The promise of precision and personalized medicine is rooted in accurate, highly sensitive, and specific disease biomarkers. This is particularly true for cancer-a disease characterized by marked tumor heterogeneity and diverse molecular signatures. Although thousands of biomarkers have been described, only a very small number have been successfully translated into clinical use. Undoubtedly, there is need for rapid, quantitative, and more cost effective biomarkers for tumor diagnosis and prognosis, to allow for better risk stratification and aid clinicians in making personalized treatment decisions. This is particularly true for cancers where specific biomarkers are either not available  or where current biomarkers tend to classify individuals into broad risk categories unable to accurately assess individual tumor aggressiveness and adverse pathology potential , thereby leading to problems of over-diagnosis and over-treatment of indolent cancer and under-treatment of aggressive cancer. This perspective highlights an emerging class of cancer biomarkers-live-single-cell phenotypic biomarkers, as compared to genomic biomarkers, and their potential application for cancer diagnosis, risk-stratification, and prognosis. Broadly, successful biomarkers can be classified into the following categories: screening–is the patient at risk of a disease; diagnostic–does the patient have a disease; prognostic–how aggressive is the patient’s disease; theranostic/companion diagnostic–which therapy or procedure will best cure or mitigate morbidity and mortality of a patient’s disease; and monitoring–has the disease been treated effectively, recurred or spread. Live cell biomarkers are poised to be a powerful addition to available biomarkers and can be assessed either via slice cultures,5 Importantly, these static biomarkers were observed in the context of formalin-fixed tissue samples that do not lend themselves to live-cell single cell analysis.5 Technological advances have improved the ability to define cellular phenotypes, and this previous narrow definition of phenotypic biomarkers to only include cell shape and proteins, is now incomplete. Analyses of live single-cell behavior suggest that dynamic biophysical cellular properties characterize both normal and disease states and represent a powerful extension and expansion of the concept of phenotypic biomarkers.6 Thus, a working definition of phenotypic biomarkers may be expanded to consider spatial and temporal considerations, including two phenotypic biomarker sub-categories: molecular and cellular , live-cell imaging, and functional magnetic resonance imaging. Each imaging modality, and related modalities employ different probes with their own binding kinetics that dictate the sensitivity and specificity of their respective temporal and spatial resolutions, engendering different ideal applications such as screening, diagnostics, prognostics, theranostics (or companion diagnostics), and monitoring, as well as potentially predicating their success in a given application.8 Given recent advances in our understanding of the tumor environment, microfluidics, high-content screening coupled with machine vision and big-data techniques (such as machine learning) it is conceivable to postulate a paradigm shift in the way patient tissue samples are analyzed9 with a transition of qualitatively assessing static, fixed tissue towards a modality of quantitatively analyzing live tissue. Further, the investigation of dynamic live-cell phenotypic biomarkers either in native micro-extracellular environments or specifically engineered extra-cellular environments may allow the level of interrogation, dissection and delineation of complex disease signaling pathways towards major clinical and therapeutic advances. As an example, probing cell-extracellular matrix (ECM) interactions or the manner in which cells interpret the adjacent microenvironment, is emerging as another ‘–omic’-like strategy to assess disease.10With that, live-cell phenotypic biomarkers represent an un-tapped wealth of biomarkers due to difficulty around keeping biopsy tissue a live in vitro, as well as automated imaging and big-data/high-content data analysis techniques.11 Integrins and focal adhesion proteins promote cellular growth and motility and are implicated in the ontogeny of epithelial tumors including prostate cancer.12 Focal adhesion protein complexes are coupled with the forward (protrusive) and rearward (retrograde) movement of actin.13 This interaction in turn regulates cell motility, growth, and proliferation.15 Thus, the number and size of focal adhesions are indicative of malignancy and metastasis, and are emerging to be informative as molecular phenotypic tumor biomarkers.18 Several studies have reported a strong association between cancer progression and aberrant expression/activity of proteins involved in cell-cell adhesion, such as Cadherins and Catenins.20 Morgan et al. report that abnormal expression of α and β Catenin, and Claudin 7 could serve as biomarkers to distinguish localized from metastatic prostate cancer.20 Broadly, as human cells transform from being benign to malignant, they demonstrate an increasingly disorganized cytoskeleton, decreased peripheral cellular actin and changes in cell adhesion properties.22 The concentrations and molecular architecture of various cytoskeletal components determine the overall mechanical response of cancer cells and therefore hold promise as phenotypic biomarkers.23Specifically, integrins, the primary mechano-transducers between cells and the ECM, recruit proteins to sites of cell–ECM contacts known as focal adhesion complexes.25 Examples include circulating proteins in blood/urine for human cancer screening, , functional proteins in fixed tissue samples for diagnostic and prognostic purposes  and specific circulating proteins to monitor response to treatment and disease progression .26 Dynamic molecular phenotypic biomarkers remain an untapped resource for cancer diagnosis and risk stratification, as well as have the potential to serve as biomarkers for screening, diagnosis, prognosis, assessment of drug response and in companion diagnostics and disease state monitoring  molecular phenotypic biomarkers have been extensively utilized in research to study the effects of genetic alterations and therapeutics in cancer cells. However, their clinical applications have been limited with only a handful in current clinical use.ing Fig. . This obImportantly, shifting from a fixed/static biomarker molecular measurement to a live/dynamic measurement may indeed provide both the spatial and temporal resolution needed to maximize molecular phenotypic biomarker information. For example, label-free quantification actin polymerization via membrane fluctuations, in that it relates to protrusive forces and motility, may be an important biomarker to characterize metastatic potential of a given cell.27 mechano-transduction28-the conversion of mechanical forces into biochemical signals29-contributes to developmental,30 physiological, and pathological processes in cancer progression.32 Work is ongoing to apply microfluidic technologies to create tools that can “squeeze” or mechanically stimulate cancer cells in suspension, derived from blood or urine samples, and thereby assess cellular mechanical properties such as contractility and force generation.33 Important recent advances in matrix biology, live single-cell imaging, microfluidics and automated image analysis have made it possible to maintain cancer cells ex vivo, and quantify their mechanical or biophysical properties as “label-free” phenotypic biomarkers.34Similar to molecular phenotypic biomarkers, cellular phenotypic biomarkers may benefit from the incorporation of dynamic data, on a single cell level, as well as a tissue-based level. For example, and as tumorigenesis is a complex biological process that involves alterations in the mechanical properties of cells,35 or from changes in complex biochemical pathways e.g., aberrant androgen signaling in prostate cancer results in altered cell migration.36 Cell spreading is another actin-dependent feature related to cellular invasiveness.37 Cell spreading is also correlated with altered DNA synthesis, motility, and differentiation.21 Cell deformability (or effective stiffness) is another cellular biomarker used to categorize cell populations and is an end result of multiple biochemical and molecular changes.34 Data from several epithelial cell carcinomas, including ovarian and breast, suggest that decreased cell stiffness or increased cell deformability is indicative of malignant transformation and may be a useful measure of metastatic potential.39 Table The aforementioned advances have made it possible to measure cellular phenotypes that take multiple snap-shots of important cellular features that are the resultant actions of multiple integrated biochemical signaling pathways, thereby interrogating a single cancer cell’s signal transduction profile to assess if it is normal or pathological. For example, cancer cells have altered migration rates compared to non-cancer cells from the same tissue. Changes in migration velocity can result from underlying cytoskeletal changes such as shortening of F-actin filamentsAs many signaling events that regulate oncogenesis and metastasis are governed by dynamic cytoskeletal events that occur on a sub-micron (sub-cellular) spatial scale and a (sub) milli-second temporal scale, the addition of quantifiable live-cell biomarkers on similar spatial and temporal scales may be useful as a powerful addition to the current suite of cellular phenotypic biomarkers towards ameliorating current clinical applications.Conventional histopathology is the current gold standard for diagnosis of most cancers . Based on the qualitative and subjective evaluation of tissue specimens, routine histopathology suffers from variability in diagnosis, suboptimal sensitivities and the inability to distinguish indolent from aggressive cancers. The relative recent adoption of immunolabeling/IHC for a handful of protein biomarkers  has improved the predictive power of histopathology by providing information about tumor subtypes, prognostication and even guiding treatment decisions. However, IHC remains a semi-quantitative method at best, is time consuming and relies on limited tissue sampling and the pathologist’s subjective visual interpretation of staining intensity.43 Genomic tests measure selected genetic loci and can be performed on static/formalin fixed paraffin-embedded tissue samples. Predictions of tumorigenicity (prognostication) are based on the quantitative gene expression data of a selected but small number of genes. Although innovative, genomic marker tests suffer from poor “signal-to-noise” ratio due to the significant tumor heterogeneity from bulk-tumor samples that occur in solid tumors such as prostate and breast cancer.46 Genetic tests are designed to detect mutational hotspots in cancer-related genes and pose a risk of false negative results when used for testing cancers with unknown mutations.47 This is reflective in the relatively low sensitivities (<0.70), or other test performance statistics (like area under the curve or AUC) reported in clinical studies for many of the marketed genomic biomarker tests.40In an attempt to make diagnosis and prognosis more standardized and accurate, several diagnostic tests based on quantification of genomic cancer biomarkers have been developed.48 A study has shown the use of a well-defined ECM allows for survival and growth of primary prostate cells derived from radical prostatectomy tissue.48 Further, the use of a defined ECM environment provides a reference standard to gage and compare how primary cells from different patients interpret a specific and consistent microenvironment. More work in this field is ongoing and shows promise towards the ability to rapidly and reproducibly culture cells derived from patient biopsy.49 Additional studies have also been successful at controlling media and ECM conditions, to establish slice cultures, organoid cultures or slow growing (~30 day) single-cell cultures.52Given the importance of maintaining in vivo cell behavior in vitro and time to culture, traditionally it has been difficult to culture single cell cultures of primary human cells derived from biopsy tissue in vitro for clinical applications. Advances in our understanding of integrin-mediated ECM interactions has enabled rapid culturing of primary human cells (such as prostate) allowing analysis of cells as early as 48 h after sample procurement with the ability to maintain primary cells in vitro for at least 7 days.Live-single-cell cellular and molecular phenotypic biomarkers represent an easily quantifiable result or biomarker representative of a concert of biochemical pathways or the total expression of the complex and inter-related cellular processes that regulate cellular homeostasis and pathogenesis , epigenetic, chromosomal, proteasome, metabolome, etc.). Imaging biophysical biomarkers and monitoring protein expression, sub-cellular localization, and modification states in living cancer cells on a single-cell level may allow direct assessment of cellular function and tumor heterogeneity by quantifying molecular and cellular behavior in individual cells.53 Assessment of phenotypic behavior at the resolution of individual cancer cells is advantageous, as it considers functionally important discrete sub-populations of cells that may dictate tumor biology that, otherwise, would be lost in bulk genomic analysis. Furthermore, single-cell analysis allow measurement of the intra-tumor heterogeneity, which itself could be used as a biomarker to predict disease progression and drug resistance.54 Moreover, live-single-cell phenotypic biomarkers provide direct measures of functional biological pathways providing vital information such as protein expression, localization or activity. Genomic tests on the other hand do not provide information about protein localization and infer data about protein expression and activity through gene expression (RNA) profiling, which have been shown to have minimal or limited correlations.55Specifically, rapid and quantitative measurement of live-cell phenotypic biomarkers using a microfluidic platform and trained, automatic machine vision, and learning algorithms could provide meaningful, objective information about the invasiveness and growth potential of tumor cells.Table 56 Cancer diagnostics is moving quickly from reliance on histopathology and immuno-histochemical tissue staining towards the use of quantitative, biomarker-based, personalized approaches , live-cell phenotypic biomarkers have potential uses in drug screening, companion diagnostics and lead compound discovery.62 a number of techniques are emerging that use high throughput microfluidic devices to discern single cell phenotypic behavior often focusing on cellular mechanics such as migration, traction force, cell tensile strength, and extra-cellular matrix / microenvironment cell interaction.73 Additionally, there are emerging technologies that apply single cell analysis techniques to for improved screening, diagnosis, prognosis, and therapy selection, as well as drug discovery and development that are extensions of more established techniques that use cellular phenotype profiling.75Indeed, motivated by the need to dissect and interrogate the heterogeneity and multiplicitous nature of many cancers,76 Live-cell phenotypic biomarkers could also be used to design and study the effect of combination therapies and the new class of immune-oncology drugs supporting the potential paradigm shift in cancer diagnosis and treatment in the era of (live-cell) personalized medicine.77Live-cell phenotypic biomarkers could be used as a theranostic (or via a companion diagnostic) approach in which an individual’s live cells could be screened against a panel of chemotherapeutic drugs (singly or in combination) to provide a direct read-out of tumor response, again, given a defined ECM, to drug treatment prior to use in oncology drug trials and clinical practice.76 Using a high-content, automated live cell imaging, phenotypic assay, Pau et al was able to screen cells treated with a small interfering RNA library and identify 2190 genomic targets that disrupted the cell cycle, mitosis, or cell death pathways.78 Indeed such large scale screening approaches utilizing live cell phenotypic assays are poised to usher in a novel approach to interrogate cancer cell biology by allowing computer generated models to be developed for studying cancer biology in-silico.79General biology is often studied using cellular phenotypic assays as screens to test gene function and compare cancer cells with non-cancer cells as a method to better understand the underpinnings of cancer cell biology.While much progress has been made in identifying biomarkers in research settings, clinical adoption has been slow. Thus, there remains a significant unmet need for specific and sensitive biomarkers of tumor aggressiveness and metastatic potential that can be used for optimal risk stratification towards treatment decision-making.Phenotypic biomarkers, particularly those evaluating live-cell behaviors, have the potential to fulfill this need for a quick and actionable biomarker platform to underpin the precision oncology paradigm. The underlying advantage of live-cell phenotypic biomarkers (molecular and cellular) over genomic biomarkers is their universal applicability in multiple solid human tumors, their ability to report on multiple, synchronous biochemical pathways, and their reflection of a cancer cell’s protein expression profile in concert and behavior in its microenvironment. Furthermore, recent technical advances allow for rapid and automated measurement of phenotypic biomarkers in large numbers of live tumor cells enabling big-data analysis at the single cell level to mitigate the challenges of tumor heterogeneity.88Precision oncology stands to benefit enormously from the development of a live-cell phenotypic biomarker (molecular and cellular) platform using high throughput automated approaches. A composite measurement of live-cell phenotypic biomarkers in cancer represents an innovative and actionable addition to personalized cancer diagnosis, risk stratification, and treatment decision-making.


Sphagnum-dominated mires, which had experienced different kinds of disturbances in the past 2000 years – fire and peat extraction, respectively. We tested the effect of disturbances on the linkages between TA community structure, functional trait composition and functional diversity using structural equation modelling. We found that traits such as mixotrophy and small hidden apertures (plagiostomic apertures) are strongly connected with disturbance, suggesting that these two traits can be used as palaeoecological proxies of peatland disturbance. We show that TA functional traits may serve as a good proxy of past environmental changes, and further analysis of trait-ecosystem relationships could make them valuable indicators of the contemporary ecosystem state.Species’ functional traits are closely related to ecosystem processes through evolutionary adaptation, and are thus directly connected to environmental changes. Species’ traits are not commonly used in palaeoecology, even though they offer powerful advantages in understanding the impact of environmental disturbances in a mechanistic way over time. Here we show that functional traits of testate amoebae (TA), a common group of palaeoecological indicators, can serve as an early warning signal of ecosystem disturbance and help determine thresholds of ecosystem resilience to disturbances in peatlands. We analysed TA traits from two  Understanding the response of ecosystems to climate and human driven processes under recent global change conditions is an important challenge for scientists145711Sphagnum and vascular vegetation cover132 emission9Anthropogenic activities, such as peat extraction, drainage and burning, are among the most common water disturbances in peatlands leading to substantial shifts in per se2129Among the different approaches used in palaeoecology, a functional trait-based approach is a useful tool to track ecosystem response to climatic and anthropogenic change2022Still, there is a scarcity of studies focusing on the long-term impact of disturbances on testate amoeba trait composition, in particular concerning effects of human-induced disturbances. Therefore, our primary goal is to examine whether the response of TA to various environmental disturbances over the past 2000 years is reflected in their trait composition. We aim to assess differences between TA trait groups in response to two disturbance agents: fire and peat extraction. We also want to define the relationship between the taxonomical and functional diversity for TA. We hypothesised that TA community composition responds to abrupt disturbance. In particular, we expected that certain traits, such as species size or aperture position, reflect shifts in peatland hydrology, based on recent findings studying the effect of water level changesSphagnum peatlands located in Poland: Linje36th centuryth to the beginning of 19th century AD (this layer does not have an absolute chronology due to radiocarbon date inversion)3638To investigate the long-term response of testate amoeba communities to disturbances, we used existing palaeoecological data sets from two Selected peat cores cover the last 2000 years, and possess TA community data and associated depth to water table (DWT) reconstructions, and macroscopic charcoal, plant macrofossil and pollen data1336diversity indices of the testate amoeba functional groups within the community; and (2) functional traits of the testate amoeba community.diversity indices for testate amoeba functional groups  along the sequences were calculated for selected TA groups  Q value, 43Q is unit-less and shows how distinct communities are functionally different, i.e. increasing Rao’s Q values indicate functionally increasingly different communities. FDQ was calculated using the synthetic functional traits  axis) defined above and the relative abundance of speciesQ.Based on the TA functional traits, we calculated the community weighted mean values (CWM) of each standardised trait, which is an index of functional composition expressed as the mean trait value of species present in the community, weighted by their relative abundancesa priori knowledge, we developed several hypothetical mechanisms to build a network of causal relationships including predicted effects of disturbancesQ). We used pollen human indicators as a proxy for peat extraction and macroscopic charcoal influx as a proxy for fire. For each core, we ran a DCA on a TA data set and extracted DCA site scores as a proxy for TA community composition. Similarly, we ran a PCA on CWM traits of TA of each core and extracted PCA site scores as a proxy for trait community composition. This model was tested for fire (n = 146) and human (n = 35) disturbances, respectively. Adequate model fits are indicated by non-significant χ2 tests (P > 0.05), a low Standardised Root Mean Square Residual index (SRMR < 0.1), and a low Root Mean Square Error of Approximation index (RMSEA < 0.1). Detailed pollen data used for both profiles can be found in Kołaczek, et al.et al.FDsemveganTo see if functional diversity of TA communities responds similarly to disturbances in both datasets, redundancy analysis (RDA) was performed using six functional traits of the TA community and DWT, macroscopic charcoal influx and pollen indicators linked to human activities, i.e. released by ruderal plants as explanatory variables , Table 2The compositional change of TA showed a drastic turnover in both peatlands in response to disturbances; however, this occurred with a delay . In LinjCompared to the community response, the response of some groups was delayed. The change of shell size in response to disturbance is clearly visible in Linje, where at the beginning of local fire activity richness and evenness of small species increased . Simultaneously, evenness and richness of large TA slightly declined. In general, richness and evenness of large TA followed the community trend, whereas in the group of small TA evenness and richness varied before the fire, and after the fire they show similar tendencies. In PK, evenness and richness of large TA followed the community trend up to the disturbed layer; together with disturbance and post-disturbance evenness and richness of small TA was in line with the community response.Species with acrostomic apertures were the most abundant in both data sets. In Linje, evenness and richness were very similar to community trends; during fires values decreased, and increased post-disturbance. Similarly, in PK richness and evenness followed community trends. In both peatlands, TA possessing axial apertures were highly variable. In pre-disturbance communities, TA with plagiostomic aperture were either not present in the community (PK), or their abundances were low (Linje).In Linje, evenness and richness of TA preferring dry and intermediate water tables were variable pre-disturbance , but during local fires indices values were more stable and began to change post fire. Evenness and richness of wet indicators were more stable post-disturbance, having the biggest variations during and after disturbances. In PK, the highest pre-disturbance  richness and evenness were observed for TA preferring intermediate and low water tables. Evenness and richness of all groups  decrease in the layer disturbed by peat extraction.Compositional change and changes in evenness and richness of TA preferring lower pH were consistent with the general community trends in both peatlands. Species preferring higher pH were less abundant and in PK compositional change was observed in the post-disturbance period. In Linje, the group of TA preferring high pH shifted during disturbances and stabilised post-disturbances.Q values decreased after disturbances started (stronger decreases are noted for PK), indicating that TA communities were functionally more similar during disturbances. In general, this shift was mainly caused by a decline in mixotrophs and body size  of TA functional diversity, while human disturbances explained 52% (R2 = 0.52) of TA functional diversity  . The effiversity . In LinjThe analysis of TA functional traits revealed similar tendencies in both peatlands , comparaHeleopera petricola323439Analysis of TA functional groups revealed that richness and abundances of small TA and species preferring dry conditions were higher just after the influence of local fire, but decreases in the abundance of large TA and those preferring wet conditions were recorded with a short delay. This delay in response of wet indicators was also recorded in the cross-correlation analysis, i.e. of macroscopic charcoal influx with wet indicator species Sphagnum peatlands the most common Euglyphids are Euglypha sp., Assulina sp., Trinema sp. and Corythion sp.Euglypha sp. are built of easily dissolvable silica platesTrinema sp. and Corythion sp. taxa were present in Linje and PK TA communities during and after disturbances, which suggests that small species do indeed colonise available niches rapidly and that they are not discouraged by tough environmental conditions.During the post-disturbance period in PK, large TA reappeared in low abundances and their evenness and richness decreased. In Linje, however, large TA got richer in the final stage of disturbances, but when fire activity decreased their evenness and richness decreased again. When disturbances were terminated, larger TA were not as abundant as before the disturbances and failed to dominate the community. This may be due to mire size, as small mires (e.g. Linje) possess low levels of variety and numbers of microhabitats. Therefore, when TA species become extinct, individuals from other habitats cannot colonise quickly. In contrast, in larger mires with variable microhabitats present and bigger and more diverse species pool for potential immigrants, such as PK, the possibility for faster colonisation from other microhabitats present on the peatland surface is higher. Another important factor is species size. Large species are heavier than smaller ones53et al.Sphagnum and increase in abundance of Eriophorum vaginatum was recorded, and later TA were affected  were recorded in both peatlands  in Linje can be connected not only with water table lowering, but also with dust deposition during local fires  in dry periods and/or local afforestationMoreover, our findings show that mixotrophy and plagiostomic aperture traits were connected with local disturbances and, hence, we suggest that these traits could be considered disturbance indicators in palaeoecology. Mixotrophy was connected with pre-disturbance communities, where in periods of high water tables high numbers of mixotrophs (mainly eatlands . MixotroTrinema sp. and Corythion sp.) were present in very low numbers during the short drought episode cal. AD 700 in Linje and were not present in PK before disturbances. They became more abundant when disturbances occurred and were established in TA communities post-disturbance. Plagiostomic apertures, small and hidden in the shell side13et al.et al.Plagiostomic apertures were clearly connected with disturbances and the post-disturbance period . Small sUnder global change conditions, ecosystems are exposed to drastic shifts; hence, recognition of useful indicators of disturbances might be essential to assess the state of ecosystem functioning. We revealed that testate amoebae morphological traits were linked to environmental conditions over a long temporal perspective: (1) testate amoebae preferring dry conditions tended to prefer lower pH, possessed small shells and hidden plagiostomic apertures which are an adaptation to low water content, whereas (2) testate amoebae preferring wet conditions tended to prefer higher pH and possessed larger shells with acrostomic aperture. We underline the sensitivity of mixotrophs to abrupt disturbances. We show that species possessing plagiostomic apertures were indicators of disturbances and that their appearance during dry phases allowed their constant presence in post-disturbance communities. Moreover, disturbances caused substantial body size decreases. The above-mentioned changes in TA communities show that human impact and fires had long-term effects on peatland communities. Our results suggest a low ecosystem resilience of peatlands that – once disturbed – may not be self-restored rapidly within a few decades. Changes in trait composition affected by fire and/or peat extraction are seen in post-disturbance communities, i.e. through the establishment of new traits (plagiostomic aperture) or a lack of previously abundant traits (mixotrophy).Our results show that testate amoebae functional traits may serve as a valuable proxy that supports conventional reconstructions of the past environmental changes in peatlands, and further analysis of trait-ecosystem relationships could make them valuable indicators of the contemporary ecosystem state. Moreover, the trait approach for testate amoebae requires more attention and more studies focused on the complex evolutionary links between testate amoebae morphology and ecosystem function.How to cite this article: Marcisz, K. et al. A novel testate amoebae trait-based approach to infer environmental disturbance in Sphagnum peatlands. Sci. Rep. 6, 33907; doi: 10.1038/srep33907 (2016).


Modeling brain disorders has always been one of the key tasks in neurobiological studies. A wide range of organisms including worms, fruit flies, zebrafish, and rodents have been used for modeling brain disorders. However, whether complicated neurological and psychiatric symptoms can be faithfully mimicked in animals is still debatable. In this review, we discuss key findings using non-human primates to address the neural mechanisms underlying stress and anxiety behaviors, as well as technical advances for establishing genetically-engineered non-human primate models of autism spectrum disorders and other disorders. Considering the close evolutionary connections and similarity of brain structures between non-human primates and humans, together with the rapid progress in genome-editing technology, non-human primates will be indispensable for pathophysiological studies and exploring potential therapeutic methods for treating brain disorders. Psychiatric disorders are a group of mental illnesses without detectable pathological symptoms such as neuronal death. They include schizophrenia, depression, bipolar disorder, and autism spectrum disorders (ASDs). The onset of most psychiatric disorders is normally in adulthood. However, the onset of ASDs is during the first 3 years of life. Therefore, ASDs are also considered to be developmental disorders, possibly caused by disruption of neural development leading to abnormal social behaviors and other symptoms .With recent advances in human genetics and neurobiological studies, ASDs have been found to include both genetic and environmental components. Particularly, mechanistic studies in animal models have provided tremendous insights into how genetic and environmental factors contribute to neural development and underlie the proper functioning of cognitive and social behaviors. A wide range of animal models including worms, fruit flies, zebrafish, and rodents have been used to study neural development and disorders. However, whether complicated neurological and psychiatric symptoms can be faithfully mimicked in animals other than primates is still debatable. In this review, we discuss technical advances for studying the neural mechanisms underlying ASDs and other brain disorders using non-human primates as animal models. Considering the close evolutionary connections and similarity of brain structures in non-human primates and humans, together with the rapid progress of genome-editing technology, non-human primates will be indispensable in pathophysiological studies and potential interventions for psychiatric disorders .The general approaches used in rodents for modeling psychiatric disorders, such as chronic restraint stress, chronic unpredictable mild stress, and chronic social defeat stress, are efficient for inducing anxiety-like behaviors for various periods of time. However, for ethical reasons, most of these approaches are not practical in non-human primates. Under these circumstances, researchers have designed mild stress models, in which monkeys are subjected to gentle stresses. Kalin and colleagues have established various stress models in monkeys, followed by assessment of physiological and neurochemical parameters . After bWith regard to the close relationships between monkeys and humans, whether complex environmental factors can be applied to monkeys is a critical question to address. Hu and colleagues have designed a paradigm in which maternal separation paradigms are mimicked in monkeys . They inMacaca mulatta) oocytes with a retrovirus harboring the green fluorescence protein (GFP) gene, followed by in vitro fertilization (IVF) with sperm. After transplantation of fertilized eggs into surrogate monkeys, researchers successfully acquired transgenic monkeys carrying the GFP gene. This pioneering work launched a new era of genetic engineering in non-human primates.Over the years, genetic approaches have been widely applied in model organisms, such as worms, fruit flies, and rodents, to establish models of brain disorders. Due to the difficulty of handling germline cells and the unavailability of embryonic stem cells, genetic engineering in monkeys did not become available until 2001 . In thisTaking advantage of the high efficiency of lentiviral infection, Li, Chan, and colleagues at Emory University used lentivirus harboring the disease-causing Huntingtin (HTT) mutant gene to infect rhesus monkey oocytes . After ICallithrix jacchus). Sasaki and colleagues also used lentiviral-based methods to produce transgenic marmosets carrying GFP transgenes [Another pioneering work in genetic engineering for non-human primates was done in marmosets  . AlthougMECP2 and examined social and motor behaviors associated with social interaction and cognition. They found that MECP2 transgenic monkeys indeed exhibit abnormal behaviors, including repetitive locomotor activity, elevated anxiety levels, and defects in social interactions, which nicely mimic ASDs in human patients [Due to the long reproductive cycle of macaques , obtaining a second generation is time-consuming and costly. In recent work on making monkey models of ASDs, Qiu and colleagues used lentivirus-based transgenic methods to produce transgenic monkeys with the autism gene patients . Remarkapatients . This woMacaca fascicularis) and transplanted them into surrogate monkeys [With the emergence of the CRISPR-Cas9 and TALEN genome-editing methods from 2011, genetic engineering in non-human primates became feasible and relatively easy. The landmark work was done by Niu and colleagues, who injected Cas9 mRNA along with single-guide RNA (sgRNA) into the fertilized eggs of cynomolgus monkeys  . AlthougMECP2 (methyl-CpG binding protein 2) gene in cynomolgus monkeys [MECP2 gene. However, whether genomic mutation of MECP2 leads to behavioral changes similar to human Rett syndrome remains to be determined [To model Rett syndrome, a developmental disorder, investigators used TALEN-based genome-editing to target the  monkeys , 16, andtermined .The rapid progress of genetic engineering in non-human primates provides researchers with a wide platform to study primate brains at an unimaginable scale. It is likely that brain imaging and cognitive studies using genetically-engineered monkeys will be available soon, and will provide causal links between genes and brain structure in the non-human primate. An elegant example is the work by Wang and colleagues, who investigated the effects of ketamine on the functional brain network using magnetic resonance imaging .Although ketamine, a non-competitive N-methyl-D-aspartate receptor antagonist, has rapid and prolonged antidepressant effects in depressed patients after a single dose, the underlying mechanisms by which ketamine affects brain structure in patients are unknown. Given the lack of models of depression in non-human primates, researchers took another approach by giving rhesus monkeys a single dose of ketamine, and examining the functional connectivity of the brain using magnetic resonance imaging. They found that ketamine treatment leads to large-scale downregulation of functional connectivity, which is precisely opposite to the brain network in depressed patients, suggesting that ketamine has antidepressant effects by affecting the global brain network. This work remarkably illustrates the value of cutting-edge technology such as brain imaging in studying the neural mechanisms underlying brain disorders.We are in an era when multiple disciplines have interacted to elucidate the mechanisms of complicated human brain disorders. Given the complexity of the brain, it has always been difficult to recapitulate the core symptoms of human brain disorders in animal models. With the rapid development of genetic-engineering technology, such as lentiviral transduction and genome-editing, it is foreseen that non-human primates will play a crucial role as animal models in the near future. It is worthwhile noting that current therapeutic methods for human brain disorders such as deep brain stimulation and transcranial magnetic stimulation will also need to be tested in non-human primates prior to human trials. Therefore, non-human primate models of brain disorders are also indispensable for translational studies. Together with the technical advances in brain imaging and non-invasive neural modulation methods such as ultrasonic stimulation, non-human primates will play more competitive roles in modeling brain disorders and serve as a platform on which researchers can seek potential therapies.


This consensus statement is directed to intensivists, hematologists, and oncologists caring for critically ill cancer patients and focuses on the management of these patients.The online version of this article (10.1007/s00277-018-3312-y) contains supplementary material, which is available to authorized users. Cancer is the second leading cause of death worldwide. According to recent publications, the incidence of neoplasms is expected to increase requiring more health care resources. Appropriate allocation of resources for cancer prevention, early diagnosis, as well as curative and palliative care remain of particular importance. This requires a detailed knowledge of the local burden of cancer .In patients with hematological or oncological malignancies, the disease itself or intensive therapy may result in critical illness in a substantial number of patients. In 1999, the American College of Critical Care Medicine stated that patients with hematological or metastasized oncological malignancies are poor candidates for ICU admission with a mortality rate of up to 90%. Thus, refusal of ICU admission of these patients was the common procedure . HoweveThe consensus group was selected to represent the expertise in the field of critical care in cancer patients in Germany and Austria. Experts were sent by the German Society of Hematology and Medical Oncology (DGHO), the Austrian Society of Hematology and Oncology (OeGHO), the German Society of Internal Intensive and Emergency Medicine (DGIIN), and the Austrian Society of Medical and General Intensive Care and Emergency Medicine (ÖGIAIN). All authors are actively supporting the “Intensive Care Medicine in Hematologic and Oncologic Patients (iCHOP)” Initiative (info@ichop.eu).Consensus statements are based on an in-depth PubMed literature retrieval of English as well as German language literature with last access February 01, 2017. For medical heading terms (MeSH) identifying potential relevant literature, please see This guideline follows the structure and definitions of the ESCMID Guideline on Candida diseases –6. It isAll recommendations are given in boldface type.This statement, of course, presumes that the acute condition may be reversed by ICU measures, and that age and comorbidities of patients do not contradict such an approach. Typically, full-code management applies to patients with curative therapeutic options, those in remission of their malignancy, as well as to patients in whom cure is not likely but the expected life-span is substantial , 9. It hintermediary prognosis , those with worsening organ failure scores after days 5 of ICU therapy had a high risk of dying. The authors suggested to offer these patients a full-code ICU trial with re-evaluation of the goals of therapy after at least 6 days [poor prognosis suggest that the optimal duration of time-limited ICU trials seems to differ largely between patients with solid tumors and hematologic malignancies [In a prospective trial on patients with t 6 days . We woulgnancies . Thus, pSeveral observational investigations on cancer patients with septic complications and respiratory failure indicate that early ICU admissions and early interventions, respectively, are associated with improved survival , 12–16. Mortality of ICU cancer patients has markedly dropped over the last decades, presumably due to improved general ICU management, improved diagnostic and therapeutic strategies for several cancer-specific situations, as well as refined admission criteria , 17, 18..A recent Brazilian investigation showed an independent correlation between daily joint rounds of cancer specialists and intensivists as well as the number of protocols with lower mortality and more efficient resource use . This isAlthough a recent study by Soares and colleagues failed to identify ICU volume as a prognostic marker for ICU survival , the majAll previous statements are summarized in Table With respect to the critical prognosis of ARF in cancer patients, especially in patients with ARF of unknown origin , 30, diaThe diagnostic strategy should encompass a comprehensive analysis of the clinical course of the underlying malignancy, including data on mechanisms of immunosuppression, antineoplastic and antimicrobial treatment, and prophylaxis , 33.Table Clinical Practice Guidelines on noninvasive mechanical ventilation in ARF” on behalf of the German Society of Pneumology and Ventilatory Medicine acknowledge that NIV can be attempted to avoid intubation in immunosuppressed (including hemato-oncologic) patients. In this regard, it is of particular importance to respect common contraindications and termination, i.e., intubation criteria [2 > 50 mmHg excluded, pre-specified intubation criteria [Noninvasive ventilation (NIV) remains the first-line standard of care in acute exacerbate chronic pulmonary disease (AECOPD) or acute cardiogenic pulmonary edema . Howevercriteria . Howevercriteria . Second,criteria . Finallycriteria ). The tr2/FiO2 < 200) and mortality (PaO2/FiO2 < 300) rates in elderly patients with pneumonia and hypoxic ARF when compared to conventional oxygen therapy or NIV [High flow nasal oxygen (HFNO) has recently been shown to be associated with reduced intubation  management are not different in cancer patients compared to critically ill patients without a malignancy as underlying disease (A-IIt). However, (i) patient’s wish, (ii) progressive (multi-) organ failure despite full-code ICU management and no chance of reversibility, or (iii) rapid progress of cancer disease without further treatment options have to be kept in mind and could change the treatment decision or treatment goals . These iIntensive care admissions often represent an additional burden for patients and families in situations without option for cure. Good communication is essential to allow patients and families to meet their preferences regarding EOL. According to the literature, too many EOL decisions are made too late . Early iTo improve quality of EOL care in critically ill patients regarding important clinical questions, i.e., preparing for withdrawal, assessment and drug management of distress, discontinuation of treatment and monitoring, or dignity conserving care , the autIn some cases, for example pulmonary involvement due to high grade non-Hodgkin lymphoma, hyperleukocytosis, etc., despite ICU treatment initiation or completion of anticancer chemotherapy, immunochemotherapy or radiotherapy is necessary to improve patient’s clinical situation. Such some decision needs intensive discussions between the responsible hemato-oncologist and intensivist. Again, we recommend joint assessments by cancer specialists and intensivists and that basic concepts of the care of critically ill cancer patients should be included into the curricula of both cancer specialists and intensivists.Published guidelines on prevention, diagnosis, and treatment of infections in cancer patients do not specifically address critically ill cancer patients. In addition, few studies and current guidelines on infections in critically ill patients specifically address the particular requirements in the treatment of cancer patients Table .Table 5RManagement of neutropenic patients in the intensive care unit Pneumocystis jirovecii pneumonia [Prophylaxis of viral , bacterineumonia Unexplained fever in neutropenic patients   Guidelines for Management of Sepsis and Septic Shock recently published . The guiIDSA guidelines on immunization in patients with cancer ; guideliProphylaxis of infectious complications with colony-stimulating factors in adult cancer patients undergoing chemotherapy .Resources on prevention and treatment of infections in cancer patients, which might be largely applicable to critically ill cancer patients, are guidelines on:The prognosis of allogeneic hematopoietic stem cell transplant (HSCT) patients admitted to an ICU has been significantly improved in the last years . It is iFor further information regarding isolation procedures, specific monitoring, as well as typical complications, please see Venous thromboembolism (VTE), including deep-vein thrombosis and pulmonary embolism, represents a major cause of morbidity and mortality in cancer patients. An increased tumor burden results in a higher risk for VTE with the highest risk in patients receiving systemic chemotherapy or being hospitalized on surgical and medical floors.Although there are no validated data on the risk assessment in cancer patients on the ICU, according to the American Society of Clinical Oncology guidelines, risk scores  are helpful tools to identify patients at highest risk for VTE –81. For Pharmacological and cellular treatment of cancer is changing dramatically not only with benefits for patient’s outcome and comfort, but also with new toxicity profiles. The vast majority of adverse events can be classified as mild or moderate but there are also some cases of severe and life-threatening complications requiring ICU admission. Diagnosis and management of severe side effects after  antibody treatment, tyrosine kinase inhibitors, immune checkpoint inhibitors, and chimeric antigen receptor-modified (CAR-) T cells were summarized recently in a concise review by the iCHOP group .In order to prevent the transfusion-associated graft versus host reaction (GvHR), all cellular blood products have to be irradiated with ≥ 30 Gy for patients with lymphatic diseases, autologous/allogeneic stem cell transplantation, and all patients treated with nucleoside analogues  . Generalwww.hlh-registry.org).Cancer patients are at risk to suffer from rare sepsis-like syndromes such as hemophagocytic lymphohistiocytosis (HLH), cytokine release syndrome (CRS), drug reaction with eosinophilia and systemic syndromes (DRESS), or capillary leak syndrome (CLS) –89. InfeDuring the last years, outcome of patients with malignant hematological or oncological diseases requiring intensive care treatment has clearly improved. From recently published data, we conclude that the severity of the acute illness is more important with regard to short-term survival than the underlying type and stage of malignancy. Patients receiving intensive treatment of their malignancy should be considered for ICU treatment as other severely ill patients without cancer. Centers should establish local admission criteria for critically ill cancer patients as well as standardized admission procedures, such as joint assessment by cancer specialists and intensivists, allowing identification of patients and standardized transfer to the ICU. In some patients, time-limited ICU trials with pre-defined do-not-escalate decisions  may be an adequate option until final decision. This implements that critically ill cancer patients should be referred to experienced intensive care units. We strongly recommend establishing a local structure with daily joint rounds of cancer specialists and intensivists as well as standard-operating procedures for the management of frequent medical conditions in critically ill cancer patients.Furthermore, centers should organize continuous medical education of cancer specialists and intensivists. Finally, basic concepts of the care of critically ill cancer patients should be included into the curricula of both cancer specialists and intensivists.In addition, the early integration of an interdisciplinary palliative care team is an urgently necessary measure.In order to improve the management of critically ill cancer patients, knowledge of current practices in German-speaking countries is of utmost interest. To produce more robust data, participation of treating centers in central patient registries, e.g., the iCHOP registry (info@ichop.eu), is highly recommended.ESM 1(DOCX 69 kb)ESM 2(DOCX 80.5 kb)


Cervical cancer is a highly prevalent cancer that affects women around the world. With the availability of new technologies, researchers have increased their efforts to develop new drug delivery systems in cervical cancer chemotherapy. In this review, we summarized some of the recent research in systematic and localized drug delivery systems and compared the advantages and disadvantages of these methods. Cervical cancer is the third most common malignancy among women, with approximately half a million newly diagnosed cases and over 200,000 deaths annually ,2. AlthoThe treatment of cervical cancer is stage-specific. While early stage disease can be cured with radiotherapy or surgery, the most effective treatment for locally advanced stage patients is concurrent chemotherapy and pelvic irradiation ,5. TypicDespites advances in treatment, patients with metastatic cancers and those with recurrent or persistent disease have limited treatment options . A numbeDrug delivery approaches can be categorized based on their route of administration  and the type of device. Systemic delivery is based on particles  with surface features that help target the desired site when injected. In contrast, localized delivery limits systemic drug toxicities by direct delivery of the drug to the tumor. Based on depot systems that are implanted either directly into or adjacent to the tumor, the later promotes the release of drug directly to the cancer site ,18. In rIn this review, we discuss the literature supporting novel drug delivery strategies for cervical cancer treatment and highlight some of the current advancements in systemic and local drug delivery systems.The entrapment of chemotherapeutic drug in nanocarriers has attracted considerable attention due to their structure, varied composition, and surface modifications . The mosHuman cancer cell lines are fundamental models, either in vitro as monolayer culture or in vivo as xenografts in mice, to study the efficacy of therapeutic agents in cancer therapy. Hela was the first cultured cancer cell line, which was derived from cervical cancer cells taken from Henrietta Lacks in 1951 . Since tNanoparticles (NPs) of noble metals have proven their efficacy in the clinical field for cancer therapy due to their unique features such as ease of synthesis, simple surface chemistry and functionalization, broad optical properties, and high surface-to-volume ratio . Noble mNoble metal NPs are mostly made by physical and chemical methods; however, this expensive synthesis can allow toxic substances to be absorbed onto their surfaces . This prMoringa olifea extract showed anticancer activity [One of the most promising biomaterials in the field of nanomedicine is silver nanoparticles (AgNPs) ,42; its activity .Unfortunately, silver was shown to be reactive with body tissues, therefore, as a strategy to reduce the high reactivity and adverse effects of silver, AgNPs were encapsulated in gelatin or polyethylene glycol to increase the biocompatibility of the surface. However, these coated particles were less effective than the non-coated ones . In anotPodophyllum hexandrum exhibited an effective in vitro anti-proliferative activity against HeLa cells by induction of DNA damage and cell cycle arrest at G2/M. Further results demonstrated that the mitochondria of AuNPs-treated cells became dysfunctional due to the activation of the caspase cascade, leading to apoptosis [Gold (Au) NPs emerged as a promising scaffold for chemotherapeutic drug and gene delivery vehicle, due to its high bioavailability and low immunogenicity . AuNPs lpoptosis . When copoptosis . Moreovepoptosis .To improve the selective delivery of therapeutic agents to specific cells or tissues, targeting ligands , which inhibited the proliferation of and induced chromatin condensation and nuclear fragmentation of SiHa cells , and copBiodegradable polymeric NPs have received considerable research interest in anticancer drug delivery due to their high drug loading capacity, self-stability, high cellular uptake, more desirable biodistribution, and capability to deliver both hydrophilic and hydrophobic drugs ,61. WhilBiodegradable polymers, either natural or synthetic, can break down through chemical or enzyme-catalyzed degradation. Biodegradable polymers offer numerous advantages in the field of drug delivery: (1) The drug release kinetics can be controlled by degradation rate of polymers, so a sustained and controlled drug release is possible; (2) the polymeric carrier would degrade into nontoxic, absorbable subunits that can be metabolized; and (3) there is no need for a follow-up surgical removal once the drug supply is depleted .co-glycolide) (PLGA) showed sustained and controlled delivery of docetaxel for cervical cancer treatment both in vitro and in vivo and demonstrated higher cellular uptake efficiency and high antitumor efficacy [Nanoparticles of various polymers have been tested. One example made of different derivatives of poly, and HeLa cells. Compared to free cisplatin, liposomal cisplatin actively inhibited cell proliferation and decreased the spheroid-forming ability of R-ME-180 cells in tumors and in nude mice, in a dose-dependent manner [Transferrin-targeted liposomes where shown to be more specific in delivery of paclitaxel to cervical cancer cell lines, compared to non-targeted liposomes . MoreoveDendrimers are spherical, highly symmetric, and greatly branched, macromolecules with a well-defined structure, surface charge, and molecular size that display a high degree of monodispersity ,85. Thei50 value, higher drug loading, faster drug release rate and more cytotoxicity compared to RGD-conjugated one in HeLa cells. This could be probably because of the higher multivalent ligand density on the surface of the IL6-conjugated dendrimers, which cause better drug delivery through receptor-mediated endocytosis [Doxorubicin loaded dendrimer was conjugated with two of cancer cell targeting moieties, IL-6 antibody and RGD (Arginyl-glycyl-aspartic acid) peptide and the drug loading capacity and release profile as well as their targeting efficiency were compared. Drug loaded dendrimers decorated with IL-6 antibody exhibited higher cellular internalization, lower ICocytosis .Another approach to improve the delivery of highly lipophilic drugs is by self-emulsifying drug delivery systems (SEDDS) . SEDDS a®) and ado-trastuzumab emtansine (Kadcyla®) [Targeted delivery of drugs by conjugating them to monoclonal antibodies, which can bind specifically to tumor-associated target antigens is an innovative approach in cancer treatment that gains considerable research interest. Antibody–drug conjugates (ADCs) can selectively deliver therapeutics to tumor cells and provide sustained clinical benefit to cancer patients with less systemic toxicity . Furtheradcyla®) ,99. Insuadcyla®) . TF-011-adcyla®) . Anotheradcyla®) ,102.Localized delivery of chemotherapeutic drugs to the cervix offers a number of advantages compared to systemic delivery such as its avoidance of systemic chemotherapeutic drug circulation, which leads to less drug waste and decreased side effects and delivery of high dose of the active agent in the cervix which improves the efficacy of the treatment  Figure . The useThis direct delivery is possible due to the easily reachable location of the cervix through the vagina . Presentco-vinyl acetate) ring device incorporated with cisplatin for local treatment of cervical cancer were recently reported. The intra-vaginal rings were found to be effective against both HPV positive and HPV negative cervical cancer cells in vitro [50 value for HeLa cells [As flexible and torus-shaped delivery systems, intra-vaginal rings can provide both sustained and controlled drug release, lasting for several weeks to several months . Recentlin vitro . Other tLa cells .Nanofibers have various applications, one of which is drug delivery, especially in local chemotherapy. Their attractive characteristics for electrospinning used in drug delivery are high encapsulation efficiency, ease of operation, cost-effectiveness, high loading capacity, and simultaneous delivery of various therapies . RecentlVaginal films made are promising delivery systems that could achieve better patient compliance and therapeutic efficacy. Curcumin-hydroxypropyl cyclodextrin complex vaginal films were shown to be retained in the vaginal mucosa for up to six hours, making it a potentially effective therapy for HPV-induced cervical cancer .® F127 combined with 5-fluorouracil and alternative mucoadhesive polymers like Carbopol 934, hydroxypropylmethylcellulose, and hyaluronic acid was created. Cytotoxicity studies with HeLa cells showed that complexes of 1% 5-fluorouracil and b-cyclodextrin or hydroxypropyl-b-cyclodextrin were just as effective as free 5-fluorouracil. These results indicate that a lower dose of this anticancer drug can achieve better therapeutic efficacy and increased patient compliance [As proven and accepted therapeutics, vaginally-applied gels have drugs and active ingredients which can restore physiological pH, moisturize and lubricate, be a contraceptive or labor inducer, and/or have microbicide activity . In regampliance . AlthougAnother approach to the cervical delivery of a cytotoxic agent is using bioadhesive cervical patches . In one Hodge et al. developed a new cervical delivery instrument (CerviPrep™) that consisted of a tubular applicator with a cervical cap on one end. When the cervical cap was positioned over the patient’s cervix, this device coulddirectly apply pharmaceuticals to the cervix with limited exposure to and absorption by the adjacent vaginal tissue. The results of clinical studies with the CerviPrep™ delivery device showed limited systemic exposure and no toxicities .Inorganic, lipidic and polymeric nanocarriers are promising candidates for the development of systemic delivery systems in cervical cancer chemotherapy. Experimental results show great potential for the widespread adoption of nanocarriers in cervical cancer treatment over conventional chemotherapy. Their attractive properties include biocompatibility, low toxicity, lower clearance rates, the ability to target specific tissues, and controlled release of chemotherapeutic agents. However, the toxicology of nanocarriers in humans still needs to be fully studied. On the other hand, localized delivery of chemotherapeutic drugs to the cervix offers a number of advantages such as increased efficacy and decreased side effects due to direct delivery to the site of cancer, which avoids systemic circulation of chemotherapeutic drugs. However, the local drug delivery systems are not effective in metastatic tumors. The therapeutic use of these drug delivery devices and formulations would vary depending on the patient’s cancer stage. Any vaginal drug delivery device or formulations could be used to provide local delivery of a chemotherapeutic drug to the cervix, and each has its own advantages like improved patient compliance and disadvantages like expensive production. Although, localized delivery of chemotherapeutic drugs directly to the cervix will improve the patient’s overall quality of life and reduce hospital presentations and cost, more research needs to be performed characterizing the local accumulation and efficacy of drugs in the cervix.


Besides being a risk factor for multiple metabolic disorders, obesity could affect female reproduction. While increased adiposity is associated with hormonal changes that could disrupt the function of the hypothalamus and the pituitary, compelling data suggest that obesity-related hormonal and inflammatory changes could directly impact ovarian function.To review the available data related to the mechanisms by which obesity, and its associated hormonal and inflammatory changes, could affect the female reproductive function with a focus on the hypothalamic-pituitary-ovarian (HPO) axis.PubMed database search for publications in English language until October 2017 pertaining to obesity and female reproductive function was performed.The obesity-related changes in hormone levels, in particular leptin, adiponectin, ghrelin, neuropeptide Y and agouti-related protein, are associated with reproductive dysfunction at both the hypothalamic-pituitary and the ovarian levels. The pro-inflammatory molecules advanced glycation end products (AGEs) and monocyte chemotactic protein-1 (MCP-1) are emerging as relatively new players in the pathophysiology of obesity-related ovarian dysfunction.There is an intricate crosstalk between the adipose tissue and the inflammatory system with the HPO axis function. Understanding the mechanisms behind this crosstalk could lead to potential therapies for the common obesity-related reproductive dysfunction. According to a recent population study, approximately 39% of the population over the age of 20 and 18% of children between the ages of 2–19 are obese –7. This The female reproductive physiology is a complex interaction between neuroendocrine and endocrine signaling affecting the hypothalamus, the pituitary gland and the ovaries. At the level of the hypothalamus, gonadotropin releasing hormone (GnRH) pulses activate the pituitary release of the two gonadotropins, follicle-stimulating hormone (FSH) and luteinizing hormone (LH) , 9. FSH The HPO axis plays a crucial role in pubertal transition. Normal pubertal development is under two main controls, adrenal and hypothalamic . The adrIt has been thought that obesity changes the expected time course of puberty and leads to earlier thelarche, adrenarche and menarche –28. AdipObesity decreases pituitary LH pulse amplitude and mean LH release without changing its frequency, leading to impaired luteal phase . AdditioLeptin is a 16-kDa adipokine secreted primarily by white adipose tissue, although it can be secreted from other tissues, such as gastric mucosa . Serum lLeptin’s energy homeostatic function in part contributes to its effect on puberty. Puberty requires a certain energy balance to proceed , 44. LepLeptin affects GnRH pulse neurons indirectly though GABA or kisspeptin , 48. BotStudies on the effect of leptin on ovarian folliculogenesis and ovulation have shown conflicting results. Leptin has been isolated in both mural and cumulus granulosa cells and within the follicular fluid of pre-ovulatory follicles in women undergoing in vitro fertilization (IVF) . Serum lGhrelin is an orexigenic enterokine composed of 28 amino acids related to the oxyntomodulin family of intestinal peptides . It is pIn the hypothalamus, ghrelin can decrease both GnRH secretion and pulsatility –73 possiStudies on the effect of ghrelin on ovarian steroidogenesis have shown mixed results , 77. AniThus, these findings suggest that ghrelin may indirectly contribute to puberty through general energy homeostasis such as insulin action, and it acts at the level of the hypothalamus, pituitary and ovary leading to alterations in normal reproductive function.The connection between peripherally circulating adipokines, enterokines and the HPO axis centers on a collection of neurons within the arcuate nucleus, which secrete the neuropeptides NPY and AgRP . NPY is Under normal physiologic conditions, NPY concentration increases in the portal capillary blood during the ovulatory surge to potentiate the action of GnRH on pituitary gonadotropin secretion , 93. SimAdiponectin is a 30-kDa adipokine secreted by the adipose tissue . OppositAdiponectin and its receptors are found in many organs including the ovaries –111. AdiThese findings suggest that adiponectin is notable for its action in mediating insulin sensitivity, with its receptors found at every level of the reproductive axis making it a great therapeutic target for ovulatory dysfunction.Insulin is a 51 amino acid protein synthesized in the beta islet cells of the pancreas. Its release is stimulated by glucose in the gastrointestinal tract from ingestion, as well as various amino acids directly . InsulinInsulin acts on the pituitary to modulate the GnRH receptor and increases LH secretion after GnRH stimulation . InsulinDisruption of insulin signaling in diet-induced obesity improves reproductive cyclicity in mice, suggesting that insulin represents a mediator for pituitary LH dysregulation in obesity . FurtherObesity is a state of chronic inflammation with macrophage infiltration into various tissues. Macrophage infiltration into adipose tissue is directly correlated to both the degree of adiposity as well as chemokine/adipokine production, such as MCP-1 . The patThe pro-inflammatory AGEs may be part of the link between diet-induced obesity and inflammation, with AGEs inducing MCP-1 gene expression  thus forKandaraki et al. has demonstrated in human immortalized granulosa cells (KGN cell line) that AGEs attenuate LH- and FSH-induced ERK signaling needed for cell proliferation and proper follicular growth , one posPopulation studies on the clinical sequelae of obesity provide a connection between obesity and subfertility. Several studies indicated that obesity is a risk factor for ovulatory dysfunction –143. ForObesity does not only affect spontaneous pregnancy rates in fertile patients, but it also confers a risk for poorer ART outcome. While earlier data could not demonstrate adequate convincing evidence of poorer ART outcome parameters, recent data supports this correlation. Large national cohort studies as well as systemic reviews and meta-analyses suggest that increasing BMI is negatively correlated with implantation, clinical pregnancy, and live birth rates –154. ObeThe poorer IVF outcomes observed in obese patients are quite intriguing. These observations suggest that the effect of obesity on ovarian function is not solely dependent on the HPO axis, since gonadotropins are supplied exogenously during IVF cycles, thus bypassing the HPO axis. Obesity adversely impacts ART outcome differently in different ethnic populations –161. HowObesity could disrupt endometrial receptivity leading to poorer implantation rates , 166, arWith the uncurbed obesity epidemic, more reproductive-aged women will face metabolic and reproductive complications. Body energy hemostasis is closely linked to reproductive function through many hormones, adipokines, cytokines, and growth factors that act at the level of the brain and the ovaries. Obesity is also a state of chronic inflammation, which may directly affect ovarian function possibly by increased macrophage infiltration in the ovaries through MCP-1 mediated pathways. The elevation of AGEs in the serum and tissues of obese women may exacerbate the reproductive dysfunction associated with adiposity and may provide, along with MCP-1, a crucial link between obesity and ovarian macrophage infiltration. Each of these molecules and their prospective pathways may represent potential therapeutic targets in order to improve the overall reproductive health of overweight/obese women. Obesity, with its alterations in the AGEs/MCP-1 axis, could disrupt the ovarian microenvironment potentially compromising oocyte competence, formation of healthy embryos and ultimately conception.Finally, this review underscores a critical need to uncover the mechanistic actions of molecules that affect almost every level of the HPO axis. Obesity, a significant and growing public health problem, is an overwhelming condition that causes reproductive disturbances in women in part due to ovarian dysfunction. Losing weight is commonly challenging and often not sustainable. Thus there is a need to establish therapies for ovarian dysfunction and to improve ovarian health in the obese patient population.


Restrictive cardiomyopathy (RCM) is the least common among cardiomyopathies. It can be idiopathic, familial, or secondary to systematic disorders. Marked increase in left and/or right ventricular filling pressures causes symptoms and signs of congestive heart failure. Electrocardiographic findings are nonspecific and include atrioventricular conduction and QRS complex abnormalities and supraventricular and ventricular arrhythmias. Echocardiography and cardiac magnetic resonance (CMR) play a major role in diagnosis. Echocardiography reveals normal or hypertrophied ventricles, preserved systolic function, marked biatrial enlargement, and impaired diastolic function, often with restrictive filling pattern. CMR offering a higher spatial resolution than echocardiography can provide detailed information about anatomic structures, perfusion, ventricular function, and tissue characterization. CMR with late gadolinium enhancement (LGE) and novel approaches  can direct the diagnosis to specific subtypes of RCM, depending on the pattern of scar formation. When noninvasive studies have failed, endomyocardial biopsy is required. Differentiation between RCM and constrictive pericarditis (CP), nowadays by echocardiography, is important since both present as heart failure with normal-sized ventricles and preserved ejection fraction but CP can be treated by means of anti-inflammatory and surgical treatment, while the treatment options of RCM are dictated by the underlying condition. Prognosis is generally poor despite optimal medical treatment. According to a recent position statement of the European Society of Cardiology (ESC) working group on myocardial and pericardial diseases, cardiomyopathy is defined as a myocardial disorder in which there is abnormality in the structure and function of the myocytes, in the absence of coronary artery disease, hypertension, valvular disease, and congenital heart disease that could justify this abnormality. In the past, cardiomyopathies were also separated from systematic disorders that could involve the myocardium. ESC classification proposes a shift from diagnosing by exclusion criteria and focusing on the morphology and function of the heart .Restrictive cardiomyopathy (RCM) is the least common of cardiomyopathies. It is characterized by impaired diastolic function with restrictive filling and reduced diastolic volume of either or both ventricles, preserved systolic function, and invariably normal or mildly increased wall thickness . Due to There are many classification criteria for the causes of RCM. According to the ESC Working group in myocardial and pericardial disease RCM can be idiopathic, familial , or secondary to systematic disorders  Table 1Table 1. 3 and S4 gallops). The ECG may reveal sinus rhythm with signs of left or biatrial enlargement or atrial fibrillation. There may be nonspecific ST and T wave abnormalities. QRS voltage may be low in the precordial leads and conductive disturbances may also occur, particularly in infiltrative diseases. Specific ECG findings may occur depending on the underlying cause and aid in the diagnostic work-up.Patients commonly present with dyspnea, fatigue and limited exercise capacity, palpitations, and syncope, while angina may also occur. Others may present with thromboembolic complications. Physical examination may reveal signs of congestive heart failure .A complete evaluation should be performed as restrictive cardiomyopathy may originate from systematic disorders or can in turn affect other organs as heart failure develops. Hematocrit (HCT), serum electrolytes, blood urea nitrogen (BUN), creatinine, 24 hr urine total protein, and liver function should be assessed. Arterial blood gas (ABG) should also be obtained to monitor hypoxia. Serum brain natriuretic peptide (BNP) or N-terminal pro-b-type natriuretic peptide (NT-proBNP) and troponin T are indicative of the heart failure and in many cases predictors of survival –7. SpeciEchocardiography is the first imaging modality for the assessment of patients with dyspnea and/or heart failure. In patients with RCM, it usually discloses nonspecific findings such as normal (not dilated) ventricles with normal or increased wall thickness Figures  althoughOne of the main utilities of echocardiography is the differential diagnosis between constrictive pericarditis (CP) and RCM. Both present as heart failure with normal-sized ventricles and preserved EF, dilated atria, and Doppler findings of increased filling pressure (often restrictive filling pattern). Differential diagnosis is important since CP can be treated by means of anti-inflammatory drugs or surgery while the treatment options of RCM are dictated by the underlying condition. For the main differences between those disease states, the reader can refer to Echocardiography is unable to establish the definite diagnosis of specific RCM subtypes due to its inherent poor tissue characterization and limited assessment of the ventricular apex and the right ventricle. Cardiac Magnetic Resonance (CMR) with a higher spatial resolution and late gadolinium enhancement (LGE) can provide detailed information about anatomic structures, perfusion, ventricular function, and tissue characterization . Late gaAs of this writing, Cardiac CT has no specific role in the work-up of patients with RCM. It can accurately exclude the presence of coronary obstructive artery disease in patients with angina symptom. Its main role is in the differential diagnosis between RCM and CP where it can give invaluable information about the thickness and composition of the pericardium .In contemporary practice, invasive catheterization and coronary angiography are rarely needed for the establishment of diagnosis. In cases where chest pain is the predominant symptom, it can reveal the presence or absence of obstructive coronary artery disease. Left and right heart catheterization can be used when findings of noninvasive tests, including the differential diagnosis between CP and RCM, are nonconclusive .Histological examination can be valuable in setting a definite diagnosis when noninvasive studies have failed, although there are periprocedural risks of endomyocardial biopsy (EMB) . MoreovePrognosis varies among the specific types of RCM but is generally poor with progressive deterioration, particularly in children, despite optimal medical treatment.β-pleated sheets antiparallel to one another, resistant to proteolysis, and they cause oxidative stress, disruption of the myocardium, and myocardial damage Treatment of CS incorporates the guidelines directed medical therapy (GDMT) of heart failure with the addition of corticosteroids to reduce the inflammatory process (except asymptomatic hilar adenopathy where no therapy is required). FDG-PET is suggested in the treatment algorithm for the follow-up , defininHemochromatosis is a group of disorders which leads to excessive accumulation of iron within the cells of liver, pancreas, heart, and several endocrine glands leading to cirrhosis, diabetes, heart failure, skin pigmentation, and so forth. It may be hereditary or secondary. In the former, properly called hemochromatosis , transfe∗) technique has been used to quantify myocardial iron overload [∗ values smaller than 20 msec are predictive of adverse cardiac events [Recently CMR with the use of T2 star  and is an uncommon cause of restrictive cardiomyopathy. HES are either primary, first described by Löffler , or seco eos/mm3 . Indepen eos/mm3 . Primaryolvement , 82.Cardiac infiltration consists of three stages: (a) acute necrotic stage, (b) thrombotic stage, and (c) fibroting stage. The first stage is characterized by inflammation where eosinophilic degranulation releases toxic proteins that induce endocardial necrosis and apoptosis. Rarely, acute necrotizing myocarditis may ensue with high early mortality without appropriate treatment. If left untreated, the first stage evolves into the thrombotic stage due to continuous eosinophilic activation. It consists of thrombus formation over the affected endocardium. Typically there is thrombotic obliteration of left and/or right ventricular apex which may extend to the ventricular outflow tracts, the basal ventricular segments, or even the atria. Thromboembolic complications dominate the clinical picture at this stage. The final stage is due to scarring and irreversible fibrosis of the endocardium  which mECG findings are nonspecific and include sinus tachycardia with conduction disturbances or nonspecific ST-T segment abnormalities. At the first stage, echocardiography may reveal increased LV wall thickness due to interstitial edema, impaired systolic function with abnormal wall kinetics, although most often ventricular systolic function remains preserved. CMR with LGE may additionally reveal endomyocardial involvement, often with patchy distribution . At the The goals of initial therapy are twofold: first to decrease the eosinophil count and attenuate the inflammatory process and second to treat the underlying cause (if any). Corticosteroids are the mainstay of treatment combined with disease specific interventions, for example, antihelminthic therapy for parasitic infections. Anticoagulation should be initiated at the second stage to prevent thromboembolic complications. Once severe EMF has developed, surgical resection with extended endocardial dissection is the treatment of choice but with poor long-term prognosis. Atrioventricular valve replacement should be undertaken at the time of operation if the valves are affected.α-galactosidase A which breaks down neutral glycosphingolipids [α-galactosidase A activity or by GLA gene sequencing, while female carriers have a normal enzyme activity and gene sequencing is needed [Anderson-Fabry disease is a rare X-linked recessive lysosomal storage disorder, caused by mutations of the GLA gene that encodes golipids . This regolipids –88 whilegolipids . Transiegolipids  or prologolipids  PR intergolipids   and enzyme replacement therapy which is effective in myocardial remodeling and improves peak systolic strain and strain rate  but its Diabetic cardiomyopathy (DMCMP) was firstly described as a dilated phenotype with reduced ejection fraction (HFrEF). Recent studies , 97 showThis is the primary restrictive cardiomyopathy, more commonly affecting women, where hemodynamic abnormalities occur without any specific histological changes and in the absence of ischemic, valvular, and congenital heart disease or hypertension. Patients usually present with overt heart failure. Echocardiographic findings are nonspecific as well, with marked dilation of the atria and normal-sized ventricles with preserved systolic function. Doppler demonstrates elevated LV filling pressures  more oftSystemic sclerosis is an autoimmune disorder that practically affects almost any organ system. Myocardial fibrosis results in heart failure symptoms, while fibrosis of the conduction system may lead to ventricular arrhythmias and sudden cardiac death. Physical examination depends on the organs involved with cardiac manifestations mainly caused by right heart failure . Echocardiographic findings are consistent with restrictive physiology.Radiotherapy used against thoracic malignancies  may provoke several cardiovascular disorders such as coronary artery disease, myocardial fibrosis, valvular heart disease, pericardial disease, and ECG conduction abnormalities. Diffuse interstitial fibrosis after relatively low doses of radiation  ultimateDanon disease is an X-linked disorder, due to primary deficiency of lysosome-associated membrane protein 2 (LAMP2). Symptoms occur in adolescence and include heart failure, mental retardation, and skeletal myopathy. Most patients die of heart failure in the third decade of life . LaboratEchocardiography reveals severe hypertrophy of both ventricles, resembling HCM but the electrophysiological abnormalities, particularly ventricular preexcitation, may give a clue for differentiating the two conditions . In DanoIt is an autosomal recessive disorder caused by repeated Guanine-Adenine-Adenine triplets in the frataxin gene on chromosome 9 . ManifesRestrictive cardiomyopathies are a group of diseases with various genetic or acquired etiologies. They are characterized by marked increase in ventricular filling pressures which cause symptoms and signs of congestive heart failure. Electrocardiogram, echocardiography, CMR with LGE, or novel radionuclide imaging can direct the diagnosis to specific subtypes of RCM which is important as treatment and prognosis differ significantly, but endomyocardial biopsy is the definite means of establishing the diagnosis. Genetic counseling and familial screening are necessary in hereditary forms.


C. Theilmann et al.  describe how one gut microbe, Lactobacillus acidophilus, catalytically unwraps plant glucosides to make deglucosylated bioactive aglycones available to human tissues. They demonstrate that understanding the metabolism of plant glycosides by intestinal bacteria is essential to appreciating how bacteria manipulate the levels of bioactive plant metabolites in the human host.The therapeutic potential of plants is widely recognized and harnessed in plant-based remedies and drug discovery. However, the factors that modulate the bioavailability and bioactivities of plant-derived phytochemicals are poorly understood. In a recent article in  Humans have always leveraged the healing power of plants. The desire to maintain health and combat disease compelled ancient populations to observe and document the effects of edible resources in their environment. As this information was passed down through generations, civilizations acquired a thorough understanding of which plants possessed therapeutic potential and which were toxic or deadly. Plants believed to possess beneficial properties were incorporated into the medicinal practices of the earliest societies. The ancient Egyptians, for example, used willow tree leaves to relieve joint pain , and in While plant-derived drugs continue to aid in the treatment and prevention of disease in the 21st century, alternative medicinal practices reminiscent of those of ancient times have also grown in popularity. People have embraced herbal supplements, functional food diets, probiotics, prebiotics, and “nutraceuticals” as a means of acquiring dietary compounds associated with anticancer, anti-inflammatory, and antioxidant activities , 4. CompLactobacillus acidophilus interacts with these compounds.The bioavailability and bioactivities of plant glycosides, such as salicin, depend greatly on their conjugation to sugar moieties; plant glycosides are more polar and often less biologically active than the deconjugated aglycone. For this reason, microbes that utilize these plant glycosides as a source of energy are key players in regulating the levels of the bioactive aglycones available to human tissues . These bLactobacillus acidophilus NCFM grows on plant glycosides commonly found in the human diet, including the monoglucosylated compounds salicin and esculin and the diglucosylated compound amygdalin. Interestingly, they found that other common bacterial strains present in the GI tract did not grow on these phytoglucosides; this finding is important because it suggests that L. acidophilus occupies a unique metabolic niche for these compounds and uses them as carbon sources.Theilmann et al.  demonstrL. acidophilus into the culture supernatant, Theilmann et al.  transporter LBA0725 and the phospho-β-glucosidase (P-Bgl) LBA0726. A second gene locus containing a related but distinct PTS, LBA0227, and a second type of the P-Bgl, LBA0225, was upregulated following growth on amygdalin but not on esculin or salicin. Theilmann et al. -diglucoside moiety. Importantly, the authors’ data provide a means of predicting the metabolism of certain plant glycosides in Lactobacilli based on the presence or absence of related gene loci.In addition to linking lleagues  determinCan this predictive power be extended to account for structural differences in these related phospho-β-glucosidases? Subtle differences in active-site architectures might provide explanations for distinct plant glucoside processing specificities among closely related phospho-β-glucosidases.Thus, we modeled the structures of the two phospho-β-glucosidases LBA0726 and LBA0225  identifiIn antiquity, humans had to rely on trial and error to determine the therapeutic properties of plant materials, and an error could have been deadly. Fortunately, our knowledge about the phytochemicals available in foods and other plants has significantly advanced. As demonstrated by Theilmann et al. , specifi


A significant increase in the incidence of cognitive impairment in HIV/AIDSpatients has been continuously observed. Consequently, three classificationcategories of cognitive impairment have been proposed: asymptomaticneurocognitive impairment (ANI) and mild neurocognitive disorder (MND), thatcorrespond to the mild and intermediate forms, and HIV-associated dementia (HAD)for the most severe cases. HIV-associated neurocognitive disorders (HAND) is abroad term that encompasses these three categories. Moreover, the application ofneuroimaging methods has led to a major breakthrough in understanding of theneurological changes in HIV, providing greater reliability in the exclusion ofassociated diseases and allowing earlier diagnosis. Therefore, abnormalitiesand/or specific neuroimaging elements may soon be incorporated into the HANDclassification criteria, which will be of great value in the management of thesediseases, including in the optimization of high CNS penetration antiretroviralregimens. With over 40 million people infected worldwide, HIVcurrently has a high prevalence of cognitive impairment, even in individuals inregular use of highly active antiretroviral combination therapy (HAART).2 . fMRI has been studied in small groups ofindividuals with HAND, and longitudinal studies with larger groups are needed tovalidate the technique for clinical use.24Positron Emission Tomography (PET) Fluorodeoxyglucose (FDG)-PET is acommonly used method in clinical practice to indirectly measure the metaboliccell/tissue activity through the use of fluorodeoxyglucose. This technique has notyet shown immediate clinical applicability in managing HAND. However, it has castlight on several pathophysiological concepts that remain unclear. This techniquedemonstrated that metabolic changes in brain tissue secondary to HIV-1 occur longbefore any measurable structural change can be detected. Other issues still understudy with PET are dopaminergic system metabolic activity interaction, the degree ofcognitive impairment, and the role that specific neurotransmitters such as serotoninplay in pathophysiological processes.2026 Despite the advancement of thesecomplementary investigation methods, there is still much to learn about theirapplication and relevance in patients with HAND. Most studies have comparedHIV-positive individuals with HIV-negative controls, focusing on the diagnosis ofHAND or exclusion of associated comorbidities.21 Further studies are needed investigating thepathophysiology of the disease in the whole brain. These studies may help to predictwhich HIV-infected patients are at increased risk for HAND, and thus improve themanagement of the disease, such as by optimization of high CNS penetrationantiretroviral regimens capable of blocking viral exposure, which could beincorporated into AIDS therapeutic guidelines.28The application of neuroimaging methods, particularly MRI, marked a major advance inunderstanding of neurological changes in HIV-1 infected patients. In face of theneed for a more accurate and earlier diagnosis, abnormalities or specificneuroimaging features may in the future be incorporated into the HAND classificationcriteria.


The stem cell-based therapy has emerged as the promising therapeutic strategies for cardiovascular diseases (CVDs). Recently, increasing evidence suggest stem cell-derived active exosomes are important communicators among cells in the heart via delivering specific substances to the adjacent/distant target cells. These exosomes and their contents such as certain proteins, miRNAs and lncRNAs exhibit huge beneficial effects on preventing heart damage and promoting cardiac repair. More importantly, stem cell-derived exosomes are more effective and safer than stem cell transplantation. Therefore, administration of stem cell-derived exosomes will expectantly be an alternative stem cell-based therapy for the treatment of CVDs. Furthermore, modification of stem cell-derived exosomes or artificial synthesis of exosomes will be the new therapeutic tools for CVDs in the future. In addition, stem cell-derived exosomes also have been implicated in the diagnosis and prognosis of CVDs. In this review, we summarize the current advances of stem cell-derived exosome-based treatment and prognosis for CVDs, including their potential benefits, underlying mechanisms and limitations, which will provide novel insights of exosomes as a new tool in clinical therapeutic translation in the future. Cardiovascular diseases (CVDs) are still the leading causes of morbidity and mortality throughout the world. Although a proportion of CVDs is largely preventable, the incidence is increasing particularly in the low- and middle-income countries  released by almost all cell types are categorized into six main classes including exosomes, microvesicles, ectosomes, membrane particles, exosome-like vesicles and apoptotic bodies based on their multiple features such as sizes, shapes, origin and main protein markers Table . Using cIn general, exosomes originate from intracellular endosomes by inward budding inside, which lead to the formation of multivesicular bodies (MVBs) and subsequently MVBs would fuse with the plasma membrane and then release outside its internal vesicles Figure . Integrihttp://www.exorbase.org/)   cell transplantation in 1968 , derived from the undifferentiated inner mass cells of an embryo, can self-renew and differentiate into many types of cells, such as cardiomyocytes. Several studies have broadly observed ESC-derived cardiomyocytes could improve cardiac regeneration and enhance function in experimental animal models  of human ESC-derived MSCs, indicating through the paracrine mechanisms . Although exosomes from both ESCs and ESC-derived cells have analogous cardioprotective effects, exosomes from the ESC-derived cardiac lineage cells theoretically exert more protective effects by transferring the endogenous specific molecules to salvage the injured neighboring cells by regulating inflammation, apoptosis, fibrosis, and angiogenesis. Accordingly, these results encourage development of a great understanding of mechanisms associated with ESC-exosome therapy for CVDs.Particularly, several miRNAs already identified in ESCs derived exosome, play important roles in CVDs and stem cell transdifferentiation , discovered by Shinya Yamanaka in 2006  function. Interestingly, iPSC-derived exosomes exhibited superior cardiac repair with regard to LV function, vascularization, and amelioration of apoptosis and hypertrophy than iPSCs transplantation  hold great cardiac regeneration potential to improve heart functions  treatment. Thereafter, they observed the highly abundant miRNA-146a was partially contributor to this effect  is one type of multipotent stromal cells that can differentiate into a variety of cell types, which are emerging as an extremely promising and major cell-based therapeutic agent for tissue regeneration and repair   have been the most widely investigated in various diseases -derived exosomes led to a markedly increase in cell viability of H9C2 cells under hypoxia/reoxygenation (H/R) Exosomes are critical cellular communicators between cells, which offer the clinical potential for both detection and treatment of various diseases. Noteworthy, exosomes are of great increasing interest for three major reasons with potential clinical applications in CVDs as following: (1) diagnostics and prognostic biomarkers, (2) therapeutic strategies, and (3) drug carriers Dykes, .Due to the relative easy and quick collection and measurement, secreted exosomes and their contents have successfully gained huge interests for their diagnostic and prognostic potentials in various diseases (Properzi et al., In addition to biomarkers, discoveries have identified that exosomes released from certain cell types exhibit therapeutic utilities in various diseases, including cancers (Inamdar et al., in vivo environment poses significant challenges to successful therapeutic outcomes. Thus, drug delivery system has been importantly developed to optimize. Recent evidence showed exosomes, unlike other vectors for gene delivery, biologically derived nano-dimensional vesicle could be used to carry different contents to adjacent or distant targeted cells (Maheshwari et al., The unstable drugs To date, the potential of stem cell exosome-based biomarkers, therapy strategies and drug delivery have largely explored specifically in the field of CVDs Figure . It thusIn conclusion, exosomes derived from stem cells exerts beneficial effects on myocardial regeneration and thus possess a huge therapeutic capacity in prevention/treatment of CVDs Table . AccordiDespite the growing amount of data have reported the beneficial effects and therapeutic potentials the field of CVDs, the exosome-based therapy in clinical application are still challenging and limited by several issues. Firstly, the large amount of time required and poor purified and isolated techniques failed to produce high quantity of exosomes limit their efficiency. (Tang et al., WD, WM, and LZ read and collect the articles involved. JL organized the tables. YY and BC choose the topic of the review and write the manuscript. BC and ZD revise and has a final check for the manuscript.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.