Binding characteristics of pheromone-binding protein 1 in Glyphodes pyloalis to organophosphorus insecticides: Insights from computational and experimental approaches.

Glyphodes pyloalis (Lepidoptera: Pyralidae) is one of the major pests in mulberry production in China, which has developed resistance to various insecticides. Chemoreception is one of the most crucial physiological tactics in insects, playing a pivotal role in recognizing chemical stimuli in the environment, including noxious stimuli such as insecticides. Herein, we obtained recombinant pheromone-binding protein 1 (GpylPBP1) that exhibited antennae-biased expression in G. pyloalis. Ligand-binding assays indicated that GpylPBP1 had the binding affinities to two organophosphorus insecticides, with a higher binding affinity to chlorpyrifos than to phoxim. Computational simulations showed that a mass of nonpolar amino acid residues formed the binding pocket of GpylPBP1 and contributed to the hydrophobic interactions in the bindings of GpylPBP1 to both insecticides. Furthermore, the binding affinities of three GpylPBP1 mutants (F12A, I52A, and F118A) to both insecticides were all significantly reduced compared to those of the GpylPBP1-wild type, suggesting that Phe12, Ile52, and Phe118 residues were crucial binding sites and played crucial roles in the bindings of GpylPBP1 to both insecticides. Our findings can be instrumental in elucidating the effects of insecticides on olfactory recognition in moths and facilitating the development of novel pest management strategies using PBPs as targets based on insect olfaction.

Odorant-Binding Protein 6 Contributes High Binding Affinity to Insecticides in a Parasitic Wasp Meteorus pulchricornis (Hymenoptera: Braconidae).

Meteorus pulchricornis is a preponderant parasitic wasp of various lepidopteran pests. The extensive application of broad-spectrum insecticides usually causes serious threats to the olfactory recognition of nontarget insects such as parasitoid wasps. However, the binding mechanism of odorant-binding proteins (OBPs) to insecticides in parasitoid wasps remains unknown. Herein, we find that the MpulOBP6 protein had a strong binding affinity to three insecticides (phoxim, chlorpyrifos, and chlorfenapyr). Results of computational simulations revealed that the hydrophobic interaction contributed by a mass of nonpolar amino acid residues was the primary driving force in the formation and stabilization of MpulOBP6-insecticide complexes. Among them, four residues (Met75, Val84, Phe121, and Pro122) and two residues (Val84 and Phe111) play an essential role in the binding of MpulOBP6 to phoxim and chlorfenapyr, respectively. Our findings could be instrumental to elucidate the effects of insecticide application toward the olfactory recognition of nontarget insects in the processes of agricultural production.

Prevalence, Risk Factors, and Prognostic Factors of Primary Malignant Bone Neoplasms with Bone Metastasis at Initial Diagnosis: A Population-Based Study.

Background: Bone metastasis (BM) has been proven to be responsible for the poor prognosis of primary malignant bone neoplasms (PMBNs). We aimed to identify the prevalence, risk factors, and prognostic factors for PMBNs patients with BM based on the Surveillance, Epidemiology, and End Results (SEER) database. Methods: 4,758 patients diagnosed with PMBNs from 2010 to 2018 were selected from the SEER database. All patients were divided into two groups: the BM group or the non-BM group. Pearson's chi-square test and Fisher's exact method were used to assess baseline characteristics, and logistic regression analysis was applied to assess risk factors. In addition, a nomogram was constructed based on the results of Cox regression analysis among 227 patients with BM. The good performance and clinical applicability of the nomogram were tested by the concordance index, operating characteristic curve, area under the curve, calibration curves, and decision curve analysis. Results: 227 (4.8%) patients had metastasis to bone at diagnosis. Primary site outside the extremities (axial: odds ratio, OR = 1.770; others: OR = 1.951), Ewing sarcoma (OR = 2.845), larger tumor size (5-8 cm: OR = 3.403; >8 cm: OR = 5.562), tumor extension beyond the periosteum (OR = 2.477), and regional lymph node metastasis (OR = 2.900) were associated with a higher risk of BM at the initial diagnosis of PMBNs. Five independent prognostic factors were found in the survival analysis: pathological type (chondrosarcoma vs. osteosarcoma: hazard ratio, HR = 0.342; Ewing sarcoma vs. osteosarcoma: HR = 0.592; and chordoma vs. osteosarcoma: HR = 0.015), marital status (HR = 2.457), pulmonary metastasis (HR = 1.934), surgery at the primary site (HR = 0.164), and chemotherapy (HR = 0.084). A nomogram based on these prognostic factors could be a good predictor of cancer-specific survival. Conclusions: We identified the prevalence, risk factors, and prognostic factors correlated with BM in PMBNs patients. The related nomogram could be a practical tool for therapeutic decision-making and individual counseling.

Identification of chemosensory genes by antennal transcriptome analysis and expression profiles of odorant-binding proteins in parasitoid wasp Aulacocentrum confusum.

The endoparasitoid wasp, Aulacocentrum confusum (Hymenoptera: Braconidae), is a preponderant natural enemy of the larvae of Glyphodes pyloalis Walker (Lepidoptera: Pyralidae), which is a destructive pest of mulberry trees. We first constructed the antennal transcriptome database of A. confusum. In total, we obtained 48,262,304 clean reads from the dataset and assembled 24,324 unigenes. A total of 12,690 (52.17%) unigenes indicated significant similarity (E-value < 10-5) compared to known protein sequences of other species from the NCBI non-redundant protein database. Gene ontology (GO) and cluster of orthologous groups (COG) analyses were used to determine the functional categories of these genes. A total of 84 putative chemosensory genes were identified from the antennal transcriptome of A. confusum, including 11 putative odorant-binding protein (OBP) genes, six chemosensory protein (CSP) genes, 44 olfactory receptor (OR) genes (including one olfactory co-receptor, Orco), 19 ionotropic receptor (IR) genes, and four sensory neuron membrane protein (SNMP) genes. Results of qPCR assays indicated that among of 11 AconOBPs, nine AconOBP genes were significantly expressed in the antennae of A. confusum adults. AconOBP8 was significantly expressed in the abdomen and AconOBP10 was highly expressed in the thorax. These findings can build a basis for further study on the processes of chemosensory perception in A. confusum at the molecular level.

Elevated levels of IL-17A and IL-35 in plasma and bronchoalveolar lavage fluid are associated with checkpoint inhibitor pneumonitis in patients with non-small cell lung cancer.

Advances in the immunology have identified that interleukin (IL)-17 and IL-35 are cytokines with diverse functions, serving important roles in autoimmune diseases and chronic inflammation. Checkpoint inhibitor pneumonitis (CIP) is focal or diffuse lung inflammation induced by immune checkpoint inhibitors and the underlying pathogenesis has not been fully explored. The aim of the present study was to investigate the roles of IL-17A and IL-35, and the correlation between their levels and different T cell subsets in CIP. The levels of IL-17A and IL-35 in peripheral blood and bronchoalveolar lavage fluid (BALF) were measured in patients with non-small cell lung cancer (NSCLC) with CIP, and the corresponding controls. The percentages of helper T lymphocyte (Th)1, Th2 and Th17 cells, and regulatory T cells (Tregs) in the peripheral blood were synchronically detected. Serum levels of IL-17A and IL-35 were significantly increased at the time of CIP diagnosis compared with the baseline, and significantly decreased upon clinical recovery or improvement. IL-17A and IL-35 were also increased in the BALF during the development of CIP compared with the baseline. Serum levels of IL-17A were positively correlated with the percentages of Th1 and Th17 cells as well as the ratio of Th17 to Tregs, but negatively associated with the frequency of Tregs in CIP. Serum levels of IL-35 were positively correlated with the percentages of Th1 and Tregs, and with the ratio of Th1 to Th2 cells in CIP. A higher frequency of Th1 and Th17 cells, as well as higher ratios of Th17 to Tregs and Th1 to Th2 cells were detected upon development of CIP comparing with the baseline. These data suggested that the activation of Th1 and Th17 cells, as well as Treg inhibition contributed to the imbalanced ratios of Th1 to Th2 and Th17 to Tregs, which resulted in increased secretion of IL-17A and IL-35 in the plasma and BALF; this may present a valuable index to monitor the development and severity of CIP in patients with NSCLC receiving immunotherapy.

Asymmetric Synthesis and Application of Chiral Spirosilabiindanes.

Reported here is the development of a class of chiral spirosilabiindane scaffolds by Rh-catalyzed asymmetric double hydrosilation, for the first time. Enantiopure SPSiOL (spirosilabiindane diol), a new type of chiral building block for the preparation of various chiral ligands and catalysts, was readily prepared on greater than 10 gram scale using this protocol. The potential of this new spirosilabiindane scaffold in asymmetric catalysis was preliminarily demonstrated by development of the corresponding monodentate phosphoramidite ligands (SPSiPhos), which were used in both a Rh-catalyzed hydrogenation and a Pd-catalyzed intramolecular carboamination.

Different screening frequencies of carbapenem-resistant Enterobacteriaceae in patients undergoing hematopoietic stem cell transplantation: which one is better?

BACKGROUND: A consensus has been reached that carbapenem-resistant Enterobacteriaceae (CRE) screening in immunosuppressed individuals can reduce the incidence of CRE bloodstream infection (BSI). METHODS: We retrospectively studied the clinical data of 395 consecutive HSCT patients from September 2017 to April 2019. From September 2017 to June 2018 (period 1), 200 patients received single CRE screening before transplantation. From July 2018 to April 2019 (period 2), 195 patients received continuous weekly CRE screening after admission. For patients colonized with CRE, targeted managements were received: (1) contact precautions and (2) preemptive CRE-targeted treatment if necessary. RESULTS: During period 1, 3 patients with CRE colonization were detected (1.5%). The CRE BSI rate was 2.0% (4 patients), and the related 30-day mortality was 50.0% (2 out of 4 patients). During period 2, 21 patients with CRE colonization were detected, and the detection rate was significantly higher than that in period 1 (P < 0.001). Of the 21 colonized patients, 4 (19.0%) patients were identified as positive for CRE at the first screening, 5 (23.8%) were identified at the second screening, and the remaining 12 (57.1%) were identified at the third or later screening. The CRE BSI rate decreased to 0.5% (1/195), and there were no CRE-related death. Fifteen colonized patients developed neutropenic fever. Thirteen colonizers were preemptively treated with tigecycline within 24 h of fever onset, and they achieved rapid temperature control. One colonizer received tigecycline later than 48 h after fever onset and ultimately survived due to the addition of polymyxin. The other received tigecycline later than 72 h after fever onset and died of septic shock. CONCLUSION: The increase in screening frequency contributed to the detection of patients with CRE colonization. Targeted managements for these colonized patients may contribute to reducing the incidence and mortality of CRE BSI, therefore improving the prognosis of patients.

Lentiviral miR30-based RNA interference against heparanase suppresses melanoma metastasis with lower liver and lung toxicity.

AIM: To construct short hairpin RNAs (shRNAs) and miR30-based shRNAs against heparanase (HPSE) to compare their safety and their effects on HPSE down-modulation in vitro and in vivo to develop a more ideal therapeutic RNA interference (RNAi) vector targeting HPSE. METHODS: First, we constructed shRNAs and miR30-based shRNAs against HPSE (HPSE-shRNAs and HPSE-miRNAs) and packed them into lentiviral vectors. Next, we observed the effects of the shRNAs on knockdown for HPSE expression, adhesion, migration and invasion abilities in human malignant melanoma A375 cells in vitro. Furthermore, we compared the effects of the shRNAs on melanoma growth, metastasis and safety in xenograft models. RESULTS: Our data showed that these artificial miRNAs targeting HPSE could be effective RNAi agents mediated by Pol II promoters in vitro and in vivo, although these miRNAs were not more potent than the HPSE-shRNAs. It was noted that obvious lung injuries, rarely revealed previously, as well as hepatotoxicity could be caused by lentivirus-mediated shRNAs (LV shRNAs) rather than lentivirus-mediated miRNAs (LV miRNAs) in vivo. Furthermore, enhanced expression of pro-inflammatory cytokines IL-6 and TGF-ß1 and endogenous mmu-miR-21a-5p were detected in lung tissues of shRNAs groups, whereas the expression of mmu-let-7a-5p, mmu-let-7b-5p and mmu-let-7c-5p were down-regulated. CONCLUSION: These findings suggest that artificial miRNAs display an improved safety profile of lowered lung injury or hepatotoxicity relative to shRNAs in vivo. The mechanism of lung injuries caused by shRNAs may be correlated with changes of endogenous miRNAs in the lung. Our data here increase the flexibility of a miRNA-based RNAi system for functional genomic and gene therapy applications.

[Effect of chronic exposure to ultraviolet on skin barrier function].

OBJECTIVE: To investigate the effect of chronic ultraviolet (UV) exposure on skin barrier function and photoaging process. METHODS: One hundred and fifty-six volunteers from Hanghzou areas were enrolled in the study. UV-exposed skin areas (neck, dorsum of hand or frontier chest) and UV-unexposed areas (waist, buttock or abdomen) were tested. Probe CM 825 of skin multi-functional detector MPA9 was applied to test the skin water content; probe TM 300 was applied to test transepidermal water loss (TEWL) and probe RVM 600 was applied to detect skin elasticity (Ur/Uf). Relative perfusion unit (PU) of the skin was detected by laser doppler flowmetry (LDF). RESULT: Skin water content value at UV-exposed skin areas was 12.78 ± 2.36 in elderly group (>50y), which was significantly lower than that of UV-unexposed skin areas(23.68 ± 3.24, P= 0.036). Highest level of TEWL (12.98 ± 2.86) g . m(-2) . h(-1) was detected at UV-exposed areas in elderly group; there were trends of increasing TEWL levels at UV-exposed areas than at UV-unexposed areas in all age groups, however, there were no statistical differences (P>0.05). The elasticity of Ur/Uf value at UV-exposed skin areas in elderly group was 0.11 ± 0.07, which was remarkably lower than that of UV-unexposed skin areas (0.32 ± 0.1, P=0.028). No significant difference of skin perfusion was observed between UV-exposed and UV-unexposed areas. CONCLUSION: Chronic exposure to UV may damage skin barrier function and therefore play a role in skin photoaging process.

[Construction of VEGF gene-targeted small interfering RNA mediated by CMV promoter and its silencing effect].

OBJECTIVE: To construct VEGF gene-targeted small interfering RNA (siRNA) and its expression vector driven by CMV promoter and to investigate its interference effect. METHODS: The VEGF gene-targeted hairpin siRNA was designed, two complementary oligonucleotide strands were synthesized. After annealing, two-strand oligonucleotide was inserted into pDC311-SV40-RC vector, which was then identified by PCR and sequenced. Then human U-2 OS cell line was transfected with the vector using lipofectamine method. Finally, ELISA was performed to evaluate the expression of VEGF protein. RESULTS: PCR-identification of positive clone and sequencing confirmed the vector containing the target siRNA. ELISA showed that compared with the control group, the expression levels of VEGF protein in transfected U-2 OS cells were decreased significantly (P<0.05). CONCLUSION: VEGF gene-targeted siRNA and its vector mediated by CMV promoter were successfully constructed, which can reduce the VEGF protein expression after transfecting.

Genistein protects against UVB-induced senescence-like characteristics in human dermal fibroblast by p66Shc down-regulation.

BACKGROUND: Genistein, as an active compound of dietary antioxidants, has shown considerable promise as an effective agent against aging process. However, the effect of genistein on skin photoaging and the associated mechanism remain unclear. OBJECTIVE: To delineate the effect of genistein on UVB-induced senescence in human dermal fibroblasts (HDFs) with emphasis on the mechanism of oxidative pathway regulated by p66Shc involved in the events. METHODS: HDFs were induced to premature senescence by repetitive subcytotoxic doses of UVB irradiation. Cellular apoptosis and DNA cell cycle were analyzed using flow cytometry. Intracellular levels of superoxide dismutase (SOD) and malondialdehyde (MDA) were detected by ELISA. Mutation levels of two large deletions of mitochondrial DNA, 4977bp and 3895bp deletion, were determined by quantitative PCR. Western blot was applied to detect the expression and activation of p66Shc (the 66-kilodalton isoform of the growth factor adapter Shc) and FKHRL1 (a forkhead protein that is intimately linked with intracellular oxidation). RESULTS: Strong activity of senescence-associated beta-galactosidase (SA-beta-gal), high percent of cell apoptosis as well as cell cycle arrest in G0/G1 phase, and increased intracellular oxidative stress were observed in HDFs irradiated by UVB. Genistein exerted dramatically protective effects on HDFs in a dose-dependent manner. Elevated copy numbers of large deletions in mitochondrial DNA were also inhibited by genistein. Down-regulation of total and phosphorylated p66Shc on Ser36, as well as FKHRL1 and its phosphorylation on Thr32, were observed after genistein treatment. CONCLUSION: The results indicate that genistein protects UVB-induced senescence-like characteristics in HDFs via maintenance of antioxidant enzyme activities and modulation of mitochondrial oxidative stress through down-regulation of a p66Shc-dependent signaling pathway, which may provide potential prevention against skin aging and even photoaging.

Matrine inhibits invasiveness and metastasis of human malignant melanoma cell line A375 in vitro.

BACKGROUND: Matrine is a traditional Chinese medicine with significant inhibitory activity against malignant tumors. Its effects on the invasiveness and metastasis of malignant tumors have rarely been reported. AIM: To investigate whether matrine can inhibit the metastasis-related activities of the human malignant melanoma cell line A375 in vitro. METHODS: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and Annexin-V-fluorescein isothiocyanate/propidium iodide (Annexin-V-FITC/PI) affinity assay were used to examine the effects of matrine on the proliferation and apoptosis induction of A375 cells. The morphologic changes of A375 cells were observed by light and electron microscopy. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting were performed to evaluate the expression of heparanase mRNA and protein. The effect of matrine on the adhesion ability and invasiveness of treated A375 cells was tested by cell-Matrigel adhesion assay and Matrigel invasion assay, respectively. RESULTS: Matrine showed significant inhibition of the proliferation of A375 cells in a dose- and time-dependent manner. It also induced apoptosis in a dose-dependent manner. Compared with the control group, the levels of heparanase mRNA and protein expression of A375 cells treated with different concentrations of matrine were decreased significantly, as were their adhesion ability and invasiveness. CONCLUSIONS: These findings indicate that matrine inhibits the invasiveness and metastasis of A375 cells in vitro. The mechanisms may be linked to the inhibition of cellular proliferation, induction of apoptosis, and downregulation of heparanase mRNA and protein expression.