The Clot Thickens: Differential Coagulotoxic and Cardiotoxic Activities of Anguimorpha Lizard Venoms.

Despite their evolutionary novelty, lizard venoms are much less studied in comparison to the intense research on snake venoms. While the venoms of helodermatid lizards have long been assumed to be for defensive purposes, there is increasing evidence of toxic activities more useful for predation than defence (such as paralytic neurotoxicity). This study aimed to ascertain the effects of Heloderma, Lanthanotus, and Varanus lizard venoms on the coagulation and cardiovascular systems. Anticoagulant toxicity was demonstrated for the Varanus species studied, with the venoms prolonging clotting times in human and bird plasma due to the destructive cleavage of fibrinogen. In contrast, thromboelastographic analyses on human and bird plasmas in this study demonstrated a procoagulant bioactivity for Heloderma venoms. A previous study on Heloderma venom using factor-depleted plasmas as a proxy model suggested a procoagulant factor was present that activated either Factor XI or Factor XII, but could not ascertain the precise target. Our activation studies using purified zymogens confirmed FXII activation. Comparisons of neonate and adult H. exasperatum, revealed the neonates to be more potent in the ability to activate FXII, being more similar to the venom of the smaller species H. suspectum than the adult H. exasperatum. This suggests potent FXII activation a basal trait in the genus, present in the small bodied last common ancestor. This also indicates an ontogenetic difference in prey preferences in the larger Heloderma species paralleing the change in venom biochemistry. In addition, as birds lack Factor XII, the ability to clot avian plasma suggested an additional procoagulant site of action, which was revealed to be the activation of Factor VII, with H. horridum being the most potent. This study also examined the effects upon the cardiovascular system, including the liberation of kinins from kininogen, which contributes to hypotension induction. This form of toxicity was previously described for Heloderma venoms, and was revealed in this study was to also be a pathophysiological effect of Lanthanotus and Varanus venoms. This suggests that this toxic activity was present in the venom of the last common ancestor of the anguimorph lizards, which is consistent with kallikrein enzymes being a shared toxin trait. This study therefore uncovered novel actions of anguimorph lizard venoms, not only contributing to the evolutionary biology body of knowledge but also revealing novel activities to mine for drug design lead compounds.

Changes in protein fluxes in skeletal muscle during sequential stages of muscle regeneration after acute injury in male mice.

Changes in protein turnover play an important role in dynamic physiological processes, including skeletal muscle regeneration, which occurs as an essential part of tissue repair after injury. The inability of muscle tissue to recapitulate this regenerative process can lead to the manifestation of clinical symptoms in various musculoskeletal diseases, including muscular dystrophies and pathological atrophy. Here, we employed a workflow that couples deuterated water (2H2O) administration with mass spectrometry (MS) to systematically measure in-vivo protein turnover rates across the muscle proteome in 8-week-old male C57BL6/J mice. We compared the turnover kinetics of over 100 proteins in response to cardiotoxin (CTX) induced muscle damage and regeneration at unique sequential stages along the regeneration timeline. This analysis is compared to gene expression data from mRNA-sequencing (mRNA-seq) from the same tissue. The data reveals quantitative protein flux signatures in response to necrotic damage, in addition to sequential differences in cell proliferation, energy metabolism, and contractile gene expression. Interestingly, the mRNA changes correlated poorly with changes in protein synthesis rates, consistent with post-transcriptional control mechanisms. In summary, the experiments described here reveal the signatures and timing of protein flux changes during skeletal muscle regeneration, as well as the inability of mRNA expression measurements to reveal changes in directly measured protein turnover rates. The results of this work described here provide a better understanding of the muscle regeneration process and could help to identify potential biomarkers or therapeutic targets.

Computational approaches identify a transcriptomic fingerprint of drug-induced structural cardiotoxicity.

Structural cardiotoxicity (SCT) presents a high-impact risk that is poorly tolerated in drug discovery unless significant benefit is anticipated. Therefore, we aimed to improve the mechanistic understanding of SCT. First, we combined machine learning methods with a modified calcium transient assay in human-induced pluripotent stem cell-derived cardiomyocytes to identify nine parameters that could predict SCT. Next, we applied transcriptomic profiling to human cardiac microtissues exposed to structural and non-structural cardiotoxins. Fifty-two genes expressed across the three main cell types in the heart (cardiomyocytes, endothelial cells, and fibroblasts) were prioritised in differential expression and network clustering analyses and could be linked to known mechanisms of SCT. This transcriptomic fingerprint may prove useful for generating strategies to mitigate SCT risk in early drug discovery.

CardioDPi: An explainable deep-learning model for identifying cardiotoxic chemicals targeting hERG, Cav1.2, and Nav1.5 channels.

The cardiotoxic effects of various pollutants have been a growing concern in environmental and material science. These effects encompass arrhythmias, myocardial injury, cardiac insufficiency, and pericardial inflammation. Compounds such as organic solvents and air pollutants disrupt the potassium, sodium, and calcium ion channels cardiac cell membranes, leading to the dysregulation of cardiac function. However, current cardiotoxicity models have disadvantages of incomplete data, ion channels, interpretability issues, and inability of toxic structure visualization. Herein, an interpretable deep-learning model known as CardioDPi was developed, which is capable of discriminating cardiotoxicity induced by the human Ether-à-go-go-related gene (hERG) channel, sodium channel (Na_v1.5), and calcium channel (Ca_v1.5) blockade. External validation yielded promising area under the ROC curve (AUC) values of 0.89, 0.89, and 0.94 for the hERG, Na_v1.5, and Ca_v1.5 channels, respectively. The CardioDPi can be freely accessed on the web server CardioDPipredictor (http://cardiodpi.sapredictor.cn/). Furthermore, the structural characteristics of cardiotoxic compounds were analyzed and structural alerts (SAs) can be extracted using the user-friendly CardioDPi-SAdetector web service (http://cardiosa.sapredictor.cn/). CardioDPi is a valuable tool for identifying cardiotoxic chemicals that are environmental and health risks. Moreover, the SA system provides essential insights for mode-of-action studies concerning cardiotoxic compounds.

Loss of adenosine A3 receptors accelerates skeletal muscle regeneration in mice following cardiotoxin-induced injury.

Skeletal muscle regeneration is a complex process orchestrated by multiple interacting steps. An increasing number of reports indicate that inflammatory responses play a central role in linking initial muscle injury responses to timely muscle regeneration following injury. The nucleoside adenosine has been known for a long time as an endogenously produced anti-inflammatory molecule that is generated in high amounts during tissue injury. It mediates its physiological effects via four types of adenosine receptors. From these, adenosine A3 receptors (A3Rs) are not expressed by the skeletal muscle but are present on the surface of various inflammatory cells. In the present paper, the effect of the loss of A3Rs was investigated on the regeneration of the tibialis anterior (TA) muscle in mice following cardiotoxin-induced injury. Here we report that regeneration of the skeletal muscle from A3R-/- mice is characterized by a stronger initial inflammatory response resulting in a larger number of transmigrating inflammatory cells to the injury site, faster clearance of cell debris, enhanced proliferation and faster differentiation of the satellite cells (the muscle stem cells), and increased fusion of the generated myoblasts. This leads to accelerated skeletal muscle tissue repair and the formation of larger myofibers. Though the infiltrating immune cells expressed A3Rs and showed an increased inflammatory profile in the injured A3R-/- muscles, bone marrow transplantation experiments revealed that the increased response of the tissue-resident cells to tissue injury is responsible for the observed phenomenon. Altogether our data indicate that A3Rs are negative regulators of injury-related regenerative inflammation and consequently also that of the muscle fiber growth in the TA muscle. Thus, inhibiting A3Rs might have a therapeutic value during skeletal muscle regeneration following injury.

Intramuscular Bleeding and Formation of Microthrombi during Skeletal Muscle Damage Caused by a Snake Venom Metalloprotease and a Cardiotoxin.

The interactions between specific snake venom toxins and muscle constituents are the major cause of severe muscle damage that often result in amputations and subsequent socioeconomic ramifications for snakebite victims and/or their families. Therefore, improving our understanding of venom-induced muscle damage and determining the underlying mechanisms of muscle degeneration/regeneration following snakebites is critical to developing better strategies to tackle this issue. Here, we analysed intramuscular bleeding and thrombosis in muscle injuries induced by two different snake venom toxins (CAMP-Crotalus atrox metalloprotease (a PIII metalloprotease from the venom of this snake) and a three-finger toxin (CTX, a cardiotoxin from the venom of Naja pallida)). Classically, these toxins represent diverse scenarios characterised by persistent muscle damage (CAMP) and successful regeneration (CTX) following acute damage, as normally observed in envenomation by most vipers and some elapid snakes of Asian, Australasian, and African origin, respectively. Our immunohistochemical analysis confirmed that both CAMP and CTX induced extensive muscle destruction on day 5, although the effects of CTX were reversed over time. We identified the presence of fibrinogen and P-selectin exposure inside the damaged muscle sections, suggesting signs of bleeding and the formation of platelet aggregates/microthrombi in tissues, respectively. Intriguingly, CAMP causes integrin shedding but does not affect any blood clotting parameters, whereas CTX significantly extends the clotting time and has no impact on integrin shedding. The rates of fibrinogen clearance and reduction in microthrombi were greater in CTX-treated muscle compared to CAMP-treated muscle. Together, these findings reveal novel aspects of venom-induced muscle damage and highlight the relevance of haemostatic events such as bleeding and thrombosis for muscle regeneration and provide useful mechanistic insights for developing better therapeutic interventions.

Myofiber directs macrophages IL-10-Vav1-Rac1 efferocytosis pathway in inflamed muscle following CTX myoinjury by activating the intrinsic TGF-ß signaling.

BACKGROUND: To explore the role of skeletal muscle specific TGF-ß signaling on macrophages efferocytosis in inflamed muscle caused by Cardiotoxin (CTX) injection. METHODS: CTX myoinjury was manipulated in TGF-ßr2flox/flox (control) mice or transgenic mice with TGF-ß receptor 2 (TGF-ßr2) being specifically deleted in skeletal muscle (SM TGF-ßr2-/-). Gene levels of TGF-ß signal molecules, special inflammatory mediators in damaged muscle or in cultured and differentiated myogenic precursor cells (MPC-myotubes) were monitored by transcriptome microarray or qRT-PCR. TGF-ß pathway molecules, myokines and embryonic myosin heavy chain in regenerating myofibers, the phenotype and efferocytosis of macrophages were evaluated by immunofluorescence, immunoblotting, Luminex, or FACS analysis. In vitro apoptotic cells were prepared by UV-irradiation. RESULTS: In control mice, TGF-ß-Smad2/3 signaling were significantly up-regulated in regenerating centronuclear myofibers after CTX-myoinjury. More severe muscle inflammation was caused by the deficiency of muscle TGF-ß signaling, with the increased number of M1, but the decreased number of M2 macrophages. Notably, the deficiency of TGF-ß signaling in myofibers dramatically affected on the ability of macrophages to conduct efferocytosis, marked by the decreased number of Annexin-V-F4/80+Tunel+ macrophages in inflamed muscle, and the impaired uptake of macrophages to PKH67+ apoptotic cells transferred into damaged muscle. Further, our study suggested that, the intrinsic TGF-ß signaling directed IL-10-Vav1-Rac1 efferocytosis signaling in muscle macrophages. CONCLUSIONS: Our data demonstrate that muscle inflammation can be suppressed potentially by activating the intrinsic TGF-ß signaling in myofibers to promote IL-10 dependent-macrophages efferocytosis. Video Abstract.

Skeletal Muscle Regeneration in Cardiotoxin-Induced Muscle Injury Models.

Skeletal muscle injuries occur frequently in daily life and exercise. Understanding the mechanisms of regeneration is critical for accelerating the repair and regeneration of muscle. Therefore, this article reviews knowledge on the mechanisms of skeletal muscle regeneration after cardiotoxin-induced injury. The process of regeneration is similar in different mouse strains and is inhibited by aging, obesity, and diabetes. Exercise, microcurrent electrical neuromuscular stimulation, and mechanical loading improve regeneration. The mechanisms of regeneration are complex and strain-dependent, and changes in functional proteins involved in the processes of necrotic fiber debris clearance, M1 to M2 macrophage conversion, SC activation, myoblast proliferation, differentiation and fusion, and fibrosis and calcification influence the final outcome of the regenerative activity.

Caffeic acid phenethyl ester protects against doxorubicin-induced cardiotoxicity and increases chemotherapeutic efficacy by regulating the unfolded protein response.

Doxorubicin (Dox) is an efficient drug used in breast cancer chemotherapy. However, the clinical application of Dox in cancer treatment is limited due to its cardiotoxicity. Caffeic acid phenethyl ester (CAPE) is a critical bioactive ingredient of honeybee propolis that possesses various beneficial pharmacological properties, such as antioxidant and anticancer activities. Here, we aimed to investigate the protective effect of CAPE on Dox-induced cardiotoxicity and its anti-breast cancer effects. CAPE significantly ameliorated Dox-induced toxicity in H9c2 cells and in a mouse model. Mechanistically, Dox caused endoplasmic reticulum (ER) dysfunction characterized by the activation of the unfolded protein response (UPR) and upregulation of Bax proteins, and CAPE attenuated the Dox-induced UPR in H9c2 cells. In contrast, CAPE significantly enhanced Dox-induced cytotoxicity in human breast cancer cells by upregulating the Dox-induced UPR; it also markedly suppressed tumor growth in 4T1 cancer-bearing BALB/c mice. In conclusion, CAPE could be used as a promising therapy for patients with cancer receiving Dox treatment.

Curcumin-coated gold nanoparticles attenuate doxorubicin-induced cardiotoxicity via regulating apoptosis in a mouse model.

Doxorubicin (DOX) is one of the most widely used chemotherapy agents; however, its nonselective effect causes cardiotoxicity. Curcumin (Cur), a well known dietary polyphenol, could exert a significant cardioprotective effect, but the biological application of this substance is limited by its chemical insolubility. To overcome this limitation, in this study, we synthesised gold nanoparticles based on Cur (Cur-AuNPs). Ultraviolet-visible (UV-Vis) absorbance spectroscopy and transmission electron microscopy (TEM) were performed for the characterisation of synthesised NPs, and Fourier transform infrared (FTIR) spectroscopy were applied to detect Cur on the surface of AuNPs. Its cytotoxicity effect on H9c2 cells was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The biological efficacy of Cur-AuNPs was assessed after acute cardiotoxicity induction in BALB/c mice with DOX injection. The serum biomarkers, myocardial histological changes, and cardiomyocyte apoptosis were then measured. The results revealed that the heart protection by Cur-AuNPs is more effective than Cur alone. Heart protective effect of Cur-AuNPs was evident both in the short-term (24 hours) and long-term (14 days) study. The results of Cur-AuNPs400 after 24 hours of toxicity induction displayed the reduction of the cardiac injury serum biomarkers (LDH, CK-MB, cTnI, ADT, and ALT) and apoptotic proteins (Bax and Caspase-3), as well as increase of Bcl-2 anti-apoptotic proteins without any sign of interfibrillar haemorrhage and intercellular spaces in the heart tissue microscopic images. Our long-term study signifies that Cur-AuNPs400 in DOX-intoxicated mice could successfully inhibit body and heart weight loss in comparison to DOX group.

Exposure to zinc oxide nanoparticles (ZnO-NPs) induces cardiovascular toxicity and exacerbates pathogenesis - Role of oxidative stress and MAPK signaling.

The precise toxico-pathogenic effects of zinc oxide nanoparticles (ZnO-NPs) on the cardiovascular system under normal and cardiovascular disease (CVD) risk factor milieu are unclear. In this study, we have investigated the dose-dependent effects of ZnO-NPs on developing chicken embryo and cell culture (H9c2 cardiomyoblast, HUVEC and aortic VSMC) models. In addition, the potentiation effect of ZnO-NPs on simulated risk factor conditions was evaluated using; 1. Reactive oxygen species (ROS) induced cardiac remodeling, 2. Angiotensin-II induced cardiac hypertrophy, 3. TNF-α induced HUVEC cell death and 4. Inorganic phosphate (Pi) induced aortic VSMC calcification models. The observed results illustrates that ZnO-NPs exposure down regulates vascular development and elevates oxidative stress in heart tissue. At the cellular level, ZnO-NPs exposure reduced the cell viability and increased the intracellular ROS generation, lipid peroxidation and caspase-3 activity in a dose-dependent manner in all three cell types. In addition, ZnO-NPs exposure significantly suppressed the endothelial nitric oxide (NO) generation, cardiac Ca2+ - ATPase activity and enhanced the cardiac mitochondrial swelling. Moreover, inhibition of p38 MAPK and JNK signaling pathways influence the cytotoxicity. Overall, ZnO-NPs exposure affects the cardiovascular system under normal conditions and it exacerbates the cardiovascular pathogenesis under selected risk factor milieu.

Is paclitaxel less cardiotoxic in the treatment of breast cancer before or after doxorubicin? / Qual estratégia terapêutica é menos cardiotóxica no tratamento do câncer de mama: uso de paclitaxel antes ou depois da doxorrubicina?

Background: The combination of doxorubicin (DOX) with paclitaxel (PTX) effectively treats breast cancer (BC). However, DOX-associated cardiotoxicity (CTX) is aggravated by the use of PTX. Consensus is lacking about which drug sequence involves the most CTX. Objectives: To evaluate whether DOX followed by PXT or the reverse sequence has the greatest cardiotoxic potential in the treatment of BC. Methods: Prospective study of women with primary BC who received four cycles of DOX and 12 infusions of PTX. Participants were divided into Group 1 (G1; PXT before DOX) and Group 2 (G2; DOX before PXT) at the discretion of the oncologist. CTX was defined as an absolute reduction in left ventricular ejection fraction (LVEF) > 10% to a value <53%. Patients underwentclinical evaluations and echocardiography before treatment (Phase 1) and one year after treatment (Phase 2). Results: Sixty-nine women were evaluated: 19 in G1 and 50 in G2. The groups had similar clinical characteristics. The doses of radiation, DOX, and PTX used were similar. Eight (11.6%) patients developed CTX: two (10.5%) in G1 and six (12.0%) in G2 (p=0.62). The mean LVEF was similar between groups in Phase 1 (G1=65.1±3.5%; G2=65.2±3.9%; p=0.96), with a significant reduction noted after one year in both groups: G1=61.4±8.1% (p=0.021) and G2=60.8±7.6% (p<0,001). Although lower, mean LVEF remained similar between groups after Phase 2 (p=0.79). Conclusions: In women with BC who underwent chemotherapy, the incidence of CTX at the end of the first year of treatment was similar regardless of whether DOX was used before or after PTX. (AU)

Taurine Has Potential Protective Effects against the Chronic Cardiotoxicity Induced by Doxorubicin in Mice.

Doxorubicin (DOX) is an effective anticancer anthracycline drug; however, the cardiotoxicity limits its application. The aim of the present study was to investigate the potential protective effect of taurine against DOX-induced chronic cardiotoxicity in mice. We found that exogenous supplementation of taurine can inhibit the weight loss of mice caused by DOX. The increased activity of myocardial enzymes creatine kinase (CK) and lactate dehydrogenase (LDH) in response to DOX treatment were significantly hampered. In addition, taurine supplementation alleviated the decrease in superoxide dismutase (SOD) activity, glutathione (GSH) content, glutathione peroxidase 4 (Gpx4) expression, and the increase in malondialdehyde (MDA) content caused by DOX. Besides, taurine alleviated myocardial myofibrillar disruption and mitochondrial edema. Furthermore, our results showed that taurine decreased the expressions of cleaved caspase-3 and Bax/Bcl2, thereby inhibiting apoptosis. These collective data demonstrated that exogenous taurine supplementation has a potentially protective effect against the myocardial damage caused by doxorubicin in mice by enhancing antioxidant capacity and reducing oxidative damage and apoptosis.

Computational determination of toxicity risks associated with a selection of approved drugs having demonstrated activity against COVID-19.

BACKGROUND: The emergence and rapid spread of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in thelate 2019 has caused a devastating global pandemic of the severe pneumonia-like disease coronavirus disease 2019 (COVID-19). Although vaccines have been and are being developed, they are not accessible to everyone and not everyone can receive these vaccines. Also, it typically takes more than 10 years until a new therapeutic agent is approved for usage. Therefore, repurposing of known drugs can lend itself well as a key approach for significantly expediting the development of new therapies for COVID-19. METHODS: We have incorporated machine learning-based computational tools and in silico models into the drug discovery process to predict Adsorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) profiles of 90 potential drugs for COVID-19 treatment identified from two independent studies mainly with the purpose of mitigating late-phase failures because of inferior pharmacokinetics and toxicity. RESULTS: Here, we summarize the cardiotoxicity and general toxicity profiles of 90 potential drugs for COVID-19 treatment and outline the risks of repurposing and propose a stratification of patients accordingly. We shortlist a total of five compounds based on their non-toxic properties. CONCLUSION: In summary, this manuscript aims to provide a potentially useful source of essential knowledge on toxicity assessment of 90 compounds for healthcare practitioners and researchers to find off-label alternatives for the treatment for COVID-19. The majority of the molecules discussed in this manuscript have already moved into clinical trials and thus their known pharmacological and human safety profiles are expected to facilitate a fast track preclinical and clinical assessment for treating COVID-19.

Proteomic profiling dataset of chemical perturbations in multiple biological backgrounds.

While gene expression profiling has traditionally been the method of choice for large-scale perturbational profiling studies, proteomics has emerged as an effective tool in this context for directly monitoring cellular responses to perturbations. We previously reported a pilot library containing 3400 profiles of multiple perturbations across diverse cellular backgrounds in the reduced-representation phosphoproteome (P100) and chromatin space (Global Chromatin Profiling, GCP). Here, we expand our original dataset to include profiles from a new set of cardiotoxic compounds and from astrocytes, an additional neural cell model, totaling 5300 proteomic signatures. We describe filtering criteria and quality control metrics used to assess and validate the technical quality and reproducibility of our data. To demonstrate the power of the library, we present two case studies where data is queried using the concept of "connectivity" to obtain biological insight. All data presented in this study have been deposited to the ProteomeXchange Consortium with identifiers PXD017458 (P100) and PXD017459 (GCP) and can be queried at https://clue.io/proteomics .

Cardiac toxicity from bisphenol A exposure in human-induced pluripotent stem cell-derived cardiomyocytes.

Bisphenol A (BPA) is a well-known endocrine-disrupting chemical that is widely used in a variety of products, including plastics, medical equipment and receipts. Hence, most people are exposed to BPA through the skin, via inhalation and via the digestive system, and such exposure has been linked to cardiovascular diseases including coronary artery disease, hypertension, atherosclerosis, and myocardial infarction. However, the underlying mechanisms of cardiac dysfunction caused by BPA remain poorly understood. In this study, we found that BPA exposure altered cardiac function in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Acute BPA exposure in hiPSC-CMs resulted in reduced field potential, as measured by multielectrode array (MEA). Furthermore, we observed that BPA dose-dependently inhibited ICa, INa or IKr channels. In addition, BPA exposure dose-dependently inhibited calcium transients and contraction in hiPSC-CMs. Our findings suggest that BPA exposure leads to cardiac dysfunction and cardiac risk factors such as arrhythmia.

Phosphocreatine attenuates doxorubicin-induced cardiotoxicity by inhibiting oxidative stress and activating TAK1 to promote myocardial survival in vivo and in vitro.

Myocardial apoptosis and necroptosis are the major etiological factor during doxorubicin (DOX) induced cardiotoxicity, and one of the important reasons that limit the drug's clinical application. Up to date, its mechanism has not been fully elucidated. The protective role of phosphocreatine (PCr) in heart surgery and medical cardiology has been observed in numerous clinical trials. This study aimed to evaluate cardioprotective actions of PCr against DOX-induced cardiotoxicity and investigate the underlying mechanism involving in transforming growth factor ß-activated kinase 1 (TAK1) mediated myocardial survive signaling pathway. Male Sprague-Dawleyrats were intraperitoneally (ip) injected with normal saline (NS) or DOX (2 mg/kg) alone or DOX with PCr (200 mg/kg) used as animal model. The data showed that DOX significantly impaired cardiac function and structure, induced oxidative stress, myocardial apoptosis and necroptosis, and dramatically down-regulated the expression level of TAK1, while the intervention of PCr obviously attenuated cardiac dysfunction, oxidative stress, myocardial apoptosis and necroptosis, especially alleviated the decrease of TAK1 expression. In vitro analysis, after H9c2 cells were pretreated with or without PCr (0.5 mM) or N-Acetyl-L-cysteine (NAC, 0.5 mM) or 5Z-7-oxozeaenol (5z-7-Ox, 1 µM) for 1 h, subsequently treated with DOX (1 µM) for 24 h. The results revealed that inhibition of TAK1 further deteriorated apoptotic and necroptotic cell death induced by DOX in H9c2 cells, but didn't affect oxidative stress. While the pretreatment of PCr or NAC enhanced antioxidant activity to reduce oxidative stress, significantly alleviated apoptotic and necroptotic cell death induced by DOX in H9c2 cells. Consistent with the results in vivo, PCr or NAC significantly inhibited the decrease of TAK1 expression induced by DOX. In conclusion, oxidative stress induced by DOX inhibits the expression of TAK1, and leads to myocardial apoptotic and necroptotic death, while the intervention of PCr increases antioxidant activity to alleviate oxidative stress, which in turn activates TAK1 signaling pathway to promote myocardial survival, and finally attenuate DOX-induced cardiotoxicity.

Characterization of the Skeletal Muscle Secretome Reveals a Role for Extracellular Vesicles and IL1α/IL1ß in Restricting Fibro/Adipogenic Progenitor Adipogenesis.

Repeated mechanical stress causes injuries in the adult skeletal muscle that need to be repaired. Although muscle regeneration is a highly efficient process, it fails in some pathological conditions, compromising tissue functionality. This may be caused by aberrant cell-cell communication, resulting in the deposition of fibrotic and adipose infiltrates. Here, we investigate in vivo changes in the profile of skeletal muscle secretome during the regeneration process to suggest new targetable regulatory circuits whose failure may lead to tissue degeneration in pathological conditions. We describe the kinetic variation of expression levels of 76 secreted proteins during the regeneration process. In addition, we profile the gene expression of immune cells, endothelial cells, satellite cells, and fibro-adipogenic progenitors. This analysis allowed us to annotate each cell-type with the cytokines and receptors they have the potential to synthetize, thus making it possible to draw a cell-cell interaction map. We next selected 12 cytokines whose receptors are expressed in FAPs and tested their ability to modulate FAP adipogenesis and proliferation. We observed that IL1α and IL1ß potently inhibit FAP adipogenesis, while EGF and BTC notably promote FAP proliferation. In addition, we characterized the cross-talk mediated by extracellular vesicles (EVs). We first monitored the modulation of muscle EV cargo during tissue regeneration. Using a single-vesicle flow cytometry approach, we observed that EVs differentially affect the uptake of RNA and proteins into their lumen. We also investigated the EV capability to interact with SCs and FAPs and to modulate their proliferation and differentiation. We conclude that both cytokines and EVs secreted during muscle regeneration have the potential to modulate adipogenic differentiation of FAPs. The results of our approach provide a system-wide picture of mechanisms that control cell fate during the regeneration process in the muscle niche.

Reduction of Superoxide Dismutase 1 Delays Regeneration of Cardiotoxin-Injured Skeletal Muscle in KK/Ta-Ins2Akita Mice with Progressive Diabetic Nephropathy.

Superoxide dismutase (SOD) is a major antioxidant enzyme for superoxide removal, and cytoplasmic SOD (SOD1) is expressed as a predominant isoform in all cells. We previously reported that renal SOD1 deficiency accelerates the progression of diabetic nephropathy (DN) via increasing renal oxidative stress. To evaluate whether the degree of SOD1 expression determines regeneration capacity and sarcopenic phenotypes of skeletal muscles under incipient and advanced DN conditions, we investigated the alterations of SOD1 expression, oxidative stress marker, inflammation, fibrosis, and regeneration capacity in cardiotoxin (CTX)-injured tibialis anterior (TA) muscles of two Akita diabetic mouse models with different susceptibility to DN, DN-resistant C57BL/6-Ins2Akita and DN-prone KK/Ta-Ins2Akita mice. Here, we report that KK/Ta-Ins2Akita mice, but not C57BL/6-Ins2Akita mice, exhibit delayed muscle regeneration after CTX injection, as demonstrated by the finding indicating significantly smaller average cross-sectional areas of regenerating TA muscle myofibers relative to KK/Ta-wild-type mice. Furthermore, we observed markedly reduced SOD1 expression in CTX-injected TA muscles of KK/Ta-Ins2Akita mice, but not C57BL/6-Ins2Akita mice, along with increased inflammatory cell infiltration, prominent fibrosis and superoxide overproduction. Our study provides the first evidence that SOD1 reduction and the following superoxide overproduction delay skeletal muscle regeneration through induction of overt inflammation and fibrosis in a mouse model of progressive DN.