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1.
mSystems ; : e0113621, 2021 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-34726486

RESUMO

Environmental monitoring in public spaces can be used to identify surfaces contaminated by persons with coronavirus disease 2019 (COVID-19) and inform appropriate infection mitigation responses. Research groups have reported detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on surfaces days or weeks after the virus has been deposited, making it difficult to estimate when an infected individual may have shed virus onto a SARS-CoV-2-positive surface, which in turn complicates the process of establishing effective quarantine measures. In this study, we determined that reverse transcription-quantitative PCR (RT-qPCR) detection of viral RNA from heat-inactivated particles experiences minimal decay over 7 days of monitoring on eight out of nine surfaces tested. The properties of the studied surfaces result in RT-qPCR signatures that can be segregated into two material categories, rough and smooth, where smooth surfaces have a lower limit of detection. RT-qPCR signal intensity (average quantification cycle [Cq]) can be correlated with surface viral load using only one linear regression model per material category. The same experiment was performed with untreated viral particles on one surface from each category, with essentially identical results. The stability of RT-qPCR viral signal demonstrates the need to clean monitored surfaces after sampling to establish temporal resolution. Additionally, these findings can be used to minimize the number of materials and time points tested and allow for the use of heat-inactivated viral particles when optimizing environmental monitoring methods. IMPORTANCE Environmental monitoring is an important tool for public health surveillance, particularly in settings with low rates of diagnostic testing. Time between sampling public environments, such as hospitals or schools, and notifying stakeholders of the results should be minimal, allowing decisions to be made toward containing outbreaks of coronavirus disease 2019 (COVID-19). The Safer At School Early Alert program (SASEA) (https://saseasystem.org/), a large-scale environmental monitoring effort in elementary school and child care settings, has processed >13,000 surface samples for SARS-CoV-2, detecting viral signals from 574 samples. However, consecutive detection events necessitated the present study to establish appropriate response practices around persistent viral signals on classroom surfaces. Other research groups and clinical labs developing environmental monitoring methods may need to establish their own correlation between RT-qPCR results and viral load, but this work provides evidence justifying simplified experimental designs, like reduced testing materials and the use of heat-inactivated viral particles.

2.
Life Sci Alliance ; 4(12)2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34663679

RESUMO

Extracellular vesicles (EVs) mediate intercellular signaling by transferring their cargo to recipient cells, but the functional consequences of signaling are not fully appreciated. RBC-derived EVs are abundant in circulation and have been implicated in regulating immune responses. Here, we use a transgenic mouse model for fluorescence-based mapping of RBC-EV recipient cells to assess the role of this intercellular signaling mechanism in heart disease. Using fluorescent-based mapping, we detected an increase in RBC-EV-targeted cardiomyocytes in a murine model of ischemic heart failure. Single cell nuclear RNA sequencing of the heart revealed a complex landscape of cardiac cells targeted by RBC-EVs, with enrichment of genes implicated in cell proliferation and stress signaling pathways compared with non-targeted cells. Correspondingly, cardiomyocytes targeted by RBC-EVs more frequently express cellular markers of DNA synthesis, suggesting the functional significance of EV-mediated signaling. In conclusion, our mouse model for mapping of EV-recipient cells reveals a complex cellular network of RBC-EV-mediated intercellular communication in ischemic heart failure and suggests a functional role for this mode of intercellular signaling.

3.
medRxiv ; 2021 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-34704096

RESUMO

Schools are high-risk settings for SARS-CoV-2 transmission, but necessary for children’s educational and social-emotional wellbeing. While wastewater monitoring has been implemented to mitigate outbreak risk in universities and residential settings, its effectiveness in community K-12 sites is unknown. We implemented a wastewater and surface monitoring system to detect SARS-CoV-2 in nine elementary schools in San Diego County. Ninety-three percent of identified cases were associated with either a positive wastewater or surface sample; 67% were associated with a positive wastewater sample, and 40% were associated with a positive surface sample. The techniques we utilized allowed for near-complete genomic sequencing of wastewater and surface samples. Passive environmental surveillance can complement approaches that require individual consent, particularly in communities with limited access and/or high rates of testing hesitancy. One sentence summary: Passive wastewater and surface environmental surveillance can identify up to 93% of on-campus COVID-19 cases in public elementary schools; positive samples can be sequenced to monitor for variants of concerns with neighborhood level resolution.

5.
Cell ; 184(19): 4939-4952.e15, 2021 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-34508652

RESUMO

The emergence of the COVID-19 epidemic in the United States (U.S.) went largely undetected due to inadequate testing. New Orleans experienced one of the earliest and fastest accelerating outbreaks, coinciding with Mardi Gras. To gain insight into the emergence of SARS-CoV-2 in the U.S. and how large-scale events accelerate transmission, we sequenced SARS-CoV-2 genomes during the first wave of the COVID-19 epidemic in Louisiana. We show that SARS-CoV-2 in Louisiana had limited diversity compared to other U.S. states and that one introduction of SARS-CoV-2 led to almost all of the early transmission in Louisiana. By analyzing mobility and genomic data, we show that SARS-CoV-2 was already present in New Orleans before Mardi Gras, and the festival dramatically accelerated transmission. Our study provides an understanding of how superspreading during large-scale events played a key role during the early outbreak in the U.S. and can greatly accelerate epidemics.


Assuntos
COVID-19/epidemiologia , Epidemias , SARS-CoV-2/fisiologia , COVID-19/transmissão , Bases de Dados como Assunto , Surtos de Doenças , Humanos , Louisiana/epidemiologia , Filogenia , Fatores de Risco , SARS-CoV-2/classificação , Texas , Viagem , Estados Unidos/epidemiologia
6.
Front Cell Dev Biol ; 9: 702046, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34540826

RESUMO

During pregnancy, conceptus-derived extravillous trophoblast (EVT) invades the endomyometrium, anchors the placenta to the maternal uterus, and remodels the spiral arteries in order to establish maternal blood supply to the fetoplacental unit. Recent reports have described early gestation EVT as polyploid and senescent. Here, we extend these reports by performing comprehensive profiling of both the genomic organization and transcriptome of first trimester and term EVT. We define pathways and gene regulatory networks involved in both initial differentiation and maturation of this important trophoblast lineage at the maternal-fetal interface. Our results suggest that like first trimester EVT, term EVT undergoes senescence and endoreduplication, is primarily tetraploid, and lacks high rates of copy number variations. Additionally, we have highlighted senescence and polyploidy-related genes, pathways, networks, and transcription factors that appeared to be important in normal EVT differentiation and maturation and validated a key role for the unfolded protein response in this context.

7.
Data Brief ; 38: 107278, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34401413

RESUMO

We present supplementary data for the published article, "Hitting the diagnostic sweet spot: Point-of-care SARS-CoV-2 salivary antigen testing with an off-the-shelf glucometer" [1]. The assay described is designed to be performed at home or in a clinic without expensive instrumentation or professional training. SARS-CoV-2 is detected by an aptamer-based assay that targets the Nucleocapsid (N) or Spike (S) antigens. Binding of the N or S protein to their respective aptamer results in the competitive release of a complementary antisense-invertase enzyme complex. The released enzyme then catalyzes the conversion of sucrose to glucose that is measured by an off-the-shelf glucometer. The data presented here describe the optimization of the assay parameters and their contribution to developing this aptamer-based assay to detect SARS-CoV-2. The assay performance was checked in a standard buffer, contrived samples, and patient samples validated with well-established scientific methods. The resulting dataset can be used to further develop glucometer-based assays for diagnosing other communicable and non-communicable diseases.

8.
bioRxiv ; 2021 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-34312621

RESUMO

Environmental monitoring in public spaces can be used to identify surfaces contaminated by persons with COVID-19 and inform appropriate infection mitigation responses. Research groups have reported detection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) on surfaces days or weeks after the virus has been deposited, making it difficult to estimate when an infected individual may have shed virus onto a SARS-CoV-2 positive surface, which in turn complicates the process of establishing effective quarantine measures. In this study, we determined that reverse transcription-quantitative polymerase chain reaction (RT-qPCR) detection of viral RNA from heat-inactivated particles experiences minimal decay over seven days of monitoring on eight out of nine surfaces tested. The properties of the studied surfaces result in RT-qPCR signatures that can be segregated into two material categories, rough and smooth, where smooth surfaces have a lower limit of detection. RT-qPCR signal intensity (average quantification cycle ( Cq )) can be correlated to surface viral load using only one linear regression model per material category. The same experiment was performed with infectious viral particles on one surface from each category, with essentially identical results. The stability of RT-qPCR viral signal demonstrates the need to clean monitored surfaces after sampling to establish temporal resolution. Additionally, these findings can be used to minimize the number of materials and time points tested and allow for the use of heat-inactivated viral particles when optimizing environmental monitoring methods. Importance: Environmental monitoring is an important tool for public health surveillance, particularly in settings with low rates of diagnostic testing. Time between sampling public environments, such as hospitals or schools, and notifying stakeholders of the results should be minimal, allowing decisions to be made towards containing outbreaks of coronavirus disease 2019 (COVID-19). The Safer At School Early Alert program (SASEA) [1], a large-scale environmental monitoring effort in elementary school and child care settings, has processed > 13,000 surface samples for SARS-CoV-2, detecting viral signals from 574 samples. However, consecutive detection events necessitated the present study to establish appropriate response practices around persistent viral signals on classroom surfaces. Other research groups and clinical labs developing environmental monitoring methods may need to establish their own correlation between RT - qPCR results and viral load, but this work provides evidence justifying simplified experimental designs, like reduced testing materials and the use of heat-inactivated viral particles.

9.
Cell ; 184(10): 2587-2594.e7, 2021 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-33861950

RESUMO

The highly transmissible B.1.1.7 variant of SARS-CoV-2, first identified in the United Kingdom, has gained a foothold across the world. Using S gene target failure (SGTF) and SARS-CoV-2 genomic sequencing, we investigated the prevalence and dynamics of this variant in the United States (US), tracking it back to its early emergence. We found that, while the fraction of B.1.1.7 varied by state, the variant increased at a logistic rate with a roughly weekly doubling rate and an increased transmission of 40%-50%. We revealed several independent introductions of B.1.1.7 into the US as early as late November 2020, with community transmission spreading it to most states within months. We show that the US is on a similar trajectory as other countries where B.1.1.7 became dominant, requiring immediate and decisive action to minimize COVID-19 morbidity and mortality.


Assuntos
COVID-19 , Modelos Biológicos , SARS-CoV-2 , COVID-19/genética , COVID-19/mortalidade , COVID-19/transmissão , Feminino , Humanos , Masculino , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidade , Estados Unidos/epidemiologia
10.
Sci Rep ; 11(1): 5877, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33723311

RESUMO

Preeclampsia (PE) is a pregnancy-specific hypertensive disorder, affecting up to 10% of pregnancies worldwide. The primary etiology is considered to be abnormal development and function of placental cells called trophoblasts. We previously developed a two-step protocol for differentiation of human pluripotent stem cells, first into cytotrophoblast (CTB) progenitor-like cells, and then into both syncytiotrophoblast (STB)- and extravillous trophoblast (EVT)-like cells, and showed that it can model both normal and abnormal trophoblast differentiation. We have now applied this protocol to induced pluripotent stem cells (iPSC) derived from placentas of pregnancies with or without PE. While there were no differences in CTB induction or EVT formation, PE-iPSC-derived trophoblast showed a defect in syncytialization, as well as a blunted response to hypoxia. RNAseq analysis showed defects in STB formation and response to hypoxia; however, DNA methylation changes were minimal, corresponding only to changes in response to hypoxia. Overall, PE-iPSC recapitulated multiple defects associated with placental dysfunction, including a lack of response to decreased oxygen tension. This emphasizes the importance of the maternal microenvironment in normal placentation, and highlights potential pathways that can be targeted for diagnosis or therapy, while absence of marked DNA methylation changes suggests that other regulatory mechanisms mediate these alterations.

11.
Biosens Bioelectron ; 180: 113111, 2021 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-33743492

RESUMO

Significant barriers to the diagnosis of latent and acute SARS-CoV-2 infection continue to hamper population-based screening efforts required to contain the COVID-19 pandemic in the absence of widely available antiviral therapeutics or vaccines. We report an aptamer-based SARS-CoV-2 salivary antigen assay employing only low-cost reagents ($3.20/test) and an off-the-shelf glucometer. The test was engineered around a glucometer as it is quantitative, easy to use, and the most prevalent piece of diagnostic equipment globally, making the test highly scalable with an infrastructure that is already in place. Furthermore, many glucometers connect to smartphones, providing an opportunity to integrate with contact tracing apps, medical providers, and electronic health records. In clinical testing, the developed assay detected SARS-CoV-2 infection in patient saliva across a range of viral loads - as benchmarked by RT-qPCR - within 1 h, with 100% sensitivity (positive percent agreement) and distinguished infected specimens from off-target antigens in uninfected controls with 100% specificity (negative percent agreement). We propose that this approach provides an inexpensive, rapid, and accurate diagnostic for distributed screening of SARS-CoV-2 infection at scale.


Assuntos
Antígenos Virais/análise , Técnicas Biossensoriais/métodos , Teste Sorológico para COVID-19/métodos , COVID-19/diagnóstico , Testes Imediatos , SARS-CoV-2/imunologia , Saliva/virologia , Adulto , Teste para COVID-19 , Proteínas do Nucleocapsídeo de Coronavírus/análise , Feminino , Humanos , Masculino , Fosfoproteínas/análise , SARS-CoV-2/isolamento & purificação , Técnica de Seleção de Aptâmeros , Sensibilidade e Especificidade , Glicoproteína da Espícula de Coronavírus/análise
12.
medRxiv ; 2021 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-33564780

RESUMO

As of January of 2021, the highly transmissible B.1.1.7 variant of SARS-CoV-2, which was first identified in the United Kingdom (U.K.), has gained a strong foothold across the world. Because of the sudden and rapid rise of B.1.1.7, we investigated the prevalence and growth dynamics of this variant in the United States (U.S.), tracking it back to its early emergence and onward local transmission. We found that the RT-qPCR testing anomaly of S gene target failure (SGTF), first observed in the U.K., was a reliable proxy for B.1.1.7 detection. We sequenced 212 B.1.1.7 SARS-CoV-2 genomes collected from testing facilities in the U.S. from December 2020 to January 2021. We found that while the fraction of B.1.1.7 among SGTF samples varied by state, detection of the variant increased at a logistic rate similar to those observed elsewhere, with a doubling rate of a little over a week and an increased transmission rate of 35-45%. By performing time-aware Bayesian phylodynamic analyses, we revealed several independent introductions of B.1.1.7 into the U.S. as early as late November 2020, with onward community transmission enabling the variant to spread to at least 30 states as of January 2021. Our study shows that the U.S. is on a similar trajectory as other countries where B.1.1.7 rapidly became the dominant SARS-CoV-2 variant, requiring immediate and decisive action to minimize COVID-19 morbidity and mortality.

13.
medRxiv ; 2021 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-33564781

RESUMO

The emergence of the early COVID-19 epidemic in the United States (U.S.) went largely undetected, due to a lack of adequate testing and mitigation efforts. The city of New Orleans, Louisiana experienced one of the earliest and fastest accelerating outbreaks, coinciding with the annual Mardi Gras festival, which went ahead without precautions. To gain insight into the emergence of SARS-CoV-2 in the U.S. and how large, crowded events may have accelerated early transmission, we sequenced SARS-CoV-2 genomes during the first wave of the COVID-19 epidemic in Louisiana. We show that SARS-CoV-2 in Louisiana initially had limited sequence diversity compared to other U.S. states, and that one successful introduction of SARS-CoV-2 led to almost all of the early SARS-CoV-2 transmission in Louisiana. By analyzing mobility and genomic data, we show that SARS-CoV-2 was already present in New Orleans before Mardi Gras and that the festival dramatically accelerated transmission, eventually leading to secondary localized COVID-19 epidemics throughout the Southern U.S.. Our study provides an understanding of how superspreading during large-scale events played a key role during the early outbreak in the U.S. and can greatly accelerate COVID-19 epidemics on a local and regional scale.

14.
Placenta ; 104: 267-276, 2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-33472134

RESUMO

INTRODUCTION: High-altitude (>2500 m) residence augments the risk of intrauterine growth restriction (IUGR) and preeclampsia likely due, in part, to uteroplacental hypoperfusion. Previous genomic and transcriptomic studies in humans and functional studies in mice and humans suggest a role for AMP-activated protein kinase (AMPK) pathway in protecting against hypoxia-associated IUGR. AMPK is a metabolic sensor activated by hypoxia that is ubiquitously expressed in vascular beds and placenta. METHODS: We measured gene expression and protein levels of AMPK and its upstream regulators and downstream targets in human placentas from high (>2500 m) vs. moderate (~1700 m) and low (~100 m) altitude. RESULTS: We found that phosphorylated AMPK protein levels and its downstream target TSC2 were increased in placentas from high and moderate vs. low altitude, whereas the phosphorylated form of the downstream target translation repressor protein 4E-BP1 was increased in high compared to moderate as well as low altitude placentas. Mean birth weights progressively fell with increasing altitude but no infants, by study design, were clinically growth-restricted. Gene expression analysis showed moderate increases in PRKAG2, encoding the AMPK γ2 subunit, and mechanistic target of rapamycin, MTOR, expression. DISCUSSION: These results highlight a differential regulation of placental AMPK pathway activation in women residing at low, moderate or high altitude during pregnancy, suggesting AMPK may be serving as a metabolic regulator for integrating hypoxic stimuli with placental function.

15.
Am J Obstet Gynecol MFM ; 2(3): 100140, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-33345877

RESUMO

BACKGROUND: Preterm birth remains a common and devastating complication of pregnancy. There remains a need for effective and accurate screening methods for preterm birth. Using a proteomic approach, we previously discovered and validated (Proteomic Assessment of Preterm Risk study, NCT01371019) a preterm birth predictor comprising a ratio of insulin-like growth factor-binding protein 4 to sex hormone-binding globulin. OBJECTIVE: To determine the performance of the ratio of insulin-like growth factor-binding protein 4 to sex hormone-binding globulin to predict both spontaneous and medically indicated very preterm births, in an independent cohort distinct from the one in which it was developed. STUDY DESIGN: This was a prospective observational study (Multicenter Assessment of a Spontaneous Preterm Birth Risk Predictor, NCT02787213) at 18 sites in the United States. Women had blood drawn at 170/7 to 216/7 weeks' gestation. For confirmation, we planned to analyze a randomly selected subgroup of women having blood drawn between 191/7 and 206/7 weeks' gestation, with the results of the remaining study participants blinded for future validation studies. Serum from participants was analyzed by mass spectrometry. Neonatal morbidity and mortality were analyzed using a composite score by a method from the PREGNANT trial (NCT00615550, Hassan et al). Scores of 0-3 reflect increasing numbers of morbidities or length of neonatal intensive care unit stay, and 4 represents perinatal mortality. RESULTS: A total of 5011 women were enrolled, with 847 included in this planned substudy analysis. There were 9 preterm birth cases at <320/7 weeks' gestation and 838 noncases at ≥320/7 weeks' gestation; 21 of 847 infants had neonatal composite morbidity and mortality index scores of ≥3, and 4 of 21 had a score of 4. The ratio of insulin-like growth factor-binding protein 4 to sex hormone-binding globulin ratio was substantially higher in both preterm births at <320/7 weeks' gestation and there were more severe neonatal outcomes. The ratio of insulin-like growth factor-binding protein 4 to sex hormone-binding globulin ratio was significantly predictive of birth at <320/7 weeks' gestation (area under the receiver operating characteristic curve, 0.71; 95% confidence interval, 0.55-0.87; P=.016). Stratification by body mass index, optimized in the previous validation study (22

Assuntos
Nascimento Prematuro , Estudos de Coortes , Feminino , Idade Gestacional , Humanos , Recém-Nascido , Gravidez , Estudos Prospectivos , Proteômica , Estados Unidos
16.
N Engl J Med ; 383(18): 1746-1756, 2020 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-33027564

RESUMO

BACKGROUND: The cause of most fetal anomalies is not determined prenatally. Exome sequencing has transformed genetic diagnosis after birth, but its usefulness for prenatal diagnosis is still emerging. Nonimmune hydrops fetalis (NIHF), a fetal abnormality that is often lethal, has numerous genetic causes; the extent to which exome sequencing can aid in its diagnosis is unclear. METHODS: We evaluated a series of 127 consecutive unexplained cases of NIHF that were defined by the presence of fetal ascites, pleural or pericardial effusions, skin edema, cystic hygroma, increased nuchal translucency, or a combination of these conditions. The primary outcome was the diagnostic yield of exome sequencing for detecting genetic variants that were classified as either pathogenic or likely pathogenic according to the criteria of the American College of Medical Genetics and Genomics. Secondary outcomes were the percentage of cases associated with specific genetic disorders and the proportion of variants that were inherited. RESULTS: In 37 of the 127 cases (29%), we identified diagnostic genetic variants, including those for disorders affecting the RAS-MAPK cell-signaling pathway (known as RASopathies) (30% of the genetic diagnoses); inborn errors of metabolism and musculoskeletal disorders (11% each); lymphatic, neurodevelopmental, cardiovascular, and hematologic disorders (8% each); and others. Prognoses ranged from a relatively mild outcome to death during the perinatal period. Overall, 68% of the cases (25 of 37) with diagnostic variants were autosomal dominant (of which 12% were inherited and 88% were de novo), 27% (10 of 37) were autosomal recessive (of which 95% were inherited and 5% were de novo), 1 was inherited X-linked recessive, and 1 was of uncertain inheritance. We identified potentially diagnostic variants in an additional 12 cases. CONCLUSIONS: In this large case series of 127 fetuses with unexplained NIHF, we identified a diagnostic genetic variant in approximately one third of the cases. (Funded by the UCSF Center for Maternal-Fetal Precision Medicine and others; ClinicalTrials.gov number, NCT03412760.).


Assuntos
Variação Genética , Hidropisia Fetal/diagnóstico , Hidropisia Fetal/genética , Diagnóstico Pré-Natal , Sequenciamento Completo do Exoma , Feminino , Humanos , Gravidez , Prognóstico
17.
Cell Rep Med ; 1(2)2020 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-32864636

RESUMO

Development of effective prevention and treatment strategies for pre-eclampsia is limited by the lack of accurate methods for identification of at-risk pregnancies. We performed small RNA sequencing (RNA-seq) of maternal serum extracellular RNAs (exRNAs) to discover and verify microRNAs (miRNAs) differentially expressed in patients who later developed pre-eclampsia. Sera collected from 73 pre-eclampsia cases and 139 controls between 17 and 28 weeks gestational age (GA), divided into separate discovery and verification cohorts, are analyzed by small RNA-seq. Discovery and verification of univariate and bivariate miRNA biomarkers reveal that bivariate biomarkers verify at a markedly higher rate than univariate biomarkers. The majority of verified biomarkers contain miR-155-5p, which has been reported to mediate the pre-eclampsia-associated repression of endothelial nitric oxide synthase (eNOS) by tumor necrosis factor alpha (TNF-α). Deconvolution analysis reveals that several verified miRNA biomarkers come from the placenta and are likely carried by placenta-specific extracellular vesicles.

18.
iScience ; 23(6): 101182, 2020 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-32512385

RESUMO

The recent discovery of extracellular RNAs in blood, including RNAs in extracellular vesicles (EVs), combined with low-input RNA-sequencing advances have enabled scientists to investigate their role in human disease. To date, most studies have been focusing on small RNAs, and methodologies to optimize long RNAs measurement are lacking. We used plasma RNA to assess the performance of six long RNA sequencing methods, at two different sites, and we report their differences in reads (%) mapped to the genome/transcriptome, number of genes detected, long RNA transcript diversity, and reproducibility. Using the best performing method, we further compare the profile of long RNAs in the EV- and no-EV-enriched RNA plasma compartments. These results provide insights on the performance and reproducibility of commercially available kits in assessing the landscape of long RNAs in human plasma and different extracellular RNA carriers that may be exploited for biomarker discovery.

20.
Nat Rev Mol Cell Biol ; 21(10): 585-606, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32457507

RESUMO

The term 'extracellular vesicles' refers to a heterogeneous population of vesicular bodies of cellular origin that derive either from the endosomal compartment (exosomes) or as a result of shedding from the plasma membrane (microvesicles, oncosomes and apoptotic bodies). Extracellular vesicles carry a variety of cargo, including RNAs, proteins, lipids and DNA, which can be taken up by other cells, both in the direct vicinity of the source cell and at distant sites in the body via biofluids, and elicit a variety of phenotypic responses. Owing to their unique biology and roles in cell-cell communication, extracellular vesicles have attracted strong interest, which is further enhanced by their potential clinical utility. Because extracellular vesicles derive their cargo from the contents of the cells that produce them, they are attractive sources of biomarkers for a variety of diseases. Furthermore, studies demonstrating phenotypic effects of specific extracellular vesicle-associated cargo on target cells have stoked interest in extracellular vesicles as therapeutic vehicles. There is particularly strong evidence that the RNA cargo of extracellular vesicles can alter recipient cell gene expression and function. During the past decade, extracellular vesicles and their RNA cargo have become better defined, but many aspects of extracellular vesicle biology remain to be elucidated. These include selective cargo loading resulting in substantial differences between the composition of extracellular vesicles and source cells; heterogeneity in extracellular vesicle size and composition; and undefined mechanisms for the uptake of extracellular vesicles into recipient cells and the fates of their cargo. Further progress in unravelling the basic mechanisms of extracellular vesicle biogenesis, transport, and cargo delivery and function is needed for successful clinical implementation. This Review focuses on the current state of knowledge pertaining to packaging, transport and function of RNAs in extracellular vesicles and outlines the progress made thus far towards their clinical applications.


Assuntos
Vesículas Extracelulares/metabolismo , Mamíferos/metabolismo , RNA/metabolismo , Animais , Transporte Biológico/fisiologia , Comunicação Celular/fisiologia , Humanos
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