Clinical Infectious Diseases

Major Article

Intravenous Injection of Coronavirus Disease 2019

(COVID-19) mRNA Vaccine Can Induce Acute
Myopericarditis in Mouse Model
Can Li,1,a Yanxia Chen,1,a Yan Zhao,1,a David Christopher Lung,2 Zhanhong Ye,1 Wenchen Song,1 Fei-Fei Liu,1 Jian-Piao Cai,1 Wan-Man Wong,1

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Cyril Chik-Yan Yip,1 Jasper Fuk-Woo Chan,1,3,4 Kelvin Kai-Wang To,1,3 Siddharth Sridhar,1,3 Ivan Fan-Ngai Hung,3,5 Hin Chu,1 Kin-Hang Kok,1 Dong-Yan Jin,6
Anna Jinxia Zhang,1,b and Kwok-Yung Yuen1,3,4,b
1
State Key Laboratory of Emerging Infectious Diseases, Carol Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong
Kong Special Administrative Region, China; 2Department of Pathology, Queen Elizabeth Hospital and Hong Kong Children’s Hospital, Hong Kong, Hong Kong Special Administrative Region, China;
3
Department of Clinical Microbiology and Infection Control, The University of Hong Kong-Shenzhen Hospital, Shenzhen, Guangdong, China; 4Hainan Medical University-The University of Hong
Kong Joint Laboratory of Tropical Infectious Diseases, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; 5Department of Medicine, Li Ka Shing Faculty of
Medicine, The University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; and 6School of Biomedical Sciences, Li Ka Shing Faculty of Medicine, The University of Hong
Kong, Pokfulam, Hong Kong Special Administrative Region, China

Background.  Post-vaccination myopericarditis is reported after immunization with coronavirus disease 2019 (COVID-19)
messenger RNA (mRNA) vaccines. The effect of accidental intravenous injection of this vaccine on the heart is unknown.
Methods.  We compared the clinical manifestations, histopathological changes, tissue mRNA expression, and serum levels of cy-
tokine/chemokine in Balb/c mice at different time points after intravenous (IV) or intramuscular (IM) vaccine injection with normal
saline (NS) control.
Results.  Although significant weight loss and higher serum cytokine/chemokine levels were found in IM group at 1–2  days
post-injection (dpi), only IV group developed histopathological changes of myopericarditis as evidenced by cardiomyocyte degener-
ation, apoptosis, and necrosis with adjacent inflammatory cell infiltration and calcific deposits on visceral pericardium, although ev-
idence of coronary artery or other cardiac pathologies was absent. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)
spike antigen expression by immunostaining was occasionally found in infiltrating immune cells of the heart or injection site, in
cardiomyocytes and intracardiac vascular endothelial cells, but not skeletal myocytes. The histological changes of myopericarditis
after the first IV-priming dose persisted for 2 weeks and were markedly aggravated by a second IM- or IV-booster dose. Cardiac
tissue mRNA expression of interleukin (IL)-1β, interferon (IFN)-β, IL-6, and tumor necrosis factor (TNF)-α increased significantly
from 1 dpi to 2 dpi in the IV group but not the IM group, compatible with presence of myopericarditis in the IV group. Ballooning
degeneration of hepatocytes was consistently found in the IV group. All other organs appeared normal.
Conclusions.  This study provided in vivo evidence that inadvertent intravenous injection of COVID-19 mRNA vaccines may
induce myopericarditis. Brief withdrawal of syringe plunger to exclude blood aspiration may be one possible way to reduce such risk.
Keywords.  mouse model; SARS-CoV-2; mRNA vaccine; intravenous; COVID-19.

Safe and effective whole-population vaccination against severe (COVID-19) pandemic [1], which has caused about 200 mil-
acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the lion cases of COVID-19 globally and over 4 million deaths by
only long-term solution to the ongoing coronavirus disease 2019 17 July 2021[2]. However, the one-dose vaccination rate in the
United States and United Kingdom was only 54.9% and 67.8%,
respectively, as of 10 July 2021[3]. Vaccine hesitancy among the
 

general public is a significant problem and is partially driven

Received 26 July 2021; editorial decision 14 August 2021; published online 18 August 2021.
a
C. L., Y. C. and Y. Z. are joint first authors. by the apprehension of rare but potentially severe side effects
b
A. J. z. and K.-Y. Y. are joint last authors. of these rapidly developed novel vaccines. An example of such
Correspondence: K.-Y. Yuen, State Key Laboratory of Emerging Infectious Diseases, Carol
Yu Centre for Infection, Department of Microbiology, Li Ka Shing Faculty of Medicine, The a side effect is myopericarditis following mRNA COVID-19
University of Hong Kong, Pokfulam, Hong Kong Special Administrative Region, China; and vaccines, which has a crude incidence of 40.6 cases per million
Department of Clinical Microbiology and Infection Control, The University of Hong Kong-
Shenzhen Hospital, Shenzhen, China ([email protected]). second doses administered to males aged 12−29 years [4]. The
Clinical Infectious Diseases®  2021;XX(XX):0–0 pathogenesis of this unexpected complication remains elusive.
© The Author(s) 2021. Published by Oxford University Press for the Infectious Diseases The World Health Organization (WHO) [5] and Centers
Society of America. This is an Open Access article distributed under the terms of the Creative
Commons Attribution-NonCommercial-NoDerivs licence (http://creativecommons.org/licenses/ for Disease Control and Prevention (CDC) [6] no longer rec-
by-nc-nd/4.0/), which permits non-commercial reproduction and distribution of the work, in any ommend aspiration of syringe plunger during intramuscular
medium, provided the original work is not altered or transformed in any way, and that the work
is properly cited. For commercial re-use, please contact [email protected]
injections, especially during vaccination when a rapid injec-
https://doi.org/10.1093/cid/ciab707 tion of a small volume may reduce discomfort [6]. However,

Intravenous COVID-19 mRNA Vaccine and Myopericarditis  •  cid 2021:XX (XX XXXX) • 1

a self-reporting study of registered nurses showed that 40% re- anti-mouse CD45, CD68, or CD3 (Abcam) were used in this
ported blood aspiration at least once, and 4% reported blood study. DNA fragmentation in cardiomyocytes was labeled using
aspiration 13 times or more during intramuscular injection. Click-iT® Plus TUNEL assay kit (Thermo Fisher Scientific) for
The finding suggests that inadvertent intravenous injection of the detection of apoptosis [10]. The slides were mounted and
vaccine is possible [7]. Recently, inadvertent intravenous injec- examined under light microscope. Representative images were
tion of adenovirus-vector based COVID-19 vaccine was im- captured with Olympus BX53 semi-motorized fluorescence mi-
plicated to trigger platelet-adenovirus aggregates taken up by croscope. The measurement of cytokine and chemokine mRNA
spleen, which mounted B-cell response of binding antibodies expression levels in different tissues, serum cytokine/chemo-
against platelets [8]. In this study, we investigated the differ- kine levels, serum troponin levels, and the statistical analysis
ences in the cardiac pathology induced by intravenous (IV) can be found in the Supplementary method.

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or intramuscular (IM) BNT162b2 mRNA COVID-19 vaccine
when compared with normal saline (NS) injection in a Balb/c RESULTS
mouse model.
Intravenous SARS-CoV-2 mRNA Vaccine Administration Induced Grossly
Visible Pathology in Heart
METHODS Groups of female Balb/c mice at age of 6–8 week were given
BNT162b2 COVID-19 mRNA vaccine either IV or IM, or the
Animal Model
same volume of NS (Figure 1A). None of the animals showed
Female Balb/c (substrain OlaHsd) mice at age of 6–8 weeks were
clinical signs of lethargy, ruffled furs, hunched back posture,
obtained from the Centre for Comparative Medicine Research
and rapid breathing throughout the course of observation.
of The University of Hong Kong and kept in Biosafety Level
Significant decrease in body weight was observed in IM mRNA
(BSL)-2 animal laboratory with 12-hour light-dark cycle and
vaccine group (mean 3.6% ± 2.1%) starting from 1 dpi; animals
free access to water and diet. The animals were randomly as-
recovered their initial weight at 7 dpi (Figure 1B). Autopsy at
signed to 3 groups for the administration of IV or IM COVID-
1–2 dpi showed white patches over the visceral pericardium in
19 mRNA vaccine, or normal saline (NS) control (Figure 1A).
37.5% (1 dpi, n = 8) to 38.5% (2 dpi, n = 13) of the IV vaccine
COVID-19 mRNA vaccine (BNT162b2 lot number 1B004A,
group but none in the IM vaccine or NS control groups (Figure
BioNTech, Germany) dose of 0.25 µg per gram of body weight
1C and 1D; Table 1; P < .05). No grossly visible changes were ob-
was injected (about 5  µg in 50  µL per mouse; dose according
served in other organs of the animals (Supplementary Figure 1).
to an immunogenicity study) via tail vein or thigh muscle, re-
spectively, with the control group having the same volume of Histopathological Changes in Mouse Heart After IV mRNA Vaccine
NS [9]. Another group of male mice was subsequently tested Administration
by the same protocol after preliminary positive results. The Low power scanning of heart sections showed blue stained
mice were monitored by clinical signs and body weight changes thickened visceral pericardium over the right atrium and ven-
for 14 days. Necropsies were performed at 1, 2, 7, and 14 days tricle at 1 dpi of IV vaccine, which became more prominent
post-injection (dpi). Organs and blood were sampled for his- at 2 dpi (Figure 2A). At higher magnification, calcific deposits
tological and real-time quantitative reverse transcription pol- were seen in these thickened pericardial tissues (Figure 2D).
ymerase chain reaction (RT-PCR), or cytokine/chemokine Multifocal pericardial and myocardial inflammatory cell infil-
levels, respectively. A group of mice received a second boosting trates and interstitial oedema were also observed (Figure 2C).
dose 14 days after the first priming dose and examined at 2 dpi Frequent foci of cardiomyocytes had degenerative changes as
after boosting. The procedures for animal experiments in this evident by the loss of the normal pattern of cross-striation and
study were approved by the HKU Committee on the Use of Live occasionally sarcoplasmic vacuolation, and necrotic changes as
Animals in Teaching and Research. distinguished by the attainment of a homogenous appearance,
sarcoplasmic fragmentation, or pyknosis (Figure 2E). These
Histopathology and Immunohistochemical Staining of Tissues changes were significantly more frequent in the IV vaccine
Formalin-fixed and paraffin-embedded mouse heart, lung, liver, group at 2 dpi (Table 1) and more often in the right atrium and
spleen, kidney, and brain tissues were cut into 4-µm sections right ventricles of the affected animals and especially promi-
and stained with hematoxylin and eosin (H&E) for histopath- nent on their pericardial side. Immunohistochemical staining
ological examination. Immunohistochemistry staining for leu- with anti-CD45 (biomarker for immune cells of lymphoid or
kocyte biomarkers and SARS-CoV-2 spike receptor binding myeloid origin) indicated that these were leukocytes, of which
domain (S-RBD), terminal deoxynucleotidyl transferase dUTP many were macrophages or histiocytes positive for CD68.
nick end labeling (TUNEL) were performed as we described CD3-positive T cells were seen less often (Figure 2G).The num-
previously [10]. Primary antibodies including rabbit anti- bers of leukocytes were more than 14 per square millimeter in
SARS-CoV-2 S-RBD used in our previous study [11], and rabbit the affected myocardial foci. These findings suggested that

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Figure 1.  Schema for vaccine administration and gross pathology of mouse after vaccination. A, Experimental schema. Groups of mice were injected with COVID-19 mRNA
vaccine via intramuscular (IM) or intravenous (IV) route. At 1, 2, 7, and 14 dpi, mice were killed for histopathological analysis. Normal saline (NS) was IV or IM injected in
parallel as control. B, Body weight changes of mice after injection. C, Representative images of gross pathology of mouse organs and heart at 1 dpi. Hearts of NS control
and IM vaccine groups appeared normal, whereas whitish patches (arrows) were seen on the visceral pericardium of hearts after IV vaccine. D, Representative images of
gross pathology of mouse organs including heart at 2 dpi. Large whitish patches (arrows) were seen on the visceral pericardium of mice receiving IV vaccine. Abbreviations:
COVID-19, coronavirus disease 2019; dpi, days post-injection; mRNA, messenger RNA.

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Figure 1.  Continued.

mice given IV mRNA COVID-19 vaccine can develop acute showed occasional positive cardiomyocytes, infiltrating im-
myopericarditis. Similar histopathological changes and severity mune cells and vascular endothelial cells within the myocar-
were found in male mice (Supplementary Figures 2B, 2C). dium and pericardium in the IV group but not IM vaccine
or NS control groups (Figures 4A, 4B, 4C). Using RT-qPCR
IV mRNA Vaccine Administration Induced Apoptosis of Cardiomyocytes, [12], the amount of COVID-19 mRNA Spike-RBD gene
Tissue mRNA, and Protein Expression of SARS-CoV-2 Spike
copies in heart tissues was significantly higher in IV than IM
To determine if apoptosis was induced in cardiomyocytes,
group at 1 dpi (Supplementary Figure 3). Although no statis-
TUNEL immunofluorescent staining was used for detection of
tically significant differences were found, the mean amount
DNA fragmentation in apoptotic cells (Figure 3A). Apoptotic
of Spike-RBD mRNA was higher in the IV group than the IM
cardiomyocytes were significantly more often found in the IV
group at all other time points.
group than the IM or NS groups. Apoptotic cardiomyocytes dis-
tributed both sporadically or as large foci were found in 75% of IM mRNA Vaccine Administration Only Induced Mild Myocardial
IV group at 1 dpi (6/8) and 38.5% at 2 dpi (5/13) (P < .05; Table Congestion and Edema
1, Figure 3B). No grossly visible change of the heart in the IM group was
To understand whether mRNA vaccine can transfect seen on autopsy or low power scan (Figure 5A). H&E sections
cardiomyocytes to express SARS-CoV-2 spike protein, we showed some myocardial vascular congestion and mild in-
used immunostaining to detect SARS-CoV-2 Spike-RBD and terstitial edema at 1 dpi (Figure 5B). Degenerative changes of

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Table 1.  Summary of the Pathological Changes in Heart and Liver After IV or IM Administration of mRNA Vaccine

Pathology a mRNA Vaccine Injection (Second Dose Given 14 days After First Dose)

2 dpi First IV 2 dpi First IM NS 2 2 dpi Second IV 2 dpi Second IM

Dose Dose dpi Dose Dose
Changes (Related Figures) n = 13 (%) n = 6 (%) n = 12 (%) n = 9 (%) n = 6 (%)

Heart Grossly visible white patches on visceral pericardium 5/13 (38.5%)† 0/6 0/12 3/9 (33.3%) 1/6 (16.7%)
(Fig 1C, 1D)
Pericardial calcific deposit 5/13 (38.5%)† 0/6 0/12 4/9 (44.4%)† 2/6 (33.3%)
(Fig. 2A, 2D)
Pericardial WBC infiltration 9/13 (69.2%)* ,††† 0/6 0/12 5/9 (55.6%)†† 2/6 (33.3%)
(Fig. 2C)

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Myocardial WBC infiltration 8/13 (61.5%)* ,††† 0/6 0/12 9/9 (100%)††† 6/6 (100%)†††
(Fig. 2C, 2G)
Cardiomyocytes degeneration 8/13 (61.5%)†† 2/6 (33.3%) 0/12 9/9 (100%)††† 6/6 (100%)†††
Fig. 2E)
Cardiomyocytes necrosis (Fig. 2F) 4/13 (30.8%) 0/6 0/12 9/9 (100%)††† 6/6 (100%)†††
Cardiomyocytes apoptosis 5/13 (38.5%)† 0/6 0/12 ND ND
(Fig. 3B)
Spike RBD expression b 3/8 (37.5%)† 0/6 0/12 ND ND
(Fig. 4A, 4B)
Serum troponin (pg/mL)c 1328.2 ± 325.8**** ,†††† 237.5 ± 121.2 215 ± 115.9 ND ND
Liver Grossly visible change at autopsy 0/6 0/6 0/6 0/9 0/6
Hepatocytes ballooning degeneration 6/6 (100%)†† 3/6 (50%) 0/6 ND ND
(Fig. 8A)
Hepatocytes necrosis 2/6 (50%) 0/6 0/6 ND ND
Abbreviations: dpi, days post-injection; IM, intramuscular; IV, intravenous; ND, not done; NS, normal saline; RBD, receptor binding domain; WBC, white blood cells.
a
Presented as the number and percentage of mice with positive gross or histological changes in the heart and liver.
b
Number heart sections showed RBD positive cells at 1 dpi.
c
Serum troponin concentration in mouse serum were determined by Enzyme Immunoassay using Mouse CTNI (Cardiac troponin-I) ELISA Kit (Mybiosource, San Diego, California, USA).
* P < .05,**** P < .0001 when compared to IM group at the same time point.
† P < .05, ††P < .01, †††P < .001, ††††P < .0001 when compared to NS 2 dpi group.

cardiomyocytes were occasionally found at 2 dpi (Figure 5B). higher serum concentrations of cytokines/chemokines at 1
No obvious immune cell infiltration, cardiomyocyte necrosis, dpi, which decreased at 2 dpi (Figure 6C). Increased serum
or TUNEL-positive apoptotic cells were found at 1 or 2 dpi cytokine/chemokine concentrations were found in the IV
(Figure 5C). The changes were not sufficient to satisfy the his- group at 1 dpi, but only CXCL10 and CCL5 were significantly
tological criteria of myocarditis. Notably, SARS-CoV-2 spike higher than the NS control group (Figure 6C). Enzyme im-
protein expression was only found in the infiltrating immune munoassay showed that serum troponin levels of the IV group
cells in the thigh muscle 1 dpi in the IM group. No degenera- (1328.2  ± 325.8 pg/mL) was significantly higher than the IM
tion, necrosis, or SARS-CoV-2 protein expression was evident group (237.5  ± 121.2 pg/mL) and NS group (215  ± 115.9 pg/
in the skeletal myocytes (Figures 5D, 5E). mL) (P < .0001; Table 1).

mRNA Vaccine Administration Induced Inflammatory Cytokine/Chemokine Histopathological Changes of the Heart at 7 dpi and 14 dpi After First Dose,
Response in the Heart, and Increased Serum Troponin and Cytokine/ and at 2 Days After the Second Dose Given at 14 Days After First Dose
Chemokine levels At 7 dpi, the heart of mice in the IV group showed persistent
RT-qPCR assay showed increased mRNA expression of in- changes of myopericarditis (Figure 7A), whereas the IM group
terferon (IFN)-α/β, interleukin (IL)-6, tumor necrosis factor only showed vascular congestion, myocardial edema, and oc-
(TNF)-α, CXCL10, and CCL3 in heart tissue homogenates casional foci of cardiomyocyte degeneration (Figure 7A). At 14
of the IV group, among which IFN-β, IL-6, and TNF-α were dpi, 4/6 (66.7%) of the mice in the IV group showed grossly vis-
significantly higher at 2 dpi than 1 dpi (Figure 6A). As for the ible white patches over the visceral pericardium, and 6/6 (100%)
IM group, all tested inflammatory cytokines/chemokines were showed changes of myopericarditis, compared with only mild
transiently upregulated in heart tissues at 1 dpi and then de- degenerative changes in the IM group (Figure 7B).
creased at 2 dpi (Figure 6B). But IL-1 β expression was signifi- Two days after the second dose of mRNA vaccine, 3/9 (33.3%)
cantly higher in cardiac tissues of male than female mice at 2 dpi and 1/6 (16.7%) of mice in the IM/IV and IV/IM groups devel-
(Supplementary Figure 2A). oped grossly visible white patches over the visceral pericardium,
Beads-based multiplex cytokine/chemokine flow respectively. Both groups showed more diffuse and severe changes
cytometry assay showed that the IM group had significantly of myopericarditis with foci of mild myocardial hemorrhage,

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Figure 2.  Representative histopathological images of heart tissues of mice receiving IV vaccine. Groups of mice were given IV injection of COVID-19 mRNA vaccine (IV
group) and NS (NS group) as control. At 1 and 2 dpi, mice were killed for histopathology analysis. A, Low magnification microscopic scanning images of heart sections (4×
magnification). Mice received IV NS showed no detectable histological changes in the heart. After IV mRNA vaccine, the scan images showed thickened and dark blue stained
visceral pericardium on the surface of right atrium and right ventricle at both 1 and 2 dpi (arrows). B, H&E stain of heart tissue of IV NS group showed no histological damage
in the myocardium and cardiomyocytes. C, H&E stained heart tissue showed inflammatory infiltrates of the myocardium at 2 dpi. Arrows indicated infiltrates of inflammatory
cells at 400× magnification. D, H&E stained heart tissue of IV vaccine group at 2 dpi showing thickened visceral pericardium with clusters of dark blue crystal-like structure
which indicated calcific deposits (arrows) with adjacent inflammatory cell infiltrates and cardiomyocytes degeneration at. 400× magnification. E, H&E stained myocardial
tissue showing cardiomyocytes degeneration at 2 dpi in IV group as indicated by arrows at 400× magnification. F, H&E image showing cardiomyocytes necrosis (arrows at
400× magnification) with immune cells infiltration in IV group at 2 dpi. G, Images of immunohistochemistry staining of white blood cells marker CD45, CD68, and CD3 in the
heart sections, showing myocardial and visceral pericardial infiltration by CD45 positive cells. Immunostaining of macrophage marker CD68 showed many positives in the
infiltrating cells, with less frequently positive CD3 biomarker for T lymphocytes. Abbreviations: COVID-19, coronavirus disease 2019; dpi, days post-injection; H&E, hematox-
ylin and eosin; IV, intravenous; mRNA, messenger RNA; NS, normal saline.

which affected both the right and left heart (Figure 7D; Table 1). and CXCL10 at 2 dpi after IV boosting, which suggested that IV
Moreover, serum cytokine/chemokine levels by beads-based flow vaccine increased inflammatory responses, although CCL2 was in-
cytometry assay showed significantly increased IFN-γ, TNF-α, creased after both IV and IM boosting (Figure 7E).

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Figure 2.  Continued.

IV mRNA Vaccine Administration Induced Histopathological Changes subclinical and the changes persisted but did not progress within
in Liver
14 dpi. But these histopathological changes of myopericarditis
H&E-stained sections of the liver tissues of the IV group deteriorated and became rather diffuse after the second dose
showed diffuse ballooning degeneration of hepatocytes and boosting with either IV or IM administration 14 days after the
focal hepatocyte necrosis at 1–2 dpi (Figure 8A), whereas the first dose of priming. Although Balb/c mice have been exten-
liver tissues of the IM group showed much milder changes sively used for modeling myocarditis due to viral, protozoal, and
without hepatocyte necrosis at 1–2 dpi (Supplementary Figure autoimmune insults [15], myocardial mineralization can occur
1B). SARS-CoV-2 spike RBD expression by immunostaining spontaneously with age in inbred laboratory rodents. However,
was occasionally seen in hepatocytes of the IV group but not we have excluded this possibility by the demonstration of highly
the IM group (Figure 8B). Except for some vascular congestion significant differences in the gross pathological and histolog-
in lungs, H&E stained sections of spleen, brain, and kidney at ical changes of myopericarditis between mice challenged by IV
1–2 dpi were unremarkable (Supplementary Figure 1C). versus IM vaccine or NS control (Table 1) [16].
Acute myocardial injury due to hypersensitivity myocarditis
DISCUSSION was reported after smallpox vaccine at a rate of 12.3–463 cases
In a Balb/c mouse model with both male and female mice, IV per 100 000 [17] and very rarely associated with other vaccines
but not IM administration of COVID-19 mRNA vaccine in- for yellow fever and influenza [17–19]. COVID-19 mRNA vac-
duced a rapid onset of multifocal myopericarditis with elevated cines were associated with myopericarditis at a rate of 12.6–24
serum troponin, cardiomyocyte degeneration, and changes of cases per million following a second dose [20]. The clinical
both necrosis and apoptosis, adjacent inflammatory infiltrate of manifestations of acute chest pain, dyspnea, arrhythmia, raised
mononuclear cells, interstitial edema, and visceral pericardial serum troponin, and electrocardiographic and gadolinium en-
calcification within 2 dpi. Moreover, the IL-1β, IFN-β, IL-6 and hanced cardiac magnetic resonance imaging (MRI) changes
TNF-α expression levels generally increased significantly from often started 3–5 days after the second dose of vaccine or oc-
1 dpi to 2 dpi in the IV group but not the IM group. Overall, the casionally after first dose [21, 22]. Two patients without meas-
findings have satisfied the Dallas and immunohistochemical urable SARS-CoV-2 spike immunoglobulin G (IgG) presented
criteria of myocarditis [13]. Similar to findings of cardiac mag- shortly after their first vaccine dose, suggesting that myocardial
netic resonance imaging in human myocarditis, the most prom- damage can happen with just 1 dose of mRNA vaccine as dem-
inent site of focal involvement was the pericardial side of atrial onstrated in our present study [23]. Moreover, the second dose
and ventricular walls [14]. Notably, the myopericarditis was of mRNA vaccine given 14  days after the first dose by either

Intravenous COVID-19 mRNA Vaccine and Myopericarditis  •  cid 2021:XX (XX XXXX) • 7

IM or IV route has markedly exacerbated the myopericarditis, damage to the endoplasmic reticulum is severe or persistent, the
which is also compatible with the clinical findings in human unfolded protein response triggers apoptosis [25]. The same in
subjects. vitro phenomenon was reported with SARS-CoV-2 [26]. More
The pathogenesis of an early onset of myopericarditis in IV studies on pathogenesis are warranted.
vaccinated mice is unclear. Intranasal administration of ioniz- Another possible causative mechanism of the mRNA vaccine
able lipid nanoparticles alone or complexed with mRNA can induced myopericarditis could be the overly activation of cy-
induce massive lung inflammation and death within 24 hours tokine production, which was also reported to cause reversible
[24]. Here we showed that SARS-CoV-2 spike protein was oc- myopericarditis and cardiomyopathy in patients treated with in-
casionally expressed in cardiomyocytes 1 dpi of IV mRNA vac- terferon for chronic myeloid leukemia, viral hepatitis, and mul-
cine, although such expression was more often in the infiltrating tiple sclerosis [27, 28]. Intravenous injection of inactivated typhoid

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immune cells in the myocardium and visceral pericardium. We vaccine was associated with progressive radiological cardiomegaly
have previously shown that the replication of SARS-CoV-1 within 2 weeks [29]. However, a sufficient degree of myocardial
leads to substantial accumulation of heavily modified trans- damage satisfying the histopathological criteria of myopericarditis
membrane viral proteins such as unfolded spike at the endo- was not observed in our IM vaccine group despite significantly
plasmic reticulum, which rapidly exceed its folding capacity higher serum proinflammatory cytokine/chemokine levels and
leading to stress and the unfolded protein response. When the body weight loss. Cardiac damage due to hypersensitivity toward

Figure 3.  Representative images of TUNEL (apoptosis biomarker) staining of heart tissues. Groups of mice received COVID-19 mRNA vaccine via intramuscular (IM) or in-
travenous (IV) route. At 1–2 dpi, the mice were killed for histopathological analysis. Control group of mice received IV NS. A, No TUNEL staining signal in heart tissue section
of NS control mice at 2 dpi (left) at 200× magnification. DNase treatment of the same tissue as positive control showed TUNEL staining in the nucleus of cardiomyocytes
(green fluorescent signal as indicated by arrows in the insert at 400× magnification). B, At 1–2 dpi of IV mRNA vaccine, TUNEL signals were shown in a large area of the
myocardial tissue (circled by dashed lines at 40× magnification, indicated by arrows in the inserts at 400× magnification). H&E staining of the same heart tissue sections are
shown on the right. Abbreviations: COVID-19, coronavirus disease 2019; dpi, days post-injection; H&E, hematoxylin and eosin; mRNA, messenger RNA; NS, normal saline;
TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.

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Figure 4.  Immunohistochemical staining for protein expression of spike receptor binding domain (RBD) in heart sections. A, Representative images of immunohistochemical
staining of heart section showed occasional spike RBD positive cardiomyocytes (arrows at 400× magnification) at 1 dpi in IV vaccine group. H&E stain of the same area of
the section is on the right. B, Representative images showing spike RBD expression in the histiocytes, and vascular endothelial cells in myocardium (arrows in 400× mag-
nification). C, Representative images of positive control for the staining of spike RBD in SARS-CoV-2 infected mouse bronchiolar epithelium (arrows in left panel), negative
expression of spike RBD by immunostaining in the myocardium of IV NS control group (middle), and immunostaining with only biotin conjugated secondary antibody in heart
sections of IV vaccine group at 1 dpi (right). Abbreviations: dpi, days post-injection; H&E, hematoxylin and eosin; IV, intravenous; mRNA, messenger RNA; NS, normal saline;
SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

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Figure 5.  Representative histopathological images of heart tissues after intramuscular administration of mRNA vaccine. Groups of mice were given IM vaccine NS (NS
group) as control. At 1–2 dpi, mice were killed for histopathology. A, Low magnification scanning images of heart sections (4× magnification). Both NS group and IM vaccine
group showed no histological changes in the heart. B, Representative H&E images of heart tissues. NS groups showed normal myocardium and cardiomyocytes. IM vaccine
group showed vascular congestion and mild degree of myocardial edema at 1 dpi. No white blood cell infiltration, cardiomyocyte degeneration, or necrosis was observed.
At 2 dpi, vascular congestion was reduced but interstitial edema could still be seen. C, TUNEL staining of heart section showed no positive signal at 2 dpi for IM group. D,
H&E images of thigh muscle showed white blood cell infiltration in the connective tissue while the adjacent skeletal muscle cells are unremarkable (magnified image in the
right) 1 dpi after IM mRNA vaccine. E, Immunohistochemistry staining of Spike RBD showed only some infiltrating white blood cells expressing spike RBD in thigh muscle of
IM group (arrows in magnified images). Abbreviations: dpi, days post-injection; H&E, hematoxylin and eosin; IM, intramuscular; IV, intravenous; mRNA, messenger RNA; NS,
normal saline; RBD, receptor binding domain; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling.

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Figure 5.  Continued.

other components of mRNA vaccine is unlikely as similar degree and arteries within the reach of a syringe needle in the deltoid
of damage should happen in both IV and IM groups. region are considered too small to allow a rapid IV injection of
Both Pfizer/BioNTech and Moderna have clearly stated vaccine without blowing out the vessel [6]. However, this spec-
that their vaccines should only be given via IM route [30, 31]. ulation also lacks supportive scientific evidence. Another pos-
However, current CDC [6] and WHO guidelines [5] no longer sibility of getting a high blood mRNA vaccine level is the rapid
recommend precautionary measures during IM vaccine admin- movement of the vaccine through the lymphatic system into
istration. Brief aspiration for blood return during intramuscular the venous circulation. Thus changing the vaccine injection site
injection of medication as a preventive measure against acci- from deltoid to the vastus lateralis muscle of lateral mid-thigh
dental IV injection was previously present in most guidelines may reduce the amount of vaccine lipid nanoparticles reaching
[32]. This practice becomes controversial as scientific evidence the venous circulation due to enhanced uptake by the dendritic
of the perceived benefit of this procedure is lacking for IM in- cells and macrophages at the regional inguinal, iliac and para-
jection of vaccine. The CDC Pink Book 2020 [6] and WHO aortic lymph nodes.
2015 position paper [5] have recommended against aspiration We note that Pfizer has conducted in vivo biodistribution
prior to vaccine injection so as to minimize pain [33]. The veins studies of IM-injected [3H]-labeled lipid-nanoparticle (LNP)

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Figure 6.  Cytokine/chemokine mRNA expression in heart and their serum levels. At 1–2 dpi of IV or IM mRNA vaccine, organs and serum were collected. A, Cytokines/
chemokines mRNA expression in heart homogenates of IV group. B, Cytokines/chemokines mRNA expression in heart homogenates of IM group; mRNA expressions were
detected by RT-qPCR with gene specific primers. House-keeping gene β-actin mRNA expression was included to normalize the amount of RNA. Data presented were relative
gene expression to NS control mice. Error bars induced mean ± standard deviation. n = 5 in each group. C, Serum cytokine/chemokine concentrations at 1–2 dpi were detected
by beads-based multiplex flow cytometer assay. Mice serum from IV and IM injection of 50 µL NS were used as control. Error bars indicated mean ± standard deviation. n = 5
each group. *P < .05, **P < .01, ***P < .001,****P < .0001. Abbreviations: dpi, days post-injection; IFN, interferon; IL, interleukin; IM, intramuscular; IV, intravenous; mRNA,
messenger RNA; NS, normal saline; RT-qPCR, reverse transcription quantitative polymerase chain reaction; TNF, tumor necrosis factor.

mRNA vaccines in rats [34]. There was some accumulation of stem cells to the myocardium by retrograde venous perfusion
the formulation in the heart at 2 dpi, although much lower than was reported to achieve a better myocardial concentration [35].
concentrations in the liver or spleen. No similar tracer study Moreover, smaller mRNA-vaccine lipid-nanoparticles(100nm
was reported when the mRNA vaccine is IV injected. Besides diameter) can be sucked into larger T tubules (diameter
the possibility of delivery to the heart through the systemic ar- >200 nm) of cardiomyocytes during diastole, but not into T tu-
terial system, the IV-injected SARS-CoV-2 mRNA could the- bules of skeletal myocyte (diameter 20–40 nm) [36]. Thus, the
oretically transfect myocardial cells through smaller cardiac T tubule system of cardiomyocytes may concentrate mRNA-
venous system (Thebesian network), which consists of a layer vaccine lipid-nanoparticles like a sponge.
of vascular endothelial cells continuous with endothelium of Interestingly, we also observed ballooning degeneration of
4 cardiac chambers without interference by valves. Delivery hepatocytes especially in our IV group, which is compatible
of pharmacologic therapy, gene therapy, growth factors, and with the heavy distribution of the mRNA vaccine formulation

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Figure 6.  Continued.

in the liver of IM injected rats in the bio-distribution study [34] younger age or male gender. Although our male mice had sim-
and microscopic vacuolation of portal hepatocytes in these rats ilar degree of myopericarditis to female mice, a lower vaccine
[37]. There have been anecdotal reports on autoimmune hepa- dose may show up differences in disease susceptibility due to im-
titis following COVID-19 mRNA vaccines in humans although munological differences or ACE2 expression driven by sex hor-
these reports are yet to be confirmed by population-level vac- mone and X chromosome. Although the histological changes
cine adverse effect monitoring systems [38, 39]. Further re- in the heart of IM group did not amount to myopericarditis, we
search into the potential association of COVID-19 mRNA cannot exclude the possibility of frank myopericarditis in indi-
vaccination and autoimmune hepatitis is required. viduals who may be more susceptible to even a slight amount
Limitations of our study included the lack of data in ex- of mRNA vaccine entering the systemic circulation from intra-
plaining the association of post-vaccination myocarditis with muscular injection. COVID-19 mRNA vaccines are safe and

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Figure 7.  Histopathological changes in the heart at 7 and 14 dpi after first dose of IV or IM mRNA vaccine and 2 dpi after second dose of vaccine. Groups of mice were
given IV and IM vaccine or NS as control. At 7–14 dpi, mice were killed for histopathology. Another 2 groups of mice were given second dose of IV or IM mRNA vaccine at
14 days after the first priming dose and sacrificed at 2 dpi after the second boosting dose. A, Representative histopathological images of mouse heart at day 7. Top panel
consisted of heart sections of IV group, which showed myocardial infiltration by white blood cells (left, arrows), interstitial edema, cardiomyocytes degeneration (middle,
arrows) and necrosis (right, arrows). Lower panel consisted of heart sections from IM group, which showed myocardial interstitial edema (left) and myocardial vascular con-
gestion (middle, arrows), with degeneration of a few cardiomyocytes (right, arrows). B, Representative histopathological images of IV and IM group at day 14. Heart in IV
group showed persistent changes of cardiomyocyte degeneration, white blood cell infiltration, and foci of necrosis (arrows). C, Heart of mice in IM group showed minimal
degeneration and infiltration but no necrosis. D, Representative histopathological images of the heart at 2 dpi after the second boosting dose given on day 14 after the first

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Figure 7.  Continued.

effective, whereas post-vaccination myopericarditis is rare and without aspiration, which carries the risk of inadvertent IV in-
self-limiting [4]. Our study indicates that IV injection of vac- jection. Increasing the size of mRNA-vaccine lipid-nanoparticle
cines might partially contribute to this clinical phenotype, thus or decreasing the vaccine dose in normal adolescents to reduce
warranting a reconsideration of the practice of IM injection risks of myopericarditis warrant further investigations. Careful

priming dose. Mouse heart in both IV and IM second dose group showed interstitial edema and diffuse cardiomyocyte degeneration on the left (arrows). Mouse heart in both
IM and IV group showed diffuse inflammatory infiltrate, focal hemorrhage and necrosis (arrows, right). E. Serum cytokine/chemokine concentrations at 2 dpi post second dose
were detected by beads-based multiplex flow cytometer assay. The NS group was used as control. Error bars indicated mean ± standard deviation. n = 5 each group. n = 9
for IV second dose boost group, n = 6 for IM second dose boost group and NS control group. *P < .05, **P < .01, ***P < .001 by multiple t test. Abbreviations: dpi, days post-
injection; IM, intramuscular; IV, intravenous; mRNA, messenger RNA; NS, normal saline; RT-qPCR, reverse transcription quantitative polymerase chain reaction.

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Figure 8.  Histopathological changes in the liver at 2 dpi after IV mRNA vaccine injection. A, Representative H&E images of liver sections. Images in the upper panel show
the liver section of NS control mice, with normal morphology of evenly distributed cords of hepatocytes. Lower panel consisted of images of liver at 2 dpi in IV group. The
liver showed diffuse degeneration of hepatocytes without a clear morphological organization of hepatic cords. A large focus of cell necrosis was circled by the dashed line.
Magnified images (400× magnification) of hepatocyte necrosis (middle panel, arrows in magnified box of left bottom panel) and ballooning degenerative changes of hepato-
cytes (right panel, arrows in magnified box of right bottom panel). B, Images of immunohistochemistry stained spike RBD in liver sections at 2 dpi after IV vaccine. A few
RBD positive cells were indicated by arrows in the inserts (400× magnification). Abbreviations: dpi, days post-injection; H&E, hematoxylin and eosin; IV, intravenous; mRNA,
messenger RNA; NS, normal saline; RBD, receptor binding domain.

16 • cid 2021:XX (XX XXXX) • Li et al

histopathological examination of the heart is required in any 8. Nicolai  L, Leunig  A, Pekayvaz  K, et  al. Thrombocytopenia and splenic platelet
directed immune responses after intravenous ChAdOx1 nCov-19 administration.
case of fatality following COVID-19 mRNA vaccines as myo- bioRxiv 2021; doi: 10.1101/2021.06.29.450356
carditis can be focal or masquerade as ischemic heart disease 9. Agency MHpR. Public assessment report authorisation for temporary supply
COVID-19 mRNA vaccine BNT162b2(BNT162b2 RNA) concentrate for solu-
in older patients. tion for injection, page 15. Available at: https://assets.publishing.service.gov.uk/
government/uploads/system/uploads/attachment_data/file/997584/COVID-
Supplementary Data 19_mRNA_Vaccine_BNT162b2__UKPAR___PFIZER_BIONTECH_ext_of_in-
Supplementary materials are available at Clinical Infectious Diseases online. dication_11.6.2021.pdf. Accessed 23 July 2021.
10. Lee  ACY, Zhang  AJX, Chu  H, et  al. H7N9 influenza A  virus activation of
Consisting of data provided by the authors to benefit the reader, the posted
necroptosis in human monocytes links innate and adaptive immune responses.
materials are not copyedited and are the sole responsibility of the authors, so
Cell Death Dis 2019; 10:442.
questions or comments should be addressed to the corresponding author. 11. Chen LL, Lu L, Choi CY, et al. Impact of SARS-CoV-2 variant-associated RBD
mutations 1 on the susceptibility to serum antibodies elicited by COVID-19 in-
Notes fection or vaccination. Clin Infect Dis 2021:ciab656. doi:10.1093/cid/ciab656.

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Author Contributions. A. J. Z. and K.-Y. Y. had roles in the study design, 12. Yip CCY, Sridhar S, Leung KH, et al. Development and evaluation of novel and
data collection, data analysis, data interpretation, and writing of the manu- highly sensitive single-tube nested real-time RT-PCR assays for SARS-CoV-2 de-
tection. Int J Mol Sci 2020; 21:5674. doi:10.3390/ijms21165674.
script. C. L., Y. C., Y. Z., Z. Y., W. S., F.-F. L., J.-P. C., W.-M. W., and C. C.-Y.
13. Aretz HT, Billingham ME, Edwards WD, et al. Myocarditis: a histopathologic def-
Y. had roles in the experiments, data collection, and/or data analysis. D. C.
inition and classification. Am J Cardiovasc Pathol 1987; 1:3–14.
L., I. F.-N. H., J. F.-W. C., K. K.-W. T., S. S., H. C., K.-H. K., and D. J. had roles 14. Mahrholdt H, Goedecke C, Wagner A, et al. Cardiovascular magnetic resonance
in experimental design, data analysis, and/or revision of the manuscript. All assessment of human myocarditis: a comparison to histology and molecular pa-
authors reviewed and approved the final version of the manuscript. thology. Circulation 2004; 109:1250–8.
Disclaimer. The funding sources had no role in the study design, data 15. Błyszczuk P. Myocarditis in humans and in experimental animal models. Front
collection, analysis, interpretation, or writing of the report. Cardiovasc Med 2019; 6:64.
Financial support. This work was supported by funding from the 16. Herman E, Eldridge S. Spontaneously occurring cardiovascular lesions in com-
Food and Health Bureau, The Government of the Hong Kong Special monly used laboratory animals. Cardiooncology 2019; 5:6.
17. Kuntz J, Crane B, Weinmann S, Naleway AL; Vaccine Safety Datalink Investigator
Administrative Region, and the Consultancy Service for Enhancing
Team. Myocarditis and pericarditis are rare following live viral vaccinations in
Laboratory Surveillance of Emerging Infectious Diseases and Research
adults. Vaccine 2018; 36:1524–7.
Capability on Antimicrobial Resistance for Department of Health of 18. Engler RJ, Nelson MR, Collins LC Jr, et al. A prospective study of the incidence
the Hong Kong Special Administrative Region Government; and do- of myocarditis/pericarditis and new onset cardiac symptoms following smallpox
nations of Richard Yu and Carol Yu, Shaw Foundation Hong Kong, and influenza vaccination. PLoS One 2015; 10:e0118283.
Michael Seak-Kan Tong, May Tam Mak Mei Yin, Lee Wan Keung Charity 19. Mei  R, Raschi  E, Poluzzi  E, Diemberger  I, De  Ponti  F. Recurrence of pericar-
Foundation Limited, Hui Ming, Hui Hoy, and Chow Sin Lan Charity ditis after influenza vaccination: a case report and review of the literature. BMC
Fund Limited, Chan Yin Chuen Memorial Charitable Foundation, Pharmacol Toxicol 2018; 19:20.
Marina Man-Wai Lee, the Hong Kong Hainan Commercial Association 20. Shay  DK, Shimabukuro  TT, DeStefano  F. Myocarditis occurring after immuni-
zation with mRNA-Based COVID-19 vaccines. JAMA Cardiol. Published online
South China Microbiology Research Fund, the Jessie & George Ho
June 29, 2021. doi:10.1001/jamacardio.2021.2821.
Charitable Foundation, Kai Chong Tong, Tse Kam Ming Laurence, Foo Oi
21. Kim HW, Jenista ER, Wendell DC, et al. Patients with acute myocarditis following
Foundation Limited, Betty Hing-Chu Lee, and Ping Cham So; Innovation mRNA COVID-19 vaccination. JAMA Cardiol 2021. June 29. doi:10.1001/
and Technology Fund, Innovation and Technology Commission, the jamacardio.2021.2828.
Government of the Hong Kong Special Administration Region. 22. Montgomery J, Ryan M, Engler R, et al. Myocarditis following immunization with
Potential conflicts of interests. J.  F.-W. C.  has received travel grants mRNA COVID-19 vaccines in members of the US military. JAMA Cardiol 2021.
from Pfizer Corporation Hong Kong and Astellas Pharma Hong Kong June 29. doi:10.1001/jamacardio.2021.2833.
Corporation Limited, and was an invited speaker for Gilead Sciences 23. Rosner  CM, Genovese  L, Tehrani  BN, et  al. Myocarditis temporally associated
Hong Kong Limited and Luminex Corporation. K. Y. Y. is the inventor an with COVID-19 vaccination. Circulation 2021; 144:502–5.
24. Ndeupen  S, Qin  Z, Jacobsen  S. The mRNA-LNP platform’s lipid nanoparticle
intranasal influenza vectored SARS-CoV-2. All other authors report no po-
component used in preclinical vaccine studies is highly inflammatory. bioRxiv
tential conflicts.
2021 Jul 23;2021.03.04.430128. doi:10.1101/2021.03.04.430128.
All authors have submitted the ICMJE Form for Disclosure of Potential 25. Chan CP, Siu KL, Chin KT, Yuen KY, Zheng B, Jin DY. Modulation of the unfolded
Conflicts of Interest. Conflicts that the editors consider relevant to the con- protein response by the severe acute respiratory syndrome coronavirus spike pro-
tent of the manuscript have been disclosed. tein. J Virol 2006; 80:9279–87.
26. Balakrishnan B, Lai K. Modulation of SARS-CoV-2 spike-induced unfolded pro-
tein response (UPR) in HEK293T cells by selected small chemical molecules.
References bioRxiv 2021: doi:10.1101/2021.02.04.429769.
1. To KK, Sridhar S, Chiu KH, et al. Lessons learned 1 year after SARS-CoV-2 emer- 27. Sonnenblick M, Rosin A. Cardiotoxicity of interferon: A review of 44 cases. Chest
gence leading to COVID-19 pandemic. Emerg Microbes Infect 2021; 10:507–35. 1991; 99:557–61.
2. World Health Organization. WHO Coronavirus (COVID-19) Dashboard. 28. Khakoo  AY, Halushka  MK, Rame  JE, Rodriguez  ER, Kasper  EK, Judge  DP.
Available at: https://covid19.who.int/. Accessed 17 July 2021. Reversible cardiomyopathy caused by administration of interferon alpha. Nat
3. Our World in Data. Statistics and research: coronavirus (COVID-19) vaccinations: Clin Pract Cardiovasc Med 2005; 2:53–7.
Share of people who received at least one dose of COVID-19 vaccine. Available at: 29. WEENS HS, HEYMAN A. Cardiac enlargement in fever therapy induced by in-
https://ourworldindata.org/covid-vaccinations. Accessed 18 July 2021. travenous injection of typhoid vaccine. Arch Intern Med 1946; 77:307–16.
4. Gargano  JW, Wallace  M, Hadler  SC, et  al. Use of mRNA COVID-19 vaccine 30. U.S. Food and Drug Administration (FDA). Emergency use authorization
after reports of myocarditis among vaccine recipients: update from the advisory (EUA) of the Pfizer-Biontech COVID-19 vaccine to prevent coronavirus di-
committee on immunization practices—United States, June 2021. MMWR Morb sease 2019 (COVID-19): fact sheet for healthcare providers administering vac-
Mortal Wkly Rep 2021; 70:977–82. cine (vaccination providers). 15 June 2021:14. Available at: https://www.fda.gov/
5. Organization WH. Reducing pain at the time of vaccination: WHO position emergency-preparedness-and-response/coronavirus-disease-2019-covid-19/
paper – September 2015. Wkly Epidemiol Rec 2015; 39:505–16. comirnaty-and-pfizer-biontech-covid-19-vaccine.
6. Centers for Disease Control and Prevention. Epidemiology and prevention of 31. U.S. Food and Drug Administration (FDA). Emergency use authorization
vaccine-preventable diseases: The Pink Book, chapter on vaccine administration. (EUA) of the Moderna COVID-19 vaccine to prevent coronavirus disease 2019
Available at: https://www.cdc.gov/vaccines/pubs/pinkbook/vac-admin.html. (COVID-19): fact sheet for healthcare providers administering vaccine. 24 June
Accessed 17 July 2021. 2021:9.
7. Thomas CM, Mraz M, Rajcan L. Blood aspiration during IM injection. Clin Nurs 32. Sepah Y, Samad L, Altaf A, Halim MS, Rajagopalan N, Javed Khan A. Aspiration
Res 2016; 25:549–59. in injections: should we continue or abandon the practice? F1000Res 2014; 3:157.

Intravenous COVID-19 mRNA Vaccine and Myopericarditis  •  cid 2021:XX (XX XXXX) • 17

33. Ipp M, Taddio A, Sam J, Gladbach M, Parkin PC. Vaccine-related pain: ran- 37. Agency MHpR. Public assessment report authorisation for temporary supply
domised controlled trial of two injection techniques. Arch Dis Child 2007; COVID-19 mRNA vaccine BNT162b2(BNT162b2 RNA) concentrate for solu-
92:1105–8. tion for injection. Page 19. Available at: https://assets.publishing.service.gov.uk/
34. SARS-CoV-2 mRNA Vaccine (BNT162, PF-0 7302048): 2.6.5.5B. government/uploads/system/uploads/attachment_data/file/997584/COVID-
Pharmacokinetics: organ distribution continued, report number: 185350, Page 6. 19_mRNA_Vaccine_BNT162b2__UKPAR___PFIZER_BIONTECH_ext_of_in-
Available at: https://www.pmda.go.jp/drugs/2021/P20210212001/672212000_303 dication_11.6.2021.pdf. Accessed 23 July 2021.
00AMX00231_I100_1.pdf. Accessed 23 July 2021. 38. Tan CK, Wong YJ, Wang LM, Ang TL, Kumar R. Autoimmune hepatitis following
35. Echeverri D, Cabrales J, Jimenez A. Myocardial venous drainage: from anatomy to COVID-19 Vaccination: true causality or mere association? J Hepatol 2021.
clinical use. J Invasive Cardiol 2013; 25:98–105. doi:10.1016/j.jhep.2021.06.009.
36. Brette F, Orchard C. T-tubule function in mammalian cardiac myocytes. Circ Res 39. Dumortier J. Liver injury after mRNA-based SARS-CoV-2 vaccination in a liver transplant
2003; 92:1182–92. recipient. Clin Res Hepatol Gastroenterol 2021: 101743. doi:10.1016/j.clinre.2021.101743.

Downloaded from https://academic.oup.com/cid/advance-article/doi/10.1093/cid/ciab707/6353927 by guest on 22 October 2021

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