We report 55 postchemotherapy resections of primary nonseminomatous mediastinal germ cell tumors with prominent vasculogenic features showing the formation of rudimentary to well-developed neoplastic vessels within primitive mesenchyme. These cases represented 25% of a cohort of 221 such specimens. The patients were 19 to 49 years old (mean, 28 y) and 98% had serological evidence of yolk sac tumor. The vasculogenic lesions, felt to represent a neoplastic reiteration of embryonic vasculogenesis in the splanchnic mesoderm of the yolk sac, were further subdivided into teratoma with vasculogenic stroma (n=9), vasculogenic mesenchymal tumor (VMT) (n=42, further classified into low grade [n=24] and high grade [n=18]), and angiosarcoma (n=4). The distinction of teratoma with vasculogenic stroma from VMT was based solely on the greater extent of VMT (exceeding 1 low power [×4 objective] microscopic field), with both categories showing a spectrum of vessels lined by atypical endothelium in a nonendothelial neoplastic stroma that often also generated vascular walls comprised of atypical smooth muscle. The angiosarcomas showed stratification of highly atypical endothelial cells or anastomosing vessels lined by nonstratified but cytologically similar endothelium. Immunohistochemical studies supported the generation of neoplastic vessels from the tumor stroma, most commonly by the development of stromal clefts showing reactivity for podoplanin, CD34, and occasionally ERG, followed by the gradual development from the clefts of thin-walled vessels that later became encircled by stromal cells showing smooth muscle differentiation by immunohistochemistry. Occasionally, round collections of stromal erythrocytes became surrounded by stromal cells to generate blood vessels. Fluorescence in situ hybridization showed chromosome 12p copy number increase in both the endothelial component and stromal component in 8/9 VMT cases and in 1/1 angiosarcoma. On follow-up, no patient with teratoma with vasculogenic stroma had evidence of a subsequent vascular tumor or sarcoma, whereas 8 of the 35 (23%) patients with VMTs (2 low grade and 6 high grade) and meaningful follow-up developed sarcoma (1 angiosarcoma, 2 rhabdomyosarcomas, and 5 not further characterized). The difference between low-grade and high-grade tumors was of borderline significance (P=0.058). Two of the 4 patients with angiosarcoma died of metastatic angiosarcoma, with the other 2 disease-free at 6.8 and 7 years. Compared with the 165 patients with follow-up and no vasculogenic lesions, there was a highly significant (P=4.3×10−5) association of any vasculogenic lesion with sarcomatoid tumors during the clinical course of VMT patients. In addition, 5/46 patients with follow-up and vasculogenic lesions (11%) died of either leukemia or myelodysplastic syndrome compared with 2 of 166 (1%) lacking them (P=0.0012). Three of the 5 patients had identifiable immature hematopoietic cells within their vasculogenic lesions, but 4 other VMT patients with these did not develop leukemia or myelodysplasia. We conclude: (1) vasculogenic lesions are frequent in postchemotherapy resections of primary mediastinal germ cell tumors with yolk sac tumor components; (2) they mostly consist of neoplastic vessels in a stroma that also generates neoplastic vascular walls of smooth muscle; (3) VMTs are associated with an increased incidence of sarcomas, even though most vasculogenic lesions in this context do not meet criteria for angiosarcoma; (4) the presence of vasculogenic lesions in postchemotherapy resections of primary mediastinal germ cell tumors place patients at increased risk for leukemia or myelodysplasia.

Primary mediastinal nonseminomatous germ cell tumors represent a rare but important malignancy that occurs in otherwise young and healthy patients. Treatment is challenging and involves cisplatin-based chemotherapy followed by surgery to remove residual disease. Avoiding bleomycin-containing chemotherapy in the treatment of primary mediastinal nonseminomatous germ cell tumors is important. Prechemotherapy and postchemotherapy pathology as well as postoperative serum tumor markers are independent predictors of long-term survival.

Primary mediastinal nonseminomatous germ cell tumors represent a rare but important malignancy that occurs in otherwise young and healthy patients. Treatment is challenging and involves cisplatin-based chemotherapy followed by surgery to remove residual disease. Avoiding bleomycin-containing chemotherapy in the treatment of primary mediastinal nonseminomatous germ cell tumors is important. Prechemotherapy and postchemotherapy pathology as well as postoperative serum tumor markers are independent predictors of long-term survival.

OBJECTIVE:Treatment of primary mediastinal nonseminomatous germ cell tumors involves cisplatin-based chemotherapy followed by surgery to remove residual disease. We undertook a study to determine short and long-term outcomes. METHODS:A retrospective analysis of patients with primary mediastinal nonseminomatous germ cell tumors who underwent surgery at our institution from 1982 to 2017 was performed. RESULTS:A total of 255 patients (mean age, 29.2 years) were identified. Acute respiratory distress syndrome occurred postoperatively in 27 patients (10.9%), which was responsible for all 11 (4.3%) postoperative deaths. Of patients who developed acute respiratory distress syndrome, more patients received bleomycin-containing chemotherapy (25 out of 169; 14.8%) than non-bleomycin regimens (2 out of 77; 2.6%) (P = .004). With respect to variables independently predictive of long-term survival, evidence of choriocarcinoma before chemotherapy (n = 12) was determined to be an adverse factor (P = .006). In contrast, biopsy-proven elements of seminoma (n = 34) were predictive of improved survival (P = .04). The worst pathology identified in the residual mediastinal mass after chemotherapy was necrosis in 61 patients (25.0%), teratoma in 84 patients (34.4%), and malignant (persistent germ cell or non-germ cell cancer) in 97 patients (39.8%), which influenced overall survival (P < .001). Additionally, teratoma with stromal atypia (n = 18) demonstrated decreased survival compared with teratoma without atypia (n = 66; P = .031). Patients with malignancy involving >50% of the residual mass (n = 47) had a 2.3-fold increased risk of death compared with ≤50% malignancy (n = 45; P = .008). Finally, elevated postoperative serum tumor markers (n = 40) was significantly predictive of adverse survival (P < .001). CONCLUSIONS:In the treatment of primary mediastinal nonseminomatous germ cell tumors, avoiding bleomycin-containing chemotherapy is important. Pre- and postchemotherapy pathology and postoperative serum tumor markers are independent predictors of long-term survival.

Prosthesis choice for aortic valve replacement (AVR) in children is frequently compromised by unavailability of prostheses in very small sizes, the lack of prosthetic valve growth, and risks associated with long-term anticoagulation. The Ross procedure with pulmonary valve autograft offers several advantages for pediatric and adult patients. We describe our current Ross AVR technique including replacement of the ascending aorta with a prosthetic graft. The procedure shown in the video involves an adult-sized male with a bicuspid aortic valve, mixed aortic stenosis and insufficiency, and a dilated ascending aorta.

RationaleProtein lysine acetylation is an important post‐translational modification that regulates cardiac metabolism. Sirtuin 3 (SIRT3) is an NAD+‐dependent deacetylase that controls mitochondrial deacetylation. Loss of SIRT3 expression causes increased mitochondrial protein acetylation along with cardiac hypertrophy, fibrosis and increased susceptibility to mechanical and ischemic stress. The childhood heart disease of Friedreich's Ataxia (FRDA) is characterized by cardiac hypertrophy and failure. There is no effective therapy. FRDA is caused by loss of frataxin (FXN), a mitochondrial protein involved in energy homeostasis. FRDA model hearts have increased mitochondrial protein acetylation and impaired SIRT3 activity, which may contribute to the pathophysiology of this heart disease and represent a therapeutic target.ObjectiveThere is evidence that NAD+ repletion by supplementation with its precursor, nicotinamide riboside (NR), can increase SIRT3 activity and improve mitochondrial function. We tested the hypothesis that NR therapy would normalize lysine acetylation and improve cardiac function in a model of FRDA heart disease.Methods and ResultsA conditional mouse model of FRDA with ablation of FXN (FXN KO) in the heart (n=6) and healthy controls (n=6) were treated with oral administration of NR (10 ml/kg/day) from wean at day 30 for 5 weeks. Results were compared to untreated age‐ and sex‐matched animals. We evaluated hearts using echocardiography, cardiac catheterization, histology, protein expression and lysine acetylation. We used the acetylation of a known target of SIRT3, superoxide dismutase 2 (SOD2), as readout for SIRT3 activity.The acetylation state of SOD2 was increased in FXN KO mice compared to controls, and did not change after NR therapy. Surprisingly, FXN KO hearts demonstrated increased global mitochondrial protein lysine acetylation in response to treatment with NR. SIRT3 protein expression did not differ between groups. Cardiac function declined steadily with age in FXN KO animals and there was no improvement after NR supplementation in FXN KO animals. Such outcome parameters included left ventricle thickness (LVPWd) (FXN KO 0.85 ±02.3 vs FXN KO +NR 0.78 ±0.12 mm, p=0.53), ejection fraction (FXN KO 26.6 ±9.7 vs. FXN KO +NR 30.6±13.5%, p=0.53), fractional shortening (FXN KO 12.4 ±4.7 vs. FXN KO +NR 14.5 ±6.7 %, p=0.51) and rates of contraction (FXN KO 5165 ±1019 vs FXN KO +NR 4683 ±760 mmHg/sec, p=0.42) and relaxation (FXN KO −4099 ±1022 vs. FXN KO +NR −3613 ±807 mmHg/sec, p=0.42). Diffuse cardiac fibrosis was evident in FXN KO hearts at 5 weeks post‐wean and there was no observable difference after NR therapy.DiscussionIn FXN KO hearts, SIRT3 may be irreversibly inactivated, possibly via lipid peroxidation from an increase in reactive oxygen species known to arise secondary to the loss of frataxin. Combining NR therapy with SIRT3 replacement, or trialing longer or larger doses of NR, may be more effective in modifying SIRT3 acetylation targets in the FXN KO heart and warrants further investigation.ConclusionIn conclusion, NAD+ supplementation with NR in the FRDA model of mitochondrial heart disease does not alter SIRT3 activity or improve cardiac function.Support or Funding InformationThis work was supported by an NHLBI NRSA fellowship grant #F31HL126489 to ARS, grants from the Friedreich's Ataxia Research Alliance (FARA) and Muscular Dystrophy Association (MDA) to RMP.

Introduction The childhood heart disease of Friedreich’s Ataxia (FRDA) is characterized by hypertrophy and failure. It is caused by loss of frataxin (FXN), a mitochondrial protein involved in energy homeostasis. FRDA model hearts have increased mitochondrial protein acetylation and impaired sirtuin 3 (SIRT3) deacetylase activity. Protein acetylation is an important regulator of cardiac metabolism and loss of SIRT3 increases susceptibility of the heart to stress-induced cardiac hypertrophy and ischemic injury. The underlying pathophysiology of heart failure in FRDA is unclear. The purpose of this study was to examine in detail the physiologic and acetylation changes of the heart that occur over time in a model of FRDA heart failure. We predicted that increased mitochondrial protein acetylation would be associated with a decrease in heart function in a model of FRDA. Methods A conditional mouse model of FRDA cardiomyopathy with ablation of FXN (FXN KO) in the heart was compared to healthy controls at postnatal days 30, 45 and 65. We evaluated hearts using echocardiography, cardiac catheterization, histology, protein acetylation and expression. Results Acetylation was temporally progressive and paralleled evolution of heart failure in the FXN KO model. Increased acetylation preceded detectable abnormalities in cardiac function and progressed rapidly with age in the FXN KO mouse. Acetylation was also associated with cardiac fibrosis, mitochondrial damage, impaired fat metabolism, and diastolic and systolic dysfunction leading to heart failure. There was a strong inverse correlation between level of protein acetylation and heart function. Conclusion These results demonstrate a close relationship between mitochondrial protein acetylation, physiologic dysfunction and metabolic disruption in FRDA hypertrophic cardiomyopathy and suggest that abnormal acetylation contributes to the pathophysiology of heart disease in FRDA. Mitochondrial protein acetylation may represent a therapeutic target for early intervention.

There is an intimate interplay between cellular metabolism and the pathophysiology of disease. Mitochondria are essential to maintaining and regulating metabolic function of cells and organs. Mitochondrial dysfunction is implicated in diverse diseases, such as cardiovascular disease, diabetes and metabolic syndrome, neurodegeneration, cancer, and aging. Multiple reversible post-translational protein modifications are located in the mitochondria that are responsive to nutrient availability and redox conditions, and which can act in protein–protein interactions to modify diverse mitochondrial functions. Included in this are physiologic redox signaling via reactive oxygen and nitrogen species, phosphorylation, O-GlcNAcylation, acetylation, and succinylation, among others. With the advent of mass proteomic screening techniques, there has been a vast increase in the array of known mitochondrial post-translational modifications and their protein targets. The functional significance of these processes in disease etiology, and the pathologic response to their disruption, are still under investigation. However, many of these reversible modifications act as regulatory mechanisms in mitochondria and show promise for mitochondrial-targeted therapeutic strategies. This review addresses the current knowledge of post-translational processing and signaling mechanisms in mitochondria, and their implications in health and disease.

We tested the hypothesis that increased mitochondrial protein acetylation is associated with impaired fatty acid metabolism and diastolic dysfunction in Friedreich’s Ataxia (FRDA). FRDA results from deficiency of the mitochondrial protein, frataxin (FXN), and causes hypertrophic cardiomyopathy. FRDA hearts show decreased ATP production. We previously showed that FXN loss results in loss of activity of the NAD+-dependent mitochondrial deacetylase, sirtuin 3 (SIRT3), and cardiac mitochondrial protein hyperacetylation in a mouse model of FRDA. Long-chain and medium chain acyl CoA dehydrogenases (LCAD, MCAD) are targets of SIRT3, suggesting that abnormal acetylation may alter fatty acid metabolism. A cardiac specific mouse model with conditional deletion of FXN in heart and skeletal muscle (FXN MCK-Cre-/-) was compared to healthy controls (FXNfl/fl). Mice underwent echocardiogram and left heart catheterization in vivo at age 30 and 65 days. Heart lysate was examined for overall lysine acetylation at ages 30, 45 and 70 days, and acetylated LCAD and MCAD. Myocardial cells were stained with oil red to detect lipids. Hearts of FXN MCK-Cre-/- animals showed mitochondrial hyperacetylation, increased LCAD and MCAD acetylation, and microsteatosis compared to controls. Diastolic dysfunction was evident in FXN MCK-Cre-/- at day 65 by increased mitral valve E/A ratios and IVRT (p<0.04 and p<0.03, respectively, n=7), prolonged tau, and decreased -dP/dt (p<0.02 and p <0.03, respectively, n=6). Cardiac mechanics at day 65 showed systolic failure with reduced EF and FS (p<0.002 and p<0.003, respectively, n=7). Comparison of FXN deficient mice and controls at day 65 trended toward flattened ESPVR and left shifted EDPVR curves, but differences were not significant. Early diastolic dysfunction was evident at day 30 in FXN MCK-Cre-/- by high E/A ratio and EDP compared to day 65 controls (p <0.03, n=7, and p<0.03, n=6, respectively). We conclude that progressive mitochondrial protein acetylation and abnormal fatty acid metabolism is associated with early diastolic dysfunction and later systolic failure. This metabolic dysregulation and pathophysiology may share characteristics with other metabolic heart disorders, such as diabetes and metabolic syndrome.