Stress cardiomyopathy (SCM) is a unique form of heart disease that primarily affects women. Described in 1990 by Sato and colleagues, it is now recognized as accounting for 5.9–7.5% of women presenting to the emergency department with acute coronary syndromes (1–4). Although the prognosis is typically good, complications such as heart failure, arrhythmias, and thromboembolic phenomena do occur (5). Recent studies have shown that SCM patients have increased long-term morbidity and mortality (5.6% risk of death per patient year) (6–8). Therefore, this disorder may not be as benign as previously thought.

After almost 30 years of study, the mechanism of SCM is still not understood. Plasma concentrations of norepinephrine, epinephrine, and dopamine are twofold higher than those of AMI (9). Myocardial biopsy in SCM reveals inflammation and myocyte contraction band necrosis, which are features of catecholamine excess (9). In contrast, acute myocardial infarction (AMI) has greater myocyte necrosis (10). Despite these different mechanisms of injury, cardiac function is impaired to an equivalent degree in both conditions (11). The exact mechanism by which LV dysfunction occurs in SCM remains poorly understood.

Due to the presenting signs and symptoms, patients with SCM are referred for urgent coronary angiography, which reveals the absence of atherosclerotic plaque rupture. Coronary angiography exposes the SCM patient to unnecessary risk without therapeutic benefit. A biomarker that could differentiate between AMI and SCM would be useful, allowing for the identification of low risk SCM patients in whom angiography could be avoided. This would be especially helpful in patients with “secondary” SCM (5). “Secondary SCM” frequently occurs in medical intensive care units and is precipitated by critical illnesses such as sepsis (12). Due to the severity of illness in these cases, cardiac interventions such as coronary angiography or the initiation of antiplatelet therapy are often deferred.

There have been no studies examining novel protein biomarkers in SCM. A ratio of serum B-type natriuretic peptide to troponin I (TnI) ≥1,272 is highly specific for SCM (95%) but has limited sensitivity (52%) (13). Soluble suppression of tumorigenicity 2 has been studied to identify SCM in medical intensive care units (12). A circulating profile of plasma microRNAs (miR-16, mir-26a, miR-1, and miR-133a) has shown good sensitivity and specificity for SCM (14). However, the use of plasma microRNAs for diagnostic purposes has not been adopted in clinical practice (15). Furthermore, these studies shed no light into the biologic differences between these two syndromes.

We hypothesized that there would be differences between circulating proteins in SCM and AMI that might elucidate the underlying mechanisms. Therefore, the purpose of our study was twofold. The first aim was to perform an unbiased comparison of circulating proteins in SCM and AMI in order to provide insight into the pathophysiology of SCM. Our second aim was to identify putative protein biomarkers of SCM.

In order to achieve these aims, we utilized recently described DNA aptamer technology to perform our screen (SOMAscan V1.3K, SOMAlogics) (16–18). This assay uses single stranded DNA aptamers to specifically bind 1,310 predetermined proteins in a single sample. The advantages of this technology include higher throughput and precision than traditional proteomic methods such as mass spectrometry or antibody based methods (16, 17).

Using this assay, we found that women presenting with either SCM or AMI have an increase in proteins related to complement, coagulation, and inflammation. The unexpected and profound activation of inflammatory and pro-thrombotic pathways in SCM may contribute to the increased risk of mortality attributed to this disorder (7, 8).

Although the pathophysiology, natural history, and outcomes of AMI and SCM are different, they do share common features. First, both are characterized by acutely decreased cardiac function (11). Second, each condition can be complicated by heart failure, arrhythmia, or thromboembolic events. Due to these similarities, women presenting with SCM are often referred for coronary angiography (5). However, in low risk cases, and in cases of “secondary SCM,” a biomarker would be useful. One of the aims of our study was to discover putative biomarkers of SCM; we identified four candidates (FSTL3, Kallikrein7, KYNU, and NET1). A second aim was to learn about the pathophysiology of SCM by comparing serum proteins to normal patients and those with AMI. We were surprised to find no differences between SCM and AMI in the acute phase. Both conditions were characterized by a robust increase in proteins related to inflammation, coagulation, and complement.

The characteristics of both study groups were consistent with prior studies. Patients with AMI had a greater peak TnI and a lower LVEF (%) on follow-up (7, 9, 11). SCM patients had smaller indexed LV diastolic and systolic volumes in both the acute and recovery phase. SCM patients had normalization of their systolic function in recovery, as demonstrated by a greater increase in LVEF (%) (Table 2).

In regards to our proteomic analysis, we wish to emphasize that the SOMAscan 1.3 K measures a predetermined panel of 1,310 proteins, which is only 6.5% of the approximately 20,000 protein encoding genes in the genome. Therefore, there are many changes that were undetected by our assay. For example, tissue inhibitor of metalloproteinase 4 has previously been shown to be increased in SCM compared to AMI (26). However, tissue inhibitor of matrix metalloproteinase (TIMP)-4 specific aptamers were not included in the version of the SOMAscan that we used; therefore, this difference was not detected. Of the proteins that were included, many were commonly elevated in both AMI and SCM, particularly those related to complement, coagulation, and innate immunity. This was unexpected, as the degree of myocardial necrosis in SCM is less than in AMI (7, 9). The trigger for this acute inflammatory response remains unclear.

Complement is an arm of the innate immune system that aids in the detection and clearance of self and bacterial antigens (27). It was demonstrated in 1990 that complement is activated in AMI (27). This is the first report of complement activation in SCM. Local hypoxia may play a role in complement activation and neutrophil chemoattraction to sites of ischemia (27). Complement activation can trigger endothelial adhesion molecule expression, and subsequent leukocyte rolling, arrest, and diapedesis. It is notable that there are several small studies indicating that patients with SCM have greater endothelial dysfunction than those with AMI even in the subacute phase (28–30). This dysfunction could be due to low-level complement activation, increased leukocyte adhesion, and release of reactive oxygen species.

Complement is known to interact with the coagulation cascade (27). The increased levels of coagulation factor V (F5) and tissue plasminogen activator (PLAT, tPA) in SCM indicate that a pro-thrombotic state may be present. Patients with SCM have inducible hyperviscosity and there are multiple reports of increased thromboembolic events in the acute phase of SCM (31–33).

Several candidate proteins demonstrated a non-significant trend to be increased in acute SCM compared to AMI and controls [NET1, FSTL3, kallikrein 7 (KLK7), KYNU] (Figure 1, cluster 3). Netrin-1 is a neuronal growth factor that was originally reported in 1991 (34). NET1 promotes endothelial cell nitric oxide production and cell growth and migration in vitro via the receptor deleted in colorectal cancer (DCC) (35). NET1 has also been shown to reduce infarct size in mice (36). There is interest in using NET1 for the treatment of postischemic conditioning in AMI.

Follistatin-like 3 (FSTL3) was increased in SCM compared to controls. FSTL3 inhibits transforming growth factor beta (TGFβ) signaling (37). FSTL3 is upregulated in the myocardium and endothelium of failing human hearts (38). Cardiac specific ablation of FSTL3 reduces cardiac hypertrophy and fibrosis in response to transverse aortic constriction (37). FSTL3 may contribute to myocyte hypertrophy in response to stress stimuli, and the increased levels of FSTL3 in SCM may promote cardiac hypertrophy and fibrosis. This is consistent with prior data demonstrating that SCM has a matrix metalloproteinase (MMP)/tissue inhibitor of matrix metalloproteinase (TIMP) profile similar to hypertensive heart disease, which is distinct from AMI (26). We observed the same MMP/TIMP profile in our study (data not shown); however, the differences did not reach statistical significance. This is likely because the prior study used ELISA to measure MMP/TIMPs (26).

Kynureninase is an enzyme involved in tryptophan metabolism. It degrades 3-hydroxy kynurenine to 3-hydroxyanthranillic acid (39). Kynurenine is an endothelium-derived relaxation factor induced in sepsis (40). Whether or not KYNU is truly elevated in SCM will be the topic of future studies. Finally, KLK7 is a serine proteinase belonging to the human kallikrein gene family, of which there are 15 members (41). The kallikrein system is broadly expressed and has also been demonstrated to play a role in cardiac remodeling (42).

In summary, despite different pathophysiologic mechanisms, the circulating proteome of AMI and SCM is similar. Activation of complement and coagulation pathways in SCM has not been previously reported and highlights the pro-thrombotic and inflammatory state of acute SCM. The stimulus for inflammation in SCM is unclear and will be the subject of future investigation. NET1, FSTL3, and KYNU, and KLK7 are putative markers of SCM that require prospective validation.

This study was carried out in accordance with the recommendations of “Committee for the Protection of Human Subjects in Research” with written informed consent from all subjects. All subjects gave written informed consent in accordance with the Declaration of Helsinki. The protocol was approved by the Institutional Review Board of UMASS Medical Center, IRB docket #00000941.

TF designed the study, interpreted the data, drafted and revised the manuscript, approved the final version, and is accountable for all aspects of the work. YE, PS, AI, RG, SS, JB, TS, and MA acquired the data, revised the manuscript, approved the final version, and are accountable for all aspects. JK, MZ, and GA made substantial contributions to the study design, revised the manuscript, approved the final version, and are accountable for all aspects of the work.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.