Diisoprenyl Cyclohexene-Type Meroterpenoids with Cytotoxic Activity from a Mangrove Endophytic Fungus Aspergillus sp. GXNU-Y85.

Five new diisoprenyl cyclohexene-type meroterpenoids, aspergienynes J-N (1-5), along with three known analogues (6-8), were obtained from the mangrove endophytic fungal strain Aspergillus sp. GXNU-Y85. The chemical structures, including their absolute configurations, were established via spectroscopic data and comparison of experimental and calculated ECD spectra. Cytotoxicity assay results indicated that compound 8 had strong cytotoxicity against HeLa cancer cells, and its IC50 value was 11.8 µM. In addition, flow cytometry analysis revealed that the cytotoxicity of 8 was due to the induction of G1 cell cycle arrest and apoptosis in HeLa cells.

Cytotoxic metabolites from the mangrove endophytic fungus Aspergillus sp. GXNU QG1a.

A new benzoquinone, guxiumasperone A (1), and a new diisoprenyl-cyclohexene-type meroterpenoids, biscognienyne M (2), along with four known diisoprenyl-cyclohexene analogues was isolated from the mangrove endophytic fungus Aspergillus QG1a. Their structures were determined by extensive spectral analysis of 1D and 2D NMR, HR-ESI- MS, and X-ray crystallography. Compound 1 was deduced by a single-crystal X-ray diffraction analysis, and the absolute configuration of 2 was further unequivocally elucidated by comparing the experimental electronic circular dichroism (ECD) data with calculated ECD spectra. Compounds 1 and 2 showed significant cytotoxic activity against selected tumour cells. Particularly, compound 2 exhibited strong activity against A2780 cancer cells with an IC50 value of 6.8 µM.

Diisoprenyl-cyclohexene-type meroterpenoids from a mangrove endophytic fungus Aspergillus sp. GXNU-Y65 and their anti-nonalcoholic steatohepatitis activity in AML12 cells.

Nine previously undescribed diisoprenyl-cyclohexene-type meroterpenoids, aspergienynes A-I, together with five known analogues, were obtained from the mangrove endophytic fungal strain Aspergillus sp. GXNU-Y65. The diisoprenyl-cyclohexene-type meroterpenoids were elucidated based on multispectroscopic analysis, and the previously undescribed compounds' absolute configurations were established via electronic circular dichroism calculations. Biological activity results indicated that aspergienyne C (compound 3) had strong anti-nonalcoholic steatohepatitis activity against AML12 cells treated with PA (Palmitic acid) + OA (Oleic acid). At the same concentration of 20 µM, 3 significantly reduced triglyceride (TG) content compared with fenofibrate (positive control) in PA + OA treated AML12 cells, and obviously increased phosphorylation of acetyl-CoA carboxylase.

A new sulphur-containing metabolite from a mangrove endophytic fungus Aspergillus sp. GXNU-MA.

A new sulphur-containing metabolite, asperiguxidione A (1), was isolated from a mangrove endophytic fungus Aspergillus sp. GXNU-MA, and four known alkaloids 2-5, were isolated together from this strain. Their structures were determined by the com-bination of 1D and 2D NMR spectroscopy, HR-ESI-MS, and ECD analysis. Compounds 1 and 2 exhibited mediate activity against Staphylococcus aureus and Enterobacter aerogenes with equal MIC values of 12.5 µg/mL. Compound 3 reduced NO production in LPS-stimulated cells with an IC50 value of 13.329 ± 0.53 µg/mL in the anti-inflammatory assay.

Liver transcriptomic and proteomic analyses provide new insight into the pathogenesis of liver fibrosis in mice.

BACKGROUND: Liver fibrosis (LF) is a kind of progressive liver injury reaction. The goal of this study was to achieve a more detailed understanding of the molecular changes in response to CCl4-induced LF through the identification of a differentially expressed liver transcriptomic and proteomic. RESULTS: A total of 1224 differentially expressed genes (DEGs) and 302 differentially expressed proteins (DEPs) were significantly identified at the transcriptomic and proteomic level, respectively, and 69 genes (hereafter "cor-DEGs-DEPs" genes) were detected at both levels. Pathway enrichment analysis showed that these cor-DEGs-DEPs genes were significantly enriched in 133 pathways. Importantly, among the cor-DEGs-DEPs genes, Gstm1, Gstm3, Ephx1 and Gstp1 were shown to be associated with metabolic pathways, and confirmed by RT-qPCR and parallel reaction monitoring (PRM) verification. CONCLUSIONS: Through the combined analysis of transcriptomic and proteomic data, this study provides valuable insights into the potential mechanism of the pathogenesis of LF, and lays a theoretical foundation for the further development of targeted therapy for LF.

Isolation and insecticidal activities of new cyclic peptides from mangrove endophytic fungus Aspergillus sp. GXNU-4QQY1a.

An investigation on bioactive metabolites from the mangrove endophytic fungus Aspergillus sp. GXNU-4QQY1a led to the isolation of two undescribed cyclic peptides, guaspertide A (1) and guaspertide B (2), together with six known compounds, 3-8. These structures and the new compounds' absolute configuration were determined by mass spectrometry analysis, nuclear magnetic resonance spectrum, electronic circular dichroism, and single-crystal X-ray diffraction. Insecticidal assays were carried out with compounds 1-8, and the results showed that compounds 1-3 and 8 exhibited good insecticidal activity against citrus psyllids.

High-Throughput Sequencing Reveals N6-Methyladenosine-modified LncRNAs as Potential Biomarkers in Mice with Liver Fibrosis.

BACKGROUND: N6-methyladenosine (m6A) is the most frequent internal modification in eukaryotic RNA. Long noncoding RNAs (lncRNAs) are a new type of noncoding regulatory molecule with multiple cellular functions. Both are closely related to the occurrence and development of liver fibrosis (LF). However, the role of m6A-methylated lncRNAs in the progression of LF remains largely unknown. METHODS: In this study, HE and Masson staining were used to observe pathological changes in the liver, m6A-modified RNA immunoprecipitation sequencing (m6A-seq) was performed to systematically evaluate the m6A modification level of lncRNAs in LF mice, meRIP-qPCR and RT-qPCR were used to detect the m6A methylation level and RNA expression level of the target lncRNAs. RESULTS: A total of 415 m6A peaks were detected in 313 lncRNAs in liver fibrosis tissues. There were 98 significantly different m6A peaks in LF, which were located on 84 lncRNAs, of which 45.2% of the lncRNA length was between 200-400 bp. At the same time, the first three chromosomes of these methylated lncRNAs were chromosomes 7, 5 and 1. RNA sequencing identified 154 differentially expressed lncRNAs in LF. The joint analysis of m6A-seq and RNA-seq found that there were three lncRNAs with significant changes in m6A methylation and RNA expression levels: lncRNA H19, lncRNA Gm16023 and lncRNA Gm17586. Subsequently, the verification results showed that the m6A methylation levels of lncRNA H19 and lncRNA Gm17586 were significantly increased, while that of lncRNA Gm16023 was significantly decreased, and the RNA expression of three lncRNAs was significantly decreased. Through the establishment of a lncRNA-miRNA-mRNA regulatory network, the possible regulatory relationships of lncRNA H19, lncRNA Gm16023 and lncRNA Gm17586 in LF were revealed. CONCLUSION: This study revealed the unique m6A methylation pattern of lncRNAs in LF mice, suggesting that the m6A methylation modification of lncRNAs is related to the occurrence and development of LF.

Early markers of neurodevelopmental disorders based on general movements for very preterm infants: study protocol for a multicentre prospective cohort study in a clinical setting in China.

INTRODUCTION: Very preterm (VPT) infants may experience varying degrees of neurodevelopmental challenges. Lack of early markers for neurodevelopmental disorders may delay referral to early interventions. The detailed General Movements Assessment (GMA) could help us to identify early markers for VPT infants at risk of atypical neurodevelopmental clinical phenotype in the very early stage of life as soon as possible. Preterm infants with high risk of atypical neurodevelopmental outcomes will have the best possible start to life if early precise intervention in critical developmental windows is allowed. METHODS AND ANALYSIS: This is a nationwide, multicentric prospective cohort study that will recruit 577 infants born <32 weeks of age. This study will determine the diagnostic value of the developmental trajectory of general movements (GMs) at writhing and fidgety age with qualitative assessment for different atypical developmental outcomes at 2 years evaluated by the Griffiths Development Scales-Chinese. The difference in the General Movement Optimality Score (GMOS) will be used to distinguish normal (N), poor repertoire (PR) and cramped sychronised (CS) GMs. We plan to build the percentile rank of GMOS (median, 10th, 25th, 75th and 90th percentile rank) in N, PR and CS of each global GM category and analyse the relationship between GMOS in writhing movements and Motor Optimality Score (MOS) in fidgety movements based on the detailed GMA. We explore the subcategories of the GMOS list, and MOS list that may identify specific early markers that help us to identify and predict different clinical phenotypes and functional outcomes in VPT infants. ETHICS AND DISSEMINATION: The central ethical approval has been confirmed from the Research Ethical Board of Children's Hospital of Fudan University (ref approval no. 2022(029)) and the local ethical approval has been also obtained by the corresponding ethics committees of the recruitment sites. Critical analysis of the study results will contribute to providing a basis for hierarchical management and precise intervention for preterm infants in very early life. TRIAL REGISTRATION NUMBER: ChiCTR2200064521.

Quantitative phosphoproteomic analysis of mice with liver fibrosis by DIA mass spectrometry analysis with PRM verification.

Liver fibrosis (LF), commonly associated with chronic liver diseases, is a major public health problem worldwide. Protein phosphorylation is not only an important form of protein modification in organisms but also the most important mechanism to regulate and control the activity and function of proteins, affecting the occurrence and development of many diseases. However, comprehensive phosphoproteomic profiling in LF has not been fully elucidated. In this study, data-independent acquisition (DIA) was used to analyse the phosphoproteomics of mice with LF. A total of 553 phosphopeptides (representing 440 phosphoproteins) had significant phosphorylation levels. Among these phosphoproteins, 49 were upregulated and 401 were downregulated, and 5 phosphoserine (P-Ser) motifs and 2 phosphothreonine (P-Thr) motifs were conserved in LF. GO and KEGG pathway enrichment analyses identified 769 significant GO terms and 49 significant KEGG pathways. Four phosphorylated proteins were selected for parallel reaction monitoring (PRM) verification, and the results were consistent with DIA data. Together, there were significantly different phosphoproteomic profiles in LF, suggesting that protein phosphorylation was related to the occurrence and progression of LF, which could pave the way for further investigation into the related regulatory mechanisms. SIGNIFICANCE: LF is a necessary stage in the development of chronic liver disease to liver cirrhosis and has attracted wide attention. To the best of our knowledge, there are few reports on the phosphorylated proteomics of LF. In this study, DIA and PRM techniques were used to study the liver tissue of mice induced by CCl4. The results showed that phosphorylation had a significant effect on the activity and function of proteins, and the PRM results were consistent with the trend observed in DIA analysis. This study will help to better reveal the relationship of phosphorylated proteins in LF and lay a foundation for further study of related regulatory mechanisms.

Asperphenyltones A and B: New Phenylfuropyridinone Skeleton from an Endophytic Aspergillus sp. GXNU-A1.

Chemical investigation of the fermentation extract of the mangrove endophytic fungus Aspergillus sp. GXNU-A1, isolated from Acanthus ilicifolius L., discovered an undescribed pair of enantiomers (asperphenyltones A and B (±1)), together with four previously described metabolites: nodulisporol (2), isosclerone (3), 2,3,4-trihydroxy-6-(hydroxymethyl)-5-methylbenzyl alcohol (4), and 4,6-dihydroxy-5-methoxy-7-methyl-1,3-dihydroisobenzofuran (5). Analyses of the 1D and 2D NMR spectroscopic data of the compounds supported their structural assignments. The presence of the asperphenyltones A and B, which are a pair of enantiomers, was established by HR-ESI-MS, 1D and 2D NMR data and confirmed by single-crystal X-ray diffraction analysis. Metabolites 1-5 were evaluated for their anti-inflammatory effects on the production of nitric oxide (NO), and 1, 3, and 4 showed significant potential inhibitory activities against NO production in activated macrophages with IC50 values of 26-40 µM, respectively.

Heat Adaptive Capacity: What Causes the Differences Between Residents of Xiamen Island and Other Areas?

Extreme heat events caused by climate change have serious adverse effects on residents' health in many coastal metropolises in southeast China. Adaptive capacity (AC) is crucial to reduce heat vulnerability in the human-environment system. However, it is unclear whether changes in individual characteristics and socioeconomic conditions likely amplify or attenuate the impacts of residents' heat adaptive capacity (HAC) changes. Moreover, which public policies can be implemented by the authorities to improve the HAC of vulnerable groups remains unknown. We conducted a questionnaire survey of 630 residents of Xiamen, a typical coastal metropolis, in 2018. The effects of individual and household characteristics, and government actions on the residents' HAC were examined by using ordinal logistic regression analysis. Results show that the majority (48.10%) of Xiamen residents had a "medium" HAC level, followed by a "high" level (37.14%). On Xiamen Island, residents who settled locally for one-three years and spent less than one hour outdoors might report weaker HAC, and their HAC would not improve with increased air conditioning units in household. In other areas of Xiamen, residents with more rooms in their households, no educational experience, and building areas <50 m2 might report better HAC. Further, vulnerable groups, such as local residents and outdoor workers on Xiamen Island, people lacking educational experience and renters in other areas of Xiamen, showed better AC to hot weather than those in previous studies. Low-income groups should be given more attention by local governments and community groups as monthly household income played a positive role in improving Xiamen residents' HAC. Rational green spaces planning and cooling services, such as street sprinkling operations, provided by municipal departments can effectively bring benefits to Xiamen residents. Identification of basic conditions of AC has significant implications for practical promoting targeted measures or policies to reduce health damages and livelihood losses of urban residents during extreme heat events.

Two-Dimensional TiVC Solid-Solution MXene as Surface-Enhanced Raman Scattering Substrate.

Two-dimensional (2D) MXenes are attractive candidates as surface-enhanced Raman scattering (SERS) substrates because of their metallic conductivity and abundant surface terminations. Herein, we report the facile synthesis of bimetallic solid-solution TiVC (MXene) and its application in SERS. The few-layered MXene nanosheets with high crystallinity were successfully prepared using a one-step chemical etching method without ultrasonic and organic solvent intercalation steps. SERS activity of the as-prepared MXene was investigated by fabricating free-standing TiVC film as the substrate. A SERS enhancement factor of 1012 and femtomolar-level detection limit were confirmed using rhodamine 6G as a model dye with 532 nm excitation. The fluorescent signal of the rhodamine 6G dye was effectively quenched, making the SERS spectrum clearly distinguishable. Furthermore, we demonstrate that the TiVC-analyte system with ultrahigh sensitivity is dominated by the chemical mechanism (CM) based on the experimental and simulation results. The abundant density of states near the Fermi level of the TiVC and the strong interaction between the TiVC and analyte promote the intermolecular charge transfer resonance in the TiVC-analyte complex, resulting in significant Raman enhancement. Additionally, several other probe molecules were used for SERS detection to further verify CM-based selectivity enhancement on the TiVC substrates. This work provides guidance for the facile synthesis of 2D MXene and its application in ultrasensitive SERS detection.

Comprehensive Analysis of the Transcriptome-Wide m6A Methylation Modification Difference in Liver Fibrosis Mice by High-Throughput m6A Sequencing.

N6-Methyladenosine (m6A), a unique and common mRNA modification method in eukaryotes, is involved in the occurrence and development of many diseases. Liver fibrosis (LF) is a common response to chronic liver injury and may lead to cirrhosis and even liver cancer. However, the involvement of m6A methylation in the development of LF is still unknown. In this study, we performed a systematic evaluation of hepatic genome-wide m6A modification and mRNA expression by m6A-seq and RNA-seq using LF mice. There were 3,315 genes with significant differential m6A levels, of which 2,498 were hypermethylated and 817 hypomethylated. GO and KEGG analyses illustrated that differentially expressed m6A genes were closely correlated with processes such as the endoplasmic reticulum stress response, PPAR signaling pathway and TGF-ß signaling pathway. Moreover, a total of 90 genes had both a significant change in the m6A level and mRNA expression shown by joint analysis of m6A-seq and RNA-seq. Hence, the critical elements of m6A modification, including methyltransferase WTAP, demethylases ALKBH5 and binding proteins YTHDF1 were confirmed by RT-qPCR and Western blot. In an additional cell experiment, we also observed that the decreased expression of WTAP induced the development of LF as a result of promoting hepatic stellate cell (HSC) activation. Therefore, this study revealed unique differential m6A methylation patterns in LF mice and suggested that m6A methylation was associated with the occurrence and course of LF to some extent.

Up-regulation of nPKC contributes to proliferation of mice pulmonary artery smooth muscle cells in hypoxia-induced pulmonary hypertension.

This study is designed to investigate the role of novel protein kinases C (nPKC) in mediating pulmonary artery smooth muscle cells (PASMCs) proliferation in pulmonary hypertension (PH) and the underlying mechanisms. Mouse PASMCs was isolated using magnetic separation technology. The PASMCs were divided into 24 h group, 48 h group and 72 h group according to different hypoxia treatment time, then detected cell proliferation rate and nPKC expression level in each group. We treated PASMCs with agonists or inhibitors of PKCdelta (PKCδ) and PKCepsilon (PKCε) and exposed them to hypoxia or normoxia for 72 h, then measured the proliferation of PASMCs. We also constructed a lentiviral vector containing siRNA fragments for inhibiting PKCδ and PKCε to transfected PASMCs, then examined their proliferation. PASMCs isolated successfully by magnetic separation method and were in good condition. Hypoxia promoted the proliferation of PASMCs, and the treatment for 72 h had the most significant effect. Hypoxia upregulated the expression of PKCδ and PKCε in mouse PASMCs, leading to PASMCs proliferation. Moreover, Our study demonstrated that hypoxia induced upregulation of PKCδ and PKCε expression resulting to the proliferation of PASMCs via up-regulating the phosphorylation of AKT and ERK. Our study provides clear evidence that increased nPKC expression contributes to PASMCs proliferation and uncovers the correlation between AKT and ERK pathways and nPKC-mediated proliferation of PASMCs. These findings may provide novel targets for molecular therapy of pulmonary hypertension.

Screening and evaluation of key genes in contributing to pathogenesis of hepatic fibrosis based on microarray data.

BACKGROUND: Hepatic fibrosis (HF), which is characterized by the excessive accumulation of extracellular matrix (ECM) in the liver, usually progresses to liver cirrhosis and then death. To screen differentially expressed (DE) long non-coding RNAs (lncRNAs) and mRNAs, explore their potential functions to elucidate the underlying mechanisms of HF. METHODS: The microarray of GSE80601 was downloaded from the Gene Expression Omnibus database, which is based on the GPL1355 platform. Screening for the differentially expressed LncRNAs and mRNAs was conducted between the control and model groups. Then, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to analyze the biological functions and pathways of the DE mRNAs. Additionally, the protein-protein interaction (PPI) network was delineated. In addition, utilizing the Weighted Gene Co-expression Network Analysis (WGCNA) package and Cytoscape software, we constructed lncRNA-mRNA weighted co-expression networks. RESULTS: A total of 254 significantly differentially expressed lncRNAs and 472 mRNAs were identified. GO and KEGG analyses revealed that DE mRNAs regulated HF by participating in the GO terms of metabolic process, inflammatory response, response to wounding and oxidation-reduction. DE mRNAs were also significantly enriched in the pathways of ECM-receptor interaction, PI3K-Akt signaling pathway, focal adhesion (FA), retinol metabolism and metabolic pathways. Moreover, 24 lncRNAs associated with 40 differentially expressed genes were observed in the modules of lncRNA-mRNA weighted co-expression network. CONCLUSIONS: This study revealed crucial information on the molecular mechanisms of HF and laid a foundation for subsequent genes validation and functional studies, which could contribute to the development of novel diagnostic markers and provide new therapeutic targets for the clinical treatment of HF.

Prognostic Nomogram for Overall Survival in Early Stage Extranodal Natural Killer/T Cell Lymphoma Treated With High-Dose Radiotherapy.

BACKGROUND: The aim of this study was to develop a prognostic nomogram for early stage extranodal natural killer/T-cell lymphoma, nasal type (ENKL) treated with high-dose radiotherapy (RT). PATIENTS AND METHODS: A total of 81 patients at 2 cancer centers with stage I to IIE ENKL who received chemotherapy (CT) and high-dose RT were retrospectively analyzed. The development of the nomogram was on the basis of the Cox proportional hazards model. We implemented the concordance index (C-index) and performed a calibration curve to determine its predictive and discriminatory capacity and compared our nomogram with the International Prognostic Index (IPI) and the Korean Prognostic Index (KPI). RESULTS: The nomogram included 4 important variables and used a multivariate analysis: lactate dehydrogenase, primary tumor invasion, tumor response, and CT regimen. The 5-year OS rate and progression-free survival were 64.7% and 57.5%, respectively for the entire group. The C-index of the nomogram for overall survival (OS) prediction was 0.87, and it was superior to the predictive power of the IPI and KPI. The calibration curve showed that the nomogram accurately predicted the 5-year OS. CONCLUSION: The proposed nomogram could provide an individualized risk estimate of the OS for early stage ENKL treated with CT and high-dose RT.

Stem-like cells of various origins showed therapeutic effect to improve the recovery of spinal cord injury.

We aimed to evaluate the therapeutic effects of exosomes, which were collected from human neuroepithelial stem cells (HNESCs) treated by miR-29b mimics, on the treatment of spinal cord injury (SCI). Computational analysis, real-time PCR, Western blot analysis and TUNEL assay, a BBB score system, the Nissl staining and IHC assay were conducted to explore the molecular signalling pathway underlying the function of exosomes in SCI. Exosomes isolated from cells treated with HNESC exhibited the strongest inhibitory effect on cell apoptosis while exhibiting the highest level of miR-29b expression and the lowest levels of PTEN and caspase-3 expression. Moreover, PTEN and caspase-3 were identified as the direct target genes of miR-29b. The exosomes isolated from the groups of HNESC and HNESC + miR-29b mimics exhibited in vivo therapeutic effects by restoring the BBB score and apoptosis index of post-SCI neuron cells to those of normal neuron cells, with the exosomes collected from the group of HNESC + miR-29b mimics showing the strongest effect. We suggested that the exosomes derived from the group of HNESC + miR-29b mimics exerted therapeutic effects on SCI by down-regulating the expression of PTEN/caspase-3 and subsequently suppressing the apoptosis of neuron cells.

The therapeutic effect of bone marrow-derived mesenchymal stem cells on osteoarthritis is improved by the activation of the KDM6A/SOX9 signaling pathway caused by exposure to hypoxia.

Abnormal expression of KDM6A and SOX9 is a key factor in the pathogenesis of osteoarthritis (OA). Cellular treatments of OA with articular cartilage chondrocytes (ACCs) and bone marrow mesenchymal stem cells (BMSCs) are promising, but their underlying mechanisms remain to be explored. The pellet size, weight and sulfated glycosaminoglycan/DNA content of ACCs were measured to evaluate the effect of BMSCs on the chondrogenic differentiation of SCCs. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to analyze the proliferation of ACCs cultured along or cocultured with BMSCs. Quantitative polymerase chain reaction (qPCR) was performed to evaluate the messenger RNA expression of KDM6A, SOX9, type2 collagen, and Aggrecan in ACCs and OA rats. Western blot and immunohistochemistry were performed to analyze the expression of KDM6A and SOX9 proteins. Bisulfite sequencing PCR was performed to assess the DNA methylation level of the SOX9 promoter. Flow cytometry was used to evaluate the apoptotic status of ACCs. The chondrogenic differentiation of ACCs was significantly enhanced by coculturing with BMSCs, especially under a hypoxic condition. The expression of KDM6A, SOX9, type2 collagen, and Aggrecan was remarkably elevated in ACCs cocultured with BMSCs. Also, the DNA methylation of SOX9 promoter was decreased in ACCs cocultured with BMSCs, along with notably reduced apoptosis. Moreover, ACCs cocultured with BMSCs could repair cartilage lesions and prevent the abnormal expression of KDM6A, SOX9, type2 collagen, and Aggrecan in OA rats. In this study, we cocultured ACCs with BMSCs and used them to treat OA rats. Our findings presented a mechanistic basis for explaining the therapeutic effect of BMSCs on OA treatment.