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Abstract In this experiment, alfalfa silage with different packing densities (500 kg/m3、600 kg/m3 and 700 kg/m3) was prepared under the conditions of outdoor high temperature and indoor room temperature, respectively. At the same time, the same lactobacillus additive was used for fermentation in each density treatment group. The chemical composition, fermentation quality and microbial community of alfalfa silage were analyzed. The results showed that the contents of dry matter (DM) and water-soluble carbohydrate (WSC) decreased with the increase of density during fermentation at high temperature. At the same time, when the density is 600 kg/m³, CP (crude protein) content is the highest, ADF (acid detergent fiber) content is the lowest. The contents and pH values of neutral detergent fiber (NDF), lactic acid (LA) and lactic acid bacteria (LAB) were significantly affected by temperature (p

Abstract This study investigated the influence of different temperatures (35℃ High temperature and average indoor ambient temperature of 25℃) and lactic acid bacterial additives (Lactiplantibacillus plantarym, Lentilactobacillus buchneri, or a combination of Lactiplantibacillus plantarym and Lentilactobacillus buchneri) on the chemical composition, fermentation quality, and microbial community of alfalfa silage feed. After a 60-day ensiling period, a significant interaction between temperature and additives was observed, affecting the dry matter (DM), crude protein (CP), acid detergent fiber (ADF), and neutral detergent fiber (NDF) of the silage feed (p < 0.05). Temperature had a highly significant impact on the pH value of the silage feed (p 0.05), while the inoculation of additives had a significant effect on lactic acid, acetic acid, and butyric acid (p > 0.05). As for the dynamic changes of microbial community after silage, the addition of three kinds of bacteria increased the abundance of lactobacillus. Among all treatment groups, the treatment group using complex bacteria had the best fermentation effect, indicating that the effect of complex lactic acid bacteria was better than that of single bacteria in high temperature fermentation. In summary, this study explained the effects of different temperatures and lactic acid bacterial additives on alfalfa fermentation quality and microbial community, and improved our understanding of the mechanism of alfalfa related silage at high temperatures.

The paper addresses the challenges of task migration and resource allocation in heterogeneous cloud–edge environments, where dynamic and stochastic conditions complicate efficient scheduling. To tackle this, the authors propose a novel scheduling algorithm combining soft actor–critic (SAC) agent with masked layer and graph convolutional network (GCN), namely MGSAC algorithm. MGSAC utilizes GCN to extract hidden structural features from the environment, enabling better adaptation to dynamic changes. Additionally, a learnable mask layer filters out ineffective actions, refining the selection of scheduling strategies and improving overall performance. By evaluating MGSAC on the real-world Bit-Brain dataset and simulating it using Cloud-Sim, experimental results demonstrate its superiority over existing algorithms in energy consumption, task response time, task migration time, and task Service-Level-Agreement violations rate, showcasing its effectiveness in real-world scenarios.

Summary: Blockade of programmed cell death 1 (PD-1) is considered a promising strategy for controlling pathogen infection by enhancing host immune cell function. Eosinophils, which play a crucial role in type 2 immune responses, are essential components of the host defense against helminth infection. Here, we investigate the role of PD-1 in eosinophilia during Trichinella spiralis infection in mice. PD-1-deficient (PD-1−/−) mice exhibit delayed expulsion of adult worms and increased muscle larva burdens compared to wild-type mice following infection. Additionally, PD-1−/− mice display impaired recruitment of eosinophils to parasite-invaded tissues, attributed to decreased upregulation of adhesion molecules on both eosinophils and vascular endothelium after infection. The compromised Th2 cytokine response further contributes to impaired adhesion interactions, affecting eosinophil migration and cytotoxicity against larvae in vitro within T. spiralis-infected PD-1−/− mice. Our findings demonstrate a positive role for PD-1 in the recruitment of eosinophils, suggesting its involvement in host defense against helminth infection.

Abstract Ischemia-reperfusion injury (IRI) is a common cause of acute kidney injury (AKI). The kidney is susceptible to IRI under several clinical conditions, including hypotension, sepsis, and surgical procedures, such as partial nephrectomy and kidney transplantation. Extensive research has been conducted on the mechanism and intervention strategies of renal IRI in past decades; however, the complex pathophysiology of IRI-induced AKI (IRI-AKI) is not fully understood, and there remains a lack of effective treatments for AKI. Renal IRI involves several processes, including reactive oxygen species (ROS) production, inflammation, and apoptosis. Mitochondria, the centers of energy metabolism, are increasingly recognized as substantial contributors to the early phases of IRI. Multiple mitochondrial lesions have been observed in the renal tubular epithelial cells (TECs) of IRI-AKI mice, and damaged or dysfunctional mitochondria are toxic to the cells because they produce ROS and release cell death factors, resulting in TEC apoptosis. In this review, we summarize the recent advances in the mitochondrial pathology in ischemic AKI and highlight promising therapeutic approaches targeting mitochondrial dysfunction to prevent or treat human ischemic AKI.

Abstract Transcription factor EB (TFEB), known as a major transcriptional regulator of the autophagy-lysosomal pathway, regulates target gene expression by binding to coordinated lysosomal expression and regulation (CLEAR) elements. TFEB are regulated by multiple links, such as transcriptional regulation, post-transcriptional regulation, translational-level regulation, post-translational modification (PTM), and nuclear competitive regulation. Targeted regulation of TFEB has been victoriously used as a treatment strategy in several disease models such as ischemic injury, lysosomal storage disorders (LSDs), cancer, metabolic disorders, neurodegenerative diseases, and inflammation. In this review, we aimed to elucidate the regulatory mechanism of TFEB and its applications in several disease models by targeting the regulation of TFEB as a treatment strategy.

Landslides, prevalent in mountainous areas, are typically triggered by tectonic movements, climatic changes, and human activities. They pose catastrophic risks, especially when occurring near settlements and infrastructure. Therefore, detecting, monitoring, and predicting landslide deformations is essential for geo-risk mitigation. The mainstream of the previous studies have often focused on deterministic models for immediate landslide prediction. However, in most of them, the aspect of prediction uncertainties are not sufficiently addressed. This paper introduces an innovative probabilistic prediction method using a Variational Autoencoder (VAE) combined with Gated Recurrent Unit (GRU) to forecast landslide deformations from a generative standpoint. Our approach consists of two main elements: firstly, training the VAE-GRU model to maximize the variational lower bound on the likelihood of historical precipitation data; secondly, using the learned approximated posterior distribution to predict imminent deformations from a generative angle. To assess the prediction quality, we use four widely-used metrics: Prediction Interval Coverage Probability (PICP), Prediction Interval Normalized Average Width (PINAW), Coverage Width-Based Criterion (CWC), and Prediction Interval Normalized Root Mean Square Width (PINRW). The results demonstrate that our proposed VAE-GRU framework surpasses traditional state-of-the-art (SOTA) probabilistic deformation prediction algorithms in terms of accuracy and reliability.

Objective: Numerous studies have highlighted the role of clock genes in diabetes disease and pancreatic β cell functions. However, whether rhythmic long non-coding RNAs involve in this process is unknown. Methods: RNA-seq and 3’ rapid amplification of cDNA ends (RACE)-PCR were used to identify the rat LncCplx2 in pancreatic β cells. The subcellular analysis with qRT-PCR and RNA-Scope were used to assess the localization of LncCplx2. The effects of LncCplx2 overexpression or knockout (KO) on the regulation of pancreatic β cell functions were assessed in vitro and in vivo. RNA-seq, immunoblotting (IB), Immunoprecipitation (IP), RNA pull-down, and chromatin immunoprecipitation (ChIP)-PCR assays were employed to explore the regulatory mechanisms through LncRNA-protein interaction. Metabolism cage was used to measure the circadian behaviors. Results: We first demonstrate that LncCplx2 is a conserved nuclear long non-coding RNA and enriched in pancreatic islets, which is driven by core clock transcription factor BMAL1. LncCplx2 is downregulated in the diabetic islets and repressed by high glucose, which regulates the insulin secretion in vitro and ex vivo. Furthermore, LncCplx2 KO mice exhibit diabetic phenotypes, such as high blood glucose and impaired glucose tolerance. Notably, LncCplx2 deficiency has significant effects on circadian behavior, including prolonged period duration, decreased locomotor activity, and reduced metabolic rates. Mechanistically, LncCplx2 recruits EZH2, a core subunit of polycomb repression complex 2 (PRC2), to the promoter of target genes, thereby silencing circadian gene expression, which leads to phase shifts and amplitude changes in insulin secretion and cell cycle genes. Conclusions: Our results propose LncCplx2 as an unanticipated transcriptional regulator in a circadian system and suggest a more integral mechanism for the coordination of circadian rhythms and glucose homeostasis.

Lactuca indica L. cv. Mengzao (LIM), acknowledged as a pivotal “One Root of Medicine and Food”, boasts dual applications in both culinary and medicinal domains. This research delves into the influence of various harvest periods (vegetative, budding, blossom, and fruiting) on distinct medicinal parts (roots, stems, leaves, flowers, and seeds) of LIM, employing plant metabolomics to assess its chemical constituents. A total of 66 chemical constituents were identified in LIM, with 11 chemical components emerging as potential markers for distinguish medicinal parts. Notably, nutritional organs exhibited elevated levels of cichoric acid, rutin and chlorogenic acid. Specifically, leaves during the budding stage displayed the highest chicoric acid content at 11.70 mg·g−1. Conversely, reproductive organs showed heightened concentrations of cichoric acid, rutin and chlorogenic acid, with seeds exhibiting the peak cichoric acid content at 4.53 mg g−1. This study enriches our understanding of LIM by offering novel insights into quality assessment and the comprehensive utilization of its diverse parts.

Lactuca indica L. contains high amounts of flavonoids and phenolic acids. However, there is limited information on the composition of these compounds in different parts of the plant. The present study analyzed the secondary metabolite profiles of the stem, leaf, flower, and seed of Lactuca indica L.cv. Mengzao (LIM) using a widely targeted metabolomic approach. A total of 576 secondary metabolites were identified, including 218, 267, 232, 286, 302, and 308 differentially accumulated metabolites (DAMs) in the stem_vs_leaf, stem_vs_flower, stem_vs_seed, leaf_vs_flower, leaf_vs_seed, and flower_vs_seed comparisons. In particular, considerable differences were detected in the flavonoids and phenolic acids, five flavonoids, five phenolic acids, one triterpenoid and one alkaloid being differentially accumulated in the four parts. Compared to the stem and flower, the leaf and seed had higher total flavonoid content and total phenolic content. Thses findings provide comprehensive insights into utilizing different parts of LIM in developing functional food products.

Abstract Ferroptosis, a nonapoptotic form of programmed cell death characterized by significant iron-dependent peroxidation of phospholipids, is regulated by cellular metabolism, redox homeostasis, and various cancer-related signaling pathways. Recently, considerable progress has been made in demonstrating the critical role of lipid metabolism in regulating ferroptosis, indicating the potential of combinational strategies for treating cancer in the future. In this study, we explored the combinational effects of lipid metabolism compounds and ferroptosis inducers on renal cell carcinoma (RCC) cells. We found potent synergy of the fatty acid amide hydrolase (FAAH) inhibitor URB597 with ferroptosis inducer (1S, 3R)-RSL3 (RSL3) in inhibiting the growth and metastasis of RCC cells both in vitro and in vivo via induction of G1 cell cycle arrest and promotion of the production of lipid peroxides, malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), and cytosolic reactive oxygen species (ROS). In addition, inhibition of FAAH increased the sensitivity of RCC cells to ferroptosis. Genome-wide RNA sequencing indicated that the combination of URB597 and RSL3 has more significant effects on regulation of the expression of genes related to cell proliferation, the cell cycle, cell migration and invasion, and ferroptosis than either single agent alone. Moreover, we found that combinational treatment modulated the sensitivity of RCC cells to ferroptosis via the phosphatidylinositol 3 kinase (PI3K)-AKT signaling pathway. These data demonstrate that dual targeting of FAAH and ferroptosis could be a promising strategy for treating RCC.

Abstract TFE3 is a member of the MiT family of the bHLH-leucine zipper transcription factor. We previously focused on the role of TFE3 in autophagy and cancer. Recently, an increasing number of studies have revealed that TFE3 plays an important role in metabolic regulation. TFE3 participates in the metabolism of energy in the body by regulating pathways such as glucose and lipid metabolism, mitochondrial metabolism, and autophagy. This review summarizes and discusses the specific regulatory mechanisms of TFE3 in metabolism. We determined both the direct regulation of TFE3 on metabolically active cells, such as hepatocytes and skeletal muscle cells, and the indirect regulation of TFE3 through mitochondrial quality control and the autophagy–lysosome pathway. The role of TFE3 in tumor cell metabolism is also summarized in this review. Understanding the diverse roles of TFE3 in metabolic processes can provide new avenues for the treatment of some metabolism-related disorders.

Abstract Organelles are functional areas where eukaryotic cells perform processes necessary for life. Each organelle performs specific functions; however, highly coordinated crosstalk occurs between them. Disorder of organelle networks often occur in various diseases. The endoplasmic reticulum (ER) and mitochondria are crucial organelles in eukaryotic cells as they are the material synthesis and oxidative metabolism centers, respectively. Homeostasis and orchestrated interactions are essential for maintaining the normal activities of cells. However, the mode and mechanism of organelle crosstalk is still a research challenge. Furthermore, the intricate association between organelle dyshomeostasis and the progression of many human diseases remains unclear. This paper systematically summarized the latest research advances in the synergistic mechanism between the endoplasmic reticulum and mitochondria and their crosstalk with other organelles based on recent literature. It also highlights the application potential of organelle homeostasis maintenance as a preventative and treatment strategy for diseases.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a tickborne bandavirus mainly transmitted by Haemaphysalis longicornis ticks in East Asia, mostly in rural areas. As of April 2022, the amplifying host involved in the natural transmission of SFTSV remained unidentified. Our epidemiologic field survey conducted in endemic areas in China showed that hedgehogs were widely distributed, had heavy tick infestations, and had high SFTSV seroprevalence and RNA prevalence. After experimental infection of Erinaceus amurensis and Atelerix albiventris hedgehogs with SFTSV, we detected robust but transitory viremias that lasted for 9–11 days. We completed the SFTSV transmission cycle between hedgehogs and nymph and adult H. longicornis ticks under laboratory conditions with 100% efficiency. Furthermore, naive H. longicornis ticks could be infected by SFTSV-positive ticks co-feeding on naive hedgehogs; we confirmed transstadial transmission of SFTSV. Our study suggests that the hedgehogs are a notable wildlife amplifying host of SFTSV in China.

In this study, the ultrasound-assisted extraction (UAE) conditions of flavonoids from Lactuca indica L.cv. Mengzao (LIM) leaves was optimized, and the flavonoids content and their antioxidant potential in different parts were analyzed. The optimal extraction parameters to obtain the highest total flavonoids content (TFC) were a a ratio of liquid to solid of 24.76 mL/g, ultrasonic power of 411.43 W, ethanol concentration of 58.86% and an extraction time of 30 min, the average TFC of LIM leaves could reach 48.01 mg/g. For the yield of flavonoids, the UAE method had the best extraction capacity compared with solvent extraction and microwave-assisted extraction (MAE). In general, the TFC in different parts of LIM followed the order flower > leaf > stem > root, the flowering period is the most suitable harvesting period. From ultra-high performance liquid chromatography-mass spectrometry (UPLC–MS) quantification, the flower samples showed significantly higher six flavonoids and had the highest radical scavenging capacities compared to other samples. A high positive correlation was observed between the antioxidant activity and TFC, luteolin-7-O-glucoside and rutin were significantly (p < 0.05) correlated with all antioxidant evaluations. This study provides valuable information for the development and utilization of flavonoids in Lactuca indica as ingredients in food, feed and nutritional health products.

Dysfunction of vascular endothelial cells (ECs) facilitates imbalanced immune responses and tissue hyperinflammation. However, the heterogeneous functions of skin ECs and their underlying mechanism in dermatoses remain to be determined. Here, focusing on the pathogenic role of skin ECs in psoriasis, we characterized the molecular and functional heterogeneity of skin ECs from healthy individuals and psoriasis patients at the single-cell level. We found that endothelial glycocalyx destruction, a major feature of EC dysfunction in psoriasis, was a driving force during the process of T cell extravasation. Interestingly, we identified a skin EC subset, IGFBP7hi ECs, in psoriasis. This subset actively responded to psoriatic-related cytokine signaling, secreted IGFBP7, damaged the endothelial glycocalyx, exposed the adhesion molecules underneath, and prepared the endothelium for immune-cell adhesion and transmigration, thus aggravating skin inflammation. More importantly, we provided evidence in a psoriasis-like mouse model that anti-IGFBP7 treatment showed promising therapeutic effects for restoring the endothelial glycocalyx and alleviating skin inflammation. Taken together, our results depict the distinct functions of EC clusters in healthy and psoriatic skin, identify IGFBP7hi ECs as an active subset modulating vascular function and cutaneous inflammation, and indicate that targeting IGFBP7 is a potential therapeutic strategy in psoriasis.

Background: The present study aimed to prove the progression of immunoglobulin A nephropathy (IgAN) patients with isolated hematuria based on repeat renal biopsy data for the first time.Methods: 29 IgAN patients with isolated hematuria who received repeat renal biopsies were analyzed retrospectively, while 29 non-isolated hematuria IgAN patients with similar age and background were randomly selected as the control group. Clinical parameters were collected at the time of biopsy. The treatment strategies (conservative treatment with RASS blocker or immunosuppressive treatment) were choosen according to the pathological results at the first renal biopsy. The activity and chronicity indexes of renal lesions were evaluated. Markers of cell inflammation and proliferation were tseted by immunochemistry. The ultrastructure of podocytes was observed by transmission electron microscopy (TEM). Podocyte and oxidative stress marker (NPHS2 and 4-HNE) were detected by immunofluorescence.Results: The IgAN patients with isolated hematuria had better clinical indicators than those with no-isolated hematuria, such as better renal function, higher albumin and lower uric acid. The interval between two biopsies in IgAN patients with isolated hematuria was 630 (interquartile range, 409.5–1,171) days. The hematuria of the patients decreased significantly from 30 (IQR, 4.00–35.00) RBC/ul in the first biopsy to 11 (IQR, 2.50–30.00) RBC/ul in the repeated biopsy (p < 0.05). The level of triglyceride decreased significantly (p < 0.05). The other clinical indicators were not statistically significant (p > 0.05). Deposits of IgA and C3 in the glomerulus were persistent. The activity index decreased, especially cellular crescent formation, while the chronicity index increased. The ultrastructure of podocytes was improved after treatment. The oxidative stress products of podocytes reduced after treatment.Conclusion: Although the clinical indicators of the IgAN patients with isolated hematuria were in the normal range, various acute and chronic pathological changes have occurred, and irreversible chronic changes have been progressing. Cell inflammation and proliferation persisted. Oxidative stress of podocytes was likely to be the therapeutic target. This study provided a strong basis for the progress of IgAN with isolated hematuria through pathological changes before and after treatment. This study will help clinicians recognize the harm of hematuria, change the traditional treatment concept, and help such patients get early treatment.

IntroductionItalian ryegrass is less studied in northern China due to high-quality forage grass has not been fully utilized. Full utilization of high-quality forage grass helps to alleviate the shortage of forage grass in winter and spring season and guarantee stable development of livestock production. Consequently, this study was aimed to evaluate the effects of different regions in northern China on the fermentative products, bacterial community compositions, and metabolic pathways and metabolites of Italian ryegrass silage.MethodsThe Italian ryegrass was harvested from three regions (Ordos-WK; Hohhot-AK; Ulanqab-SYK) and ensiled for 60 days. Single molecule real-time (SMRT) sequencing and ultra-high performance liquid chromatography-mass spectrometry (UHPLC–MS/MS) were used to analyze bacterial communities and metabolites, respectively.ResultsAfter 60 d of fermentation, the SYK group had the lowest pH (4.67), the highest lactic acid contents (95.02 g/kg DM) and largest lactic acid bacteria populations (6.66 log10 cfu/g FM) among the treatment groups. In addition, the SYK group had the highest abundance of Lactiplantibacillus plantarum (63.98%). In SYK group, isoquinoline alkaloid biosynthesis was the significantly enriched (p

IntroductionThis study aimed to investigate the effects of different growth stages (booting period-SYK; initial flowering-SCK; full flowering-SSK) on the fermentation quality, microbial community, metabolic pathways and metabolomic characteristics of Italian ryegrass silage.MethodsSingle molecule real-time (SMRT) sequencing and ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS) were used to analyze bacterial communities and metabolites, respectively.ResultsAfter 60 d of fermentation, SYK had the lowest pH and the highest lactic acid content, which were significantly different from the other groups. The bacteria with the highest abundance in SYK, SCK and SSK groups were Lactiplantibacillus plantarum (63.98%), Weissella minor (28.82%) and Levilactobacillus brevis (64.81%), respectively. In addition, among the main differential metabolites in different growth stages, the number of amino acids was the most, and the corresponding metabolic pathways were mainly amino acid metabolic pathways. The biosynthesis of phenylalanine, tyrosine and tryptophan was significantly enriched (p

ObjectiveThe objective of this study was to determine the effect of Lactobacillus plantarum (L. plantarum) and propionic acid (PA) on the microbial community and fermentation performance of high-moisture amaranth silage.MethodsAmaranth silages were rown without addition (AhGCK) as a control and with L. plantarum JYLP-002 (AhGLP) or propionic acid (AhGPA) and then were opened after 60 days of ensiling to determine the microbial community and fermentation quality.ResultsCrude protein (CP) content, lactic acid (LA) content, and lactic acid bacteria (LAB) counts were significantly higher in AhGLP and AhGPA compared with those in AhGCK (p < 0.05). In contrast, pH, acetic acid (AA) content, and yeast and aerobic bacteria counts were significantly lower in AhGLP and AhGPA compared with those in AhGCK (p < 0.05). In addition, propionic acid (PA) levels were markedly higher in AhGPA (p < 0.05). In terms of microbial communities, the silage in the additive groups showed an increased relative abundance of Lactiplantibacillus plantarum and Lentilactobacillus buchneri and a reduced relative abundance of Enterobacter cloacae and Clostridium tyrobutyricum. The abundance of Xanthomonas oryzae was significantly increased in AhGPA, but completely inhibited in the silage supplemented with L. plantarum. Spearman’s correlation analysis revealed that Lentilactobacillus buchneri and Levilactobacillus brevis were positively associated with LA and negatively associated with pH. Conversely, Clostridium tyrobutyricum and Enterobacter cloacae were negatively associated with LA, but positively associated with pH and AA content. AA content was inversely correlated with Lentilactobacillus buchneri. Functional prediction analysis showed that LAB dominated the three groups of silage and the silages containing additives had improved carbohydrate and amino acid metabolism compared with the control silage; in particular, the AhGLP group had more heterotypic fermentation processes and a richer metabolic pathway. Furthermore, the epiphytic Lactiplantibacillus plantarum and Lentilactobacillus buchneri could inhibit the reproductive activity of undesirable microorganisms to a certain extent, thus slowing the spoilage process of the silage.ConclusionIn conclusion, L. plantarum can improve fermentation characteristics by modulating the microbial community attached to high-moisture amaranth silage and will prove useful for preserving high-moisture silage.

Abstract SARS-CoV-2 and SARS-CoV are genetically related coronavirus and share the same cellular receptor ACE2. By replacing the VSV glycoprotein with the spikes (S) of SARS-CoV-2 and SARS-CoV, we generated two replication-competent recombinant viruses, rVSV-SARS-CoV-2 and rVSV-SARS-CoV. Using wild-type and human ACE2 (hACE2) knock-in mouse models, we found a single dose of rVSV-SARS-CoV could elicit strong humoral immune response via both intranasal (i.n.) and intramuscular (i.m.) routes. Despite the high genetic similarity between SARS-CoV-2 and SARS-CoV, no obvious cross-neutralizing activity was observed in the immunized mice sera. In macaques, neutralizing antibody (NAb) titers induced by one i.n. dose of rVSV-SARS-CoV-2 were eight-fold higher than those by a single i.m. dose. Thus, our data indicates that rVSV-SARS-CoV-2 might be suitable for i.n. administration instead of the traditional i.m. immunization in human. Because rVSV-SARS-CoV elicited significantly stronger NAb responses than rVSV-SARS-CoV-2 in a route-independent manner, we generated a chimeric antigen by replacing the receptor binding domain (RBD) of SARS-CoV S with that from the SARS-CoV-2. rVSV expressing the chimera (rVSV-SARS-CoV/2-RBD) induced significantly increased NAbs against SARS-CoV-2 in mice and macaques than rVSV-SARS-CoV-2, with a safe Th1-biased response. Serum immunized with rVSV-SARS-CoV/2-RBD showed no cross-reactivity with SARS-CoV. hACE2 mice receiving a single i.m. dose of either rVSV-SARS-CoV-2 or rVSV-SARS-CoV/2-RBD were fully protected against SARS-CoV-2 challenge without obvious lesions in the lungs. Our results suggest that transplantation of SARS-CoV-2 RBD into the S protein of SARS-CoV might be a promising antigen design for COVID-19 vaccines.

In this study, we analyzed the fermentation quality, microbial community, and metabolome characteristics of ryegrass silage from different harvests (first harvest-AK, second harvest-BK, and third harvest-CK) and analyzed the correlation between fermentative bacteria and metabolites. The bacterial community and metabolomic characteristics were analyzed by single-molecule real-time (SMRT) sequencing and ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS/MS), respectively. After 60 days of ensiling, the pH of BK was significantly lower than those of AK and CK, and its lactic acid content was significantly higher than those of AK and CK. Lactiplantibacillus and Enterococcus genera dominate the microbiota of silage obtained from ryegrass harvested at three different harvests. In addition, the BK group had the highest abundance of Lactiplantibacillus plantarum (58.66%), and the CK group had the highest abundance of Enterococcus faecalis (42.88%). The most annotated metabolites among the differential metabolites of different harvests were peptides, and eight amino acids were dominant in the composition of the identified peptides. In the ryegrass silage, arginine, alanine, aspartate, and glutamate biosynthesis had the highest enrichment ratio in the metabolic pathway of KEGG pathway enrichment analysis. Valyl-isoleucine and glutamylvaline were positively correlated with Lactiplantibacillus plantarum. D-Pipecolic acid and L-glutamic acid were positively correlated with Levilactobacillus brevis. L-phenylalanyl-L-proline, 3,4,5-trihydroxy-6-(2-methoxybenzoyloxy) oxane-2-carboxylic acid, and shikimic acid were negatively correlated with Levilactobacillus brevis. In conclusion, this study explains the effects of different harvest frequencies on the fermentation quality, microbial community, and metabolites of ryegrass, and improves our understanding of the ensiling mechanisms associated with different ryegrass harvesting frequencies.

In this study, the triterpenoids in the leaves of Lactuca indica L.cv. Mengzao (LIM) were extracted via microwave-assisted ethanol extraction, and the optimum extraction conditions for triterpenoids were determined through single-factor experiments and the Box–Behnken method. The effects of three factors (solid–liquid ratio, microwave power and extraction time) on the total triterpenoids content (TTC) were evaluated. The TTC of different parts (roots, stems, leaves and flowers) of LIM in different growth stages was studied, and the scavenging effects of the highest TTC parts on DPPH, ABTS and hydroxyl free radicals were investigated. The results showed that the optimum extraction conditions for microwave-assisted extraction of total triterpenoids from LIM leaves were as follows: solid–liquid ratio of 1:20 g/mL; microwave power of 400 W; and extraction time of 60 min. Under these conditions, the TTC was 29.17 mg/g. Compared with the fresh raw materials, the TTC of the materials increased after freeze drying. The leaves of LIM had the highest TTC, and the flowering stage was the best time. The triterpenoids from the leaves had a strong ability to eliminate DPPH and ABTS free radicals, and the elimination effect of dried leaves was better than that of fresh leaves, while the elimination effect of hydroxyl free radicals was not obvious. The tested method was used to extract total triterpenoids from LIM using a simple process at low cost, which provides a reference for developing intensive processing methods for L. indica.

Several mRNA-based vaccines for SARS-CoV-2 are in late phase clinical development. Here, the authors show that a single immunization with a mRNA vaccine expressing SARS-CoV-2 spike RBD induces neutralizing antibodies that are maintained for at least 6.5 months and confer protection in a sera transfer study in mice.

BackgroundMacrophages are the most abundant infiltrating immune-related stromal cells present in and around tumors, showing different phenotypes and functions. M2 macrophages mainly exert immunosuppressive functions and promote tumor growth. Exosomes are emerging as important mediators of cross-talk between tumor cells and the microenvironment. CircRNAs are novel members of non-coding RNAs that regulate cancer proliferation and progression. However, the mechanism by which exosomal circRNA regulates macrophage polarization in renal cell carcinoma (RCC) is still largely unknown.MethodsRCC-derived exosomes were characterized using transmission electron microscopy and nanoparticle tracking analysis (NTA). CCK-8, wound healing, and Transwell assays were performed to assess whether exosomes would affect the proliferation, migration, and invasion of RCC. Furthermore, we performed a bioinformatics analysis to identify circRNAs in RCC serum-derived exosomes from the GEO database. The fluorescence in situ hybridization (FISH) assay was used to detect the cellular distribution of circSAFB2. Bioinformatics analyses (StarBase 2.0) were used to pool the miRNA targets of circSAFB2. Luciferase assays were performed to verify the direct interactions. Western blotting was used to detect markers of macrophage M2 polarization. Lastly, mouse xenograft and bioluminescence imaging were used to examine the clinical relevance of exosomal circSAFB2 in vivo.ResultsWe report the circRNA derived from SAFB2 and evaluate its biological function in promoting the immune escape of RCC. We found that circSAFB2 was highly expressed in RCC tissues and RCC-derived exosomes. Furthermore, we demonstrated that exosomal circSAFB2 mediates the polarization of M2 macrophages through the miR-620/JAK1/STAT3 axis to promote RCC metastasis.ConclusionsOur data first demonstrated that circSAFB2 leads to immune escape from RCC by mediating M2 macrophage polarization via the miR-620/JAK1/STAT3 axis. These findings indicate a novel molecular mechanism of exosomal circSAFB2 in the progression of RCC and implicate circSAFB2 as a target for exosome-mediated tumor immune evasion.

Summary: Helminths and helminth-derived products hold promise for treating joint bone erosion in rheumatoid arthritis (RA). However, the mechanisms of helminths ameliorating the osteoclastic bone destruction are incompletely understood. Here, we report that Trichinella spiralis infection or treatment with the excreted/secreted products of T. spiralis muscle larvae (MES) attenuated bone erosion and osteoclastogenesis in mice with collage-induced arthritis (CIA) through inhibiting M1 monocyte/macrophage polarization and the production of M1-related proinflammatory cytokines. In vitro, MES inhibited LPS-induced M1 macrophage activation while promoting IL-4-induced M2 macrophage polarization. Same effects of MES were also observed in monocytes derived from RA patients, wherein MES treatment suppressed LPS-induced M1 cytokine production. Moreover, MES treatment attenuated LPS and RANKL co-stimulated osteoclast differentiation from the RAW264.7 macrophages through inhibiting activation of the NF-κB rather than MAPK pathway. This study provides insight into the M1 subset as a potential target for helminths to alleviate osteoclastic bone destruction in RA.

Ensiling native grass is an effective method to protect the nutritional quality of forage and alleviate feed shortages in the cold winter of the Inner Mongolian Plateau. To improve the usability of native grass resources as feed in China, the effects of lactic acid bacteria and molasses additions on the microbial population, fermentation quality, and nutritional quality of native grass during silage were investigated. Treatments were a control treatment with no additive (CK), lactic acid bacteria (L), molasses (M), and lactic acid bacteria in combination with molasses (L+M), all of which were stored at ambient temperature (17–28°C) for 7, 14, 30, and 60 days. The results showed that all additives improved nutritional value and fermentation quality with low pH and ammonia nitrogen (NH3–N) and high crude protein (CP) and water soluble carbohydrate (WSC) than control silage over the ensiling period. Compared with L or M silage, the L+M silage combination improved fermentability, as evidenced by higher LA content and a faster pH drop during the first 7 days of ensiling. With prolonged ensiling time, the combined addition of L and M could increase the count of desirable Lactobacillus, decrease microbial diversity, and inhibit the growth of undesirable microorganism, such as Clostridia, Escherichia, and Enterobacter abundance compared with silage treated with CK, L. or M. Application of L together with M could further improve the silage quality of native grass by altering bacterial community structure. In summary, the addition of lactic acid bacteria and molasses increased the relative abundance of Lactobacillus of native grass silage and improved fermentation quality.

Electronic tongues and artificial gustation for crucial analytes in the environment, such as metal ions, are becoming increasingly important. In this contribution, we propose a multi-level fusion framework for a hybrid impedimetric and voltammetric electronic tongue to enhance the accuracy of K+, Mg2+, and Ca2+ detection in an extensive concentration range (100.0 nM–1.0 mM). The proposed framework extracts electrochemical-based features and separately fuses, in the first step, impedimetric features, which are characteristic points and fixed frequency features, and the voltammetric features, which are current and potential features, for data reduction by LDA and classification by kNN. Then, in a second step, a decision fusion is carried out to combine the results for both measurement methods based on Dempster–Shafer (DS) evidence theory. The classification results reach an accuracy of 80.98% and 81.48% for voltammetric measurements and impedimetric measurements, respectively. The decision fusion based on DS evidence theory improves the total recognition accuracy to 91.60%, thus realizing significantly high accuracy in comparison to the state-of-the-art. In comparison, the feature fusion for both voltammetric and impedimetric features in one step reaches an accuracy of only 89.13%. The proposed hierarchical framework considers for the first time the fusion of impedimetric and voltammetric data and features from multiple electrochemical sensor arrays. The developed approach can be implemented for several further applications of pattern fusion, e.g., for electronic noses, measurement of environmental contaminants such as heavy metal ions, pesticides, explosives, and measurement of biomarkers, such as for the detection of cancers and diabetes.

Increasing evidence has shown that the metabolism and clearance of molecular targeted agents, such as sorafenib, plays an important role in mediating the resistance of HCC cells to these agents. Metabolism of sorafenib is performed by oxidative metabolism, which is initially mediated by CYP3A4. Thus, targeting CYP3A4 is a promising approach to enhance the sensitivity of HCC cells to chemotherapeutic agents. In the present work, we examined the association between CYP3A4 and the prognosis of HCC patients receiving sorafenib. Using the online tool miRDB, we predicted that has-microRNA-4277 (miR-4277), an online miRNA targets the 3’UTR of the transcript of cyp3a4. Furthermore, overexpression of miR-4277 in HCC cells repressed the expression of CYP3A4 and reduced the elimination of sorafenib in HCC cells. Moreover, miR-4277 enhanced the sensitivity of HCC cells to sorafenib in vitro and in vivo. Therefore, our results not only expand our understanding of CYP3A4 regulation in HCC, but also provide evidence for the use of miR-4277 as a potential therapeutic in advanced HCC.

Harvest time and storage time had significant effects on nutrient quality and Volatile Organic Compounds (VOCs) of alfalfa. The objective of this study was to use headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) to analyze alfalfa at different harvest periods (budding stage, early blooming, full blooming), and storage for 0 d, 90 d, 180 d, 270 d, and 360 d, the dynamic changes in VOCs and nutritional quality. Results indicated that ketones, aldehydes, alcohols and esters were the main volatile components of alfalfa VOCs, accounting for 87.41%, 88.57% and 90.85% of the total volatile components at budding stage, early blooming and full blooming, respectively. VOCs and nutrient quality of alfalfa varied significantly in different harvest periods; delayed harvesting significantly reduced alfalfa aldehydes, alcohols, crude protein (CP), and total digestible nutrients (TDN) and significantly increased ketone, dry matter (DM), acid detergent fiber (ADF) and neutral detergent fiber (NDF) contents (p < 0.05). After Comprehensive consideration ofthe yield, nutritional quality and VOCs of alfalfa, the best harvest time was determined to be the budding stage. The VOCs and nutrient quality of alfalfa were significantly different at different storage durations, and ketones, ADF and NDF were significantly increased and significantly reduced DM, CP and TDN (p < 0.05). Ketone content and neutral detergent fiber (NDF) content had a significant positive correlation, (NDF F = 5.5, p = 0.024). Storage 360 d 2-methylbutyraldehyde (musty), 1-octen-3-one (earthy musty odor) content increased. These may be the key compounds that causes the smell of fresh alfalfa to fade away and a musty smell to emerge.

Background: Sirtuin 5 (SIRT5) is a NAD+-dependent lysine deacylase. The SIRT5 deficiency mouse model shows that it is dispensable for metabolic homeostasis under normal conditions. However, the biological role of SIRT5 and acylation in pathological states such as obesity and type 2 diabetes (T2D) remains elusive. Methods: The hepatic SIRT5-overexpressing ob/ob mouse model (ob/ob-SIRT5 OE) was established by CRISPR/Cas9 gene editing tool Protein malonylation and succinylation lysine sites were identified by immunoprecipitation coupled lipid chromatography - tandem mass spectrometry (LC-MS/MS) methods. Findings: The ob/ob-SIRT5 OE mice showed decreased malonylation and succinylation, improved cellular glycolysis, suppressed gluconeogenesis, enhanced fatty acid oxidation, and attenuated hepatic steatosis. A total of 955 malonylation sites on 434 proteins and 1377 succinylation sites on 429 proteins were identified and quantitated. Bioinformatics analysis revealed that malonylation was the major SIRT5 target in the glycolysis/gluconeogenesis pathway, whereas succinylation was the preferred SIRT5 target in the oxidative phosphorylation pathway. Interpretation: Hepatic overexpression of SIRT5 ameliorated the metabolic abnormalities of ob/ob mice, probably through demalonylating and desuccinylating proteins in the main metabolic pathways. SIRT5 and related acylation might be potential targets for metabolic disorders. Fund: National Key R&D Program of China, the National Natural Science Foundation of China, the Strategic Priority Research Programs (Category A) of the Chinese Academy of Sciences, the Interdisciplinary Medicine Seed Fund of Peking University and the National Laboratory of Biomacromolecules. Keywords: Sirtuins, SIRT5, Malonylation, Succinylation, Metabolism, Proteomics

Embryonic stem cell pluripotency depends upon precise regulation by a core transcription network. Here the authors show that polyglutamylation mediated stabilization of the transcription factor Klf4 by TTLL1 and TTLL4 promotes reprogramming, pluripotency and preimplantation embryonic development.

We investigated functionalized graphene materials to create highly sensitive sensors for volatile organic compounds (VOCs) such as formaldehyde, methanol, ethanol, acetone, and isopropanol. First, we prepared VOC-sensitive films consisting of mechanically exfoliated graphene (eG) and chemical graphene oxide (GO), which have different concentrations of structural defects. We deposited the films on silver interdigitated electrodes on Kapton substrate and submitted them to thermal treatment. Next, we measured the sensitive properties of the resulting sensors towards specific VOCs by impedance spectroscopy. We obtained the eG- and GO-based electronic nose composed of two eG films- and four GO film-based sensors with variable sensitivity to individual VOCs. The smallest relative change in impedance was 5% for the sensor based on eG film annealed at 180 °C toward 10 ppm formaldehyde, whereas the highest relative change was 257% for the sensor based on two-layers deposited GO film annealed at 200 °C toward 80 ppm ethanol. At 10 ppm VOC, the GO film-based sensors were sensitive enough to distinguish between individual VOCs, which implied excellent selectivity, as confirmed by Principle Component Analysis (PCA). According to a PCA-Support Vector Machine-based signal processing method, the electronic nose provided identification accuracy of 100% for individual VOCs. The proposed electronic nose can be used to detect multiple VOCs selectively because each sensor is sensitive to VOCs and has significant cross-selectivity to others.

L. indica L. cv. Mengzao, a medicinal plant of the Ixeris genus, is rich in flavonoids. In order to thoroughly analyze the the distribution and dynamic change of major flavonoids in its various parts from different growth periods, the flavonoids extracted from L. indica L. cv. Mengzao were identified and quantitatively analyzed by ultra-high-performance liquid chromatography mass spectrometer (LC-MS/MS). Results indicated that 15 flavonoids were identified from L. indica L. cv. Mengzao, and rutin, luteolin, luteolin-7-O-glucoside, kaempferol, quercetin, and apigenin are the major flavonoids in L. indica L. cv. Mengzao. In general, the total flavonoids’ content in different parts of L. indica L. cv. Mengzao followed the order flowers > leaves > stems > roots. Flowers and leaves are the main harvesting parts of L. indica L. cv. Mengzao, and the flowering period is the most suitable harvesting period. This study provides valuable information for the development and utilization of L. indica L. cv. Mengzao and determined the best part to harvest and the optimal time for harvesting.

Background/Aims: Renal tubular epithelial-mesenchymal transition (EMT) is regarded as an important factor leading to renal interstitial fibrosis. Erythropoietin (EPO) has been reported to attenuate renal fibrosis. The mechanism underlying this protective effect of EPO remains unclear. In this study, we aim to identify possible mechanisms of the EPO renoprotective effect. Methods: Hypoxia was induced in vitro by incubating human proximal tubular epithelial cell line HK-2 cells in 1% O2 and 5% CO2. Western blotting and reverse transcription polymerase chain reaction analyses were used to evaluate the expression of epithelial and mesenchymal markers in the cell samples. The expression of miR-200b in the HK-2 cells under hypoxia or treatment with EPO was examined. Results: EPO represses hypoxia-induced EMT by upregulating miR-200b in HK-2 cells. Overexpression of miR-200b represses the effect of ETS proto-oncogene 1 (Ets-1)-induced EMT in HK-2 cells. Conclusion: miR-200 mediates the protective effects of EPO on EMT in hypoxic HK-2 cells. EPO attenuated hypoxia-induced EMT by increasing miR-200 expression via the repression of Ets-1.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-021-22820-x

Lumican (LUM) is a candidate gene for myopia in the MYP3 locus. In this study, a mutant lumican (L199P) transgenic mouse model was established to investigate the axial length changes and ultrastructural features of the sclera. The mouse model was established by pronuclear microinjection. Transgenic mice and wild-type B6 mice were killed at eight weeks of age. Gene expression levels of LUM and collagen type I (COL1) in the sclera were analyzed by quantitative real-time polymerase chain reaction (qPCR), and the protein levels were assessed by Western blot analysis. Ocular axial lengths were measured on the enucleated whole eye under a dissecting microscope. Ultrastructural features of collagen fibrils in the sclera were examined with transmission electron microscopy (TEM). Lumican and collagen type I were both elevated at the transcriptional and protein levels. The mean axial length of eyes in the transgenic mice was significantly longer than that in the wild-type mice (3,231.0 ± 11.2 μm (transgenic group) vs 3,199.7 ± 11.1 μm (controls), p