A high-throughput pipeline for scalable kit-free RNA extraction

An overreliance on commercial, kit-based RNA extraction in the molecular diagnoses of infectious disease presents a challenge in the event of supply chain disruptions and can potentially hinder testing capacity in times of need. In this study, we adapted a well-established, robust TRIzol-based RNA extraction protocol into a high-throughput format through miniaturization and automation. The workflow was validated by RT-qPCR assay for SARS-CoV-2 detection to illustrate its scalability without interference to downstream diagnostic sensitivity and accuracy. This semi-automated, kit-free approach offers a versatile alternative to prevailing integrated solid-phase RNA extraction proprietary systems, with the added advantage of improved cost-effectiveness for high volume acquisition of quality RNA whether for use in clinical diagnoses or for diverse molecular applications

Identification of BPIFA1/SPLUNC1 as an epithelium-derived smooth muscle relaxing factor

Asthma is a chronic airway disease characterized by inflammation, mucus hypersecretion and abnormal airway smooth muscle (ASM) contraction. Bacterial permeability family member A1, BPIFA1, is a secreted innate defence protein. Here we show that BPIFA1 levels are reduced in sputum samples from asthmatic patients and that BPIFA1 is secreted basolaterally from healthy, but not asthmatic human bronchial epithelial cultures (HBECs), where it suppresses ASM contractility by binding to and inhibiting the Ca(2+) influx channel Orai1. We have localized this effect to a specific, C-terminal α-helical region of BPIFA1. Furthermore, tracheas from Bpifa1(−/−) mice are hypercontractile, and this phenotype is reversed by the addition of recombinant BPIFA1. Our data suggest that BPIFA1 deficiency in asthmatic airways promotes Orai1 hyperactivity, increased ASM contraction and airway hyperresponsiveness. Strategies that target Orai1 or the BPIFA1 deficiency in asthma may lead to novel therapies to treat this disease

Design of thermosensitive polymer‐coated magnetic mesoporous silica nanocomposites with a core‐shell‐shell structure as a magnetic/temperature dual‐responsive drug delivery vehicle

A stimuli-responsive nanocomposite with a core-shell-shell structure consisting of iron oxide (Fe3O4) nanoparticles as core, mesoporous silica as middle shell, and poly(N-isopropyl acrylamide-co-acrylic acid) (P[NIPAAm-co-AAc]) as an exterior shell with thermo-responsivity properties was synthesized to be used as a magnetic/temperature responsive drug delivery system. The structure, morphology, and size of P(NIPAAm-co-AAc)-coated mesoporous silica embedded magnetite nanoparticles (P(NIPAAm-co-AAc)@mSiO2@Fe3O4) were characterized by XRD, FTIR, and TEM analyses. Also, the heating ability of mesoporous silica-coated Fe3O4 nanoparticles, and P(NIPAAm-co-AAc)@mSiO2@Fe3O4 nanocomposites was investigated under the exposure of an alternating magnetic field (AMF). The results indicated that the prepared nanocomposites could generate enough heat for hyperthermia applications. Moreover, the magnetic/temperature-responsive drug release behavior of P(NIPAAm-co-AAc)@mSiO2@Fe3O4 nanocomposites loaded with fluorouracil (5-FU) was studied under the exposure of the AMF (frequency = 120 kHz, and amplitude = 22 kA m−1), as well as two different temperatures (37°C and 45°C). The results showed that only 7.8% of the drug could be released after 20 h at 37°C (below the LCST of the copolymer). In contrast, by increasing the temperature of release medium up to 45°C (above the LCST of the copolymer), the amount of released drug was increased up to 47%. Moreover, by exposing the prepared nanocomposite to a safe AMF, a burst release of drug was observed, indicating the excellent responsivity of the carrier to an external magnetic field. These results proved that the obtained nanocomposite has a great performance to be used as a magnetic/temperature-sensitive drug carrier for advanced drug delivery applications