Asparagine Synthetase Marks a Distinct Dependency Threshold for Cardiomyocyte Dedifferentiation.

BACKGROUND: Adult mammalian cardiomyocytes have limited proliferative capacity, but in specifically induced contexts they traverse through cell-cycle reentry, offering the potential for heart regeneration. Endogenous cardiomyocyte proliferation is preceded by cardiomyocyte dedifferentiation (CMDD), wherein adult cardiomyocytes revert to a less matured state that is distinct from the classical myocardial fetal stress gene response associated with heart failure. However, very little is known about CMDD as a defined cardiomyocyte cell state in transition. METHODS: Here, we leveraged 2 models of in vitro cultured adult mouse cardiomyocytes and in vivo adeno-associated virus serotype 9 cardiomyocyte-targeted delivery of reprogramming factors (Oct4, Sox2, Klf4, and Myc) in adult mice to study CMDD. We profiled their transcriptomes using RNA sequencing, in combination with multiple published data sets, with the aim of identifying a common denominator for tracking CMDD. RESULTS: RNA sequencing and integrated analysis identified Asparagine Synthetase (Asns) as a unique molecular marker gene well correlated with CMDD, required for increased asparagine and also for distinct fluxes in other amino acids. Although Asns overexpression in Oct4, Sox2, Klf4, and Myc cardiomyocytes augmented hallmarks of CMDD, Asns deficiency led to defective regeneration in the neonatal mouse myocardial infarction model, increased cell death of cultured adult cardiomyocytes, and reduced cell cycle in Oct4, Sox2, Klf4, and Myc cardiomyocytes, at least in part through disrupting the mammalian target of rapamycin complex 1 pathway. CONCLUSIONS: We discovered a novel gene Asns as both a molecular marker and an essential mediator, marking a distinct threshold that appears in common for at least 4 models of CMDD, and revealing an Asns/mammalian target of rapamycin complex 1 axis dependency for dedifferentiating cardiomyocytes. Further study will be needed to extrapolate and assess its relevance to other cell state transitions as well as in heart regeneration.

Chemoproteomic Profiling of Signaling Metabolite Fructose-1,6-Bisphosphate Interacting Proteins in Living Cells.

Fructose-1,6-bisphosphate (FBP), a cellular endogenous sugar metabolite in the glycolytic pathway, has recently been reported to act as a signaling molecule to regulate various cellular events through the engagement of important proteins. Though tremendous progress has been made in identifying specific FBP-protein interactions, the comprehensive identification of FBP-interacting proteins and their regulatory mechanisms remains largely unexplored. Here, we describe a concise synthetic approach for the scalable preparation of a photoaffinity FBP probe that enables the quantitative chemoproteomic profiling of FBP-protein interactions based on photoaffinity labeling (PAL) directly in living cells. Using such a protocol, we captured known FBP targets including PKM2 and MDH2. Furthermore, among unknown FBP-interacting proteins, we identified a mitochondrial metabolic enzyme aldehyde dehydrogenase 2 (ALDH2), against which FBP showed inhibitory activity and resulted in cellular ROS upregulation accompanied by mitochondrial fragmentation. Our findings disclosed a new mode of glucose signaling mediating by the FBP-ALDH2-ROS axis.

Catestatin enhances ATP-induced activation of glial cells mediated by purinergic receptor P2X4.

The activation of glial cells and its possible mechanism play an extremely important role in understanding the pathophysiological process of some clinical diseases, and catestatin (CST) is involved in regulating this activation. In this project, we found that CST could enhance the activation of satellite glial cells (SGCs) and microglial cells and that the expression of P2X4 was increased; the co-expression of the P2X4 receptor with glial fibrillary acidic protein (GFAP) and the P2X4 receptor with CD11b was also increased significantly in glial cells of the ATP + CST group, and TNF-α and IL-1ß also showed a rising trend; the expression of phosphorylated ERK1/2 was also increased in the ATP + CST group. In summary, we conclude that CST could enhance ATP-induced activation of SGCs and microglial cells mediated by the P2X4 receptor and that the ERK1/2 signaling pathway may be involved in this activation process.

Quality of force fields and sampling methods in simulating pepX peptides: a case study for intrinsically disordered proteins.

Intrinsically disordered proteins (IDPs) are a group of proteins that lack well-defined structures under native conditions and carry out crucial physiological functions in various biochemical pathways. Due to the heterogeneous nature of IDPs, molecular dynamics simulations have been extensively adopted to investigate the conformational ensembles and dynamic properties of these proteins. However, their accuracy remains limited by the development of force fields and sampling algorithms. Here, we evaluated the quality of both force fields and enhanced sampling algorithms based on five short pepX peptides. Our results show that the more extended conformational ensembles sampled by the AMOEBA polarizable force field present a higher ability to reproduce experimental NMR observables than AMBER and CHARMM classical force fields. Moreover, a better agreement with experiments is achieved in the simulation of IaMD (integrated accelerated molecular dynamics) than in aMD (accelerated molecular dynamics). The results together indicate that the combination of AMOEBA force field and IaMD enhanced sampling might be a better choice for simulating IDPs. This work may provide important clues for developments and applications of force fields and enhanced sampling methods in future simulations of IDPs.

A double bilayer to study the nonequilibrium environmental response of GIRK2 in complex states.

G protein-gated inwardly rectifying potassium (GIRK) channels play essential roles in electrical signaling in neurons and muscle cells. Nonequilibrium environments provide crucial driving forces behind many cellular events. Here, we apply the antiparallel alignment double bilayer model to study GIRK2 in response to the time-dependent membrane potential. Using molecular dynamics and umbrella sampling, we examined the time-dependent environmental impact on the ion conduction, energy basis, and primary motions of GIRK2 in different complex states with phosphatidylinositol-4,5-bisphosphate (PIP2) and G-protein ßγ subunits (Gßγ). The antiparallel alignment double bilayer model enables us to study the transport performance in inward and outward K+ and mixed K+ and Na+. We obtained the recoverable discharge process of GIRK2 complexed with both PIP2 and Gßγ, compared with occasional conduction under PIP2-only regulation. Calculations of potential of mean force suggest different regulation by the helix bundle crossing (HBC) gate and G-loop gate regarding different complex states and under a membrane potential. In a nonequilibrium environment, distinct functional rocking motions of GIRK2 were identified under strengthened correlations between the transmembrane helices and downstream cytoplasmic domains with binding of PIP2, cations, and Gßγ. The findings suggest the potential domain motions and dynamics associated with a nonequilibrium environment and highlight the application of the antiparallel alignment double bilayer model to investigate factors in an asymmetric environment.

Higher Accuracy Achieved for Protein-Ligand Binding Pose Prediction by Elastic Network Model-Based Ensemble Docking.

Molecular docking plays an indispensable role in predicting the receptor-ligand interactions in which the protein receptor is usually kept rigid, whereas the ligand is treated as being flexible. Because of the inherent flexibility of proteins, the binding pocket of apo receptors might undergo significant conformational rearrangement upon ligand binding, which limits the prediction accuracy of docking. Here, we present an iterative anisotropic network model (iterANM)-based ensemble docking approach, which generates multiple holo-like receptor structures starting from the apo receptor and incorporates protein flexibility into docking. In a validation data set consisting of 233 chemically diverse cyclin-dependent kinase 2 (CDK2) inhibitors, the iterANM-based ensemble docking achieves higher capacity to reproduce native-like binding poses compared with those using single apo receptor conformation or conformational ensemble from molecular dynamics simulations. The prediction success rate within the top5-ranked binding poses produced by the iterANM can further be improved through reranking with the molecular mechanics-Poisson-Boltzmann surface area method. In a smaller data set with 58 CDK2 inhibitors, the iterANM-based ensemble shows a higher success rate compared with the flexible receptor-based docking procedure AutoDockFR and other receptor conformation generation approaches. Further, an additional docking test consisting of 10 diverse receptor-ligand combinations shows that the iterANM is robustly applicable for different receptor structures. These results suggest the iterANM-based ensemble docking as an accurate, efficient, and practical framework to predict the binding mode of a ligand for receptors with flexibility.

Risk factors for hospital-acquired influenza A and patient characteristics: a matched case-control study.

BACKGROUND: While hospital-acquired influenza A results in an additional cost burden and considerable mortality in patients, its risk factors are unknown. We aimed to describe the characteristics of patients vulnerable to hospital-acquired influenza A and to identify its risk factors to assist clinicians control hospital-acquired infections and reduce the burden of treatment. METHODS: A case-control study was conducted among hospitalized patients aged ≥18 years at a tertiary level teaching hospital during the 2018-2019 influenza A season. Patient data were retrieved from hospital-based electronic medical records. Hospital-acquired influenza A was defined as a case of influenza A diagnosed 7 days or more after admission, in a patient with no evidence of influenza A infection on admission. The controls without influenza A were selected among patients exposed to the same setting and time period. We identified risk factors using conditional logistic regression and described the characteristics of hospital-acquired influenza A by comparing the clinical data of infected patients and the controls. RESULTS: Of the 412 hospitalized patients with influenza A from all the departments in the study hospital, 93 (22.6%) cases were classified as hospital-acquired. The most common comorbidities of the 93 cases were hypertension (41.9%), coronary heart disease (21.5%), and cerebrovascular disease (20.4%). Before the onset of hospital-acquired influenza A, patients presented more lymphocytopenia (51.6% vs 35.5%, P = 0.027), hypoalbuminemia (78.5% vs 57.0%, P = 0.002), and pleural effusion (26.9% vs 9.7%, P = 0.002) than the matched controls. Infected patients also had longer hospital stays (18 days vs 14 days, P = 0.002), and higher mortality rates (10.8% vs 2.2%, P = 0.017) than the matched controls. Lymphocytopenia (odds ratio [OR]: 3.11; 95% confidence interval [CI]: 1.24-7.80; P = 0.016), hypoalbuminemia (OR: 2.24; 95% CI: 1.10-4.57; P = 0.027), and pleural effusion (OR: 3.09; 95% CI: 1.26-7.58; P = 0.014) were independently associated with hospital-acquired influenza A. CONCLUSIONS: Lymphocytopenia, hypoalbuminemia and pleural effusion are independent risk factors that can help identify patients at high risk of hospital-acquired influenza A, which can extend hospital stay and is associated with a high mortality.

Quantitative Lysine Reactivity Profiling Reveals Conformational Inhibition Dynamics and Potency of Aurora A Kinase Inhibitors.

Small-molecule inhibitors of protein kinases attract widespread interest in the field of disease therapy because of their high specificity and ease of administration. However, dissecting the conformational inhibition dynamics of kinase inhibitors is still challenging. Here, simultaneously monitoring the conformational inhibition details and potency of Aurora A kinase inhibitors has been achieved by active isotope dimethyl labeling coupled with mass spectrometry-based quantitative lysine reactivity profiling. The conformational effects of inhibitors on lysine reactivity can be globally quantified to feasibly reveal the regions involved in the kinase dynamic inhibition. The half-maximum disturbance concentrations (DC50 values) of the conformation-specific lysine residues could directly represent the conformational selectivity and potency of kinase inhibitors. Further, K309 is discovered as a novel hotspot contributing to the inhibition of Aurora A kinase via the specific rotation of kinase activation loop. This quantitative lysine reactivity profiling strategy might greatly promote the development of targeted drugs.

The role of connexin43 in neuropathic pain induced by spinal cord injury.

Neuropathic pain is caused by the damage or dysfunction of the nervous system. In many neuropathic pain models, there is an increase in the number of gap junction (GJ) channels, especially the upregulation of the expression of connexin43 (Cx43), leading to the secretion of various types of cytokines and involvement in the formation of neuropathic pain. GJs are widely distributed in mammalian organs and tissues, and Cx43 is the most abundant connexin (Cx) in mammals. Astrocytes are the most abundant glial cell type in the central nervous system (CNS), which mainly express Cx43. More importantly, GJs play an important role in regulating cell metabolism, signaling, and function. Many existing literatures showed that Cx43 plays an important role in the nervous system, especially in the CNS under normal and pathological conditions. However, many internal mechanisms have not yet been thoroughly explored. In this review, we summarized the current understanding of the role and association of Cx and pannexin channels in neuropathic pain, especially after spinal cord injury, as well as some of our own insights and thoughts which suggest that Cx43 may become an emerging therapeutic target for future neuropathic pain, bringing new hope to patients.

Nannochloropsis genomes reveal evolution of microalgal oleaginous traits.

Oleaginous microalgae are promising feedstock for biofuels, yet the genetic diversity, origin and evolution of oleaginous traits remain largely unknown. Here we present a detailed phylogenomic analysis of five oleaginous Nannochloropsis species (a total of six strains) and one time-series transcriptome dataset for triacylglycerol (TAG) synthesis on one representative strain. Despite small genome sizes, high coding potential and relative paucity of mobile elements, the genomes feature small cores of ca. 2,700 protein-coding genes and a large pan-genome of >38,000 genes. The six genomes share key oleaginous traits, such as the enrichment of selected lipid biosynthesis genes and certain glycoside hydrolase genes that potentially shift carbon flux from chrysolaminaran to TAG synthesis. The eleven type II diacylglycerol acyltransferase genes (DGAT-2) in every strain, each expressed during TAG synthesis, likely originated from three ancient genomes, including the secondary endosymbiosis host and the engulfed green and red algae. Horizontal gene transfers were inferred in most lipid synthesis nodes with expanded gene doses and many glycoside hydrolase genes. Thus multiple genome pooling and horizontal genetic exchange, together with selective inheritance of lipid synthesis genes and species-specific gene loss, have led to the enormous genetic apparatus for oleaginousness and the wide genomic divergence among present-day Nannochloropsis. These findings have important implications in the screening and genetic engineering of microalgae for biofuels.

Fragment-Based Design, Synthesis, and Biological Evaluation of 1-Substituted-indole-2-carboxylic Acids as Selective Mcl-1 Inhibitors.

Based on a known selective Mcl-1 inhibitor, 6-chloro-3-(3-(4-chloro-3,5-dimethylphenoxy)propyl)-1H-indole-2-carboxylic acid, we applied a fragment-based approach to obtain new molecules that extended into the p1 pocket of the BH3 groove and then exhibited binding selectivity for the Mcl-1 over the Bcl-2 protein. After we deconstructed the 1H-indole-2-carboxylic acid from the parental molecule, a benzenesulfonyl was substituted at the 1-position to adopt a geometry preferred for accessing the p1 pocket according to the binding mode of the parental molecule identified by X-ray crystallography. A linear relationship between the free energy of ligand binding (ΔG) and the count of non-hydrogen heavy atoms (HAC) was maintained during the molecular growing to occupy the p1 pocket. Finally, we not only obtained compound 12 with a 7.5-fold selectivity to Mcl-1 (Ki = 0.48 µM by fluorescence polarization) over Bcl-2 (Ki = 3.6 µM), but also provided evidence that additional occupation of the p1 pocket is more favorable for Mcl-1 than for Bcl-2 binding, and contributes more to Mcl-1 inhibition than occupation of the p2 pocket. Compound 12 exhibited a selective killing ability on Mcl-1-dependent cancer cells.

Genomic Foundation of Starch-to-Lipid Switch in Oleaginous Chlorella spp.

The ability to rapidly switch the intracellular energy storage form from starch to lipids is an advantageous trait for microalgae feedstock. To probe this mechanism, we sequenced the 56.8-Mbp genome of Chlorella pyrenoidosa FACHB-9, an industrial production strain for protein, starch, and lipids. The genome exhibits positive selection and gene family expansion in lipid and carbohydrate metabolism and genes related to cell cycle and stress response. Moreover, 10 lipid metabolism genes might be originated from bacteria via horizontal gene transfer. Transcriptomic dynamics tracked via messenger RNA sequencing over six time points during metabolic switch from starch-rich heterotrophy to lipid-rich photoautotrophy revealed that under heterotrophy, genes most strongly expressed were from the tricarboxylic acid cycle, respiratory chain, oxidative phosphorylation, gluconeogenesis, glyoxylate cycle, and amino acid metabolisms, whereas those most down-regulated were from fatty acid and oxidative pentose phosphate metabolism. The shift from heterotrophy into photoautotrophy highlights up-regulation of genes from carbon fixation, photosynthesis, fatty acid biosynthesis, the oxidative pentose phosphate pathway, and starch catabolism, which resulted in a marked redirection of metabolism, where the primary carbon source of glycine is no longer supplied to cell building blocks by the tricarboxylic acid cycle and gluconeogenesis, whereas carbon skeletons from photosynthesis and starch degradation may be directly channeled into fatty acid and protein biosynthesis. By establishing the first genetic transformation in industrial oleaginous C. pyrenoidosa, we further showed that overexpression of an NAD(H) kinase from Arabidopsis (Arabidopsis thaliana) increased cellular lipid content by 110.4%, yet without reducing growth rate. These findings provide a foundation for exploiting the metabolic switch in microalgae for improved photosynthetic production of food and fuels.

Association of the glucocorticoid receptor D641V variant with steroid-resistant asthma: a case-control study.

OBJECTIVES: Several mutations of the glucocorticoid receptor (GR) gene cause malfunction of the protein, resulting in steroid resistance. In diseases other than asthma, the GR variants I559N, D641V, and V729I have been linked to steroid resistance. The aim of this study was to evaluate the link of these GR variants in steroid-resistant (SR) asthma in the Chinese Han population. METHODS: GR polymorphisms were determined in 64 SR asthma patients, 217 steroid-sensitive (SS) asthma patients and 221 healthy control (CTR) individuals. The analysis of the GR variants was performed using PCR-sequence specific primers according to the European Molecular Biology Laboratory database (NC_000005.8). In addition, ligand binding and serum cortisol levels were determined. RESULTS: Compared with SS asthma patients and CTRs, a significant lower frequency of the GR D641V variant AA genotype (P=0.003, 0.014, respectively) and the A allele (P=0.001, 0.009, respectively) was found in SR asthma patients. Furthermore, the equilibrium dissociation constant (Kd) of GR ligand binding in SR asthma patients with the GR D641V variant AA genotype was significantly lower compared with the AT or the TT genotype carriers (P=0.006, 0.016, respectively). There was no significant difference between the I559N and V729I GR variants on comparing SR asthma patients with SS asthma patients or CTRs. CONCLUSION: This study suggests that the D641V variant of the GR is probably associated with SR asthma in the Chinese Han population.

Late Positivity Correlates with Subjective Reports: Evidence from the Low-frequency and High-frequency Reporting Tasks.

Identifying the neural correlates of consciousness (NCCs) is an important way to understand the fundamental nature of consciousness. By recording event-related potentials (ERPs) using EEG, researchers have found three potential electrophysiological NCCs: early positive correlate of consciousness (enhanced P1), visual awareness negativity (VAN), and late positivity (LP). However, LP may reflect post-perceptual processing associated with subjective reports rather than consciousness per se. The present experiment investigated the relationship between LP and subjective reports. We adopted two subjective reporting tasks that differed in the requirement for subjective reports. In the low-frequency reporting task, participants needed to report whether they saw the target picture in 25% of trials, whereas in the high-frequency reporting task, participants needed to report whether they saw the target picture in each trial. Behavioral results showed that the hit rates were lower and false alarm rates were higher on reporting trials in low-frequency reporting tasks than on reporting trials in high-frequency reporting tasks. Unexpectedly, VAN was larger on reporting trials in the low-frequency reporting task than on reporting trials in the high-frequency reporting task. Importantly, our ERP results showed that LP was larger on reporting trials in the high-frequency reporting task than on reporting trials in the low-frequency reporting task. Thus, our findings indicated that when the frequency of reports was increased, the task relevance of the stimuli increased, which led to larger LP amplitudes. These findings suggest that LP correlates with subjective reports.

LC-MS/MS profiling of colon oxysterols and cholesterol precursors in mouse model of ulcerative colitis.

Oxysterols and cholesterol precursors are being increasingly investigated in humans and laboratory animals as markers for various diseases in addition to their important functions. However, the quantitative analysis of these bioactive molecules is obstructed by high structural similarity, poor ionization efficiency and low abundance. The current assay methods are still cumbersome to be of practical use, and their applicability in different bio-samples needs to be evaluated and optimized as necessary. In the present work, chromatographic separation conditions were carefully studied to achieve baseline separation of difficult-to-isolate compound pairs. On the other hand, an efficient sample purification method was established for colon tissue samples with good recoveries of sterols, demonstrating negligible autoxidation of cholesterol into oxysterols. The developed UPLC-APCI-MS/MS method was thoroughly validated and applied to measure oxysterols and cholesterol precursors in colon tissue of dextran sulfate sodium (DSS)-induced mouse colitis models, and it is expected to be successfully applied to the quantitative determination of such components in other tissue samples.

Protective Efficacy on Adult Ischemic Myocardium under Bypass: Del Nido vs. St. Thomas Blood Cardioplegia.

PURPOSE: To compare the myocardial protective efficacy of del Nido cardioplegia (DNC) with St. Thomas blood cardioplegia (SBC) in adult cardiac surgery. METHODS: From January to December 2021, all the patients who underwent elective cardiac operation were randomly divided into two cohorts based on the type of cardioplegia: DNC group and SBC group. Three categories of variables were compared: patient demographics, clinical variables, and laboratory variables. RESULTS: A total of 133 patients were enrolled in this study: DNC group, n = 65; and SBC group, n = 68. Except that the volume of cardioplegia administration were obvious less in the DNC group (P <0.01), no significant difference was found in the other postoperative clinical variables (P >0.05). No statistical significance was proved (P >0.05) in postoperative troponin I, creatine kinase, and B-type natriuretic peptide. The malondialdehyde concentration was higher in the SBC group, whether it is at 4 hours (P <0.05) or 24 hours (P >0.05) after operation. At the same two points in time, the change in superoxide dismutase activity was more significant in the SBC group (P <0.05). CONCLUSION: The DNC cardioplegia was safe and effective on adult myocardium protection. The potential antioxidant stress effect in DNC may provide a direction for further improvement on the formula of cardioplegic solution.

Study on Long Term Property of Soft Soil Solidified with Industrial Waste Residue and Regenerated Fine Aggregate.

The long-term properties of solidified soft soil, including an immersion test, the dry-wet cycle and the freeze-thaw cycle, were systematically studied. Firstly, the immersion stability of solidified soft soil was confirmed. The appearance of soft soil solidified by a solidified agent and raw fine aggregate did not change significantly, and it was still intact without damage when the soaking time increased up to 28 d. Secondly, the mass and compressive strength loss of solidified soft soil were determined. When the number of dry-wet cycles was one, three, five and seven, the accumulated-mass loss rate was 1.4%, 3.0%, 4.5% and 6.0%, respectively, and the compressive-strength loss rate was -10.3%, 13.9%, 41.2% and 53.6%, respectively. Compared with solidified soft soil under standard curing environments, solidified soft soil after seven dry-wet cycles showed small cracks, and the structural compactness began to decline. Finally, the influence of the freeze-thaw cycle on the mass, compressive strength and microstructure of solidified soft soil was confirmed. When the number of freeze-thaw cycles was 5, 10, 15 and 20, the accumulated-mass loss rate was 12.6%, 16.7%, 17.9% and 18.8%, respectively. The microstructure of the solidified soft soil was damaged, and the increase in porosity was the main reason for its strength reduction or even failure. Nevertheless, soft soil with a solidified agent and recycled fine aggregate had no obvious damage to the microstructure, and the freeze-thaw resistance was relatively superior.

A Reactive Force Field for Molecular Dynamics Simulations of Glucose in Aqueous Solution.

To expand the capabilities of reactive force field (ReaxFF) in simulations of biological processes involving glucose, in this work, using Metropolis Monte Carlo algorithm, new ReaxFF parameters for glucose have been developed to better describe the properties of glucose in water during molecular dynamics (MD) simulations. With the newly trained ReaxFF, the mutarotation of glucose in water can be better described, as suggested by our metadynamics simulations. In addition, the newly trained ReaxFF can better describe the distributions of the three stable conformers along the key dihedral angle of α-anomer and ß-anomer. With better descriptions of hydration around glucose, the Raman and Raman optical activity spectra can be more accurately calculated. In addition, the infrared spectra obtained from simulations with the new glucose ReaxFF are more accurate than those obtained with the original ReaxFF. We note that although our trained ReaxFF performs better than the original ReaxFF, it is not generally applicable to all carbohydrates, which require further parametrization. We also find that the absence of explicit water molecules in the training sets may lead to inaccurate descriptions of water-water interactions around the glucose, implicating that it is necessary to optimize the water ReaxFF parameters together with the target molecule. The improved ReaxFF makes it possible to explore interesting biological processes involving glucose more accurately and efficiently.

RNA Folding Based on 5 Beads Model and Multiscale Simulation.

RNA folding prediction is very meaningful and challenging. The molecular dynamics simulation (MDS) of all atoms (AA) is limited to the folding of small RNA molecules. At present, most of the practical models are coarse grained (CG) model, and the coarse-grained force field (CGFF) parameters usually depend on known RNA structures. However, the limitation of the CGFF is obvious that it is difficult to study the modified RNA. Based on the 3 beads model (AIMS_RNA_B3), we proposed the AIMS_RNA_B5 model with three beads representing a base and two beads representing the main chain (sugar group and phosphate group). We first run the all atom molecular dynamic simulation (AAMDS), and fit the CGFF parameter with the AA trajectory. Then perform the coarse-grained molecular dynamic simulation (CGMDS). AAMDS is the foundation of CGMDS. CGMDS is mainly to carry out the conformation sampling based on the current AAMDS state and improve the folding speed. We simulated the folding of three RNAs, which belong to hairpin, pseudoknot and tRNA respectively. Compared to the AIMS_RNA_B3 model, the AIMS_RNA_B5 model is more reasonable and performs better.

Chemical index components and quality control of Traditional Chinese Medicine: "Never change a winning team"? -A case study of volatile oil from Bupleuri radix.

Chemical index components, especially those defined as quality control (QC) markers through spectrum-effect relationship approach, are commonly suggested and adopted as indicator for quality control of Traditional Chinese Medicines (TCMs). However, are chemical index components and quality control of TCMs "never change a winning team"? In this study, under the ponderation of the applicability of QC markers strategy, spectrum-effect relationship and OPLS-DA between GC×GC-MS fingerprint and inhibitory effect on the expression of extracellular secretory TNF-α of volatile oil from Bupleuri radix (BVO) was studied with the purpose of discovery of QC markers and establish a bioactive compounds-based QC method. 290 compounds of BVO were identified by GC×GC-MS. Besides, BVO had significant inhibitory effects on the expression of extracellular secretory TNF-α in a dose-dependent manner. The potency of different batches of BVOs could be distinguished with this bioassay-based method, which has been validated in terms of intermediate precision, repeatability, linearity, range and credibility tests. The QC markers of BVO were investigated by Spearman's correlation test and OPLS-DA. It is regrettable that there were no ideal QC markers of BVO could be found. In conclusion, quality control method relayed on chemical QC markers is not feasible for TCMs with complex composition but lack of ingredients that dominate in content, just like BVO. Alternatively, a bioassay-based method established in our study is suitable for quality control of BVO.