Multiple cell death modalities and immune response in pulpitis.

AIM: To investigate the level and distribution of apoptosis, pyroptosis, necroptosis, and NETosis in pulpitis with or without necrosis on a basis of histological classification. Additionally, to examine the effect of pulpitis with necrosis (PWN) on the number and activation of peripheral and bone marrow (BM) neutrophils, as well as spleen lymphocytes, in a mouse model of pulpitis. METHODOLOGY: The material comprised 20 permanent teeth, with or without caries, which were classified into three histological categories based on the distribution of inflammatory cells and the presence or absence of necrosis: (i) healthy pulp (HP), (ii) pulpitis without necrosis (PWON), and (iii) PWN. The levels of the four regulated cell death (RCD) pathways were detected by immunohistochemical and immunofluorescent staining with specific markers: apoptosis (caspase-8, cleaved caspase-3), pyroptosis (cleaved caspase-1, membrane-binding gasdermin D), necroptosis (receptor-interacting kinase 3, phosphorylated MLKL), and NETosis (myeloperoxidase, citrullinated histone H3). Acute pulpitis was induced in C57BL/6J mice via pulp exposure, and the mice were divided into four groups: (i) control (no tooth preparation, n = 6), (ii) Day 1 (sacrificed at 1 day after pulp exposure, n = 3), (iii) Day 3 (n = 3), and (iv) Day 5 (n = 7). The control and Day 5 groups were used for further immunofluorescent analysis to assess the levels of RCD and flow cytometry to monitor the changes in peripheral and BM neutrophils, as well as spleen lymphocytes. Human dental pulp stem cells (hDPSCs) were isolated and cultured from extracted healthy third molars. Apoptosis and necroptosis in hDPSCs were induced by staurosporine, whilst pyroptosis was induced by lipopolysaccharide and nigericin. One-way analysis of variance (ANOVA) with Tukey's test, Welch's ANOVA with Tamhane's test, and Student's t-tests were used to compare immunohistochemical labelling and flow cytometric data amongst groups (p < .05). RESULTS: The pulpal tissue of PWN can be divided into the abscess core (PWN-AC) and fibrous tissue (PWN-FT). The ratio of total necrotic cells (TUNEL-positive) in PWN-AC was significantly higher than in PWN-FT and PWON (both p < .01). Compared with HP, the expression levels of markers for apoptosis and pyroptosis were increased in PWON, whilst the expression levels of markers for apoptosis, pyroptosis, and NETosis were elevated in PWN, primarily detected in PWN-AC. Interestingly, myeloperoxidase (MPO) was exclusively observed in PWN-AC, with minimal detection in PWN-FT and PWON. Additionally, the frequency of MPO+ cells was significantly higher than that of MB-GSDMD+ cells and Cl-cas3+ cells in PWN-AC (both p < .01). Histological observation and TUNEL staining showed abundant necrotic cells in mouse pulpal tissue after pulp exposure, indicating a simulation of human PWN. In mouse pulpitis tissue, markers of apoptosis, pyroptosis, and NETosis were detected. In vitro, various cell deaths including apoptosis, pyroptosis, and necroptosis were also triggered in hDPSCs under various cell death treatments. Furthermore, in terms of systemic changes, pulp exposure-induced pulpitis could increase the number (p < .05) and cellular activity (p < .01) of neutrophils from BM in a mouse model. No significant changes in peripheral blood neutrophils, spleen T cells, B cells, or the CD4/CD8 ratio were detected between the control and pulpitis mice. CONCLUSIONS: Our findings uncover distinct patterns of mixed cell death at different histological stages of human pulpitis and the impact of pulpitis on the number and activity of BM neutrophils. Notably, NETosis occurs specifically and predominates in the abscess area of pulpitis, suggesting a potential effect of neutrophil extracellular traps (NETs) on pulpitis progression and NETs-targeted diagnostic strategy may play a role in decision making for vital pulp therapy.

Genomic Analysis of a Novel Torradovirus "Rehmannia Torradovirus Virus": Two Distinct Variants Infecting Rehmannia glutinosa.

Rehmannia glutinosa, a crucial medicinal plant native to China, is extensively cultivated across East Asia. We used high-throughput sequencing to identify viruses infecting R. glutinosa with mosaic, leaf yellowing, and necrotic symptoms. A novel Torradovirus, which we tentatively named "Rehmannia torradovirus virus" (ReTV), was identified. The complete sequences were obtained through reverse-transcription polymerase chain reaction (RT-PCR), 5' and 3' rapid amplification of cDNA ends, and Sanger sequencing. The amino acid sequence alignment between the ReTV-52 isolate and known Torradovirus species in the Pro-Pol and coat protein regions were 51.3-73.3% and 37.1-68.1%, respectively. Meanwhile, the amino acid sequence alignment between the ReTV-8 isolate and known Torradovirus species in the Pro-Pol and coat protein regions were 52.7-72.8% and 36.8-67.5%, respectively. The sequence analysis classified ten ReTV strains into two variants. The ReTV-52 genome has two RNA segments of 6939 and 4569 nucleotides, while that of ReTV-8 consists of two RNA segments containing 6889 and 4662 nucleotides. Sequence comparisons and phylogenetic analysis showed ReTV strains clustered within the Torradovirus, exhibiting the closet relation to the squash chlorotic leaf spot virus. The RT-PCR results showed a 100% ReTV detection rate in all 60 R. glutinosa samples. Therefore, ReTV should be classified as a novel Torradovirus species. ReTV is potentially dangerous to R. glutinosa, and necessitating monitoring this virus in the field.

Design, synthesis and structure-activity relationships of novel non-ketolides: 9-Oxime clarithromycin featured with seven-to thirteen-atom-length diamine linkers at 3-OH.

We report here on the structure-activity relationships of hybrids combining 3-descladinosyl clarithromycin with quinolones linked by extended diamine connectors. Several hybrids, exemplified by 23Bc, 23Be, 23Bf, 26Be, and 30Bc, not only restored potency against inducibly resistant pathogens but also exhibited significantly enhanced activities against constitutively resistant strains of Staphylococcus pneumoniae and Staphylococcus pyogenes, which express high-level resistance independent of clarithromycin or erythromycin induction. Additionally, the novel hybrids showed susceptibility against Gram-negative Haemophilus influenzae. Notably, hybrid 23Be demonstrated dual modes of action by inhibiting both protein synthesis and DNA replication in vitro and in vivo. Given these promising characteristics, 23Be emerges as a potential candidate for the treatment of community-acquired bacterial pneumonia.

Cerebral Endothelial CXCR2 Promotes Neutrophil Transmigration into Central Nervous System in LPS-Induced Septic Encephalopathy.

Septic encephalopathy (SE) represents a severe inflammatory syndrome linked to elevated septic mortality rates, lacking specific therapeutic interventions, and often resulting in enduring neurological sequelae. The present investigation endeavors to elucidate the involvement of C-X-C Motif Chemokine Receptor 2 (CXCR2) in the pathogenesis of SE and to explore the potential of CXCR2 modulation as a therapeutic avenue for SE. Employing a murine SE model induced by lipopolysaccharide (LPS) administration, CXCR2 knockout mice and the CXCR2 inhibitor SB225002 were utilized to assess neutrophil recruitment, endothelial integrity, and transendothelial migration. Our findings substantiate that either CXCR2 deficiency or its inhibition curtails neutrophil recruitment without impacting their adhesion to cerebral endothelial cells. This phenomenon is contingent upon endothelial CXCR2 expression rather than CXCR2's presence on neutrophils. Furthermore, the CXCR2 blockade preserves the integrity of tight junction protein ZO-1 and mitigates F-actin stress fiber formation in cerebral endothelial cells following septic challenge. Mechanistically, CXCL1-mediated CXCR2 activation triggers cerebral endothelial actin contraction via Rho signaling, thereby facilitating neutrophil transmigration in SE. These observations advocate for the potential therapeutic efficacy of CXCR2 inhibition in managing SE.

The Microbiome Protocols eBook initiative: Building a bridge to microbiome research.

The Microbiome Protocols eBook (MPB) serves as a crucial bridge, filling gaps in microbiome protocols for both wet experiments and data analysis. The first edition, launched in 2020, featured 152 meticulously curated protocols, garnering widespread acclaim. We now extend a sincere invitation to researchers to participate in the upcoming 2nd version of MPB, contributing their valuable protocols to advance microbiome research.

PTPRO inhibits LPS-induced apoptosis in alveolar epithelial cells.

Acute lung injury (ALI) and its severe manifestation, acute respiratory distress syndrome (ARDS), represent critical clinical syndromes with multifactorial origins, notably stemming from sepsis within intensive care units (ICUs). Despite their high mortality rates, no selective cure is available beside ventilation support. Apoptosis plays a complex and pivotal role in the pathophysiology of acute lung injury. Excessive apoptosis of alveolar epithelial and microvascular endothelial cells can lead to disruption of lung epithelial barrier integrity, impairing the body's ability to exchange blood and gas. At the same time, apoptosis of damaged or dysfunctional cells, including endothelial and epithelial cells, can help maintain tissue integrity and accelerate recovery from organ pro-inflammatory stress. The balance between pro-survival and pro-apoptotic signals in lung injury determines patient outcomes, making the modulation of apoptosis an area of intense research in the quest for more effective therapies. Here we found that protein tyrosine phosphatase receptor type O (PTPRO), a poorly understood receptor-like protein tyrosine phosphatase, is consistently upregulated in multiple tissue types of mice under septic conditions and in the lung alveolar epithelial cells. PTPRO reduction by its selective short-interfering RNA (siRNA) leads to excessive apoptosis in lung alveolar epithelial cells without affecting cell proliferation. Consistently PTPRO overexpression by a DNA construct attenuates apoptotic signaling induced by LPS. These effects of PTPTO on cellular apoptosis are dependent on an ErbB2/PI3K/Akt/NFκB signaling pathway. Here we revealed a novel regulatory pathway of cellular apoptosis by PTPRO in lung alveolar epithelial cells during sepsis.

First Report of Aspergillus westerdijkiae Causing Leaf Blight on Rehmannia glutinosa in China.

Rehmannia glutinosa (also known as Chinese foxglove) is a perennial dicotyledonous herb, which plays an important role in traditional Chinese medicine. Its active ingredients have a wide range of pharmacological effects on the blood system, endocrine system, immune system, cardiovascular system, and nervous system (Zhang et al. 2008). In May 2022, leaf blight was observed on 45-day-old R. glutinosa in a seedling nursery in Jiaozuo City (35°01'44.20″N, 113°05'30.63″E), Henan Province, China with an approximate disease incidence up to 54% (~1,300 plants). Irregular brown lesion initially appeared on the tips of basal leaves, then progressed to the entire leaf causing leaf drying out (Supple. Fig. 1-A, B, C). The same symptoms appeared successively in the leaves from the base to the top of the plant, which eventually caused the whole plant to die. To identify the pathogen, eight symptomatic leaves were randomly collected from eight individual plants, and cut into small pieces (5 × 5 mm) at the border of lesions. The pieces were surface disinfected in 75% ethanol for 15 s, followed by 1% NaClO for 1 min, rinsed in sterile water three times, and placed on potato dextrose agar (PDA) medium in the dark for 3 days at 25℃. Finally, 12 purified isolates (DHY1-DHY12) were obtained by using single spore method. Leaves of R. glutinosa seedlings were inoculated with conidial suspension (106 conidia/ml), three plants were inoculated per isolate. Controls were treated with sterilized water. All inoculated and control plants were incubated in a greenhouse at 25℃ under 80 ± 10% humidity and a 8-h/16-h dark/light cycle. This experiment was repeated three times. After 5 days, similar symptoms to those of diseased leaves in the seedling nursery appeared on leaves inoculated with DHY4-DHY10, while plants inoculated with DHY1-DHY3, DHY11-DHY12, and the controls remained asymptomatic (Supple. Fig.1-D, E). The same fungi were re-isolated from diseased leaves, fulfilling Koch's postulates. The causal agents DHY4 to DHY10, showed similar morphology, which were morphologically identified as Aspergillus sp. (Visagie et al. 2014). Isolate DHY5 was selected for further study. On PDA plates, the colonies were covered with white velutinous mycelia (Supple. Fig.1-F). Conidia were ochre yellow and outwards concentric circles. Vesicles were globose, and about 20.1-26.6 µm in diameter (Supple. Fig.1-G). Conidiophore stipes were smooth walled and hyaline, with conidial heads radiating. The conidia were light yellow to orange, exudate clear to orange droplets. The conidia were (2.53-3.25) µm × (2.58-3.47) µm in diameter (n=50) (Supple. Fig.1-H). For further molecular identification, the ITS and TUB gene sequences were amplified with primer pairs ITS1/ITS4 and BT2a/BT2b (Glass and Donaldson. 1995), respectively. BLASTn searches of the ITS (PP355445) and TUB (PP382788) sequences showed 100% and 98.42% similarity to those of A. westerdijkiae (OP237108 and OP700424), respectively. Phylogenetic analysis based on the concatenated sequences of ITS and TUB confirmed that the fungus was A. westerdijkiae, (Supple. Fig.2). A. westerdijkiae was mainly reported on its secondary metabolite ochratoxin A contamination of agricultural products, fruits, and various food products, such as coffee beans (Alvindia et al 2016), grapes (Díaz et al. 2009), oranges and fruit juice (Marino et al. 2009), etc. To our knowledge, this is the first report of A. westerdijkiae causing leaf blight on R. glutinosa in China.

Genomic Characterization and Molecular Detection of Rehmannia Allexivirus Virus, a Novel Allexivirus Infecting Rehmannia glutinosa.

Rehmannia glutinosa is one of the most important medicinal plants in China and is affected by viral diseases. In this study, a new virus tentatively named Rehmannia Allexivirus virus (ReAV) was identified through high-throughput sequencing, reverse-transcription polymerase chain reaction (RT-PCR), and Sanger sequencing. The complete genome length was 7297 nt and it contained five open reading frames (ORFs) encoding replicase, triple gene block 1(TGB1), TGB2, TGB3, and coat protein (CP). The replicase and CP presented nucleotide homology ranges of 59.9-65.2% and 47.5-55.5% between the nine ReAV isolates and the other 12 species of the genus Allexivirus. In the nine isolates, ReAV-20 and ReAV-31 isolates showed breakpoints in the replicase and CP regions, respectively. The other isolates shared 87.2-96.5% nt with the whole genome nucleotide identity. The phylogenetic tree showed that seven ReAV isolates based on replicase, CP, and whole genome sequences were clustered in the same branch and were related to the genus Allexivirus. The ReAV detection rates for 60 R. glutinosa samples were 73.3-81.7% through RT-PCR using primers targeting the replicase or CP genes. These results demonstrate that ReAV is the dominant virus in R. glutinosa. This study provides important evidence for understanding viruses infecting R. glutinosa and for establishing efficient strategies to prevent viral spread.

Cadmium exposure activates mitophagy through downregulating thyroid hormone receptor/PGC1α signal in preeclampsia.

Gestational cadmium exposure increases the risk of preeclampsia. Placenta mitophagy was activated in preeclampsia. The aim of present study was to explore the mechanism of cadmium-induced mitophagy activation and its association with preeclampsia. Mitophagy markers expression levels were detected by quantitative real-time PCR, Western blot, immunofluorescence and immunochemistry in preeclampsia placenta. JEG3 cells were treated with CdCl2, iopanoic acid (IOP), 3-methyladenine and PGC1α SiRNA to verify mechanism of cadmium-induced mitophagy. Mitophagy marker LC3BII/I and P62 expression were increased and mitochondrial membrane receptor protein TOM20 and FUNDC1 expression were decreased in preeclampsia placenta as compared with that in normotension control. Mitophagy marker LC3BII/I and P62 expression were increased and TOM20 and FUNDC1 expression was decreased in CdCl2-treated JEG3 cells. Meanwhile, mitochondrial biogenesis regulator, PGC1α expression was decreased in preeclampsia and CdCl2-treated JEG3 cells. The expressions of LC3B and P62 were increased and the expressions of TOM20, FUNDC1 and PGC1α were decreased in IOP-treated cell. PGC1α SiRNA transfection led to increased expression of LC3BII/I and P62 and decreased expression of TOM20 and FUNDC1. The expression of sFlt1 was increased in preeclampsia placenta, CdCl2-treated cells, in IOP-treated cells and in PGC1α SiRNA transfected cells. 3-methyladenine treatment protected the increased expression of sFlt1 in CdCl2-treated cells, in IOP-treated cells and in PGC1α SiRNA transfected cells. Meanwhile, co-treatment of cadmium and IOP or PGC1αSiRNA led to a reduce expressions of OPA1, MFN1, MFN2 and FUNDC1 as compared to cadmium-treated, IOP-treated and PGC1α SiRNA-treated cells. These results elucidated that maternal cadmium exposure activated placenta mitophagy through downregulation of thyroid hormone receptor signal mediated decreased expression of PGC1α and was associated with the occurrence of preeclampsia.

Clinical Manifestation, Risk Factors, and Immune Checkpoint Inhibitor Rechallenge of Checkpoint Inhibitor-Associated Pneumonitis in Patients With Lung Cancer.

SUMMARY: Immune-related adverse effects can lead to damage to various systems of the body, checkpoint inhibitor-associated pneumonitis (CIP) is one of the potentially lethal immune-related adverse effects. However, evidence regarding the risk factors associated with CIP is limited. To timely and accurate identification and prompt treatment of CIP, understanding the risk factors for multimorbidity among diverse study populations becomes crucial. We retrospectively analyzed the clinical data of 1131 patients with lung cancer receiving immunotherapy to identify 110 patients with CIP, the clinical characteristics and radiographic features of patients with CIP were analyzed. A case-control study was subsequently performed to identify the risk factors of CIP. The median treatment cycle was 5 cycles and the median time to onset of CIP was 4.2 months. CIP was mainly grade I or II. Most cases improved after discontinuation of immune checkpoint inhibitors (ICIs) or hormone therapy. Severe CIP tended to occur earlier in comparison to mild to moderate cases. The recurrence rate was 20.6% in ICI-rechallenged patients, and patients with relapsed CIP were usually accompanied by higher-grade adverse events than at first onset. Among the 7 patients with relapse, ICI-associated deaths occurred in 2 patients (28.6%). For rechallenging with ICIs after recovery from CIP, caution should be practiced. Male [odds ratio (OR): 2.067; 95% CI: 1.194-3.579; P = 0.009], history of chest radiation (OR: 1.642; 95% CI: 1.002-2.689; P = 0.049) and underlying lung disease (OR: 2.347; 95% CI: 1.008-5.464; P =0.048) was associated with a higher risk of CIP.

First report of basal stem rot caused by three Fusarium species on Chinese yam in China.

The cultivated variety of Chinese yam (Dioscorea polystachya Turcz. cv. Tiegun) is an economically important plant, capable of producing tubers that are used as food and traditional Chinese medicine. The basal stem rot was found on approximately 65% of yam (tuber expansion stage) in a total of 10 ha field in Wuzhi, Wen, and Hua counties, Henan, China (Sep 2021). Dark brown fusiform lesions initially occurred at the stems basal, irregularly extending to join together and leading to loop-stem necrotic indentation. Three diseased samples from Wuzhi county were collected, cut into 5 × 5 mm pieces, surface sterilized in 75% ethanol (30 s) and 1% NaClO (1 min), washed in sterile water 3 times, and placed on PDA in the dark for 3 days at 28℃. A total of 44 isolates forming three groups of Fusarium colonies were obtained using monosporic isolation, of which 19, 8, and 17 isolates were identified as F. oxysporum, F. solani, and F. proliferatum based on colony morphology, respectively. Typical isolates SYJJ6, 9, and 10 for each group were further studied. The SYJJ6 colonies showed gray white abundant fluffy aerial mycelium with rough edges, formation of ellipsoid, unicellular microconidia without septa, 5.6 to 13.4 × 2.4 to 4.7 µm (n = 50), and sickle-shaped, slightly curved macroconidia with 2 to 4 septa, 14.0 to 23.9 × 3.4 to 5.1 µm (n = 50). Isolate SYJJ9 produced flocculent white colonies, grew in a circular pattern with a sharp edge, forming oval or oblong microconidia with zero or one septum, 11.2 to 18.8 × 3.4 to 6.2 µm (n = 50), and slightly curved macroconidia with 2 to 3 septa, 27.6 to 44.0 × 3.9 to 7.4 µm (n = 50). SYJJ10 produced whitish or pinkish white colonies with fluffy aerial mycelium and a red pigmentation, produced renal or oval microconidia with no septa, 5.1 to 11.8 × 1.8 to 4.2 µm (n = 50), and falcate, slightly curved macroconidia with 3 to 4 septa, 16.1 to 30.2 × 3.1 to 5.9 µm (n = 50). Additionally, TUB, EF-1α, and RPB2 genes were amplified with primers BT2a/BT2b, EF1/EF2, and 5f2/-7cr, respectively (Glass and Donaldson 1995; O'Donnell et al. 1998, 2010). BLASTn analysis on SYJJ6 (OR047663, OR047666, OR047669), SYJJ9 (OR047665, OR047667, OR047670), and SYJJ10 (OR047664, OR047668, OR047671) gene sequences were over 99% identical to those of F. oxysporum (100%, MK432917; 100%, MN417196; 99.61%, MN457531), F. solani (100%, MF662662; 100%, MN223440; 99.80%, CP104055), and F. proliferatum (100%, ON557521; 100%, ON458137; 99.90%, LT841266), respectively. Pathogenicity tests of three isolates were separately performed on 60-day-old yam seedlings. The basal stems were wounded using needle, and the wounds were wrapped with cotton balls soaked with conidial suspension (1 mL, 3×106 conidia/mL) or water (control). Each isolate treated three plants and repeated three times. All plants were grown at 28℃ under a 16/8-h light/dark cycle. Typical symptoms emerged on basal stems at 16, 13, and 17 days after inoculation with the conidia of isolates SYJJ6, 9, and 10, while the control basal stems appeared healthy. The re-isolated fungi were identical to the original three isolates. Fusarium species (F. oxysporum, F. commune, F. humuli, etc.)were previously reported to cause wilt or stem rot on different D. polystachya cultivars (Fang et al. 2020; Li et al. 2023; Zhao et al. 2013), or basal stem rot on Panax ginseng (Ma et al. 2020). This is the first report of Chinese yam basal stem rot caused by Fusarium species, which threatens the production of Chinese yam 'Tiegun' and should be further studied.

A Qualitative Inquiry into the Challenges in Overcoming Heroin Addiction Among Chinese Individuals Who Experience Voluntary Rehabilitation.

This study explores the intentions and experiences of individuals seeking abstinence from heroin use through voluntary rehabilitation, as well as challenges to achieving a positive outcome. This study used semi-structured interviews with 37 individuals (22 female) who used heroin and utilized rational choice theory to guide data analysis. This study found that participants tried both medical and non-medical treatment methods but faced structural and personal barriers that hindered their efforts. While ceasing heroin use would presumably benefit their health and family relationships, individuals perceived the physical, psychological, and social costs of abstinence as outweighing the potential benefits and ultimately resumed heroin use. More targeted voluntary rehabilitation programs addressing specific structural and personal barriers are therefore recommended to benefit Chinese people seeking abstinence from drug use.

Absolute Stimulated Raman Cross Sections of Molecules.

Stimulated Raman scattering (SRS) is a fundamental optical process that was discovered more than 60 years ago. While the early SRS spectroscopy studies have provided valuable insights into materials systems, the advent of SRS microscopy has launched a rapidly growing field in biological imaging. However, a fundamental understanding of the molecular response under SRS is still lacking. Herein we present a new framework to introduce molecule-intrinsic stimulated Raman scattering cross sections, σSRS, in the unit of Göppert-Mayer (GM). The absolute SRS cross sections determined for real molecular systems challenge the conventional wisdom that Raman spectroscopy is always a weak process. The enormous rate acceleration of SRS, captured by an apparent SRS cross section, stems from a synergistic effect between the field and the molecule. Our new framework goes beyond the conventional optics-centric view and presents a molecule-inclusive perspective, thus offering a comprehensive foundation for the future growth of SRS spectroscopy and microscopy.

[CXCL1 induces the contraction of endothelial cytoskeleton and increases permeability in mouse cerebral endothelium bEND.3 cells by promoting protein kinase B phosphorylation].

Objective To investigate the effects of C-X-C motif chemokine ligand 1 (CXCL1) and its receptor CXCR2 on the cerebral endothelial cytoskeleton rearrangement and permeability in the inflammation of septic encephalopathy. Methods The murine model of septic encephalopathy was established by intraperitoneal injection of LPS (10 mg/kg). The levels of TNF-α and CXCL1 in the whole brain tissue were detected by ELISA. The expression of CXCR2 was detected by Western blot analysis after bEND.3 cells were stimulated with 500 ng/mL LPS and 200 ng/mL TNF-α. After treated with CXCL1(150 ng/mL), the changes of endothelial filamentous actin (F-actin) rearrangement in bEND.3 cells were observed by immuno-fluorescence staining. In the cerebral endothelial permeability test, bEND.3 cells were randomly divided into PBS control group, CXCL1 group, and CXCL1 combined with CXCR2 antagonist SB225002 group. Then endothelial transwell permeability assay kit was used to detect the endothelial permeability changes. After stimulated with CXCL1 in bEND.3 cells, Western blot analysis was used to detect the expression of protein kinase B (AKT) and phosphorylated-AKT (p-AKT). Results Intraperitoneal injection of LPS significantly increased the levels of TNF-α and CXCL1 in the whole brain. LPS and TNF-α both upregulated the expression of CXCR2 protein in bEND.3 cells. CXCL1 stimulation induced the endothelial cytoskeleton contraction, increased paracellular gap formation and elevated endothelial permeability in bEND.3 cells, which was inhibited by the pretreatment with SB225002(CXCR2 antagonist). Furthermore, CXCL1 stimulation also enhanced the phosphorylation of AKT in bEND.3 cells. Conclusion CXCL1 induces the cytoskeleton contraction and increased permeability through AKT phosphorylation in bEND.3 cells, which can be effectively inhibited by CXCR2 antagonist SB225002.

Modulation of metal species as control point for Ni-catalyzed stereodivergent semihydrogenation of alkynes with water.

A base-assisted metal species modulation mechanism enables Ni-catalyzed stereodivergent transfer semihydrogenation of alkynes with water, delivering both olefinic isomers smoothly using cheap and nontoxic catalysts and additives. Different from most precedents, in which E-alkenes derive from the isomerization of Z-alkene products, the isomers were formed in orthogonal catalytic pathways. Mechanistic studies suggest base as a key early element in modulation of the reaction pathways: by adding different bases, nickel species with disparate valence states could be accessed to initiate two catalytic cycles toward different stereoisomers. The practicability of the method is showcased with nearly 70 examples, including internal and terminal triple bonds, enynes and diynes, affording semi-hydrogenated products in high yields and selectivity.

[Protein tyrosine phosphatase receptor type O (PTPRO) promotes phagocytic activity of alveolar epithelial cells via activating AKT signal pathway in LPS induced acute lung injury].

Objective To investigate the effect of protein tyrosine phosphatase receptor type O (PTPRO) on the phagocytic activity of alveolar epithelial cells in LPS-induced acute lung injury. Methods Mice were randomly divided into the normal control group and LPS stimulation group. The infiltration of inflammatory cells was detected by HE staining. The cytokine TNF-α level in lung was analyzed by ELISA. Western blotting was performed to detect the effect of LPS on PTPRO protein expression in lung. After the expression of PTPRO in MLE-12 cells was silenced by siRNA in vitro, flow cytometry was used to detect the effects of LPS and PTPRO siRNA on the phagocytic activity of MLE-12 cells, and the effects of LPS and PTPRO siRNA on the expression of PTPRO, AKT and phosphorylated AKT protein were measured by Western blotting. Results After the establishment of murine acute lung injury model by LPS injection(1 mg/kg), the infiltrated polymorphonuclear leukocytes were markedly increased. The level of TNF-α in lung tissue and the expression of PTPRO in MLE-12 cells were both significantly increased after LPS stimulation. However, the activity of MLE-12 cells to phagocytose fluorescent microbeads was evidently decreased after silencing PTPRO. Furthermore, silencing PTPRO induced a remarkable decrease in the phosphorylation of AKT in MLE-12 cells. Conclusion PTPRO can promote phagocytic activity of MLE-12 cells via activating AKT signaling pathway.

Immunomodulatory roles of selenium nanoparticles: Novel arts for potential immunotherapy strategy development.

Current chemotherapy strategies used in clinic appear with lots of disadvantages due to the low targeting effects of drugs and strong side effects, which significantly restricts the drug potency, causes multiple dysfunctions in the body, and even drives the emergence of diseases. Immunotherapy has been proved to boost the body's innate and adaptive defenses for more effective disease control and treatment. As a trace element, selenium plays vital roles in human health by regulating the antioxidant defense, enzyme activity, and immune response through various specific pathways. Profiting from novel nanotechnology, selenium nanoparticles have been widely developed to reveal great potential in anticancer, antibacterial, and anti-inflammation treatments. More interestingly, increasing evidence has also shown that functional selenium nanoparticles can be applied for potential immunotherapy, which would achieve more effective treatment efficiency as adjunctive therapy strategies for the current chemotherapy. By directly interacting with innate immune cells, such as macrophages, dendritic cells, and natural killer cells, selenium nanoparticles can regulate innate immunity to intervene disease developments, which were reported to boost the anticancer, anti-infection, and anti-inflammation treatments. Moreover, selenium nanoparticles can also activate and recover different T cells for adaptive immunity regulations to enhance their cytotoxic to combat cancer cells, indicating the potential of selenium nanoparticles for potential immunotherapy strategy development. Here, aiming to enhance our understanding of the potential immunotherapy strategy development based on Se NPs, this review will summarize the immunological regulation effects of selenium nanoparticles and the application of selenium nanoparticle-based immunotherapy strategies. Furthermore, we will discuss the advancing perspective of selenium nanoparticle-based potential immunotherapy as a kind of novel adjunctive therapy to enhance the efficiency of current chemotherapies and also introduce the current obstacles for the development of selenium nanoparticles for potential immunotherapy strategy development. This work is expected to promote the future research on selenium nanoparticle-assisted immunotherapy and finally benefit the more effective disease treatments against the threatening cancer and infectious and chronic diseases.

First Report of Youcai Mosaic Virus Infecting Yam in China.

Yam (Dioscorea opposita Thunb.) is cultivated mainly as a functional food and for nutritional and medicinal purposes in China (1). It is propagated through tubers and this facilitates the spread and accumulation of viruses in the crop, eventually leading to yield losses (2). At present, different virus species belonging to the genera Aureusvirus, Badnavirus, Carlavirus, Comovirus, Cucumovirus, Fabavirus, Macluravirus, Potexvirus and Potyvirus have been reported in yams (3) and fifteen viruses in these genera have been detected in China. In July 2020, a survey of viral diseases on yam was conducted in plantations of Wenxian and Mengzhou counties in Henan Province, China. Fifty-four leaf samples of Dioscorea opposite showing mosaic and leaf discoloration (Supplementary Fig1) were collected from eight fields (five to ten plants per field). These leaf samples were ground in liquid nitrogen and total RNA was extracted from a portion of the mixed powder using RNAprep Pure Plant Plus Kit (TIANGEN Biotech, Beijing, China). A cDNA library was constructed using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB, USA) after ribosomal RNA depletion using Ribo-off rRNA Depletion Kit (Vazyme Biotech, Nanjing, China), and sequenced on the Illumina NovaSeq 6000 system at the Berry Genomics Corporation (Beijing, China). A total of 87,075 contigs (>200 bp) were generated from de novo assembly (CLC Genomic Workbench 10.0) from a total of 34,656,172 paired-end reads. After BLASTn analysis, three contigs with the length of 1009, 1340 and 1859 nucleotides shared 96.33%, 96.72% and 96.29% nt identity respectively with youcai mosaic virus SX isolate, a tobamovirus (YoMV GenBank accession no. JX422022). In addition to YoMV, broad bean wild virus 2 and yam latent virus were also identified, which had previously been reported in yams in China. To confirm the NGS result, total RNAs were extracted from fifty-four above-mentioned samples and RT-PCR was carried out to amplify a 528 bp fragment of the coat protein (CP) of YoMV by using a pair of specific primers CP gene. PCR products with expected size were obtained from 26 out of 54 samples, and seventeen amplicons of YoMV-CP were sequenced (accession nos. ON052726 to ON052742). The nt sequence identities of CP gene among these seventeen isolates were 99.6%-100%. Furthermore, the near-full-length genomic sequence of YoMV-Do41 isolate was obtained from sample 41 by RT-PCR amplification of four overlapping fragments using the following primer pairs: YoMV-15F/YoMV-1910R, YoMV-1770F/YoMV-3750R, YoMV-3645F/YoMV-5404R and YoMV-4921F/YoMV-6280R (Supplementary Table1). The YoMV-Do41 isolate was 6, 274 nt in length (accession no. ON149803) and shared 89.65% and 97.31% nt identities to As1-2 isolate (GenBank accession no. MW307290) and to SX isolate (accession no. JX422022), respectively.To the best of our knowledge, this is the first report of YoMV infecting yam in China. YoMV has a wide host range including genera Impatiens, Rehmannia, Brassica, Chelidonium, Trifolium, Crossandro, Alstroemeria, Stellaria. This study will serve as an important reference for the host range of YoMV. According to the detection rate infections with YoMV in yam are common in these producing regions. Further studies will be required to determine the infection rate in other producing regions and the potential threat posed by YoMV on yam production should be considered.

Genome-Based Analysis of a Multidrug-Resistant Hypervirulent Klebsiella pneumoniae.

Reports on multidrug-resistant hypervirulent Klebsiella pneumoniae (MDR-hvKP) in recent years indicate the wide-spreading trend of MDR-hvKP. The co-occurrence of hypervirulence and drug resistance poses a great challenge to clinical treatment. In this study, molecular characteristics of an MDR strain hvKP247 and 30 clinically isolated hvKP strains were characterized. The whole genome of hvKP247 belonging to sequence type (ST) 5214 and capsule serotype K1 was sequenced and analyzed. Conjugation experiments were performed to evaluate transferability of the plasmids in hvKP247. We found two new STs among our isolates, ST5570 and ST5571. The ST5214 hvKP247 contained two transferable plasmids: a hybrid virulence plasmid (pHvKP247-vir) carrying transfer-related modules that had self-transferable ability, and a drug-resistant plasmid (pHvKP247-MDR) that could be indirectly transferred with the help of pHvKP247-vir. The virulence-related genes were located on the pHvKP247-vir and chromosomal ICEKp1 mobile genetic element. In conclusion, the hybrid virulence plasmid and the drug-resistant plasmid are co-transferred, which emphasizes the importance of raising public awareness of the simultaneous spread of virulence and resistance genes of MDR-hvKP strains.

Binding mechanism of oseltamivir and influenza neuraminidase suggests perspectives for the design of new anti-influenza drugs.

Oseltamivir is a widely used influenza virus neuraminidase (NA) inhibitor that prevents the release of new virus particles from host cells. However, oseltamivir-resistant strains have emerged, but effective drugs against them have not yet been developed. Elucidating the binding mechanisms between NA and oseltamivir may provide valuable information for the design of new drugs against NA mutants resistant to oseltamivir. Here, we conducted large-scale (353.4 µs) free-binding molecular dynamics simulations, together with a Markov State Model and an importance-sampling algorithm, to reveal the binding process of oseltamivir and NA. Ten metastable states and five major binding pathways were identified that validated and complemented previously discovered binding pathways, including the hypothesis that oseltamivir can be transferred from the secondary sialic acid binding site to the catalytic site. The discovery of multiple new metastable states, especially the stable bound state containing a water-mediated hydrogen bond between Arg118 and oseltamivir, may provide new insights into the improvement of NA inhibitors. We anticipated the findings presented here will facilitate the development of drugs capable of combating NA mutations.