CEACAM6 as a Novel Therapeutic Target to Boost HO-1-mediated Antioxidant Defense in COPD.

Rationale: Tobacco smoking and air pollution are primary causes of chronic obstructive pulmonary disease (COPD). However, only a minority of smokers develop COPD. The mechanisms underlying the defense against nitrosative/oxidative stress in nonsusceptible smokers to COPD remain largely unresolved. Objectives: To investigate the defense mechanisms against nitrosative/oxidative stress that possibly prevent COPD development or progression. Methods: Four cohorts were investigated: 1) sputum samples (healthy, n = 4; COPD, n = 37), 2) lung tissue samples (healthy, n = 13; smokers without COPD, n = 10; smoker+COPD, n = 17), 3) pulmonary lobectomy tissue samples (no/mild emphysema, n = 6), and 4) blood samples (healthy, n = 6; COPD, n = 18). We screened 3-nitrotyrosine (3-NT) levels, as indication of nitrosative/oxidative stress, in human samples. We established a novel in vitro model of a cigarette smoke extract (CSE)-resistant cell line and studied 3-NT formation, antioxidant capacity, and transcriptomic profiles. Results were validated in lung tissue, isolated primary cells, and an ex vivo model using adeno-associated virus-mediated gene transduction and human precision-cut lung slices. Measurements and Main Results: 3-NT levels correlate with COPD severity of patients. In CSE-resistant cells, nitrosative/oxidative stress upon CSE treatment was attenuated, paralleled by profound upregulation of heme oxygenase-1 (HO-1). We identified carcinoembryonic antigen cell adhesion molecule 6 (CEACAM6) as a negative regulator of HO-1-mediated nitrosative/oxidative stress defense in human alveolar type 2 epithelial cells (hAEC2s). Consistently, inhibition of HO-1 activity in hAEC2s increased the susceptibility toward CSE-induced damage. Epithelium-specific CEACAM6 overexpression increased nitrosative/oxidative stress and cell death in human precision-cut lung slices on CSE treatment. Conclusions: CEACAM6 expression determines the hAEC2 sensitivity to nitrosative/oxidative stress triggering emphysema development/progression in susceptible smokers.

SPARC, a Novel Regulator of Vascular Cell Function in Pulmonary Hypertension.

BACKGROUND: Pulmonary hypertension (PH) is a life-threatening disease, characterized by excessive pulmonary vascular remodeling, leading to elevated pulmonary arterial pressure and right heart hypertrophy. PH can be caused by chronic hypoxia, leading to hyper-proliferation of pulmonary arterial smooth muscle cells (PASMCs) and apoptosis-resistant pulmonary microvascular endothelial cells (PMVECs). On reexposure to normoxia, chronic hypoxia-induced PH in mice is reversible. In this study, the authors aim to identify novel candidate genes involved in pulmonary vascular remodeling specifically in the pulmonary vasculature. METHODS: After microarray analysis, the authors assessed the role of SPARC (secreted protein acidic and rich in cysteine) in PH using lung tissue from idiopathic pulmonary arterial hypertension (IPAH) patients, as well as from chronically hypoxic mice. In vitro studies were conducted in primary human PASMCs and PMVECs. In vivo function of SPARC was proven in chronic hypoxia-induced PH in mice by using an adeno-associated virus-mediated Sparc knockdown approach. RESULTS: C57BL/6J mice were exposed to normoxia, chronic hypoxia, or chronic hypoxia with subsequent reexposure to normoxia for different time points. Microarray analysis of the pulmonary vascular compartment after laser microdissection identified Sparc as one of the genes downregulated at all reoxygenation time points investigated. Intriguingly, SPARC was vice versa upregulated in lungs during development of hypoxia-induced PH in mice as well as in IPAH, although SPARC plasma levels were not elevated in PH. TGF-ß1 (transforming growth factor ß1) or HIF2A (hypoxia-inducible factor 2A) signaling pathways induced SPARC expression in human PASMCs. In loss of function studies, SPARC silencing enhanced apoptosis and reduced proliferation. In gain of function studies, elevated SPARC levels induced PASMCs, but not PMVECs, proliferation. Coculture and conditioned medium experiments revealed that PMVECs-secreted SPARC acts as a paracrine factor triggering PASMCs proliferation. Contrary to the authors' expectations, in vivo congenital Sparc knockout mice were not protected from hypoxia-induced PH, most probably because of counter-regulatory proproliferative signaling. However, adeno-associated virus-mediated Sparc knockdown in adult mice significantly improved hemodynamic and cardiac function in PH mice. CONCLUSIONS: This study provides evidence for the involvement of SPARC in the pathogenesis of human PH and chronic hypoxia-induced PH in mice, most likely by affecting vascular cell function.

Fibroblast growth factor 10 reverses cigarette smoke- and elastase-induced emphysema and pulmonary hypertension in mice.

BACKGROUND: COPD is an incurable disease and a leading cause of death worldwide. In mice, fibroblast growth factor (FGF)10 is essential for lung morphogenesis, and in humans, polymorphisms in the human FGF10 gene correlate with an increased susceptibility to develop COPD. METHODS: We analysed FGF10 signalling in human lung sections and isolated cells from healthy donor, smoker and COPD lungs. The development of emphysema and PH was investigated in Fgf10+/- and Fgfr2b+/- (FGF receptor 2b) mice upon chronic exposure to cigarette smoke. In addition, we overexpressed FGF10 in mice following elastase- or cigarette smoke-induced emphysema and pulmonary hypertension (PH). RESULTS: We found impaired FGF10 expression in human lung alveolar walls and in primary interstitial COPD lung fibroblasts. In contrast, FGF10 expression was increased in large pulmonary vessels in COPD lungs. Consequently, we identified impaired FGF10 signalling in alveolar walls as an integral part of the pathomechanism that leads to emphysema and PH development: mice with impaired FGF10 signalling (Fgf10+/- and Fgfr2b+/- ) spontaneously developed lung emphysema, PH and other typical pathomechanistic features that generally arise in response to cigarette smoke exposure. CONCLUSION: In a therapeutic approach, FGF10 overexpression successfully restored lung alveolar and vascular structure in mice with established cigarette smoke- and elastase-induced emphysema and PH. FGF10 treatment triggered an initial increase in the number of alveolar type 2 cells that gradually returned to the basal level when the FGF10-mediated repair process progressed. Therefore, the application of recombinant FGF10 or stimulation of the downstream signalling cascade might represent a novel therapeutic strategy in the future.

Nicotine promotes e-cigarette vapour-induced lung inflammation and structural alterations.

BACKGROUND: Electronic cigarette (e-cigarette) vapour is gaining popularity as an alternative to tobacco smoking and can induce acute lung injury. However, the specific role of nicotine in e-cigarette vapour and its long-term effects on the airways, lung parenchyma and vasculature remain unclear. RESULTS: In vitro exposure to nicotine-containing e-cigarette vapour extract (ECVE) or to nicotine-free e-cigarette vapour extract (NF ECVE) induced changes in gene expression of epithelial cells and pulmonary arterial smooth muscle cells (PASMCs), but ECVE in particular caused functional alterations (e.g. a decrease in human and mouse PASMC proliferation by 29.3±5.3% and 44.3±8.4%, respectively). Additionally, acute inhalation of nicotine-containing e-cigarette vapour (ECV) but not nicotine-free e-cigarette vapour (NF ECV) increased pulmonary endothelial permeability in isolated lungs. Long-term in vivo exposure of mice to ECV for 8 months significantly increased the number of inflammatory cells, in particular lymphocytes, compared to control and NF ECV in the bronchoalveolar fluid (BALF) (ECV: 853.4±150.8 cells·mL-1; control: 37.0±21.1 cells·mL-1; NF ECV: 198.6±94.9 cells·mL-1) and in lung tissue (ECV: 25.7±3.3 cells·mm-3; control: 4.8±1.1 cells·mm-3; NF ECV: 14.1±2.2 cells·mm-3). BALF cytokines were predominantly increased by ECV. Moreover, ECV caused significant changes in lung structure and function (e.g. increase in airspace by 17.5±1.4% compared to control), similar to mild tobacco smoke-induced alterations, which also could be detected in the NF ECV group, albeit to a lesser degree. In contrast, the pulmonary vasculature was not significantly affected by ECV or NF ECV. CONCLUSIONS: NF ECV components induce cell type-specific effects and mild pulmonary alterations, while inclusion of nicotine induces significant endothelial damage, inflammation and parenchymal alterations.

Effect of clinical isolate or cleavage site mutations in the SARS-CoV-2 spike protein on protein stability, cleavage, and cell-cell fusion.

The trimeric severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein (S) is the sole viral protein responsible for both viral binding to a host cell and the membrane fusion event needed for cell entry. In addition to facilitating fusion needed for viral entry, S can also drive cell-cell fusion, a pathogenic effect observed in the lungs of SARS-CoV-2-infected patients. While several studies have investigated S requirements involved in viral particle entry, examination of S stability and factors involved in S cell-cell fusion remain limited. A furin cleavage site at the border between the S1 and S2 subunits (S1/S2) has been identified, along with putative cathepsin L and transmembrane serine protease 2 cleavage sites within S2. We demonstrate that S must be processed at the S1/S2 border in order to mediate cell-cell fusion and that mutations at potential cleavage sites within the S2 subunit alter S processing at the S1/S2 border, thus preventing cell-cell fusion. We also identify residues within the internal fusion peptide and the cytoplasmic tail that modulate S-mediated cell-cell fusion. In addition, we examined S stability and protein cleavage kinetics in a variety of mammalian cell lines, including a bat cell line related to the likely reservoir species for SARS-CoV-2, and provide evidence that proteolytic processing alters the stability of the S trimer. This work therefore offers insight into S stability, proteolytic processing, and factors that mediate S cell-cell fusion, all of which help give a more comprehensive understanding of this high-profile therapeutic target.

Myeloid-cell-specific deletion of inducible nitric oxide synthase protects against smoke-induced pulmonary hypertension in mice.

BACKGROUND: Pulmonary hypertension (PH) is a common complication of COPD, associated with increased mortality and morbidity. Intriguingly, pulmonary vascular alterations have been suggested to drive emphysema development. Previously, we identified inducible nitric oxide synthase (iNOS) as an essential enzyme for development and reversal of smoke-induced PH and emphysema, and showed that iNOS expression in bone-marrow-derived cells drives pulmonary vascular remodelling, but not parenchymal destruction. In this study, we aimed to identify the iNOS-expressing cell type driving smoke-induced PH and to decipher pro-proliferative pathways involved. METHODS: To address this question we used 1) myeloid-cell-specific iNOS knockout mice in chronic smoke exposure and 2) co-cultures of macrophages and pulmonary artery smooth muscle cells (PASMCs) to decipher underlying signalling pathways. RESULTS: Myeloid-cell-specific iNOS knockout prevented smoke-induced PH but not emphysema in mice. Moreover, iNOS deletion in myeloid cells ameliorated the increase in expression of CD206, a marker of M2 polarisation, on interstitial macrophages. Importantly, the observed effects on lung macrophages were hypoxia-independent, as these mice developed hypoxia-induced PH. In vitro, smoke-induced PASMC proliferation in co-cultures with M2-polarised macrophages could be abolished by iNOS deletion in phagocytic cells, as well as by extracellular signal-regulated kinase inhibition in PASMCs. Crucially, CD206-positive and iNOS-positive macrophages accumulated in proximity of remodelled vessels in the lungs of COPD patients, as shown by immunohistochemistry. CONCLUSION: In summary, our results demonstrate that iNOS deletion in myeloid cells confers protection against PH in smoke-exposed mice and provide evidence for an iNOS-dependent communication between M2-like macrophages and PASMCs in underlying pulmonary vascular remodelling.

Role reversal of functional identity in host factors: Dissecting features affecting pro-viral versus antiviral functions of cellular DEAD-box helicases in tombusvirus replication.

Positive-stranded (+)RNA viruses greatly exploit host cells to support viral replication. However, unlike many other pathogens, (+)RNA viruses code for only a limited number of genes, making them highly dependent on numerous co-opted host factors for supporting viral replication and other viral processes during their infections. This excessive dependence on subverted host factors, however, renders (+)RNA viruses vulnerable to host restriction factors that could block virus replication. Interestingly, cellular ATP-dependent DEAD-box RNA helicases could promote or inhibit the replication of Tomato bushy stunt virus (TBSV) replication. However, it is currently unknown what features make a particular DEAD-box helicase either pro-viral or antiviral. In this work, we succeeded in reversing the viral function of the antiviral DDX17-like RH30 DEAD-box helicase by converting it to a pro-viral helicase. We also turned the pro-viral DDX3-like RH20 helicase into an antiviral helicase through deletion of a unique N-terminal domain. We demonstrate that in the absence of the N-terminal domain, the core helicase domain becomes unhinged, showing altered specificity in unwinding viral RNA duplexes containing cis-acting replication elements. The discovery of the sequence plasticity of DEAD-box helicases that can alter recognition of different cis-acting RNA elements in the viral genome illustrates the evolutionary potential of RNA helicases in the arms race between viruses and their hosts, including key roles of RNA helicases in plant innate immunity. Overall, these findings open up the possibility to turn the pro-viral host factors into antiviral factors, thus increasing the potential antiviral arsenal of the host for the benefit of agriculture and health science.

[Molecular mechanism of Spatholobi Caulis in treatment of ovarian cancer based on network pharmacology and experimental verification].

The present study explored the main active ingredients and the underlying mechanism of Spatholobi Caulisin the treatment of ovarian cancer(OC) by network pharmacology, molecular docking, and in vitro cell experiments. The active ingredients and their predicted targets(AITs) were first acquired online with the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). Theoretical disease targets(DTs) were obtained through professional databases including GeneCards, OMIM, PharmGkb, TTD, and DrugBank. The common targets in the intersection of AITs and DTs were used for the construction of a "drug-ingredient-disease-target" network by Cytoscape 3.7.1. STRING database was used to construct a protein-protein interaction(PPI) network. R 4.0.5 was used for GO and KEGG functional enrichment analyses. Schr9 dinger Maestro was used to perform and optimize the molecular docking and virtual screening.Twenty-three active ingredients of Spatholobi Caulis were screened out, involving 75 OC targets and 178 signaling pathways.Network analysis revealed that Spatholobi Caulis presumedly exerted an anti-OC effect by acting on key protein targets such as GSK-3ß, Bcl-2, and Bax. Molecular docking showed that GSK-3ß possessed goodbinding activity to prunetin. In vitro cell experiments preliminarily verified the core targets and pathways of prunetin, the active ingredient of Spatholobi Caulis against human OC SKOV3 cells.CCK-8 assay was used to detect the cell proliferation, and flow cytometry was used to detect the effect of prunetin on apoptosis of human OC SKOV3 cells.The expression of prunetin targets and related regulatory proteins was detected by Western blot.In vitro cell experiments demonstrated that prunetindisplayed significant inhibitory effects on the proliferation of OC cells and could induce apoptosis of SKOV3 cells. Western blot showed that prunetin could induce SKOV3 cell apoptosis by inhibiting GSK-3ß phosphorylation and regulating the expression of downstream Bcl-2 and Bax proteins. This study reveals the scientific nature of network pharmacology in the prediction and guidance of experimental design, confirming that prunetin can treat OC by blocking the GSK-3ß/Bcl-2/Bax cell signal transduction pathway. The findings are expected to provide a basis for the investigation of the mechanism of Spatholobi Caulis in the treatment of OC.

The effect of long-term doxycycline treatment in a mouse model of cigarette smoke-induced emphysema and pulmonary hypertension.

Chronic obstructive pulmonary disease (COPD) is a major cause of death and a still incurable disease, comprising emphysema and chronic bronchitis. In addition to airflow limitation, patients with COPD can suffer from pulmonary hypertension (PH). Doxycycline, an antibiotic from the tetracycline family, in addition to its pronounced antimicrobial activity, acts as a matrix metalloproteinase (MMP) inhibitor and has anti-inflammatory properties. Furthermore, doxycycline treatment exhibited a beneficial effect in several preclinical cardiovascular disease models. In preclinical research, doxycycline is frequently employed for gene expression modulation in Tet-On/Tet-Off transgenic animal models. Therefore, it is crucial to know whether doxycycline treatment in Tet-On/Tet-Off systems has effects independent of gene expression modulation by such systems. Against this background, we assessed the possible curative effects of long-term doxycycline administration in a mouse model of chronic CS exposure. Animals were exposed to cigarette smoke (CS) for 8 mo and then subsequently treated with doxycycline for additional 3 mo in room air conditions. Doxycycline decreased the expression of MMPs and general pro-inflammatory markers in the lungs from CS-exposed mice. This downregulation was, however, insufficient to ameliorate CS-induced emphysema or PH. Tet-On/Tet-Off induction by doxycycline in such models is a feasible genetic approach to study curative effects at least in established CS-induced emphysema and PH. However, we report several parameters that are influenced by doxycycline and use of a Tet-On/Tet-Off system when evaluating those parameters should be interpreted with caution.

Blocking tombusvirus replication through the antiviral functions of DDX17-like RH30 DEAD-box helicase.

Positive-stranded RNA viruses replicate inside cells and depend on many co-opted cellular factors to complete their infection cycles. To combat viruses, the hosts use conserved restriction factors, such as DEAD-box RNA helicases, which can function as viral RNA sensors or as effectors by blocking RNA virus replication. In this paper, we have established that the plant DDX17-like RH30 DEAD-box helicase conducts strong inhibitory function on tombusvirus replication when expressed in plants and yeast surrogate host. The helicase function of RH30 was required for restriction of tomato bushy stunt virus (TBSV) replication. Knock-down of RH30 levels in Nicotiana benthamiana led to increased TBSV accumulation and RH30 knockout lines of Arabidopsis supported higher level accumulation of turnip crinkle virus. We show that RH30 DEAD-box helicase interacts with p33 and p92pol replication proteins of TBSV, which facilitates targeting of RH30 from the nucleus to the large TBSV replication compartment consisting of aggregated peroxisomes. Enrichment of RH30 in the nucleus via fusion with a nuclear retention signal at the expense of the cytosolic pool of RH30 prevented the re-localization of RH30 into the replication compartment and canceled out the antiviral effect of RH30. In vitro replicase reconstitution assay was used to demonstrate that RH30 helicase blocks the assembly of viral replicase complex, the activation of the RNA-dependent RNA polymerase function of p92pol and binding of p33 replication protein to critical cis-acting element in the TBSV RNA. Altogether, these results firmly establish that the plant DDX17-like RH30 DEAD-box helicase is a potent, effector-type, restriction factor of tombusviruses and related viruses. The discovery of the antiviral role of RH30 DEAD-box helicase illustrates the likely ancient roles of RNA helicases in plant innate immunity.

Adenylate Kinase 4-A Key Regulator of Proliferation and Metabolic Shift in Human Pulmonary Arterial Smooth Muscle Cells via Akt and HIF-1α Signaling Pathways.

Increased proliferation of pulmonary arterial smooth muscle cells (PASMCs) in response to chronic hypoxia contributes to pulmonary vascular remodeling in pulmonary hypertension (PH). PH shares numerous similarities with cancer, including a metabolic shift towards glycolysis. In lung cancer, adenylate kinase 4 (AK4) promotes metabolic reprogramming and metastasis. Against this background, we show that AK4 regulates cell proliferation and energy metabolism of primary human PASMCs. We demonstrate that chronic hypoxia upregulates AK4 in PASMCs in a hypoxia-inducible factor-1α (HIF-1α)-dependent manner. RNA interference of AK4 decreases the viability and proliferation of PASMCs under both normoxia and chronic hypoxia. AK4 silencing in PASMCs augments mitochondrial respiration and reduces glycolytic metabolism. The observed effects are associated with reduced levels of phosphorylated protein kinase B (Akt) as well as HIF-1α, indicating the existence of an AK4-HIF-1α feedforward loop in hypoxic PASMCs. Finally, we show that AK4 levels are elevated in pulmonary vessels from patients with idiopathic pulmonary arterial hypertension (IPAH), and AK4 silencing decreases glycolytic metabolism of IPAH-PASMCs. We conclude that AK4 is a new metabolic regulator in PASMCs interacting with HIF-1α and Akt signaling pathways to drive the pro-proliferative and glycolytic phenotype of PH.

Lung developmental arrest caused by PDGF-A deletion: consequences for the adult mouse lung.

PDGF-A is a key contributor to lung development in mice. Its expression is needed for secondary septation of the alveoli and deletion of the gene leads to abnormally enlarged alveolar air spaces in mice. In humans, the same phenotype is the hallmark of bronchopulmonary dysplasia (BPD), a disease that affects premature babies and may have long lasting consequences in adulthood. So far, the knowledge regarding adult effects of developmental arrest in the lung is limited. This is attributable to few follow-up studies of BPD survivors and lack of good experimental models that could help predict the outcomes of this early age disease for the adult individual. In this study, we used the constitutive lung-specific Pdgfa deletion mouse model to analyze the consequences of developmental lung defects in adult mice. We assessed lung morphology, physiology, cellular content, ECM composition and proteomics data in mature mice, that perinatally exhibited lungs with a BPD-like morphology. Histological and physiological analyses both revealed that enlarged alveolar air spaces remained until adulthood, resulting in higher lung compliance and higher respiratory volume in knockout mice. Still, no or only small differences were seen in cellular, ECM and protein content when comparing knockout and control mice. Taken together, our results indicate that Pdgfa deletion-induced lung developmental arrest has consequences for the adult lung at the morphological and functional level. In addition, these mice can reach adulthood with a BPD-like phenotype, which makes them a robust model to further investigate the pathophysiological progression of the disease and test putative regenerative therapies.

Multi-walled carbon-nanotube-decorated tungsten ditelluride nanostars as anode material for lithium-ion batteries.

Multi-walled carbon-nanotube (MWCNT)-decorated WTe2 nanostars (WTe2@CNT nanocomposites) are to be employed for the first time as anode candidates in the development of lithium-ion (Li-ion) batteries. WTe2@CNT nanocomposites deliver a high discharge capacity of 1097, 475, 439, 408, 395 and 381 mA h g-1 with an increasing current density of 100, 200, 400, 600, 800 and 1000 mA g-1, respectively, while WTe2 nanostars exhibit a reversible capacity of 655, 402, 400, 362, 290 and 197 mA h g-1 with the aforementioned current densities. Furthermore, WTe2@CNT nanocomposites exhibit a superior reversible capacity of 592 mA h g-1 at 500 mA g-1 with a capacity retention of 100% achieved over 500 cycles, while bare WTe2 nanostars deliver ∼85 mA h g-1 over 350 cycles. This remarkable Li cycling performance is attributed to MWCNTs interconnected with WTe2 nanostars. In addition, the exposed active interlayers of the WTe2 nanostars, which are responsible for maintaining the structural integrity of the electrodes, buffer the large volume expansion within the WTe2 nanostars, avoiding the agglomeration of the particles. The layered WTe2 nanostars were synthesized via the solution-phase method, and present extremely good possibilities for the scaling-up of Li-ion battery storage systems.

Persistent development of adomavirus and aquareovirus in a novel cell line from marbled eel with petechial skin haemorrhage.

In Taiwan, a petechial haemorrhage disease associated with mortality has affected marbled eels (Anguilla marmorata). The eels were revealed to be infected with adomavirus (MEAdoV, previously recognized as a polyoma-like virus). In this study, cell line DMEPF-5 was established from the pectoral fin of a diseased eel. DMEPF-5 was passaged >70 times and thoroughly proliferated in L-15 medium containing 2%-15% foetal bovine serum at 20-30°C. Transcripts of neural cell adhesion molecule 1 and nestin genes, and nucleic acids of MEAdoV and a novel reovirus (MERV) in the cells were demonstrated by reverse transcription-polymerase chain reaction analysis. Phylogenetic analysis revealed that the AdoV LO8 proteins mostly relate to adenovirus adenain, whereas MERV is close to American grass carp reovirus in Aquareovirus G, based on a partial VP2 nucleotide sequence. DMEPF-5 cells are susceptible to additional viral infection. Taken together, the marbled eels with the haemorrhagic disease have coinfection with MEAdoV and MERV, and the pathogenic role of MEAdoV and MERV warrants research. DMEPF-5 has gene expression associated with mesenchymal stem and progenitor cells and is the first cell line persistently infected with adomavirus and aquareovirus. DMEPF-5 can facilitate studies of such viruses and haemorrhagic disease.

Synthesis and two-photon absorption property characterizations of small dendritic chromophores containing functionalized quinoxaliniod heterocycles.

A series of star-shaped multi-polar chromophores (compounds 1-3) containing functionalized quinoxaline and quinoxalinoid (indenoquinoxaline and pyridopyrazine) units has been synthesized and characterized for their two-photon absorption (2PA) properties both in the femtosecond and the nanosecond time domain. Under our experimental conditions, these model fluorophores are found to manifest strong and wide-dispersed two-photon absorption in the near-infrared region. It is demonstrated that molecular structures with multi-branched π frameworks incorporating properly functionalized quinoxalinoid units would possess large molecular nonlinear absorptivities within the studied spectral range. Effective optical-power attenuation and stabilization behaviors in the nanosecond time domain of a selected representative dye molecule (i.e., compound 2) from this model compound set were also investigated and the results indicate that such structural motif could be a useful approach for the molecular design toward strong two-photon-absorbing material systems for quick-responsive and broadband optical-suppressing-related applications, particularly to confront long laser pulses.

The implantable and biodegradable PHBHHx 3D scaffolds loaded with protein-phospholipid complex for sustained delivery of proteins.

PURPOSE: PHBHHx (poly(3-hydroxybutyrate-co-3-hydroxyhexanoate)) is an excellent biomaterial for tissue repair. Here, we aim to develop a PHBHHx-based three-dimensional (3D) scaffold system for sustained delivery of proteins (insulin serves as a model protein). METHODS: The insulin-phospholipid complex (INS-PLC) was prepared to enhance the insulin lipophilicity. INS-PLC loaded PHBHHx 3D scaffolds (INS-PLC-SCAs) containing PEG-2000 were fabricated by lyophilization. In vitro release was performed in the medium with or without lipase. The bioactivity of INS-PLC-SCAs was measured in diabetic rats. RESULTS: In vitro release shows that the release rate of INS-PLC-SCAs was very slow (~6% of total insulin was released within 120 days), and PEG-2000 or lipase had no effect on its release pattern. The bioactivity test shows that the hypoglycaemic effect of insulin was maintained after formulated into scaffolds. After subcutaneous (s.c.) implantation, its therapeutic effect lasted for over 130 h, and its bioavailability was enhanced by 4-fold. CONCLUSIONS: PHBHHx based 3D scaffold has a great potential for sustained delivery of proteins, especially growth factors. When growth factors are incorporated, it can serve as a bifunctional system that provides a porous skeleton for cells attachment and proliferation, as well as a matrix for long term release of the loaded growth factors.

Effect of mask coverage on face identification in Taiwanese men and women.

Mask wearing is the easiest and most effective way to avoid COVID-19 infection; however, it affects interpersonal activities, especially face identification. This study examined the effects of three mask coverage levels (full coverage, FC; coverage up to the middle [MB] or bottom of the nose bridge [BB]) on face identification accuracy and time. A total of 115 university students (60 men and 55 women) were recruited to conduct a computer-based simulation test consisting of 30 questions (10 questions [five face images each of men and women] for the three mask coverage levels). One unmasked target face and four face images with a specified mask coverage level were designed for each question, and the participants were requested to select the same face from the four covered face images on the basis of the target face. The ANOVA results indicated that identification accuracy was significantly affected by sex (p < 0.01) and the mask coverage level (p < 0.001), whereas identification time was only influenced by sex (p < 0.05). The multiple comparison results indicated that the identification accuracy rate for faces wearing a mask with FC (90.3%) was significantly lower than for those wearing masks with coverage up to the MB (93.7%) and BB (94.9%) positions; however, no difference in identification accuracy rate was observed between the MB and BB levels. Women exhibited a higher identification accuracy rate than men (94.1% vs. 91.9%) in identifying unfamiliar faces, even though they may spend less time identifying the images. A smaller mask coverage level (i.e., the BB level) does not facilitate face identification. The findings can be served as a reference for people to trade-off between wearing a mask and interpersonal interaction in their daily activities.

Imaging analysis reveals budding of filamentous human metapneumovirus virions and direct transfer of inclusion bodies through intercellular extensions.

IMPORTANCE: Human metapneumovirus is an important respiratory pathogen that causes significant morbidity and mortality, particularly in the very young, the elderly, and the immunosuppressed. However, the molecular details of how this virus spreads to new target cells are unclear. This work provides important new information on the formation of filamentous structures that are consistent with virus particles and adds critical new insight into the structure of extensions between cells that form during infection. In addition, it demonstrates for the first time the movement of viral replication centers through these intercellular extensions, representing a new mode of direct cell-to-cell spread that may be applicable to other viral systems.

Toward controllable and predictable synthesis of high-entropy alloy nanocrystals.

High-entropy alloy (HEA) nanocrystals have attracted extensive attention in catalysis. However, there are no effective strategies for synthesizing them in a controllable and predictable manner. With quinary HEA nanocrystals made of platinum-group metals as an example, we demonstrate that their structures with spatial compositions can be predicted by quantitatively knowing the reduction kinetics of metal precursors and entropy of mixing in the nanocrystals under dropwise addition of the mixing five-metal precursor solution. The time to reach a steady state for each precursor plays a pivotal role in determining the structures of HEA nanocrystals with homogeneous alloy and core-shell features. Compared to the commercial platinum/carbon and phase-separated counterparts, the dendritic HEA nanocrystals with a defect-rich surface show substantial enhancement in catalytic activity and durability toward both hydrogen evolution and oxidation. This quantitative study will lead to a paradigm shift in the design of HEA nanocrystals, pushing away from the trial-and-error approach.

Proprietary Medicines Containing Bupleurum chinense DC. (Chaihu) for Depression: Network Meta-Analysis and Network Pharmacology Prediction.

Background and Aims: The rapid development of society has resulted in great competitive pressures, leading to the increase in suicide rates as well as incidence and recurrence of depression in recent years. Proprietary Chinese medicines containing Bupleurum chinense DC. (Chaihu) are widely used in clinical practice. This study aimed at evaluating the efficacy and safety of oral proprietary Chinese medicines containing Chaihu for treating depression by network meta-analysis (NMA) and exploring the potential pharmacological mechanisms of the optimal drugs obtained based on NMA. Methods: This study searched for clinical randomized controlled trial studies (RCTs) about Chaihu-containing products alone or in combination with selective serotonin reuptake inhibitors (SSRI), serotonin-norepinephrine reuptake inhibitors (SNRI), and cyclic antidepressants (CAS) for depression in eight databases. The search deadline is from data inception to April 2021. For efficacy assessment, the clinical response rate, the Hamilton Depression Scale-17 (HAMD-17), and adverse reactions were calculated. The methodological quality of the included studies was assessed for risk of bias following the Cochrane Handbook for Systematic Reviews of Interventions, and the data were subjected to NMA via the Stata version 16.0 software. Subsequently, the optimal drug obtained from the NMA results, Danzhi Xiaoyao pill (DZXY), was used to conduct network pharmacology analysis. We searched databases to acquire bioactive and potential targets of DZXY and depression-related targets. The protein-protein interaction (PPI) network, component-target network, the Gene Ontology (GO), and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were performed by the STRING database, Cytoscape 3.9.0 software, and R version 4.1.2, respectively. Results: Thirty-seven RCTs, with a total of 3,263 patients, involving seven oral proprietary Chinese medicines containing Chaihu, were finally included. The results of the NMA demonstrated that the top four interventions with the best efficiency were Jiawei Xiaoyao + SSRI, DZXY + SNRI, Xiaoyao pill + SSRI, and Jieyu pill + SNRI; the top four interventions reducing HAMD score were DZXY + SNRI, Jiawei Xiaoyao, Jieyu pill, and Puyu pill + SNRI; the top four interventions with the least adverse effects were Jieyu pill, Anle pill + SSRI, DZXY + SNRI, and Puyu pill + SNRI. In the aspects above, DZXY + SNRI performed better than other treatments. After network meta-analysis, we conducted a network pharmacology-based strategy on the optimal drugs, DZXY, to provide the pharmacological basis for a conclusion. A total of 147 active compounds and 248 targets in DZXY were identified, of which 175 overlapping targets related to depression. Bioinformatics analysis revealed that MAPK3, JUN, MAPK14, MYC, MAPK1, etc. could become potential therapeutic targets. The MAPK signaling pathway might play an essential role in DZXY against depression. Conclusion: This is the very first systematic review and network meta-analysis evaluating different oral proprietary Chinese medicines containing Chaihu in depressive disorder. This study suggested that the combination of proprietary Chinese medicines containing Chaihu with antidepressants was generally better than antidepressant treatment. The incidence of adverse reactions with antidepressants alone was higher than that with proprietary Chinese medicines containing Chaihu alone or in combination with antidepressants. DZXY + SNRI showed significantly better results in efficacy, HAMD scores, and safety. The antidepressant effect of DZXY may be related to its regulation of neuroinflammation and apoptosis.