Carbon nanotube (CNT) field-effect transistor (FET)-based biosensors have shown great potential for ultrasensitive biomarker detection, but challenges remain, which include unsatisfactory ...sensitivity, difficulty in stable functionalization, incompatibility with scalable fabrication, and nonuniform performance. Here, we describe ultrasensitive, label-free, and stable FET biosensors built on polymer-sorted high-purity semiconducting CNT films with wafer-scale fabrication and high uniformity. With a floating gate (FG) structure using an ultrathin Y2O3 high-κ dielectric layer, the CNT FET biosensors show amplified response and improved sensitivity compared with those sensors without Y2O3, which is attributed to the chemical gate-coupling effect dominating the sensor response. The CNT FG-FETs are modified to selectively detect specific disease biomarkers, namely, DNA sequences and microvesicles, with theoretical record detection limits as low as 60 aM and 6 particles/mL, respectively. Furthermore, the biosensors exhibit highly uniform performance over the 4 in. wafer as well as superior bias stress stability. The FG CNT FET biosensors could be extended as a universal biosensor platform for the ultrasensitive detection of multiple biological molecules and applied in highly integrated and multiplexed all CNT-FET-based sensor architectures.
Introduction
Brain cells secrete extracellular microvesicles (EVs) that cross the blood‐brain barrier. Involved in cell‐to‐cell communication, EVs contain surface markers and a biologically active ...cargo of molecules specific to their tissue (and cell) of origin, reflecting the tissue or cell's physiological state. Isolation of brain‐secreted EVs (BEVs) from blood provides a minimally invasive way to sample components of brain tissue in Alzheimer's disease (AD), and is considered a form of “liquid biopsy.”
Methods
We performed a comprehensive review of the PubMed literature to assess the biomarker and therapeutic potential of blood‐isolated BEVs in AD.
Results
We summarize methods used for BEV isolation, validation, and novel biomarker discovery, as well as provide insights from 26 studies in humans on the biomarker potential in AD of four cell‐specific BEVs isolated from blood: neuron‐, neural precursor‐, astrocyte‐, and brain vasculature–derived BEVs. Of these, neuron‐derived BEVs has been investigated on several fronts, and these include levels of amyloid‐β and tau proteins, as well as synaptic proteins. In addition, we provide a synopsis of the current landscape of BEV‐based evaluation/monitoring of AD therapeutics based on two published trials and a review of registered clinical trials.
Discussion
Blood‐isolated BEVs have emerged as a novel player in the study of AD, with enormous potential as a diagnostic, evaluation of therapeutics, and treatment tool. The literature has largely concentrated on neuron‐derived BEVs in the blood in AD. Given the multifactorial pathophysiology of AD, additional studies, in neuron‐derived and other brain cell–specific BEVs are warranted to establish BEVs as a robust blood‐based biomarker of AD.
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•Aβ induced neuroinflammation, altered hippocampal oxidative status, led to MVs release.•Total number of MVs increased in the CSF of Aβ-treated group.•The majority of CSF MVs were ...annexin V-positive in Aβ group.•There was no significant change in Annexin V-negative MVs.
Extracellular microvesicles (MVs) as a specific signaling molecule have received much attention in nervous system studies. Alterations in the tissue redox status in pathological conditions, such as Alzheimer’s disease (AD), facilitate the translocation of cell membrane phosphatidylserine to the outer leaflet and lead to the MVs shedding. Annexin V binds with high affinity to phosphatidylserine. Some arguments exist about whether Annexin V-negative MVs should be considered in pathological conditions.
We compared the kinetics of two phenotypes of Annexin V-positive and Annexin V-negative MVs in the cerebrospinal fluid (CSF) of amyloid-β (Aβ)-treated male Wistar rats with flow cytometry technique. The Aβ was injected bilaterally into the cerebral ventricles. Thioflavin T staining was used to confirm the presence of hippocampal Aβ fibrils two weeks post-Aβ injection. Levels of hippocampal interleukin-1β were assessed as an inflammatory index. The CSF malondialdehyde (MDA) concentration was determined. The cognitive impairment and anxiety behaviors were assessed by object recognition and elevated plus maze tests, respectively.
Elevation of MDA levels and a significant rise in the scoring of IL-1β staining were found in the Aβ group. The Aβ induced anxiogenic behavior, impaired novel object recognition memory, and increased the CSF levels of the total number of MVs. The number of Annexin V-positive MVs was significantly higher than Annexin V-negative MVs in all groups.
Data showed that Annexin V-positive MVs potentially have a significant contribution to the pathophysiology of the Aβ-induced cognitive impairment. To catch a clear image of microvesicle production in pathological conditions, both phenotypes of Annexin V-positive and Annexin V-negative MVs should be analyzed and reported.
Alzheimer's disease (AD) is characterized by extracellular amyloid-β (Aβ) deposition, which activates microglia, induces neuroinflammation and drives neurodegeneration. Recent evidence indicates that ...soluble pre-fibrillar Aβ species, rather than insoluble fibrils, are the most toxic forms of Aβ. Preventing soluble Aβ formation represents, therefore, a major goal in AD. We investigated whether microvesicles (MVs) released extracellularly by reactive microglia may contribute to AD degeneration. We found that production of myeloid MVs, likely of microglial origin, is strikingly high in AD patients and in subjects with mild cognitive impairment and that AD MVs are toxic for cultured neurons. The mechanism responsible for MV neurotoxicity was defined in vitro using MVs produced by primary microglia. We demonstrated that neurotoxicity of MVs results from (i) the capability of MV lipids to promote formation of soluble Aβ species from extracellular insoluble aggregates and (ii) from the presence of neurotoxic Aβ forms trafficked to MVs after Aβ internalization into microglia. MV neurotoxicity was neutralized by the Aβ-interacting protein PrP and anti-Aβ antibodies, which prevented binding to neurons of neurotoxic soluble Aβ species. This study identifies microglia-derived MVs as a novel mechanism by which microglia participate in AD degeneration, and suggest new therapeutic strategies for the treatment of the disease.
BACKGROUNDThe rate of cesarean deliveries is steadily growing worldwide as a result of increasing maternal age at first delivery. Ensuring optimal recovery after surgery, specifically the development ...of a functionally competent uterine scar to facilitate vaginal birth after a cesarean delivery (VBAC), is one of the challenges in modern obstetrics. Extracellular microvesicles (EMVs) are secreted by multiple cell types and act as mediators of intercellular interaction during tissue reparation. The immunomodulatory and regenerative effects of EMVs of mesenchymal stromal cells (MSCs) have been studied shown in pre-clinical studies. AIM OF THE STUDYTo evaluate the safety profile of EMVs of mesenchymal stromal placental cells (MSPCs) injected during the cesarean delivery and the impact of this pilot approach on post-surgery recovery. MATERIALS AND METHODSThis pilot study included 53 women undergoing cesarean delivery with (n = 23) or without (n = 30) an injection of 500 µl of MSC EMVs after closing the uterine incision with a single continuous Vicryl suture. RESULTSAll study participants had uncomplicated post-surgery period. The mean inpatient stay duration in women receiving the EMV injection was 4.26 ± 0.09 days vs. 5.33 ± 0.38 in the control group (p<.05). There were no postpartum inflammatory complications in the study group compared with two cases (6.7%) by postpartum endometritis/myometrial infection and one case (3.3%) of lochiometra in the control group. SUMMARYIntra-surgery injection of MSC EMVs was well-tolerated and associated with a lower rate of infectious post-partum complications in women undergoing cesarean delivery.
Abnormal glycosylation is a hallmark of tumor development, and tumor-associated carbohydrate antigens are potential immune targets for tumor therapy. Tumor-associated extracellular microvesicles are ...subcellular vesicles released from cell membranes that have immunogenicity similar to that of precursor cells. However, unmodified tumor-derived microvesicles have weaknesses, such as low immunogenicity, poor biostability, and short half-life in vivo. For the first time, we herein generated extracellular microvesicles containing modified tumor-associated carbohydrate antigens by constructing a cell line with highly expressed antigen-processing enzymes utilizing fluorine-modified monosaccharide substrates via a metabolic oligosaccharide engineering strategy. The microvesicles were applied to tumor immunity, achieving enhanced immunoprophylaxis and immunotherapy effects. Furthermore, the mechanisms of antitumor immunity were explored. Our findings may provide new insights into the adhibition of suitably modified extracellular microvesicles and the development of more effective carbohydrate-based anticancer vaccines.
Osteoarthritis is a prevalent worldwide joint disease, which demonstrates a remarkable adverse effect on the patients' life modality. Medicinal agents, exclusively nonsteroidal anti-inflammatory ...drugs (NSAIDs), have been routinely applied in the clinic. But, their effects are restricted to pain control with insignificant effects on cartilage renovation, which would finally lead to cartilage destruction. In the field of regenerative medicine, many researchers have tried to use stem cells to repair tissues and other human organs. However, in recent years, with the discovery of extracellular microvesicles, especially exosomes, researchers have been able to offer more exciting alternatives on the subject. Exosomes and microvesicles are derived from different types of bone cells such as mesenchymal stem cells, osteoblasts, and osteoclasts. They are also recognized to play substantial roles in bone remodeling processes including osteogenesis, osteoclastogenesis, and angiogenesis. Specifically, exosomes derived from a mesenchymal stem cell have shown a great potential for the desired purpose. Exosomal products include miRNA, DNA, proteins, and other factors. At present, if it is possible to extract exosomes from various stem cells effectively and load certain products or drugs into them, they can be used in diseases, such as rheumatoid arthritis, osteoarthritis, bone fractures, and other diseases. Of course, to achieve proper clinical use, advances have to be made to establish a promising regenerative ability for microvesicles for treatment purposes in the orthopedic disorders. In this review, we describe the exosomes biogenesis and bone cell derived exosomes in the regenerate process of bone and cartilage remodeling.
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ABSTRACT
Endothelial microparticles (EMPs) are endothelium‐derived submicron vesicles that are released in response to diverse stimuli and are elevated in cardiovascular disease, which is correlated ...with risk factors. This study investigates the effect of EMPs on endothelial cell function and dysfunction in a model of free fatty acid (FFA) palmitate‐induced oxidative stress. EMPs were generated from TNF‐α‐stimulated HUVECs and quantified by using flow cytometry. HUVECs were treated with and without palmitate in the presence or absence of EMPs. EMPs were found to carry functional eNOS and to protect against oxidative stress by positively regulating eNOS/Akt signaling, which restored NO production, increased superoxide dismutase and catalase, and suppressed NADPH oxidase and reactive oxygen species (ROS) production, with the involvement of NF‐erythroid 2‐related factor 2 and heme oxygenase‐1. Conversely, under normal conditions, EMPs reduced NO release and increased ROS and redox‐sensitive marker expression. In addition, functional assays using EMP‐treated mouse aortic rings that were performed under homeostatic conditions demonstrated a decline in endothelium‐dependent vasodilatation, but restored the functional response under lipid‐induced oxidative stress. These data indicate that EMPs harbor functional eNOS and potentially play a role in the feedback loop of damage and repair during homeostasis, but are also effective in protecting against FFA‐induced oxidative stress; thus, EMP function is reflected by the microenvironment.— Mahmoud, A. M., Wilkinson, F. L., McCarthy, E. M., Moreno‐Martinez, D., Langford‐Smith, A., Romero, M., Duarte, J., Alexander, M. Y. Endothelial microparticles prevent lipid‐induced endothelial damage via Akt/eNOS signaling and reduced oxidative stress. FASEB J. 31, 4636–4648 (2017). www.fasebj.org
Liquid biopsy is becoming a new source of biomarkers that complement and resolve some of the most important limitations of surgical biopsy, which are the accessibility to the diseased tissue and its ...heterogeneity, especially relevant for tumors. The diseased tissues release their molecule content to the bloodstream in free form, inside a cell or within extracellular vesicles (EVs). While the identification of molecular alterations in total DNA isolated from peripheral blood is already in use for some tumors that secrete large amounts of DNA, it is challenging to assay those secreting lower amounts of molecules as well as for many other non-tumoral pathologies like immunological and cardiovascular diseases. In this scenery, the compartment of diseased tissue-derived EVs will be one of the best alternatives for the detection and identification of current and new biomarkers and targets in the clinical management of these diseases. Here, we review the mechanisms of molecular internalization as well as the correlation of EV's cargo with clinical parameters in tumor and non-tumor diseases, with special emphasis in clinical application.
•Prostate derived extracellular microvesicles transport PSA at their surface.•PSA in extracellular microvesicles can be measured with conventional methods.•Complexed and free PSA are loaded ...differentially in extracellular microvesicles.•The concentrations of complexed and free PSA in extracellular microvesicles could help in the management of PCa patients.
Characterization of PSA in extracellular microvesicles (EVs) and its reactivity to commercial methods.
EVs derived from serum of 47 prostate cancer (PCa) patients, 27 benign prostatic hyperplasia (BPH) patients and 42 healthy controls were analyzed. EVs isolation and quantification of PSA immunoreactive to total (ev-T-PSA) or free (ev-F-PSA) PSA immunoassays, were performed using commercial assays. PSA in CD81+ or CD63+ EVs was determined directly in serum by an immunocapture-ELISA (IC-ELISA).
Ev-T-PSA immunoreactive to Elecsys assay was detected in all samples. Median T-PSA ev/srm ratio was 2.20 % (Q1-Q3: 0.80–4.00 %), although in some samples this ratio reached 59 %. T-PSA ev/srm ratio was higher in those samples with serum T-PSA below 4 µg/L than in those exceeding that cut-off (p < 0.001). T-PSA ev/srm ratio was lower in PCa patients compared to healthy controls and BPH patients (p < 0.001). Elecsys immunoassays detected higher concentrations of ev-T-PSA and ev-F-PSA than Immulite (p < 0.001). PSA was detected by IC-ELISA more intensely in CD81+ EVs than in CD63+ EVs, and ev-T-PSA correlated with PSA+ CD63+ (p < 0.001) but not with PSA+ CD81+.
EVs-bound PSA is another form of circulating PSA whose measurement could be easily performed in clinical laboratories by automated immunoassays.
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