Sodium Tanshinone IIA Sulfonate Protects Primary Cardiomyocytes Against Radiation-Induced Myocardial Injury via the p38 Pathway.

Sodium tanshinone IIA sulfonate (STS), which is extracted from a Chinese medicinal herb, possesses many pharmacologic functions, such as coronary dilation, anti-inflammatory properties, and antiapoptotic and antioxidant effects. It remains unknown whether STS can protect cardiomyocytes injured after radiation therapy. An in vitro Sprague-Dawley (SD) rat neonatal cardiomyocyte system was established. Primary cardiomyocytes (PCMs) from neonatal SD rats were isolated under sterile conditions. PCM cells were divided into a control group (0 Gy/hour) and 5 experimental radiation therapy groups (0.25 Gy/hour, 0.5 Gy/hour, 1 Gy/hour, 2 Gy/hour, and 4 Gy/hour). Cell viability, the content of malondialdehyde (MDA), the lactate dehydrogenase (LDH) leakage rate, and superoxide dismutase (SOD) and glutathione (GSH) activities were recorded separately in each group after 7 days of culture. Western blot was used to detect the levels of p38, caspase-3 protein, and X protein (BAX) associated with B-cell lymphoma 2 (Bcl-2) in PCMs. X-rays inhibited cell growth, decreased cell viability, and induced an oxidative stress response in PCMs. STS and SB203580 (the inhibitor of P38 mitogen-activated protein kinase pathway) alleviated X-ray-induced damage to PCMs. An enzyme-linked immunosorbent assay showed that X-rays increased the cTnT level. STS and SB203580 ameliorated the X-ray-induced increase in cTnT leakage. X-rays enhanced the expression of p38/p-p38 and caspase-3 while reducing the expression of Bcl-2/BAX in PCMs, as demonstrated by western blotting. STS and SB203580 mitigated the changes in protein expression triggered by X-ray radiation. In conclusions, STS was shown to exert significant cardioprotective, anti-inflammatory, and antioxidant effects in PCMs by inhibiting the p38 mitogen-activated protein kinase pathway.

Photobiomodulation improves cell survival and death parameters in cardiomyocytes exposed to hypoxia/reoxygenation.

INTRODUCTION: Cardiovascular diseases are the leading cause of morbidity and mortality worldwide. Ischemic heart disease is one of the most harmful conditions to cellular structure and function. After reperfusion treatment, a spectrum of adverse effects becomes evident, encompassing altered cell viability, heightened oxidative stress, activated autophagy, and increased apoptosis. Photobiomodulation (PBM) has been utilized in experimental models of cardiac hypoxia to enhance mitochondrial response and ameliorate biochemical changes in injured tissue. However, the effects of PBM on cultured cardiomyocytes subjected to hypoxia/reoxygenation are not yet well established. METHOD: H9C2 cardiomyocytes were exposed to hypoxia with concentrations of 300 µM CoCl2 for 24 h, followed by 16 h of reoxygenation through incubation in a normoxic medium. Treatment was conducted using GaAIAs Laser (850 nm) after hypoxia at an intensity of 1 J/cm2. Cells were divided into three groups: Group CT (cells maintained under normoxic conditions), Group HR (cells maintained in hypoxia and reoxygenation conditions without treatment), Group HR + PBM (cells maintained in hypoxia and reoxygenation conditions that underwent PBM treatment). Cell viability was analyzed using MTT, and protein expression was assessed by western blot. One-way ANOVA with the Tukey post hoc test was used for data analysis. Differences were significant when p < 0.05. RESULTS: PBM at an intensity of 1 J/cm2 mitigated the alterations in cell survival caused by hypoxia/reoxygenation. Additionally, it significantly increased the expression of proteins Nrf2, HSP70, mTOR, LC3II, LC3II/I, and Caspase-9, while reducing the expression of PGC-1α, SOD2, xanthine oxidase, Beclin-1, LC3I, and Bax. CONCLUSION: PBM at intensities of 1 J/cm2 reverses the changes related to oxidative stress, mitochondrial biogenesis, autophagy, and apoptosis caused by hypoxia and reoxygenation in a culture of cardiomyocytes.

Total extracts from Abelmoschus manihot (L.) alleviate radiation-induced cardiomyocyte ferroptosis via regulating redox imbalances mediated by the NOX4/xCT/GPX4 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Radiation-induced heart disease (RIHD) is one of the most serious complications in patients receiving chest radiotherapy, partially offsetting its benefits. At present, there is a lack of effective treatments for RIHD. Ferroptosis is a newly discovered type of cell death that results from iron-dependent lipid peroxide accumulation. It was recently shown that irradiation generates severe ferroptosis, providing new insights for the treatment of RIHD. Abelmoschus manihot (L.) possesses excellent pharmacological properties and is widely used in treating various ischemic heart and brain diseases; however, its efficacy and mechanism in treating RIHD are unknown. AIM: This study aimed to investigate the efficacy and mechanism of total extracts from A. manihot (L.) (TEA) in treating RIHD. MATERIALS AND METHODS: C57BL/6 mice and H9C2 cells were exposed to irradiation to induce RIHD in vivo and in vitro, respectively. In vivo, we evaluated the protective effects of TEA (150 and 300 mg/kg) on RIHD. Body and heart weight changes of mice were calculated in each group, and malondialdehyde (MDA) level, glutathione/oxidized glutathione (GSH/GSSH) and nicotinamide adenine dinucleotide phosphate (NADPH/NADP+) ratios, western blot, heart histology, and immunohistochemistry were used to evaluate TEA effectiveness. We identified the potential mechanism of radiation-induced cardiomyocyte injury in H9C2 cells treated with small interfering RNA. We determined the effective dose of TEA (0.6 mg/mL) using a Cell Counting Kit-8 assay. Intracellular Fe2+ and lipid peroxidation levels were detected by Phen Green™ SK diacetate probe, BODIPY 581/591 C11 staining, and MDA, GSH, and NADPH kits, and the level of target protein was evaluated by immunofluorescence and western blot. RESULTS: Radiation inhibited system Xc-cystine (xCT)/glutathione peroxidase 4 (GPX4) expression and activity in cardiomyocytes in a time and dose-dependent manner. After silencing xCT/GPX4, MDA significantly increased and GSH/GSSH and NADPH/NADP+ ratios were reduced. xCT/GPX4 inhibition drove ferroptosis in radiation-induced H9C2 injury. Oxidative stress in H9C2 was significantly enhanced by irradiation, which also significantly increased NADPH oxidase 4 (NOX4) expression and inhibited nuclear factor E2-related factor 2 (Nrf2) expression in vivo and in vitro. Inhibition of xCT/GPX4 drove ferroptosis in radiation-induced H9C2 injury, which was aggravated by inactivation of Nrf2 and alleviated by inhibition of NOX4. Compared with the ionizing radiation-only group, TEA improved body weight loss, MDA levels, and histological changes induced by irradiation in mice hearts, and increased the ratio of GSH/GSSH and NADPH/NADP+in vivo; it also reduced lipid peroxidation and intracellular Fe2+ accumulation, restored MDA levels, and elevated the ratios of GSH/GSSH and NADPH/NADP+ in irradiation-injured H9C2 cells. TEA up-regulated Nrf2, xCT, and GPX4 expression and inhibited NOX4 expression in vivo and in vitro. CONCLUSIONS: Ferroptosis induced by redox imbalance mediated through the NOX4/xCT/GPX4 axis is a potential mechanism behind radiation-induced cardiomyocyte injury, and can be prevented by TEA.

Study and optimization of the photobiomodulation effects induced on mitochondrial metabolic activity of human cardiomyocytes for different radiometric and spectral conditions.

Photobiomodulation (PBM) represents a promising and powerful approach for non-invasive therapeutic interventions. This emerging field of research has gained a considerable attention due to its potential for multiple disciplines, including medicine, neuroscience, and sports medicine. While PBM has shown the ability to stimulate various cellular processes in numerous medical applications, the fine-tuning of treatment parameters, such as wavelength, irradiance, treatment duration, and illumination geometry, remains an ongoing challenge. Furthermore, additional research is necessary to unveil the specific mechanisms of action and establish standardized protocols for diverse clinical applications. Given the widely accepted understanding that mitochondria play a pivotal role in the PBM mechanisms, our study delves into a multitude of PBM illumination parameters while assessing the PBM's effects on the basis of endpoints reflecting the mitochondrial metabolism of human cardiac myocytes (HCM), that are known for their high mitochondrial density. These endpoints include: i) the endogenous production of protoporphyrin IX (PpIX), ii) changes in mitochondrial potential monitored by Rhodamine 123 (Rhod 123), iii) changes in the HCM's oxygen consumption, iv) the fluorescence lifetime of Rhod 123 in mitochondria, and v) alterations of the mitochondrial morphology. The good correlation observed between these different methods to assess PBM effects underscores that monitoring the endogenous PpIX production offers interesting indirect insights into the mitochondrial metabolic activity. This conclusion is important since many approved therapeutics and cancer detection approaches are based on the use of PpIX. Finally, this correlation strongly suggests that the PBM effects mentioned above have a common "fundamental" mechanistic origin.

Dexmedetomidine ameliorates x-ray-induced myocardial injury via alleviating cardiomyocyte apoptosis and autophagy.

BACKGROUND: Radiotherapy is a primary local treatment for tumors, yet it may lead to complications such as radiation-induced heart disease (RIHD). Currently, there is no standardized approach for preventing RIHD. Dexmedetomidine (Dex) is reported to have cardio-protection effects, while its role in radiation-induced myocardial injury is unknown. In the current study, we aimed to evaluate the radioprotective effect of dexmedetomidine in X-ray radiation-treated mice. METHODS: 18 male mice were randomized into 3 groups: control, 16 Gy, and 16 Gy + Dex. The 16 Gy group received a single dose of 16 Gy X-ray radiation. The 16 Gy + Dex group was pretreated with dexmedetomidine (30 µg/kg, intraperitoneal injection) 30 min before X-ray radiation. The control group was treated with saline and did not receive X-ray radiation. Myocardial tissues were collected 16 weeks after X-ray radiation. Hematoxylin-eosin staining was performed for histopathological examination. Terminal deoxynucleotidyl transferase dUTP nick-end labeling staining was performed to assess the state of apoptotic cells. Immunohistochemistry staining was performed to examine the expression of CD34 molecule and von Willebrand factor. Besides, western blot assay was employed for the detection of apoptosis-related proteins (BCL2 apoptosis regulator and BCL2-associated X) as well as autophagy-related proteins (microtubule-associated protein 1 light chain 3, beclin 1, and sequestosome 1). RESULTS: The findings demonstrated that 16 Gy X-ray radiation resulted in significant changes in myocardial tissues, increased myocardial apoptosis, and activated autophagy. Pretreatment with dexmedetomidine significantly protects mice against 16 Gy X-ray radiation-induced myocardial injury by inhibiting apoptosis and autophagy. CONCLUSION: In summary, our study confirmed the radioprotective effect of dexmedetomidine in mitigating cardiomyocyte apoptosis and autophagy induced by 16 Gy X-ray radiation.

The Proteoglycans Biglycan and Decorin Protect Cardiac Cells against Irradiation-Induced Cell Death by Inhibiting Apoptosis.

Radiation-induced heart disease (RIHD), a common side effect of chest irradiation, is a primary cause of mortality among patients surviving thoracic cancer. Thus, the development of novel, clinically applicable cardioprotective agents which can alleviate the harmful effects of irradiation on the heart is of great importance in the field of experimental oncocardiology. Biglycan and decorin are structurally related small leucine-rich proteoglycans which have been reported to exert cardioprotective properties in certain cardiovascular pathologies. Therefore, in the present study we aimed to examine if biglycan or decorin can reduce radiation-induced damage of cardiomyocytes. A single dose of 10 Gray irradiation was applied to induce radiation-induced cell damage in H9c2 cardiomyoblasts, followed by treatment with either biglycan or decorin at various concentrations. Measurement of cell viability revealed that both proteoglycans improved the survival of cardiac cells post-irradiation. The cardiocytoprotective effect of both biglycan and decorin involved the alleviation of radiation-induced proapoptotic mechanisms by retaining the progression of apoptotic membrane blebbing and lowering the number of apoptotic cell nuclei and DNA double-strand breaks. Our findings provide evidence that these natural proteoglycans may exert protection against radiation-induced damage of cardiac cells.

Ultrasonic-Assisted Enzymolysis Extraction and Protective Effect on Injured Cardiomyocytes in Mice of Flavonoids from Prunus mume Blossom.

Prunus mume blossom is an edible flower that has been used in traditional Chinese medicine for thousands of years. Flavonoids are one of the most active substances in Prunus mume blossoms. The optimal ultrasonic-assisted enzymatic extraction of flavonoids from Prunus mume blossom (FPMB), the components of FPMB, and its protective effect on injured cardiomyocytes were investigated in this study. According to our results, the optimal extraction process for FPMB is as follows: cellulase at 2.0%, ultrasonic power at 300 W, ultrasonic enzymolysis for 30 min, and an enzymolysis temperature of 40 °C. FPMB significantly promoted the survival rate of cardiomyocytes and reduced the concentration of reactive oxygen species (ROS). FPMB also improved the activities of proteases caspase-3, caspase-8, and caspase-9 in cardiomyocytes. The cardiomyocyte apoptosis rate in mice was significantly reduced by exposure to FPMB. These results suggest that the extraction rate of FPMB may be improved by an ultrasonic-assisted enzymatic method. FPMB has a protective effect on the injured cardiomyocytes.

Evaluation of laser induced sarcomere micro-damage: Role of damage extent and location in cardiomyocytes.

Whereas it is evident that a well aligned and regular sarcomeric structure in cardiomyocytes is vital for heart function, considerably less is known about the contribution of individual elements to the mechanics of the entire cell. For instance, it is unclear whether altered Z-disc elements are the reason or the outcome of related cardiomyopathies. Therefore, it is crucial to gain more insight into this cellular organization. This study utilizes femtosecond laser-based nanosurgery to better understand sarcomeres and their repair upon damage. We investigated the influence of the extent and the location of the Z-disc damage. A single, three, five or ten Z-disc ablations were performed in neonatal rat cardiomyocytes. We employed image-based analysis using a self-written software together with different already published algorithms. We observed that cardiomyocyte survival associated with the damage extent, but not with the cell area or the total number of Z-discs per cell. The cell survival is independent of the damage position and can be compensated. However, the sarcomere alignment/orientation is changing over time after ablation. The contraction time is also independent of the extent of damage for the tested parameters. Additionally, we observed shortening rates between 6-7% of the initial sarcomere length in laser treated cardiomyocytes. This rate is an important indicator for force generation in myocytes. In conclusion, femtosecond laser-based nanosurgery together with image-based sarcomere tracking is a powerful tool to better understand the Z-disc complex and its force propagation function and role in cellular mechanisms.

An antibody against L1 cell adhesion molecule inhibits cardiotoxicity by regulating persistent DNA damage.

Targeting the molecular pathways underlying the cardiotoxicity associated with thoracic irradiation and doxorubicin (Dox) could reduce the morbidity and mortality associated with these anticancer treatments. Here, we find that vascular endothelial cells (ECs) with persistent DNA damage induced by irradiation and Dox treatment exhibit a fibrotic phenotype (endothelial-mesenchymal transition, EndMT) correlating with the colocalization of L1CAM and persistent DNA damage foci. We demonstrate that treatment with the anti-L1CAM antibody Ab417 decreases L1CAM overexpression and nuclear translocation and persistent DNA damage foci. We show that in whole-heart-irradiated mice, EC-specific p53 deletion increases vascular fibrosis and the colocalization of L1CAM and DNA damage foci, while Ab417 attenuates these effects. We also demonstrate that Ab417 prevents cardiac dysfunction-related decrease in fractional shortening and prolongs survival after whole-heart irradiation or Dox treatment. We show that cardiomyopathy patient-derived cardiovascular ECs with persistent DNA damage show upregulated L1CAM and EndMT, indicating clinical applicability of Ab417. We conclude that controlling vascular DNA damage by inhibiting nuclear L1CAM translocation might effectively prevent anticancer therapy-associated cardiotoxicity.

Cardiac Shock Wave Therapy Alleviates Hypoxia/Reoxygenation-Induced Myocardial Necroptosis by Modulating Autophagy.

Regulated necrosis (necroptosis) is crucially involved in cardiac ischaemia-reperfusion injury (MIRI). The aim of our study is to investigate whether shock wave therapy (SWT) is capable of exerting protective effects by inhibiting necroptosis during myocardial ischaemia-reperfusion (I/R) injury and the possible role of autophagy in this process. We established a hypoxia/reoxygenation (H/R) model in vitro using HL-1 cells to simulate MIRI. MTS assays and LDH cytotoxicity assay were performed to measure cell viability and cell damage. Annexin V/PI staining was used to determine apoptosis and necrosis. Western blotting was performed to assess the changes in cell signaling pathways associated with autophagy, necroptosis, and apoptosis. Reactive oxygen species (ROS) production was detected using DHE staining. Autophagosome generation and degradation (autophagic flux) were analysed using GFP and RFP tandemly tagged LC3 (tfLC3). HL-1 cells were then transfected with p62/SQSTM1 siRNA in order to analyse its role in cardioprotection. Our results revealed that SWT increased cell viability in the H/R model and decreased receptor-interacting serine/threonine-protein kinase 1 (RIPK1) and RIPK3 expression. ROS production was also inhibited by SWT. Moreover, SWT decreased Beclin1 expression and the ratio of LC3-II/LC3-I following H/R. Simultaneously, in the tfLC3 assay, the SWT provoked a decrease in the cumulative autophagosome abundance. siRNA-mediated knockdown of p62 attenuated H/R-induced necroptosis, and SWT did not exert additive effects. Taken together, SWT ameliorated H/R injury by inhibiting necroptosis. SWT also relieved the blockade of autophagic flux in response to H/R injury. The restoration of autophagic flux by SWT might contribute to its cardioprotective effect on necroptosis following H/R injury.

Impact of High-Dose Irradiation on Human iPSC-Derived Cardiomyocytes Using Multi-Electrode Arrays: Implications for the Antiarrhythmic Effects of Cardiac Radioablation.

Cardiac radioablation is emerging as an alternative option for refractory ventricular arrhythmias. However, the immediate acute effect of high-dose irradiation on human cardiomyocytes remains poorly known. We measured the electrical activities of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) upon irradiation with 0, 20, 25, 30, 40, and 50 Gy using a multi-electrode array, and cardiomyocyte function gene levels were evaluated. iPSC-CMs showed to recover their electrophysiological activities (total active electrode, spike amplitude and slope, and corrected field potential duration) within 3-6 h from the acute effects of high-dose irradiation. The beat rate immediately increased until 3 h after irradiation, but it steadily decreased afterward. Conduction velocity slowed in cells irradiated with ≥25 Gy until 6-12 h and recovered within 24 h; notably, 20 and 25 Gy-treated groups showed subsequent continuous increase. At day 7 post-irradiation, except for cTnT, cardiomyocyte function gene levels increased with increasing irradiation dose, but uniquely peaked at 25-30 Gy. Altogether, high-dose irradiation immediately and reversibly modifies the electrical conduction of cardiomyocytes. Thus, compensatory mechanisms at the cellular level may be activated after the high-dose irradiation acute effects, thereby, contributing to the immediate antiarrhythmic outcome of cardiac radioablation for refractory ventricular arrhythmias.

Sodium tanshinone IIA sulfonate prevents radiation-induced damage in primary rat cardiac fibroblasts.

This study investigated the effects of X-ray irradiation on primary rat cardiac fibroblasts (CFs) and its potential mechanism, as well as whether sodium tanshinone IIA sulfonate (STS) has protective effect on CFs and its possible mechanism. Our data demonstrated that X-rays inhibited cell growth and increased oxidative stress in CFs, and STS mitigated X-ray-induced injury. Enzyme-linked immuno-sorbent assay showed that X-rays increased the levels of secreted angiotensin II (Ang II) and brain natriuretic peptide (BNP). STS inhibited the X-ray-induced increases in Ang II and BNP release. Apoptosis and cell cycle of CFs were analyzed using flow cytometry. X-rays induced apoptosis in CFs, whereas STS inhibited apoptosis in CFs after X-ray irradiation. X-rays induced S-phase cell cycle arrest in CFs, which could be reversed by STS. X-rays increased the expression of phosphorylated-P38/P38, cleaved caspase-3 and caspase-3 as well as decreased the expression of phosphorylated extracellular signal-regulated kinase 1/2 (ERK 1/2)/ERK 1/2 and B cell lymphoma 2 (Bcl-2)/Bcl-2 associated X protein (BAX) in CFs, as shown by Western blotting. STS mitigated the X-ray radiation-induced expression changes of these proteins. In conclusion, our results demonstrated that STS may potentially be developed as a medical countermeasure to mitigate radiation-induced cardiac damage.

The circadian clock protects against ionizing radiation-induced cardiotoxicity.

Radiation therapy (RT) is commonly used to treat solid tumors of the breast, lung, and esophagus; however, the heart is an unintentional target of ionizing radiation (IR). IR exposure to the heart results in chronic toxicities including heart failure. We hypothesize that the circadian system plays regulatory roles in minimizing the IR-induced cardiotoxicity. We treated mice in control (Day Shift), environmentally disrupted (Rotating Shift), and genetically disrupted (Per 1/2 mutant) circadian conditions with 18 Gy of IR to the heart. Compared to control mice, circadian clock disruption significantly exacerbated post-IR systolic dysfunction (by ultrasound echocardiography) and increased fibrosis in mice. At the cellular level, Bmal1 protein bound to Atm, Brca1, and Brca2 promoter regions and its expression level was inversely correlated with the DNA damage levels based on the state of the clock. Further studies with circadian synchronized cardiomyocytes revealed that Bmal1 depletion increased the IR-induced DNA damage and apoptosis. Collectively, these findings suggest that the circadian clock protects from IR-induced toxicity and potentially impacts RT treatment outcome in cancer patients through IR-induced DNA damage responses.

Cardiac pacing using transmural multi-LED probes in channelrhodopsin-expressing mouse hearts.

Optogenetics enables cell-type specific monitoring and actuation via light-activated proteins. In cardiac research, expressing light-activated depolarising ion channels in cardiomyocytes allows optical pacing and defibrillation. Previous studies largely relied on epicardial illumination. Light penetration through the myocardium is however problematic when moving to larger animals and humans. To overcome this limitation, we assessed the utility of an implantable multi light-emitting diode (LED) optical probe (IMLOP) for intramural pacing of mouse hearts expressing cardiac-specific channelrhodopsin-2 (ChR2). Here we demonstrated that IMLOP insertion needs approximately 20 mN of force, limiting possible damage from excessive loads applied during implantation. Histological sections confirmed the confined nature of tissue damage during acute use. The temperature change of the surrounding tissue was below 1 K during LED operation, rendering the probe safe for use in situ. This was confirmed in control experiments where no effect on cardiac action potential conduction was observed even when using stimulation parameters twenty-fold greater than required for pacing. In situ experiments on ChR2-expressing mouse hearts demonstrated that optical stimulation is possible with light intensities as low as 700 µW/mm2; although stable pacing requires higher intensities. When pacing with a single LED, rheobase and chronaxie values were 13.3 mW/mm2 ± 0.9 mW/mm2 and 3 ms ± 0.6 ms, respectively. When doubling the stimulated volume the rheobase decreased significantly (6.5 mW/mm2 ± 0.9 mW/mm2). We have demonstrated IMLOP-based intramural optical pacing of the heart. Probes cause locally constrained tissue damage in the acute setting and require low light intensities for pacing. Further development is necessary to assess effects of chronic implantation.

Hyperacetylation of Cardiac Mitochondrial Proteins Is Associated with Metabolic Impairment and Sirtuin Downregulation after Chronic Total Body Irradiation of ApoE -/- Mice.

Chronic exposure to low-dose ionizing radiation is associated with an increased risk of cardiovascular disease. Alteration in energy metabolism has been suggested to contribute to radiation-induced heart pathology, mitochondrial dysfunction being a hallmark of this disease. The goal of this study was to investigate the regulatory role of acetylation in heart mitochondria in the long-term response to chronic radiation. ApoE-deficient C57Bl/6J mice were exposed to low-dose-rate (20 mGy/day) gamma radiation for 300 days, resulting in a cumulative total body dose of 6.0 Gy. Heart mitochondria were isolated and analyzed using quantitative proteomics. Radiation-induced proteome and acetylome alterations were further validated using immunoblotting, enzyme activity assays, and ELISA. In total, 71 proteins showed peptides with a changed acetylation status following irradiation. The great majority (94%) of the hyperacetylated proteins were involved in the TCA cycle, fatty acid oxidation, oxidative stress response and sirtuin pathway. The elevated acetylation patterns coincided with reduced activity of mitochondrial sirtuins, increased the level of Acetyl-CoA, and were accompanied by inactivation of major cardiac metabolic regulators PGC-1 alpha and PPAR alpha. These observations suggest that the changes in mitochondrial acetylation after irradiation is associated with impairment of heart metabolism. We propose a novel mechanism involved in the development of late cardiac damage following chronic irradiation.

Modulation of cardiac optogenetics by vitamin A.

Cardiac optogenetics is an emergent research area and refers to the delivery of light-activated proteins to excitable heart tissue and the subsequent use of light for controlling the electrical function with high spatial and temporal resolution. Channelrhodopsin-2 (ChR2) is a light-sensitive ion channel with the chromophore, all trans retinal, derived from vitamin A (all-trans-retinol; retinol). In this study, we explored whether exogenous vitamin A can be a limiting factor in the light responsiveness of cardiomyocytes-expressing ChR2. We showed that in cardiomyocytes virally transduced with ChR2 (H134R)-enhanced yellow fluorescent protein, vitamin A supplements lower than 10 µM significantly increased ChR2 expression. Adding 1 µM vitamin A changed light-induced transmembrane potential difference significantly, whereas 5 µM dramatically induced membrane depolarization and triggered intracellular calcium elevation. We concluded that vitamin A supplementation can modulate the efficiency of ChR2 and provide a complementary strategy for improving the performance of optogenetic tools.

Optical Pacing of Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Mediated by a Conjugated Polymer Interface.

The use of light for triggering skeletal and cardiac muscles allows lower invasiveness higher selectivity and unprecedented possibility to target individual cells or even subcellular compartments in a temporally and spatially precise manner. Because cells are in general transparent, this requires the development of suitable interfaces that bestow light sensitivity to living matter. In the present work, successfully demonstrated is the use of conjugated polymer films as transducer to optically enhance the contraction rate of a human and patient-specific cardiac in vitro cell model. By different experimental approaches, the coupling mechanism to the photothermal effect is assigned. This work extends the range of application of the polymer-mediated cell photostimulation phenomenon to cardiac muscle cells, opening up possible applications in cardiac therapy and for implementation of in vitro studies.

Biological Cardiac Tissue Effects of High-Energy Heavy Ions - Investigation for Myocardial Ablation.

Noninvasive X-ray stereotactic treatment is considered a promising alternative to catheter ablation in patients affected by severe heart arrhythmia. High-energy heavy ions can deliver high radiation doses in small targets with reduced damage to the normal tissue compared to conventional X-rays. For this reason, charged particle therapy, widely used in oncology, can be a powerful tool for radiosurgery in cardiac diseases. We have recently performed a feasibility study in a swine model using high doses of high-energy C-ions to target specific cardiac structures. Interruption of cardiac conduction was observed in some animals. Here we report the biological effects measured in the pig heart tissue of the same animals six months after the treatment. Immunohistological analysis of the target tissue showed (1.) long-lasting vascular damage, i.e. persistent hemorrhage, loss of microvessels, and occurrence of siderophages, (2.) fibrosis and (3.) loss of polarity of targeted cardiomyocytes and wavy fibers with vacuolization. We conclude that the observed physiological changes in heart function are produced by radiation-induced fibrosis and cardiomyocyte functional inactivation. No effects were observed in the normal tissue traversed by the particle beam, suggesting that charged particles have the potential to produce ablation of specific heart targets with minimal side effects.

Optogenetic stimulation of Gs-signaling in the heart with high spatio-temporal precision.

The standard technique for investigating adrenergic effects on heart function is perfusion with pharmaceutical agonists, which does not provide high temporal or spatial precision. Herein we demonstrate that the light sensitive Gs-protein coupled receptor JellyOp enables optogenetic stimulation of Gs-signaling in cardiomyocytes and the whole heart. Illumination of transgenic embryonic stem cell-derived cardiomyocytes or of the right atrium of mice expressing JellyOp elevates cAMP levels and instantaneously accelerates spontaneous beating rates similar to pharmacological ß-adrenergic stimulation. Light application to the dorsal left atrium instead leads to supraventricular extrabeats, indicating adverse effects of localized Gs-signaling. In isolated ventricular cardiomyocytes from JellyOp mice, we find increased Ca2+ currents, fractional cell shortening and relaxation rates after illumination enabling the analysis of differential Gs-signaling with high temporal precision. Thus, JellyOp expression allows localized and time-restricted Gs stimulation and will provide mechanistic insights into different effects of site-specific, long-lasting and pulsatile Gs activation.

GeneLab Database Analyses Suggest Long-Term Impact of Space Radiation on the Cardiovascular System by the Activation of FYN Through Reactive Oxygen Species.

Space radiation has recently been considered a risk factor for astronauts' cardiac health. As an example, for the case of how to query and identify datasets within NASA's GeneLab database and demonstrate the database utility, we used an unbiased systems biology method for identifying key genes/drivers for the contribution of space radiation on the cardiovascular system. This knowledge can contribute to designing appropriate experiments targeting these specific pathways. Microarray data from cardiomyocytes of male C57BL/6 mice followed-up for 28 days after exposure to 900 mGy of 1 GeV proton or 150 mGy of 1 GeV/n 56Fe were compared to human endothelial cells (HUVECs) cultured for 7 days on the International Space Station (ISS). We observed common molecular pathways between simulated space radiation and HUVECs flown on the ISS. The analysis suggests FYN is the central driver/hub for the cardiovascular response to space radiation: the known oxidative stress induced immediately following radiation would only be transient and would upregulate FYN, which in turn would reduce reactive oxygen species (ROS) levels, protecting the cardiovascular system. The transcriptomic signature of exposure to protons was also much closer to the spaceflight signature than 56Fe's signature. To our knowledge, this is the first time GeneLab datasets were utilized to provide potential biological indications that the majority of ions on the ISS are protons, clearly illustrating the power of omics analysis. More generally, this work also demonstrates how to combine animal radiation studies done on the ground and spaceflight studies to evaluate human risk in space.