Characterization of Serum Extracellular Vesicles with Human Age
血清细胞外囊泡与人类年龄的表征
基本信息
- 批准号:10688899
- 负责人:
- 金额:$ 57.55万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:
- 资助国家:美国
- 起止时间:至
- 项目状态:未结题
- 来源:
- 关键词:AddressAffectAfrican American populationAgeAgingAlzheimer&aposs DiseaseAngiogenic FactorAntigensB-Cell ActivationB-LymphocytesBiologicalBiological AgingBiological AssayBiological MarkersBloodBlood CirculationBody mass indexCD14 geneCancer PatientCardiovascular DiseasesCell AgingCell SurvivalCell physiologyCellsChronic DiseaseCommunicationCommunitiesComplications of Diabetes MellitusCultured CellsDataDevelopmentDiabetes MellitusDiagnosisDiagnosticDiseaseElderlyEncapsulatedEndothelial CellsEpidemiologyErythrocytesFree RadicalsFunctional disorderFutureGene ExpressionHumanImmuneImmune responseImmunofluorescence ImmunologicIn VitroIncubatedIndividualInflammationInflammatoryInsulin ResistanceLaboratoriesLeukocytesLipid BindingLipidsLongevityLongitudinal StudiesLongitudinal cohortMalignant NeoplasmsMeasuresMessenger RNAMetabolic DiseasesMethodsMicroRNAsMinorityMitochondriaMitochondrial DNAModelingMolecularMorbidity - disease rateMothersMucin 1 proteinMutationNeighborhoodsNeurodegenerative DisordersNon-Insulin-Dependent Diabetes MellitusNucleic AcidsObesityOrganOxidative StressPaperParticipantPathologicPathway interactionsPeripheral Vascular DiseasesPhenotypePhysiologicalPlasmaPlayPopulationPopulation SciencesPrecipitationPrediabetes syndromePreparationProteinsPublicationsRNARaceReportingResearchRespirationRoleSamplingSerumSignal TransductionSignaling ProteinSmoking StatusSystemTechnologyTestingTherapeuticTimeTranslatingUntranslated RNAVEGFA geneVesicleVisitWorkage groupage relatedagedbasecell motilitycohortcytokinedesigndiabeticdiagnostic toolexosomeextracellular vesiclesgenetic informationhealth disparityhealthy agingimmune functionin uteroinsulin signalingintercellular communicationmicrovesiclesmiddle agemigrationmonocytemortalitynanoparticlenormal agingoxidative damagepopulation basedsextheoriesuptake
项目摘要
We characterized EV attributes across selected age groups of participants from the population based, longitudinal study, Healthy Aging in Neighborhoods of Diversity across the Life Span (HANDLS) in LEPS. We analyzed circulating plasma EVs from the HANDLS study using both a cross-sectional and longitudinal approach to address age-related changes in community-dwelling individuals. This sub-cohort consisted of 30 young individuals, 30 middle-aged individuals, and 14 old individuals, who had contributed plasma at two different time points (Visit 1 and Visit 2) approximately 5 years apart. The participants in each age group were matched by race and sex. We isolated EVs from plasma using a precipitation method and analyzed size and concentration using Nanoparticle Tracking Analysis and linear mixed-model regression. We found that EV concentration decreases with advancing age. We also analyzed whether EV concentration was related to demographic and anthropometric measures. BMI and smoking status were significantly associated with EV concentration only at visit 2. There were no significant changes in EV concentration with sex or race. The major finding that EV concentration decreases with advancing age was further explored by examining uptake of EVs by immune cells. EVs from older individuals were more readily internalized by B cells and increased MHC-II expression on monocytes compared with EVs from younger individuals, indicating that the decreased concentration of EVs with age may be due in part to increased internalization. EVs activated both monocytes and B cells, and activation of B cells by LPS enhanced EV internalization. We found a relative stability of EV concentration and protein amount in individual subjects over time. Our data provide information towards establishing a profile of EVs with human age, which will further aid in the development of EV-based diagnostics for aging and age-related diseases. We found that several immune-related antigens like MUCIN16, MUC1, NY-ESO, CD14 and PDL increase with age. The reduction in EV concentration may be a consequence of aging-related phenotypic changes like cellular senescence or part of altered intercellular signaling; both are important hallmarks of aging. EVs may be part of the aging mechanism and may change because of aging-related mechanisms and thus serve as biological aging indicators. While the current study shows that EVs change with age, further research is required to clarify their role in aging.
As a follow-on study we examine the relationship between EVs and the presence of diabetes and pre-diabetes. To address this, we have designed a longitudinal sub-cohort of HANDLS that consists of 58 participants who have donated plasma at two time points approximately 5 years apart. Of these 58 participants, 19 were euglycemic at both time points, 19 participants were euglycemic at time point 1 and developed type 2 diabetes mellitus by the second time point and 20 were pre-diabetic at time point 1 and diagnosed with diabetes mellitus at time point 2. Individuals in this sub-cohort have been matched on obesity status. we designed cross-sectional and longitudinal cohorts of euglycemic, pre-diabetic and diabetic participants. In our recent publication in Diabetes (Freeman DW, Noren Hooten N, Eitan E et al 2018) diabetic individuals had significantly higher levels of EVs in their circulation than euglycemic controls. Using a cell-specific EV assay, we identified that erythrocyte-derived EVs are higher with diabetes. We found that insulin resistance increases EV secretion. Furthermore, the levels insulin signaling proteins were altered in EVs from individuals with high levels of insulin resistance and -cell dysfunction. Moreover, EVs from diabetic individuals were preferentially internalized by circulating leukocytes. Cytokine levels in the media and in EVs were higher from monocytes incubated with diabetic EVs. Microarray of these leukocytes revealed altered gene expression pathways related to cell survival, oxidative stress and immune function. Collectively, these results suggest that insulin resistance increases the secretion of EVs, which are preferential internalized by leukocytes and alters leukocyte function. Future work will focus on the influence of race and mortality on EV cargo and the biologic effects of EV cargo on cellular function in the setting of chronic disease and life span. Inflammation-related atherosclerotic peripheral vascular disease is a major end organ complication of diabetes mellitus that results in devastating morbidity and mortality. We examined EV protein cargo from diabetic individuals and whether these EVs cause functional changes in endothelial cells. We quantified inflammatory protein levels in plasma-derived EVs from a longitudinal cohort of euglycemic and diabetic individuals and used in vitro endothelial cell biological assays to assess the functional effects of these EVs with samples from a cross-sectional cohort. We found several significant associations between EV inflammatory protein levels and diabetes status. The angiogenic factor, vascular endothelial growth factor A (VEGF-A), was associated with diabetes status in our longitudinal cohort. Those with diabetes mellitus had higher EV VEGF-A levels compared to euglycemic individuals. Additionally, EV levels of VEGF-A were significantly associated with homeostatic model assessment of insulin resistance (HOMA-IR) and -cell function (HOMA-B). To test whether EVs with different inflammatory cargo can demonstrate different effects on endothelial cells, we performed cell migration and immunofluorescence assays. We observed that EVs from diabetic individuals increased cell lamellipodia formation and migration when compared to EVs from euglycemic individuals. Higher levels of inflammatory proteins were found in EVs from diabetic individuals. Our data implicate EVs as playing important roles in peripheral vascular disease that occur in individuals with diabetes mellitus and suggest that EVs may serve as an informative diagnostic tool for the disease.
We also investigated the mitochondrial free radical theory of aging which suggests that accumulating oxidative damage to mitochondria and mitochondrial DNA (mtDNA) plays a central role in aging. Circulating cell-free mtDNA (ccf-mtDNA) isolated from blood may be a biomarker of disease. Extracellular vesicles (EVs) are small (30-400 nm), lipid-bound vesicles capable of shuttling proteins, nucleic acids, and lipids as part of intercellular communication systems. Here, we report that a portion of ccf-mtDNA in plasma is encapsulated in EVs. To address whether EV mtDNA levels change with human age, we analyzed mtDNA in EVs from individuals aged 30-64 years cross-sectionally and longitudinally. EV mtDNA levels decreased with age. Furthermore, the maximal mitochondrial respiration of cultured cells was differentially affected by EVs from old and young donors. Our results suggest that plasma mtDNA is present in EVs, that the level of EV-derived mtDNA is associated with age, and that EVs affect mitochondrial energetics in an EV age-dependent manner.
我们对 LEPS 中基于人口的纵向研究“跨生命周期多样性社区健康老龄化”(HANDLS) 中选定的参与者年龄组的 EV 属性进行了描述。我们使用横断面和纵向方法分析了 HANDLS 研究中的循环血浆 EV,以解决社区居住个体与年龄相关的变化。该子队列由 30 名年轻人、30 名中年人和 14 名老年人组成,他们在相隔约 5 年的两个不同时间点(访视 1 和访视 2)贡献了血浆。每个年龄组的参与者都按种族和性别进行匹配。 我们使用沉淀法从血浆中分离出 EV,并使用纳米颗粒跟踪分析和线性混合模型回归分析了尺寸和浓度。我们发现 EV 浓度随着年龄的增长而降低。我们还分析了 EV 浓度是否与人口和人体测量指标相关。仅在第 2 次访视时,BMI 和吸烟状况与 EV 浓度显着相关。EV 浓度随性别或种族没有显着变化。通过检查免疫细胞对 EV 的摄取,进一步探讨了 EV 浓度随着年龄增长而降低的主要发现。与年轻个体的 EV 相比,老年个体的 EV 更容易被 B 细胞内化,并且单核细胞上的 MHC-II 表达增加,表明 EV 浓度随着年龄的增长而降低,部分原因可能是内化增加。 EV 激活单核细胞和 B 细胞,LPS 激活 B 细胞可增强 EV 内化。我们发现随着时间的推移,个体受试者的 EV 浓度和蛋白质含量相对稳定。我们的数据为建立电动汽车与人类年龄的关系提供了信息,这将进一步有助于开发基于电动汽车的衰老和年龄相关疾病诊断方法。我们发现一些免疫相关抗原,如 MUCIN16、MUC1、NY-ESO、CD14 和 PDL 随着年龄的增长而增加。 EV 浓度的降低可能是与衰老相关的表型变化(例如细胞衰老或细胞间信号传导改变的一部分)的结果;两者都是衰老的重要标志。 EV可能是衰老机制的一部分,并且可能因衰老相关机制而发生变化,因此可以作为生物衰老指标。虽然目前的研究表明电动汽车随着年龄的增长而变化,但还需要进一步的研究来阐明它们在衰老中的作用。
作为后续研究,我们研究了 EV 与糖尿病和糖尿病前期之间的关系。 为了解决这个问题,我们设计了一个 HANDLS 纵向子队列,由 58 名参与者组成,他们在相隔大约 5 年的两个时间点捐赠了血浆。在这 58 名参与者中,有 19 名参与者在两个时间点血糖均正常,19 名参与者在时间点 1 血糖正常,并在第二时间点发展为 2 型糖尿病,20 名参与者在时间点 1 为糖尿病前期,并在时间点 2 诊断为糖尿病。该亚队列中的个体已在肥胖状态上进行匹配。我们设计了血糖正常、糖尿病前期和糖尿病参与者的横断面和纵向队列。在我们最近在《糖尿病》杂志上发表的文章(Freeman DW、Noren Hooten N、Eitan E 等人 2018 年)中,糖尿病患者循环中的 EV 水平显着高于血糖正常的对照组。使用细胞特异性 EV 测定,我们发现糖尿病患者的红细胞来源的 EV 较高。我们发现胰岛素抵抗会增加 EV 分泌。此外,具有高水平胰岛素抵抗和细胞功能障碍的个体的 EV 中胰岛素信号蛋白的水平发生了改变。此外,来自糖尿病个体的EV优先被循环白细胞内化。与糖尿病 EV 一起孵育的单核细胞中,培养基和 EV 中的细胞因子水平较高。这些白细胞的微阵列揭示了与细胞存活、氧化应激和免疫功能相关的基因表达途径的改变。总的来说,这些结果表明胰岛素抵抗会增加 EV 的分泌,EV 优先被白细胞内化并改变白细胞功能。 未来的工作将集中于种族和死亡率对 EV 货物的影响以及 EV 货物在慢性疾病和寿命背景下对细胞功能的生物学影响。炎症相关的动脉粥样硬化性外周血管疾病是糖尿病的主要终末器官并发症,导致毁灭性的发病率和死亡率。我们检查了来自糖尿病个体的 EV 蛋白货物以及这些 EV 是否引起内皮细胞的功能变化。 我们对来自血糖正常和糖尿病个体的纵向队列的血浆来源的 EV 中的炎症蛋白水平进行了定量,并使用体外内皮细胞生物学测定来评估这些 EV 与来自横截面队列的样本的功能影响。我们发现 EV 炎症蛋白水平与糖尿病状态之间存在一些显着关联。在我们的纵向队列中,血管生成因子血管内皮生长因子 A (VEGF-A) 与糖尿病状态相关。与血糖正常的个体相比,糖尿病患者的 EV VEGF-A 水平较高。此外,VEGF-A 的 EV 水平与胰岛素抵抗(HOMA-IR)和细胞功能(HOMA-B)的稳态模型评估显着相关。为了测试具有不同炎症货物的 EV 是否可以对内皮细胞表现出不同的影响,我们进行了细胞迁移和免疫荧光测定。我们观察到,与血糖正常个体的 EV 相比,糖尿病个体的 EV 增加了细胞板状伪足的形成和迁移。 在糖尿病患者的 EV 中发现了更高水平的炎症蛋白。我们的数据表明 EV 在糖尿病患者发生的外周血管疾病中发挥着重要作用,并表明 EV 可以作为该疾病的信息诊断工具。
我们还研究了线粒体自由基衰老理论,该理论表明,线粒体和线粒体 DNA (mtDNA) 氧化损伤的累积在衰老过程中发挥着核心作用。从血液中分离出的循环游离线粒体 DNA (ccf-mtDNA) 可能是疾病的生物标志物。细胞外囊泡 (EV) 是一种小型(30-400 nm)脂质结合囊泡,能够作为细胞间通讯系统的一部分运输蛋白质、核酸和脂质。在这里,我们报告血浆中的一部分 ccf-mtDNA 被封装在 EV 中。为了确定 EV mtDNA 水平是否随人类年龄而变化,我们对 30-64 岁个体 EV 中的 mtDNA 进行了横断面和纵向分析。 EV mtDNA 水平随着年龄的增长而下降。此外,培养细胞的最大线粒体呼吸受到年老和年轻供体的 EV 的不同影响。我们的结果表明血浆 mtDNA 存在于 EV 中,EV 衍生的 mtDNA 水平与年龄相关,并且 EV 以 EV 年龄依赖性方式影响线粒体能量。
项目成果
期刊论文数量(0)
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michele k evans其他文献
michele k evans的其他文献
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{{ truncateString('michele k evans', 18)}}的其他基金
Oxidative DNA Damage And Repair In Prostate Cancer
前列腺癌中的氧化 DNA 损伤和修复
- 批准号:
7132274 - 财政年份:
- 资助金额:
$ 57.55万 - 项目类别:
Effects of race and socioeconomic status on the epigenetic aging clock
种族和社会经济地位对表观遗传衰老时钟的影响
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10473355 - 财政年份:
- 资助金额:
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Proteolytic disregulation of the S326C mutant OGG1 DNA repair enzyme
S326C 突变体 OGG1 DNA 修复酶的蛋白水解失调
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8552417 - 财政年份:
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$ 57.55万 - 项目类别:
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8335872 - 财政年份:
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$ 57.55万 - 项目类别:
Interplay between Mitochondrial DNA Haplogroups, Mitochondrial Function, Oxidative Stress, and Hypertension
线粒体 DNA 单倍群、线粒体功能、氧化应激和高血压之间的相互作用
- 批准号:
10250869 - 财政年份:
- 资助金额:
$ 57.55万 - 项目类别:
Measuring DNA Damage and Repair Capacity in Human Popula
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7327074 - 财政年份:
- 资助金额:
$ 57.55万 - 项目类别:
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7327033 - 财政年份:
- 资助金额:
$ 57.55万 - 项目类别:
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