scholarly journals Conserved and Divergent Modulation of Calcification in Atherosclerosis and Aortic Valve Disease by Tissue Extracellular Vesicles

2020 ◽  
Author(s):  
Mark C. Blaser ◽  
Fabrizio Buffolo ◽  
Arda Halu ◽  
Florian Schlotter ◽  
Hideyuki Higashi ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Intracellular Signaling ◽  
Target Genes ◽  
Rho Gtpase ◽  
Enzymatic Digestion ◽  
Immunogold Electron Microscopy ◽  
Aortic Valves ◽  
Mirna Sequencing ◽  
Network Approaches ◽  
Gradient Separation

AbstractBackgroundFewer than 50% of patients develop calcification of both atherosclerotic plaques and aortic valves, implying differential pathogenesis. While circulating extracellular vesicles (EVs) act as biomarkers of cardiovascular diseases, tissue-entrapped EVs associate with early mineralization, but their contents, function, and contributions to disease remain unknown.ResultsGlobal proteomics of human carotid artery endarterectomies and calcified aortic valves from a total of 27 donors/patients revealed significant over-representation of proteins with vesicle-associated pathways/ontologies common to both diseases. We exploited enzymatic digestion, serial (ultra)centrifugation and OptiPrep density-gradient separation to isolate EV populations from diseased arteries and valves. Mass spectrometry found 22 EV marker proteins to be highly enriched in the four least-dense OptiPrep fractions while extracellular matrix proteins predominated in denser fractions, as confirmed by CD63 immunogold electron microscopy and nanoparticle tracking analysis. Proteomics and miRNA-sequencing of OptiPrep-enriched tissue EVs quantified 1,104 proteins and 123 miR cargoes linked to 5,182 target genes. Pathway networks of proteins and miR targets common to artery and valve tissue EVs revealed a shared regulation of Rho GTPase and MAPK intracellular signaling cascades. 179 proteins and 5 miRs were significantly altered between artery and valve EVs; multi-omics integration determined that EVs differentially modulated cellular contraction and p53-mediated transcriptional regulation in diseased vascular vs. valvular tissue.ConclusionsOur findings delineate a strategy to isolate, purify, and study protein and RNA cargoes from EVs entrapped in fibrocalcific tissues. Multi-omics and network approaches implicated tissue-resident EVs in human cardiovascular disease.

scholarly journals Integration of miRNA and mRNA Co-Expression Reveals Potential Regulatory Roles of miRNAs in Developmental and Immunological Processes in Calf Ileum during Early Growth

Cells ◽  
2018 ◽  
Vol 7 (9) ◽  
pp. 134 ◽  
Author(s):  
Duy Do ◽  
Pier-Luc Dudemaine ◽  
Bridget Fomenky ◽  
Eveline Ibeagha-Awemu
Keyword(s):  
Protein Metabolism ◽  
Intracellular Signaling ◽  
Early Life ◽  
Growth And Development ◽  
Target Genes ◽  
Early Growth ◽  
Mirna Sequencing ◽  
Significant Enrichment ◽  
Mrna Analysis ◽  
Functional Analyses

This study aimed to investigate the potential regulatory roles of miRNAs in calf ileum developmental transition from the pre- to the post-weaning period. For this purpose, ileum tissues were collected from eight calves at the pre-weaning period and another eight calves at the post-weaning period and miRNA expression characterized by miRNA sequencing, followed by functional analyses. A total of 388 miRNAs, including 81 novel miRNAs, were identified. A total of 220 miRNAs were differentially expressed (DE) between the two periods. The potential functions of DE miRNAs in ileum development were supported by significant enrichment of their target genes in gene ontology terms related to metabolic processes and transcription factor activities or pathways related to metabolism (peroxisomes), vitamin digestion and absorption, lipid and protein metabolism, as well as intracellular signaling. Integration of DE miRNAs and DE mRNAs revealed several DE miRNA-mRNA pairs with crucial roles in ileum development (bta-miR-374a—FBXO18, bta-miR-374a—GTPBP3, bta-miR-374a—GNB2) and immune function (bta-miR-15b—IKBKB). This is the first integrated miRNA-mRNA analysis exploring the potential roles of miRNAs in calf ileum growth and development during early life.

The Pleiotropic Intricacies of Hedgehog Signaling: From Craniofacial Patterning to Carcinogenesis

FACE ◽  
2021 ◽  
pp. 273250162110243
Author(s):  
Mikhail Pakvasa ◽  
Andrew B. Tucker ◽  
Timothy Shen ◽  
Tong-Chuan He ◽  
Russell R. Reid
Keyword(s):  
Intracellular Signaling ◽  
Limb Development ◽  
Hedgehog Signaling ◽  
Target Genes ◽  
Craniofacial Development ◽  
Signaling Cascade ◽  
Signaling Mechanisms ◽  
Intracellular Signaling Cascade ◽  
And Function

Hedgehog signaling was discovered more than 40 years ago in experiments demonstrating that it is a fundamental mediator of limb development. Since that time, it has been shown to be important in development, homeostasis, and disease. The hedgehog pathway proceeds through a pathway highly conserved throughout animals beginning with the extracellular diffusion of hedgehog ligands, proceeding through an intracellular signaling cascade, and ending with the activation of specific target genes. A vast amount of research has been done elucidating hedgehog signaling mechanisms and regulation. This research has found a complex system of genetics and signaling that helps determine how organisms develop and function. This review provides an overview of what is known about hedgehog genetics and signaling, followed by an in-depth discussion of the role of hedgehog signaling in craniofacial development and carcinogenesis.

scholarly journals Circulating or tissue microRNAs and extracellular vesicles as potential lung cancer biomarkers: a systematic review

2017 ◽  
Vol 33 (1) ◽  
pp. 3-9 ◽  
Author(s):  
Yan Song ◽  
Xiuli Yu ◽  
Zongmei Zang ◽  
Guijuan Zhao
Keyword(s):  
Lung Cancer ◽  
Systematic Review ◽  
Cancer Patients ◽  
Extracellular Vesicles ◽  
Target Genes ◽  
Enrichment Analysis ◽  
Cancer Biomarkers ◽  
Biological Processes ◽  
Lung Cancer Patients ◽  
Prediction Of Prognosis

For both lung cancer patients and clinical physicians, tumor biomarkers for more efficient early diagnosis and prediction of prognosis are always wanted. Biomarkers in circulating serum, including microRNAs (miRNAs) and extracellular vesicles, hold the greatest possibilities to partially substitute for tissue biopsy. In this systematic review, studies on circulating or tissue miRNAs and extracellular vesicles as potential biomarkers for lung cancer patients were reviewed and are discussed. Furthermore, the target genes of the miRNAs indicated were identified through the miRTarBase, while the relevant biological processes and pathways of miRNAs in lung cancer were analyzed through MiRNA Enrichment Analysis and Annotation (MiEAA). In conclusion, circulating or tissue miRNAs and extracellular vesicles provide us with a window to explore strategies for diagnosing and assessing prognosis and treatment in lung cancer patients.

scholarly journals Human Bone Marrow Mesenchymal Stromal Cell-Derived Extracellular Vesicles Promote Proliferation of Degenerated Nucleus Pulposus Cells and the Synthesis of Extracellular Matrix Through the SOX4/Wnt/β-Catenin Axis

2021 ◽  
Vol 12 ◽  
Author(s):  
Haoyu Wang ◽  
Fei Li ◽  
Wenrui Ban ◽  
Jing Zhang ◽  
Guiqi Zhang
Keyword(s):  
Extracellular Matrix ◽  
Nucleus Pulposus ◽  
Extracellular Vesicles ◽  
Target Genes ◽  
Intervertebral Disk ◽  
Human Bone ◽  
Nucleus Pulposus Cells ◽  
Synthetic Genes ◽  
Downstream Target ◽  
Transcription Factor 4

Objective: Intervertebral disk degeneration (IDD) is a major cause of pain in the back, neck, and radiculus. Mesenchymal stem cells (MSCs)-derived extracellular vesicles (EVs) are therapeutic in musculoskeletal degenerative diseases such as IDD. This study explored the effect and functional mechanism of human bone MSCs (hBMSCs)-derived EVs in proliferation and apoptosis of degenerated nucleus pulposus cells (DNPCs) and extracellular matrix (ECM) synthesis.Methods: Extracellular vesicles were isolated from hBMSCs and identified. DNPCs were induced by TNF-α. EVs were incubated with DNPCs for 24h. Internalization of EVs by DNPCs, DNPCs proliferation, apoptosis, and expressions of ECM synthetic genes, degrading genes and miR-129-5p were assessed. Downstream target genes of miR-129-5p were predicted. Target relation between miR-129-5p and SRY-box transcription factor 4 (SOX4) was verified. DNPCs proliferation, apoptosis, and ECM synthesis were measured after treatment with EVs and miR-129-5p inhibitor or SOX4 overexpression. Expressions of SOX4 and Wnt/β-catenin pathway-related proteins were determined.Results: hBMSC-EVs promoted DNPCs proliferation, inhibited apoptosis, increased expressions of ECM synthetic genes, and reduced expressions of ECM degrading genes. hBMSC-EVs carried miR-129-5p into DNPCs. Silencing miR-129-5p in EVs partially inverted the effect of EVs on DNPCs proliferation and ECM synthesis. miR-129-5p targeted SOX4. SOX4 overexpression annulled the effect of EVs on DNPCs proliferation and ECM synthesis. Expressions of Wnt1 and β-catenin were decreased in EVs-treated DNPCs, while silencing miR-129-5p in EVs promoted expressions of Wnt1 and β-catenin.Conclusion: hBMSC-EVs promoted DNPCs proliferation and ECM synthesis by carrying miR-129-5p into DNPCs to target SOX4 and deactivating the Wnt/β-catenin axis.

scholarly journals Differential Expression Pattern of Goat Uterine Fluids Extracellular Vesicles miRNAs during Peri-Implantation

Cells ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 2308
Author(s):  
Yanshe Xie ◽  
Guangbin Liu ◽  
Xupeng Zang ◽  
Qun Hu ◽  
Chen Zhou ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Small Rnas ◽  
Target Genes ◽  
Embryo Implantation ◽  
Mature Embryo ◽  
Luciferase Reporter ◽  
Small Rna Sequencing ◽  
Transmission Electron ◽  
Differential Expression Pattern ◽  
Endometrial Epithelium

Early pregnancy failure occurs when a mature embryo attaches to an unreceptive endometrium. During the formation of a receptive endometrium, extracellular vesicles (EVs) of the uterine fluids (UFs) deliver regulatory molecules such as small RNAs to mediate intrauterine communication between the embryo and the endometrium. However, profiling of small RNAs in goat UFs’ EVs during pregnancy recognition (day 16) has not been carried out. In this study, EVs were isolated from UFs on day 16 of the estrous cycle or gestation. They were isolated by Optiprep™ Density G radient (ODG) and verified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blotting. Immunostaining demonstrated that CD63 was present both in the endometrial epithelium and glandular epithelium, and stain intensity was greater in the pregnant endometrium compared to the non-pregnant endometrium. Small RNA sequencing revealed that UFs’ EVs contained numerous sRNA families and a total of 106 differentially expressed miRNAs (DEMs). Additionally, 1867 target genes of the DEMs were obtained, and miRNA–mRNA interaction networks were constructed. GO and KEGG analysis showed that miRNAs were significantly associated with the formation of a receptive endometrium and embryo implantation. In addition, the fluorescence in situ hybridization assay (FISH) showed that chi-miR-451-5p was mainly expressed in stromal cells of the endometrium and a higher level was detected in the endometrial luminal epithelium in pregnant states. Moreover, the dual-luciferase reporter assay showed that chi-miR-451-5p directly binds to PSMB8 and may play an important role in the formation of a receptive endometrium and embryo implantation. In conclusion, these results reveal that UFs’ EVs contain various small RNAs that may be vital in the formation of a receptive endometrium and embryo implantation.

scholarly journals Cyclic AMP response element-binding protein is required in excitatory neurons in the forebrain to sustain wakefulness

SLEEP ◽  
2020 ◽  
Author(s):  
Mathieu E Wimmer ◽  
Rosa Cui ◽  
Jennifer M Blackwell ◽  
Ted Abel
Keyword(s):  
Cyclic Amp ◽  
Intracellular Signaling ◽  
Target Genes ◽  
Binding Protein ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Response Element ◽  
Creb Phosphorylation ◽  
Element Binding Protein ◽  
And Control

Abstract The molecular and intracellular signaling processes that control sleep and wake states remain largely unknown. A consistent observation is that the cyclic adenosine monophosphate (AMP) response element-binding protein (CREB), an activity-dependent transcription factor, is differentially activated during sleep and wakefulness. CREB is phosphorylated by the cyclic AMP/protein kinase A (cAMP/PKA) signaling pathway as well as other kinases, and phosphorylated CREB promotes the transcription of target genes. Genetic studies in flies and mice suggest that CREB signaling influences sleep/wake states by promoting and stabilizing wakefulness. However, it remains unclear where in the brain CREB is required to drive wakefulness. In rats, CREB phosphorylation increases in the cerebral cortex during wakefulness and decreases during sleep, but it is not known if this change is functionally relevant to the maintenance of wakefulness. Here, we used the Cre/lox system to conditionally delete CREB in the forebrain (FB) and in the locus coeruleus (LC), two regions known to be important for the production of arousal and wakefulness. We used polysomnography to measure sleep/wake levels and sleep architecture in conditional CREB mutant mice and control littermates. We found that FB-specific deletion of CREB decreased wakefulness and increased non-rapid eye movement sleep. Mice lacking CREB in the FB were unable to sustain normal periods of wakefulness. On the other hand, deletion of CREB from LC neurons did not change sleep/wake levels or sleep/wake architecture. Taken together, these results suggest that CREB is required in neurons within the FB but not in the LC to promote and stabilize wakefulness.

scholarly journals The Mediation of miR-34a/miR-449c for Immune Cytokines in Acute Cold/Heat-Stressed Broiler Chicken

Animals ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 2168
Author(s):  
Tao Li ◽  
Yiping Song ◽  
Xiuyu Bao ◽  
Jianqin Zhang
Keyword(s):  
Broiler Chicken ◽  
Target Genes ◽  
Growth Period ◽  
Study Groups ◽  
Interferon Γ ◽  
Luciferase Reporter ◽  
Receptor Interaction ◽  
Acute Heat Stress ◽  
Transcriptional Regulatory ◽  
Mirna Sequencing

An increasing amount of evidence has revealed that microRNAs (miRNAs) participated in immune regulation and reaction to acute cold and heat stresses. As a new type of post-transcriptional regulatory factor, miRNA has received widespread attention; However, the specific mechanism used for this regulation still needs to be determined. In this study, thirty broilers at the same growth period were divided into three groups and treated with different temperature and humidity of CS (10–15 °C and 90% Relative Humidity (RH)), HS (39 °C and 90% RH), and NS (26 °C and 50–60% RH) respectively. After 6 h, splenic tissues were collected from all study groups. miRNA sequencing was performed to identify the differentially expressed miRNAs (DEMs) between HS, CS, and NS. We found 33, 37, and 7 DEMs in the HS-NS, HS-CS, CS-NS group. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that DEMs were significantly enriched in cytokine–cytokine receptor interaction and functioned as the cellular responders to stress. We chose two miRNA, miR-34a and miR-449c, from the same family and differential expressed in HS-CS and HS-NS group, as the research objects to predict and verify the target genes. The dual-luciferase reporter assay and quantitative real-time PCR (qRT-PCR) confirmed that two cytokines, IL-2 and IL-12α, were the direct target genes of miR-34a and miR-449c. To further understand the mediation mechanism of miRNAs in acute cold/heat-stressed broiler chicken, a splenic cytokines profile was constructed. The results showed that IL-1β was strongly related to acute heat stress in broiler chicken, and from this we predicted that the increased expression of IL-1β might promote the expression of miR-34a, inducing the upregulation of interferon-γ (INF-γ) and IL-17. Our finds have laid a theoretical foundation for the breeding of poultry resistance and alleviation of the adverse effects of stress.

scholarly journals Small Extracellular Vesicles from Human Fetal Dermal Cells and Their MicroRNA Cargo: KEGG Signaling Pathways Associated with Angiogenesis and Wound Healing

2020 ◽  
Vol 2020 ◽  
pp. 1-18
Author(s):  
Cinzia Maria Chinnici ◽  
Giandomenico Amico ◽  
Alessia Gallo ◽  
Gioacchin Iannolo ◽  
Nicola Cuscino ◽  
...  
Keyword(s):  
Wound Healing ◽  
Signaling Pathways ◽  
Extracellular Vesicles ◽  
Target Genes ◽  
Fold Increase ◽  
Target Cells ◽  
Published Data ◽  
Dermal Cell ◽  
Thrombospondin 1

The use of cell secreted factors in clinical settings could be an alternative to conventional cell therapy, with the advantage of limiting concerns generally associated with traditional cell transplantation, such as tumorigenicity, immunoreactivity, and carrying of infections. Based on our published data, we predict a potential role for extracellular vesicles (EVs) in contributing to the proangiogenic activity of human fetal dermal cell secretome. Depletion of nanosized EVs from secretome significantly impaired its ability to induce formation of mesh-like structures in vitro. The isolated EVs were characterized for size and concentration by nanoparticle tracking analysis, and for protein markers (Rab5+, Alix+, CD63+, and calnexin-). The microRNA profile of EVs revealed 87 microRNAs significantly upregulated (≥15-fold increase) in fetal compared to adult dermal cell-derived EVs. Interestingly, these upregulated microRNAs included microRNAs with a validated role in angiogenesis according to literature. Moreover, the DIANA-TarBase v7.0 analysis confirmed enrichment in the KEGG signaling pathways associated with angiogenesis and wound healing, with the identification of putative target genes including thrombospondin 1. To validate the in silico data, EVs were also characterized for total protein contents. When tested in in vitro angiogenesis, fetal dermal cell-derived EVs were more effective than their adult counterpart in inducing formation of complete mesh-like structures. Furthermore, treatment of fibroblasts with fetal dermal-derived EVs determined a 4-fold increase of thrombospondin 1 protein amounts compared with the untreated fibroblasts. Finally, visualization of CSFE-labeled EVs in the cytosol of target cells suggested a successful uptake of these particles at 4-8 hours of incubation. We conclude that EVs are important contributors of the proangiogenic effect of fetal dermal cell secretome. Hence, EVs could also serve as vehicle for a successful delivery of microRNAs or other molecules of therapeutic interest to target cells.

scholarly journals Circulating MicroRNAs in Extracellular Vesicles as Potential Biomarkers of Alcohol-Induced Neuroinflammation in Adolescence: Gender Differences

2020 ◽  
Vol 21 (18) ◽  
pp. 6730
Author(s):  
Francesc Ibáñez ◽  
Juan R. Ureña-Peralta ◽  
Pilar Costa-Alba ◽  
Jorge-Luis Torres ◽  
Francisco-Javier Laso ◽  
...  
Keyword(s):  
Gender Differences ◽  
Extracellular Vesicles ◽  
Target Genes ◽  
Alcohol Intoxication ◽  
Peripheral Circulation ◽  
Female Adolescents ◽  
Circulating Micrornas ◽  
Male Adolescents ◽  
First Time

Current studies evidence the role of miRNAs in extracellular vesicles (EVs) as key regulators of pathological processes, including neuroinflammation and neurodegeneration. As EVs can cross the blood–brain barrier, and EV miRNAs are very stable in peripheral circulation, we evaluated the potential gender differences in inflammatory-regulated miRNAs levels in human and murine plasma EVs derived from alcohol-intoxicated female and male adolescents, and whether these miRNAs could be used as biomarkers of neuroinflammation. We demonstrated that while alcohol intoxication lowers anti-inflammatory miRNA (mir-146a-5p, mir-21-5p, mir-182-5p) levels in plasma EVs from human and mice female adolescents, these EV miRNAs increased in males. In mice brain cortices, ethanol treatment lowers mir-146a-5p and mir-21-5p levels, while triggering a higher expression of inflammatory target genes (Traf6, Stat3, and Camk2a) in adolescent female mice. These results indicate, for the first time, that female and male adolescents differ as regards the ethanol effects associated with the inflammatory-related plasma miRNAs EVs profile, and suggest that female adolescents are more vulnerable than males to the inflammatory effects of binge alcohol drinking. These findings also support the view that circulating miRNAs in EVs could be useful biomarkers for screening ethanol-induced neuroinflammation and brain damage in adolescence.

scholarly journals Differential Response of Progesterone Receptor Isoforms in Hormone-Dependent and -Independent Facilitation of Female Sexual Receptivity

2006 ◽  
Vol 20 (6) ◽  
pp. 1322-1332 ◽  
Author(s):  
Shaila K. Mani ◽  
Andrea M. Reyna ◽  
Jian Zhong Chen ◽  
Biserka Mulac-Jericevic ◽  
Orla M. Conneely
Keyword(s):  
Signaling Pathways ◽  
Reproductive Behavior ◽  
Intracellular Signaling ◽  
Target Genes ◽  
Critical Role ◽  
Progesterone Receptors ◽  
Protein Isoforms ◽  
Sexual Receptivity ◽  
Progesterone Receptor Isoforms ◽  
Null Mutant Mice

Abstract Neurobehavioral effects of progesterone are mediated primarily by its interaction with neural progesterone receptors (PRs), expressed as PR-A and PR-B protein isoforms. Whereas the expression of two isoforms in the neural tissues is suggestive of their selective cellular responses and modulation of distinct subsets of PR-induced target genes, the role of individual isoforms in brain and behavior is unknown. We have previously demonstrated a critical role for PRs as transcriptional mediators of progesterone (ligand-dependent), and dopamine (ligand-independent)-facilitated female reproductive behavior in female mice lacking both the isoforms of PR. To further elucidate the selective contribution of the individual PR isoforms in female sexual receptive behavior, we used the recently generated PR-A and PR-B isoform-specific null mutant mice. We present evidence for differential responses of each isoform to progesterone and dopamine agonist, SKF 81297 (SKF), and demonstrate a key role for PR-A isoform in both hormone-dependent and -independent facilitation of sexual receptive behavior. Interestingly, whereas both the isoforms were essential for SKF-facilitated sexual behavior, PR-A appeared to play a more important role in the 8-bromo-cAMP-facilitated lordosis response, raising the possibility of distinct intracellular signaling pathways mediating the responses. Finally, we also demonstrate that antiprogestin, RU38486, was an effective inhibitor of PR-A-mediated, progesterone-dependent, but not SKF or 8-bromo-cAMP-dependent sexual receptivity. The data reveal the selective contributions of individual isoforms to the signaling pathways mediating female reproductive behavior.

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