Development, Histological, and Ultrastructural Evaluation of Sheep Hyperimmune Serum Therapy for COVID-19 / Desarrollo, Evaluación Histológica y Ultraestructural de la Terapia con Suero Hiperinmune de Oveja para COVID-19

SUMMARY: In response to the threat posed by new variants of SARS-CoV-2 and the urgent need for effective treatments in the absence of vaccines, the aim of this study was to develop a rapid and cost-effective hyperimmune serum (HS) derived from sheep and assess its efficacy. The utilization of a halal-certified, easily maintained in certain geographic regions, easy-to-handle animal such as sheep could provide a viable alternative to the expensive option of horses. Sheep were immunized with a whole inactivated SARS-CoV- 2 antigen to produce HS, which was evaluated for neutralizing potency using the PRNT50 assay. K18-hACE2 transgenic mice (n=35) were divided into three groups: control, SARS-CoV-2 exposure through inhalation, and SARS-CoV-2 exposed mice treated with HS. HS efficacy was assessed through serum proinflammatory cytokine levels, qRT-PCR analysis, histopathological examination of lungs and hearts, and transmission electron microscopy. Purified HS exhibited significant neutralizing activity (1/24,576). The SARS-CoV-2+HS group showed lower levels of TNF-α, IL-10, and IL-6 (P<0.01) and relatively lower levels of MCP-1 compared to the SARS-CoV-2 group. HS prevented death, reduced viral RNA levels in the lungs and hearts, protected against severe interstitial pneumonia, preserved lung tissue integrity, and prevented myocyte damage, while the SARS-CoV-2 group exhibited viral presence in the lungs. This study successfully developed a sheep-derived HS against the entire SARS-CoV-2 virus, resulting in a significant reduction in infection severity, inflammation, and systemic cytokine production. The findings hold promise for treating severe COVID-19 cases, including emerging viral variants, and immunocompromised patients.

En respuesta a la amenaza que suponen las nuevas variantes del SARS-CoV-2 y la urgente necesidad de tratamientos eficaces en ausencia de vacunas, el objetivo de este estudio fue desarrollar un suero hiperinmune (HS) rápido y rentable derivado de ovejas. y evaluar su eficacia. La utilización de un animal con certificación halal, de fácil mantenimiento en determinadas regiones geográficas y de fácil manejo, como las ovejas, podría proporcionar una alternativa viable a la costosa opción de los caballos. Las ovejas fueron inmunizadas con un antígeno de SARS-CoV-2 completamente inactivado para producir HS, cuya potencia neutralizante se evaluó mediante el ensayo PRNT50. Los ratones transgénicos K18-hACE2 (n = 35) se dividieron en tres grupos: control, exposición al SARS-CoV-2 mediante inhalación y ratones expuestos al SARS-CoV-2 tratados con HS. La eficacia de HS se evaluó mediante niveles de citoquinas proinflamatorias en suero, análisis qRT-PCR, examen histopatológico de pulmones y corazones y microscopía electrónica de transmisión. El HS purificado exhibió una actividad neutralizante significativa (1/24,576). El grupo SARS-CoV-2+HS mostró niveles más bajos de TNF-α, IL-10 e IL-6 (P<0,01) y niveles relativamente más bajos de MCP-1 en comparación con el grupo SARS-CoV-2. HS evitó la muerte, redujo los niveles de ARN viral en los pulmones y el corazón, protegió contra la neumonía intersticial grave, preservó la integridad del tejido pulmonar y evitó el daño de los miocitos, mientras que el grupo SARS-CoV-2 exhibió presencia viral en los pulmones. Este estudio desarrolló con éxito un HS derivado de ovejas contra todo el virus SARS-CoV-2, lo que resultó en una reducción significativa de la gravedad de la infección, la inflamación y la producción sistémica de citocinas. Los hallazgos son prometedores para el tratamiento de casos graves de COVID- 19, incluidas las variantes virales emergentes y los pacientes inmunocomprometidos.

Structures from intact myofibrils reveal mechanism of thin filament regulation through nebulin.

In skeletal muscle, nebulin stabilizes and regulates the length of thin filaments, but the underlying mechanism remains nebulous. In this work, we used cryo-electron tomography and subtomogram averaging to reveal structures of native nebulin bound to thin filaments within intact sarcomeres. This in situ reconstruction provided high-resolution details of the interaction between nebulin and actin, demonstrating the stabilizing role of nebulin. Myosin bound to the thin filaments exhibited different conformations of the neck domain, highlighting its inherent structural variability in muscle. Unexpectedly, nebulin did not interact with myosin or tropomyosin, but it did interact with a troponin T linker through two potential binding motifs on nebulin, explaining its regulatory role. Our structures support the role of nebulin as a thin filament "molecular ruler" and provide a molecular basis for studying nemaline myopathies.

ROS and iron homeostasis dependent ferroptosis play a vital role in 5-Fluorouracil induced cardiotoxicity in vitro and in vivo.

5-Fluorouracil (5-FU) was a key chemotherapeutic agent in the treatment of different solid tumors. However, cardiotoxicity was included among the therapeutic strategies of 5-FU. The molecular mechanism of cardiotoxicity induced by 5-FU remains unclear. The aim of the study was to investigate whether ferroptosis was involved in 5-FU-induced cardiotoxicity in vivo and in vitro. The in vivo cardiotoxicity model was induced by intraperitoneal injection of 5-FU at the dose of 15, 30, 60 mg/kg for 7 days. Body weight, general condition and plasma enzyme activities of the mice were observed to evaluate heart function. In addition, HE staining, MASSON staining and TEM technology was used. Western-blot analysis were performed to evaluate the protein level of iron transport, iron storage and reactive oxygen species (ROS) of ferroptosis. In H9c2 cardiomyocyte cells, cell viability, generation of ROS, mitochondrial activity and cellular Fe2+ levels were measured. The in vivo results showed that 5-FU significantly impaired cardiac function and structure. The serum creatine kinase (CK) and lactate dehydrogenase (LDH) levels were significantly increased in 5-FU group. HE and MASSON staining showed that 5-FU caused structural injuries. In addition, 5-FU increased the level of ferroptosis markers involving malonaldehyde (MDA) and Fe2+ content. Ferrostatin-1 (Fer-1) was an aromatic amine that specifically binds with lipid ROS and protects cells against lipid peroxidation. Furthermore, 5-FU markedly induced ferroptosis in H9c2 cardiomyocyte cells, which mainly embodied as declined cell vitality, accumulated iron, elevated lipid peroxides. Conversely, inhibition of ferroptosis by Fer-1 completely abolished 5-FU-induced effects. Both in vivo and in vitro experiments indicated that 5-FU increased the expression of ferroptosis, mainly by reducing the expression of glutathione peroxidase 4 (GPX4) and ferritin heavy chain 1 (FTH1), but enhancing the expression of transferrin receptor 1 (TfR1). In conclusion, the present study suggested that ROS and iron homeostasis dependent ferroptosis played a vital role in 5-FU induced cardiotoxicity.

Fasting promotes acute hypoxic adaptation by suppressing mTOR-mediated pathways.

Rapid adaptation to a hypoxic environment is an unanswered question that we are committed to exploring. At present, there is no suitable strategy to achieve rapid hypoxic adaptation. Here, we demonstrate that fasting preconditioning for 72 h reduces tissue injuries and maintains cardiac function, consequently significantly improving the survival rates of rats under extreme hypoxia, and this strategy can be used for rapid hypoxic adaptation. Mechanistically, fasting reduces blood glucose and further suppresses tissue mTOR activity. On the one hand, fasting-induced mTOR inhibition reduces unnecessary ATP consumption and increases ATP reserves under acute hypoxia as a result of decreased protein synthesis and lipogenesis; on the other hand, fasting-induced mTOR inhibition improves mitochondrial oxygen utilization efficiency to ensure ATP production under acute hypoxia, which is due to the significant decrease in ROS generation induced by enhanced mitophagy. Our findings highlight the important role of mTOR in acute hypoxic adaptation, and targeted regulation of mTOR could be a new strategy to improve acute hypoxic tolerance in the body.

Myocardial ultrastructure can augment genetic testing for sporadic dilated cardiomyopathy with initial heart failure.

AIMS: The aim of the present study was to consider whether the ultrastructural features of cardiomyocytes in dilated cardiomyopathy can be used to guide genetic testing. METHODS AND RESULTS: Endomyocardial biopsy and whole-exome sequencing were performed in 32 consecutive sporadic dilated cardiomyopathy patients [51.0 (40.0-64.0) years, 75% men] in initial phases of decompensated heart failure. The predicted pathogenicity of ultrarare (minor allele frequency ≤0.0005), non-synonymous variants was determined using the American College of Medical Genetics guidelines. Focusing on 75 cardiomyopathy-susceptibility and 41 arrhythmia-susceptibility genes, we identified 404 gene variants, of which 15 were considered pathogenic or likely pathogenic in 14 patients (44% of 32). There were five sarcomeric gene variants (29% of 17 variants) found in five patients (16% of 32), involving a variant of MYBPC3 and four variants of TTN. A patient with an MYBPC3 variant showed disorganized sarcomeres, three patients with TTN variants located in the region encoding the A-band domain showed sparse sarcomeres, and a patient with a TTN variant in encoding the I-band domain showed disrupted sarcomeres. The distribution of diffuse myofilament lysis depended on the causal genes; three patients with the same TMEM43 variant had diffuse myofilament lysis near nuclei (P = 0.011), while two patients with different DSP variants had lysis in the peripheral areas of cardiomyocytes (P = 0.033). CONCLUSIONS: Derangement patterns of myofilament and subcellular distribution of myofilament lysis might implicate causal genes. Large-scale studies are required to confirm whether these ultrastructural findings are related to the causative genes.

Ultrastructural aberrations, histological disruption and upregulation of the VEGF, CD34 and ASMA immunoexpression in the myocardium of anemic albino rats.

Iron deficiency anemia (IDA) is a global health problem affecting various body systems and tissues including the cardiovascular system. Several literatures described the associated physiological and clinical changes in the cardiovascular system and heart. However, the associated structural changes were poorly investigated. Therefore, the main aim of the present work was to elucidate whether IDA induces structural changes and alterations in the VEGF, CD34 and ASMA immunoexpression in the myocardium of albino rats. Thirty adult male albino rats were divided into two groups (fifteen rats each); control and anemic. Hematological data for all animals were assessed weekly and statistically analyzed. Three weeks later, animals were sacrificed, and heart specimens were obtained and processed for light and electron microscopy. All hematological parameters showed a statistically significant decrease in the anemic group. Structurally, the anemic group showed markedly degenerated, disrupted and disorganized cardiomyocytes in addition to markedly congested blood vessels, fibroblasts, collagen fibers deposition and perivascular cellular infiltration were noted. Also, positive immunostaining for VEGF, CD34 and ASMA was observed. Ultra-structurally, the myocardium of the anemic group showed disrupted and degenerated myofibrils with degenerated nuclei, perinuclear edema, widened interstitial spaces and marked collagen deposition. Mitochondria markedly increased with abnormal shapes. IDA induced myocardial injury that may propagate to regeneration through activated CD34 progenitor cells and increased VEGF or to degeneration and fibrosis through collagen fibers deposition and enhanced ASMA. So, early diagnosis and treatment of IDA is mandatory to avoid the associated myocardial structural changes.

Early Changes in Rat Heart After High-Dose Irradiation: Implications for Antiarrhythmic Effects of Cardiac Radioablation.

Background Noninvasive cardiac radioablation is employed to treat ventricular arrhythmia. However, myocardial changes leading to early-period antiarrhythmic effects induced by high-dose irradiation are unknown. This study investigated dose-responsive histologic, ultrastructural, and functional changes within 1 month after irradiation in rat heart. Methods and Results Whole hearts of wild-type Lewis rats (N=95) were irradiated with single fraction 20, 25, 30, 40, or 50 Gy and explanted at 1 day or 1, 2, 3, or 4 weeks' postirradiation. Microscopic pathologic changes of cardiac structures by light microscope with immunohistopathologic staining, ultrastructure by electron microscopy, and functional evaluation by ECG and echocardiography were studied. Despite high-dose irradiation, no myocardial necrosis and apoptosis were observed. Intercalated discs were widened and disrupted, forming uneven and twisted junctions between adjacent myocytes. Diffuse vacuolization peaked at 3 weeks, suggesting irradiation dose-responsiveness, which was correlated with interstitial and intracellular edema. CD68 immunostaining accompanying vacuolization suggested mononuclear cell infiltration. These changes were prominent in working myocardium but not cardiac conduction tissue. Intracardiac conduction represented by PR and QTc intervals on ECG was delayed compared with baseline measurements. ST segment was initially depressed and gradually elevated. Ventricular chamber dimensions and function remained intact without pericardial effusion. Conclusions Mononuclear cell-related intracellular and extracellular edema with diffuse vacuolization and intercalated disc widening were observed within 1 month after high-dose irradiation. ECG indicated intracardiac conduction delay with prominent ST-segment changes. These observations suggest that early antiarrhythmic effects after cardiac radioablation result from conduction disturbances and membrane potential alterations without necrosis.

Adverse cardiovascular effects of exposure to cadmium and mercury alone and in combination on the cardiac tissue and aorta of Sprague-Dawley rats.

The aim of this study was to identify cardiovascular effects of relevant concentrations of Cd and Hg alone and in combination as a mixture in water. This was achieved by administering to male Sprague-Dawley rats via gavage 0.62 mg/kg Cd or 1.23 mg/kg Hg, or a combination of 0.62 mg/kg Cd and 1.23 mg/kg Hg in the co-exposure group for 28 days. Concentrations were the rat equivalence dosages of 1,000 times the World Health Organization's limits of 0.003 mg/L and 0.006 mg/L for Cd and Hg, respectively, for water. With termination, blood levels of the metals were increased. For all metal exposed groups, histological evaluation and transmission electron microscopy of the myocardium revealed myofibrillar necrosis, increased fibrosis, vacuole formation and mitochondrial damage. Cd caused the most mitochondrial damage while Hg to a greater degree induced fibrosis. In the aorta, both Cd and Hg also increased collagen deposition adversely altering the morphology of the fenestrated elastic fibers in the tunica media. Co-exposure resulted in increased cardiotoxicity with increased mitochondrial damage, fibrosis and distortion of the aortic wall as a result of increased collagen deposition, as well as altered elastin deposition, fragmentation and interlink formation. These are typical features of oxidative damage that correlates with a phenotype of premature ageing of the CVS that potentially can lead to hypertension and premature cardiac failure.

Hydrogen sulfide ameliorates doxorubicin­induced myocardial fibrosis in rats via the PI3K/AKT/mTOR pathway.

The present study aimed to determine the role and regulatory mechanism of hydrogen sulfide (H2S) in the amelioration of doxorubicin­induced myocardial fibrosis in rats. It is hypothesized that the PI3K/AKT/mTOR signaling pathway is regulated to inhibit endoplasmic reticulum stress (ERS) and autophagy to reduce myocardial fibrosis. A total of 40 adult male Sprague Dawley rats were randomly divided into 4 groups (n=10/group). The 4 groups included the normal control group (control group), model group [doxorubicin (Dox) group], H2S intervention model group (H2S+Dox group) and H2S control group (H2S group). The model used in the present study was constructed by administering intraperitoneal injections of doxorubicin (3.0 mg/kg every other day; total of 6 injections). In addition, the intervention factor, NaHS and the donor of H2S, was also administered by intraperitoneal injection (56 µmol/kg/day), which lasted a month. Pathological changes in the rats were observed using Masson staining and transmission electron microscopy, while the protein expression levels of MMPs/TIMPs, transforming growth factor­ß1, cystathionine lyase and PI3K/AKT/mTOR, which are autophagy­related and ERS­related proteins were detected in myocardial tissues using western blot analysis. The gene expression levels of collagen type I α­2 chain and collagen type III α­1 chain were detected using reverse transcription­quantitative PCR and the quantification of myocardial H2S content was performed using ELISA. In the Dox group compared with that in the control group, myocardial fibers were significantly disordered, while the protein expression levels of ERS­related and autophagy­related proteins were increased markedly, and the expression levels of PI3K/AKT/mTOR proteins were reduced markedly. The aforementioned changes were markedly reversed following H2S intervention, which indicated that H2S exerts a positive protective effect on doxorubicin­induced myocardial fibrosis. The protective mechanism of H2S intervention in myocardial fibrosis is hypothesized to be associated with the inhibition of overactivation of the ER and that of autophagy via upregulation of the PI3K/AKT/mTOR pathway.

Thyroid Hormone Receptor α Mutations Cause Heart Defects in Zebrafish.

Background: Mutations of thyroid hormone receptor α1 (TRα1) cause resistance to thyroid hormone (RTHα). Patients exhibit growth retardation, delayed bone development, anemia, and bradycardia. By using mouse models of RTHα, much has been learned about the molecular actions of TRα1 mutants that underlie these abnormalities in adults. Using zebrafish models of RTHα that we have recently created, we aimed to understand how TRα1 mutants affect the heart function during this period. Methods: In contrast to human and mice, the thra gene is duplicated, thraa and thrab, in zebrafish. Using CRISPR/Cas9-mediated targeted mutagenesis, we created C-terminal mutations in each of two duplicated thra genes in zebrafish (thraa 8-bp insertion or thrab 1-bp insertion mutations). We recently showed that these mutant fish faithfully recapitulated growth retardation as found in patients and thra mutant mice. In the present study, we used histological analysis, gene expression profiles, confocal fluorescence, and transmission electron microscopy (TEM) to comprehensively analyze the phenotypic characteristics of mutant fish heart during development. Results: We found both a dilated atrium and an abnormally shaped ventricle in adult mutant fish. The retention of red blood cells in the two abnormal heart chambers, and the decreased circulating blood speed and reduced expression of contractile genes indicated weakened contractility in the heart of mutant fish. These abnormalities were detected in mutant fish as early as 35 days postfertilization (juveniles). Furthermore, the expression of genes associated with the sarcomere assembly was suppressed in the heart of mutant fish, resulting in abnormalities of sarcomere organization as revealed by TEM, suggesting that the abnormal sarcomere organization could underlie the bradycardia exhibited in mutant fish. Conclusions: Using a zebrafish model of RTHα, the present study demonstrated for the first time that TRα1 mutants could act to cause abnormal heart structure, weaken contractility, and disrupt sarcomere organization that affect heart functions. These findings provide new insights into the bradycardia found in RTHα patients.

High-Resolution Cryo-EM Structure of the Cardiac Actomyosin Complex.

Heart contraction depends on a complicated array of interactions between sarcomeric proteins required to convert chemical energy into mechanical force. Cyclic interactions between actin and myosin molecules, controlled by troponin and tropomyosin, generate the sliding force between the actin-based thin and myosin-based thick filaments. Alterations in this sophisticated system due to missense mutations can lead to cardiovascular diseases. Numerous structural studies proposed pathological mechanisms of missense mutations at the myosin-myosin, actin-tropomyosin, and tropomyosin-troponin interfaces. However, despite the central role of actomyosin interactions a detailed structural description of the cardiac actomyosin interface remained unknown. Here, we report a cryo-EM structure of a cardiac actomyosin complex at 3.8 Å resolution. The structure reveals the molecular basis of cardiac diseases caused by missense mutations in myosin and actin proteins.

Melatonin alleviates sepsis-induced heart injury through activating the Nrf2 pathway and inhibiting the NLRP3 inflammasome.

Melatonin improved the outcome of septic cardiomyopathy by inhibiting NLRP3 priming induced by reactive oxygen species. To get insights into these events, we studied the melatonin/Nrf2 antioxidant pathways during sepsis in the heart of NLRP3-deficient mice. Sepsis was induced by cecal ligation and puncture and melatonin was given at a dose of 30 mg/kg. Nuclear turnover of Nrf2 and p-Ser40 Nrf2 and expression of ho-1 were enhanced in nlrp3+/+ and nlrp3-/- mice during sepsis. Sepsis caused higher mitochondria impairment, apoptotic and autophagic events in nlrp3+/+ mice than in nlrp3-/- animals. These findings were accompanied by greater levels of Parkin and PINK-1, and lower Mfn2/Drp-1 ratio in nlrp3+/+ than in nlrp3-/- mice during sepsis, supporting less mitophagy in the latter. Ultrastructural analysis of myocardial tissue further confirmed these observations. The activation of NLRP3 inflammasome accounted for most of the deleterious effects of sepsis, whereas the Nrf2-dependent antioxidative response activation in response to sepsis was unable to neutralize these events. In turn, melatonin further enhanced the Nrf2 response in both mice strains and reduced the NLRP3 inflammasome activation in nlrp3+/+ mice, restoring myocardial homeostasis. The data support that the anti-inflammatory efficacy of melatonin against sepsis depends, at least in part, on Nrf2 activation.

Chronic cardiac structural damage, diastolic and systolic dysfunction following acute myocardial injury due to bromine exposure in rats.

Accidental bromine spills are common and its large industrial stores risk potential terrorist attacks. The mechanisms of bromine toxicity and effective therapeutic strategies are unknown. Our studies demonstrate that inhaled bromine causes deleterious cardiac manifestations. In this manuscript we describe mechanisms of delayed cardiac effects in the survivors of a single bromine exposure. Rats were exposed to bromine (600 ppm for 45 min) and the survivors were sacrificed at 14 or 28 days. Echocardiography, hemodynamic analysis, histology, transmission electron microscopy (TEM) and biochemical analysis of cardiac tissue were performed to assess functional, structural and molecular effects. Increases in right ventricular (RV) and left ventricular (LV) end-diastolic pressure and LV end-diastolic wall stress with increased LV fibrosis were observed. TEM images demonstrated myofibrillar loss, cytoskeletal breakdown and mitochondrial damage at both time points. Increases in cardiac troponin I (cTnI) and N-terminal pro brain natriuretic peptide (NT-proBNP) reflected myofibrillar damage and increased LV wall stress. LV shortening decreased as a function of increasing LV end-systolic wall stress and was accompanied by increased sarcoendoplasmic reticulum calcium ATPase (SERCA) inactivation and a striking dephosphorylation of phospholamban. NADPH oxidase 2 and protein phosphatase 1 were also increased. Increased circulating eosinophils and myocardial 4-hydroxynonenal content suggested increased oxidative stress as a key contributing factor to these effects. Thus, a continuous oxidative stress-induced chronic myocardial damage along with phospholamban dephosphorylation are critical for bromine-induced chronic cardiac dysfunction. These findings in our preclinical model will educate clinicians and public health personnel and provide important endpoints to evaluate therapies.

Remote ischaemic preconditioning ameliorates anthracycline-induced cardiotoxicity and preserves mitochondrial integrity.

AIMS: Anthracycline-induced cardiotoxicity (AIC) is a serious adverse effect among cancer patients. A central mechanism of AIC is irreversible mitochondrial damage. Despite major efforts, there are currently no effective therapies able to prevent AIC. METHODS AND RESULTS: Forty Large-White pigs were included. In Study 1, 20 pigs were randomized 1:1 to remote ischaemic preconditioning (RIPC, 3 cycles of 5 min leg ischaemia followed by 5 min reperfusion) or no pretreatment. RIPC was performed immediately before each intracoronary doxorubicin injections (0.45 mg/kg) given at Weeks 0, 2, 4, 6, and 8. A group of 10 pigs with no exposure to doxorubicin served as healthy controls. Pigs underwent serial cardiac magnetic resonance (CMR) exams at baseline and at Weeks 6, 8, 12, and 16, being sacrifice after that. In Study 2, 10 new pigs received 3 doxorubicin injections (with/out preceding RIPC) and were sacrificed at week 6. In Study 1, left ventricular ejection fraction (LVEF) depression was blunted animals receiving RIPC before doxorubicin (RIPC-Doxo), which had a significantly higher LVEF at Week 16 than doxorubicin treated pigs that received no pretreatment (Untreated-Doxo) (41.5 ± 9.1% vs. 32.5 ± 8.7%, P = 0.04). It was mainly due to conserved regional contractile function. In Study 2, transmission electron microscopy (TEM) at Week 6 showed fragmented mitochondria with severe morphological abnormalities in Untreated-Doxo pigs, together with upregulation of fission and autophagy proteins. At the end of the 16-week Study 1 protocol, TEM revealed overt mitochondrial fragmentation with structural fragmentation in Untreated-Doxo pigs, whereas interstitial fibrosis was less severe in RIPC+Doxo pigs. CONCLUSION: In a translatable large-animal model of AIC, RIPC applied immediately before each doxorubicin injection resulted in preserved cardiac contractility with significantly higher long-term LVEF and less cardiac fibrosis. RIPC prevented mitochondrial fragmentation and dysregulated autophagy from AIC early stages. RIPC is a promising intervention for testing in clinical trials in AIC.

Coexistence of amyloidosis and light chain deposition disease in the heart.

There are few reports on the coexistence of cardiac amyloid light-chain (AL) amyloidosis and light chain deposition disease (LCDD), despite their similar pathophysiologies caused by plasma-cell dyscrasia. Herein, we report the coexistence of these diseases. A 59-year-old man was referred to our hospital because of exertional dyspnea and hypotension. Renal dysfunction of unknown etiology had been present for 4 years and hemodialysis had been introduced. Severe systolic and diastolic cardiac dysfunction was apparent, accompanied with dilatation and granular sparkling, but not with left ventricular hypertrophy. The plasma-free light chain κ was found to be extremely high, with a κ/λ ratio of 1,919. Light microscopic examination of the endomyocardial biopsy revealed spotty and homogenous deposits, which positively stained with Congo red, and exhibited a blazing apple-green color under polarized light. Based on these results, cardiac amyloidosis was diagnosed. In specimens prepared for electron microscopy, no amyloid fibrils could be found. Instead, we observed amorphous nonfibrillar deposits around several small vessels including capillaries and small arteries, which were consistent with light-chain deposits. LCDD was diagnosed based on the systemic increase in κ light chain and the ultrastructural findings of the endomyocardial biopsy specimens. Coexistence of cardiac amyloidosis and LCDD was thus confirmed in our patient. An electron microscopic assessment in addition to Congo red staining may be useful to diagnose latent LCDD in patients with suspected cardiac light-chain amyloidosis.

Echinacoside reverses myocardial remodeling and improves heart function via regulating SIRT1/FOXO3a/MnSOD axis in HF rats induced by isoproterenol.

Myocardial remodelling is important pathological basis of HF, mitochondrial oxidative stress is a promoter to myocardial hypertrophy, fibrosis and apoptosis. ECH is the major active component of a traditional Chinese medicine Cistanches Herba, plenty of studies indicate it possesses a strong antioxidant capacity in nerve cells and tumour, it inhibits mitochondrial oxidative stress, protects mitochondrial function, but the specific mechanism is unclear. SIRT1/FOXO3a/MnSOD is an important antioxidant axis, study finds that ECH binds covalently to SIRT1 as a ligand and up-regulates the expression of SIRT1 in brain cells. We hypothesizes that ECH may reverse myocardial remodelling and improve heart function of HF via regulating SIRT1/FOXO3a/MnSOD signalling axis and inhibit mitochondrial oxidative stress in cardiomyocytes. Here, we firstly induce cellular model of oxidative stress by ISO with AC-16 cells and pre-treat with ECH, the level of mitochondrial ROS, mtDNA oxidative injury, MMP, carbonylated protein, lipid peroxidation, intracellular ROS and apoptosis are detected, confirm the effect of ECH in mitochondrial oxidative stress and function in vitro. Then, we establish a HF rat model induced by ISO and pre-treat with ECH. Indexes of heart function, myocardial remodelling, mitochondrial oxidative stress and function, expression of SIRT1/FOXO3a/MnSOD signalling axis are measured, the data indicate that ECH improves heart function, inhibits myocardial hypertrophy, fibrosis and apoptosis, increases the expression of SIRT1/FOXO3a/MnSOD signalling axis, reduces the mitochondrial oxidative damages, protects mitochondrial function. We conclude that ECH reverses myocardial remodelling and improves cardiac function via up-regulating SIRT1/FOXO3a/MnSOD axis and inhibiting mitochondrial oxidative stress in HF rats.

Cardioprotective Effect of Tangeretin by Inhibiting PTEN/AKT/mTOR Axis in Experimental Sepsis-Induced Myocardial Dysfunction.

Sepsis aggregates undesirable immune response causing depression of ventricular myocardium and diastolic dysfunction. This present study examined the effect of a plant-derived flavone tangeretin (TG) on autophagy and reduction in myocardial dysfunction. The sepsis was induced by cecum ligation and puncture (CLP) in male Sprague-Dawley rats. Abnormal changes were seen in the heart after the sepsis induction. These abnormalities were analyzed based on the cardiac markers, namely Cardiac myosin light chain-1 (cMLC1) and Cardiac troponin I (cTnl), echocardiography, and plasma parameters, like Lactate dehydrogenase (LDH) and Creatinine kinase (CK). Microanatomy of the heart was studied using hematoxylin and eosin stained histopathological samples of cardiac tissue. Western blot technique was used to detect the nature and extent of protein with the amount of a specific RNA (gene expression) in the cardiac homogenate. Oxidative damage was analyzed using redox marker, reduced glutathione. This study successfully showed that TG attenuated sepsis-induced myocardial dysfunction by inhibiting myocardial autophagy via silencing the Phosphatase and tensin homolog (PTEN) expression and acting on the AKT/mTOR pathway. The present findings supported that TG is a novel cardioprotective therapeutic target for sepsis induced myocardial dysfunction.

Electronic cigarettes cause alteration in cardiac structure and function in diet-induced obese mice.

In spite of the widespread use of electronic cigarettes, also known as e-cigarettes, and the proposed adverse cardiac effects of nicotine, the detrimental effects of e-cigarettes on the heart are not well known. This study examines the detrimental effects of e-cigarettes with nicotine at doses that yield circulating nicotine and cotinine in the ranges similar to the levels found in habitual smokers, and a high fat diet (HFD) on cardiac structure and function in a commonly used model of diet-induced obesity (DIO). C57BL/6J mice on an HFD were exposed to e-cigarette in the presence (2.4% nicotine) or absence (0% nicotine) of nicotine and saline aerosol for 12 weeks. Echocardiographic data demonstrated a decrease in left ventricular (LV) fractional shortening, LV ejection fraction, and velocity of circumferential fiber shortening (VCF) in mice treated with e-cigarette (2.4% nicotine) compared to e-cigarette (0% nicotine) or saline exposed mice. Cardiomyocytes (CMs) of mice treated with e-cigarette (2.4% nicotine) exhibited LV abnormalities, including lipid accumulation (ventricular steatosis), myofibrillar derangement and destruction, and mitochondrial hypertrophy, as revealed by transmission electron microscopy. The detrimental effects of e-cigarettes (2.4% nicotine) on cardiac structure and function was accompanied by increased oxidative stress, plasma free fatty acid levels, CM apoptosis, and inactivation of AMP-activated protein kinase and activation of its downstream target, acetyl-CoA-carboxylase. Our results indicate profound adverse effects of e-cigarettes (2.4% nicotine) on the heart in obese mice and raise questions about the safety of the nicotine e-cigarettes use.

Histopathologic and Ultrastructural Findings in Human Myocardium After Stereotactic Body Radiation Therapy for Recalcitrant Ventricular Tachycardia.

BACKGROUND: Stereotactic body radiation therapy (SBRT) is a novel treatment for refractory ventricular tachycardia (VT). While outcomes have been described in small studies, histological findings after SBRT for VT are unknown. METHODS: We identified 4 explanted hearts in the context of transplant that received prior SBRT as part of an 11-patient compassionate use series at our institution. Clinical VTs and computed tomography-defined target volume areas of SBRT were correlated to the anatomic specimens. Gross pathological, histological, and ultrastructural examination of tissue in the target area of SBRT was performed. RESULTS: All 4 patients had nonischemic cardiomyopathy, and 3 had left ventricular assist devices. In all cases, patients had recurrent sustained VT and had failed multiple antiarrhythmics and radiofrequency ablations. Four patients underwent 5 total SBRT therapy sessions with 25-Gy single-fraction dose delivered to the area of culprit scar. The time from SBRT to explant ranged from 12 to 250 days. Histopathologic features following radiation were comparable in all patients and were characterized by areas of subendocardial necrosis surrounded by a rim of fibrosis. In 1 patient, the surrounding myocardium showed cytoplasmic vacuolization in myocytes and in another patchy interstitial fibrosis. Vascular changes consisted of myointimal thickening with prominence of endothelial cells. Electron microscopy of myocardium showed irregular, convoluted intercalated disc regions, loss of contractile elements with disrupted and haphazardly arranged myofibrils, and edematous mitochondria with loss of cisternae. CONCLUSIONS: Here, we report the first series of findings in human tissue in 4 patients after SBRT. Histopathologic features were consistent across all 4 patients and were indicative of cell injury, death, and to a lesser extent, fibrosis. Electron microscopy demonstrated features consistent with acute injury. These specimens provide radiobiological mechanisms of acute cellular injury during SBRT for VT, which may have an antiarrhythmic effect before the onset of fibrosis.

Regenerative Potential of a Suspension and Spheroids of Multipotent Mesenchymal Stromal Cells from Human Umbilical Cord on the Model of Myocardial Infarction in Rats.

Regenerative potential of multipotent mesenchymal stromal cells from the human umbilical cord (MMSC-UC) in the suspension and spheroid form was revealed during the progression of experimental small focal myocardial infarction in rats. In isoproterenol-induced myocardial infarction, foci of necrosis and inflammatory infiltrate and at later terms fibrosis foci were found mainly in the left ventricle of rat heart. In rats receiving MMSC-UC, destructive changes in the myocardium, fibrous scars, and inflammatory process were less pronounced. MMSC-UC also contributed to normalization of the morphofunctional parameters of the heart. Spheroids exhibited higher efficiency in comparison with cell suspension.