Single-molecule techniques are being developed with the exciting prospect of revolutionizing the healthcare industry by generating vast amounts of genetic and proteomic data. One exceptionally ...promising route is in the use of nanopore sensors. However, a well-known complexity is that detection and capture is predominantly diffusion limited. This problem is compounded when taking into account the capture volume of a nanopore, typically 10(8)-10(10) times smaller than the sample volume. To rectify this disproportionate ratio, we demonstrate a simple, yet powerful, method based on coupling single-molecule dielectrophoretic trapping to nanopore sensing. We show that DNA can be captured from a controllable, but typically much larger, volume and concentrated at the tip of a metallic nanopore. This enables the detection of single molecules at concentrations as low as 5 fM, which is approximately a 10(3) reduction in the limit of detection compared with existing methods, while still maintaining efficient throughput.
The purpose of these guidelines is to assist physicians in recommending, performing, interpreting and reporting the results of FDG PET/CT for oncological imaging of adult patients. PET is a ...quantitative imaging technique and therefore requires a common quality control (QC)/quality assurance (QA) procedure to maintain the accuracy and precision of quantitation. Repeatability and reproducibility are two essential requirements for any quantitative measurement and/or imaging biomarker. Repeatability relates to the uncertainty in obtaining the same result in the same patient when he or she is examined more than once on the same system. However, imaging biomarkers should also have adequate reproducibility, i.e. the ability to yield the same result in the same patient when that patient is examined on different systems and at different imaging sites. Adequate repeatability and reproducibility are essential for the clinical management of patients and the use of FDG PET/CT within multicentre trials. A common standardised imaging procedure will help promote the appropriate use of FDG PET/CT imaging and increase the value of publications and, therefore, their contribution to evidence-based medicine. Moreover, consistency in numerical values between platforms and institutes that acquire the data will potentially enhance the role of semiquantitative and quantitative image interpretation. Precision and accuracy are additionally important as FDG PET/CT is used to evaluate tumour response as well as for diagnosis, prognosis and staging. Therefore both the previous and these new guidelines specifically aim to achieve standardised uptake value harmonisation in multicentre settings.
The cellular cytoskeleton is crucial for many cellular functions such as cell motility and wound healing, as well as other processes that require shape change or force generation. Actin is one ...cytoskeleton component that regulates cell mechanics. Important properties driving this regulation include the amount of actin, its level of cross-linking, and its coordination with the activity of specific molecular motors like myosin. While studies investigating the contribution of myosin activity to cell mechanics have been performed on cells attached to a substrate, we investigated mechanical properties of cells in suspension. To do this, we used multiple probes for cell mechanics including a microfluidic optical stretcher, a microfluidic microcirculation mimetic, and real-time deformability cytometry. We found that nonadherent blood cells, cells arrested in mitosis, and naturally adherent cells brought into suspension, stiffen and become more solidlike upon myosin inhibition across multiple timescales (milliseconds to minutes). Our results hold across several pharmacological and genetic perturbations targeting myosin. Our findings suggest that myosin II activity contributes to increased whole-cell compliance and fluidity. This finding is contrary to what has been reported for cells attached to a substrate, which stiffen via active myosin driven prestress. Our results establish the importance of myosin II as an active component in modulating suspended cell mechanics, with a functional role distinctly different from that for substrate-adhered cells.
Neuron-targeted electrical modulation Otto, Kevin J; Schmidt, Christine E
Science (American Association for the Advancement of Science),
03/2020, Volume:
367, Issue:
6484
Journal Article
Peer reviewed
Engineering neurons to make conductive polymers enables cell type–specific behaviors
Conductive polymers have been widely studied and used for biomedical applications—including as biosensors, neural ...prostheses, and bioactuators—and for drug delivery and tissue engineering (
1
). Conductive polymers are organic chains of alternating single and double bonds, which endow the polymers with metal-like semiconductive properties. Exogenous application of electrical stimulation to these polymers can promote cellular activities such as proliferation, adhesion, migration, differentiation, and protein secretion. Because many cells and tissues, particularly neurons, are responsive to electrical fields, conductive polymers are attractive for biological and medical applications. On page 1372 of this issue, Liu
et al.
(
2
) report a genetically targeted approach to assemble conductive polymers in neurons. This in turn remodels membrane electrical properties and enables cell type–specific cellular and behavioral modulation, such as control of neuronal firing, as demonstrated in cultures of rat hippocampal neurons, mouse brain slices, human cortical spheroids, and in living
Caenorhabditis elegans
worms.
Cyclins and cyclin-dependent kinases (CDKs) are hyperactivated in numerous human tumors. To identify means of interfering with cyclins/CDKs, we performed nine genome-wide screens for human microRNAs ...(miRNAs) directly regulating cell-cycle proteins. We uncovered a distinct class of miRNAs that target nearly all cyclins/CDKs, which are very effective in inhibiting cancer cell proliferation. By profiling the response of over 120 human cancer cell lines, we derived an expression-based algorithm that can predict the response of tumors to cell-cycle-targeting miRNAs. Using systemic administration of nanoparticle-formulated miRNAs, we inhibited tumor progression in seven mouse xenograft models, including three treatment-refractory patient-derived tumors, without affecting normal tissues. Our results highlight the utility of using cell-cycle-targeting miRNAs for treatment of refractory cancer types.
•Characterization of human microRNAs which target the cell-cycle machinery•Profiling cell-cycle-targeting miRNAs against 122 human cancer cell lines from CCLE•Algorithm to predict the response of tumors to cell-cycle-targeting miRNAs•In vivo delivery of cell-cycle-targeting miRNAs inhibits cancer growth
By performing screens for miRNAs targeting cell-cycle proteins, Hydbring et al. identify a class of miRNAs that target multiple cyclins and CDKs. Nanoparticle delivery of these miRNAs inhibits tumor growth in several xenograft models, including treatment-refractory patient-derived xenografts.
Electrical stimulation using implantable electrodes is widely used to treat various neuronal disorders such as Parkinson’s disease and epilepsy and is a widely used research tool in neuroscience ...studies. However, to date, devices that help better understand the mechanisms of electrical stimulation in neural tissues have been limited to opaque neural electrodes. Imaging spatiotemporal neural responses to electrical stimulation with minimal artifact could allow for various studies that are impossible with existing opaque electrodes. Here, we demonstrate electrical brain stimulation and simultaneous optical monitoring of the underlying neural tissues using carbon-based, fully transparent graphene electrodes implanted in GCaMP6f mice. Fluorescence imaging of neural activity for varying electrical stimulation parameters was conducted with minimal image artifact through transparent graphene electrodes. In addition, full-field imaging of electrical stimulation verified more efficient neural activation with cathode leading stimulation compared to anode leading stimulation. We have characterized the charge density limitation of capacitive four-layer graphene electrodes as 116.07–174.10 μC/cm2 based on electrochemical impedance spectroscopy, cyclic voltammetry, failure bench testing, and in vivo testing. This study demonstrates the transparent ability of graphene neural electrodes and provides a method to further increase understanding and potentially improve therapeutic electrical stimulation in the central and peripheral nervous systems.
Flexible neural probes are hypothesized to reduce the chronic foreign body response (FBR) mainly by reducing the strain-stress caused by an interplay between the tethered probe and the brain's ...micromotion. However, a large discrepancy of Young's modulus still exists (3-6 orders of magnitude) between the flexible probes and the brain tissue. This raises the question of whether we need to bridge this gap; would increasing the probe flexibility proportionally reduce the FBR?
Using novel off-stoichiometry thiol-enes-epoxy (OSTE+) polymer probes developed in our previous work, we quantitatively evaluated the FBR to four types of probes with different softness: silicon (~150 GPa), polyimide (1.5 GPa), OSTE+
(300 MPa), and OSTE+
(6 MPa).
We observed a significant reduction in the fluorescence intensity of biomarkers for activated microglia/macrophages and blood-brain barrier (BBB) leakiness around the three soft polymer probes compared to the silicon probe, both at 4 weeks and 8 weeks post-implantation. However, we did not observe any consistent differences in the biomarkers among the polymer probes.
The results suggest that the mechanical compliance of neural probes can mediate the degree of FBR, but its impact diminishes after a hypothetical threshold level. This infers that resolving the mechanical mismatch alone has a limited effect on improving the lifetime of neural implants.
Introduction
The vagus nerve facilitates bidirectional communication between the gut and brain. A low vagal tone has been described in many conditions characterized by inflammation and gut dysbiosis, ...including hypertension (HTN). While linked with clinical observations, the mechanisms of vagal gut‐brain communication are not fully understood. Moreover, the role of the sub‐diaphragmatic gut‐projecting vagal nerve branches in control of blood pressure (BP) remains unclear. The objective of this study was to examine the systemic and central effects of augmenting vagus nerve activity using subdiaphragmatic vagus nerve stimulation (sdVNS) during the development of HTN in spontaneously hypertensive rat (SHR).
Methods
Male SHR (8 wo) were implanted with BP telemeters and either a silicone cuff electrode (STIM, N=5) or sham cuff (SHAM, N=5) on the ventral subdiaphragmatic vagal trunk. Following recovery and baseline BP recordings, STIM rats were connected to a stimulus generator, and subjected to biphasic (600 μA, 500 us/phase, 25 Hz) vagus nerve stimulation (10 minutes/ session, 3 sessions/ day, 5 days/ week) for 7 weeks. SHAM rats were subjected to similar handling but with no stimulation. Blood pressure was recorded with a 24‐hour continuous recording once per week for eight weeks in all rats. Rats were sacrificed and cecal content, plasma, proximal colon, and nucleus tractus solitarius (NTS) were collected. Bacterial 16s sequencing was used to determine abundance and composition of gut bacteria. Inflammatory cytokines including INF‐γ, IL‐1β, IL‐4, IL‐5, IL‐6, KC/GRO, IL‐10, IL‐13, TNF‐α, IL‐17α, and corticosterone were measured in serum and proximal colon. To further discern mechanisms, the NTS transcriptome was measured at endpoint using RNA‐seq.
Results
In SHAM SHR, both the mean BP (MBP) and diastolic BP (DBP) were significantly elevated in week one (ΔMBP= +13.6 mmHg, p = 0.01; ΔDBP= +11.5 mmHg, p = 0.03) when compared to baseline BP. In contrast, in STIM SHR, the MBP and DBP were not significantly elevated until week five of sdVNS (ΔMBP= +25.0 mmHg, p < 0.001; ΔDBP= +20.8 mmHg, p < 0.001) when compared to baseline BP. Systolic BP (SBP) values were similarly elevated compared to baseline at all time points examined. At endpoint, we observed no difference in any of the 11 proinflammatory cytokines measured in either plasma or colonic lysates between SHAM and STIM SHR. In addition, no difference was observed in the composition or abundance of major gut bacterial phyla. However, chronic intermittent sdVNS resulted in 387 up‐regulated and 236 down‐regulated genes in the NTS. Gene set enrichment analysis revealed differential regulation of central inflammatory and neural signaling pathways in the NTS of SHR following sdVNS.
Conclusions
Chronic intermittent sdVNS can delay but not prevent the onset of HTN in the SHR. This delay does not coincide with changes in systemic inflammatory markers or gut bacterial composition; however, changes in the NTS transcriptome indicate afferent rather than efferent effects of chronic intermittent sdVNS in the SHR. Future studies will dissect precise central effects of afferent gut‐brain communication.
Objective: This study aims to identify the impact of using edge sites over center sites on a planar silicon microelectrode array. Methods: We used custom-designed, silicon-substrate multisite ...microelectrode arrays with sites on the center, edge, and tip. We compared their single unit recording capability, noise level, impedance, and histology to identify the differences between each site location. Wide and narrow devices were used to evaluate if the differences are consistent and meet theoretical expectations. Results: On the wide device, significantly more number of edge sites were functional than center sites over the course of 8 weeks with generally higher signal-to-noise amplitude ratio. On the narrow device, edge sites also performed generally better than center sites, but the differences were not significant and smaller than wide devices. The data from the tip sites were inconclusive. Conclusion: Edge sites outperformed center sites in terms of single unit recording capability. This benefit decreased as the device gets narrower and the distance to center sites decreases. Significance: We showed that a simple alteration to the site placement can greatly enhance the functionality of silicon microelectrodes. This study promotes the idea that not only the substrate but also the site architecture needs attention to lengthen the lifetime of neural implants.
During type 1 diabetes, an autoimmune attack destroys pancreatic β-cells leading to the inability to maintain glucose homeostasis. These β-cells are neuroresponsive endocrine cells which normally ...secrete insulin partially in response to input from the vagus nerve. This neural pathway can be utilized as a point of therapeutic intervention by delivering exogenous stimulation to drive increased insulin secretion. In this study, a cuff electrode was implanted on the pancreatic branch of the vagus nerve just prior to pancreatic insertion in rats, and a continuous glucose meter was implanted into the descending aorta. Streptozotocin (STZ) was used to induce a diabetic state, and changes in blood glucose were assessed using various stimulation parameters. Stimulation driven changes in hormone secretion, pancreatic blood flow, and islet cell populations were assessed. We found increased changes in the rate of blood glucose change during stimulation which subsided after stimulation ended paired with increased concentration of circulating insulin. We did not observe increased pancreatic perfusion, which suggests that the modulation of blood glucose was due to the activation of b-cells rather than changes in the extra-organ transport of insulin. Pancreatic neuromodulation showed potentially protective effects by reducing deficits in islet diameter, and ameliorating insulin loss after STZ treatment.
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