Trace detection of SARS-CoV-2 N-protein by diamond solution-gate field-effect transistor.

In this study, we introduced H-terminated diamond solution-gate field-effect transistor (H-diamond SGFET) to detect trace SARS-CoV-2 N-protein, which plays an important role in replication and transcription of viral RNA. 1-Pyrenebutyric acid-N-hydroxy succinimide ester (Pyr-NHS) was modified on H-diamond surface as linker, on which the specific antibody of SARS-CoV-2 N-protein was catenated. Fourier transform infrared spectrum, scanning electron microscope and energy dispersive spectrum were utilized to demonstrate the modification of H-diamond with Pyr-NHS and antibody. Shifts of IDS(max) at VGS = -500 mV in transfer characteristics of H-diamond SGFET was observed to determine N-protein concentration in phosphate buffer solution. Good linear relationship between IDS(max) and log10(N-protein) was observed from 10-14 to 10-5 g/mL with goodness of fit R2 = 0.90 and sensitivity of 1.98 µA/Log10 [concentration of N-protein] at VDS = -500 mV, VGS = -500 mV. Consequently, this prepared H-diamond SGFET biosensor may provide a new idea for diagnosis of SARS-CoV-2 due to a wide detection range from 10-14 to 10-5 g/mL and low limit of detection 10-14 g/mL.

ROS feedback regulates the microRNA-19-targeted inhibition of the p47phox-mediated LPS-induced inflammatory response.

In acute lung injury/acute respiratory distress syndrome (ALI/ARDS), pathogenesis is associated with the regulation of macrophage-generated oxidative stress, and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX)-derived reactive oxygen species(ROS) are key to regulating oxidative stress. In the present study, we found that miR-19 inhibited the expression of p47phox in macrophages, resulting in the alleviation of the lipopolysaccharides(LPS)-induced inflammatory response. In a mouse LPS-induced model of lung injury, miR-19-deficient murine lung tissue was more susceptible to inflammatory responses and exhibited a higher infiltration rate, a higher number of inflammatory cells in the lungs, a higher level of inflammatory cytokines in the Bronchoalveolar lavage fluid (BALF), and more severe pathological damage in lung tissues. Moreover, following stimulation with LPS, p47phox was expressed at lower levels in miR-19-deficient murine pulmonary inflammatory cells than in those in wild-type rats. In LPS-treated Raw264.7 macrophages, miR-19 mimics blocking the down-regulation of LPS-induced p47phox expression, the accumulation of ROS, and the release of inflammatory cytokines. When siRNA was used to interfere with p47phox expression following stimulation with LPS, a lower level of ROS-mediated inflammatory cytokines were released. We found that the accumulation of ROS inhibited the LPS-induced release of inflammatory cytokines, the upregulation of miR-19 and the down-regulation of LPS-induced p47phox expression. Finally, we constructed a p47phox 3'UTR luciferase reporter plasmid to provide direct confirmation that miR-19 targets p47phox expression. The results of this study indicate the presence of a mechanism by which miR-19 regulates oxidative stress in macrophages. These data also provide potential targets for studies aimed at developing therapies for ARDS.