platelet count continued decreasing with a nadir of 23,000 mm3 on
day 2. Intravenous high-dose (1 g/kg) immunoglobulins (IVIG)
were administered on days 4 and 5. Platelet count increased up to
344,000 mm3 within 3 days, but it restarted decreasing shortly after,
reaching a platelet count of 32,000 mm3 13 days after the IVIG
administration (Fig. 2). Hence, plasma exchange was performed on

three consecutively days (3 L exchange with 5% albumin replace-
ment) with a low response (Fig. 2).

Treatment with fondaparinux 2.5 mg subcutaneously u.i.d.
was started when platelet count had reached 50,000 mm3. The
patient had never been treated with heparin both in the past
and recently prior to stroke. Blood gas analysis worsened on
day 11. A control thorax CT scan was performed, which showed
a widespread ground-glass attenuation suggestive of a severe
acute respiratory distress syndrome. Patient is still hospitalized
in a critical condition.

Patient 2 was a 55-year-old healthy woman with no preexisting
conditions except for mild hypothyroidism. She had never been
treated with heparin. Seven days after receiving the first dose of
ChAdOx1 nCoV-19 vaccine she started complaining of abdom-
inal pain and presented to a first aid on the morning of day 10th.
Routine blood examinations were normal except for the high level
of D-dimer (5441 ng/mL) and mild thrombocytopenia (PLT
133,000). Abdominal ultrasound was normal. In the afternoon,
during her hospital stay, she experienced a transient episode of
aphasia and right hemiparesis, followed 2 h later by generalized
seizures and coma. Orotracheal intubation was performed. Brain
CT scan, angio-CT, and perfusion CT showed occlusion of the
right internal carotid artery terminus and of the left MCA,
extensive ischemic cores and severe bilateral hypoperfusion,
without treatable penumbra (Fig. 3).

The patient was transferred to our ED. Blood examination
repeated 10 h later showed worsening of thrombocytopenia
(PLT 97,000 mm3) with a further decrease on the following day
(PLT 59,000 mm3). Treatment with IVIG (1 g/kg per day) and
dexamethasone 40 mg u.i.d. were started on day 1. TTE was
normal. Twelve-hour post-stroke a total body CT revealed
extensive portal vein thrombosis with occlusion of the left
intrahepatic branches
subsegmental
pulmonary arteries thrombosis. Brain CT scan showed bilateral
malignant MCA infarct with uncal herniation (Fig. 3). Brain
death was declared 24 h later. Autopsy was not performed due
to consent denial by relatives.

and left

lower

lobe

Laboratory testing. D-dimer levels were elevated in both patients
at admission. Coagulation parameters were normal. No signs of
hemolysis were evident.

A peripheral blood smear showed giant platelets in patient 1

and platelet anisocytosis in patient 2.

Patient 1 developed a progressive severe normochromic
normocytic anemia with a nadir hemoglobin level of 5.4 g/dL
on day 4 when red blood cell transfusion was performed. Patient
received a second transfusion on day 14 for a subsequent
reduction of hemoglobin level (nadir 7.4 g/dL).

Screening for thrombophilia was negative in both patients.
Lupus anticoagulant, anticardiolipin and beta-2 glycoprotein
antibodies and antinuclear antibodies tested negative.

Coagulation Factor VIII was found increased in both the
patients. Factor XIII was markedly decreased in patient 1 and not
dosed in patient 2. We observed increased circulating levels of
von Willebrand Factor Antigen (VWF:Ag) and an increase in the
levels of VWF-Ristocetin Cofactor (VWF:RCo).

Platelet testing. Serum of both patients showed high levels of pan
antibodies (IgG, IgM, and IgA) to PF4–polyanion complexes.
However, patient 1 was initially not positive but high levels of
antibodies were found at day 15 (1.68 optical density—OD405).
Patient 2 resulted positive since day 1 (1.29 OD405). Functional
activity test showed that platelets from healthy donors were
clearly activated by patient 1 and 2 sera in the absence of added
heparin (saline), with a percent of ATP release after 20 min of

2

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

 
 
 
 
 
 
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

ARTICLE

Fig. 1 Radiological findings in Patient 1. a Computed tomograghy angiography (CTA) showed proximal M1 segment occlusion of the right middle cerebral
artery (MCA) (yellow arrow); b, c CT perfusion (CTP) maps: mean transit time (MTT) (b) and cerebral blood volume (CBV) (c) showed a large area of
mismatch indicating salvageable penumbra; d, e endovascular mechanical thrombectomy achieved completed recanalization (e) of the MCA occlusion
(white arrow in d); f, g MCA reocclusion on M2 segment 2 h after the procedure, as showed by 3D time of flight (TOF) magnetic resonance imaging
(MRI) sequence (white arrow in f), with ischemic penumbra on mean transit time map in perfusion MRI (g); h, i second complete endovascular
recanalization (h before and i after the mechanical thrombectomy); j, k fluid attenuated inversion recovery MRI at 12 h showed the extension of ischemia to
superficial and deep right MCA territory with mass effect on the ventricular system (j), with occlusion of MCA at TOF magnetic resonance angiography
(white arrow in k). Extension of the arterial occlusion to the right internal carotid artery (yellow asterisk in l and m).

22% and 14%, respectively. Platelet activation was not inhibited
efficiently by high-dose heparin in both the patients (percent of
ATP release after 20 min of 19% and 9%, respectively). Serum
from healthy volunteers did not induce platelet secretion.

Table 1.

COVID-19 serological testing. Serum antibodies to spike protein
of Sars-CoV-2 (IgG) were positive in patient 1 and negative in
patient 2. Both the patients had negative COVID-19 real-time
reverse transcriptase-polymerase chain reaction (rRT-PCR) test
on nasopharyngeal swab. Patient 2 bronchoalveolar lavage (BAL)
tested on day 1 was also negative.

Clot analysis. Clot collected from patient 1 (Supplementary
Fig. 1) during the first thrombectomy was mainly composed of
platelets (85% of the total material examined) and was massively
infiltrated by neutrophils with scarce evidence of karyorrhexis.
Histological features consistent with the presence of neutrophil
DNA extracellular traps (NETs) were observed.

Clot collected during the second endovascular procedure was a
red-blood-cell-rich thrombus (90% of red blood cells and 10% fibrin
and platelets) with scarce neutrophils (Supplementary Fig. 2).

For detailed description of laboratory findings see Supplementary

Discussion
infarct, systemic
We present two cases of malignant cerebral
venous thrombosis and concomitant thrombocytopenia in two
young healthy adult women within 10 days from vaccination
against SARS-CoV-2 with ChAdOx1 nCov-19. High serum levels
of antibodies to PF4–polyanion complexes were found in both
patients.

Some relevant findings from these two cases need considerations.
Arterial thrombosis in VITT seems to be a rare event com-
paring to venous thrombosis. A population-based cohort study
in Norway and Denmark described an increased rate of venous
thromboembolic events but not of arterial events among reci-
pients of ChAdOx1 nCoV-19 (ref. 11). Greinacher et al.2
reported 1 patient, out of 11, with aortoiliac thrombosis whereas
Scully et al.4 described two patients, out of 23, affected by MCA
occlusion. In Shultz’s case series no arterial thrombosis was
reported3. More recently, Blauenfeld et al.5 have reported
the first case of VITT in Denmark who developed bilateral

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

3

 
 
 
 
 
 
ARTICLE

NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

Fig. 2 Platelet count course in Patient 1. AIS acute ischemic stroke, MT mechanical thrombectomy, DC decompressive craniectomy, PVT portal vein
thrombosis, ARDS acute respiratory distress syndrome. Source data are provided as a Source Data file.

Fig. 3 Radiological findings in Patient 2. a Computed tomograghy angiography (CTA) showed the proximal M1 segment occlusion of the left middle
cerebral artery (MCA) (black arrow) and the occlusion of the right internal carotid artery terminus (yellow arrow); b CT perfusion (CTP) showed bilateral
infarct core (cerebral blood volume—CBV—map in c) and hypoperfusion (mean transit time—MTT—map in b), without treatable penumbra; d 24 h after,
brain CT revealed bilateral malignant MCA infarcts; e portal vein thrombosis with extension to the left intrahepatic branches (black arrow); f subsegmental
pulmonary artery thrombosis (white arrow).

adrenal hemorrhage and a fatal massive ischemic stroke treated
with decompressive craniectomy, 7 days after receiving the
ChAdOx1 nCoV-19 vaccination. Three other cases of VITT
presenting with ischemic stroke after the administration of the
ChAdOx1 nCoV-19 vaccine in United Kingdom have been
described6.

VITT appears to be similar to autoimmune HIT2. Range of
thrombotic complications in HIT is broad with 15–20% of
patients suffering arterial events, and 30–60% developing venous
clots (some patients get a combination of venous and arterial
thrombosis)12. The endothelium has been identified as a potential
target for HIT antibodies. Recently, it has been demonstrated that

4

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

 
 
 
 
 
 
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

ARTICLE

VWF strings released from Weibel–Palade bodies activated
endothelium, are capable of binding PF4 released from activated
platelets. The resulting PF4/VWF complexes forming antigen
sites are recognized by HIT antibodies with consequent greater
accumulation of platelets to the injured arterial endothelium,
thrombus formation, and propagation13.

Patient 1 had a right MCA stroke due to occlusion of the
M1 segment. Despite two successful endovascular treatments, the
patient experienced a repeated occlusion of the same vessel and
developed a malignant MCA infarct. We can hypothesize that the
second and the third occlusion at the same vascular site might be
due to the PF4/VWF complexes recognized by HIT antibodies on
the post-thrombectomy injured arterial endothelium. A high level
of VWF and VWF:RCo, D-dimer, and coagulation Factor VIII
support this hypothesis, suggesting a high prothrombotic state
and endothelium dysfunction. Notably, at the time patient 1 was
admitted, we were not yet aware of VITT, as the first paper
reporting this syndrome was published a few days later. Hence,
finding the low platelet count, a platelet transfusion was pre-
scribed before the first thrombectomy, probably exacerbating the
hypercoagulable state.

Histology of the first removed thrombus showed a platelet-
rich clot with neutrophils, while the second one showed a red-
blood-cell-rich clot. Platelet-rich thrombi are formed by VWF,
NETs, and fibrin14. Platelet/fibrin thrombi were also found in
the experimental model of HIT in veins and arteries of multiple
organs15 and HIT is also known for this reason, as “white clot
syndrome”16.

We cannot say whether histological differences of the two
thrombi account for different pathophysiology. The main dif-
ference between clots was their age. Thrombus formation is a
highly dynamic process: first layers of red blood cells and fibrin
are main components of thrombotic material, and then platelets
and white blood cells become prevalent. The second thrombus
was retrieved earlier (about 2 h after its formation), so we can
assume that it was fresher than the first one17.

Pathogenesis of the severe normochromic normocytic anemia
developed by patient 1 was not defined. We ruled out active
bleeding through abdomen CT scan, no melena, or hematemesis
were evident. Laboratory findings excluded hemolytic anemia.
Bone marrow examination was not done due to the critical
conditions of the patient and because of the stability of the red
blood cells count after transfusion.

We found a high level of Sars-cov-2 IgG serum antibodies
suggestive of a previously COVID-19 asymptomatic infection in
patient 1. No Sars-CoV-2 IgG serum antibodies were present in
serum of patient 2. These data would suggest that anti-spike
protein antibodies might not be relevant in the pathogenesis of
to PF4–polyanion
this
complexes were initially negative in serum of patient 1, while a
high antibody level was found in serum collected and tested
2 weeks later.

Interestingly, antibodies

syndrome.

The serologic pattern of these two cases is similar to that of the
so-called autoimmune HIT, since serum from our patients strongly
activated platelets in the absence of heparin (heparin-independent
platelet activation)18. Moreover, platelet secretion was scarcely
sensitive to inhibition with high-dose heparin, as observed in two
patients described by Schultz et al.3 and in one described by
Greinacher et al.2. A disseminated intravascular coagulation was not
observed in our patients.

Both the patients had extensive arterial thrombosis of the lung
and venous thrombosis of the splanchnic district. A hypercoa-
gulable state was observed in both the patients with a high level of
VWF and VWF:RCo, D-dimer, and coagulation Factor VIII.
Factor XIII was markedly decreased in patient 1, probably due to
excessive consumption within thrombi19.

Treatment of this syndrome is challenging. Administration of
high-dose IVIG has been suggested in an attempt to inhibit Fcγ
receptor-mediated platelet activation, since in the treatment of
severe autoimmune HIT this treatment resulted in rapid increase
in platelet count18,20. Unfortunately, platelet response of patient 1
to high-dose IVIG was striking but
transient. Poor platelet
increase was observed after three sessions of PEx. Early non-
heparin anticoagulation,
in addition to high-dose IVIG and
steroids, is warranted in VITT to prevent thrombosis21,22 and, in
view of the patient 1 clinical course, it should have been con-
sidered soon after thrombectomy in order to prevent reocclusion
of the damaged vessel. However, early anticoagulation of patients
affected by large brain infarct increases the risk of hemorrhagic
transformation23. Moreover,
the patient underwent decom-
pressive craniectomy on day 3, which is an additional contra-
indication to early full-dose anticoagulation. Hence, we started
with Fondaparinux 2.5 mg per day and we increased the dosage to
7.5 mg per day after 2 weeks from stroke onset.

Many scientific questions regarding VITT are still unanswered
as highlighted by a recently published editorial24. Although lim-
ited to two patients, data collected in this report strengthen the
hypothesis that platelets could be initially activated by unknown
factors related to the ChAdOx1 nCov-19 vaccine/host interaction.
An atypical autoimmune HIT may be the consequent event that
could amplify platelet activation and coagulation cascade. The
rare occurrence of VITT also in patients who received the Ad26.
COV2.S vaccine (Johnson & Johnson/Janssen)7–9 suggests a
possible pathogenic role of the adenoviral vector. In addition, a
potential role for the ChAdOx1 nCov-19 vaccine cell-culture-
derived proteins and components (i.e. EDTA) has been recently
postulated25. More investigations are warranted to elucidate this
and other hypotheses26 and clarify the pathogenesis of this cat-
astrophic syndrome.

One limitation of the present paper is that we have not tested
the PF4 antibody titer immediately after plasma exchange, and
hence we do not know whether the recurrent thrombocytopenia
was due to a persistent high level of PF4 antibody titer. Another
limitation is that we have not dosed soluble DNA or citrullinated
in the patients’ sera. NETs are
histones, markers of NETs,
released from the neutrophils in response to extracellular stimuli
to contain infection, but their excessive formation can be detri-
mental by inducing microvascular thrombosis and propagating
inflammation27. Greinacher et al.25 observed that VITT patient
serum incubated with PF4 and purified neutrophils induces NETs
formation (NETosis) with strongly enhanced response in the
presence of platelets. NETosis could play a role in amplifying
thrombi formation in VITT, but further studies are needed to
confirm this issue.

In conclusion, our two cases of young adult women with massive
brain artery thrombosis in addition to extensive systemic venous
thrombocytopenia, and PF4–polyanion antibodies,
thrombosis,
developed within 10 days from ChAdOx1 nCov-19 vaccination,
might represent a stroke variant of the recently named VITT syn-
drome. Ischemic stroke may be the first serious symptom of VITT.
Physicians should be aware of this possibility and should carefully
investigate patient medical history asking for any previous vacci-
nation (within 4–28 days) especially in young female stroke
patients. If low platelet count would be present (it could also be
mild initially), a chest and abdominal CT scan should be performed
to rule out venous thrombosis, together with an anti-PF4/heparin
IgG enzyme immunoassay and functional platelet assay. If diagnosis
of VITT is confirmed (but also if the suspect of VITT is high,
without waiting for specific tests), non-heparin anticoagulation
should be promptly started, if platelet count is up to 50 × 109/L, as
well as therapy with high dose of steroids and IVIG. Platelet
transfusion should be avoided.

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

5

 
 
 
 
 
 
ARTICLE

NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

Methods
Functional and serologic testing. Both the patients were tested for COVID-19
through a nose/throat rRT-PCR test. A molecular test for COVID-19 on BAL was
also performed for patient 2.

Serum antibodies (IgG) to the spike protein of Sars-cov-2 were measured with

the Liaison-CLIA-DiaSorin system on blood samples of both the patients.

Coagulation Factors VIII and XIII, total levels of VWF:Ag, and its capability to

adhere to platelet glycoprotein complex GPIb-IX-V (VWF:RCo) were dosed.

Antibodies to PF4/polyanion in serum were tested twice in patient 1 (on day 1
and on day 15) and once in patient 2 (on day 1), by using a commercial enzyme
immunoassay (IgG/IgA/IgM, Immucor, Lifecodes, Waukesha, WI).

Functional test with high concentration heparin (100 IU/ml) was performed28.

Briefly, 73 μL of washed platelet suspension was dispensed in microplates and
activated with heat-inactivated (56 °C for 30 min) patient serum (27 μL) in the
presence or absence of high heparin dose (100 U/mL) and left for 20 min at room
temperature to allow activation. To avoid any possible thrombin effect Hirudin
(5 U/ml) was added to the patient sera. ATPlite (PerkinElmer, Waltham, MA), and
ATP monitoring system based on luciferin/luciferase method, (25 μL) was then
added to the microplates. Emitted light was measured with a plate luminometer
(Victor 3, PerkinElmer). All dosages were performed in duplicate. Effective ATP
release was calculated according to the company’s instructions. Platelet suitability
was evaluated with U-46619 plus epinephrine (2 + 20 μM)-induced ATP release.
Data are reported as a percentage of ATP release induced by serum/total
intraplatelet ATP amount.

Clot analysis. Clot material from patient 1 was removed from the stent retriever
during the first endovascular procedure and collected by the thromboaspiration
technique during the second procedure. Histological sections obtained from the
formalin-fixed paraffin-embedded material were stained with hematoxylin–eosin
and phosphotungstic acid–hematoxylin. Immunohistochemistry for CD61 was also
performed to highlight the platelet component.

Informed consent. Written informed consent was obtained from all participants
according to Italian regulation, and the experimental procedure was approved by
the Institutional Review Board at Sapienza University of Rome and was conducted
in accordance with the Declaration of Helsinki.

Reporting summary. Further information on research design is available in the Nature
Research Reporting Summary linked to this article.

Data availability
The data of this report are available from the corresponding author upon reasonable
requests. Source data are provided with this paper.

Received: 8 June 2021; Accepted: 18 July 2021;

References
1. Voysey, M. et al. Safety and efficacy of the ChAdOx1 nCoV-19 vaccine

(AZD1222) against SARS-CoV-2: an interim analysis of four randomised
controlled trials in Brazil, South Africa, and the UK. Lancet 397, 99–111
(2021).

2. Greinacher, A. et al. Thrombotic thrombocytopenia after ChAdOx1 nCov-19

Vaccination. N. Engl. J. Med. 384, 2092–2101 (2021).
Schultz, N. H. et al. Thrombosis and thrombocytopenia after ChAdOx1
nCoV-19 vaccination. N. Engl. J. Med. 384, 2124–2130 (2021).
Scully, M. et al. Pathologic antibodies to platelet factor 4 after ChAdOx1
nCoV-19 vaccination. N. Engl. J. Med. https://doi.org/10.1056/
NEJMoa2105385 (2021).
Blauenfeldt, R. A. et al. Thrombocytopenia with acute ischemic stroke and
bleeding in a patient newly vaccinated with an adenoviral vector-based
COVID-19 vaccine. J. Thromb. Haemost. https://doi.org/10.1111/jth.15347
(2021).

6. Al-Mayhani, T. et al. Ischaemic stroke as a presenting feature of

ChAdOx1 nCoV-19 vaccine-induced immune thrombotic thrombocytopenia.
J. Neurol. Neurosurg. Psychiatry https://doi.org/10.1136/jnnp-2021-326984
(2021).

7. Muir, K. L., Kallam, A., Koepsell, S. A. & Gundabolu, K. Thrombotic

thrombocytopenia after Ad26.COV2.S vaccination. N. Engl. J. Med. 384,
1964–1965 (2021).
Sadoff, J., Davis, K. & Douoguih, M. Thrombotic thrombocytopenia after
Ad26.COV2.S vaccination—response from the manufacturer. N. Engl. J. Med.
384, 1965–1966 (2021).

3.

4.

5.

8.

6

9.

See, I. et al. US Case Reports of Cerebral Venous Sinus Thrombosis with
Thrombocytopenia after Ad26.COV2.S Vaccination March 2 to April 21 2021.
JAMA 325, 2448–2456 (2021).

10. Hacke, W. et al. ‘Malignant’ middle cerebral artery territory infarction: clinical

course and prognostic signs. Arch. Neurol. 53, 309–315 (1996).
11. Pottegård, A. et al. Arterial events, venous thromboembolism,

thrombocytopenia, and bleeding after vaccination with Oxford-AstraZeneca
ChAdOx1-S in Denmark and Norway: population based cohort study. BMJ
373, n1114 (2021).

12. Warkentin, T. E. HIT: still stringing us along. Blood 135, 1193–1194

13.

(2020).
Johnston, I. et al. Recognition of PF4-VWF complexes by heparin-induced
thrombocytopenia antibodies contributes to thrombus propagation. Blood
135, 1270–1280 (2020).

14. Staessens, S. et al. Structural analysis of ischemic stroke thrombi: histological
indications for therapy resistance. Haematologica 105, 498–507 (2020).
15. Reilly, M. P. et al. Heparin-induced thrombocytopenia/thrombosis in a

transgenic mouse model requires human platelet factor 4 and platelet
activation through FcgammaRIIA. Blood 98, 2442–2447 (2001).
16. Towne, J. B., Bernhard, V. M., Hussey, C. & Garancis, J. C. White clot

syndrome. Peripheral vascular complications of heparin therapy. Arch. Surg.
114, 372–377 (1979).

17. Bacigaluppi, M., Semerano, A., Gullotta, G. S. & Strambo, D. Insights from

thrombi retrieved in stroke due to large vessel occlusion. J. Cereb. Blood Flow
Metab. 39, 1433–1451 (2019).

18. Greinacher, A., Selleng, K. & Warkentin, T. E. Autoimmune heparin-induced

thrombocytopenia. J. Thromb. Haemost. 15, 2099–2114 (2017).

19. Ząbczyk, M., Natorska, J. & Undas, A. Factor XIII and fibrin clot properties in
acute venous thromboembolism. Int. J. Mol. Sci. 22, https://doi.org/10.3390/
ijms22041607 (2021).

20. Warkentin, T. E. High-dose intravenous immunoglobulin for the treatment
and prevention of heparin-induced thrombocytopenia: a review. Expert Rev.
Hematol. 12, 685–698 (2019).

21. Thaler, J. et al. Successful treatment of vaccine-induced prothrombotic

immune thrombocytopenia (VIPIT). J. Thromb. Haemost. https://doi.org/
10.1111/jth.15346 (2021).

22. Graf, T. et al. Immediate high-dose intravenous immunoglobulins followed by
direct thrombin-inhibitor treatment is crucial for survival in Sars-Covid-19-
adenoviral vector vaccine-induced immune thrombotic thrombocytopenia
VITT with cerebral sinus venous and portal vein thrombosis. J. Neurol. 1–3
(2021).

23. Paciaroni, M. et al. Hemorrhagic transformation in patients with acute

ischemic stroke and atrial fibrillation: time to initiation of oral anticoagulant
therapy and outcomes. J. Am. Heart Assoc. 7, e010133 (2018).

24. Cines, D. B. & Bussel, J. B. SARS-CoV-2 vaccine-induced immune thrombotic

thrombocytopenia. N. Engl. J. Med. https://doi.org/10.1056/NEJMe2106315
(2021).

25. Greinacher, A. et al. Towards understanding ChAdOx1 nCov-19 vaccine-induced
immune thrombotic thrombocytopenia (VITT). Preprint at https://doi.org/
10.21203/rs.3.rs-440461/v1 (2021).

26. Douxfils, J. et al. Hypotheses behind the very rare cases of thrombosis with
thrombocytopenia syndrome after SARS-CoV-2 vaccination. Thromb. Res.
203, 163–171 (2021).

27. Papayannopoulos, V. Neutrophil extracellular traps in immunity and disease.

Nat. Rev. Immunol. 18, 134–147 (2018).

28. Guarino, M. L. et al. New platelet functional method for identification of
pathogenic antibodies in HIT patients. Platelets 28, 728–730 (2017).

Acknowledgements
This research did not receive a grant from funding agency.

Author contributions
M.D.M. conceived and designed the study, enrolled the patients, interpreted the results,
and prepared the original manuscript. M.I. is the neurointerventional radiologist who
retrieved the thrombi of patient 1, reviewed, and edited Figs. 1 and 3 of the manuscript.
A.C. is the hematologist consultant who provided treatment decision making and coa-
gulation parameters’ analysis; E.N. edited the table and Fig. 2 and critically reviewed the
manuscript., O.G.S., E.S., I.B., L.P. and N.C. participated in data patients collection; B.C.
and E.M. performed the histopathological examination of retrieved thrombi and edited
Supplementary Fig. 1; F.P. performed the functional and serological platelets analysis and
contributed to the interpretation of results; M.C., S.T. and D.T. critically reviewed and
edited the manuscript.

Competing interests
The authors declare no competing interests.

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

 
 
 
 
 
 
NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-021-25010-x

ARTICLE

Additional information
Supplementary information The online version contains supplementary material
available at https://doi.org/10.1038/s41467-021-25010-x.

Correspondence and requests for materials should be addressed to M.D.M.

Peer review information Nature Communications thanks Massimo Radin and the other,
anonymous, reviewer(s) for their contribution to the peer review of this work. Peer
reviewer reports are available.

Reprints and permission information is available at http://www.nature.com/reprints

Open Access This article is licensed under a Creative Commons
Attribution 4.0 International License, which permits use, sharing,
adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative
Commons license, and indicate if changes were made. The images or other third party
material in this article are included in the article’s Creative Commons license, unless
indicated otherwise in a credit line to the material. If material is not included in the
article’s Creative Commons license and your intended use is not permitted by statutory
regulation or exceeds the permitted use, you will need to obtain permission directly from
the copyright holder. To view a copy of this license, visit http://creativecommons.org/
licenses/by/4.0/.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.

© The Author(s) 2021

NATURE COMMUNICATIONS |  

 (2021) 12:4663  | https://doi.org/10.1038/s41467-021-25010-x | www.nature.com/naturecommunications

7