COVID-19 and the Heart: A Systematic Review of Cardiac Autopsies

Importance: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-associated cardiac injury has been postulated secondary to several mechanisms. While tissue diagnosis is limited during the acute illness, postmortem studies can help boost our understanding and guide management. Objective: To report the cardiac tissue autopsy findings in coronavirus disease 2019 (COVID-19) decedents. Evidence Review: Articles published in PubMed and Embase reporting postmortem cardiac pathology of COVID-19 decedents till September 2020. We included adult studies excluding preprints. The Joanna Briggs Institute Critical Appraisal Checklist for Case Reports was used to assess quality. We extracted gross and histology data as well as the incidence of myocarditis, cardiac ischemia, thrombosis, and dilatation. We also looked at the reported cause of death (PROSPERO registration CRD42020190898). Findings: Forty-one relevant studies identified including 316 cases. The deceased were mostly male (62%) and elderly (median age, 75; range, 22–97 years). The most common comorbidities were hypertension (48%) and coronary artery disease (33%). Cardiac pathologies contributed to the death of 15 cases. Besides chronic cardiac pathologies, postmortem examination demonstrated cardiac dilatation (20%), acute ischemia (8%), intracardiac thrombi (2.5%), pericardial effusion (2.5%), and myocarditis (1.5%). SARS-CoV-2 was detected within the myocardium of 47% of studied hearts. Conclusions and Relevance: SARS-CoV-2 can invade the heart, but a minority of cases were found to have myocarditis. Cardiac dilatation, ischemia, mural, and microthrombi were the most frequent findings. The systematic review was limited by the small number of cases and the quality of the studies, and there is a need to standardize the cardiac postmortem protocols.


INTRODUCTION
While coronavirus disease 2019 (COVID-19) primarily affects the lungs, it is increasingly recognized as a multiorgan disease. The underlying mechanism may be direct viral invasion or secondary to the systematic effect of the infection (e.g., hypoperfusion, hypoxia, massive inflammatory response/cytokine storm).
Cardiac comorbidity and standard coronary risk factors (e.g., obesity, diabetes, and hypertension) are associated with adverse outcomes among patients with COVID-19 (1). COVID-19 is also associated with release of the highly specific marker of myocardial cell death-Troponin. Where this is tested in all hospitalized patients, the prevalence of elevated Troponin has been reported in up to 71% and is a predictor of outcome (40% mortality vs. 8% in those without myocardial injury) (2). A recent meta-analysis of published retrospective observational studies identified a positive troponin in 27% of 1,550 patients, with a similar impact on increased mortality and increased probability of needing intensive care (3).
Acute setting cardiac imaging (mainly echocardiography), while a valuable tool to assess the cardiac function and structure, suffers many limitations (4). Endomyocardial biopsies (EMBs) are rarely performed due to logistics and infection control reasons.
Postmortem examination (PM) is a valuable resource to understand the pathophysiology, cause of death, and the extent of organ involvement. Lessons from previous infectious diseases [e.g., human immunodeficiency virus (HIV)] have demonstrated the benefit of PMs (5).
To date, single case reports to modest-sized autopsy series have failed to clarify the nature of cardiac involvement. Histological findings vary from interstitial edema with or without myocarditis (6), lymphocytic endothelialitis (7), microvascular microthrombi and venous thrombosis (8), to extensive interstitial fibrosis with no endothelialitis (9), and no evidence of myocarditis (10). Optimal management depends on knowledge of the mechanism of myocardial injury, as the treatment and required follow-up will differ among the various pathologies outlined above.
To gain a better understanding of the prevalent cardiac findings in patients dying of COVID-19-we undertook a systematic review of all reported autopsies that included cardiac findings.

METHODOLOGY
A protocol of a systematic review was registered on PROSPERO database (CRD42020190898) on the 23rd June 2020. The aim was to investigate autopsy findings for patients who died from a confirmed COVID-19 infection (https://www.crd.york.ac.uk/ prospero/display_record.php?RecordID=190898).
An initial systematic search was conducted through the NHS Healthcare Databases Advanced Search tool (HDAS) on 7th of June 2020 for published articles in PubMed and Embase databases. The search strategy is shown in Table 1. An electronic search alert was set to identify any new study on the EMBASE database through Healthcare Databases Advanced Search (HDAS) (option not available for PubMed) till the 21st of September 2020. The search was done by AR and included the period from 1st January 2019 to the search date. AR screened the references for additional articles. We identified 88 articles that reported PM tissue pathology. AR reviewed the full-text to retrieve articles which reported PM cardiac pathology. We reviewed only published articles in journals (excluding preprints) in the English language and included humans since 2019 (Figure 1: PRISMA diagram). Articles or cases with duplicate reporting have been excluded to the best of our knowledge. AR assessed the quality of the case series studies using the Joanna Briggs Institute Critical Appraisal Checklist for Case Reports (12) (Supplementary Table 2). SZ and AR extracted the data from the included studies. Any conflict was resolved by discussion and mutual agreement.

Patient, Intervention, Comparison, and Outcome (PICO) Statement
Patient Adult patients (≥18 years old) who died and had a laboratory confirmation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Intervention
None.

Comparison
None or other patients who died from another cause.

Outcome
Pathological description of PM cardiac involvement.

Search Strategy
The search resulted in 226 titles. After duplicate removal of and title screening, we screened the full text of 108 articles (52 from weekly alerts) that yielded 88 articles reporting PM tissue pathology. Among those, 41 studies reported PM heart examination and included 336 cases (Figure 1). Studies were mostly case reports (n = 13) or case series (n = 24), while three studies compared cases to controls (6-10, 13-48) (Table 2, Figure 2). Authors reported cases from 14 countries, mostly developed westernized ones (Supplementary Table 1). Two studies reported on the same population, with one mainly focusing on PM cardiac examination (22,23). The quality of the included studies was mostly moderate (Supplementary Table 2).
We analyzed the PM cardiac histopathology for 316 cases [after excluding cases unconfirmed as COVID-19 (n = 6) or with no PM cardiac tissue examination (n = 14)].

Cause of Death
The cause of death was reported for 190 cases and, for the majority of these, was respiratory in origin (Supplementary Tables 1, 2). However, cardiac contribution to death was mentioned for 15 cases while pulmonary embolism (PE) was mentioned in eight cases.

DISCUSSION
Our review confirms that among patients dying from COVID-19, cardiac abnormalities are prevalent, but that specific changes of acute myocarditis are uncommon (1.5% of cases). Myocardial ischemia, thrombosis, and cardiac dilatation were the most dominant acute findings (Figure 2). Prevalence of the non-specific myocardial edema (ME) was 100% in the six studies reporting it (6,15,19,35,44,45). The highly prevalent chronic cardiac pathologies not only reflect the impact of cardiac comorbidities but also complicated the histopathological interpretation.   • All inflammatory cells positive for CD68; but none stained with CD3 • No myocarditis • No evidence of myocyte necrosis • Ischemic process of cardiac muscle highly suggested P5: • Severe interstitial infiltration of LCA-positive inflammatory cells with predominance of CD68 positive macrophages and focal aggregation of CD3 positive T cells • Histologic evidence of myocyte necrosis including hyper-eosinophilia and enucleation • Ischemic necrosis of myocardium should be considered P6: • No interstitial inflammation P7: • Majority of inflammatory cells showed immunoreactivity for CD68 and rare cells positive for CD3.

Role of Ischemia, Endotheliitis, and Hypercoagulability
The most alarming finding is the intracardiac, coronary arterial, and venous thrombosis, which may be in part explained by the COVID-19-associated coagulopathy (CAC). Myocardial ischemia can be further aggravated by the frequent pre-existing CAD and myocardial supply-demand mismatch. By means of its receptor, SARS-CoV-2 can directly invade the endothelium leading to endothelial cell (EC) inflammation (i.e., endothelialitis), dysfunction, and death (49). Endothelial dysfunction can also result from an inappropriate immune and cytokine response. Endothelialitis, and hence EC dysfunction, subsequently induces a procoagulant state (CAC), loss of barrier function, inflammatory tissue infiltration, edema, and injury (49,50). Cardiovascular comorbidities are usually associated with chronic EC dysfunction, which can explain the worse outcome when further acute insult is superadded.
However, endothelialitis was not a consistent finding in our reviewed studies but, when detected, was associated with microthrombi and had multiorgan distribution. Varga et al. showed multiorgan endotheliitis in all three studied cases (7). Ackermann et al. showed widespread endotheliitis and capillary thrombosis in COVID-19-affected lungs in a much more common prevalence than in non-COVID acute respiratory distress syndrome (ARDS) lungs (51). In contrast, Bradley et al. concluded not only no evidence of endothelialitis but also little evidence of cardiac microthrombi (9). Rapkiewics et al. noted no endothelial abnormalities but a platelet-rich microthrombi in all seven hearts examined, despite anticoagulation (8). It appears that alternate mechanisms of ischemia overlap, and while anticoagulation may be highly relevant in limiting pulmonary thrombosis, this may be less likely to significantly ameliorate any cardiac contribution to poor outcomes. Nicolai et al. highlighted thrombi to be rich in platelets, fibrin, and neutrophil extracellular traps (NETs), while Jensen et al. described platelet-rich cerebral microangiopathy (36,46). The role of NET and platelets may be significant and could support other potential therapies (e.g., antiplatelet therapy).

Chamber Dilatation and Myocardial Edema
Heart weight exceeded the normal range in 90% of cases reflecting a combination of chronic pathologies (e.g., hypertrophy), myocardial edema (marker of injury), and chambers dilatation. The observed cardiac dilatation (especially of the right heart) may be long standing or acute and hence relate to preload or afterload (pulmonary hypertension) changes occurring during the acute illness and its treatment. ME reflects myocardial tissue response to most types of injury and hence its nonspecificity. Ischemia, septic cardiomyopathy, viral, or inflammatory infiltration can all contribute to it. Schmittinger et al. showed ME in 90% of PM septic hearts in a patchy distribution (median of 25% of tissue sections) (52). Of note, ME can reflect an early tissue change after insult (as early as 3 min in the setting of ischemia due to the disruption of the Na+/K+ pump) (53). Detecting ME has therapeutic implications, as it causes less energetic efficiency, arrhythmias, and reduced cardiac wall compliance. All of these are expected to impair systolic and diastolic function and can ultimately lead to fibrosis (53,54). While cardiac MRI (CMR) can detect it in vivo, histological diagnosis remains technically challenging (53). This challenge, combined with the lack of standardized protocol guidelines for PM cardiac pathology reporting, may mean that ME was overlooked in many of the published reports.

Myocardial Fibrosis
Myocardial fibrosis was reported in nearly a quarter of cases. It is the end result of cardiac injury arising from different acute or chronic mechanisms. Cytokines were also implicated in cardiac fibroblast activation (55,56).
The interpretation in COVID-19 is difficult and depends on many factors. It can reflect a chronic or a de novo subacute process. Aging and many reported comorbidities are strongly associated with fibrosis (56). Of note, amyloidosis (a pathology associated with fibrosis) was described in 11 cases and was significantly more prevalent when compared to a historical agematched cohort (18,37,48).
Myocardial fibrosis can be divided into two types: interstitial fibrosis and replacement fibrosis, with considerable overlap between the two (55). While interstitial fibrosis is considered reactive and potentially reversible, replacement fibrosis is not (55). Interstitial fibrosis was previously detected in 100% of PM septic hearts but in a patchy nature (52). Such focal nature means that an extensive PM cardiac pathological examination is necessary. In fact, CMR may be superior as a diagnostic modality despite the difficultly to perform in unstable patients (55,57).
Myocardial fibrosis represents the structural equivalent of heart failure. While ME is expected in the "reversible" septic cardiomyopathy, increased fibrous deposition (i.e., replacement fibrosis) would not be a likely finding in such reversible pathology (52,57,58).

Viral Invasion, Inflammatory Infiltrate, and Myocarditis
Studies investigating the presence of SARS-CoV-2 within the myocardium were positive in about half the cases. In 1986, The Dallas criteria were proposed for the histopathological categorization and diagnosis of myocarditis based on endomyocardial biopsies. The "Dallas criteria" defines acute myocarditis as "an inflammatory infiltrate associated with myocyte necrosis or damage not characteristic of myocardial ischemia." Borderline myocarditis requires a less intense inflammatory infiltrate with no light microscopic signs of myocyte destruction (59). In COVID-19 PM studies, inflammatory infiltrate (mainly lymphocytic) was observed in a minor proportion (about 10%) and was limited in extent for the majority of cases. As such, when interstitial edema and inflammatory infiltrate were observed, they did not meet the diagnostic criteria of myocarditis, except in five cases. In fact, some authors attributed such inflammatory infiltrate to an ischemic process (28). This suggests that contrary to early conjectures, acute and fulminant myocarditis are rare during the acute illness.

Clinical and Imaging Correlation
Correlating the histopathological data to the clinical, imaging, and investigational data can provide more insights into the likely mechanisms of cardiac involvement in COVID-19. Clinical presentation varies from ST elevation MI due to thrombotic occlusion of epicardial coronaries, to ischemia and/or infarction without obstructive coronary disease, through to tachy and brady arrhythmias, depressed left and right ventricular function, and occasional pericardial involvement (60). A review of published literature suggests that elevated Troponin and heart failure dominate the clinical presentations (61).
Echocardiography is readily performed in the acute setting but provides limited insights into the cause when compared to CMR. In a large multinational survey, Dweck et al. reported the echocardiographic findings in 1,216 studies performed over 17 days (62). Fifty-five percent of scans were abnormal. Impaired LV function or dilation (39%) followed by RV abnormalities (33%) dominated. These findings are non-specific, but clear wall motion abnormalities suggesting infarction were rare (3%). The RV abnormalities most likely relate to increased afterload given the high prevalence of pulmonary thromboembolism and extensive lung damage associated with COVID-19 infection (63). The LV abnormalities are non-specific but provide further evidence of the high prevalence of cardiac damage.
CMR-based studies have focused on patients post recovery (too late for confirmation of myocarditis) but have shown a high prevalence of abnormalities. The largest to date is a German study of relatively young patients (mean age, 49 years), largely managed at home (67 of 100), studied a median of 71 days post infection. Seventy-eight percent were reported to show abnormalities, including reduction in LV function, elevated T1 and T2 (the latter suggesting ME) and late gadolinium enhancement (LGE) (nonischemic pattern in 20, ischemic in 12). Three patients with very elevated T2 were referred for endomyocardial biopsy and typical features of myocarditis reported. The T1 and T2 abnormalities suggest ongoing myocardial edema, and the LGE enhancement suggests fibrosis-both of which are common in the autopsy data (64).
The second CMR-based study included only patients in whom Troponin had been elevated during hospital admission. Fiftyone patients were studied 27 days post hospital discharge. In 22 patients, pulmonary embolism and/or coronary ischemia were identified before scanning as the most likely cause of troponin leakage. Among 29 patients (mean age, 64 years) with no clinically identified cause for myocardial injury, an ischemic pattern injury (LGE) was identified in 5, dual pathology (ischemic and non-ischemic) in 4, and non-ischemic in 11. Intriguingly, T1 and T2 were not abnormal in this study. This study thus also supports the histological finding of significant myocardial fibrosis but suggests that edema clears fairly quickly in those that recover (2). Again, Rajpal et al. performed CMR on 26 athletes with a history of mild COVID-19 infection. Four of them (15%) had criteria of myocarditis despite mild or no symptoms, and 30% showed signs of previous cardiac injury (65).

What Can We Conclude From Integrating All Available Data?
Merging the clinical, investigational, and autopsy data, we are presented with a picture that demonstrates a high prevalence of cardiac abnormalities, in part due to exacerbation of underlying cardiac pathology and partly coagulation disorders affecting the pulmonary and coronary vessels. Direct cardiac involvement mainly takes the form of non-coronary myocyte death, myocyte dysfunction, and interstitial fibrosis without substantial inflammatory infiltration or clear ischemia.
The role of direct viral cellular damage remains to be fully explored, and if this is the driving force, it is intriguing that the inflammatory response appears muted. However, it is possible that while the virus is rarely causing a fulminant or acute myocarditis, it can cause a persistent chronic myocardial inflammation with significant long-term implications. It is also important to note the reporting of a delayed immune response in the form of Kawasaki's disease in pediatric patients supporting the issue of long-term sequelae of the SARS-CoV-2 infection (66). Whether immunosuppressive treatment (e.g., dexamethasone and Tocilizumab) during the acute illness is of benefit or causes more harm to the heart should await randomized controlled studies including long-term follow-up.
Thus, on balance, the data strongly suggest significant viral replication in the myocardium without true acute myocarditis in most instances, with frequent non-MI pattern fibrosis-consistent with microvascular ischemia/thrombi and, in some cases, endothelial inflammation. Given the frequent presence of fibrosis associated with cell death, it is likely that complete recovery is unlikely-a clear distinction from septic cardiomyopathy. In addition, the exacerbation of underlying disease would appear to frequently unmask coronary disease, further increasing the benefit of careful cardiological follow-up.
As the vast majority of studied patients in this review died during the acute illness and cardiac abnormality was prevalent in the population studied, we can conclude that myocarditis was not a dominant cause of cardiac dysfunction identified premortem in COVID-19 patients, while the role of endothelialitis needs further clarification.

Limitations
Our work delineates the importance of PM to guide the understanding of COVID-19. However, the small number of published PM cases in a disease, which has caused more than 1 million fatalities, highlights a hugely missed opportunity. Cardiac pathological changes are more likely to be focal in nature and hence easily missed if the heart is not examined in its entirety. Furthermore, the high prevalence of myocardial fibrosis, myocyte damage, or viral RNA in some studies but not others suggest a need to standardize histological reporting to establish common ground between pathologists and clinicians. There is also a genuine need for an international case register to gather the largest possible data in the shortest interval.
While our work is limited by the quality and small number of cases per study, we think it can contribute to a better understanding of COVID-19-associated cardiac injury. Other limits include the probable selection and reporting bias. PM is performed for patients who died during the acute illness and for certain subgroups of patients due to clinical or legal reasons. The longest duration of illness in our cohort is 52 days, which means that the long-term evolution or complications of the disease cannot be covered by this review.

CONCLUSIONS
To conclude, our review confirmed the high prevalence of cardiac pathological findings in COVID-19 patients. Cardiac dilatation, ischemia, and thrombosis were the most prevalent findings. SARS-CoV-2 was present in nearly half of the examined hearts, but true myocarditis was evident in just 1.5% of the deceased patients.

DATA AVAILABILITY STATEMENT
The original contributions presented in the study are included in the article/Supplementary Material, further inquiries can be directed to the corresponding author/s.

AUTHOR CONTRIBUTIONS
AR: conceptualization and design, registration of the protocol, conduct of the search, quality assessment, data extraction, data interpretation, and manuscript drafting. SZ: conceptualization, design, and writing of the protocol, extraction and interpretation of the data, and manuscript drafting. HF: data analysis and interpretation and writing and revising the manuscript. JC: data analysis and interpretation and writing and reviewing the manuscript. All authors: contributed to the article and approved the submitted version.