Quantitative analysis of respiratory viruses based on lab-on-a-chip platform.

The quantitative analysis of respiratory viruses is of great importance for rapid diagnosis, precision medicine, and prognosis. Several current quantitative analysis systems have been proposed and commercialized. Although they have been proven in trials, quantitative analyzes based on real samples are still complex, time-consuming, and expensive. Therefore, they are not able to directly quantify real samples. In this work, we presented a lab-on-a-chip platform combined with an automated control system to achieve quantitative analysis from samples to results. We developed a multilayer integrated chip to rapidly extract and quantify RNA of coronavirus disease 2019 (COVID-19) pseudovirus from large-volume nasal swab samples. The dependence of the magnetic bead size and the interfacial effect was studied for the first time, and the conditions of immiscible filtration assisted by surface tension (IFAST) method for nucleic acid extraction were optimized to increase the nucleic acid recovery rate up to 85%. Inside the chip, a pneumatic valve was developed for automatic opening and closing of the liquid channel. The integrated chip platform and automatic control system presented here are advantageous for use in resource-limited settings (RLS). In addition, our method can be extended to other respiratory viruses and other sample types.

CircPSMC3 Inhibits Colorectal Cancer Cell Growth and Migration by Controlling miR-31-5p/YAP/ß-Catenin.

INTRODUCTION: Colorectal cancer (CRC) is hackneyed cancer and a major lethiferous cancer. Circular RNAs (CircRNAs) have been discovered to own important roles in controlling CRC progression. CircPSMC3 is known to exhibit lower expression in diversified cancers. However, the regulatory function of CircPSMC3 in CRC keeps unclear. METHODS: The expression of CircPSMC3 and miR-31-5p was confirmed through RT-qPCR. The cell proliferation was measured through CCK-8 and EdU assays. The protein expression of genes was examined through a western blot. The cell invasion and migration were tested through Transwell and wound healing assays. The binding ability between CircPSMC3 and miR-31-5p was confirmed through the luciferase reporter assay. RESULTS: CircPSMC3 exhibited lower expression in CRC tissues and cell lines. Additionally, CircPSMC3 was revealed to suppress cell proliferation in CRC. Moreover, through Transwell and wound healing assays, CircPSMC3 was discovered to repress CRC cell invasion and migration. In CRC tissues, miR-31-5p expression was up-regulated and negatively correlated with CircPSMC3 expression. Further mechanism exploration experiments disclosed that CircPSMC3 is bound with miR-31-5p to modulate the YAP/ß-catenin axis in CRC. At last, through rescue assays, CircPSMC3 inhibited cell proliferation, invasion and migration through sponging miR-31-5p in CRC. CONCLUSION: Our work was the first time to probe the potential regulatory effects of CircPSMC3 in CRC, and these above results uncovered that CircPSMC3 inhibited CRC cell growth and migration through regulating miR-31-5p/YAP/ß-catenin. This discovery hinted that CircPSMC3 may serve as a useful therapeutic candidate for CRC.

Fabrication of planar monolayer microreactor array for visual statistical analysis and droplet-based digital quantitative analysis in situ.

Planar monolayer microreactor arrays (PMMRAs) make droplet-based numerical measurements and statistical analysis cheap and easy. However, PMMRAs are typically produced in complex microfluidic devices and, moreover, still requires stringent control to reduce droplet loss during heating. In this paper, a simple, reliable, and flexible method for fabricating PMMRAs in a 96-well plate is described in detail by using simple materials and low-cost equipment. The partitioned droplets spontaneously assemble into PMMRAs in the plates, and this distribution is maintained even after incubation. This is advantageous for in situ analysis based on an individual droplet in droplet digital loop-mediated isothermal amplification (ddLAMP) and does not require the transfer of positive droplets. Precise and reproducible quantification of classical swine fever virus (CSFV) extracts was executed in these PMMRAs to verify its availability. Our results demonstrate that the proposed approach not only provides a flexible and controllable execution scheme for droplet-based nucleic acid quantification in resource-limited laboratories but also opens new perspectives for numerous analytical and biochemical applications using droplets as versatile plastic microreactors.

KIF2A participates in the progression of hepatocellular carcinoma and angiogenesis by interacting with Notch1.

Kinesin family member 2A (KIF2A) serves a vital role in the development of hepatocellular carcinoma (HCC); however, the biological effect of KIF2A on the malignant progression of HCC remains unclear. Therefore, the present study was conducted to systematically determine the biological role of KIF2A in HCC and to better understand the molecular mechanism. The differences of KIF2A expression in HHL-5 normal human hepatocytes and the HCC cell lines Li-7, Huh7 and MHCC97 were assessed by reverse transcription-quantitative PCR and western blotting analysis. Moreover, viability, proliferation, migration and invasion of HCC cells were assessed by performing CCK-8, 5-ethynyl-2'-deoxyuridine staining, wound healing and Transwell assays. Additionally, the tube formation assay was performed to evaluate angiogenesis of HUVECs incubated with the conditioned media of HCC cells in vitro. The interaction between KIF2A and Notch1 was analyzed through co-immunoprecipitation assay. KIF2A was revealed to be highly expressed in HCC cells. KIF2A knockdown suppressed HCC cell proliferation, migration and invasion, and impaired in vitro angiogenesis. Furthermore, it was revealed that KIF2A interacted with Notch1 and positively regulated Notch1 expression. The suppressive effects of KIF2A knockdown on HCC cell proliferation, migration, invasion and in vitro angiogenesis were partially reversed by Notch1 overexpression. Overall, KIF2A may act as an oncogene in HCC via activation of the Notch1 signaling pathway.

Nuclear receptor 4A1 (NR4A1) silencing protects hepatocyte against hypoxia-reperfusion injury in vitro by activating liver kinase B1 (LKB1)/AMP-activated protein kinase (AMPK) signaling.

The nuclear receptor 4A1 (NR4A1) is widely involved in the regulation of cell survival and is related to ischemic injury in several organs. This research examined the emerging role and mechanism of NR4A1 in hepatocyte ischemia-reperfusion injury (IRI). BRL-3A cells were subjected to hypoxia-reperfusion (H/R) to simulate an IRI model in vitro. The expression of NR4A1 and liver kinase B1 (LKB1)/AMP-activated protein kinase (AMPK) pathway-related proteins (LKB1, AMPK, and ACC) was detected by western blotting or RT-qPCR under H/R condition after NR4A1 overexpression or silencing. Then, radicicol, an inhibitor of LKB1 pathway, was used to determine the role of NR4A1 in hepatocyte H/R injury by regulating LKB1. Under the help of CCK-8 assay, cell viability was assessed. The levels of ROS, MDA, and SOD were determined with corresponding kits to evaluate oxidative stress. Additionally, RT-qPCR was employed to analyze the releases of the inflammatory factors. Flow cytometry was applied to estimate the apoptosis and its related proteins, and autophagy-associated proteins were assayed by western blotting. Results indicated that NR4A1 was highly expressed, while proteins in LKB1/AMPK signaling was downregulated in BRL-3A cells exposed to H/R. The activation of LKB1/AMPK pathway could be negatively regulated by NR4A1. Moreover, NR4A1 depletion conspicuously promoted cell viability, inhibited oxidative stress as well as inflammation, and induced apoptosis and autophagy in H/R-stimulated BRL-3A cells, which were reversed after radicicol intervention. Collectively, NR4A1/LKB1/AMPK axis is a new protective pathway involved in hepatocyte IRI, shedding new insights into the improvement of hepatocyte IRI.

Polydatin Exerts an Antitumor Effect Through Regulating the miR-382/PD-L1 Axis in Colorectal Cancer.

Background: Colorectal cancer (CRC) is considered as one of the most lethal malignancies worldwide. However, the effective therapies remain limited. Polydatin, a main effective component of the Chinese herb Polygonum cuspidatum, has multiple antitumor activities; however, whether Polydatin has anti-CRC activity is not fully understood. Materials and Methods: CRC cell proliferation and apoptosis were measured after treatment of Polydatin using Cell Counting Kit-8 assay, colony formation assay, and flow cytometer assay. The expression of miR-382 and programmed cell death ligand 1 (PD-L1) were determined in CRC cell lines by quantitative real-time polymerase chain reaction and Western blot, respectively. Furthermore, dual-luciferase reporter assay was conducted to determine the target of miR-382. Moreover, loss-of-functional experiments were used to identify the effect of Polydatin on miR-382. Finally, tumor xenograft experiments were conducted to determine the effect of Polydatin in vivo. Results: As a result, Polydatin effectively inhibited cell proliferation and promoted cell apoptosis in CRC cell lines. PD-L1 was confirmed as a direct target of miR-382. Furthermore, Polydatin could suppress the expression of PD-L1 by upregulating miR-382. Moreover, Polydatin inhibits proliferation and promotes apoptosis of CRC cells by regulating miR-382 and suppressing CRC tumor growth in vivo. Conclusion: Polydatin inhibits CRC cell proliferation and promotes apoptosis by regulating miR-382/PD-L1 axis. Polydatin could be a potential compound to synthesize novel antitumor drugs.

Effect of AMPK/Akt/mTOR pathway on cytokine-induced killer cells immunotherapy on colorectal cancer cells.

PURPOSE: To investigate the effect of cytokine-induced killer cells (CIKs) immunotherapy on colorectal cancer cells and the possible mechanisms using co-culture model of CIKs and human colorectal cancer cells. METHODS: CIKs were induced by INF-γ, CD3 and recombinant human IL-2 culture system in vitro. A co-culture model of CIKs and SW480 colorectal cancer cell line was established. The proliferation, invasion and apoptosis of SW480 cells were detected by CCK-8 (cell counting kit-8), transwell assay and Hoechst staining, respectively. The contents of IL-2, IL-10, insulin-like growth factor (IGF)-ß and vascular endothelial growth factor (VEGF) in the culture medium was detected by ELISA (enzyme-linked immunosorbent assay). The expression levels of AMPK/Akt/mTOR signaling-related proteins were detected by Western blot. RESULTS: CCK-8 assay showed that the proliferative ability of co-cultured CIK+SW480 cells was remarkably lower than that of SW480 cells (p<0.05). Transwell assay showed that the number of invasive cells in co-cultured CIK+SW480 cells was less than that of SW480 cells (p<0.05). Hoechst staining showed that the apoptosis rate of co-cultured CIK+SW480 cells was higher compared with that of SW480 cells (p<0.05). Increased levels of IL-2 and IL-10 and decreased levels of IGF-ß and VEGF were found in the co-culture medium by ELISA (p<0.05). Western blot results indicated that p-AMPK and p-Akt were upregulated, whereas FoxM1 and p-mTOR were downregulated in co-cultured CIK+SW480 cells compared with SW480 cells (p<0.05). CONCLUSIONS: CIKs inhibit the proliferation and invasion of colorectal cancer cells through downregulating FoxM1 and mTOR via AMPK/Akt/mTOR pathway.

Bifunctional Luminomagnetic Rare-Earth Nanorods for High-Contrast Bioimaging Nanoprobes.

Nanoparticles exhibiting both magnetic and luminescent properties are need of the hour for many biological applications. A single compound exhibiting this combination of properties is uncommon. Herein, we report a strategy to synthesize a bifunctional luminomagnetic Gd2-xEuxO3 (x = 0.05 to 0.5) nanorod, with a diameter of ~20 nm and length in ~0.6 µm, using hydrothermal method. Gd2O3:Eu(3+) nanorods have been characterized by studying its structural, optical and magnetic properties. The advantage offered by photoluminescent imaging with Gd2O3:Eu(3+) nanorods is that this ultrafine nanorod material exhibits hypersensitive intense red emission (610 nm) with good brightness (quantum yield more than 90%), which is an essential parameter for high-contrast bioimaging, especially for overcoming auto fluorescent background. The utility of luminomagnetic nanorods for biological applications in high-contrast cell imaging capability and cell toxicity to image two human breast cancer cell lines T47D and MDA-MB-231 are also evaluated. Additionally, to understand the significance of shape of the nanostructure, the photoluminescence and paramagnetic characteristic of Gd2O3:Eu(3+) nanorods were compared with the spherical nanoparticles of Gd2O3:Eu(3+).

Ventriculoperitoneal shunt placement in poor-grade patients with chronic normal pressure hydrocephalus after aneurysmal subarachnoid haemorrhage.

OBJECTIVE: The aim of this study was to analyse the shunt placement in patients who had normal pressure hydrocephalus after poor-grade aneurysmal subarachnoid haemorrhage (aSAH). METHODS: Patients diagnosed with NPH after poor-grade aSAH were divided into a treatment group and control group, based on whether they had received ventriculoperitoneal shunt placement. The treatment group was then divided into an improvement group and non-improvement group according to their recovery. The Glasgow Outcome Scale and Mini Mental Scale Examination were used for 3 month and 1 year follow-up rehabilitation measures. RESULTS: Of the 46 total patients, significant improvement was observed at the 3 month and 1 year follow-ups (p < 0.01) after shunt implantation in the treatment group compared to the control group. Furthermore, patients who were younger (p = 0.022), had better neurological function (higher Glasgow Coma Score, p < 0.01) and less severe hydrocephalus (lower EI, p < 0.01) appears to be more likely to benefit from the shunt. CONCLUSIONS: Patients who had NPH due to poor-grade aSAH would benefit from shunt placement when given the correct candidates and timely management of shunt malfunction. Additionally, the curative effect of the shunt should have been regarded as a long-term goal of rehabilitation in these patients.

Three-dimensionally engineered porous silicon electrodes for Li ion batteries.

The ultimate goal of Li ion battery design should consist of fully accessible metallic current collectors, possibly of nanoscale dimensions, intimately in contact with high capacity stable electrode materials. Here we engineer three-dimensional porous nickel based current collector coated conformally with layers of silicon, which typically suffers from poor cycle life, to form high-capacity electrodes. These binder/conductive additive free silicon electrodes show excellent electrode adhesion resulting in superior cyclic stability and rate capability. The nickel current collector design also allows for an increase in silicon loading per unit area leading to high areal discharge capacities of up to 0.8 mAh/cm(2) without significant loss in rate capability. An excellent electrode utilization (∼85%) and improved cyclic stability for the metal/silicon system is attributed to reduced internal stresses/fracture upon electrode expansion during cycling and shorter ionic/electronic diffusion pathways that help in improving the rate capability of thicker silicon layers.

Functionalization of nitrogen-doped carbon nanotubes with gallium to form Ga-CN(x)-multi-wall carbon nanotube hybrid materials.

In an effort to combine group III-V semiconductors with carbon nanotubes, a simple solution-based technique for gallium functionalization of nitrogen-doped multi-wall carbon nanotubes has been developed. With an aqueous solution of a gallium salt (GaI(3)), it was possible to form covalent bonds between the Ga(3+) ion and the nitrogen atoms of the doped carbon nanotubes to form a gallium nitride-carbon nanotube hybrid at room temperature. This functionalization was evaluated by x-ray photoelectron spectroscopy, energy dispersive x-ray spectroscopy, Raman spectroscopy, scanning electron microscopy and transmission electron microscopy.

3D nanoporous nanowire current collectors for thin film microbatteries.

Conventional thin film batteries are fabricated based on planar current collector designs where the high contact resistance between the current collector and electrodes impedes overall battery performance. Hence, current collectors based on 3D architectures and nanoscale roughness has been proposed to dramatically increase the electrode-current collector surface contact areas and hence significantly reduce interfacial resistance. The nanorod-based current collector configuration is one of several 3D designs which has shown high potential for the development of high energy and high power microbatteries in this regard. Herein we fabricate a nanoporous nanorod based current collector, which provides increased surface area for electrode deposition arising from the porosity of each nanorods, yet keeping an ordered spacing between nanorods for the deposition of subsequent electrolyte and electrode layers. The new nanostructured 3D current collector is demonstrated with a polyaniline (PANI)-based electrode system and is shown to deliver improved rate capability characteristics compared to planar configurations. We have been able to achieve stable capacities of ~32 µAh/cm(2) up to 75 cycles of charge/discharge even at a current rate of ~0.04 mA/cm(2) and have observed good rate capability even at high current rates of ~0.8 mA/cm(2).

Graphene quantum dots derived from carbon fibers.

Graphene quantum dots (GQDs), which are edge-bound nanometer-size graphene pieces, have fascinating optical and electronic properties. These have been synthesized either by nanolithography or from starting materials such as graphene oxide (GO) by the chemical breakdown of their extended planar structure, both of which are multistep tedious processes. Here, we report that during the acid treatment and chemical exfoliation of traditional pitch-based carbon fibers, that are both cheap and commercially available, the stacked graphitic submicrometer domains of the fibers are easily broken down, leading to the creation of GQDs with different size distribution in scalable amounts. The as-produced GQDs, in the size range of 1-4 nm, show two-dimensional morphology, most of which present zigzag edge structure, and are 1-3 atomic layers thick. The photoluminescence of the GQDs can be tailored through varying the size of the GQDs by changing process parameters. Due to the luminescence stability, nanosecond lifetime, biocompatibility, low toxicity, and high water solubility, these GQDs are demonstrated to be excellent probes for high contrast bioimaging and biosensing applications.

Building energy storage device on a single nanowire.

Hybrid electrochemical energy storage devices combine the advantages of battery and supercapacitors, resulting in systems of high energy and power density. Using LiPF(6) electrolyte, the Ni-Sn/PANI electrochemical system, free of Li-based electrodes, works on a hybrid mechanism based on Li intercalation at the anode and PF(6)(-) doping at the cathode. Here, we also demonstrate a composite nanostructure electrochemical device with the anode (Ni-Sn) and cathode (polyaniline, PANI) nanowires packaged within conformal polymer core-shell separator. Parallel array of these nanowire devices shows reversible areal capacity of ∼3 µAh/cm(2) at a current rate of 0.03 mA/cm(2). The work shows the ultimate miniaturization possible for energy storage devices where all essential components can be engineered on a single nanowire.

Direct synthesis of lithium-intercalated graphene for electrochemical energy storage application.

A novel approach for bulk synthesis of lithium-intercalated graphene sheets through the reduction of exfoliated graphene oxide in liquid ammonia and lithium metal is reported. It is demonstrated here that as-synthesized lithiated graphite oxide sheets (Li-RGO) can be directly used as an electrode material in lithium batteries. The electrochemical studies on Li-RGO electrodes show a significant enhancement in the specific capacity of the lithium battery over commercially available graphite electrodes. Partial intercalation of lithium ions in between graphene layers makes this material a good candidate for electrochemical energy storage applications.

Conformal coating of thin polymer electrolyte layer on nanostructured electrode materials for three-dimensional battery applications.

Various three-dimensional (3D) battery architectures have been proposed to address effective power delivery in micro/nanoscale devices and for increasing the stored energy per electrode footprint area. One step toward obtaining 3D configurations in batteries is the formation of core-shell nanowires that combines electrode and electrolyte materials. One of the major challenges however in creating such architectures has been the coating of conformal thin nanolayers of polymer electrolytes around nanostructured electrodes. Here we show conformal coatings of 25-30 nm poly(methyl methacralate) electrolyte layers around individual Ni-Sn nanowires used as anodes for Li ion battery. This configuration shows high discharge capacity and excellent capacity retention even at high rates over extended cycling, allowing for scalable increase in areal capacity with electrode thickness. Our results demonstrate conformal nanoscale anode-electrolyte architectures for an efficient Li ion battery system.