Ultrastructural cardiac pathology: the wide (yet so very small) world of cardiac electron microscopy.

Electron microscopy (EM) was a popular diagnostic tool in the 1970s and early 80s. With the adoption of newer, less expensive techniques, such as immunohistochemistry, the role of EM in diagnostic surgical pathology has dwindled substantially. Nowadays, even in academic centers, EM interpretation is relegated to renal pathologists and the handful of (aging) pathologists with experience using the technique. As such, EM interpretation is truly arcane-understood by few and mysterious to many. Nevertheless, there remain situations in which EM is the best or only ancillary test to ascertain a specific diagnosis. Thus, there remains a critical need for the younger generation of surgical pathologists to learn EM interpretation. Recognizing this need, cardiac EM was made the theme of the Cardiovascular Evening Specialty Conference at the 2023 United States and Canadian Academy of Pathology (USCAP) annual meeting in New Orleans, Louisiana. Each of the speakers contributed their part to this article, the purpose of which is to review EM as it pertains to myocardial tissue and provide illustrative examples of the spectrum of ultrastructural cardiac pathology seen in storage/metabolic diseases, cardiomyopathies, infiltrative disorders, and cardiotoxicities.

Mass spectrometry in IgG4-related disease diagnosis.

We compared liquid chromatography tandem mass spectrometry (LC-MS/MS) against Binding Site immunonephelometry (BSIN) with regards to these methods' abilities to diagnose IgG4-related disease (IgG4-RD). IgG subclasses were gathered from laboratory from December 2011 to December 2020. The IgG4-RD positive and negative patients were diagnosed according to the ACR/EULAR classification criteria by extensive chart review. Both methods' results were compared in terms of test characteristics. For BSIN, there were 43 IgG4-RD positive cases and 174 disease negative cases, while for LC-MS/MS, there were 102 IgG4-RD positive cases and 562 disease negative cases. The majority of IgG4-RD patients by BSIN and LC-MS/MS had an elevated IgG4 level, 81% and 86%, respectively. For BSIN, the ROC curve, cut-off value of 1.25 g/L, had a sensitivity of 81% and a specificity of 84%. For LC-MS/MS, the ROC curve, cut-off value of 1.25 g/L, had a sensitivity of 86% and a specificity of 84%. The responder index score to IgG4 level r-correlation value for BSIN and LC-MS/MS was 0.5 and 0.6, respectively. In our center, LC-MS/MS and BSIN are equivalent test methods in IgG4-RD diagnosis. IgG4 level does correlate with disease activity by the responder index. LC-MS/MS is a valid and equally reliable alternative to BSIN in the diagnosis of IgG4-related disease.

Consensus statement on the processing, interpretation and reporting of temporal artery biopsy for arteritis.

Giant cell arteritis (GCA) is the most common systemic vasculitis in adults in Europe and North America, typically involving the extra-cranial branches of the carotid arteries and the thoracic aorta. Despite advances in noninvasive imaging, temporal artery biopsy (TAB) remains the gold standard for establishing a GCA diagnosis. The processing of TAB depends largely on individual institutional protocol, and the interpretation and reporting practices vary among pathologists. To address this lack of uniformity, the Society for Cardiovascular Pathology formed a committee tasked with establishing consensus guidelines for the processing, interpretation, and reporting of TAB specimens, based on the existing literature. This consensus statement includes a discussion of the differential diagnoses including other forms of arteritis and noninflammatory changes of the temporal artery.

A Novel Tissue Preservation and Transport Solution as a Substitute for Formalin.

Formalin, a common laboratory fixative, is a type 1 carcinogen; a biohazard with risks, environmental, disposal, and legal costs; and a chemical modifier of protein epitopes in tissues. A less-toxic tissue preservation method is therefore badly needed. We have developed a novel tissue preservation medium, Amber, composed of low-potassium dextran glucose, 10% honey, and 1% coconut oil. This study investigates Amber as compared with formalin with respect to the following aspects: (1) histologic preservation, (2) epitope integrity with immunohistochemistry (IHC) and immunofluorescence (IF), and (3) integrity of tissue RNA. Rat and human lung, liver, kidney, and heart tissues were collected and stored for 24 hours at 4 °C in Amber or formalin. The tissues were evaluated with hematoxylin and eosin; IHC: thyroid transcription factor, muscle-specific actin, hepatocyte-specific antigen, and common acute lymphoblastic leukemia antigen; and IF: VE-cadherin, vimentin, and muscle-specific actin. RNA quality upon extraction was also assessed. Amber demonstrated superior and/or noninferior performance in rat and human tissue evaluation with respect to standard techniques of histology, IHC, IF, and extracted RNA quality. Amber maintains high-quality morphology without compromising the ability to perform IHC and nucleic acid extraction. As such, Amber could be a safer and superior substitute to formalin for clinical tissue preservation for contemporary pathological examination.

Myocarditis Following COVID-19 Vaccination.

Myocarditis is an established but rare adverse event following administration of messenger RNA-based coronavirus disease 2019 (COVID-19) vaccines and is most common in male adolescents and young adults. Symptoms typically develop within a few days of vaccine administration. Most patients have mild abnormalities on cardiac imaging with rapid clinical improvement with standard treatment. However, longer term follow-up is needed to determine whether imaging abnormalities persist, to evaluate for adverse outcomes, and to understand the risk associated with subsequent vaccination. The purpose of the review is to evaluate the current literature related to myocarditis following COVID-19 vaccination, including the incidence, risk factors, clinical course, imaging findings, and proposed pathophysiologic mechanisms.

Cardiac Magnetic Resonance Imaging T1 and T2 Mapping in Systemic Lupus Erythematosus in Relation to Antimalarial Treatment.

PURPOSE: Patients with systemic lupus erythematosus (SLE) are at risk of cardiac disease including antimalarial-induced cardiomyopathy (AMIC). The purpose of this study is to evaluate cardiac magnetic resonance imaging parametric mapping findings in SLE patients with AMIC and investigate the relationship of T1/T2 mapping to antimalarial (AM) treatment duration. MATERIALS AND METHODS: All patients with SLE who had undergone cardiac magnetic resonance imaging with T1/T2 mapping for evaluation of suspected cardiac disease between 2018 and 2021 were evaluated and compared with healthy controls. To facilitate comparison between scanners, T1/T2 values were converted to a z -score using scanner-specific local reference values. Patients were classified into 3 groups: AMIC, myocarditis, and other (no AMIC or myocarditis). RESULTS: Forty-five SLE patients (47±17 y, 80% female; 8 [18%] with AMIC and 7 [16%] with myocarditis) and 30 healthy controls (39±15 y, 60% female) were included. Patients with AMIC had higher T1 and T2 compared with controls ( z -score 1.1±1.3 vs. 0±0.6, P =0.01 and 1.7±1.1 vs. 0±1.0, P <0.01, respectively) and lower values compared with those with myocarditis (3.7±1.6, P <0.01 and 4.0±2.0, P <0.01, respectively). T1 correlated negatively with AM treatment duration in patients without AMIC or myocarditis ( r =-0.36, P =0.048) and positively in patients with AMIC ( r =0.92, P =0.001). AM treatment duration did not correlate significantly with T1 in patients with myocarditis or with T2 in any group. CONCLUSIONS: The relationship between T1 and AM treatment duration differed between groups. Native T1 decreases with longer treatment in patients without AMIC or myocarditis, possibility due to glycosphingolipid accumulation. In patients with AMIC, increasing T1 with longer treatment could reflect fibrosis.

Comparison of modes of failure and clinical outcomes between explanted porcine and bovine pericardial bioprosthetic valves.

OBJECTIVE: To compare pathological and hemodynamic modes of failure and operative outcomes between explanted porcine and bovine pericardial bioprosthetic valves. METHODS: Patients who underwent explantation of their bioprosthetic valves at Toronto General Hospital from 2007 to 2019 were identified. Retrospective chart review was conducted to attain demographic information, operative outcomes, and echocardiography and pathology reports. RESULTS: A total of 278 patients underwent explantation of their porcine (n=183) or bovine pericardial (n=95) valves. A greater proportion of the porcine group had severe regurgitation, compared to the bovine group (45.3% vs. 19.8%, p<.001). Porcine valves had higher rates of cusp flail (19.4% vs. 3.3%, p<.001). The rates of moderate or worse stenosis were higher among bovine pericardial valves (37.9% vs. 15.8%, p<.001). On pathologic examination, the porcine valves exhibited more cusp tears (67.6% vs. 50.5%, p=.006), while higher incidences of calcification were found in the bovine group (p<.001). Rate of stroke was higher during the explantation procedure of the bovine valves (5.3% vs. 0.5%, p=.040). CONCLUSIONS: The primary mode of failure was regurgitation in porcine valves due to cusp tears and stenosis in bovine valves due to calcification. Establishing a clear understanding of failure modes based on valve material may improve design and guide valve selection at the time of surgery.

Outcomes of Reimplantation of the Aortic Valve in Patients With Aortic Cusp Fenestration.

BACKGROUND: Aortic cusp fenestrations are common in patients with aortic root aneurysm, and their management during aortic valve repair remains controversial. We believe that fenestrations in the area of the commissures may rupture after reimplantation of the aortic valve because this operation increases the mechanical stress on the cusps. For this reason we have reinforced the free margin of the aortic cusp with fenestration with fine Gore-Tex sutures (WL Gore). This study examines the outcomes of reimplantation of the aortic valve in patients who had cusp fenestration reinforced with Gore-Tex sutures. METHODS: A review of all patients who had reimplantation of the aortic valve for aortic root aneurysm disclosed 111 patients who had at least 1 cusp fenestration reinforced with a double layer of a fine Gore-Tex suture. The outcomes of these patients were examined and compared with a group of patients without fenestration using propensity score analysis. All patients were followed prospectively with images of the heart. RESULTS: The median follow-up was 8.3 years. Overall the cumulative incidence of aortic valve reintervention at 15 years was 4.8% and the cumulative incidence of aortic insufficiency of moderate or severe degree was 9.2%. Comparison of outcomes of patients with and without fenestrations showed similar results up to 15 years of follow-up. CONCLUSIONS: Reinforcement of the free margins of cusps with fenestrations using Gore-Tex sutures is safe and does not seem to adversely affect the durability of reimplantation of the aortic valve.

Cardiac Imaging in Myocarditis: Current Evidence and Future Directions.

Myocarditis is defined as a non-ischemic inflammatory disease of the myocardium. It remains a challenge to diagnose given non-specific symptoms and lack of specific blood biomarkers. Cardiac imaging plays an important role in the evaluation of myocarditis with unique strengths and limitations of different imaging modalities, including cardiac magnetic resonance imaging, echocardiography, cardiac computed tomography, and positron emission tomography. The purpose of this review is to discuss the strengths and limitations of various cardiac imaging techniques in the evaluation of myocarditis, review imaging findings in specific causes of myocarditis including COVID-19 and after vaccination, evaluate the role of imaging in differentiating myocarditis from potential mimics and differential considerations, identify current gaps in knowledge, and propose future directions.

Myocarditis Following COVID-19 Vaccination.

Myocarditis is an established but rare adverse event following administration of messenger RNA-based coronavirus disease 2019 (COVID-19) vaccines and is most common in male adolescents and young adults. Symptoms typically develop within a few days of vaccine administration. Most patients have mild abnormalities on cardiac imaging with rapid clinical improvement with standard treatment. However, longer term follow-up is needed to determine whether imaging abnormalities persist, to evaluate for adverse outcomes, and to understand the risk associated with subsequent vaccination. The purpose of the review is to evaluate the current literature related to myocarditis following COVID-19 vaccination, including the incidence, risk factors, clinical course, imaging findings, and proposed pathophysiologic mechanisms.

Microcalcification and Thoracic Aortopathy: A Window Into Disease Severity.

BACKGROUND: Patients with thoracic aortopathy are at increased risk of catastrophic aortic dissection, carrying with it substantial mortality and morbidity. Although granular medial calcinosis (medial microcalcification) has been associated with thoracic aortopathy, its relationship to disease severity has yet to be established. METHODS: One hundred one thoracic aortic specimens were collected from 57 patients with thoracic aortopathy and 18 control subjects. Standardized histopathologic scores, immunohistochemistry, and nanoindentation (tissue elastic modulus) were compared with the extent of microcalcification on von Kossa histology and 18F-sodium fluoride autoradiography. RESULTS: Microcalcification content was higher in thoracic aortopathy samples with mild (n=28; 6.17 [2.71-10.39]; P≤0.00010) or moderate histopathologic degeneration (n=30; 3.74 [0.87-11.80]; P<0.042) compared with control samples (n=18; 0.79 [0.36-1.90]). Alkaline phosphatase (n=26; P=0.0019) and OPN (osteopontin; n=26; P=0.0045) staining were increased in tissue with early aortopathy. Increasingly severe histopathologic degeneration was related to reduced microcalcification (n=82; Spearman ρ, -0.51; P<0.0001)-a process closely linked with elastin loss (n=82; Spearman ρ, -0.43; P<0.0001) and lower tissue elastic modulus (n=28; Spearman ρ, 0.43; P=0.026).18F-sodium fluoride autoradiography demonstrated good correlation with histologically quantified microcalcification (n=66; r=0.76; P<0.001) and identified areas of focal weakness in vivo. CONCLUSIONS: Medial microcalcification is a marker of aortopathy, although progression to severe aortopathy is associated with loss of both elastin fibers and microcalcification.18F-sodium fluoride positron emission tomography quantifies medial microcalcification and is a feasible noninvasive imaging modality for identifying aortic wall disruption with major translational promise.

Pleiotropic activation of endothelial function by angiotensin II receptor blockers is crucial to their protective anti-vascular remodeling effects.

There are no therapeutics that directly enhance chronic endothelial nitric oxide (NO) release, which is typically associated with vascular homeostasis. In contrast, angiotensin II (AngII) receptor type 1 (AT1R) blockers (ARBs) can attenuate AngII-mediated oxidative stress, which often leads to increased endothelial NO bioavailability. Herein, we investigate the potential presence of direct, AngII/AT1R-independent ARB class effects on endothelial NO release and how this may result in enhanced aortic wall homeostasis and endothelial NO-specific transcriptome changes. Treatment of mice with four different ARBs induced sustained, long-term inhibition of vascular contractility by up to 82% at 16 weeks and 63% at 2 weeks, an effect reversed by L-NAME and absent in endothelial NO synthase (eNOS) KO mice or angiotensin converting enzyme inhibitor captopril-treated animals. In absence of AngII or in tissues with blunted AT1R expression or incubated with an AT2R blocker, telmisartan reduced vascular tone, supporting AngII/AT1R-independent pleiotropism. Finally, telmisartan was able to inhibit aging- and Marfan syndrome (MFS)-associated aortic root widening in NO-sensitive, BP-independent fashions, and correct aberrant TGF-ß signaling. RNAseq analyses of aortic tissues identified early eNOS-specific transcriptome reprogramming of the aortic wall in response to telmisartan. This study suggests that ARBs are capable of major class effects on vasodilatory NO release in fashions that may not involve blockade of the AngII/AT1R pathway. Broader prophylactic use of ARBs along with identification of non-AngII/AT1R pathways activated by telmisartan should be investigated.

18F-NaF PET/MRI for Detection of Carotid Atheroma in Acute Neurovascular Syndrome.

Background MRI and fluorine 18-labeled sodium fluoride (18F-NaF) PET can be used to identify features of plaque instability, rupture, and disease activity, but large studies have not been performed. Purpose To evaluate the association between 18F-NaF activity and culprit carotid plaque in acute neurovascular syndrome. Materials and Methods In this prospective observational cohort study (October 2017 to January 2020), participants underwent 18F-NaF PET/MRI. An experienced clinician determined the culprit carotid artery based on symptoms and record review. 18F-NaF uptake was quantified using standardized uptake values and tissue-to-background ratios. Statistical significance was assessed with the Welch, χ2, Wilcoxon, or Fisher test. Multivariable models were used to evaluate the relationship between the imaging markers and the culprit versus nonculprit vessel. Results A total of 110 participants were evaluated (mean age, 68 years ± 10 [SD]; 70 men and 40 women). Of the 110, 34 (32%) had prior cerebrovascular disease, and 26 (24%) presented with amaurosis fugax, 54 (49%) with transient ischemic attack, and 30 (27%) with stroke. Compared with nonculprit carotids, culprit carotids had greater stenoses (≥50% stenosis: 30% vs 15% [P = .02]; ≥70% stenosis: 25% vs 4.5% [P < .001]) and had increased prevalence of MRI-derived adverse plaque features, including intraplaque hemorrhage (42% vs 23%; P = .004), necrotic core (36% vs 18%; P = .004), thrombus (7.3% vs 0%; P = .01), ulceration (18% vs 3.6%; P = .001), and higher 18F-NaF uptake (maximum tissue-to-background ratio, 1.38 [IQR, 1.12-1.82] vs 1.26 [IQR, 0.99-1.66], respectively; P = .04). Higher 18F-NaF uptake was positively associated with necrosis, intraplaque hemorrhage, ulceration, and calcification and inversely associated with fibrosis (P = .04 to P < .001). In multivariable analysis, carotid stenosis at or over 70% (odds ratio, 5.72 [95% CI: 2.2, 18]) and MRI-derived adverse plaque characteristics (odds ratio, 2.16 [95% CI: 1.2, 3.9]) were both associated with the culprit versus nonculprit carotid vessel. Conclusion Fluorine 18-labeled sodium fluoride PET/MRI characteristics were associated with the culprit carotid vessel in study participants with acute neurovascular syndrome. Clinical trial registration no. NCT03215550 and NCT03215563 © RSNA, 2022 Online supplemental material is available for this article.

Alpha-synuclein seeding shows a wide heterogeneity in multiple system atrophy.

BACKGROUND: Multiple system atrophy (MSA) is a neurodegenerative condition characterized by variable combinations of parkinsonism, autonomic failure, cerebellar ataxia and pyramidal features. Although the distribution of synucleinopathy correlates with the predominant clinical features, the burden of pathology does not fully explain observed differences in clinical presentation and rate of disease progression. We hypothesized that the clinical heterogeneity in MSA is a consequence of variability in the seeding activity of α-synuclein both between different patients and between different brain regions. METHODS: The reliable detection of α-synuclein seeding activity derived from MSA using cell-free amplification assays remains challenging. Therefore, we conducted a systematic evaluation of 168 different reaction buffers, using an array of pH and salts, seeded with fully characterized brain homogenates from one MSA and one PD patient. We then validated the two conditions that conferred the optimal ability to discriminate between PD- and MSA-derived samples in a larger cohort of 40 neuropathologically confirmed cases, including 15 MSA. Finally, in a subset of brains, we conducted the first multi-region analysis of seeding behaviour in MSA. RESULTS: Using our novel buffer conditions, we show that the physicochemical factors that govern the in vitro amplification of α-synuclein can be tailored to generate strain-specific reaction buffers that can be used to reliably study the seeding capacity from MSA-derived α-synuclein. Using this novel approach, we were able to sub-categorize the 15 MSA brains into 3 groups: high, intermediate and low seeders. To further demonstrate heterogeneity in α-synuclein seeding in MSA, we conducted a comprehensive multi-regional evaluation of α-synuclein seeding in 13 different regions from 2 high seeders, 2 intermediate seeders and 2 low seeders. CONCLUSIONS: We have identified unexpected differences in seed-competent α-synuclein across a cohort of neuropathologically comparable MSA brains. Furthermore, our work has revealed a substantial heterogeneity in seeding activity, driven by the PBS-soluble α-synuclein, between different brain regions of a given individual that goes beyond immunohistochemical observations. Our observations pave the way for future subclassification of MSA, which exceeds conventional clinical and neuropathological phenotyping and considers the structural and biochemical heterogeneity of α-synuclein present. Finally, our methods provide an experimental framework for the development of vitally needed, rapid and sensitive diagnostic assays for MSA.

The Role of Machine Learning in Cardiovascular Pathology.

Machine learning has seen slow but steady uptake in diagnostic pathology over the past decade to assess digital whole-slide images. Machine learning tools have incredible potential to standardise, and likely even improve, histopathologic diagnoses, but they are not yet widely used in clinical practice. We describe the principles of these tools and technologies and some successful preclinical and pretranslational efforts in cardiovascular pathology, as well as a roadmap for moving forward. In nonhuman animal models, one proof-of-principle application is in rodent progressive cardiomyopathy, which is of particular significance to drug toxicity studies. Basic science successes include screening the quality of differentiated stem cells and characterising cardiomyocyte developmental stages, with potential applications for research and toxicology/drug safety screening using derived or native human pluripotent stem cells differentiated into cardiomyocytes. Translational studies of particular note include those with success in diagnosing the various forms of heart allograft rejection. For fully realising the value of these tools in clinical cardiovascular pathology, we identify 3 essential challenges. First is image quality standardisation to ensure that algorithms can be developed and implemented on robust, consistent data. The second is consensus diagnosis; experts don't always agree, and thus "truth" may be difficult to establish, but the algorithms themselves may provide a solution. The third is the need for large-enough data sets to facilitate robust algorithm development, necessitating large cross-institutional shared image databases. The power of histopathology-based machine learning technologies is tremendous, and we outline the next steps needed to capitalise on this power.

A Tale of 2 Aneurysms: Cardiogenic Shock Secondary to Vascular Behçet's Syndrome.

A patient with vascular Behçet's syndrome (BS), a subtype of BS with mainly venous/arterial manifestations, presented with a left main aneurysm/thrombus and cardiogenic shock. The clinical diagnosis of BS includes mucocutaneous, vascular, and neurologic criteria. It is important to consider vascular BS as a nonatherosclerotic cause of coronary aneurysms. (Level of Difficulty: Intermediate.).