Harnessing Natural Products by a Pharmacophore-Oriented Semisynthesis Approach for the Discovery of Potential Anti-SARS-CoV-2 Agents.

Natural products possessing unique scaffolds may have antiviral activity but their complex structures hinder facile synthesis. A pharmacophore-oriented semisynthesis approach was applied to (-)-maoelactone A (1) and oridonin (2) for the discovery of anti-SARS-CoV-2 agents. The Wolff rearrangement/lactonization cascade (WRLC) reaction was developed to construct the unprecedented maoelactone-type scaffold during semisynthesis of 1. Further mechanistic study suggested a concerted mechanism for Wolff rearrangement and a water-assisted stepwise process for lactonization. The WRLC reaction then enabled the creation of a novel family by assembly of the maoelactone-type scaffold and the pharmacophore of 2, whereby one derivative inhibited SARS-CoV-2 replication in HPA EpiC cells with a low EC50 value (19±1 nM) and a high TI value (>1000), both values better than those of remdesivir.

Chaetolactam A, an Azaphilone Derivative from the Endophytic Fungus Chaetomium sp. g1.

Chaetolactam A (1), an unprecedented azaphilone derivative bearing a unique 9-oxa-7-azabicyclo[4.2.1]octan-8-onering system, together with two new compounds, 11-epi-chaetomugilide B (2) and chaetomugilide D (3) was isolated from an endophytic fungus, Chaetomium sp. g1. Notably, extensive NMR data analyses, NMR calculations with DP4 and DP4+ analyses, ECD calculations, and the RDC method were employed to establish the structure of 1. Furthermore, 2 exhibited potent apoptosis induction activity by mediating caspase-3 activation and PARP degradation at 3 µM in HL-60.

3-Hydroxy-4-methyldecanoic Acid-Containing Cyclotetradepsipeptides from an Endolichenic Beauveria sp.

An investigation of an endolichenic Beauveria sp. led to the discovery of seven new cyclotetradepsipeptides, beauveamides A-G (2-8), along with the known beauverolide Ka (1). All incorporate a 3-hydroxy-4-methyldecanoic acid (HMDA) moiety in their structures. Their configuration was determined through Marfey's, J-based configuration analysis, and NMR computational methods, representing the first time that the stereostructures of HMDA-moiety-containing cyclotetradepsipeptides have been established. Compounds 1 and 2 exhibited protecting effects on HEI-OC1 cells at 10 µM, while 1, 4, and 5 could stimulate glucose uptake in cultured rat L6 myoblasts at 50 µM. Compound 1 showed dose-dependent activity in both L6 myoblasts and myotubes.

Zfp217 mediates m6A mRNA methylation to orchestrate transcriptional and post-transcriptional regulation to promote adipogenic differentiation.

A complex and highly orchestrated gene expression program chiefly establishes the properties that define the adipocyte phenotype, in which the vast majority of factors are involved in transcriptional regulation. However, the mechanisms by post-transcriptional modulation are poorly understood. Here, we showed that zinc finger protein (Zfp217) couples gene transcription to m6A mRNA modification to facilitate adipogenesis. Zfp217 modulates m6A mRNA methylation by activating the transcription of m6A demethylase FTO. Consistently, depletion of Zfp217 compromises adipogenic differentiation of 3T3L1 cells and results in a global increase of m6A modification. Moreover, the interaction of Zfp217 with YTHDF2 is critical for allowing FTO to maintain its interaction with m6A sites on various mRNAs, as loss of Zfp217 leads to FTO decrease and augmented m6A levels. These findings highlight a role for Zfp217-dependent m6A modification to coordinate transcriptional and post-transcriptional regulation and thus promote adipogenic differentiation.

Circulating Exosomal miR-221 from Maternal Obesity Inhibits Angiogenesis via Targeting Angptl2.

Maternal obesity disrupts both placental angiogenesis and fetus development. However, the links between adipocytes and endothelial cells in maternal obesity are not fully understood. The aim of this study was to characterize exosome-enriched miRNA from obese sow's adipose tissue and evaluate the effect on angiogenesis of endothelial cells. Plasma exosomes were isolated and analyzed by nanoparticle tracking analysis (NTA), electron morphological analysis, and protein marker expression. The number of exosomes was increased as the gestation of the sows progressed. In addition, we found that exosomes derived from obese sows inhibited endothelial cell migration and angiogenesis. miRNA detection showed that miR-221, one of the miRNAs, was significantly enriched in exosomes from obese sows. Further study demonstrated that exosomal miR-221 inhibited the proliferation and angiogenesis of endothelial cells through repressing the expression of Angptl2 by targeting its 3' untranslated region. In summary, miR-221 was a key component of the adipocyte-secreted exosomal vesicles that mediate angiogenesis. Our study may be a novel mechanism showing the secretion of "harmful" exosomes from obesity adipose tissues causes placental dysplasia during gestation.

(+)- and (-)-Alternarilactone A: Enantiomers with a Diepoxy-Cage-like Scaffold from an Endophytic Alternaria sp.

The enantiomers (+)- and (-)-alternarilactone A (1), the first examples of dibenzo-α-pyrones bearing a diepoxy-cage-like moiety, were isolated from the endophytic fungus Alternaria sp. hh930. The deficiency in 1H-1H COSY and HMBC correlations caused by the highly oxidized caged system of 1 and the deceptive and ambiguous signals such as "W" couplings in NMR data increased the risk of structure misassignment of 1. By performing a quantum chemical calculation of the NMR chemical shifts together with a DP4+ probability analysis and single-crystal X-ray crystallographic experiment, their structures were unambiguously determined, and their absolute configurations were determined by ECD calculations.

miR-221 negatively regulates inflammation and insulin sensitivity in white adipose tissue by repression of sirtuin-1 (SIRT1).

It is well known that obesity-induced white adipose tissue inflammation is an important reason for insulin-resistance and type 2 diabetes mellitus. Sirtuin-1 (SIRT1) is an important regulator of inflammtion response pathways in white adipose tissue. Here, we found that miR-221 negatively regulated SIRT1 in white adipose tissue during inflammation and HFD-induced obesity. MiR-221 is a putative oncogene which has been found overexpressed in a number of human tumors. Recently, it has also found that miR-221 was increased in obese adipose tissue and may be involved in inflammation and insulin-resistance. However the specific mechanism remains to be elucidated. In our present study, we found that overexpression of miR-221 decreased the protein abundance of SIRT1 and caused inflammation and insulin-resistance in differentiated 3T3-L1 cells. Conversely, miR-221 inhibition increased the protein levels, ameliorated inflammation, and improved insulin sensitivity. Moreover, inhibition of SIRT1 by EX527 significantly diminished the downregulation of the inflammation and insulin-resistance levels induced by the miR-221 inhibitor. In conclusion, our data suggest that miR-221 promotes white adipose tissue inflammation and decreases insulin sensitivity in obesity, at least in part, through suppressing SIRT1.

GPR120: a critical role in adipogenesis, inflammation, and energy metabolism in adipose tissue.

It is well known that adipose tissue has a critical role in the development of obesity and metabolic diseases and that adipose tissue acts as an endocrine organ to regulate lipid and glucose metabolism. Accumulating in the adipose tissue, fatty acids serve as a primary source of essential nutrients and act on intracellular and cell surface receptors to regulate biological events. G protein-coupled receptor 120 (GPR120) represents a promising target for the treatment of obesity-related metabolic disorders for its involvement in the regulation of adipogenesis, inflammation, glucose uptake, and insulin resistance. In this review, we summarize recent studies and advances regarding the systemic role of GPR120 in adipose tissue, including both white and brown adipocytes. We offer a new perspective by comparing the different roles in a variety of homeostatic processes from adipogenic development to adipocyte metabolism, and we also discuss the effects of natural and synthetic agonists that may be potential agents for the treatment of metabolic diseases.

Lower dietary n-6 : n-3 ratio and high-dose vitamin E supplementation improve sperm morphology and oxidative stress in boars.

A 2×2 factorial experiment (10 boars per treatment) was conducted for 16 weeks to evaluate the effects of the dietary n-6:n-3 ratio (14:1 vs 6:1) and vitamin E (200 vs 400mg kg-1) on boar sperm morphology and oxidative stress. Sperm mitochondrial membrane potential (MMP), reactive oxygen species (ROS), DNA damage (8-hydroxydeoxyguanosine; 8-OHdG), seminal lipoperoxidation (malondialdehyde; MDA) and antioxidant capacity in the serum, spermatozoa and seminal plasma were assessed as indicators of oxidative stress. Sperm production was similar among groups but increased (P<0.05) throughout the 16 weeks of the study. Although sperm α-tocopherol content, ROS and seminal MDA did not differ between the two dietary n-6:n-3 ratio treatments, enhanced antioxidant enzyme activity and MMP, but decreased 8-OHdG, were found in spermatozoa from boars consuming the 6:1 diet. The diet with the 6:1 ratio positively affected sperm morphology at Weeks 12 and 16 (P<0.05). The α-tocopherol content and antioxidant capacity increased in boars with increasing levels of vitamin E supplementation. Compared with low-dose vitamin E, high-dose vitamin E supplementation improved sperm morphology. Overall, the results indicate that an n-6:n-3 ratio of 6:1 and 400 mg/kg vitamin E have beneficial effects on sperm morphology by improving antioxidative stress.

Gypmacrophin A, a Rare Pentacyclic Sesterterpenoid, Together with Three Depsides, Functioned as New Chemical Evidence for Gypsoplaca macrophylla (Zahlbr.) Timdal Identification.

The phytochemical investigation on 1 g of materials from Gypsoplacamacrophylla (Zahlbr.) Timdal resulted in the discovery of gypmacrophin A, a rare pentacyclic sesterterpenoid; brialmontin III, a new polysubstituted depside and two known ones, brialmontins I and II. The structure and absolute configurations of gypmacrophin A were elucidated by spectroscopic analyses and computational methods. Gypmacrophin A showed weak inhibition of AchE with an IC50 value of 32.03 µM. The four compounds provided new chemical evidence for G. macrophylla identification.

Methionine Regulates mTORC1 via the T1R1/T1R3-PLCß-Ca2+-ERK1/2 Signal Transduction Process in C2C12 Cells.

The mammalian target of rapamycin complex 1 (mTORC1) integrates amino acid (AA) availability to support protein synthesis and cell growth. Taste receptor type 1 member (T1R) is a G protein-coupled receptor that functions as a direct sensor of extracellular AA availability to regulate mTORC1 through Ca2+ stimulation and extracellular signal-regulated kinases 1 and 2 (ERK1/2) activation. However, the roles of specific AAs in T1R1/T1R3-regulated mTORC1 are poorly defined. In this study, T1R1 and T1R3 subunits were expressed in C2C12 myotubes, and l-AA sensing was accomplished by T1R1/T1R3 to activate mTORC1. In response to l-AAs, such as serine (Ser), arginine (Arg), threonine (Thr), alanine (Ala), methionine (Met), glutamine (Gln), and glycine (Gly), Met induced mTORC1 activation and promoted protein synthesis. Met also regulated mTORC1 via T1R1/T1R3-PLCß-Ca2+-ERK1/2 signal transduction. Results revealed a new role for Met-regulated mTORC1 via an AA receptor. Further studies should be performed to determine the role of T1R1/T1R3 in mediating extracellular AA to regulate mTOR signaling and to reveal its mechanism.

Recent Advances in Understanding Amino Acid Sensing Mechanisms that Regulate mTORC1.

The mammalian target of rapamycin (mTOR) is the central regulator of mammalian cell growth, and is essential for the formation of two structurally and functionally distinct complexes: mTORC1 and mTORC2. mTORC1 can sense multiple cues such as nutrients, energy status, growth factors and hormones to control cell growth and proliferation, angiogenesis, autophagy, and metabolism. As one of the key environmental stimuli, amino acids (AAs), especially leucine, glutamine and arginine, play a crucial role in mTORC1 activation, but where and how AAs are sensed and signal to mTORC1 are not fully understood. Classically, AAs activate mTORC1 by Rag GTPases which recruit mTORC1 to lysosomes, where AA signaling initiates. Plasma membrane transceptor L amino acid transporter 1 (LAT1)-4F2hc has dual transporter-receptor function that can sense extracellular AA availability upstream of mTORC1. The lysosomal AA sensors (PAT1 and SLC38A9) and cytoplasmic AA sensors (LRS, Sestrin2 and CASTOR1) also participate in regulating mTORC1 activation. Importantly, AAs can be sensed by plasma membrane receptors, like G protein-coupled receptor (GPCR) T1R1/T1R3, and regulate mTORC1 without being transported into the cells. Furthermore, AA-dependent mTORC1 activation also initiates within Golgi, which is regulated by Golgi-localized AA transporter PAT4. This review provides an overview of the research progress of the AA sensing mechanisms that regulate mTORC1 activity.

Molecular cloning, expression pattern analysis of porcine Rb1 gene and its regulatory roles during primary dedifferentiated fat cells adipogenic differentiation.

Adipocytes are the main constituent of adipose tissue and are considered to be a corner stone in the homeostatic control of whole body metabolism. Recent reports evidenced that retinoblastoma 1 (Rb1) gene plays an important role in fat development and adipogenesis in mice. Here, we cloned the partial cDNA sequences of the porcine Rb1 gene which contains the complete coding sequences (CDS) of 2820bp encoding a protein of 939 amino acids. Bioinformatic analysis revealed that the CDS of porcine Rb1 was highly identical with those of cattle, human and mice. The porcine Rb1 has three typical conserved structural domains, including Rb-A pocket domain, CYCLIN domain and C-terminus domain, and the phylogenetic tree indicates a closer genetic relationship with cattle and human. Tissue distribution analysis showed that Rb1 expression appeared to be ubiquitously in various tissues, being higher in heart, liver, muscle, and stomach. Furthermore, significant downregulation of Rb1 was found at the initial stage of dedifferentiated fat (DFAT) cells adipogenic differentiation. With the knockdown of the Rb1 expression by siRNA, the number of DFAT cells recruited to white rather than brown adipogenesis was promoted, and mRNA levels of adipogenic markers, such as PPARγ, aP2, LPL and adiponectin and protein expression of PPARγ and adiponectin were increased after hormone stimulation. The underlying mechanisms may be that knockdown of Rb1 promotes the mitotic clonal expansion and PPARγ expression by derepressing the transcriptional activity of E2F so as to facilitate the first steps of adipogenesis. In summary, we cloned and characterized an important negative regulator in adipogenic commitment of porcine DFAT cells.

Intervention effects of fructooligosaccharide and astragalus polysaccharide, as typical antibiotic alternatives, on antibiotic resistance genes in feces of layer breeding: advantages and defects.

Although antibiotic alternatives are widely used in livestock and poultry breeding industry after in-feed antibiotics ban, their intervention effects on antibiotic resistance genes (ARGs) in these food animals' feces remain poorly understood. Here effects of fructooligosaccharide (FOS) and astragalus polysaccharide (APS), as typical antibiotic alternatives in China, on ARGs in layer feces were estimated by performing metagenomic sequencings and fluorescence quantitative PCR. Fructooligosaccharide significantly reduced sum abundance of ARGs and mobile genetic elements (MGEs) by increasing Lactobacillus clones and reducing Escherichia clones which had relatively higher abundances of ARG subtypes and MGE subtypes in layer feces. However, at least parts of core ARGs and MGEs categories were not reduced by FOS, such as aminoglycosides- and tetracyclines-resistant genes, Tn916, Integrase, and so on. MGEs and microbiome, especially Escherichia genus and Lactobacillus genus, were the key factors affecting ARGs' sum abundance. MGEs had a higher correlation coefficient with ARGs' sum abundance than Escherichia genus and Lactobacillus genus. These findings firstly reveal the defects of antibiotic alternatives in controlling bacterial resistance in livestock and poultry breeding after in-feed antibiotics ban, and more strategies are needed to control pollutions and risks of core ARGs and MGEs in food animals' feces under a special environment.

Discovery and bioinspired total syntheses of unprecedented sesquiterpenoid dimers unveiled bifurcating [4 + 2] cycloaddition and target differentiation of enantiomers.

[4 + 2] cycloaddition has led to diverse polycyclic chiral architectures, serving as novel sources for organic synthesis and biological exploration. Here, an unprecedented class of cadinane sesquiterpene [4 + 2] dimers, henryinins A-E (1-5), with a unique 6/6/6/6/6-fused pentacyclic system, were isolated from Schisandra henryi. The divergent total syntheses of compounds 1-5 and their enantiomers (6-10) were concisely accomplished in eight linear steps using a protection-free approach. Mechanistic studies illustrated the origin of selectivity in the key [4 + 2] cycloaddition as well as the inhibition of reaction pathway bifurcation via desymmetrization. The chemical proteomics results showed that a pair of enantiomers shared common targets (PRDX5 C100 and BLMH C73) and had unique targets (USP45 C588 for 4 and COG7 C419 for 9). This work provides experimental evidence for the discovery of unprecedented cadinane dimers from selective Diels-Alder reaction and a powerful strategy to explore the biological properties of natural products.

Feeding a DHA-enriched diet increases skeletal muscle protein synthesis in growing pigs: association with increased skeletal muscle insulin action and local mRNA expression of insulin-like growth factor 1.

Dietary n-3 PUFA have been demonstrated to promote muscle growth in growing animals. In the present study, fractional protein synthesis rates (FSR) in the skeletal muscle of growing pigs fed a DHA-enriched (DE) diet (DE treatment) or a soyabean oil (SO) diet (SO treatment) were evaluated in the fed and feed-deprived states. Feeding-induced increases in muscle FSR, as well as the activation of the mammalian target of rapamycin and protein kinase B, were higher in the DE treatment as indicated by the positive interaction between diet and feeding. In the fed state, the activation of eIF4E-binding protein 1 in the skeletal muscle of pigs on the DE diet was higher than that in pigs on the SO diet (P<0·05). Feeding the DE diet increased muscle insulin-like growth factor 1 (IGF-1) expression (P<0·05) and insulin action (as demonstrated by increased insulin receptor (IR) phosphorylation, P<0·05), resulting in increased IR substrate 1 activation in the fed state. However, no difference in plasma IGF-1 concentration or hepatic IGF-1 expression between the two treatments was associated. The increased IGF-1 expression in the DE treatment was associated with increased mRNA expression of the signal transducer and activator of transcription 5A and decreased mRNA expression of protein tyrosine phosphatase, non-receptor type 3 in skeletal muscle. Moreover, mRNA expression of protein tyrosine phosphatase, non-receptor type 1 (PTPN1), the activation of PTPN1 and the activation of NF-κB in muscle were significantly lower in the DE treatment (P<0·05). The results of the present study suggest that feeding a DE diet increased feeding-induced muscle protein synthesis in growing pigs, and muscle IGF-1 expression and insulin action were involved in this action.

5-Aminolevolinic acid mitigates the cadmium-induced changes in Brassica napus as revealed by the biochemical and ultra-structural evaluation of roots.

In the present study, the ameliorating effects of 5-aminolevulinic acid (ALA) under cadmium (Cd) stress conditions were studied with special emphasis on root morphology and ultra-structure in oilseed rape. For this purpose, plants were treated hydroponically at three different Cd levels (0, 100, 500µM) and foliar spray of ALA with three concentrations (0, 12.5, 25mg/l) simultaneously. The results showed that foliar application of ALA improved the plant growth, root morphology and reduced the reactive oxygen species and malondialdehyde contents in roots under Cd stress conditions. The higher concentration of Cd (500µM) decreased the activities of antioxidants enzymes like catalase (CAT), superoxide dismutase (SOD), peroxidase (POD) and glutathione reductase (GR) and also reduced the oxidized glutathione and total glutathione contents in roots. Application of ALA at 25mg/l dosage significantly enhanced the antioxidant activities e.g. APX, SOD, POD, and GSH contents under Cd stress. The microscopic micrographs showed that application of exogenous ALA improved the cell structure under Cd toxicity. A whole cell with developed nucleus, nuclear membrane, smooth cell wall, continuous endoplasmic reticulum, and well shaped mitochondria was observed under the combine application of ALA and Cd. These results suggest that, application of ALA helped the plants to improve root growth, root antioxidant enzymes, and ultra-structural changes in root tip cells under fifteen days Cd-induced stress.

[Morphological characteristics and gene mapping of a novel bent pedicel branch (bpb1) mutant in rice].

Rice pedicels are tightly associated with the yield of grain. In the present study, a novel and stable pedicel mutant bpb1 (bent pedicel branch 1) was obtained from the wild type "Zhenong 7" after 60Co γ-ray treatment. The mutant had the typical phenotype of bent pedicel branches with multiple abnormal phenotypes, such as longer pedicels, short panicles, and dwarfism. Detail examination using scanning electron microscopy revealed that the pedicel epidermal hairs and stomas in the mutant were smaller than those in the wild type. The epidermal and sclerenchymatous cells were arranged irregularly, and the cells in the bend region of pedicels became smaller and arranged closely. The transverse observation of the mutant pedicel branches showed that the small vascular bundles arranged differently from those of the wild type. Genetic analysis indicated that the abnormal phenotypes were controlled by a single recessive gene. Using the F2 mapping population from the bpb1 mutant crossed with the japonica rice variety "Zhenongda 104", the bpb1 gene was mapped in a 343 kb region between two SSR markers, RM21537 and RM21552, at the long arm of chromosome 7. Because no homologous gene was found in this region until now, bpb1 might be a novel gene related to the pedicel development and growth. This study could be beneficial to future cloning and functional analysis of the bpb1 gene.

Heavy metal contamination in Shanghai agricultural soil.

As heavy metals in soil could enrich in biomass and pose health risk to human, it is vital to monitor their contaminations to ensure qualified agricultural production. In this study, we collected >4000 soil samples from agricultural fields in Shanghai during 2010∼2020, and unveiled heavy metal contamination status in this metropolitan. We found that although Shanghai has a long industrialization history, the heavy metal levels in agricultural soil are within safe ranges according to national standard. Specifically, the median levels of Cd, Hg, As, Pb, Cr and Cu are 0.11, 0.13, 7.47, 23.80, 41.00 and 28.30 mg/kg, respectively, which are as good as, or even better than national averages. However, there are spatial and temporal heterogeneities for heavy metal contaminations in Shanghai. For example, the levels of Cd, Hg and Cr are relatively higher in some districts with high industry density, which should be further monitored in the future. Moreover, while the levels for Cd, Cr and Pb have decreased, the level for Hg has mildly increased during this period which needs counteractive measures. Correlation analysis of heavy metal levels and soil fertility parameters suggested overuse of fertilizers may be related to heavy metal contamination in some regions. In summary, our study present by far the largest and most comprehensive landscape of heavy metal contamination in Shanghai agricultural soil, which will be useful for future policy-design and land use planning to ensure safe agricultural production.

Optimizing feeding regimen of replacement gilts to improve their reproductive performance and retention rate of their first 2 parities.

The effects of age, body weight (BW), and backfat thickness (BF) of replacement gilts at first estrus and first mating on their subsequent reproductive performance and retention of their first 2 parities as sows was evaluated. A total of 3,025 Danish replacement gilts were categorized by farm (allocated to 4 farms), cross combination, age, BW and BF at first estrus and first mating, estrous cycle number at first mating, and flush feeding before first mating. The result shows that all the factors mentioned above were significantly associated with reproductive performance and retention rates of the first 2 parities. Farm 3 had more piglets born alive per litter (BA) (P < 0.05). Farms 3 and 4 had more healthy piglets per litter (HP) (P < 0.05). Farm 4 had the most piglets weaned per litter (PW) (P < 0.05). Landrace × Landrace × Yorkshire (L × L × Y) replacement gilts had the most total piglets born per litter (TB), BA, HP, PW and a higher retention rate of the 2 parities than Landrace × Yorkshire (L × Y) replacement gilts (P < 0.05). In addition, flush feeding before first mating had the most TB, BA, HP, PW, and a higher retention rate of the 2 parities than no flush feeding (P < 0.05). Because the effects of replacement gilts rearing parameters on reproductive performance traits differed, we used 100 replacement gilts as a unit and the total number of weaned piglets from the first 2 parities as a new index. Replacement gilts undergoing their first estrus between 180 and 210 d of age at 115 to 124.9 kg BW and 14 to 15 mm BF had significantly higher reproductive indexes for their first 2 parities per 100 replacement gilts. Replacement gilts that mated between 210 and 230 d of age at 140 to 149.9 kg BW and 15 to 16 mm BF had optimal reproductive indexes. These results provide a new insight into the complex relationships among these reproductive performance traits and may help guide successful management of replacement gilts as a pivotal starting point for future fertility and longevity of rearing herds.