However, clinical evidence assigns a detrimental role to MR activation in the pathogenesis of severe cardiac diseases such as congestive heart failure. This mini review is meant to present and briefly discuss some recent discoveries about locally triggered cardiac MR signals with the objective of shedding some light on its physiological but potentially pathological consequences in the heart.Current evidence reveals that cardiac mineralocorticoid receptor (MR) activation following myocardial stretch plays an important physiological role in adapting developed force to sudden changes in hemodynamic conditions. Its underlying mechanism involves a previously unknown nongenomic effect of the MR that triggers redox-mediated Na Mineralocorticoids are steroid hormones mainly involved in electrolyte/water balance regulation. Aldosterone (ALD) is the main hormone in this group. It is synthesized in the zona glomerulosa of the adrenal gland cortex by the action of P-450 ALD-synthase from 11-deoxycorticosterone through a process mainly controlled by angiotensin II (AngII) and potassium. The effects of ALD on extracellular fluid volume regulation and sodium/potassium homeostasis result from activation of cytosolic mineralocorticoid receptor (MR) on polarized epithelia of kidney distal nephron, salivary and sweat glands, and colon. Once activated, MR translocates to the nucleus and operates as a ligand-dependent transcription factor, mediating classic genomic effects. Interestingly, rapid nongenomic actions of ALD, probably linked to the plasma membrane MR subpopulation –5. As wiIt seems reasonable to imagine that the impact of ALD on cardiovascular systems may arise from its effects on electrolyte/water balance. However, the striking salutary effects of MR antagonism in heart failure (HF) patients –18 invitThe MR is a member of the steroid/thyroid hormone receptor superfamily of ligand-inducible transcription factors, which also includes the glucocorticoid receptor (GR). Although increased ALD concentration constitutes the best-recognized stimulus for MR activation, it can also be stimulated under normal or even low ALD. Furthermore, it has high affinity for ALD and also for cortisol and 11-deoxycorticosterone . The lat+ for one extracellular Na+, being the main responsibility of intracellular pH (pHi) regulation in the myocardium. It possesses high sensitivity to changes in pHi by allosterically sensing H+ and also to phosphorylation or interaction with associated proteins that modulate its activity , due to an increase in Ca2+ transient amplitude, and accounting for ~20% of total increase in force. Ex vivo studies in isolated papillary muscles isometrically contracting serve to describe a complex signaling cascade underlying the SFR, in which stretch-triggered NHE1 activation is crucial. The NHE1 could potentially increase force by two mechanisms: an increase in pHi that would increase myofilament Ca2+ sensitivity and/or an increase in intracellular Na+ leading to Ca2+ increase. We have described a mechanism that comprises a sequential activation of receptors  leading to redox-mediated NHE1 stimulation, which increases intracellular Na+, and thus Ca2+ transient amplitude through reverse Na+/Ca2+ exchange. The scheme of −/+, leading to Ca2+ increase.Increased left ventricular end-diastolic volume caused either by increasing aortic resistance or venous return stretches the ventricular wall and immediately leads to a more powerful contraction. This adaptive response occurs in two consecutive phases: the immediate Frank-Starling mechanism attributed to enhanced myofilament Cadothelin –35, whicThe demonstration that MR activation is crucial to increase mitochondrial ROS, leading to SFR development , 36, 37 HF is a chronic progressive condition in which the heart is unable to pump enough blood to meet organ/tissue requirements. It constitutes one of the most important global health problems. Current treatment is primarily based on inhibition of hormones, mainly AngII, ALD, and catecholamines. The pathophysiological role of ALD in cardiovascular disease has been classically attributed to sodium/water retention and hypertension due to MR activation in renal tubular cells. However, growing evidence suggests that local MR activation in the cardiovascular system may play a role. Although positive correlation between plasma ALD and cardiovascular damage has been described , completThe first MR antagonist accepted for clinical use ~60 years ago was spironolactone. Beyond some tolerability problems, it was approved for the Randomized Aldactone Evaluation Study (RALES) clinical trial in patients with severe HF (Class III-IV NYHA). This study was terminated prematurely due to an interim analysis revealing ~30% reduction in relative risk of death in spironolactone-treated patients together with an impressive reduction in hospitalization for cardiac reasons . AfterwaA new generation of nonsteroid MR blockers with more selectively target benefits, reduced risks of altered receptor affinity and tissue tropism, and minimized side-effects such as hyperkalemia has been recently developed: finerenone, apararenone, and esaxerenone . FinerenIn contrast to the beneficial effects observed in all these studies, two clinical trials failed to show benefits after MR antagonism. In the Treatment of Preserved Cardiac Function Heart Failure with an Aldosterone Antagonist (TOPCAT) study, spironolactone did not improve clinical outcomes in patients with symptomatic HF with relatively preserved ejection fraction ≥45%) (% (45). I2+ transient amplitude that underlies the SFR  are also involved in the progression of severe cardiac pathologies such as hypertrophy and HF  emerge as suitable alternatives.ILE and NGP contributed equally to planning, writing, and critical reading of this manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Antibody repertoire sequencing provides a molecular fingerprint of current and past pathogens encountered by the immune system. Most repertoire studies in humans require measuring the B cell response in the blood, resulting in a large bias to the IgM isotype. The extent to which the circulating IgM antibody repertoire correlates to lymphoid tissue-resident B cells in the setting of viral infection remains largely uncharacterized. Therefore, we compared the IgM repertoires from both blood and bone marrow (BM) plasma cells (PCs) following acute or chronic lymphocytic choriomeningitis virus (LCMV) infection in mice. Despite previously reported serum alterations between acute and chronic infection, IgM repertoire signatures based on clonal diversity metrics, public clones, network, and phylogenetic analysis were largely unable to distinguish infection cohorts. Our findings, however, revealed mouse-specific repertoire fingerprints between the blood and PC repertoires irrespective of infection status. The possibility of personalized medicine is becoming increasingly possible due to the revolution in high-throughput sequencing (HTS) technologies  , with the exception of two time points  due to failed library amplification  of LCMV clone 13, respectively  for 15 animals  for the top 50 clones either before infection (−10 dpi) or 10 dpi for an exemplary mouse from each cohort . We imme2-fold change for all V genes uncovered signatures of differential germline expression between these two time points   found in at least two mice. Quantifying the percent of clones found in multiple mice revealed that both the pooled blood and the BM PC repertoires were largely private, with <7% of clones found within multiple mice . Despite × 1016) . TogetheBoth the large discordance of V gene usage for all cohorts  and the Until now, our analysis had relied on describing the antibody repertoire based on identical CDRH3 amino acid sequences, which inherently ignores any relationship between highly similar antibody sequences. This led us to next ask whether patterns of convergent selection following LCMV infection could be detected if we examined the presence of closely-related antibody sequences. To this end, we first quantified the mean pairwise edit distance, which quantifies the number of amino acid mutations separating two sequences, for each mouse at each time point. The edit distance remained constant throughout the 80 days for all cohorts in both blood and BM PC repertoires , suggestWe finally questioned asked how frequent class-switched IgG clones were detected with identical IgM clones within the same repertoire. We thereby calculated the percentage of IgM clones with an identical IgG CDRH3 sequence in the blood repertoire for each mouse across the 80 days . This quAs the amount of high-throughput antibody repertoire sequencing data increases, attention is being drawn toward the possibility of using information extracted from repertoires as a possible immunodiagnostic. In the context of human patients, this would rely upon sampling circulating lymphocytes in the blood, subsequently sequencing antibody repertoires, and finally deconvoluting immune histories in a potentially pathogen specific manner. As it is difficult to connect clonal information from circulating repertoires to tissue-resident B cells, significant weight is given to the snapshot presented by the blood repertoire. Here we have leveraged an extensively-studied murine viral infection model that allowed for both the deconstruction of antibody repertoires in both circulating and tissue-resident B cells. It has been previously described that cohort-specific signatures arise in the IgG repertoires following LCMV infection, in addition to a higher level of congruence between the blood and BM PC IgG repertoires following chronic LCMV infection relative to acutely-infected and uninfected animals  was taken from accession number E-MTAB-8585 . Wilcoxon-matched pairs signed rank test was used for paired data . One-way ANOVA with Tukey's multiple comparison test was performed for hypothesis testing involving all three infection cohorts with a single experimental variable, with comparisons performed against the uninfected cohort . Two-way ANOVA with Tukey's multiple comparison test was performed for hypothesis testing involving all three infection cohorts and different time points, with comparisons performed against the uninfected cohort . All analyses were performed using GraphPad Prism 8. All error bars represent the standard error of the mean.All statistical significance for unpaired data was calculated using the Kolmogorov-Smirnov test (*All data used in this study and corresponding sample annotations are located on the European Bioinformatics Institute repository under the accession number E-MTAB-8585.AY, AO, and SR wrote the manuscript. AY performed the computational analysis. All authors contributed to study and manuscript design.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.


Medical research is changing into direction of precision therapy, thus, sophisticated preclinical models are urgently needed. In human pathogenic virus research, the major technical hurdle is not only to translate discoveries from animals to treatments of humans, but also to overcome the problem of interspecies differences with regard to productive infections and comparable disease development. Transgenic mice provide a basis for research of disease pathogenesis after infection with human-specific viruses. Today, humanized mice can be found at the very heart of this forefront of medical research allowing for recapitulation of disease pathogenesis and drug mechanisms in humans. This review discusses progress in the development and use of transgenic mice for the study of virus-induced human diseases towards identification of new drug innovations to treat and control human pathogenic infectious diseases. In times of a global pandemic, it is more obvious than ever that human pathogenic viruses are a major threat to global health. The emergence of human pathogenic viruses such as SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) or Zika virus highlights the urgent need of representative preclinical models to study virus entry, replication and spread within an organism as tools to develop efficient antiviral therapies and vaccinations. Aside from that, also several well-known human pathogenic viruses that are characterized in molecular detail like influenza virus, human immunodeficiency virus (HIV) or hepatitis viruses still remain a huge threat to public health.Mechanisms of infection, replication and disease pathology differ widely among different viruses. Suitability of cell culture systems or organoids to obtain comprehensive knowledge about viral characteristics or to test antiviral approaches is highly limited due to the lack of a functional immune system and vasculature. Thus, animal models still represent a unique opportunity to study viral replication processes and antiviral agents in vivo. Rodent models, and especially mice, are widely accepted and the preferred preclinical systems as they are readily available and exhibit low maintenance costs. However, viruses coevolve with their host organisms and, thus, often show a restricted species specificity frequently complicating the research of human pathogenic viruses in mouse models. This narrow host range is often based on the lack of surface entry receptors necessary for virus infection or due to remarkable differences in murine and human innate immune responses upon viral infection. Accompanying this, tissue tropism of human pathogenic viruses can differ significantly in mice causing differences in virus-induced pathology . Hence, Genetically modified and transgenic mouse models represent a sophisticated possibility to overcome interspecies differences in human pathogenic virus research such as species specificity and tissue tropism, allowing to investigate the function of specific human entry receptors, host factors or innate and adaptive immune responses in immunocompetent organisms. These models were of significant importance for scientific progress within the last decades and provided immeasurable insights into virus biology and disease pathogenesis. A decisive milestone was the development of immune system or tissue-humanized mouse models facilitating infection and replication of human pathogenic viruses in a humanized system. With recent advances in human cell or tissue engraftment in transgenic humanized mice, antiviral strategies and vaccines against many human pathogenic viruses already have been or will be evaluated in the future, towards identification of new drug innovations to treat and control human pathogenic infectious diseases. This review aims to provide an overview of different options to genetically engineer and humanize mice. We will discuss several examples of human pathogenic viruses that have been studied in translational, human-relevant preclinical systems see .Due to the absence of permissive mouse models for several human viruses, transgenic mouse strains were developed incorporating essential individual viral genes or even whole virus genomes within their own genome to investigate virus pathogenicity and characterize specific properties of viral genes.Some of the most intensively studied transgenic mouse models expressing viral genomes have been generated to analyze hepatitis B and C virus -induced liver pathology and virus replication. Hepatitis B and C infection share a common phenotype of liver pathology induced by chronic necroinflammation resulting in liver cirrhosis and hepatocellular carcinoma (HCC). HBV cccDNA  persists in infected host cells and the virus can uniquely integrate its genome into the DNA of the infected cell. Most integration events retain the HBx antigen . In sharenvelope [core [precore [X [In order to study the role of individual HBV genes in the development of hepatopathology, immunotolerant HBV-transgenic mice were generated by germline modifications. While the expression of the viral envelope protein did not induce hepatocellular cytopathology, adoptive transfer of virus–antigen primed splenocytes resulted in hepatocellular injury and immunopathology caused by MHC I-restricted antigen-presentation to CD8+ T lymphocytes and envelope-specific antibodies . These denvelope ,6,7, cor[precore  and X [1ecore [X ,11 genesecore [X . Using tecore [X ,14,15,16ecore [X ,18 or byecore [X ,20. Moreecore [X ,22. Several groups have generated transgenic mice expressing individual HCV proteins or viral polyproteins to study the effect on liver pathology, steatosis and HCC manifestations ,27,28,29Since murine hepatocytes do not express the respective virus entry receptors for HBV and HCV, the abovementioned studies did neither allow for virus infection studies nor resemble the whole infection process via the natural route and virus spread as observed within human tissues . Though In line with HBV and HCVtg mice, several attempts have been made to generate HIVtg mice resulting in substantial understanding of virus–host interactions and clinical manifestation of acquired immune deficiency syndrome (AIDS) symptoms or the development of Kaposi’s sarcoma ,34,35,36Further examples of transgenic mouse models expressing HBV, HCV or HIV transgenes can be found in Human pathogenic virus’ species specificity and tissue tropism is often a result of host-specific unique expression patterns of cell surface receptors needed for virus entry. Due to genetic differences of humans and mice, genes encoding necessary cell surface receptors can be lacking or significantly differ in mice. This often results in decreased susceptibility of mice for human pathogenic viruses. This drawback can be overcome by transgenic mice expressing humanized virus entry receptors as a result of genetic modification. ACE2 gene under a human cytokeratin 18 (K18) promoter in the epithelium of lung, colon, liver, and kidneys [ACE2 mice by either microinjection of the hACE gene under the murine ACE2 promoter [ACE2 with the human ACE2 [The current SARS-CoV-2 pandemic is an excellent example of the urgent need of susceptible mouse models for emerging human pathogenic viruses to gain insights into virus biology and to develop antiviral strategies and vaccinations. Earlier studies suggested angiotensin-converting enzyme 2 (ACE2) as essential entry receptor for SARS-CoV viruses . Sequenc kidneys  were ori kidneys ,90. Othepromoter  or by usman ACE2 . These thACE2 and hTMPRSS2 under the murine TMPRSS2 promoter in epithelial cells by applying CRISPR/Cas9 technology [Recently, studies demonstrated interaction of ACE2 with the transmembrane protease/serine subfamily member 2 (TMPRSS2) which proteolytically cleaves ACE2 and the viral S protein. This interaction seems to be essential for viral infection efficiency . Thus, rchnology .PVR gene were successfully generated by germline modifications to allow in vivo studies of PV infection and manifestation of poliomyelitis [hPVR-tg mice after oral administration, which is the primary route of infection in humans [hPVR was expressed under the promoter of the murine orthologue Tage4, mice were more susceptible to oral PV infection. However, only mice of an age of up to three weeks developed symptoms of poliomyelitis even though these mice were more susceptible to oral infection compared to mice expressing hPVR under the human promoter [PVR under the control of diverse promoters [hPVR in the intestine is not solely responsible for infection establishment and indicate that other host factors might be involved in efficient viral infection and tissue permissiveness. Another successful and well-studied example of how to facilitate infection of nonpermissive mice with human pathogenic viruses, is the generation of transgenic mice expressing the human entry receptors for poliovirus (PV) infection. PVs are highly pathogenic for humans and are known to be the causative agent of human paralytic poliomyelitis, which is developed in approximately 1% of infected individuals ,117. In myelitis ,60. Trann humans ,60,61. Wpromoter . Severalromoters ,64. UnfoPVR and lacking the alpha/beta interferon receptor (IFNAR) gene were generated [Interestingly, it was shown that PV is not able to interfere with the murine interferon (IFN) system. Thus, transgenic mice expressing the human enerated . These menerated . This isenerated ,70. NeveNot only the expression and presence of surface receptors have a decisive influence on the permissiveness of mice for human pathogenic viruses. Differences in host determinants and intracellular antiviral responses also play an important role in the different outcome of virus replication, susceptibility of the organism to certain viruses as well as disease pathology in mice versus humans.per se permissive for various, though not all, influenza viruses, although mice do not belong to the group of natural hosts of IAV. Within the last decade, mouse models have played a significant role in understanding pathogenesis and molecular mechanisms underlying IAV infection. Nevertheless, it was shown that infection selectivity based on individual patterns of 2,3- and 2,6-linked sialic acid residues, serving as entry receptors for avian and human IAVs respectively, varies in respiratory cell populations in mice and humans [Several human pathogenic viruses such as influenza A viruses (IAV) have the capability to infect laboratory inbred strains of mice, such as BALB/c, C57BL/6 or DBA/2j . Amongstd humans ,122. Mx gene plays a major role in innate immune responses to control IAV infection. While the majority of wild type mice express a functional Mx gene and, thus, show resistance to IAV infection, several inbred mouse strains demonstrate large deletions or nonsense mutations within the Mx locus [Mx gene survived otherwise lethal doses of 1918 H1N1 pandemic and avian H5N1 IAV [Mx in the restriction and regulation of IAV infections. Additionally, depending on the infectious dose given and the strains used, IAV infection shows a more severe disease phenotype and lethality in mice compared to humans. Studies suggested that the IFN-regulated Mx locus ,124. ComH5N1 IAV  . ThMx genes do not necessarily reflect human antiviral intracellular immune responses. These innate restriction factors represent effective species barriers that need to be overcome by viruses to invade the human host. IAV poses a constant threat to humans since zoonotic viral transmissions can cause severe disease and may give rise to pandemics. Thus, transgenic mice expressing the entire human Mx locus could be explicitly used to investigate the role of human MxA as a restriction factor that must be overcome by zoonotic IAV strains. Interestingly, these mice exhibited a higher resistance against avian H5 an H7 viruses but were readily susceptible to seasonal human IAVs of H1N1 or H3N2 origin that already efficiently adapted to the human host [However, murine and human antiviral host factors can essentially differ and mice expressing murine man host . HLA-A*0201 [HLA-DR3 [Not only cell intrinsic immune response factors, but also factors involved in cellular innate and adaptive immune responses are essential for IAV-induced pathology and present a critical component of antiviral immunity. Especially the emergence of zoonotic IAV with surface glycoproteins that are distinct from seasonal human strains pose a risk and demand preclinical systems to investigate vaccination approaches and their efficacy in humans. Human transgenic mice expressing the human MHC class I allele A-A*0201 ,95 or th[HLA-DR3  provided[HLA-DR3 ,95,96. F[HLA-DR3 ,129,130.Humanized mice with functional human cell or tissue engraftment have advanced research in the field of human pathogenic virus infections. Engraftment of human hematopoietic stem cells (HSCs) that develop into functional human immune systems even allows the study of bloodborne virus infections as well as virus-induced immune responses and immunopathology-mediated disease pathogenesis, bridging the gap between rodent models and the clinics.protein kinase, DNA-activated, catalytic polypeptide (PRKDC) gene, resulting in severe combined immunodeficiency (scid), or knockout of recombination-activating genes (RAG) lead to impaired development of T and B cells [interleukin-2 receptor common gamma chain (IL-2rγ) allows for even more efficient human cell and tissue engraftment due to severe impairments in multiple cytokine complexes and a profound defect in natural killer (NK) and NKT cell development [scid gamma), NRG (NOD Rag gamma) and NOG (NOD scid gamma) mice. The latter contain a IL2rg null mutation that does not even allow for cytokine binding. There are three conventional ways to engraft immunodeficient mice with functional human cells: (i) intravenous injection of human bone marrow, cord blood, or granulocyte-colony-stimulating factor (G-CSF) cytokine-mobilized peripheral blood mononuclear cells (PBMCs) which are all rich in human HSC activity, leads to the engraftment of a naive human hemopoietic system in the murine recipient (HSC model) [Currently, there are few major mouse models used with varying degrees of immunodeficiency resulting in different levels of graft acceptance and tolerance for human cells . Th B cells ,134,135.elopment . Immunodelopment ,139,140.C model) ,142. (iiC model) ,144. (iiC model) ,146. IL-3, GM-CSF and stem cell factor (SCF), therefore combining the features of the highly immunodeficient NSG mouse with the expression of cytokines that support the stable engraftment of myeloid lineages and regulatory T cell populations [signal-regulatory protein alpha (SIRPα), protecting human cells from being phagocytosed. Finally, cytokine-encoding genes M-CSF, IL-3, GM-CSF and thrombopoietin are humanized by knock-in replacement, resulting in defects in mouse cell populations dependent on the respective mouse cytokine and, thus, support for engraftment of transplanted human cells [HLA-A2/HHD immunodeficient mice express human HLA class 1 heavy and light chains allowing for the generation of functional human T cell subsets with HLA-restricted immune responses [HLA-DR4 genotype) fused to the I-Ed MHC class II molecule. The presence of these HLA-DR4-IE transgenes allows enhanced HLA-DR-matched HSC engraftment and subsequent human T and B cell development [HLA-A2 and HLA-DR1 providing a platform for the generation of humanized models that better resist graft versus host disease [Aforementioned humanized mouse models with fully functional human immune responses have been applied for studies with different human pathogenic viruses. However, although humanized mice with engrafted human cells provided suitable tools to study human-specific pathogens in the past, these models have been shown to possess some major drawbacks such as development of graft versus host disease, the absence of erythrocytes and neutrophils within reconstituted human immune systems, low numbers and functionally impaired human myeloid cells, dominance of immature B cells as well as minimal production of antigen-specific IgG class antibodies ,148,149.ulations . Anotheran cells . NSG-HLAesponses . DRAG mielopment . Intereselopment . Most re disease . These different humanized transgenic mice are currently in use to model the human immune system in scenarios of health and pathology, and serve as a basis for the analysis of human pathogenic viruses, enabling the evaluation of therapeutic candidates in an in vivo setting more relevant to human physiology, in the future.Improvement of humanized mouse models has opened up a new avenue for studying bloodborne viruses such as HIV as well as hepatitis B and C viruses, where knowledge was previously gained in viral gene/genome transgenic mice. In addition, hemorrhagic fever-inducing viruses such as dengue virus (DENV) or Ebola could be finally investigated in vivo by application of the aforementioned mouse models that allow for human immune system induced pathology. The most extensively studied virus in transgenic immune system humanized mice is HIV. HIV research is confronted with various challenges such as the species barrier that was overcome by engraftment of human fetal thymic or lymph node implants allowing for acute infection. However, the RNA genome of HIV is reverse-transcribed into DNA and integrated into the host genome, resulting in latent infections difficult to clear by the host immune system. Additional humanization and especially the use of BLT mice finally allowed for the modelling of HIV transmission via vaginal and rectal routes, latency and a dynamic interplay of HIV-specific cellular immunity and viral escape from immune pressure . FoPreclinical studies on mosquito-borne DENV causing systemic disease have proven to be a difficult task because the virus demonstrates a tropism for different subsets of human cells . In addiAFC8), which induces mouse liver cell death after administration of synthetic drug AP20187, were used. AFC8-hu HSC/Hep mice supported HCV infection in the liver and generated a human immune T cell response against HCV antigens. HCV infection induced liver inflammation, hepatitis, and fibrosis, which correlated with activation of stellate cells and expression of human fibrogenic genes [In the past years, human-associated viruses have been extensively studied in mice with humanized immune systems. Nevertheless, study of hepatotropic pathogens such as hepatitis B and C viruses remained elusive due to the lack of human hepatocytes in the mouse liver. Human liver chimeric mice are still the gold standard for allowing hepatotropic infections, even though the highly immunocompromised status of these engrafted mice precluded liver pathogenesis mediated by human immune surveillance. Thus, transgenic mice with dual humanization of immune system and liver were established . Hoic genes ,77. NeveHLA-A2 transgenic mice with human hematopoietic stem cells and liver progenitor cells, supported HBV infection and establishment of persistent infection. Human immune responses, albeit impaired in the liver, chronic liver inflammation and liver fibrosis were detected in infected animals. In addition, application of an HBV neutralizing antibody efficiently inhibited HBV infection and associated liver diseases in humanized mice. Furthermore, HBV-mediated liver disease was found to be associated with high levels of infiltrated human macrophages with an M2-like activation phenotype, which has been shown to be associated with accelerated liver fibrosis and necrosis in patients with acute HBV-induced liver failure [The A2/NSG-hu HSC/Hep mouse, a humanized mouse model with both human immune system and human liver cells by reconstituting immunodeficient human  failure .Although these transgenic humanized mouse models have not been used extensively in research on human pathogenic viruses yet, their benefit especially in drug and vaccination studies is undeniable. Interestingly, it already has been shown that DRAGA mice can be used to generate cross-reactive, human anti-influenza monoclonal antibodies (hu-mAbs). DRAGA mice were found to be suitable for influenza virus infection, as they can clear a sublethal infection and sustain a lethal infection with influenza virus. A single administration of viral hemagglutinin protein-specific hu-mAbs in IAV-infected DRAGA mice significantly delayed lethality by reducing lung damage . Successful establishment of transgenic and especially transgenic humanized mice provided immeasurable opportunities to advance medical research on infectious diseases that were previously inaccessible. The transgenic mouse models described in this review are, amongst others, groundbreaking for the understanding of human pathogenic virus infections and their disease pathology. Along that line, these mouse models provide the possibility to study emerging virus-induced diseases, as discussed for SARS-CoV-2. Since some viruses have the unique ability to cross species boundaries and adapt to new hosts, humanized mouse models will continue to play a crucial role in understanding newly emerging or human-adapting viruses, in the future. However, existing models are still far from being perfect. A mouse model that can be applied to study all known and newly emerging human pathogenic viruses while completely mimicking virus-induced disease pathogenesis does not exist and researchers still need to consider the scientific questions in the respective areas of the disease to select the best suitable animal model. While virus entry receptor or antiviral host factor humanized mice are a great model to study virus infection, replication and the role of single human genes involved in antiviral immune responses, they are not suitable to evaluate disease pathogenesis or efficacy of antiviral approaches as the complexity of human immune responses is not necessarily reflected. Especially the analysis of possible vaccination candidates or antiviral approaches additionally requires immune system humanized mice. Amongst these, HUMAMICE are highly sophisticated humanized models that allow e.g., to study antigen specificity of human immune cells, however, without additional humanization, these animals are not susceptible for infection with human pathogenic viruses such as SARS-CoV-2.The ideal future model for most human pathogenic virus infections will be an animal model with a reconstituted human immune system presenting antigens in the human HLA context, constitutively expressing human cytokines necessary for efficient immune cell development. In addition, specific human organs or humanized cell types would be needed to mimic viral transmission routes and disease pathology. Such a model does not exist yet, but with the current advances in technology it might be only a matter of time to have it available to study human specific virus infection complexity and to develop more efficient vaccine and treatment strategies. Furthermore, there still is the sustained need to identify and replace human-specific factors that are absent in mice but are needed for optimal human cell differentiation and function to decrease or even prevent the development of graft versus host disease that still occurs in many of the human immune system engrafted models.Thus, key improvements in specific areas are still required, however, with careful characterization, the future will yield more appropriate preclinical models, supporting more rapid and relevant translational drug discovery and development.


Nine species-level potential human pathogens were identified in riverine sediments including Acinetobacer lwoffi and Arcobacter skirrowii, two pathogens associated with the cattle microbiome. Microbial community composition at locations in close proximity to intensive agriculture was not resistant nor resilient to agricultural runoff disturbance within 5 months post-disturbance but did reach a new, stable microbial composition. From this data, we conclude that sediment microbial community composition is sensitive and shifts in response to chemical and microbial pollution from intensive agriculture, has a low capacity to resist infiltration by non-native, harmful bacteria and, overall, the natural buffering capacity of freshwater ecosystems is unable to fully resist the impacts from agricultural press disturbance.Microorganisms are critically important for the function of surface water ecosystems but are frequently subjected to anthropogenic disturbances at either acute (pulse) or long-term (press) scales. Response and recovery of microbial community composition and function following pulse disturbance is well-studied in controlled, laboratory scale experiments but is less well-understood in natural environments undergoing continual press disturbance. The objectives of this study were to determine the drivers of sediment microbial compositional and functional changes in freshwaters receiving continual press disturbance from agricultural land runoff and to evaluate the ability of the native microbial community to resist disturbance related changes as a proxy for freshwater ecosystem health. Freshwater sediments were collected seasonally over 1 year in Kewaunee County, Wisconsin, a region impacted by concentrated dairy cattle farming, manure fertilization, and associated agricultural runoff which together serve as a press disturbance. Using 16S rRNA gene amplicon sequencing, we found that sediments in locations strongly impacted by intensive agriculture contain significantly higher abundances ( Freshwater ecosystems are a vital natural resource, providing sources of drinking water, food, animal habitats, and recreation. Sediment microbial communities within freshwater ecosystems support important functions including carbon and nitrogen cycling due to their high abundance, diversity, and stability . SedimenArcobacter and Acinetobacter and more well-known pathogens including methicillin resistant Staphylococcus aureus and toxic Escherichia coli  pollution and degraded water quality in rivers and streams . Livestohia coli . Input on events . The resn events . Severaln events  providesn events . Understn events .In our study, the primary research question is: how does agricultural land runoff impact microbial community composition and function in freshwater sediments in a region of intensive agriculture that suffers from reduced freshwater quality? We have addressed this question in Kewaunee County, Wisconsin, which is home to 16 Confined Animal Feeding Operations  and more than 160 smaller farms. Each year, approximately 151,000 tons of manure solids and 784 million gallons of liquified manure are applied onto land in the county between the mandated spreading dates of April 15–December 31 , servingCyanobacteria and members of the genera Pseudomonas and Novosphingobium); and (3) potential pathogens associated with the cattle microbiome will be detected more frequently in freshwater sediments at river locations impacted by manure runoff from intensive agriculture. We tested these hypotheses using 16S rRNA gene sequencing combined with exact sequence matching and functional inferences.We hypothesized that (1) sediment microbial communities in the highly impacted Kewaunee River watershed will be resilient to press disturbance from manure fertilization runoff due to long-term stress and adaptation to pollutants in the region; (2) microbial community composition will shift seasonally during the manure fertilization period toward communities enriched with copiotrophs  from a dairy CAFO farm was collected in May 2017 and two composite manure samples from a < 200 head beef farm were collected in May and September 2017 for reference. A total of 68 sediment samples were collected for this study.The three major watersheds within Kewaunee County, Wisconsin have been fully described elsewhere . BrieflySediment and manure were subsampled in the laboratory for a total of two separate DNA extractions. DNA was extracted from 0.5 g of sediment or manure using the DNeasy PowerSoil Kit (Qiagen) according to manufacturer instructions. Duplicate DNA extractions were pooled for downstream analyses.PRJNA555250.Universal primers were used to amplify the V3–V4 hypervariable region of the 16S rRNA gene followed by next generation sequencing (NGS) . Paired-Potential pathogens were identified at the species level using the DADA2 extension for 100% exact sequence matching of sample sequences to sequenced reference strains within the Silva database . To iden, phosphate , total coliform bacteria, and E. coli were measured in river water from sediment collection sites. Nitrate and phosphate were measured using Ion Chromatography in unaltered water samples . Total coliform bacteria and E. coli were measured in 100 mL of river water within 24 h of collection using the USEPA approved Colilert Quanti-Tray/2000 Method , and a portion of these results have been reported in previous work . Agricul0 Method . Metal cphyloseq package in R was used to calculate relative abundances of raw ASV counts based on taxonomic classification  and the intensity of manure fertilization at this timepoint. We postulate that no significant difference in alpha or beta-diversity between pre-and post-disturbance samples indicates community resistance while a significant difference pre- and post- disturbance with a return to the pre- disturbance state indicates community resilience. Resistance and resilience indices at the alpha-diversity level were calculated using estimators of sample diversity, richness, and evenness including Chao1, ACE, Shannon, and Inverse Simpson diversity indices. Resistance and resilience at the beta-diversity level were estimated by calculating the difference in beta-diversity between pre- and post-disturbance microbial communities using both PERMANOVA  and ANOSIM  tests, as in Resistance and resilience measurements of sediment alpha diversity (within sample) and beta-diversity (between samples) pre- and post-manure fertilization disturbance were calculated with February samples considered “pre-disturbance,” May samples considered “disturbance,” September samples considered “4 months post-disturbance,” and October samples “5 months post-disturbance.” We considered May sample” disturbance” due to the start of the fertilization period  is comprised of members of the phyla analysis . Additiomicrobia , both ofudomonas .p > 0.05). However, the diversity of microbial communities within sediment samples from highly impacted locations was significantly higher (p < 0.05) 4 months post-disturbance compared to their pre-disturbance state  was estimated before, during and after manure fertilization disturbance using Chao1, ACE, Shannon, and Inverse Simpson diversity metrics within samples clustered by sampling location . When coce state  and fresce state  have alsp = 0.001 main test, p < 0.01 post hoc t-tests; p = 0.83 post hoc t-test, Spatial differences in the abundance of dominant taxa are evident from p < 0.01) in Branch 2 sites of the Kewaunee River compared to OKW sites; the effect size estimate displayed in Thiobacillus, Methylotenera, Crenotrhix, Nitrospira, and Rhodoferax among others. Of ASVs mapped to the species level, those that were more abundant in Branch 2 sites include Arenimonas aquatica, Leptothrix cholodnii, Nitrospira defluvii, Flavobacterium flevense, Polaromonas naphthalenivorans, Zoogloea oleivorans, Flavobacterium psychrolimnae, and Flavobacterium xinjiangense. Members of these genera and associated higher level taxonomy are commonly found in freshwater environments  and the locations least impacted by agricultural pollution (OKW sites) using differential abundance analysis . A totalronments  but are ronments . In part systems , and Flae manure , suggestE. coli, and 12 metals previously reported  in Branch 2 samples than OKW samples, indicating potential differences in microbial community function in areas highly impacted by intensive agriculture compared to unimpacted locations. This finding was also reflected by the ASV taxa mapped to these functions including Rhodoferax and Nitrospira which were found to be both more abundant in highly impacted sites , microbial community composition of the post-disturbance samples were more similar to one another than samples pre- or during manure fertilization disturbance . This result suggests sediment microbial communities impacted by manure fertilization disturbance reach a new steady state by 5 months post-disturbance  and beta diversity (community composition). Within sample sediment microbial richness and evenness did not differ significantly pre- and post-disturbance at any sampling location ; howeverturbance .As stated previously, the branches of the Kewaunee River watershed are differentially impacted by manure fertilization and agricultural runoff; thus, we analyzed if each individual river branch displayed the same temporal pattern and lack of resistance/resilience identified within the watershed as a whole. We discovered that the lack of resistance and resilience identified in Kewaunee River sediments was driven by only two of the four branches: the highly impacted Branch 2 and Downstream. The other two, less impacted branches of the Kewaunee River were not dissimilar over time and were instead resistant to microbial community compositional changes . Additioonly identified in branches of the Kewaunee River that are highly impacted by intensive agriculture disturbance. This temporal trend disappears entirely in less impacted branches and the unimpacted OKW sites, supporting our hypothesis that agricultural runoff disturbance and associated spatial differences drive changes in microbial community composition in these sediments, not seasonal differences. These results highlight the complexity of analyzing the impacts of press disturbance in natural environments.One limitation of this study is that samples were collected over 1 year; thus, additional sample collection over multiple years is necessary to fully tease apart the relationship between disturbance, resilience, and seasonal temporal turnover in freshwater sediments. It is important to note that the temporal changes in microbial community composition associated with agricultural runoff disturbance in this study could also be associated with seasonal changes that occur in temperate regions such as Kewaunee County, WI. For example, the concentration of dissolved oxygen in surface waters is known to change seasonally with temperature  but alsoUsing manure as cropland fertilizer introduces a number of human health risks into the environment including potential pathogenic bacteria . Due to Acinetobacter lwoffii, Aeromonas sobria, and Arcobacter skirrowii  under accession: KH and RB designed the sampling regimen and identified collection locations. RB collected and processed samples and wrote the initial manuscript draft. RB and AB analyzed sequence data and performed statistical analysis. KH, AB, and SS edited and provided suggestions for the final version of the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The perceived threat of a contagious virus may lead people to be distrustful of immigrants and out-groups. Since the COVID-19 outbreak, the salient politicized discourses of blaming Chinese people for spreading the virus have fueled over 2000 reports of anti-Asian racial incidents and hate crimes in the United States.The study aims to investigate the relationships between news consumption, trust, intergroup contact, and prejudicial attitudes toward Asians and Asian Americans residing in the United States during the COVID-19 pandemic. We compare how traditional news, social media use, and biased news exposure cultivate racial attitudes, and the moderating role of media use and trust on prejudice against Asians is examined.A cross-sectional study was completed in May 2020. A total of 430 US adults  participated in an online survey through Amazon’s Mechanical Turk platform. Respondents answered questions related to traditional news exposure, social media use, perceived trust, and their top three news channels for staying informed about the novel coronavirus. In addition, intergroup contact and racial attitudes toward Asians were assessed. We performed hierarchical regression analyses to test the associations. Moderation effects were estimated using simple slopes testing with a 95% bootstrap confidence interval approach.P=.02), had a personal infection history , and interacted with Asian people frequently in their daily lives  reported more negative attitudes toward Asians after controlling for sociodemographic variables. Relying more on traditional news media  and higher levels of trust in social media  were positively associated with prejudice against Asians. In contrast, consuming news from left-leaning outlets  and neutral outlets  was linked to less prejudicial attitudes toward Asians. Among those who had high trust in social media, exposure had a negative relationship with prejudice. At high levels of trust in digital websites and apps, frequent use was related to less unfavorable attitudes toward Asians.Participants who identified as conservatives (β=.08, Experiencing racial prejudice among the Asian population during a challenging pandemic can cause poor psychological outcomes and exacerbate health disparities. The results suggest that conservative ideology, personal infection history, frequency of intergroup contact, traditional news exposure, and trust in social media emerge as positive predictors of prejudice against Asians and Asian Americans, whereas people who get COVID-19 news from left-leaning and balanced outlets show less prejudice. For those who have more trust in social media and digital news, frequent use of these two sources is associated with lower levels of prejudice. Our findings highlight the need to reshape traditional news discourses and use social media and mobile news apps to develop credible messages for combating racial prejudice against Asians. Since the COVID-19 outbreak, the United States has become an epicenter, surpassing 5 million confirmed cases as of August 2020. The behavioral immune system theory posits that people’s tendency to avoid the risk of pathogen contagion is driven by cognitive and affective responses to informational cues . EvidencTwo psychological mechanisms are relevant in explaining the sharp increase in prejudice against out-groups when people cope with high levels of anxiety about infectious diseases. First, the contact hypothesis -11 postuSecond, consuming information in news media and social media offers indirect forms of mediated or vicarious contact, thus shaping attitudes toward out-groups when people have minimal direct interaction with minority groups during shelter-in-place orders ,14. EarlTo date, little research has linked the two aforementioned mechanisms to bias toward stigmatized groups in the context of a global pandemic. Based on a cross-sectional online survey of 430 participants, we jointly examined two psychological mechanisms underlying the activation of prejudice against Asians and Asian Americans in the United States during the COVID-19 pandemic: intergroup contact and indirect contact of media consumption. To reflect the hyper-choice and hyper-partisan news environment, we differentiated three functionally different concepts of media consumption: media use, media trust, and media bias. Next, we compared how traditional news, social media use, and news apps cultivated racial attitudes. Last, this research lent empirical support to the moderating role of media use and trust in prejudice against Asians. Together, the timeliness of our results will yield theoretical and practical insights into developing public health interventions for reducing racial discrimination linked to the COVID-19 pandemic.Literature on the behavioral immune system has documented that humans’ subjective risk perceptions of contagious diseases will trigger a series of cognitive, affective, and behavior mechanisms to avoid pathogen infection ,2. The tDespite a lower fatality rate than Middle East respiratory syndrome (MERS) and Ebola, the SARS-CoV-2 virus has caused one of the highest death tolls worldwide, and there is still no vaccine available . As the In addition to serving as situational cues to activate aversion of minority groups, news reports offer indirect mediated contact that possibly shapes prejudice toward ethnic minorities ,14. DiffDecades of cultivation theory and news framing research have proved the direct influence of using traditional media, mostly newspapers and TV newscasts, on cultivating audiences’ prejudicial views of out-groups . PersistAlthough the aforementioned studies are insightful, relying on one average scale to measure individuals’ exposure to traditional media outlets  limits our understanding of how American audiences receive information from hyper-choice, hyper-partisan, networked news environments ,14,23,29In addition to the well-documented influence of traditional media exposure on prejudice, the role of social media and digital outlets in shaping users’ views of ethnic minorities cannot be overlooked. Social media was and is an essential source for coping with the MERS outbreak and the ongoing pandemic -33. At tMedia trust and media use are correlated but conceptually different . A worldLast, relying on conservative or liberal media could also be a significant determinant of negative attitudes toward out-groups. An online survey with 422 Italians demonstrated a positive relationship between exposure to right wing newspapers and TV newscasts, and prejudice against immigrants after controlling for participants’ political ideology. Conversely, using left-leaning news outlets showed a negative association with prejudice .Guided by the behavioral immune system theory -5, assocH1: The frequency of direct intergroup contact will be positively associated with prejudicial attitudes toward Asians.As no prior research distinguishes how modalities of news, news trust, and media bias relate to virus-activated prejudice, we propose the following research questions (RQs):RQ1: How will media use, media trust, and media bias be associated with prejudicial attitudes toward Asians?RQ2: How will media trust moderate the relationship between media use and prejudicial attitudes toward Asians?In May 2020, we recruited participants from Amazon’s Mechanical Turk (Mturk) program, a dominant internet-based platform for gathering online samples in behavioral science fields . A shortR2=0.13) with 99% power and 18 predictors indicated a minimum sample of 275 [To determine the required sample size to achieve desired statistical power, we performed an a priori estimate using G*Power version 3.1 (Heinrich-Heine-Universität Düsseldorf) before data collection . A mediue of 275 . To ensuThe Institutional Review Board office of Renmin University of China approved the research protocol, and responses were collected via Qualtrics software . The first author at a US university in the southwest was provided with an anonymized data set containing no identifiable private information connected to participants. Ethics exemptions were obtained at coauthors’ institutions before launching the survey. At the beginning of the survey, the respondents agreed to participate in the research voluntarily for receiving compensation. Next, respondents answered questions related to traditional news exposure, social media use, perceived trust, and their top three news channels for staying informed about the novel coronavirus. In addition, the frequency of intergroup contact and racial attitudes toward Asians were assessed. The survey took approximately 16 minutes to complete .We used a common measure ,37,51 toRespondents rated how often they get news about COVID-19 from five traditional sources including print newspaper or magazine, radio, local TV, national network television, and cable TV on a 4-point scale with endpoints labeled “never” and “often” . Two single 4-point scales measured social media use  and news websites or app use .To gauge different levels of media trust in news sources, we asked participants to indicate how much they trust information about COVID-19 from the aforementioned media outlets  . Trust iExposure to biased news sources was assessed by asking respondents to select the top three news channels they frequently visited for gathering COVID-19 news. We obtained a list of 31 popular news brands across a wide range of platforms from the Pew Research Center’s survey of US media polarization and the 2020 election . We thenWe modified a measure from  to assesAs news consumption is associated with sociodemographic variables, we measured common characteristics for statistical control ,37,43. RF8,421=17.65, P<.001). Specifically, participants who were married , held conservative beliefs , and had a personal infection  reported more negative attitudes toward Asians.P<.001) after controlling for demographic characteristics, supporting H1. Respondents who interacted with Asian people frequently in their daily lives were more inclined to hold unfavorable attitudes toward Asians.Turning to the first hypothesis, we found a positive association between intergroup contact and prejudice . Reading print newspapers and watching TV newscasts about COVID-19 was positively associated with higher levels of prejudice . Regarding the main effects of media trust, the results suggested that the more people trusted news circulated on social media, the higher their prejudicial attitudes were . Trust in traditional media and digital channels had no significant associations with negative feelings about Asians. Notably, relying on liberal media  and balanced news sources  for staying informed about the COVID-19 epidemic was significantly associated with lower levels of negative attitudes toward Asians.For RQ1, media use, trust, and reliance on biased sources showed significantly divergent relationships with prejudicial attitudes toward Asians . The interaction term of trust and social media use was statistically significant . We also found a similar significant interaction effect of websites and apps trust and use on prejudice . To explore how variation in media trust changes the relationship between use of social media or digital news and prejudice, we conducted simple slopes analyses. As depicted in The second RQ explored whether news trust moderated the relationship between media use and prejudice against Asians. Interaction terms explained a 3% increase in variance in prejudice (Drawing from the behavioral immune system theory, intergroup relations, and media psychology literature, this study is a novel attempt to investigate the joint mechanisms of direct intergroup contact and mediated contact offered by diverse news sources on associating with prejudicial attitudes toward Asians during the COVID-19 outbreak. In accordance with previous studies ,24,26, pWe empirically examined the associations between three forms of indirect mediated contact and prejudice. The findings reveal the divergent role of news exposure, trust, and political-leaning media in racial attitudes when political orientation and intergroup contact are controlled for. Relying more on traditional news media was positively related to prejudice toward Asians, supporting the TV news’ capability to activate or reinforce prevalent racial stereotypes -30. In lRegression results suggest habitual selections of news outlets with ideological leanings are a more driven factor than the modalities of media in associating with disease-activated racial attitudes. Consuming news from left-leaning and neutral outlets was associated with less prejudicial attitudes toward Asians holding other factors constant. Journalistic powerhouses with liberal and centrist perspectives are more critical of the current administration’s effort in curbing the pandemic and attribute the blame to uncoordinated policy responses at all levels of governments. Although conservative news such as Fox and liberal outlets such as CNN shared similar keyword use of mentioning China and the novel virus, a recent study found CNN news stories featured more scientific statements from Dr Anthony Fauci and Dr Deborah Birx, and emphasized more protective responses such as social distancing and lockdowns than their conservative counterparts did . LiberalIt is worth highlighting the positive link between social media trust and prejudice. Trust in social media was positively associated with prejudicial attitudes toward Asians. The nature of filter algorithms and selective exposure patterns might afford social media users to be exposed to or spread unfiltered discriminating language about Asians. Using social media as news sources increases users’ probability to encounter trending anti-Asian sentiment, even when users do not actively seek such information. People tend to trust information created and shared by friends and family members within homogenous networks . Recent Notably, this study contributes to existing literature by identifying trust in social media and digital websites and apps as a boundary condition of the news exposure-prejudice relationship. Among those who used social media infrequently, those who trusted social media more reported higher prejudice against Asians than those who trusted social media less. However, this difference became less pronounced among heavy users. Likewise, for those who had high levels of trust in digital websites and apps, frequent use was related to less unfavorable attitudes toward Asians. It could be possible that heavy users of social media and digital news have advanced media literacy of filter algorithms, equipping them with higher efficacy to move beyond personalized feeds and search for less politicized information and diverse perspectives. In other words, frequent use of social media and digital sources buffers the relationship between trust in social media and digital apps, and prejudice linked to COVID-19. Together, these results suggest that researchers should consider using nuanced measures of media exposure, media trust, and media bias to generate more accurate conclusions. In a hyper-choice networked news environment, Americans’ use of and trust in traditional media, social media, and well-known news brands with distinct political leanings have more complex relationships with their disease-ignited prejudice toward minority groups than previously assumed.We acknowledge limitations of this study. First, the self-selected sample might reflect respondents’ heightened concerns about COVID-19–related topics. In addition, the cross-sectional study did not guarantee the causal effects of intergroup contact and news use on influencing racial attitudes. Hence, it is possible that participants who held negative views about the Chinese government or Chinese people selected to consume anti-Asian coverage on traditional outlets and social media because those news reports confirmed their existing viewpoints. Given our correlational findings, longitudinal studies will expand upon the results to establish the causal relationships. Second, the study did not directly measure the prevalence of anti-Asian discourses circulated via all media outlets since the major outbreak in the United States. However, a recent study analyzing Google Trends data between December 2019 to March 2020 showed that the “Chinese virus” rhetoric has led to a sharp increase in search rates for anti-Chinese slurs. Therefore, we can confidently infer that an increase in search rates reflected trending news reports and social media discussion related to anti-Asian sentiment and racist attacks .First, in light of these limitations, future research should content analyze the prevalence of anti-Asian sentiment on different types of media and correlate with public attitudes to replicate these findings. Second, as citizens can access the same news content from multiple modalities, conventional categorizations of media use might not capture a full picture of people’s news consumption. A granular examination of website-tracking data to pinpoint what types of news coverage individuals frequently visit for coping with the pandemic will be informative. Last, future inquiries can compare the influence of negative portrayals of Asians circulated through traditional media and social networking sites on out-group attitudes and behavioral avoidance using experimental designs.Experiencing racial discrimination among the Asian population during a challenging pandemic could cause poor psychological outcomes and exacerbate health disparities . The res


After the first emotional improvement, patients experienced six emotional deteriorations , while their function consistently increased . Clinicians should be aware of the relationship between emotional changes and functional progression as well as the occurrence of emotional fluctuations when supervising and consulting patients during postoperative rehabilitation. A larger study is warranted to generalize the results.An interrelationship between psychological and physical health is generally accepted in the field of sports medicine. This preliminary study explored the association between emotional changes and functional outcomes and aimed to describe how each aspect progresses during postoperative rehabilitation. Four collegiate student-athletes  who underwent supervised postoperative rehabilitation due to a lower-extremity injury volunteered for participation in the study. Emotion was quantified using a visual analogue scale prior to and after each session while self-reported function using the Lower-extremity Functional Scale was assessed every eight sessions throughout rehabilitation. There was a moderate correlation between emotional changes and functional outcomes ( Sports injuries often require surgical intervention. During postoperative rehabilitation, patients (athletes) experience negative psychological responses such as frustration and self-doubt ,2. The qThe interactive relationship between emotional status and functional outcomes is generally accepted in the field of sports medicine. For example, individuals with higher degrees of psychological well-being display a better immune function  or physiThe major components of postoperative rehabilitation generally consist of aerobic, flexibility, resistance, and balance exercises in conjunction with therapeutic modalities . The rehTherefore, in this preliminary study, we sought to describe the association between emotional changes and functional outcomes and to describe how each aspect progresses during the course of postoperative rehabilitation. Emotion was quantified using a 100 mm VAS  before aCollegiate student-athletes  who underwent an orthopedic surgery due to a lower-extremity injury were recruited through convenient sampling between February 2019 and January 2020. A lower-extremity injury was defined as a lower-extremity musculoskeletal condition that required surgery. Surgery was defined as a medical intervention performed by an orthopedic surgeon, aiming to fix or repair injured structures. Once a surgery date was scheduled after consulting with an orthopedic surgeon, patients were informed about the study. After a full understanding of the eligibility criteria and the testing procedures, patients gave written informed consent and determined to participate in the study. The informed consent form and the study procedures were approved by the Institutional Review Board of the Kyung Hee University (KHSIRB-19-001). Patients who had surgery in the preceding 48 h prior to obtaining the baseline data collection were excluded. Patients who had the following conditions (either pre-existing or current) for the last six months or who were taking medications due to these conditions were also excluded: (1) any medical conditions  that may influence emotional and functional status and (2) mental disturbance, defined as any health condition affecting emotional and/or behavioral change, other than one from the musculoskeletal condition that met the inclusion criteria.Eleven patients initially volunteered but seven were subsequently excluded—four were excluded because the baseline data were not obtained within 48 h from surgery, two were discontinued from rehabilitation because of facility shutdown due to the coronavirus disease 2019 pandemic and one transferred to another clinic due to the long distance between home and the rehabilitation clinic. Therefore, four patients were finally analyzed , who performed all dependent measurements. Each patient’s number of rehabilitation sessions (number of clinic visits) and time for each rehabilitation session (in minutes) were recorded. Specific therapeutic interventions as classified by six categories  were also obtained.Emotion was recorded using a modified VAS  before aDifferent numbers of clinic visits (number of rehabilitation sessions) for each patient yielded different amounts of data points. To conduct statistical analysis, data for emotion and function were session-normalized using a polynomial function . The mear) was calculated to determine the bivariate relationships (p < 0.05) among the datasets of the emotion  and that of function. To examine the presence of ups and downs in each dependent measurement throughout the rehabilitation period, two-way (time × session interaction: emotion) and one-way (session effect: emotion before and after each session and function) analyses of variance (p < 0.05) were performed. To determine ups and downs in reference to the values at the first session (and to reduce the type I error), Dunnett’s comparisons were performed as post-hoc tests (p < 0.10). To test differences in therapeutic interventions  across patients, a chi-square test using binary variables was conducted (p < 0.05). To determine practical significance, Cohen’s d effect sizes (ES) were calculated when statistical differences existed [Pearson correlation coefficient  and function. For overall emotion, the r value was 0.58 (p < 0.0001).All comparisons in the bivariate correlations were statistically significant . Regardless of session , however, overall emotion was improved by 6% after each session of rehabilitation . Meanwhile, regardless of time , patient emotions were altered over the rehabilitation period , patients experienced six emotional downs throughout rehabilitation . There was no statistically significant difference among emotions recorded after sessions  consistently progressed after the first improvement (33 points) at the 9th session  and there was no functional deterioration . Therapeutic modality (29%) and resistance exercise (28%) were the two most frequent treatments, followed by aerobic (16%), balance (11%), flexibility (11%), and plyometrics (5%) exercises. On average, our patients began rehabilitation 2.5 days after their surgery, visited the clinic 50 times (50 sessions of rehabilitation) during the rehabilitation period of 17.2 weeks (2.9 times per week) and participated for approximately two hours (122-min) per rehabilitation session . The sixOur hypotheses were supported in that emotional changes were moderately correlated with functional progressions; a single-session of rehabilitation resulted in a slight emotional improvement and there was emotional fluctuation along with consistent functional progression. The results of our descriptive data offer a better understanding of how collegiate student-athletes psychologically and physically progress during postoperative rehabilitation. Our study supports that, when patients appear to experience emotional struggle or remain at a low emotional status for a long time period, clinicians could initiate open communication with them in a manner that may have a positive effect. Explaining the interactive relationship between emotion and function and that emotional ups and downs are common during the rehabilitation process could help patients adhere to rehabilitation, achieve emotional improvement and stay at a higher emotional status.r = 0.41; emotion before session vs. function: r = 0.62) correlation (time-collapsed: r = 0.58). Our observation is consistent with those of previous reports [r-values indicate that at least 17% up to 38% of the variation in functional progression can be explained by variation in emotional change. Emotion is difficult to define and/or measure. Overtime, researchers have tested many different approaches with various measurement tools. Emotion can be measured by physiological responses such as autonomic nervous system [Between emotion and function, there was a moderate , the amount of emotional improvement measured on a VAS was 3 mm (51 to 54 mm). Although limited to quantifying pain, previously established minimal differences on VAS were at least 15 mm  or 18 mmWhile function consistently improved, emotional fluctuation was observed during the rehabilitation period. The first emotional improvement (time-collapsed) appeared at the time point corresponding to 24% of the rehabilitation period. Afterwards, patients experienced six emotional deteriorations , which is corresponding to 28% of the entire rehabilitation period C. The emn = 4), which limits generalization of the results. When considering an expected change in the VAS of 16.8 mm with a standard deviation of 19.1 mm (an ES of 0.88) based on our data, the minimum number of subjects need to detect clinically meaningful effect was calculated as ten . Along with the internal and external psychosocial factors above, degree of heterogeneity in terms of gender, family, sport, performance level, previous history of surgery and rehabilitation, characteristics  of the lower-extremity pathology or other medical condition in our patients certainly impact our results. Based on the results of this preliminary analysis, data from a larger number of patients from a narrow population should be analyzed in the future. Since emotion is complex, as stated above, it should be measured and analyzed by a multidimensional approach. There are emotionally mediated contributing factors that can potentially affect clinical function. For example, level of certain hormones  due to emotional changes can influence the healing process and functional outcomes. Therefore, we acknowledge that our assessment using the VAS cannot fully reflect emotional response. The nature of rehabilitative treatment and communication with clinicians in terms of quantity and quality across patients may have varied, which could have affected the study results. According to the chi-square test, specific therapeutic rehabilitative interventions were different. However, maintaining similar proportions of therapeutic interventions in patients with different pathologies is not possible. The sampling frequency in main outcomes was different in that emotion measurements were obtained each session while function was assessed every eight sessions; as such, the average numbers of measurements for emotion and function were 50 and seven, respectively. For functional outcome measures, however, the time course of change should be considered for determining the optimal measurement frequency [Several important limitations and assumptions of our study should be addressed. First, our study has a small sample size (requency .n = 4), which has a limitation for generalizing the results. Future researchers should conduct a study with a larger number of participants from the homogenous clinical population.We observed a moderate linear relationship between emotional changes and functional progression in collegiate student-athletes during the postoperative rehabilitation period. A single session of rehabilitation resulted in slight emotional improvement. After the first emotional improvement, patients experienced six instances of emotional deterioration (37% of the time), while function consistently progressed. The results of our preliminary data have several clinical implications: clinicians should be aware of the relationship between emotion and function and the potential existence of emotional ups and downs throughout the rehabilitation period; psychological consultation focusing on these topics could help patients advance to an emotional improvement and remain at positive emotional status. It should be noted that we only provided preliminary data (


Chronic Chagas cardiomyopathy (CCC) is the most common cause of nonischemic cardiomyopathy in endemic Latin American countries. Immigrants to the United States suffer from this disease, but it is underrecognized. We describe the three hallmark clinical presentations: stroke, ventricular arrhythmias, and heart failure, which should prompt suspicion for CCC. Chagas disease is a chronic systemic parasitic infection caused by the protozoan Trypanosoma cruzi that is transmitted to humans by triatomine insects in endemic areas of Latin America .CCC typically appears 15-30 years after the initial infection, and 20-30% of infected individuals present with manifestations such as heart failure, stroke, and brady- and tachyarrhythmias [In this case series, we describe the experience of a high-volume referral center with different presentations of Chagas cardiomyopathy, highlighting presentations that should raise suspicion for Trypanosoma cruzi infection.A 28-year-old El Salvadorian male was referred to cardiology for palpitations after an electrocardiogram (ECG) showed bifascicular block. A transthoracic echocardiogram (TTE) showed normal biventricular size and function, and a cardiac magnetic resonance imaging (MRI) was suggested because of the ECG abnormalities. This showed a small area of transmural myocardial fibrosis with focal dyskinesis of the true apex, muscular diverticula, and midapical lateral hypokinesis. Because of the myocardial scar, a coronary computed tomography angiography was suggested to evaluate for coronary atherosclerosis, but the patient was lost to follow-up.2 years later, he presented with new right hemiparesis of indeterminate duration. Brain MRI revealed acute ischemic infarct in the left middle cerebral artery territory . ThromboElectrocardiogram (ECG) showed normal sinus rhythm with right bundle branch block (RBBB), left anterior fascicular block (LAFB), and secondary T wave changes . TTE witGiven the patient's country of origin, ECG, and TTE findings, CCC was considered. Commercial Hemagen followed by CDC confirmatory assays were positive for Trypanosoma cruzi infection, which confirmed the diagnosis.Patient was discharged to a rehabilitation facility and continued to follow up with our clinic. Family screening was discussed as an outpatient.A 46-year-old El Salvadorian female with a history of alcohol abuse presented to our hospital with hemodynamically unstable ventricular tachycardia requiring cardioversion.She presented with chest tightness and dizziness of 2-hour duration and was found to be in monomorphic ventricular tachycardia requiring cardioversion. Laboratory investigations showed low potassium and magnesium levels which were repleted. Thyroid function was normal. Troponin I was 0.581 ng/ml (reference range < 0.045 ng/ml). ECG after cardioversion showed normal sinus rhythm with RBBB and LAFB .Coronary angiography showed no coronary artery disease. TTE showed normal biventricular function without regional wall motion abnormalities. Cardiac MRI showed extensive transmural basal lateral wall gadolinium enhancement suggestive of fibrosis . CommercThe patient received a single-chamber implantable cardioverter defibrillator for secondary prevention. Family screening was discussed as an outpatient; the patient was made aware that all children of an infected mother must receive testing to evaluate for congenital transmission.A 43-year-old El Salvadorian male with heart failure from CCC required left ventricular assist device (LVAD) implantation and subsequently orthotopic heart transplantation.The patient presented at the age of 22 with a syncopal event for which a dual-chamber pacemaker was placed. After developing symptoms of heart failure in his early 30's, he was found by TTE to have severe systolic dysfunction with an ejection fraction of 10%. Coronary angiography showed no obstructive coronary artery disease. Nonischemic lab analysis was unrevealing.He underwent right heart catheterization with biopsy which showed multifocal chronic inflammation with fibrosis and myocardial necrosis. Given his country of origin, there was high suspicion of Chagas disease as the precipitating cause of his nonischemic cardiomyopathy. Trypanosoma cruzi serologic assays were obtained and were positive.Despite guideline-directed medical therapy, he required inotropic support and subsequently a HeartWare LVAD was placed. Two years later, the patient received a heart transplant and within 2 weeks of transplant was diagnosed with acute reactivation of Chagas by polymerase chain reaction assay. He responded well to benznidazole therapy.Chagas Cardiomyopathy is the most important clinical presentation of Chagas disease. While only 20-30% of infected patients will develop cardiomyopathy , 3.ECG findings that are highly suggestive of Chagas cardiomyopathy include first-degree atrioventricular block, RBBB, LAFB, bifascicular block, or premature ventricular contractions . NeverthPatients with Chagas cardiomyopathy with preserved or mildly depressed ejection fraction are still at risk for lethal ventricular arrhythmias as well as strokes, and strong consideration should be given to risk to stratify those patients with cardiac imaging , 4.All children of seropositive women and any additional family members of an infected individual should be screened for the disease because of the risk of maternal-fetal transmission and the higher rates of Chagas disease in immediate family members , 6.Most importantly, physicians in the United States who work with Latin American immigrants should be aware that this disease in not uncommon in patients presenting with unexplained cardiac symptoms and/or suspicious electrocardiographic features. Seroprevalence studies have shown high rates of Chagas disease in Latin American immigrants from Chagas-endemic countries to the U.S.: at the Olive View Medical Center in Los Angeles County, 5% of patients with conduction abnormalities , 7.5% ofChagas disease should be strongly suspected in Latin American immigrant patients from endemic countries who present with stroke, ventricular arrhythmias, or heart failure.


Obesity and overweight are common modern health challenges. Caloric intake greater than that needed for energy production results in excess storage of fat in the abdomen. Visceral fat secretes a wide spectrum of adipokines, and increased adiposity is associated with a higher risk of development of metabolic disorders. In addition, adipose tissue secretes extracellular vesicles (EVs) to communicate with peripheral cells and distant organs, and regulate whole-body metabolism. Furthermore, clinical evidence has shown that adipose tissue-derived EVs are present at low levels in the circulation of healthy individuals. In contrast, individuals with metabolic syndrome have significantly higher levels of circulating adipose-derived EVs. The composition of the contents of EVs is dynamic, and closely mirrors individual daily habits and fasting-fed state metabolic characteristics. In this mini-review, we aimed to elucidate the role of adipocyte-derived EVs in regulation of whole-body metabolism under physiological and pathophysiological conditions. Studies have shown that adipose tissue may be a major source of circulating exosomal miRNAs that regulate metabolic homeostasis and directly promote insulin-resistance in other organs. Furthermore, the composition of adipocyte-derived circulating miRNAs in EVs may change prior to development of metabolic disorder. Adipocyte-derived miRNAs in EVs may also induce obesity-related changes such as M1 polarization and inhibition of M2 polarization of macrophages, which may affect the biological behaviors of surrounding tumor cells. Obesity and overweight are serious public health concerns linked to complications such as cardiovascular disease , type 2 Extracellular vesicles are membrane structures released by cells that play a role to exchange components between cells. Based on the current research those were comprehended by transmission and immuno-electron microscopy and by biochemical means, EVs typically are classified into microvesicles and exosomes.Exosomes are 30–150 nm in diameter, generated through inward budding of endosomal membranes which leads to form multivesicular endosomes (MVEs) . The carMultiple reports have implicated EVs as a vehicle transferring miRNAs between cells . StudiesThe prevalence of metabolic syndrome is directly associated with the incidence of being overweight or obese . DiagnosIn healthy individuals, EVs primarily derive from platelets, smaller population derive from monocytes, and vascular endothelial cells . ConnollLarge numbers of non-circulation-derived EVs may indicate a pathological state. Eguchi et al. showed that obese individuals had greater levels of circulating EVs than lean individuals. Moreover, the abundance of perilipin A in circulating EVs of obese individuals was higher than that of lean individuals, which suggested that these EVs might have originated in adipose tissue . HoweverKobayashi et al. analyzed circulating EVs in 203 participants with or without risk factors for metabolic diseases. The results showed that the number of circulating EVs correlated most strongly with risk factors for metabolic diseases. They further isolated circulating EVs and found that perilipin A was correlated with serum triglyceride (TG) levels, asialoglycoprotein receptor 1 (ASGPR1) levels were significantly increased in individuals with metabolic risk factors. In contrast, levels of annexin V and CD9, which are common EV markers, did not differ between individuals with or without metabolic risk factors . PerilipObesity occurs as a result of increased food intake or decreased metabolism. Adipose tissue is critical to systemic energy homeostasis because it participates in the regulation of metabolic organs. Previous studies have described how the functions of adipocyte-derived EVs are integrated and coordinated in accordance with system-wide metabolic demands.Clair et al. found that fasting enhanced the abundance of signaling pathways in adipose tissue secreting EVs compared to the fed stage. For example, proteins involved in Wnt signaling and in the mitochondrial respiratory chain were increased in the fasted state. Adipocytes may receive EVs that contain membrane proteins from endothelial cells and then transport these proteins into adipocyte-derived EVs. Glucagon signaling enhanced endothelial cell uptake and packaging of extracellular cargos into EVs without enhancing endothelial cell EV secretion in mice fed a high-fat diet .In vitro results showed that EV−associated MALAT1, which was significantly increased in adipocytes of obese mice, regulated mTOR signaling through miR−181b and miR−144. Adipocyte-derived EVs from obese mice increased appetite and weight gain in lean mice. In contrast, adipocyte−derived EVs from lean mice attenuated appetite and reduced body weight of obese mice  have been identified in adipocyte-derived EVs , these pmiRNAs in adipocyte-derived EVs may play a more important role in the circulation than proteins such as leptin, adiponectin, or FABP4. Thomou found that the levels of 422 of 653 detectable exosomal miRNAs were altered in the serum of the mice with an adipose-tissue-specific knockout of the miRNA-processing enzyme Dicer (ADicerKO) which exhibits a defect in miRNA processing in adipose tissue, comparing with the wild type. The circulating levels of exosomal miRNAs were rescued following transplantation of brown adipose tissue (BAT) into the ADicerKO mice. In addition, Thomou showed that lipodystrophic patients exhibited drastically reduced levels of exosomal miRNAs in the bloodstream compared to those in controls. However, the numbers of exosomes did not differ between these two groups. This finding suggested that reduced levels of miRNAs were due to altered miRNA processing in adipocytes, and not the number of circulating exosomes . These rin vivo. Lean mice that were administered adipose tissue macrophage (ATM)-derived exosomes with increased levels of miR-155 from wild type obese mice by high-fat feeding were found to exhibit glucose intolerance and insulin resistance  of mice exposed to cold stimulation. In addition, the exposure of 10 people to a 10-day cold acclimation period resulted in decreased levels of miR-92a .Daily dietary habits such as different glycemic load dietaries, and physical activities such as acute or chronic exercise also induce changes in the expression levels of miRNAs in the circulation . Severalin vivo.Hubal et al. evaluated global miRNA profiles of circulating adipocyte-derived exosomes before and 1 year after gastric bypass surgery of obese patients. The results showed that changes in 46 known mature miRNAs correlated with changes in HOMA . This fiObesity is closely related to chronic inflammation, as evidenced by the accumulation of immune cells in adipose tissue. Recent studies found that adipocyte-derived EVs play an important role in macrophage infiltration. For example, exosomes derived from visceral adipose tissue of high fat-fed animals promoted macrophage M1 polarization through activation of NF-κB .Obesity induces accumulation of M1 polarized macrophages in adipose tissue. Studies have shown that M1 polarization of ATMs in obesity is associated with inflammation and insulin resistance in adipose tissue . ExosomeLean adipose tissue contains predominantly M2-polarized macrophages, which play a key role in maintaining homeostasis. However, during the development of obesity, ATMs populated in adipose tissue shift from anti-inflammatory M2 to proinflammatory M1, which results in production of proinflammatory factors that exacerbate insulin resistance . Pan et Adipose tissue is capable of producing and secreting a variety of adipokines and cytokines that regulate metabolic homeostasis. Adiponectin is a representative adipocytokine, which circulates at very high concentrations, constituting approximately 0.01–0.05% of total serum proteins. The secretion of adiponectin in the form of classical or/and exosome pathways by adipocyte is influenced by hormones   and metaOn the other hand, adipocyte-derived EVs could also regulate or protect target cells or organs, responding to circular hormones. Melatonin administration has been shown to increase cellular α−ketoglutarate in adipocytes of obese mice and to increase levels of α−ketoglutarate in exosomes secreted by adipocytes. These exosomes then promote M2 macrophage activation through increased α−ketoglutarate levels in macrophages . MelatonObesity may promote tumor development through enhanced secretion of proinflammatory molecules, altered energy metabolism, secretion of pro-angiogenic factors, and immune cell recruitment and macrophage polarization. Recent studies have shown that adipose-derived EVs influence the biological behavior of tumor cells through transport of circRNAs to hepatocellular carcinoma  and MMP3Adipose tissue is thought to play an important role in regulation of endocrine function, and can modulate metabolism in other organs. Recent studies suggest that adipose cells regulate metabolism in liver, skeletal muscle, even tumors through release of exosomes into the circulatory system. Many studies have shown that the miRNA in exosomes changes significantly during metabolic disease development, may serve as ideal markers for disease diagnosis. The roles of miRNAs contained in adipocyte-derived EVs in regulating metabolism and the biological behavior of tumors, and the variation under metabolic stress are listed in However, exosomes in the circulatory system may originate from many cell types. Adipose tissue is supposed a major source of circulating exosomal miRNAs, even to enhance secreting amount of EVs under metabolic stress. Therefore, monitoring adipocyte-derived miRNAs in circulatory exosomes may represent a novel approach to prevention and treatment of obesity-related diseases. Thomou have shown levels of miRNAs in circulating adipocyte-derived EVs change prior to the onset of metabolic disorder . Global Some hormone drugs, such as melatonin, alter the levels of specific proteins or miRNAs in fat cells. These changes result in changes in the levels of protein or miRNA in exosomes secreted by fat cells. Titration of hormone drug dosing may benefit from monitoring the levels of target proteins or miRNAs in circulating EVs. This may result in better disease control and reduced adverse reactions.YZ contributed to drafting the review. CT contributed to designing and composing the review. Both authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.