Spike substitution T813S increases Sarbecovirus fusogenicity by enhancing the usage of TMPRSS2.

SARS-CoV Spike (S) protein shares considerable homology with SARS-CoV-2 S, especially in the conserved S2 subunit (S2). S protein mediates coronavirus receptor binding and membrane fusion, and the latter activity can greatly influence coronavirus infection. We observed that SARS-CoV S is less effective in inducing membrane fusion compared with SARS-CoV-2 S. We identify that S813T mutation is sufficient in S2 interfering with the cleavage of SARS-CoV-2 S by TMPRSS2, reducing spike fusogenicity and pseudoparticle entry. Conversely, the mutation of T813S in SARS-CoV S increased fusion ability and viral replication. Our data suggested that residue 813 in the S was critical for the proteolytic activation, and the change from threonine to serine at 813 position might be an evolutionary feature adopted by SARS-2-related viruses. This finding deepened the understanding of Spike fusogenicity and could provide a new perspective for exploring Sarbecovirus' evolution.

Cucumis sativus PHLOEM PROTEIN 2-A1 like gene positively regulates salt stress tolerance in cucumber seedlings.

PHLOEM PROTEIN 2-A1 like (PP2-A1) gene is a member of the PP2 multigene family, and the protein encoded by which has the function of stress defense. Based on our previous proteomic study of cucumber phloem sap, CsPP2-A1 protein expression was significantly enriched under salt stress. In this paper, we obtained CsPP2-A1 interfering (CsPP2-A1-RNAi) cucumber by Agrobacterium tumefaciens-mediated method. The phenotypic changes of wild-type (WT) cucumber, CsPP2-A1-overexpressing (OE) cucumber, and CsPP2-A1-RNAi cucumber under salt treatment were observed and compared. Furthermore, physiological indicators were measured in four aspects: osmoregulation, membrane permeability, antioxidant system, and photosynthetic system. The analysis of contribution and correlation for each variable were conducted by principal component analysis (PCA) and Pearson's correlation coefficient. The above results showed that CsPP2-A1-RNAi cucumber plants exhibited weaker salt tolerance compared to WT cucumber and CsPP2-A1-OE cucumber plants in terms of phenotype and physiological indicators in response to salt stress, while CsPP2-A1-OE cucumber always showed the robust salt tolerance. Together, these results indicated that CsPP2-A1 brought a salinity tolerance ability to cucumber through osmoregulation and reactive oxygen species (ROS) homeostasis. The results of the study provided evidence for the function of CsPP2-A1 in plant salt tolerance enhancement, and they will serve as a reference for future salt-tolerant cucumber genetic manipulation.

Roles of ß-catenin, TCF-4, and survivin in nasopharyngeal carcinoma: correlation with clinicopathological features and prognostic significance.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is a common malignant tumor of the head and neck region with poorly understood progression and prognosis. The present study aims at exploring whether the expression of ß-catenin, TCF-4, and survivin affects clinicopathological features and prognostic significance in NPC. METHODS: We enrolled 164 patients with NPC and 70 patients with chronic nasopharyngitis (CNP) in this study. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC) were conducted to evaluate the expression of ß-catenin, TCF-4, and survivin. Spearman's rank correlation analysis and Pearson correlation analysis were used to measure the correlation of ß-catenin, TCF-4, and survivin. Risk factors for prognosis and survival conditions of NPC patients were analyzed by Cox proportional hazards model and Kaplan-Meier curves. RESULTS: The results obtained revealed that mRNA and protein expression of ß-catenin, TCF-4, and survivin was higher in NPC tissues than in CNP tissues. Positive correlations amongst ß-catenin, TCF-4, and survivin were identified by Spearman's rank correlation analysis and Pearson correlation analysis. There was a significant correlation in expression of ß-catenin, TCF-4, and survivin with EBV DNA, EBV-VCA-IgA, EBV-EA-IgA, T stage, N stage, and clinicopathological stages. Lower overall survival (OS), distant metastasis-free survival (DMFS), local recurrence-free survival (LRFS), and disease-free survival (DFS) rates were detected in NPC patients with positive expression of ß-catenin, TCF-4, and survivin, in contrast to those with negative expression. Cox proportional hazards model demonstrated that ß-catenin, TCF-4, and survivin protein positive expression were independent risk factors for OS and DFS of NPC prognosis; there was an evident correlation between clinicopathological stages, TCF-4, and EBV-EA-IgA and OS, DMFS, LRFS, and DFS of NPC. CONCLUSIONS: The aforementioned results indicate that ß-catenin, TCF-4, and survivin proteins are highly expressed in NPC, which can be used as factors to predict the malignancy of NPC. In addition, positive expression of ß-catenin, TCF-4, and survivin are potential risk factors that lead to an unfavorable prognosis of OS and DFS in NPC patients.

Evaluation of the peripheral analgesic effect of sufentanil lipid nanoparticles.

OBJECTIVE: We wished to evaluate the effect of sufentanil lipid nanoparticles on peripheral analgesia of inflammatory pain model rats. METHODS: Ninety SD rats were randomly divided into an inflammatory model group (group A, n = 54) and a blank control group (group B, n = 36). Group A was further divided into the sufentanil lipid nanoparticles group (group A1, n = 18), the sufentanil group (group A2, n = 18), and the inflammatory pain model group (group A3, n = 18); group B was divided into the sufentanil lipid nanoparticles group (group B1, n = 18) and the sufentanil group (group B2, n = 18). Rats of group A were given a formalin injection in the foot to produce the inflammatory pain model. Group B rats were given a normal saline foot injection of the same dosage. Then, groups A1 and B1 were given sufentanil lipid nanoparticles (0.82 µg/kg) treatment. Groups A2 and B2 were given sufentanil of the same dosage, and group A3 were given normal saline. Pain scores of Group A rats were recorded and analyzed. The ELISA method was adopted to determine drug concentration in rat brain, plasma, and the inflammatory pain/subcutaneous area. RESULTS: Pain scores of rats in group A3 were always higher than those in groups A1 and A2, and the pain scores of group A2 were higher than in group A1 0-30 min after administration (P < 0.05). The brain drug concentration in groups A2 and B1 fluctuated over time; the brain drug concentrations of groups A2 and B2 were respectively higher than those of groups A1 and B1 (P < 0.05). There was no significant difference between the plasma drug concentrations of different groups at the same time point (P > 0.05); however, there was a notable difference within each group at different time points (P < 0.05), and the drug concentration of the inflammatory tissues in group A1 changed significantly over time (P < 0.05). Thirty minutes after administration, drug concentration in the inflammatory site of group A1 was higher than that of groups A2, B1, and B2 (P < 0.05). CONCLUSION: Sufentanil lipid nanoparticles had a comparatively weak effect on the central nervous system because of their features such as large particle size and targeted and controlled release. They have shown a remarkable analgesic effect in the peripheral inflammatory pain areas.

Comparative global B cell receptor repertoire difference induced by SARS-CoV-2 infection or vaccination via single-cell V(D)J sequencing.

Dynamic changes of the paired heavy and light chain B cell receptor (BCR) repertoire provide an essential insight into understanding the humoral immune response post-SARS-CoV-2 infection and vaccination. However, differences between the endogenous paired BCR repertoire kinetics in SARS-CoV-2 infection and previously recovered/naïve subjects treated with the inactivated vaccine remain largely unknown. We performed single-cell V(D)J sequencing of B cells from six healthy donors with three shots of inactivated SARS-CoV-2 vaccine (BBIBP-CorV), five people who received the BBIBP-CorV vaccine after having recovered from COVID-19, five unvaccinated COVID-19 recovered patients and then integrated with public data of B cells from four SARS-CoV-2-infected subjects. We discovered that BCR variable (V) genes were more prominently used in the SARS-CoV-2 exposed groups (both in the group with active infection and in the group that had recovered) than in the vaccinated groups. The VH gene that expanded the most after SARS-CoV-2 infection was IGHV3-33, while IGHV3-23 in the vaccinated groups. SARS-CoV-2-infected group enhanced more BCR clonal expansion and somatic hypermutation than the vaccinated healthy group. A small proportion of public clonotypes were shared between the SARS-CoV-2 infected, vaccinated healthy, and recovered groups. Moreover, several public antibodies had been identified against SARS-CoV-2 spike protein. We comprehensively characterize the paired heavy and light chain BCR repertoire from SARS-CoV-2 infection to vaccination, providing further guidance for the development of the next-generation precision vaccine.

Influence of Cu on the Improvement of Magnetic Properties and Structure of L10 FePt Nanoparticles.

L10 ordered FePt and FePtCu nanoparticles (NPs) with a good dispersion were successfully fabricated by a simple, green, one-step solid-phase reduction method. Fe (acac)3, Pt (acac)2, and CuO as the precursors were dispersed in NaCl and annealed at different temperatures with an H2-containing atmosphere. As the annealing temperature increased, the chemical order parameter (S), average particle size (D), coercivity (Hc), and saturation magnetization (Ms) of FePt and FePtCu NPs increased and the size distribution range of the particles became wider. The ordered degree, D, Hc, and Ms of FePt NPs were greatly improved by adding 5% Cu. The highest S, D, Hc, and Ms were obtained when FePtCu NPs annealed at 750 °C, which were 0.91, 4.87 nm, 12,200 Oe, and 23.38 emu/g, respectively. The structure and magnetic properties of FePt and FePtCu NPs at different annealing temperatures were investigated and the formation mechanism of FePt and FePtCu NPs were discussed in detail.

microRNA-136 inhibits proliferation and promotes apoptosis and radiosensitivity of cervical carcinoma through the NF-κB pathway by targeting E2F1.

AIMS: Several microRNAs (miRs) are expressed aberrantly and associated with progression, tumorigenesis, and prognosis of haematological and solid tumors. The study aimed to identify the effects involved with microRNA-136 (miR-136) on the concurrent enhancement of proliferation, apoptosis and radiosensitivity of cervical carcinoma through the NF-κB signaling pathway by targeting E2F1. MAIN METHODS: Totally 338 patients with cervical carcinoma were recruited in this study. The expressions of miR-136, E2F1, p65, CyclinD1, Atm, Chk2, Bcl-2, Survivin and Bax were detected using RT-qPCR and Western blot analysis. Cells with highest miR-136 expression were subsequently assigned into different groups. Cell survival and apoptosis rate were detected by colony formation assay and flow cytometry, respectively. KEY FINDINGS: Compared to the sensitivity group, E2F1, p65, Bcl-2 and Survivin exhibited increased levels, while expression of CyclinD1, Atm, Chk2, Bax and miR-136 was reduced in the confrontation group. Cell survival rate was declined at 6 and 8 Gy of X-ray irradiation compared with 0, 2 and 4 Gy. Compared with the blank and NC groups, expression of E2F1, p65, Bcl-2 and Survivin was increased, while that of CyclinD1, Atm, Chk2, Bax and miR-136 was all decreased. The cell survival rate was increased; while apoptosis rate was decreased in the miR-136 inhibitor group. The trends observed in the miR-136 mimics and siRNA-E2F1 groups were contradictory to the miR-136 inhibitor group. SIGNIFICANCE: Based on our results, miR-136 inhibits proliferation, while acting to promote apoptosis and radiosensitivity in cervical carcinoma by targeting E2F1 through the NF-κB signaling pathway, resulting in improved prognoses.

Mechanism of MicroRNA-708 Targeting BAMBI in Cell Proliferation, Migration, and Apoptosis in Mice With Melanoma via the Wnt and TGF-ß Signaling Pathways.

OBJECTIVE: The aim of this study was to evaluate the mechanisms involved with miRNA-708 and its targeting of bone morphogenetic protein and activin membrane-bound inhibitor in cell proliferation, migration, and apoptosis in mice with melanoma via the Wnt and transforming growth factor ß signaling pathways. METHODS: Sixty mice were recruited of which 40 were subsequently assigned into the experimental group (22 mice were successfully established as melanoma model and 18 mice used in tumor xenograft), and the normal control group consisted of 20 mice. B16 cells were assigned to the normal, blank, and negative control, miR-708 mimics, miR-708 inhibitors, si-BAMBI, and miR-708 inhibitors + si-bone morphogenetic protein and activin membrane-bound inhibitor groups. Western blotting and reverse transcription quantitative polymerase chain reaction were employed to detect the expression levels within the tissues and cell lines. TCF luciferase reporter (TOP-FLASH) or a control vector (FOP-FLASH) was applied to detect the activity of the Wnt signaling pathway. MTT3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide assay, flow cytometry, scratch test, and Transwell assay were conducted, respectively, for cell proliferation, apoptosis, migration, and invasion, while tumor xenograft procedures were performed on the nude mice recruited for the study. RESULTS: Compared to the normal control group, the model group displayed increased expressions of bone morphogenetic protein and activin membrane-bound inhibitor, Wnt10B, P53, and Bcl-2; TOPflash activity; ß-catenin expression; cell proliferation; migration; and invasion capabilities while decreased expressions of miR-708, vascular endothelial growth factor, Fas, Bax, Caspase-3, and cleaved Caspase-3 and apoptosis rate. Compared to the blank and negative control groups, the miR-708 mimics and small-interfering RNA-bone morphogenetic protein and activin membrane-bound inhibitor groups exhibited decreases expressions of bone morphogenetic protein and activin membrane-bound inhibitor, Wnt10B, P53, and Bcl-2 and decreased proliferation, migration, and invasion capabilities, while increases in the apoptosis rate, expressions of vascular endothelial growth factor, Fas, Bax, Caspase-3, and cleaved Caspase-3; however, downregulated levels of TOPflash activity and ß-catenin expression were recorded. The miR-708 inhibitors group displayed an opposite trend. CONCLUSION: Downregulation of miR-708-targeted bone morphogenetic protein and activin membrane-bound inhibitor inhibits the proliferation and migration of melanoma cells through the activation of the transforming growth factor ß pathway and the suppression of Wnt pathway.

MicroRNA-152 inhibits tumor cell growth while inducing apoptosis via the transcriptional repression of cathepsin L in gastrointestinal stromal tumor.

OBJECTIVE: MicroRNAs are widely thought to play a regulatory role in gene expression. Although the more unique microRNA expression profiles have been reported in several tumors, there remains a scarcity of knowledge in relation to microRNA expression profiles in GISTs. During this study, through the alteration in the expression of microRNA-152 (miR-152) in gastrointestinal stromal tumor (GIST) cells, we subsequently evaluated its ability to influence the processes associated with cancer, including proliferation, migration, invasion, and apoptosis, as well as the associated mechanisms. METHODS: The expression of miR-152 and cathepsin L (CTSL) in GIST cell lines (GIST882, GIST430, GIST48 and GIST-T1) and normal gastric mucosal cell line RGM-1 were determined. A series of miR-152 mimics, miR-152 inhibitors, and siRNA against CTSL were introduced to treat GIST-T1 cells with the lowest miR-152 and the highest CTSL were assessed. Cell viability, cell cycle entry, apoptosis, and cell migration/invasion were all evaluated by means of CCK-8 assay, flow cytometry analyses of Annexin V-FITC/PI staining, and transwell assays. RESULTS: The target prediction program and luciferase reporter gene assay verified CTSL is the target of miR-152. Regarding the biological significance of miR-152, siRNA knockdown and ectopic expression studies revealed that miR-152 mimic or siRNA against CTSL exposure reduced cell viability and migration/invasion, which resulted in more cells arrested at the S stage, and induced apoptosis. MiR-152 inhibitor exposure was observed to have induced effects on CTSL cells as opposed to those induced by that of the miR-152 mimics. In contrast, miR-152 downregulation abrogated the effects induced by siRNA against CTSL treatment. CONCLUSION: The key findings of this study provided evidence suggesting that miR-152 functions by means of binding to CTSL to induce GIST cell apoptosis and inhibit proliferation, migration, and invasion. The anti-tumor role of miR-152 makes it an attractive therapeutic target for GIST.

Efficient multicyclic sorption and desorption of lead ions on facilely prepared poly(m-phenylenediamine) particles with extremely strong chemoresistance.

Nitric acid, hydrochloric acid and EDTA were carefully chosen as desorbent to systematically evaluate the adsorption/desorption performance of the Pb(2+)-adsorbing fine microparticles of poly(m-phenylenediamine). The sorption/desorption efficiency was maximized by optimizing desorption condition including the desorbent concentration, contact time, and desorption mode. The variation of the solution pH with Pb(2+) desorption was recorded to speculate the desorption mechanism. The practical reusability of the microparticles was elaborated through the sorption-desorption cycle experiments in an optimum condition. It was found that the desorption was very rapid with an equilibrium time of several minutes. A strong dependence of the desorbability on the species and concentration of the desorbents was observed. When 20 mM EDTA was chosen as the desorbent, the highest desorptivity was up to 94.2% that was much higher than those using nitric and hydrochloric acids. A successive sorption-desorption study employing nitric acid indicated that the microparticles could be simply regenerated and reutilized for more than 5 cycles together with Pb(2+) re-adsorption efficiency of about 50% and accumulative Pb(2+) adsorption capacity of up to 720.4 mg L(-1). Facilely prepared, extremely chemoresistant and cost-effective PmPD microparticles would be potentially used for multicyclic sorption of lead ions from aqueous solution.

The effect of DNA-PKcs gene silencing on proliferation, migration, invasion and apoptosis, and in vivo tumorigenicity of human osteosarcoma MG-63 cells.

The purpose of this study was to explore the role by which the DNA-dependent protein kinase complex catalytic subunit (DNA-PKcs) influences osteosarcoma MG-63 cell apoptosis, proliferation, migration and invasion. Osteosarcoma tissues and adjacent normal tissues were obtained from 57 osteosarcoma patients. Human osteosarcoma MG-63 cells were assigned into designated groups including the blank, siRNA-negative control (NC) and siRNA-DNA-PKcs groups. RT-qPCR and Western blotting methods were employed to evaluate the mRNA and protein expressions of DNA-PKcs. A cell counting kit-8 (CCK-8) assay was performed to assess cell viability. The evaluation of cell migration and invasion were conducted by means of Scratch test and Transwell assay. Flow cytometry with PI and annexin V/PI double staining was applied for the analysis of the cell cycle and apoptosis. Twenty-Four Balb/c nude mice were recruited and randomly divided into the blank, siRNA-NC and siRNA-DNA-PKcs groups. Tumorigenicity of the Balb/c nude mice was conducted to evaluate the rate of tumor formation, as well as for the assessment of tumor size and weight, and confirm the number of lung metastatic nodules in the mice post transfection. Osteosarcoma tissues were found to possess greater expression of DNA-PKcs than that of the adjacent normal tissues. DNA-PKcs expression in osteosarcoma tissues were correlated with the clinical stage and metastasis. Compared with the blank and siRNA-NC groups, proliferation, miration, as well as the invasion abilities of the MG-63 cells increased. Furthermore, an increase in apoptosis and cells at the G1 stage in the MG-63 cells was observed, while there were reductions in the cells detected at the S stage. The mRNA and protein expressions of CyclinD1, PCNA, Bcl-2 decreased while those of Bax increased in the siRNA-DNA-PKcs group. The tumor formation rate, tumor diameter, weight and lung metastatic nodules among the nude mice in the siRNA-DNA-PKcs group were all lower than those in the blank and siRNA-NC groups. The observations and findings of the study suggested that the silencing of DNA-PKcs inhibits the proliferation, migration and invasion, while acting to promote cell apoptosis in MG-63 cells and osteosarcoma growth in nude mice.