SARS-CoV-2 envelope protein causes acute respiratory distress syndrome (ARDS)-like pathological damages and constitutes an antiviral target

Cytokine storm and multi-organ failure are the main causes of SARS-CoV-2-related death. However, the origin of excessive damages caused by SARS-CoV-2 remains largely unknown. Here we show that the SARS-CoV-2 envelope (2-E) protein alone is able to cause acute respiratory distress syndrome (ARDS)-like damages in vitro and in vivo. 2-E proteins were found to form a type of pH-sensitive cation channels in bilayer lipid membranes. As observed in SARS-CoV-2-infected cells, heterologous expression of 2-E channels induced rapid cell death in various susceptible cell types and robust secretion of cytokines and chemokines in macrophages. Intravenous administration of purified 2-E protein into mice caused ARDS-like pathological damages in lung and spleen. A dominant negative mutation lowering 2-E channel activity attenuated cell death and SARS-CoV-2 production. Newly identified channel inhibitors exhibited potent anti-SARS-CoV-2 activity and excellent cell protective activity in vitro and these activities were positively correlated with inhibition of 2-E channel. Importantly, prophylactic and therapeutic administration of the channel inhibitor effectively reduced both the viral load and secretion of inflammation cytokines in lungs of SARS-CoV-2-infected transgenic mice expressing human angiotensin-converting enzyme 2 (hACE-2). Our study supports that 2-E is a promising drug target against SARS-CoV-2.Subject terms: Cell death, Molecular biology     

3D-QSAR studies of checkpoint kinase 1 inhibitors based on molecular docking and CoMFA

Three-dimensional quantitative structure–activity relationship (3D-QSAR) studies were performed on a series of substituted 1,4-dihydroindeno[1,2-c]pyrazoles inhibitors, using molecular docking and comparative molecular field analysis (CoMFA). The docking results from GOLD 3.0.1 provide a reliable conformational alignment scheme for the 3D-QSAR model. Based on the docking conformations and alignments, highly predictive CoMFA model was built with cross-validated q ² value of 0.534 and non-cross-validated partial least-squares analysis with the optimum components of six showed a conventional r ² value of 0.911. The predictive ability of this model was validated by the testing set with a conventional r ² value of 0.812. Based on the docking and CoMFA, we have identified some key features of the 1,4-dihydroindeno[1,2-c]pyrazoles derivatives that are responsible for checkpoint kinase 1 inhibitory activity. The analyses may be used to design more potent 1,4-dihydroindeno[1,2-c]pyrazoles derivatives and predict their activity prior to synthesis.     

Liquid-Solid Phase-Inversion PLGA Implant for the Treatment of Residual Tumor Tissue after HIFU Ablation

BackgroundHIFU has been shown to be a more suitable alternative for the treatment of primary solid tumors and metastatic diseases than other focal heat ablation techniques due to its noninvasive and extracorporeal nature. However, similar to other focal heat ablation techniques, HIFU is still in need of refinements due to tumor recurrence.MethodsIn this work, we investigated the effectiveness of an adjunct treatment regimen using doxorubicin (DOX)-loaded, injectable, in situ-forming, and phase-inverting PLGA as the second line of defense after HIFU ablation to destroy detrimental residual tumors and to prevent tumor recurrence. All of the statistical analyses were performed using the Statistical Package for the Social Sciences 18.0(SPSS, Inc., Chicago, IL, USA), and p< 0.05 was considered statistically significant. All of the results are presented as the means ± STDEV (standard deviation). For multiple comparisons, ANOVA (differences in tumor volumes, growth rates, apoptosis, proliferation indexes, and Bcl-2 and Bax protein levels) was used when the data were normally distributed with homogenous variance, and rank sum tests were used otherwise. Once significant differences were detected, Student-t tests were used for comparisons between two groups.ResultsOur results revealed that DOX diffused beyond the ablated tissue regions and entered tumor cells that were not affected by the HIFU ablation. Our results also show that HIFU in concert with DOX-loaded PLGA led to a significantly higher rate of tumor cell apoptosis and a lower rate of tumor cell proliferation in the areas beyond the HIFU-ablated tissues and consequently caused significant tumor volume shrinkage (tumor volumes:0.26±0.1,1.09±0.76, and 1.42±0.9cm³ for treatment, sham, and no treatment control, respectively).ConclusionsFrom these results, we concluded that the intralesional injection of DOX-loaded PLGA after HIFU ablation is significantly more effective than HIFU alone for the treatment of solid tumors.     

In vitro concentration dependent transport of phenytoin and phenobarbital,but not ethosuximide,by human P-glycoprotein

AimsOne possible mechanism for epilepsy drug resistance is overexpression of P-glycoprotein in the blood–brain barrier, but whether (or which) antiepileptic drugs (AEDs) are transported by P-gp remains unclear. We evaluated AEDs as P-gp substrates using cell monolayers.Main methodsBi-directional transport assays and concentration equilibrium transport assays (CETAs) were performed for phenytoin (PHT), phenobarbital (PB), and ethosuximide (ESM) using wildtype Madin–Darby Canine Kidney II cell line MDCKII and porcine renal endothelial cell line LLC–PK1 cells and these cells transfected with human MDR1 cDNA to express P-gp.Key findingsWildtype cells demonstrated no efflux transport of PHT, PB, or ESM. In CETAs, both MDR1-transfected cell lines transported PHT from basolateral to apical when PHT loading concentrations were 5 or 10, but not 20 µg/ml. MDCK–MDR1 cells transported PB when initial concentrations were 10 or 20, but not 5 µg/ml. LLC–MDR1 did not transport PB. P-gp inhibitor verapamil blocked efflux transport. MDR1-transfected cells did not transport ESM at 5.6 or 56 µg/ml. Bi-directional transport assays demonstrated weak transport for PHT but not PB or ESM.SignificanceHuman P-gp transports PHT and PB, but not ESM, in a concentration dependent manner. CETA may be more sensitive than bi-directional assays to detect transport of drugs with high passive diffusion. Potential P-gp substrates should be tested at clinically relevant concentration ranges.     

Studies on the Free Amino Acid Content in Anthers of Malesterile and Fertile Plants of Taigu Wheat,with Special Reference to Free Proline Content

Biochemical analyses were made on anthers and pistils at various developmental stages of both male-sterne and fertile plants of Taigu wheat. Analyses ineluded total free amino aeids and free proline. The following results were obtained: 1. There was no significant difference between the content of free proline in anthers of male-sterile and fertile plants at reduction division of mierospore mother cells. 2. In anthers with early uninucleate miorospores, the content of free proline of fertile plants was remarkably higher than that of male-sterile plants. It is interesting to note that at this stage the content of free proline in fertile plants rose to 1.65% of the dry weight of the anther, constituting 50% of the total free amino acids, and amounted to 7-fold of that in male-sterile plants. This result is in line with the results obtained with most cytoplasmic malesterile plants reported by other workers, although malesterility in Taigu wheat is controlled by the nueleus. 3. In pistils, at the stages eorresponding to the early uninneleate and the binueleate stages of the pollen, the free proline content of fertile plants was twice as much as that of the male-sterile plants. This differenee disappeared gradually after fertilization. 4. Tile content of total free amino aoids did not fluetuate as much as the free proline content. There was no differenee in anthers of both types of plants during reduction division of mierospore mother cells. In anthers with early uninueleate pollen grains, total free amino acid content of fertile plants exceeded that of male-sterile plant, the difference levelled off at latter stages. In pistils, before fertilization, the content of total free amino acids of the fertile phmts was slightly higher than that of the male-sterile plants. After fertilization t}fis difference was no nmre noticeable.     

Synthesis of 2H-benzo[b][1,4]oxazin-3(4H)-one derivatives as platelet aggregation inhibitors

Novel 2H-benzo[b][1,4]oxazin-3(4H)-ones have been synthesized by condensation, reduction, O-alkylation and Smiles rearrangement using 3-bromo-4-hydroxy benzaldehyde, anilines, and chloroacetyl chloride as starting materials. All the synthesized compounds have been characterized by (1)H NMR, (13)C NMR, and HRMS, and tested for the inhibitory ability on platelet aggregation. The results have shown that the ADP (adenosine 5'-diphosphate)-induced platelet aggregation was inhibited by 7a-g with the IC(50) value at 10.14-18.83 μmol/L. Compound 7a exhibited the most potent inhibitory effect (IC(50)=10.14 μmol/L) among all the compounds, but less potent than the control drug ticlopidine (3.18 μmol/L) and aspirin (6.07 μmol/L). The preliminary structure-activity relationship (SAR) was initially investigated in the study.     

Epitope Tags beside the N-Terminal Cytoplasmic Tail of Human BST-2 Alter Its Intracellular Trafficking and HIV-1 Restriction

BST-2 blocks the particle release of various enveloped viruses including HIV-1, and this antiviral activity is dependent on the topological arrangement of its four structural domains. Several functions of the cytoplasmic tail (CT) of BST-2 have been previously discussed, but the exact role of this domain remains to be clearly defined. In this study, we investigated the impact of truncation and commonly-used tags addition into the CT region of human BST-2 on its intracellular trafficking and signaling as well as its anti-HIV-1 function. The CT-truncated BST-2 exhibited potent inhibition on Vpu-defective HIV-1 and even wild-type HIV-1. However, the N-terminal HA-tagged CT-truncated BST-2 retained little antiviral activity and dramatically differed from its original protein in the cell surface level and intracellular localization. Further, we showed that the replacement of the CT domain with a hydrophobic tag altered BST-2 function possibly by preventing its normal vesicular trafficking. Notably, we demonstrated that a positive charged motif “KRXK” in the conjunctive region between the cytotail and the transmembrane domain which is conserved in primate BST-2 is important for the protein trafficking and the antiviral function. These results suggest that although the CT of BST-2 is not essential for its antiviral activity, the composition of residues in this region may play important roles in its normal trafficking which subsequently affected its function. These observations provide additional implications for the structure-function model of BST-2.     

Robust stability analysis of delayed Takagi-Sugeno fuzzy Hopfield neural networks with discontinuous activation functions

In this paper, the global robust stability problem of delayed Takagi–Sugeno fuzzy Hopfield neural networks with discontinuous activation functions (TSFHNNs) is considered. Based on Lyapunov stability theory and M-matrices theory, we derive a stability criterion to guarantee the global robust stability of TSFHNNs. Compared with the existing literature, we remove the assumptions on the neuron activations such as Lipschitz conditions, bounded, monotonic increasing property or the assumption that the right-limit value is bigger than the left one at the discontinuous point. Finally, two numerical examples are given to show the effectiveness of the proposed stability results.