Contrast-enhanced ultrasound (CEUS) is increasingly being used in children. One of the most common referrals for CEUS performance is characterization of indeterminate focal liver lesions and follow-up of known liver lesions. In this setting, CEUS is performed with intravenous administration of ultrasound contrast agents (UCAs). When injected into a vein, UCA microbubbles remain confined within the vascular network until they dissipate. Therefore, visualization of UCA within the tissues and lesions corresponds to true blood flow. CEUS enables continuous, real-time observation of the enhancement pattern of a focal liver lesion, allowing in most cases for a definite diagnosis and obviating the need for further cross-sectional imaging or other interventional procedures. The recent approval of Lumason  for pediatric liver CEUS applications has spurred the widespread use of CEUS. In this review article we describe the role of CEUS in pediatric liver applications, focusing on the examination technique and interpretation of main imaging findings of the most commonly encountered benign and malignant focal liver lesions. We also compare the diagnostic performance of CEUS with other imaging modalities for accurate characterization of focal liver lesions.The online version contains supplementary material available at 10.1007/s00247-021-04976-2. Common benign solid liver lesions in the pediatric population include infantile and congenital hemangiomas and focal nodular hyperplasia, whereas hepatic adenoma and regenerative nodular hyperplasia occur infrequently . Th. Th60]. The CEUS enhancement pattern of the congenital hemangioma is similar to that of infantile hemangioma, demonstrating typically peripheral nodular enhancement during the arterial phase, with centripetal continuous filling during the late portal venous phase Fig.  57]. In. In57]. Focal nodular hyperplasia (FNH) is a benign tumor that is most frequently found in young to middle-age adults, though it is also encountered in the pediatric population, typically between the ages of 2 years and 5 years. It can also be seen in oncology patients following radiation and chemotherapy , 61. PeoFocal nodular hyperplasia is thought to represent a proliferative hepatocellular response caused by increased blood flow resulting from a dystrophic artery. Histologically, it is composed of benign-appearing hepatocytes arranged in nodules that are usually partially delineated by fibrous bands originating from the central fibrous scar that contains the dystrophic arterial vessels , 63. AtyBecause this lesion is composed of normal hepatocytes, on US it appears well-circumscribed and nearly isoechoic with the adjacent normal liver, although it can also be hypo- or, rarely, hyperechoic . The cenIn a recent pediatric CEUS study, classic FNH showed the presence of a central feeding artery that is visible in the early arterial phase and demonstrates a centrifugal filling pattern, with stellate (spoke-wheel) hyperenhancement in the arterial to early portal venous phases, and iso- or hyperenhancement in the late portal venous phase Fig.  7, 66]., 66.7, 6There are some contradictory results regarding the correlation between the lesion size and the identification of the central stellate scar on CEUS. Some investigators showed that the classic spoke-wheel pattern depended on the size of the lesion and was more frequently seen in larger FNHs, typically those greater than 3 cm , 68. OthHepatic adenomas are benign, typically solitary lesions that are frequently diagnosed in women ages 35–40 years. Although hepatic adenomas are very rare in children, they are encountered more often in young girls who take oral contraception, in children who are on steroid therapy and in those with underlying metabolic disease such as galactosemia and glycogen storage disease , 71. A h, intralesional necrosis and hemorrhage). Uncomplicated hepatic adenomas on US tend to be homogeneous lesions with similar reflectivity to the adjacent normal liver, or they appear relatively hyperechoic if there is high intralesional lipid content. However, on a background of diffuse fatty liver infiltration or glycogen storage disease, adenomas might appear hypoechoic .Fig.Fig76].FOne previous study evaluating the characteristics of focal fatty infiltration of 25 lesions in adults reported a variable frequency pattern in the arterial phase, including isoenhancement (44%), hyperenhancement (12%) and hypoenhancement (44%). In the portal venous phase all lesions (100%) demonstrated isoenhancement, and these areas could not be distinguished from adjacent liver tissue . This vaRegenerative nodules, also known as “nodular regenerative hyperplasia,” occur as a hyperplastic response of hepatocytes to an insult. In the rare cases that they occur in children, they are often associated with conditions leading to chronic liver disease  or to previous use of steroids and antineoplastic medication .The appearance of regenerative nodules on US is variable and depends on the size of the nodules. Small nodules might not be apparent, but if there are many of them the liver appears with diffusely heterogeneous echotexture or distortion of normal architecture. If nodules are visible, they are generally well-circumscribed, homogeneous, and hypoechoic or hyperechoic compared with the normal liver , 81.On CEUS these nodules show similar enhancement with the background liver parenchyma throughout all the enhancement phases. They are distinct from their malignant counterparts in that they do not show washout during the portal venous or late phases .Among the primary and secondary malignant hepatic tumors known in children are hepatoblastoma, hepatocellular carcinoma, undifferentiated  sarcoma, angiosarcoma, embryonal rhabdomyosarcoma, lymphoma and metastasis. Their appearance is variable on gray-scale US and can overlap with that of benign lesions. A recent meta-analysis of 57 studies showed that CEUS has excellent diagnostic accuracy in differentiating malignant from benign focal liver lesions, with a pooled sensitivity and specificity of 0.92  and 0.87 (95% CI: 0.86–0.88), respectively . The aveThe arterial-phase enhancement pattern of malignant lesions can be variable. For example, hypervascular metastasis and hepatocellular carcinoma exhibit hyperenhancement during the arterial phase, while some metastases appear as hypoenhanced or demonstrate a rim-like enhancement in the arterial phase. Larger malignant lesions might show a disorganized internal vasculature, which becomes more visible on CEUS studies than it does on non-contrast ultrasound, CT or MRI.It is important to consider that the enhancement pattern of hepatic inflammatory and infectious lesions on CEUS can be like those of primary liver malignancy and metastatic disease. Because of the hyperemia associated with infection, abscesses have a peripheral rim of hyperenhancement during the arterial phase of CEUS. If the central component is replaced by purulent material, then the center does not enhance. However, depending on the degree of liquefaction the central aspect might also be hypoenhancing or have a well-defined non-enhancing region surrounded by enhancing septations. During the venous phase, inflammatory and infectious lesions show hypoenhancement. These patterns can be indistinguishable from malignant lesions, and biopsy might be required if the history, clinical signs and symptoms are not clear , 85.Hepatoblastoma is the most common primary malignant pediatric liver tumor, accounting for more than 90% of liver cancers in children younger than 5 years. It usually occurs sporadically, but it is sometimes associated with prematurity, Beckwith–Wiedemann syndrome, hemihypertrophy and familial adenomatous polyposis coli , 86. It Hepatoblastoma is a well-circumscribed, encapsulated, large solid mass with a lobulated contour. Epithelial hepatoblastomas typically demonstrate a relatively more homogeneous appearance, while mixed epithelial and mesenchymal tumors appear markedly heterogeneous. Teratoid components appear cystic. Areas of hemorrhage or necrosis within the tumor are common . Coarse On CEUS hepatoblastoma can show early peripheral enhancement during the arterial phase and marked contrast washout during the late portal venous phase Fig.  4]. It . It 4]. Hepatocellular carcinoma (HCC) is the second most common primary malignant liver tumor in children, accounting for 20–30% of primary hepatic malignancies. It is usually found in older children and adolescents (10–14 years) but has also been found in children younger than 5 years. Several diseases in children can lead to cirrhosis, which increases the risk of developing HCC  . HoweverThe enhancement patterns of HCC on CEUS have been detailed in adults using the CEUS Liver Imaging Reporting and Data System (LI-RADS) . This syOn CEUS, HCC typically demonstrates arterial-phase hyperenhancement, which can involve diffusely the whole lesion or a portion of the lesion (in what is known as nodule-in-nodule or mosaic appearance). Another typical feature is washout, which is late in onset (beginning 60 s after contrast injection) and mild in degree (the lesion becomes less enhanced than the surrounding liver but still demonstrates some degree of persistent contrast enhancement within 2 min after contrast injection) . The CEUA common feature of all metastatic liver lesions on CEUS is contrast washout, which is typically rapid (beginning within 60 s after contrast injection) and marked (the lesion is completely devoid of contrast within 2 min after contrast injection), resulting in a “punched-out” appearance  and contrast (left) images. In the arterial phase, the lesion demonstrates a peripheral, nodular, discontinuous enhancement pattern. In the portal venous phase, the lesion appears homogeneously hyperenhancing, whereas in the late phase, the lesion remains hyperenhancing compared to the adjacent liver parenchyma A 2-week-old boy with elevated liver enzymes. Same patient as Fig. Online Supplementary Material 2left) and contrast (right) images. During the early arterial phases, rapid enhancement of the lesion originates from the center and progresses centrifugally in in a spoke-wheel pattern. In the portal venous phase, the lesion is homogeneously hyperenhancing and remains hyperenhancing during the late phase, where a central hypoenhancing scar is evident. This enhancement pattern is in keeping with focal nodular hyperplasiaA 17-year-old girl with abdominal pain. Focal liver lesion was identified on US that was performed for appendicitis. Sagittal contrastenhanced ultrasound (CEUS) cinematic clip of the liver with dual display of gray-scale  and contrast-enhanced (right) images. There is hyperenhancement of the lesion in the early arterial phase followed by isoenhancement and slow washout. These contrast-enhanced US features are typical of HCC A 14-year-old boy with fibrolamellar hepatocellular carcinoma (HCC). Sagittal contrast-enhanced ultrasound (CEUS) cinematic clip of the right liver lobe with dual display of gray-scale  and contrast-enhanced (right) images. There is early washout of the lesion beginning at about 10 s. Early washout is typical of liver metastases on CEUS A 7-year-old girl with Wilms tumor and metastatic liver disease. Sagittal contrast-enhanced ultrasound (CEUS) cinematic clip of the liver with dual display of gray-scale (


From our mapping, key regional differences are apparent in fine-scale valley network attributes including morphologic type, planimetric form, density, and links to candidate paleolakes. This information, combined with crater retention age (inferred exposure age), provides new details on the relative timing and nature of aqueous processes in the region. Newly identified pitted-type valley networks have morphological similarity to terrestrial pitted landforms in an evaporite setting. We interpret the pitted valley networks to reflect late-stage groundwater processes concentrated along the former fluvial conduits. Evidence from this study indicates that localized reactivation of valley networks occurred during or after exhumation of eastern Nhc1 unit.Morphological characterization of valley networks in three exposures of ancient cratered highlands (N Valley networks are the most common drainage feature on Mars and are frequently cited as evidence of former climate conditions conducive to sustained surface overland flow of water [e.g., Some of the ancient cratered terrain appears to be devoid of fluvial dissection [e.g., Fine-scale valley networks  were first identified in Mars Global Surveyor Mars Orbiter Camera images (0.5–12 m/pixel) and include ridges interpreted as inverted channels where the surrounding substrate has been removed by differential erosion [e.g., hc1), the lowest exposed stratigraphic unit in the region [hc1 exposures surround hematite-bearing plains where the Mars Exploration Rover (MER) opportunity is currently conducting surface observations, providing additional geologic context for rover observations.In order to characterize the aqueous history of the GMP, we examine the fine-scale valley networks in three areas that are mapped as Noachian ancient cratered highlands , as degraded layered deposits in meter-scale images [6 km2 and burying the three Nhc1 areas by an average of ~360 m [Mh) with a hematite mineralogical signature. Opportunity observations determined the hematite to be extensive lag deposits containing iron-bearing spherical concretions that eroded out from sedimentary outcrops [Within the study region, there is a clear stratigraphic succession of rock units from the lowermost ancient cratered terrains to intermediate layered deposits and a capping plains unit  [Hynek af ~360 m . At the hc1 in this study are based on the 1:2,000,000 scale geologic map of Meridiani Planum by hc1 , we assume that the mapped tributaries extend to the drainage divide, a conservative estimate of former extent, and use a convex-hull method [To quantify the spatial coverage of valley networks, we define regional valley network density as the ratio of total mapped valley network length to the area of the map unit [hc1, noting those craters that clearly superpose the unit.  Craters greater than 100 m in diameter were mapped on CTX images, and counts excluded obvious secondary clusters. We determine the crater size-frequency populations for surface units using standard crater counting methodology as detailed in hc1 provide relative age constraints in the geologic reconstruction. Results are shown in Crater counts were conducted on the western and eastern exposures of unit NFour principle fine-scale valley network morphologic types are observed: pitted, channel, ridges, and knobs . TransitPitted-type valley networks, documented for the first time in this study, are circular- to oval-shaped depressions that often intersect one another to form a chain that connects with a channel segment . Pits vahc1 regions, which, in general, show basaltic compositions reflecting the pervasive sand cover of this unit. Only one valley network segment, located in the eastern Nhc1 unit, has a strong sulfate signature , albeit over a much smaller area than in the southern Nhc1. Pitted-type valley networks, with a greater number of closely spaced low-order segments, appear to be a more representative record of the original planimetric pattern than classic valley networks that show no microdissection  [Regional differences are also apparent in the attributes of valley networks. Both eastern and southern Nhc1, the fine-scale valley networks mapped in these regions are an important record of past aqueous conditions. We interpret the fine-scale valley network morphologic types to mark the former fluvial conduit (~100 m wide). Inner channels are rarely observed for Martian valley networks because the canyon floor is typically covered by wind-blown sediment, but where seen, it is typically 90–350 m wide [Although large-scale valley networks are generally absent in eastern and western NTo understand the processes that may have been involved in forming the fine-scale valley network morphologic types, and their interconnected configuration, documented in this study, we draw comparisons with a terrestrial analog site with denuded evaporite deposits in a hyperarid, wind-eroded setting. Features in the Pampa del Tamarugal region of the Chilean Atacama Desert have similarities to fine-scale valley networks in the GMP based on parallels in the scale and arrangement of landforms, the environment setting, and the nature of materials (composition and grain size), to the extent known.The analog site is on a high-elevation plateau (~1000 m) situated between the Pacific coastal mountains and the higher Andes Mountains to the east. This hyperarid region receives less than 10 mm/yr of annual precipitation and almost no locally generated runoff [In Salar de Llamara [Wind is a prominent force in eroding and sculpting deposits in the Atacama Desert. In Salar de Llamara, the prevailing winds (~15 m/s) are to the northeast [hc1. These ridges are continuous over several kilometers and have few branching segments, consistent with terrestrial inverted channels which generally lack tributaries [e.g., hc1 have continuity relationships with candidate paleolake deposits    and S8b.hannels) , is simihc1 units in our study area by this time. However, from the morphological attributes of fine-scale valley networks, the pattern of fluvial dissection appears to differ regionally, possibly reflecting the availability of water [e.g., hc1, while the lower elevation and shallow gradient setting of western Nhc1 was a local sedimentary sink with single-thread fluvial systems and ponded water.Using the development sequence of GMP from prior studies, the mapped regional attributes of fine-scale valley networks in this study, and the Salar de Llamara analog, we discuss the nature and relative timing of aqueous processes associated with valley networks on the ancient cratered terrain. The overwhelming majority of classic valley networks in Arabia Terra appear to have been carved by the Noachian-Hesperian boundary (~3.7 Ga) [Etched; see section 2). Fluctuating water tables resulted in sulfate precipitation with thicknesses >50 m in areas [e.g., hc1 at mapping scale [Near the time of the Noachian-Hesperian boundary (~3.7 Ga), the GMP was buried by layered sedimentary materials forming the etched units  and therefore postdates both (1) the majority of regional exhumation and (2) formation of the capping hematite-bearing Meridiani plains (opportunity field site). Reactivation of the former valley network conduits by groundwater-fed dissolution/precipitation processes generated pit chains, curvilinear depressions, and ridges that are analogous to those observed in Salar de Llamar, Chile. Localized reactivation of valley networks likely occurred during or after exhumation of eastern Nhc1 unit. Exposure ages of the Nhc1 units constrain the timing of these late-stage groundwater processes (In contrast, the valley network types in eastern Nrocesses .Results from this study indicate that surface conditions in Arabia Terra must have been warm enough to support liquid water, at least at times, in the Hesperian or later. This is consistent with previous studies that have identified aqueous activity that postdates regional widespread fluvial incision in Arabia Terra: deposition of hydrates following erosion of the etched deposits [In conclusion, this study provides details on the history of GMP valley networks: an early phase of sustained surface overland flow, followed by a period dominated by groundwater processes associated with widespread evaporite deposition and ultimately transitioned to localized dissolution of solutes that reactivated ancient valley networks. Further constraining the relative timing and duration of these aqueous periods is of fundamental importance to an accurate characterization of past climate conditions on Mars [


Soil heavy metal (HM) pollution, which arises from natural and anthropogenic sources, is a prime threat to the environment due to its accumulative property and non-biodegradability. Ectomycorrhizal (ECM) symbiosis is highly efficient in conferring enhanced metal tolerance to their host plants, enabling their regeneration on metal-contaminated lands for bioremediation programs. Numerous reports are available regarding ECM fungal potential to colonize metal-contaminated lands and various defense mechanisms of ECM fungi and plants against HM stress separately. To utilize ECM–plant symbiosis successfully for bioremediation of metal-contaminated lands, understanding the fundamental regulatory mechanisms through which ECM symbiosis develops an enhanced metal tolerance in their host plants has prime importance. As this field is highly understudied, the present review emphasizes how plant’s various defense systems and their nutrient dynamics with soil are affected by ECM fungal symbiosis under metal stress, ultimately leading to their host plants ameliorated tolerance and growth. Overall, we conclude that ECM symbiosis improves the plant growth and tolerance against metal stress by (i) preventing their roots direct exposure to toxic soil HMs, (ii) improving plant antioxidant activity and intracellular metal sequestration potential, and (iii) altering plant nutrient uptake from the soil in such a way to enhance their tolerance against metal stress. In some cases, ECM symbiosis promotes HM accumulation in metal stressed plants simultaneous to improved growth under the HM dilution effect. Beta vulgaris is higher in summers than in winter due to the relatively high transpiration rates  pollution. HMs can also leach down into groundwater or can be transferred to the successive levels of the food chain , causingon rates . From plon rates . HM toxion rates .Eucalyptus, Pinus, Acacia, and Picea  derived from tree Picea can transfer to the surrounding trees through ECM mycelia networks, where the exchange is found to be higher with phylogenetically related trees. The ECM communities among the phylogenetically related trees are very similar in composition  typosition , host agposition , and hosposition ; and   reduces the growth of Willow plants upon ECM inoculation in Betulapubescens  Inl stress . Furtherl stress . Zong etl stress . (iii) Al stress . In the l stress . The enhl stress . The varl stress . While il stress . Under Cd stress . The ECMd stress . To enhad stress .Oryza sativa OsMT1b and OsMT2c under Cr stress (Oryza sativa OsMT1e under Cd stress (Physcomitrella patens PpMT2 under Cd and Cu stress (Phytolacca americana PaMT3-1 under Cd stress (LbMT1 and LbMT2 in Laccaria bicolor under Cd and Cu stress, respectively (PaMT1 in Pisolithus albus under excess Cd and Cu stress (Hebeloma mesophaeum under Cd stress; and SuiMT1 and SuiMT2 in Suillus indicus under Cu stress (Metallothioneins are cysteine-rich low–molecular weight proteins that strongly bind to metal ions through their thiol groups of cysteine residues, thus playing an essential role in metal sequestration and detoxification . Besidesr stress ; Oryza sd stress ; Physcomu stress ; and Phyd stress . On the ectively ; PaMT1 iu stress ; HmMT3 iu stress , leads tu stress . Althougu stress .Glutathiones are well known to prevent oxidative stress and xenobiotics detoxification in cells . The upr2O2, whereas peroxidase and catalase mediate the H2O2 conversion to H2O (Quercus acutissima roots inoculated with Suillus luteus compared to non-ECM plants (Pinus massoniana under Al stress has also been reported upon their inoculation with Suillus luteus compared to non-ECM plants (The excess concentration of plant nutrient Zn can lead to increased production of ROS, which, if it breaks its balance with ROS destruction, results in oxidative stress . With evn to H2O . The actn to H2O . The ECMM plants . The lowM plants . It thusThe resource allocation between ECM fungal and plant partners strongly regulates the maintenance of long-term symbiotic association among both the partners. This long-term cooperation of nutrient transfer is maintained by transcriptional and translational regulations of transporter systems at regulatory checkpoints as shown in Further, the Na and P enrichment highly promotes Cd uptake and its sequestration with proline in plant roots which further decreases Cd transfer from sources to stem and helps to enhance the phytoextraction potential of the plant . On the P. albus symbiosis reduces Mg uptake in their host plants growing on ultramafic substrate rich in HMs to check excess Mg transfer. In contrast, host uptake for ultramafic soil deficient in N, P, K, and Ca increased, thus improving plant growth and creating a barrier for HMs present in excess (Pinus sylvestris roots inoculated with different ECM fungal species under HM stress was reported compared to non-mycorrhizal plants. In shoots, reduced Fe content and increased Mg and Ca content were observed upon ECM symbiosis compared to non-mycorrhizal plants under metal stress (On the other hand, n excess . The enhl stress . The varl stress .At present, the restoration of abandoned mining lands is highly required to improve soil quality, microorganisms and plants growth for ecological rehabilitation . The useSedum alfredii have more Cd accumulation in roots than non-hyperaccumulators (Robinia pseudoacacia seedlings under As stress enhanced the Indole-3-acetic acid and abscisic acid content, decreasing the zeatin riboside gibberellic acid concentrations and altering the ratios of various phytohormones in the host plant. These results suggest that the mycorrhiza-mediated phytohormones are essential factors behind host-alleviated metal toxicity (The different species of ECM fungi affect host tolerance efficiency against metal stress differently , therebymulators . Among tmulators . It is nmulators . Inoculatoxicity . Althougtoxicity . Severaltoxicity . It woultoxicity . To thestoxicity . The biotoxicity . The AM toxicity . Similartoxicity .The ECM symbiosis improves the HM tolerance and growth of their metal stressed host plants through several mechanisms, which thus help in regenerating the metal-contaminated lands with plants. First, the ECM fungi can either act as a physical barrier between soil HM and plant roots or enhance HM accumulation in host plants depending on several factors such as HM type, external concentrations, and plant and fungal species. ECM fungi can also prevent HM transfer to plant roots by sequestering them on fungal cell walls or intracellularly with MTs and glutathione. ECM symbiosis ameliorates plant growth and their tolerance to oxidative stress under HM stress. The ECM fungi can change the nutrition dynamic of plants with soil in such a way to create HM dilution effects and to prevent HM transfer from roots to shoots. The role of ECM promoted or inhibited release plant root exudates in HM stress tolerance needs more studies for clarification. On the basis of the collected evidences, the ECM symbiosis proves to be beneficial for promoting plant growth on metal-contaminated lands and enhancing soil HM phytoextraction in host plants to reduce soil HM content. This field requires more extensive studies to understand the nutrient dynamics of ECM plants under metal stress and how it affects their tolerance.EC conducted the literature review and designed and wrote the manuscript. MR supervised, corrected, improved, and accepted the final version of the manuscript. Both authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


One of the most important features of striated cardiac muscle is the excitability that turns on the excitation-contraction coupling cycle, resulting in the heart blood pumping function. The function of the heart pump may be impaired by events such as myocardial infarction, the consequence of coronary artery thrombosis due to blood clots or plaques. This results in the death of billions of cardiomyocytes, the formation of scar tissue, and consequently impaired contractility. A whole heart transplant remains the gold standard so far and the current pharmacological approaches tend to stop further myocardium deterioration, but this is not a long-term solution. Electrically conductive, scaffold-based cardiac tissue engineering provides a promising solution to repair the injured myocardium. The non-conductive component of the scaffold provides a biocompatible microenvironment to the cultured cells while the conductive component improves intercellular coupling as well as electrical signal propagation through the scar tissue when implanted at the infarcted site. The in vivo electrical coupling of the cells leads to a better regeneration of the infarcted myocardium, reducing arrhythmias, QRS/QT intervals, and scar size and promoting cardiac cell maturation. This review presents the emerging applications of intrinsically conductive polymers in cardiac tissue engineering to repair post-ischemic myocardial insult. Cardiovascular diseases (CVDs) represent the leading cause of death within the European Society of Cardiology member countries with 108.7 million active cases as of 2017 and 1.67 million lives lost to ischemic heart disease the same year . MyocardIn recent years, the parallel progresses in cell biology, materials science, and advanced nano-manufacturing procedures have allowed us to envision the possibility to set up novel strategies (collectively dubbed “tissue engineering”) to combine cells and biomaterials to fabricate myocardium-like structures in vitro to be engrafted into the heart to repair the damaged parts. This approach is characterized by an extreme level of complexity and, as a matter of fact, after more than two decades of extensive efforts, many issues remain to be answered before tissue engineering products can be used at the bedside. Indeed, the maturation of CMs from stem cells is orchestrated by biological and physical signals released through the extracellular matrix (ECM) to the cells. ECM is a composite material made of a gel matrix and different fiber types. The interplay among the different ECM components releases physical signals relevant to determine the cell fate. Several studies have been performed to define protocols helpful to manufacture a structure (scaffold) emulating the native ECM on which cells can grow and differentiate. Natural or synthetic biomaterials, such as cardiac patches , injectaThe present review is aimed at analyzing the state-of-the-art use of intrinsically electroconductive polymeric materials in the fabrication of functional cardiac constructs to envision future directions and potential clinical applications.Polyaniline (PANI) is a widely used material in tissue engineering and biomedical applications  due to il-lactide-co-ε-caprolactone)/PANI electrospun membranes [l-lactide)/aniline pentamer films also demonstrated a high proliferation rate and, after six days under the electrical stimulation, cells demonstrated a 1.57-fold increase in the intracellular calcium concentration compared to the unstimulated group [The presence of PANI in these constructs improves the intercellular signaling pathways between cultured cells, which enhances the cell viability and various cellular behaviours as well. Good viability and proliferation rate were observed when H9c2 rat cardiac myoblasts were cultured on pristine and gelatin-blended polyaniline substrates. The cell population doubled after 54 h of culture  with smoembranes . H9c2 raed group .l-lactide acid)/PANI sheets generated myotubes with their length, myotube density, and fusion index increased 2–3 fold compared to when cultured on non-conductive PLA membranes. Furthermore, neonatal rat CMs cultured on these nanofibrous conductive PLA/PANI sheets maintained synchronous beating patterns for up to 21 days and exhibited striated morphology with enhanced expression of sarcomeric α-actinin and Cx43. Conversely, non-conductive PLA membranes induced irregular contractions in the cultured CMs [Pristine polyaniline , or whenured CMs .l-lactide) blended with aniline tetramer demonstrated good biodegradability, high ductility and high electroactivity. After seven days of culture, these conductive copolymers promoted the myogenic differentiation of C2C12 cells to myotubes with increased myotube density, length, diameter, and maturation index also evident from the elevated expression of MyoD and cTnT2 [Aligned conductive mesh of PANI and Poly(lactide-co-glycolic acid) attracted fibronectin and laminin from the culture medium and led to better cell attachment. Three days post culture, neonatal rat CMs developed rhythmic beating patterns with enhanced expression of cardiac troponin I (cTnI) and Cx43 . Fibrousnd cTnT2 .More recently, conductive injectable hydrogels based on dextran grafted aniline tetramer blended with chitosan demonstrated adequate in vivo injectability and degradability when implanted subcutaneously in rats. Different cell lines, such as C2C12 and L929, encapsulated in these conductive hydrogels maintained vitality and proliferation capacities when released in a rat volumetric muscle loss injury model. Therefore, these conductive hydrogels could also be used as a cell delivery vehicle, promoting muscle tissue regeneration .2+ transient propagation in cultured neonatal rat CMs and improved myocardial electrical signal conduction with elevated cardiac functions in the rat MI model [Polypyrrole (PPy) is another intrinsically conductive polymer that has been extensively studied in the field of biomedical and tissue engineering, as shown in MI model . Human-iMI model  while thMI model . This deMI model .2+ influx called calcium transient is the link between the electrical signals that run throughout the heart and myocyte contractions to pump blood [2+ levels) was reduced by 1.24-fold compared to cells cultured on non-conductive PCL films [v1.5 encoded by the SCN5A gene [+ ions then depolarised the plasma membrane, leading to the synchronous beating of the CMs.Calcium is a critical regulator of the cardiac contraction and relaxation cycle. Intracellular Camp blood . HL-1 muCL films ,76. SimiCL films . In addiCL films . The actN5A gene . The infPEDOT-based electroconductive scaffolds represent a good electrical interface with biological cells/tissues owing to their intrinsic capability to conduct electrical currents, good stability, mechanical properties and biocompatibility, thus providing promising platforms for tissue engineering applications ,80,81. TA highly porous PEDOT/alginate conductive scaffold was prepared to form a chemically crosslinked alginate network, using adipic acid hydrazide as the crosslinker, in which simultaneously PEDOT was synthesized in situ. Brown adipose-derived stem cells (BADSCs) differentiated to cardiomyocyte-like cells seven days post-culturing on these conductive porous scaffolds. Almost 21% and 60% of the adherent cells demonstrated enhanced expression of Cx43 and cTnT2, respectively, with most cells exhibiting enhanced α-actinin expression. The electrical stimulation improved the differentiation of BADSC to cardiomyocytes on these porous scaffolds . LikewisTo further mimic the micro/nanofibrous architecture of the native myocardium as well as the myocardial electrical conductivity, a multiscale conductive scaffold was fabricated via a layer-by-layer deposition approach combining solution-based and melt-based electrohydrodynamic (EHD) printing techniques. PCL microfibers were printed to mimic collagenous fibres and sub-micron scale conductive PEDOT:PSS-PEO fibres were printed to mimic the electrically conductive Purkinje fibres of native cardiac tissue. Fibre orientation was controlled in a layer-by-layer manner. Primary rat cardiomyocytes demonstrated good adhesion with PEDOT:PSS-PEO/PCL micro/nanofibrous conductive scaffolds and, after eight days, cells exhibited enhanced expression of α-actinin and Cx43. Cells started to beat synchronously and the beating frequency increased by 1.46-fold compared to cells cultured on non-conductive PCL scaffolds . Similarpeak) which indicates a strong electrical coupling between AG and PG. The passive group will fire an action potential once the threshold (Upeak) crosses the resting membrane potential of −90 mV [Though the electroconductive scaffolds provide better platforms for the tissue engineering of nerve, cardiac, and skeletal muscle tissues, the mechanism by which these constructs influence cellular behaviour is yet to be known . To uncof −90 mV ,92. In tf −90 mV ,93,94,95f −90 mV .Electroconductive scaffolds represent a leap forward in the effort to manufacture supports to properly address cell fate towards a cardiomyocytic phenotype. The scaffold fabrication is a complex endeavour finalised to replicate in vitro the ECM microenvironment to foster proper cell-cell and cell-matrix interactions driving the cardiomyocyte maturation process. An optimal scaffold must be biocompatible , with (i) adequate porosity  and (ii) mechano-structural design , and (iii) with electroconductivity appropriate to mimic the ECM structure and function. Finally, these characteristics must allow the integration of the engineered tissue with the native injured myocardium to restore heart function. Studies on experimental animals have shown that scaffolds endowed with appropriate electroconductivity substantially improve the beneficial effects after implantation in injured hearts. Ejection fraction (EF) and fraction shortening (FS) that give a measure of the blood pumping ability of the heart ,97 were These encouraging results, however, do not clarify the mechanisms through which the engineered tissue is integrated into the injured myocardium. After an ischemic attack, the deprivation of the oxygen and other blood-borne factors determine the impairment of the myocardial machinery  subverting the ECM composition and function. Among others, a complex array of inflammatory factors is released that induces the recruitment of different immune cell populations. Macrophages are accumulated in the damaged tissue either to remove the debris (M1 population) or to activate the limited native regenerative potential (M2 population). As a final result, myocardial bioarchitecture is wrecked, the vascular beds are no more hierarchically juxtaposed, and fibrosis alters the tissue stiffness and other mechano-structural characteristics. In this inhospitable context, it is hard to imagine that the grafted cells/structure viability can be adequate to be integrated with the host tissue. This requires that novel strategies are implemented to modulate the processes occurring in the injured myocardial tissue. The electroconductive scaffold can be used as a device to release novel drugs and to apply mechano-structural and electric signals able to restrain the overwhelming inflammatory processes. At the same time, an electroconductive scaffold can enhance the cardiomyocytic differentiation potential of the MSC, hiPSCs, and CPC (cardiac progenitor cells) used to manufacture the engineered tissue. In fact, MSCs do not express the MHC class II molecules and thus evade immune system recognition. They also depress the inflammatory reaction modifying the polarisation of M1 macrophages to reparative M2 macrophages via miR-182 shuttling  and decrHopes that the adult human heart can regenerate after infarction remain scarce due to the low turnover rate of the cardiomyocytes. Therefore, cardiac tissue engineering could provide promising solutions to repair the injured myocardium. Culturing stem/progenitor cells on electroconductive scaffolds can increase their differentiation by potentially several folds due to the electrical properties of such scaffolds. As the intrinsic electric field of the conductive scaffold encourages the differentiation of stem cells to cardiomyocyte-like cells, this eventually leads to the expression of the key cardiac proteins, such as Cx43, and subsequently facilitates the electrical coupling and contractility of cells. Cardiac functions of the infarcted heart are significantly alleviated as well when the conductive scaffold is implanted in vivo, as shown in rat MI models . This si


Circular RNA (circRNA), a relatively new member of the non-coding RNA family, has spurred great interest among researchers following its discovery as a ubiquitous class within the RNA world. Rapid progress in circRNA biology has coincided with its identification in a plethora of diverse roles including regulation of gene expression and probable coding potential, as well as competing interactions with proteins and microRNAs in various pathological conditions. Emerging evidence suggests that circRNAs also function in viral infections. The deregulation of circRNAs during viral infection has prompted investigations into the possibilities of circRNA as a competing endogenous RNA (ceRNA) that modulates response to infection. Recently, viruses have been shown to encode circRNAs with proviral functions, providing a strong impetus for focused efforts to elucidate the networks coaxed by circRNAs during infection. This review elaborates on recent insights gained on the roles of circRNAs during virus infection and immunity. The success of a pathogen is derived from its ability to subvert the host intracellular mechanisms effectively. Viruses, being obligatory parasites, are adept at subverting various host mechanisms for their benefit. Hijacking the host intracellular machinery by employing a set of virulence factors, viruses promote an environment within the host cell to produce millions of their copies and deplete the host resources. This sudden conflict must be solved, and the recurrence also has to be prevented from time to time. Hence, vertebrates have developed intricate antiviral signaling mechanisms with the help of which such intruders are kept in check. By augmenting the expression of antiviral factors comprising proteins and other endogenous non-coding RNAs (ncRNAs), the host strives to fight the incoming virus. Various viral and host strategies shape this battle. While central to this host-virus conflict has been proteinaceous effectors, recent evidence also suggests circRNAs as an arsenal employed by the host as well as the virus.CircRNA is the class of RNA formed by a non-canonical splicing event, termed as back-splicing. Among other ncRNAs, the archaic perception of circRNA as transcriptional junk was a significant impediment to pioneering research on this topic . HoweverHuman cells have various proteins with antiviral functions that are augmented during the infection. These antiviral effectors target the viral life cycle and disrupt various virion components by sensing them as the virus tries to replicate. RNA viruses are detected by key RNA binding proteins such as retinoic acid-inducible gene I (RIG-I), melanoma differentiation-associated gene 5 (MDA5), laboratory of genetics and physiology 2 (LGP2), Protein Kinase R (PKR) and Toll-like receptor 3 (TLR3) , includivia a downstream cascade  , PIWI-inFor instance, Epstein-Barr Virus (a Herpesvirus) encodes circRPMS1 that was earlier shown to promote metastasis by escalating cell proliferation, cell invasion, and inhibiting apoptosis in EBV-positive Nasopharyngeal carcinoma (NPC) cells. It is also known to regulate gene expression by sponging miRNAs such as miR-203, miR-31, and miR-451 –24. A feCircRNA was also found to regulate gene expression in the case of CHB (Chronic hepatitis B), promoting the pathogenesis of HBV (Hepatitis B virus) and associated liver disease. By establishing a bioinformatics pipeline for detecting circRNA associated with CHB and performing an in silico analysis of the circRNA-miRNA-mRNA axis, Zhou and colleagues have shown that the circRNA hsa_circ_0000650 promotes TGFβ2 expression by negatively impacting the miRNA, miR-6873-3p . AnotherTo identify the role of circRNA in the host and viral interactions, researchers have developed competing endogenous RNA (ceRNA) networks  to explovia ceRNA networking showed that circRNA might serve as potential therapeutic targets in Middle East respiratory syndrome coronavirus (MERS-CoV) infection. An siRNA mediated knockdown of host-derived circRNAs, circFNDC3B and circCNOT1, significantly reduced viral load suggesting the pro-viral activity of circRNAs. Furthermore, circFNDC3B and circCNOT1 regulated target mRNA expression involved in ERK/MAPK pathway and RIG-I-mediated antiviral signaling (In Ebolavirus (EBOV) infection, both the immune system and the vascular system are hampered, which leads to severe hemorrhagic symptoms . To undeignaling . HoweverOur discussion until this point suggests that circRNAs are both liabilities as well as an asset for the host. Although the fate of virus-host interaction may be altered by circRNA, the probable answer would lie in the mechanism of action. It is very well conceivable that alternate mechanisms exist which presently remain unearthed.The discovery of circRNAs and associated functions has enabled a paradigm shift in the field of non-coding RNA biology, which has now extended to viral infections. Even considering just a handful of validated circRNAs, the interacting network seems intricately intertwined. Changes in the levels of circRNAs could have far-reaching ramifications on infection outcomes by modulating immune effectors. A wealth of intriguing early results on virally encoded circRNAs and those regulating immune responses provides a strong impetus for focused efforts in the future to elucidate their functions. Such analysis will require biochemical characterizations using relevant infection models to identify the interactions which are not only binary but include a network of potentially intertwined interactions coaxed by the circRNA.In summary, an analysis of the myriad circRNA interactions in viral infections may represent a robust platform to uncover regulatory networks that protein-focused studies have overlooked. Development of circRNA-centric molecular assays and improvement in the functional assays could help unveil new roles of circRNA in the foreseeable future. Comprehensive insights into host-virus interactions obtained by such analyses can have exciting implications for developing an improved, enhanced, and effective therapeutic arsenal.ACho and PM contributed equally. Conceptualization: ACha. All authors contributed to the article and approved the submitted version.ACha thanks DBT/Wellcome Trust India Alliance [grant number IA/I/18/2/504006] for the financial support. ACho, PM, and VB are supported by a fellowship from the MHRD, UGC, and CSIR, respectively.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The expansion of the nuclear cardiology armamentarium to encompass a wider range of tracers targeted to specific molecular processes offers opportunities for precise monitoring of disease progression and guidance of therapy.Journal of Nuclear Cardiology, Stendahl and colleagues describe a novel diagnostic device for minimally invasive tissue-level detection of tracer distribution in the myocardium, with a vision toward guided regional therapeutic intervention.18F-fluorodeoxyglucose (18F-FDG) was observed by ex vivo PET imaging, thought to reflect regional inflammatory cell infiltration to the infarct and border zone territories. The regional disparity in signal was confirmed by spatial mapping using the β-detection catheter, providing a matrix distribution of activity content. Segment-to-segment activity established reasonable correlations between ex vivo PET, gamma well counting, and catheter-based measurements supporting the accuracy of the latter.In the current issue of the The strength of the manuscript lies in the ingenuity of the approach, which tackles the challenge of regional tracer uptake heterogeneity using a minimally invasive local measurement. While the analysis is limited to only two animals and selected areas in these infarct hearts, validation by conventional  PET and well counting indicates the capacity of the β-detector catheter to define spatial disparities of tracer accumulation. However, it is disappointing that this potential is not explored further, e.g., by confirmation of cellular substrate by histopathology. Indeed, while typical investigation requires alignment of adjacent tissue sections for ex vivo autoradiography, histology, and immunohistochemistry,18F-FDG distribution for proof-of-concept. 18F-FDG is notoriously promiscuous, which hampers the interpretation of the meaning of the signal. Indeed, numerous studies have demonstrated the inefficacy of typical suppression methods, especially in actively remodeling or acute infarct myocardium, generating a mixed substrate for the 18F-FDG signal including inflammatory leukocytes and metabolically compromised cardiomyocytes,This limitation in regional characterization is compounded by the reliance on A theoretical benefit of the β-detection catheter is the possibility to selectively distinguish β+ decay from γ decay. Such detection might provide a more accurate localization of the signal by identifying the emitted particle rather than the annihilation event. With a rising prevalence of transition metal radiochemistry in cardiovascular molecular imaging,Sensitivity poses a further challenge, wherein conventional PET imaging of inflammation after myocardial infarction has generally not attempted to discern between regionally robust or subtle inflammatory content. In conditions like myocardial infarction where the imaging target  is also elevated in the circulation, delineation of subtle tissue inflammation can be challenging, especially with transition metal isotopes. While the total inflammatory signal on PET imaging can predict functional outcome,Indeed, it is problematic that the analysis is by necessity conducted entirely ex vivo. Clearly one benefit for the β-detection catheter is the possibility for minimally invasive measurement, providing a more accurate virtual biopsy on the miniature scale. It will be essential to demonstrate the equivalence of in vivo measurements, replete with cardiac motion to confirm the accuracy of catheter-based measurements in the real clinical situation. This will further allow characterization of β-detector sensitivity for activity in tissue sections against blood pool.Nonetheless, the potential applications for such a device are broad, as many etiologies of cardiovascular disease exhibit regional heterogeneity. As intimated by Stendahl and colleagues, one potential application is localized inflammatory cell infiltration after acute myocardial infarction, where the severity and persistence of adverse inflammation can contribute to worse prognosis and remodeling.Ultimately, the greatest strength of the non-invasive or minimally invasive approach is the capability to combine regional diagnostics with regional therapy. Regional inflammation could be directly treated by anti-inflammatory agents, gene therapy to promote endogenous repair, or immunomodulatory therapy to target adverse remodeling processes.Miniaturization to explore pathogenesis remains the realm of science fiction, the capability to visualize distinct regional differences in tissue substrate within the injured myocardium on a miniatured scale is an initial step toward tissue-level quantification and substrate delineation. The potential combination of such tissue-level imaging techniques with targeted delivery of cell-, gene-, or drug-based therapy bears clear potential for future modification of image-guided therapy. This catheter-based fantastic voyage provides a template for accurate study of regional tracer distribution, but many steps remain to bring such methodology to clinical practice.


Gut microbiota has emerged as a major metabolically active organ with critical functions in both health and disease. The trillions of microorganisms hosted by the gastrointestinal tract are involved in numerous physiological and metabolic processes including modulation of appetite and regulation of energy in the host spanning from periphery to the brain. Indeed, bacteria and their metabolic byproducts are working in concert with the host chemosensory signaling pathways to affect both short- and long-term ingestive behavior. Sensing of nutrients and taste by specialized G protein-coupled receptor cells is important in transmitting food-related signals, optimizing nutrition as well as in prevention and treatment of several diseases, notably obesity, diabetes and associated metabolic disorders. Further, bacteria metabolites interact with specialized receptors cells expressed by gut epithelium leading to taste and appetite response changes to nutrients. This review describes recent advances on the role of gut bacteria in taste perception and functions. It further discusses how intestinal dysbiosis characteristic of several pathological conditions may alter and modulate taste preference and food consumption via changes in taste receptor expression. The gastrointestinal tract is the main location site that harbors approximately 4 × 10articles . Along wHumans can recognize five primary distinct taste qualities: sweet, bitter, sour, salty and savory (or umami) and are also able to detect fat properties, although this is debatable ,5. This Sugars, minerals, organic acids, alkaloids and amino acids in foods act as chemical messengers by binding to their corresponding taste receptors . This inSince identifying the chemical composition of foods is vital to homeostatic processes, it is not surprising that the same taste receptors of the tongue are also found in the gastrointestinal tract. These receptors then transduce nutrient signals into neuropeptide hormones, vagus nerve activation, or nutrient utilization . InteresThe complexity of the gut microbiome such as richness and abundance have been used as a predictive factor of the host metabolic health. Diet and dietary habits are the main modifiers of the gut microbiome composition profile. Not surprisingly some determinants of obesity such as taste perceptions that influence changes in appetite regulation and energy metabolism have been linked with the gut microbiome. This relationship between taste and oral and intestinal microbiome is reviewed in the next sections.Staphylococci and Streptococci [The oral microbiome is one of the most stable and diverse ecosystems in the body due to the variety of niches present in the mouth and the abundance of exogenous nutrients during feeding and endogenous nutrients in salivary production ,31. Specptococci . Since tptococci ,34. Micrptococci  , Actinomyces (Actinobacteria), and Lactobacillus (Firmicutes) species degrade carbohydrates into organic acids while Prevotella (Bacteroidetes) and Porphyromonas (Bacteroidetes) species break down proteins into amino acids and SCFAs [Streptococci [Another possibility by which oral microbiome influences taste perception is through the consumption of tastants  or synthesis of bioactive metabolites  before receptor interaction, ultimately inducing sensorial adaptation . Takahasnd SCFAs . While tnd SCFAs  used metptococci .Lactobacilli growth [Actinobacteria and Bacteroidetes in tongue film are linked to an increase in taste sensitivity, especially to bitterness [Actinobacteria produce phenols which can enhance the sensation of astringency and the bitter taste in food products [Actinomyces (Actinobacteria), Oribacterium (Firmicutes), Solobacterium (Firmicutes) and Catonella (Firmicutes); and the Gram-negative Campylobacter (Proteobacteria) [While bacterial metabolism may deplete sucrose, certain species can also enrich sensory-active molecules like acids in the medium near the taste receptors. This microbial production of acid does not induce a conscious sour sensation but rather increases sour detection thresholds through the sensory adaptation phenomenon. In this context, taste ability was reduced in the acutely hospitalized elderly, particularly those with high i growth . By anali growth . In thistterness . The mecproducts . When exacteria) . Taken tDue to the high environmental variations in the distal gut, the density and composition of intestinal microbial communities differ greatly from those in the oral microbiota. Whereas oral microbes must endure periods of time without exogenous nutrients, gut microbes have a continual source of nutrients in the form of dietary fiber . AlthougTaste receptors such as TAS2R are responsive to microbial-quorum sensing molecules and toxins by protecting against harmful microbes. For example, activation of TAS38 receptor increases the release of anti-microbial B-defensin and the hormone cholecystokinin. In response to changes in intestinal luminal environment, specialized EEC cells secrete a myriad of gut peptides such as CCK, GLP-1, GLP-2 and PYY. This complex regulatory process is mediated by a large family of cell-surface G-protein-coupled receptors (GPRC), expressed on the apical domain of EEC with a high degree of specificity for nutrient and taste sensing as well as microorganisms and toxic compounds. Interestingly, the GPCRs responsible for taste sensing and found in the lingual epithelium are also present in the intestine. For example, T1R2, T1R3, gustducin and GLP-1 are co-expressed in the same enteroendocrine cell. Therefore, the intestinal epithelium contains GPCR for detection of all nutrients, including fatty acids. Of interest is the fact that, indole, a tryptophan derivate and bacteria byproduct that modulates gut microbiota stimulate enteroendocrine cells to release GLP-1. It is known that GLP-1 receptors are expressed in gustatory neurons and are involved in transmission of taste signals, particularly sweet taste . FurtherThe link between gut microbiota metabolites and enteroendocrine cells has been well documented . TherefoStreptococci Mutans counts [In order to provide further insight into the complexities of human eating preference, a growing number of studies are seeking to characterize oral microbiota composition against the backdrop of taste sensitivity. A recent study by Cattaneo  found ths counts .While these correlations between bacteria taxa and gustatory function do not yet have systematic explanations, a potential mechanism of receptor modulation by bacteria does arise from germ-free murine studies. In 2011, Swartz et al. found that germ-free mice had both a higher preference for sweets and a greater number of T1R2/3 sweet taste receptors in the proximal intestine compared to normal mice. While the absence of intestinal bacteria did not change lingual expression of T1R2/3, the increase of intestinal receptors seemed to have promoted long-term acceptance and preference for nutritive sweet stimuli. This phenomenon was primarily attributed to the post-oral nutrient feedback, reinforcing oral cues or taste associations, thus stimulating further consumption [This role of gut microbes is not limited to sweet tastants. In a similar study, germ-free mice showed an increased preference for intralipid emulsion that was associated with changes in lingual and proximal intestine fatty-acid receptors . In abseTaste buds face a unique challenge against pathogens being exposed to the oral cavity without a strong physical barrier. The presence of commensal microbes in proximity to taste and nutrient-sensing cells is complex and may also lead to an immune response in some instances. Indeed, microbial elements like lipopolysaccharide (LPS) and flagellin can induce inflammatory processes that have effects both locally and systemically. While these inflammatory expression patterns normally protect taste bud and nutrient-sensing cells from pathogens, it may also play a role in the modulating taste perception as well as the pathogenesis of taste dysfunctions .Inflammation is initiated by the activation of Toll-like receptors (TLRs) by microbial pathogen-associated molecular patterns (PAMPs), as well as other inflammatory agents from damaged tissues or stress . TLR sigActivation of the immune system can modulate taste perception by two means: acuity and sensitivity. The average turnover rate of taste cells is 8–12 days; therefore, a continuous supply of differentiated taste receptor cells is crucial for normal taste function . A decreAdministration of LPS is an effective method for mimicking bacterial infection. Depending on where and how LPS is introduced, the immune response in taste buds can vary from hours to days later . Therefo+ taste function, which may lead to heightened Na+ sensitivity during infection [Another method of measuring taste change has been through the systemic injection of LPS. For example, LPS injected intraperitoneally finds its way through mesenteric absorption and circulation to taste bud cells. This route represents an acute bacterial infection and has been shown to lead to the expression of inflammatory cytokine expression  around circumvallate and foliate papillae. This LPS-induced inflammation was shown to inhibit the proliferation of taste progenitor cells and reduce the number of newly born cells entering taste buds. This also moderately shortened the average lifespan of mature taste bud cells . Intrapenfection . Interesnfection .Lastly, the ingestion of LPS is a multifactorial vehicle that mimics consumption of harmful microbes and has been shown to affect taste. The timing of response following ingestion becomes a critical variable to consider. For instance, mice who ingested Gram-positive LPS bacteria exhibited a decrease in neural responses to sucrose during a single overnight period. A decrease in sucrose sensitivity was observed 7 days after injection, in parallel with decreased expression of sweet taste receptors T1R2+T1R3. These results did not occur in acute-, and lingual-treated LPS treated mice nor in TLR4 knockout mice, indicating that ingestion and proper immune responses were necessary to suppress sucrose response. Modulation of certain gut hormones known to affect peripheral taste function was listed as the potential mechanism of communication between the gut and taste bud cells . In addiC. jejuni, produced a rapid anxiety-like response in mice through a vagal pathway [While there are many protective mechanisms preventing LPS from crossing the gut barrier and entering systemic circulation, permeability has been shown to increase from gut dysbiosis induced by high-fat diets . It has  pathway . Since tToll-like receptors play a crucial role in sensing the intestinal microbes via recognition of pathogen-associated molecular patterns (MMAPs) that are derived from various microbes, triggering inflammatory and immune responses. Some TLR, such as TLR4 has been shown to influence anorexigenic signals . For exaHuman taste bud cells secrete a number of diverse peptides as well as express cognate receptors . These pNumerous studies over the years have sought to characterize the gustatory effects of these peptides  and showE. coli proteins also leads to an increase in plasma PYY and GLP-1 levels [i/o signaling; however, FFAR2 also signals Gαq/11 to release intracellular calcium leading to secretion of gut hormones [As stated above, the gut is the largest hormone-producing organ in the body  and hous1 levels . Further1 levels  and actihormones  Recentlyhormones .The model of the gut microbiota as a stimulator of the immune system intersects with its role in hormone regulation . LPS is There are many known factors that drive food choices and habits, with taste considered as one of the main predictors . This rePopular nutrition advice often claims that one can “retrain taste buds” by adhering long enough to a diet low in sugar, salt and fat . Indeed,E. coli) leading to dysbiosis (increased Firmicutes) [Turner et al.  recentlymicutes) . Furthermicutes) ,103,104.micutes) .Another indication that the habitual consumption of certain foods leads to changes in gut microbiota and nutrient-sensing ability is evident from studies on consumption of inulin, a dietary fiber found in plants. A recent murine study by Weninger et al.  found siEating disorders associated with taste responsiveness might lend itself to the influence of gut microbiota . These mInflammatory diseases such as bowel disease (IBD) are associated with alterations in taste sensitivity. For example, in a human IBD case-control study, taste sensitivity was significantly reduced in all tastes except for sour . SimilarClostridium hathewayi, Actinomyces viscosus, Bacteroides nordii, Streptococcus, Rothia, Erysipelatoclostridium and Veillonella along with significant depletion of beneficial commensals such as Lachnospiraceae bacterium, Eubacterium rectale, Ruminococcus obeum, Fusicatenibacter, Eubacterium hallii, Anaerostipes, Agathobacter, Roseburia, Dorea formicigenerans, Clostridium butyricum, Clostridium leptum and Faecalibacterium prausnitzii [Lactobacillus and Bifidobacterium are also decreased in COVID-19 patients. It is not known whether changes in the gut microbiota environment due to COVID-19 are linked with taste changes in infected patients. It is widely known that a significant number of COVID-19 patients report taste changes, as well as changes in the overall oral sensitivity to commonly used condiments and spices. Recently, however, Doyle et al. demonstrated that ACE2 receptors are also present on a subpopulation of Type II cells, PLCβ2 positive, in taste buds, thus facilitating the entry of SARS-CoV-2 virus in the oral cavity [Coprobacillus, Clostridium ramosum and Clostridium hathewayi, correlated with COVID-19 severity. Taste sensitivity is reduced in inflammatory conditions [The SARS-CoV-2 spike glycoprotein (S) binds to angiotensin-converting enzyme-2 (ACE2) receptors that is abundantly expressed in the intestinal enterocytes’ brush border and colonic epithelial cells, supporting SARS-CoV-2 replication . This reusnitzii ,126. Soml cavity . Furtherl cavity ,129. Oranditions . WhetherTaste changes occur in up to 84% of patients undergoing chemotherapy treatments . Recent Several studies have shown that obesity is associated with taste changes, particularly a reduction in taste acuity and taste bud abundance. For example, hypothalamic and brainstem T1R3 and T2R116 taste receptors as well as signaling molecules such as Gα14 and TRPM5 were downregulated by obesity in mice ,138. In bacteroides/lactobacillus ratio compared to high lipid tasters [In a systematic review, Ahmed et al.  conclude tasters  that migTaste plays a significant role in food choices and is the most important driver of food consumption . The inc


A 75-year-old-male with a history of multiple basal cell carcinomas visited our clinic for aesthetic treatment of a large pore in the nasal tip that he noticed 4 months ago. Following pressure application, keratinous material emerged from the center of the lesion . DermoscPore-like or large pore basal cell carcinoma is an atypical and very infrequent clinical presentation of BCC. Both dilated pores and BCC, are tumors of follicular origin. Some authors have postulated that the same stem cell could originate them, so they may coexist in a single cutaneous lesion. Dermatologists must take this into account when making differential diagnoses of solitary enlarged pores, especially when localized on the face. Dermoscopy is an excellent tool for its recognition , 2.


Soft tissue sarcomas (STSs) are rare mesenchymal tumors. With more than 80 histological subtypes of STSs, data regarding novel biomarkers of strong prognostic and therapeutic value are very limited. To date, the most important prognostic factor is the tumor grade, and approximately 50% of patients that are diagnosed with high-grade STSs die of metastatic disease within five years. Systemic chemotherapy represents the mainstay of metastatic STSs treatment for decades but induces response in only 15–35% of the patients, irrespective of the histological subtype. In the era of immunotherapy, deciphering the immune cell signatures within the STSs tumors may discriminate immunotherapy responders from non-responders and different immunotherapeutic approaches could be combined based on the predominant cell subpopulations infiltrating the STS tumors. Furthermore, understanding the immune diversity of the STS tumor microenvironment (TME) in different histological subtypes may provide a rationale for stratifying patients according to the TME immune parameters. In this review, we introduce the most important immune cell types infiltrating the STSs tumors and discuss different immunotherapies, as well as promising clinical trials, that would target these immune cells to enhance the antitumor immune responses and improve the prognosis of metastatic STSs patients. Soft tissue sarcomas (STSs) are a heterogeneous group of rare tumors arising from mesenchymal tissues . STSs caThe American Joint Committee on Cancer (AJCC) staging system for STSs relies on the histologic grade, the tumor size and depth, and the presence of distant or nodal metastases ,8,9. To Patients with STSs are managed according to the generally accepted guidelines and those with localized and resectable diseases are treated by surgery . The maiMetastatic STSs have very limited treatment options . SystemiCancer immunotherapy has changed the treatment landscape in oncology, modified the therapeutic algorithms in multiple malignancies and, furthermore, became the leading treatment for metastatic diseases . With thWhile molecular characteristics of STSs are being urgently investigated among studies, the understanding of the events that occur within the tumor-immune system interplay in STSs are far from satisfactory . The pheIn this review, we introduce the most important immune cell types infiltrating the STSs tumors and discuss different immunotherapies that would target these immune cells to enhance the anti-tumor immune responses and improve the prognosis of metastatic STSs patients.clinicaltrials.gov and search databases. Excluded were clinical trials of unknown status and withdrawn clinical trials. Only English written and peer-reviewed articles published in indexed international journals until June 2021 were reviewed. Databases used for search included Medline/Pubmed, Scopus, and Web of Science. The selection process is summarized in A comprehensive review of the literature on diverse immune cell populations infiltrating the tumor microenvironment (TME) of STSs and a review of therapeutic approaches targeting these cell populations was conducted. Soft tissue sarcoma, T cells, CD8 T (cytotoxic) cells, CD4 T (helper) cells, natural killer (NK) cells, macrophages, T regulatory cells, Tregs, and FOXP3 T cells were used as the key words in the search strategy. The diagnosis of osteosarcoma, as well as Kaposi sarcoma, was excluded. Therapies targeting the particular immune cell population were searched through the official registry at +) and cytotoxic T cells (CD8+) are the two main subpopulations of T cells that control and shape the immune responses in the tumor microenvironment (TME) . To. To66]. + TILs infiltration in the TME and on the expression of immune checkpoint molecules [clinicaltrials.gov). Only one CPI study has reached phase III of clinical testing and is currently recruiting (NCT04741438). In this French randomized prospective multicentre study, STSs patients will receive a combination of nivolumab (3 mg/kg) and ipilimumab (1 mg/kg) for a maximum treatment period 12 months (NCT04741438). The inclusion criteria cover patients with metastatic or unresectable advanced sarcomas of rare subtype.Response to immunotherapy with CPIs largely depends on the level of CD8olecules . Since SOut of the most commonly applied CPIs, nivolumab, pembrolizumab, and ipilimumab entered phase II clinical trials with a great deal of promise .In an observational phase II clinical trial, pembrolizumab was administered in monotherapy to 42 STSs patients with diverse tumor histology (NCT02301039). In this study, one complete response was observed in a patient with undifferentiated pleomorphic sarcoma and a total of seven patients experienced objective responses . To noteclinicaltrial.gov). Nivolumab monotherapy was tested in patients with locally advanced, unresectable, or metastatic sarcoma in a randomized multicenter, open-label phase II study (NCT02500797). In this study, only 5% of the patients receiving nivolumab responded to the treatment whereas addition of ipilimumab to nivolumab therapy led to an increased response rates highlighting a promising efficacy of the combination therapy [Nivolumab is currently being evaluated for the treatment of STSs mostly in combination therapies , and an interesting combination of nivolumab with trabectedin  and oncolytic virus Talimogene Laherparepvec is to be evaluated in sarcoma patients in a currently recruiting phase II clinical trial (NCT03886311). Another phase I/II study in STSs patients aims to evaluate the efficacy and safety of nivolumab, together with ipilimumab and trabectedin as a first line treatment (NCT03138161). Nivolumab is further being evaluated in a combination therapy with chemotherapy , radiation therapy (NCT03307616), and targeted therapy .Efficacy of ipilimumab therapy in STSs is being assessed in phase II clinical trials, including mostly combination therapies . A singlACT is a particular form of cell-based anticancer immunotherapy where both peripheral T cells and TILs can be used for ex vivo expansion and therapeutic administration to the patient . This hiclinicaltrials.gov; July 1st 2021).A modification of traditional ACT is based on the infusion of the patients’ ex vivo expanded T cells after previous genetic modification of T-cells to express a chimeric antigen receptor (CAR) specific for a tumor antigen . This th+CD25+FOXP3-expressing T cells) regulate and suppress other cell types of the immune system. Their modulatory functions include production of cytokines, expression of inhibitory molecules, cytolytic functions, disruption of Ca2+ supply to the effector CD8+ lymphocytes, and multiple other mechanisms causing T cell anergy [+ cells were relatively low as compared to CD4+ and CD8+ cells [+ infiltration while superficial tumors had relatively low FOXP3+ infiltration [+ Tregs infiltration and PD-L1/FOXP3 were significantly associated with overall survival in patients with undifferentiated pleomorphic sarcoma [Regulatory T cells  and, in the context of cancer, NK cells are believed to be the main effector cells of the innate immune system . NK cellranzymes ,82. NK cranzymes . Previouranzymes . In STSsral area .Bücklein et al. have provided a detailed analysis of NK cell presence and function in STSs patients and, unlike in other malignancies, peripheral blood NK cells exhibited a profound impairment in cell function, especially in the intermediate and high-grade STSs . Judge eIL-15 is a potent activator of NK cells with a major importance for NK cell proliferation and survival . The advclinicaltrials.gov, July 1st 2021).NK cells are known for their expression of a wide variety of activation and inhibitory receptors . NKG2A iclinicaltrials.gov, July 1st 2021).TIGIT and CD96 are other NK cell receptors that serve as an optimal target for NK-mediated cancer immunotherapy [Another NK receptor targeting agent is lirilumab, a pan-KIR2D blocker, which also has no ongoing clinical trials in STSs . TIM-3 aotherapy . While Cotherapy . Severalotherapy .Adoptive cell transfer of NK cells has also become a promising treatment approach for advanced and/or metastatic diseases . A phaseMacrophages differentiated from circulating monocytes that efficiently migrate within the tissue and continue trafficking into the TME are called tumor-associated macrophages (TAMs) . TAMs arIn different cancer types, TAMs were associated with poor prognosis and reduced overall survival (OS) . Even thIn a recent study by Dancsok et al., TAMs were investigated in 1242 sarcoma specimens . The stuIn leiomyosarcoma, authors Lee et al. described a significant association between high density of TAMs and worse disease-specific survival . This waSince macrophages represent the predominant cell type in most STSs, different clinical trials have been initiated to either suppress the pro-tumorigenic functions of TAMs or to modulate the natural ability of TAMs to eliminate target cells .CD47 is a transmembrane protein that is highly expressed on neoplastically transformed cells . CD47 bi+ and CD8+ T cells, thus contributing to the tumor regression [A multikinase inhibitor Pexidartinib (PLX3397) is currently being evaluated in a phase I/II clinical trial for the treatment of multiple sarcomas including liposarcoma, leiomyosarcoma, synovial sarcoma, and rhabdomyosarcoma (NCT02584647). Pexidartinib is an inhibitor of proto-oncogene receptor tyrosine kinase (KIT), colony-stimulating factor-1 receptor (CSF1R) and FMS-like tyrosine kinase 3 (FLT3), and was shown to decrease the load of intratumoral TAMs and increase the proportions of CD4gression . Due to gression ,115. ResTrabectedin is a chemotherapeutic drug which was recently approved in the treatment of advanced STSs . TrabectOther possible therapeutic targets include chemokine and chemokine receptors regulating the macrophage trafficking towards the tumor . CCL2-CC+ T cells belong to the main antitumor players in the TME, the infiltration of the TME with CD8+ T cells, and the expression of immune checkpoint molecules have become prerequisites for an effective immunotherapy with CPIs [+ T cells and the highest expression of PD-1 [+ T cells tended to become excluded from the TME which could imply a limited efficacy of CPIs in tumors of greater size [+ T cell infiltration among different studies which is reflected in the greatly disappointing results in clinical trials [Soft tissue sarcomas are rare but mostly lethal mesenchymal tumors . STSs arith CPIs . Pleomor of PD-1 . Therefo of PD-1 . In addi of PD-1 . Interester size . Leiomyol trials . Out of l trials ,123. Thel trials .+ T cells in addition to Tregs [Poor immunotherapy responses in patients with leiomyosarcomas may also be associated with their relatively high loads of Tregs as compared to pleomorphic sarcomas . TME infto Tregs ,80. Datato Tregs ,87. Patito Tregs ,126. Macto Tregs . TAMs arto Tregs . Therapito Tregs ,120.With the current knowledge of diverse immune cell populations infiltrating STS tumors, the ongoing clinical practice could fundamentally change and allow stratification of patients based on the immune landscape of the TME. As distinct histotypes of STSs are recognized to be infiltrated with immune cells in diverse proportions and are only sensitive to specific cytotoxic drugs, it is likely that also preclinical research will focus on deciphering the optimal histology-driven therapeutic regimens.It is already clear that clinical trials aiming at multiple immune cell populations, such as T cells and macrophages that are triggering both the innate and adaptive immunity, bring a great deal of promise. As shown in clinical trials, immunotherapies can be easily combined and chemotherapy can serve as a powerful tool to sensitize TME to immunotherapy in these mostly chemoresistant tumors.


We found that increasing levels of immune infiltration and activation were associated with a generally lower probability of cancer developing ex-PA, suggesting that immune surveillance may constrain the malignant transformation of benign salivary tumors. More immunologically infiltrated tumors were more likely to have developed de novo. Taken together, these data suggest a role for tumor escape from immune surveillance in the development of CA exPA. The immune-cold microenvironments of CA ex PA tumors may in part explain their more aggressive clinical behavior.Pleomorphic adenoma (PA), the most common salivary gland tumor, is a benign tumor that carries a risk of malignant transformation to various histologies of carcinoma ex pleomorphic adenoma (CA exPA). Recently, genomic analyses have provided deeper insights into the molecular biology of salivary gland cancers. However, the molecular processes that underlie the progression from PA to CA exPA are largely unknown. In this study, we used RNAseq data from CA ex PA of myoepithelial ( Salivary gland cancers (SGCs) are rare, aggressive human malignancies that account for fewer than 5% of all carcinomas arising within the head and neck. These difficult-to-treat cancers exhibit a wide spectrum of morphological and histopathological characteristics: more than 20 distinct SGC subtypes arise within salivary gland tissue . TherapePIK3CA and BRAF, ETV6-NTRK3 fusions, and overexpression of Her-2 and androgen receptors [Over the past several years, genomic analyses have provided deeper insights into the molecular biology of SGCs and led to the identification of potentially druggable genetic alterations including mutations in eceptors . Howeverde novo salivary gland malignancies, with a reported 5-year overall survival-rate as low as 25% [Carcinoma ex pleomorphic adenoma (CA exPA) is a rare malignant neoplasm of the major salivary glands, accounting for about 3–15% of all SGCs , 5, withw as 25% .TP53 as well as high expression levels of cyclin D1, p16, COX-2, HGF-A, c-Met, and EGFR [Recent molecular analysis of CA exPA have generated new data shedding light on mechanisms of carcinogenesis. These studies have identified prevalent mutations of and EGFR , leavingn = 40), ACC (n = 20), and SDC (n = 16) [Recently we analyzed RNA sequencing (RNAseq) data from SGC tumors to better understand the immune tumor microenvironment (TME) of these cancers. These analyses have shown that the immune TME varies widely across the different histologic subtypes of SGC. We deconvolved and analyzed the immune microenvironment and neoantigen landscape in 76 SGCs representing the three most lethal histologies: MECA ((n = 16) .On one end of the spectrum, SDCs generally exhibit high levels of immune infiltration with concomitant higher levels of T-cell dysfunction, as well as higher mutational load. In stark contrast, ACCs tended to have an immune-excluded microenvironment, with immunosuppressive cellular populations such as M2-polarized macrophages and myeloid-derived suppressor cells both prevalent, in addition to harboring a very low mutational load. MECAs had a more heterogeneous, but generally intermediate, pattern, with both immune-low and immune-high phenotypes. Taken together, these data provided new insights into the immune microenvironment and neoantigen landscape of SGCs, showing that mechanisms of immune escape appear to differ by histology.To our knowledge, no studies have examined the potential role of the immune microenvironment in modulating the evolution of PA to CA exPA. Here, to evaluate a possible association between immune surveillance and the development of CAexPA, we used RNA sequencing data from the above SGC cohort with a focus on 24 MECA and 6 SDC cases that developed from PA.We used bulk RNAseq data for immune deconvolution applying several orthogonal tools including CIBERSORT, ssGSEA and ESTIMATE as described in detail previously .n = 24) and SDC ex PA (n = 6) and de novo MECA (n = 16) and SDC (n = 10). In general, we observed directionally consistent associations between all aggregate immune metrics and the probability of cancer developing ex-PA (de novo. While these data cannot confirm any specific causal mechanism, they suggest that immune surveillance may constrain the transformation of longstanding benign PA tumors to CA exPA, and that this phenomenon is less relevant in de novo carcinogenesis, where immune escape may be required early on in tumor development.In a logistic regression model controlling for histology type as covariate, we analyzed the association between immunological tumor microenvironment metrics and cancer origin in MECA (ng ex-PA . These dWe then performed an exploratory analysis to determine whether any individual immunological cellular populations or processes were potentially associated with this phenotype . Becausede novo. Because prior studies have indicated that the mutational landscape in PA and CA exPA are largely similar, it is quite possible that immune escape is a necessary step in the malignant transformation of PA tumors, which do not transform so long as they remain under effective adaptive immune surveillance.Taken together, these data reveal a more immune-depleted or immune-excluded microenvironment in MECAs and SDCs that developed from PA compared to MECAs and SDCs that developed These data are consistent with the prior findings reported by Furuse and colleagues, who found high levels of TGFß expression in 10/10 cases of CA exPA; in contrast to absent expression in the epithelial and myoepithelial cells of PA [Interestingly, CA exPA patients tend to be much older than PA patients . Since ide novo counterparts which may, in part, explain their more aggressive clinical behavior.In conclusion, adaptive immune surveillance may constrain the evolution of pleomorphic adenomas to malignancy. CA ex PA tumors have immunologically cold microenvironments compared with their Further research will shed light on the specific immunological mechanisms and cellular populations that mediate this phenotype. As our patient cohort only comprised carcinomas and not adenomas, we can only speculate on the composition of the immune microenvironment in PA and how it evolves over time; ultimately, separate sampling of the PA and carcinoma compartments of a CA exPA tumor may be a more informative technique to dissecting genetic and immunologic factors mediating cancer evolution.de novo carcinomas are more likely to respond to immunotherapeutic approaches, having more infiltrated immune microenvironments; or less likely, having undergone immune escape earlier in their development. As therapies evolve, these data also suggest there may be opportunities for immunotherapeutic interception of PA tumors—if malignant transformation could be prevented or predicted, the practice of immediate surgical resection for all benign PA tumors in the salivary gland may prove unnecessary in the future.We also await further clinical data from ongoing clinical trials that may inform whether SGCs that are


Cardiovascular diseases and cancer are the leading cause of morbidity and mortality globally. Cardiotoxicity from chemotherapeutic agents results in substantial morbidity and mortality in cancer survivors and patients with active cancer. Cardiotoxicity induced by 5-fluorouracil (5-FU) has been well established, yet its incidence, mechanisms, and manifestation remain poorly defined. Ischemia secondary to coronary artery vasospasm is thought to be the most frequent cardiotoxic effect of 5-FU. The available evidence of 5-FU-induced epicardial coronary artery spasm and coronary microvascular dysfunction suggests that endothelial dysfunction or primary vascular smooth muscle dysfunction  are the possible contributing factors to this form of cardiotoxicity. In patients with 5-FU-related coronary artery vasospasm, termination of chemotherapy and administration of nitrates or calcium channel blockers may improve ischemic symptoms. However, there are variable results after administration of nitrates or calcium channel blockers in patients treated with 5-FU presumed to have myocardial ischemia, suggesting mechanisms other than impaired vasodilatory response. Clinicians should investigate whether chest pain and ECG changes can reasonably be attributed to 5-FU-induced cardiotoxicity. More prospective data and clinical randomized trials are required to understand and mitigate potentially adverse outcomes from 5-FU-induced cardiotoxicity. In recent years, important progress in the oncological field has led to an increase in cancer survival rates in high-income countries . CardiovFluoropyrimidines, such as 5-fluorouracil (5-FU) and its oral prodrug capecitabine, are the third most common drug used for the treatment of solid malignancies such as breast, colorectal, head, neck, gastrointestinal, and bladder cancer ,5,6. TheRegrettably, fluoropyrimidines are strongly associated with cardiac adverse events and are the second most common drug associated with cardiotoxicity after anthracyclines . The cliAlthough the pathophysiological mechanisms behind fluoropyrimidines-induced cardiotoxicity are yet to be fully understood, coronary vasospasm and microcirculatory dysfunction most likely play a role in the development of myocardial ischemia and chest pain, which is the predominant manifestation of 5-FU induced cardiotoxicity.This review aims to examine the characteristics and the role of vasospasm and the potential role of microcirculatory dysfunction in patients with fluoropyrimidines-induced ischemic heart disease. The included studies consisted of both prospective and retrospective studies .The most common clinical manifestation of 5-FU cardiotoxicity is chest pain, which can be either nonspecific or anginal and is often, but not always, associated with electrocardiographic (ECG) changes  Table 1Table 1. p-value = 0.0023) in a small cohort in which cardiotoxicity occurred in 7 of the 209 patients receiving their first course of 5-FU [Several potential risk factors for 5-FU chest pain have been suggested, including underlying CAD, older age, and concomitant use of other treatments with cardiac side effects. Patients with pre-existing cardiac disease were at elevated risk of cardiotoxicity  and had fewer cardiovascular risk factors  when compared to those not developing coronary vasospasm. No sex- or race-dependent differences were observed between the two groups. Although in the analysis by Zafar and colleagues, no differences in prognosis were observed between patients with and those without coronary vasospasm, other investigations have observed patients with of severe outcomes, ranging from development of myocardial infarction to sudden cardiac death [Incidence of coronary vasospasm in 2021, a large cohort study by Zafar and colleagues , aimed tac death . The risk of coronary vasospasm depends on the cumulative dose , the schCoronary vasospasm is not a homogeneous entity. The term “vasospastic angina” refers to a condition that may be caused by focal coronary artery spasm or by diffuse spasm of the entire coronary arterial tree, namely vasotonic angina. These two major categories are distinct entities and should be managed differently .Focal epicardial coronary artery spasm may be identified in some patients with angiographically normal or near normal coronary arteries. Focal epicardial coronary artery spasm is often considered a synonymous of variant angina, also called Prinzmetal’s angina. However, it should be recognized that coronary artery spasm is not specific to Prinzmetal angina. On selective coronary angiography, 0.5–0.8% of patients could show evidence of coronary artery spasm stimulated either by the tip of the catheter or the contrast medium . Focal eDiffuse spasm is a manifestation of endothelial dysfunction that alters and enhances vessel reactivity to normal sympathetic stimulation, partly due to reduced shear-mediated nitric oxide (NO) release and excess of reactive oxygen species ,70. A heA study conducted by a research group of the University of Bologna is crucial to demonstrate that vasotonic angina actually is a disorder of the entire coronary arterial tree ,73. PatiIn the case of fluoropyrimidine-induced cardiotoxicity, several case reports and case series reported clinical pictures compatible with variant angina in patients who had a history of angiographically documented coronary artery lesions . HoweverThe evidence of fluoropyrimidine-induced diffuse spasm of the coronary arterial tree is compliant with the hypothesis that endothelial dysfunction or primary vascular smooth muscle dysfunction  is a possible contributing factor to this form of cardiotoxicity. Fluoropyrimidines exert their toxic effect on the endothelium through a plethora of different mechanisms. Cwikiel and colleagues  observedMicrovascular dysfunction appears to be involved in fluoropyrimidine-induced coronary vasospasm. This hypothesis is supported by the evidence of global akinesia or dyskinesia at echocardiography in myocardial regions not supplied by stenotic coronary arteries . HoweverFocal coronary artery spasm can be effectively treated by CCBs and nitrates ,47,48,86A further caveat should be noted. Shimokawa et al.  demonstrIn summary, the heterogeneity in the response to CCBs and to nitrates could explain why there are variable results after administration of CCBs or nitrates in some patients with 5-FU presumed to have myocardial ischemia. Induced chest pain persisted after the administration of some types of CCBs or nitrates. One point is critical. Testing that separates those patients whose symptoms are due to myocardial ischemia from those whose pain is non-ischemic is strictly necessary. Even minimal atherosclerotic disease on angiography (or intravascular ultrasound imaging) warrants risk-factor modification and prevention therapies.Cardiotoxicity related to 5-FU administration is a poorly understood but relatively common clinical entity that deserves special consideration given the frequent use of this agent and the cardiac complications associated with its use. The mechanism of 5-FU-related cardiotoxicity may occur from a combination of ischemia related to epicardial coronary vasospasm and microvascular dysfunction. Further clinical studies in humans are required to clarify these mechanisms. Patients with 5-FU based chemotherapy who develop vasospasm are difficult to manage. Patients who present with chest-pain symptoms should be routinely referred to cardio-oncology for optimization of their medications. Patients should receive a baseline ECG. Patients should also be instructed to stop therapy and to call the emergency department if experiencing any chest discomfort at home. Clinicians should determine if further 5-FU is required or whether acceptable alternative treatments can be safely used. When further doses of 5-FU are necessary, clinicians should proceed cautiously. Management of the affected patients should focus on separating patients whose symptoms are due to myocardial ischemia from those whose pain is non-ischemic. Coronary angiography or computed tomography angiography is required as it would seem prudent to perform a sensitive screen for significant fixed coronary disease prior to performing any provocative testing for a diagnosis of coronary spasm. Patients with normal or near-normal coronary arteries may undergo ergonovine or acetylcholine tests, which would result in a better characterization of the vasospastic disease . The posIn summary, the evidence of coronary vasospasm and microcirculatory dysfunction in 5-FU-induced ischemic heart disease is still lacking. Physicians would consider urgent coronarography for patients presenting with suspected acute coronary syndrome and computed tomography angiography for low-risk patients with persistent minor or mild symptoms. Additional provocative tests should be performed in patients with unobstructed coronary arteries. Such an approach permits discrimination of variant angina and vasotonic angina along with coronary microvascular disorders versus non-cardiac chest pain, which permits distinct treatments outlined in consensus practice guidelines. When further doses of 5-FU are required, clinicians should proceed cautiously and wait for a definitive diagnosis of coronary artery disease.