The 5-Aminolevulinic Acid (5-ALA) Supplement Enhances PSII Photochemical Activity and Antioxidant Activity in the Late Growth Promotion of Pseudostellaria heterophylla.

This study focused on the physiological regulation and mechanism of exogenous 5-aminolevulinic acid (5-ALA) in the late growth of P. heterophylla. In the middle of May, different concentrations of 5-ALA (0, 10, 20, 50 mg·L-1) were sprayed on the leaves. The effects of 5-ALA on tuberous root growth, antioxidant enzyme system, gas exchange, photosynthetic pigment contents and photosynthetic characteristics were measured from 23 May to 13 June. A concentration of 20 mg·L-1 of 5-ALA led to a significant increase in the yield of fresh root and biomass allocation at 38.12% and 25.07%, respectively, in comparation with the control (0 mg·L-1). The moderate concentration of 5-ALA statistically stimulated antioxidation activities. 5-ALA treatment enhanced photosynthetic activity and reduced photodamage. Compared to the control, there were increases in the chlorophyll fluorescence parameters of P. heterophylla under 5-ALA treatment. Moreover, 20 mg·L-1 of 5-ALA significantly changed the kinetic parameters of fluorescence. It enhanced the light absorption and distribution efficiency of PSII and the activities of leaves, resulting in alleviating photoinhibition by the excess excitation energy. The correlation indicated that there was a significant positive correlation between the yield of tuberous roots and biomass allocation, Pn and catalase (CAT), and a negative correlation between the yield of tuberous roots and malondialdehyde (MDA). The appropriate 5-ALA concentration in the late growth stage of P. heterophylla effectively enhanced the net photosynthetic capacity, mainly resulting from the enhancement of PSII photochemical activity to promote the increases in excitation energy absorption, capture and electron transfer efficiency of the leaves. Finally, 5-ALA treatment can increase the photochemical activity of PSII in the whole leaf and ultimately delay the senescence of P. heterophylla.

Diagnostic Value of Multimodal Magnetic Resonance Imaging in Discriminating Between Metastatic and Non-Metastatic Pelvic Lymph Nodes in Cervical Cancer.

Background: The status of pelvic lymph node (PLN) metastasis affects treatment and prognosis plans in patients with cervical cancer. However, it is hard to be diagnosed in clinical practice. Purpose: The present study aimed to evaluate the diagnostic value of multimodal magnetic resonance imaging (MRI) in discriminating between metastatic and non-metastatic pelvic lymph nodes (PLNs) in cervical cancer. Methods: This retrospective study analyzed MRIs of 209 PLNs in 25 women with pathologically proven cervical cancer. All PLNs had been assessed by pre-treatment multimodal MRIs, and their status was finally confirmed by histopathology. In conventional MRI, lymph node characteristics were compared between metastatic and non-metastatic PLNs. Signal intensity, time-intensity curve (TIC) patterns minimal and mean apparent diffusion coefficients (ADC) were compared between them in DWI. In DCE-MRI, quantitative (Ktrans, Kep and Ve) analyses were performed on DCE-MRI sequences, and their predictive values were analyzed by ROC curves. Results: Of 209 PLNs, 22 (10.53%) were metastases and 187 (89.47%) were non-metastases at histopathologic examination. Considering a comparison of lymph node characteristics, the short axis size, the long axis size, and the boundary differed significantly between the two groups (P<0.05).The differences in ADCmin, TIC types, Ktrans and Ve between metastatic and non-metastatic PLNs were significant as well (P<0.05). The good diagnostic performance of multimodal MRI was shown in discriminating between metastatic and non-metastatic PLNs, with the sensitivity of 85.0% (17/20), specificity of 97.3% (184/189), and accuracy of 96.2% (201/209). ROC analyses showed that the diagnostic accuracy of ADCmin, Ktrans and Ve for discriminating between metastatic and non-metastatic PLNs in cervical cancer was 83.7%, 91.4%, and 92.4% with the cut-off values of 0.72 × 10-3mm2/s, 0.52 min-1, and 0.53 min-1, respectively. Conclusion: Multimodal MRI showed good diagnostic performance in determining PLN status in cervical cancer.

Observation on the Efficacy of Ginkgo Ketone Ester Drop Pill in Improving Hypertension Combined with Carotid Atherosclerotic Plaque.

Purpose: To observe and analyze the efficacy of Ginkgo ketone ester drop pill intervention in patients with hypertension combined with carotid atherosclerotic plaque. Methods: The subjects were 300 patients with hypertension complicated with carotid atherosclerotic plaque treated in our hospital from January 2019 to September 2021. The grouping was done by the random number table method and 300 patients were divided equally into 2 groups. One group was treated with Western medicine alone (clopidogrel sulfate tablets, phenyl amlodipine tablets, irbesartan tablets, and resorvastatin) as the Western medicine group (WM group, n = 150), and one group was added to this intervention with Ginkgo ketone ester drop pill as the Chinese medicine group (CM group, n = 150). The observation indexes were the improvement of blood pressure (systolic blood pressure (SBP) and diastolic blood pressure (DBP)), blood lipids (low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglyceride (TG), and total cholesterol (TC)), vascular endothelial function (nitric oxide (NO) and endothelin-1 (ET-1)), inflammatory factors (C-reactive protein (CRP) and interleukin-6 (IL-6)), plaque (intimal medial thickness (IMT) of carotid artery and plaque area), and efficacy after intervention and adverse effects during intervention in both groups. Results: After intervention, SBP, DBP, LDL-C, TG, and TC levels were lower and HDL-C levels were higher in both groups than before intervention in the same group, and both CM groups improved significantly compared with the WM group (P < 0.05). After intervention, NO levels were higher and ET-1 levels were lower in both groups than before the intervention in the same group, and both CM groups improved significantly compared with the WM group (P < 0.05). After intervention, CRP and IL-6 levels were lower in both groups than before intervention in the same group, and both CM groups improved significantly compared with the WM group (P < 0.05). After intervention, IMT and plaque area were lower in both groups than before intervention in the same group and both CM groups improved significantly compared with the WM group (P < 0.05). The total effective number of the CM group was better than the WM group (P < 0.05), and there was no significant difference in the adverse reactions number in both groups (P > 0.05). Conclusions: The treatment of hypertension combined with carotid atherosclerotic plaque with Ginkgo ketone ester drop pill helps to improve the blood pressure, blood lipid, and vascular endothelial function of patients and helps to inhibit the inflammation level and atherosclerotic plaque of patients, with significant efficacy and no significant adverse effects in patients, which is worthy of clinical promotion.

LncRNA NEAT1/microRNA-124 regulates cell viability, inflammation and fibrosis in high-glucose-treated mesangial cells.

Long non-coding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) has been frequently found to be dysregulated, which contributes to diabetes-related complications. The present study aimed to explore the effect of knockdown on mouse mesangial cell (MMC) viability, apoptosis, inflammation and fibrosis in an in vitro model of diabetic nephropathy (DN). The SV40 MES13 MMC cell line was first cultured with high glucose to establish an in vitro MMC DN cell model. Lnc-NEAT1 shRNA or the negative control shRNA were transfected into MMC DN cells, followed by the measurement of cell viability, apoptosis, inflammation, fibrosis and microRNA (miR)-124 expression, a known target of lnc-NEAT1, using Cell Counting Kit-8, flow cytometry, ELISA, western blotting [Capain1 (capn1), ß-catenin (CTNNB1), cleaved caspase 3, cleaved poly-(ADP ribose) polymerase, fibronectin and Collagen] and reverse transcription-quantitative PCR (Capn1, CTNNB1, lnc-NEAT1, fibronectin, collagen and miR-124), respectively. In rescue experiments, the miR-124 and negative control inhibitor were co-transfected into lnc-NEAT1-downregulated cells, following which cell viability, apoptosis, inflammation, fibrosis, capn1 and CTNNB1 expression were measured. Lnc-NEAT1 expression was increased in high glucose-treated cells compared with that in normal glucose-treated cells and osmotic control cells, suggesting that lnc-NEAT1 is overexpressed in the MMC DN cell model. In the MMC DN cell model, lncRNA-NEAT1 knockdown enhanced cell apoptosis but reduced cell viability and the secretion of inflammatory cytokines in the supernatant (IL-1ß, IL-8, monocyte chemotactic protein 1 and TNF-α), in addition to reducing the expression of fibrosis markers fibronectin and collagen I in the lysates. Lnc-NEAT1 knockdown increased miR-124 expression. Furthermore, transfection with the miR-124 inhibitor reduced cell apoptosis but increased cell viability, inflammation and fibrosis in lnc-NEAT1-downregulated MMC DN cells. miR-124 inhibitor transfection also increased the expression levels of Capn1 and CTNNB1. Taken together, the findings of the present study demonstrated that lnc-NEAT1 knockdown was able to attenuate MMC viability, inflammation and fibrosis by regulating miR-124 expression and the Capn1/ß-catenin signaling pathway downstream. Therefore, Lnc-NEAT1 may serve as a potential therapeutic target for DN.

Structural insights into ligand recognition and selectivity of somatostatin receptors.

Somatostatin receptors (SSTRs) play versatile roles in inhibiting the secretion of multiple hormones such as growth hormone and thyroid-stimulating hormone, and thus are considered as targets for treating multiple tumors. Despite great progress made in therapeutic development against this diverse receptor family, drugs that target SSTRs still show limited efficacy with preferential binding affinity and conspicuous side-effects. Here, we report five structures of SSTR2 and SSTR4 in different states, including two crystal structures of SSTR2 in complex with a selective peptide antagonist and a non-peptide agonist, respectively, a cryo-electron microscopy (cryo-EM) structure of Gi1-bound SSTR2 in the presence of the endogenous ligand SST-14, as well as two cryo-EM structures of Gi1-bound SSTR4 in complex with SST-14 and a small-molecule agonist J-2156, respectively. By comparison of the SSTR structures in different states, molecular mechanisms of agonism and antagonism were illustrated. Together with computational and functional analyses, the key determinants responsible for ligand recognition and selectivity of different SSTR subtypes and multiform binding modes of peptide and non-peptide ligands were identified. Insights gained in this study will help uncover ligand selectivity of various SSTRs and accelerate the development of new molecules with better efficacy by targeting SSTRs.

APLNR Regulates IFN-γ signaling via ß-arrestin 1 mediated JAK-STAT1 pathway in melanoma cells.

The apelin receptor (APLNR) regulates many biological processes including metabolism, angiogenesis, circulating blood volume and cardiovascular function. Additionally, APLNR is overexpressed in various types of cancer and influences cancer progression. APLNR is reported to regulate tumor recognition during immune surveillance by modulating the IFN-γ response. However, the mechanism of APLNR cross-talk with intratumoral IFN-γ signaling remains unknown. Here, we show that activation of APLNR up-regulates IFN-γ signaling in melanoma cells through APLNR mediated ß-arrestin 1 but not ß-arrestin 2 recruitment. Our data suggests that ß-arrestin 1 directly interacts with STAT1 to inhibit STAT1 phosphorylation to attenuate IFN-γ signaling. The APLNR mutant receptor, I109A, which is deficient in ß-arrestins recruitment, is unable to enhance intratumoral IFN-γ signaling. While APLNR N112G, a constitutively active mutant receptor, increases intratumoral sensitivity to IFN-γ signaling by enhancing STAT1 phosphorylation upon IFN-γ exposure. We also demonstrate in a co-culture system that APLNR regulates tumor survival rate. Taken together, our findings reveal that APLNR modulates IFN-γ signaling in melanoma cells and suggest that APLNR may be a potential target to enhance the efficacy of immunotherapy.

Optimization of Taxol Extraction Process Using Response Surface Methodology and Investigation of Temporal and Spatial Distribution of Taxol in Taxus mairei.

Taxus mairei is an important source for industrial extraction of taxol in China. However, the standard and steps of extraction are currently not uniform, which seriously affects the taxol yield. In the present study, the influence of four factors (methanol concentration, solid-liquid ratio, ultrasonic extraction temperature, and ultrasonic extraction time) on the taxol yield was successively explored in T. mairei. A response surface methodology (RSM) was used to optimize the extraction process based on the single-factor experiments above. The optimal conditions were as follows: methanol concentration was 90%, solid-liquid ratio was 1:15 (g/mL), ultrasonic extraction temperature was 40 °C and ultrasonic extraction time was 60 min. Moreover, the twigs and needles from T. mairei with different tree ages were treated by the optimum extraction process, which further revealed temporal and spatial distribution of taxol in the reproducible tissues. Interestingly, the taxol content was relatively higher in needles of T. 'Jinxishan' (a cultivar from T. mairei with yellow aril, FY), but was less in FY twigs. The accumulation of taxol in twigs and leaves of females (with red aril, FR) was significantly higher than that of males (M); however, the content showed a decreasing trend with the increasing tree ages. Therefore, it is suitable to increase the proportion of female trees especially the FY leaves as raw materials for the industrial production of taxol from T. mairei, and the tree ages should be better controlled at 3-7 years.

A self-reported Frailty Index predicts long-term mortality in hospitalized patients with cirrhosis.

BACKGROUND: Frailty is a syndrome that diminishes the potential for functional recovery in liver cirrhosis (LC). However, its utility is limited due to sole reliance on physical performance, especially in hospitalized patients. We investigate the predictive value of a modified self-reported Frailty Index in cirrhotics, and identify which health deficits play more important roles. METHODS: Consecutive LC patients were assessed by our frailty scale. Outcomes of interest were mortality for 90-day, 1-year and 2-year. Independent predictors were identified by multivariate Cox regression. Receiver operating characteristic curve (ROC) was performed to evaluate discriminative ability. We used a combination of stepwise selection, best subset selection, and Akaike information criteria (AIC) to identify pivotal frailty components. RESULTS: The study cohort consisted of 158 patients, in which 37 expired during follow-up. Compared with non-frail groups, the frail group had higher 1- and 2-year mortality. The area under ROC of the Child-Turcotte-Pugh classification (CTP) and Frailty Index were 0.66 and 0.68, while 0.72 for CTP + Frailty Index (P=0.034), respectively. The optimal predictors comprised instrumental activities of daily living (IADL) limitation, falls and loss of weight (AIC =170, C-statistic =0.67). CONCLUSIONS: It is plausible for incorporating Frailty Index to improve prognostication in cirrhotics. IADL limitation, falls and loss of weight play more crucial roles on mortality determination.

Chlorophyll a Fluorescence Transient and 2-Dimensional Electrophoresis Analyses Reveal Response Characteristics of Photosynthesis to Heat Stress in Malus. 'Prairifire'.

Flowering crabapples are a series of precious ornamental woody plants. However, their growth and development are inhibited in the subtropical regions due to the weak photosynthesis under high-temperature environment in the summer. Chlorophyll a fluorescence transient and 2-dimensional electrophoresis (2-DE) analyses were conducted to investigate the response characteristics of photosynthesis under simulated 38 °C heat stress in leaves of Malus. 'Prairifire', a spring-red leaf cultivar of flowering crabapple with strong thermal adaptability. In the present study, the net photosynthetic rate (Pn) was significantly decreased during the heat shock process, which showed a similar trend to the stomatal conductance (Gs), indicating a sensitive stomatal behavior to heat stress. Moreover, an efficient reaction center in photosystem II (PSII), and a functionally intact oxygen-evolving complex (OEC) conferred strong photosynthetic adaptability under heat stress. The higher level of transketolase (TK) under 48-h heat shock treatment was considered a protective mechanism of photosynthetic apparatus. However, heat stress inhibited the functions of light harvesting complex II (LHCII), electron transport in PSII, and the levels of key enzymes in the Calvin cycle, which were considered as the reasons causing an increase in the proportion of non-stomatal restrictions.

Loss of APJ mediated ß-arrestin signalling improves high-fat diet induced metabolic dysfunction but does not alter cardiac function in mice.

Apelin receptor (APJ) is a G protein-coupled receptor that contributes to many physiological processes and is emerging as a therapeutic target to treat a variety of diseases. For most disease indications the role of G protein vs ß-arrestin signalling in mitigating disease pathophysiology remains poorly understood. This hinders the development of G protein biased APJ agonists, which have been proposed to have several advantages over balanced APJ signalling agonists. To elucidate the contribution of APJ ß-arrestin signalling, we generated a transgenic mouse harbouring a point mutation (APJ I107A) that maintains full G protein activity but fails to recruit ß-arrestin following receptor activation. APJ I107A mutant mice did not alter cardiac function at rest, following exercise challenge or in response to pressure overload induced cardiac hypertrophy. Additionally, APJ I107A mice have comparable body weights, plasma glucose and lipid levels relative to WT mice when fed a chow diet. However, APJ I107A mice showed significantly lower body weight, blood insulin levels, improved glucose tolerance and greater insulin sensitivity when fed a high-fat diet. Furthermore, loss of APJ ß-arrestin signalling also affected fat composition and the expression of lipid metabolism related genes in adipose tissue from high-fat fed mice. Taken together, our results suggest that G protein biased APJ activation may be more effective for certain disease indications given that loss of APJ mediated ß-arrestin signalling appears to mitigate several aspects of diet induced metabolic dysfunction.

Function-based high-throughput screening for antibody antagonists and agonists against G protein-coupled receptors.

Hybridoma and phage display are two powerful technologies for isolating target-specific monoclonal antibodies based on the binding. However, for complex membrane proteins, such as G protein-coupled receptors (GPCRs), binding-based screening rarely results in functional antibodies. Here we describe a function-based high-throughput screening method for quickly identifying antibody antagonists and agonists against GPCRs by combining glycosylphosphatidylinositol-anchored antibody cell display with ß-arrestin recruitment-based cell sorting and screening. This method links antibody genotype with phenotype and is applicable to all GPCR targets. We validated this method by identifying a panel of antibody antagonists and an antibody agonist to the human apelin receptor from an immune antibody repertoire. In contrast, we obtained only neutral binders and antibody antagonists from the same repertoire by phage display, suggesting that the new approach described here is more efficient than traditional methods in isolating functional antibodies. This new method may create a new paradigm in antibody drug discovery.

Author Correction: Cryo-EM structures of PAC1 receptor reveal ligand binding mechanism.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Cryo-EM structures of PAC1 receptor reveal ligand binding mechanism.

The pituitary adenylate cyclase-activating polypeptide type I receptor (PAC1R) belongs to the secretin receptor family and is widely distributed in the central neural system and peripheral organs. Abnormal activation of the receptor mediates trigeminovascular activation and sensitization, which is highly related to migraine, making PAC1R a potential therapeutic target. Elucidation of PAC1R activation mechanism would benefit discovery of therapeutic drugs for neuronal disorders. PAC1R activity is governed by pituitary adenylate cyclase-activating polypeptide (PACAP), known as a major vasodilator neuropeptide, and maxadilan, a native peptide from the sand fly, which is also capable of activating the receptor with similar potency. These peptide ligands have divergent sequences yet initiate convergent PAC1R activity. It is of interest to understand the mechanism of PAC1R ligand recognition and receptor activity regulation through structural biology. Here we report two near-atomic resolution cryo-EM structures of PAC1R activated by PACAP38 or maxadilan, providing structural insights into two distinct ligand binding modes. The structures illustrate flexibility of the extracellular domain (ECD) for ligands with distinct conformations, where ECD accommodates ligands in different orientations while extracellular loop 1 (ECL1) protrudes to further anchor the ligand bound in the orthosteric site. By structure-guided molecular modeling and mutagenesis, we tested residues in the ligand-binding pockets and identified clusters of residues that are critical for receptor activity. The structures reported here for the first time elucidate the mechanism of specificity and flexibility of ligand recognition and binding for PAC1R, and provide insights toward the design of therapeutic molecules targeting PAC1R.

Structure-guided discovery of a single-domain antibody agonist against human apelin receptor.

Developing antibody agonists targeting the human apelin receptor (APJ) is a promising therapeutic approach for the treatment of chronic heart failure. Here, we report the structure-guided discovery of a single-domain antibody (sdAb) agonist JN241-9, based on the cocrystal structure of APJ with an sdAb antagonist JN241, the first cocrystal structure of a class A G protein-coupled receptor (GPCR) with a functional antibody. As revealed by the structure, JN241 binds to the extracellular side of APJ, makes critical contacts with the second extracellular loop, and inserts the CDR3 into the ligand-binding pocket. We converted JN241 into a full agonist JN241-9 by inserting a tyrosine into the CDR3. Modeling and molecular dynamics simulation shed light on JN241-9-stimulated receptor activation, providing structural insights for finding agonistic antibodies against class A GPCRs.

Fragment-Based Computational Method for Designing GPCR Ligands.

G protein-coupled receptors (GPCRs) are the largest family of cell surface receptors, which is arguably the most important family of drug target. With the technology breakthroughs in X-ray crystallography and cryo-electron microscopy, more than 300 GPCR-ligand complex structures have been publicly reported since 2007, covering about 60 unique GPCRs. Such abundant structural information certainly will facilitate the structure-based drug design by targeting GPCRs. In this study, we have developed a fragment-based computational method for designing novel GPCR ligands. We first extracted the characteristic interaction patterns (CIPs) on the binding interfaces between GPCRs and their ligands. The CIPs were used as queries to search the chemical fragments derived from GPCR ligands, which were required to form similar interaction patterns with GPCR. Then, the selected chemical fragments were assembled into complete molecules by using the AutoT&T2 software. In this work, we chose ß-adrenergic receptor (ß-AR) and muscarinic acetylcholine receptor (mAChR) as the targets to validate this method. Based on the designs suggested by our method, samples of 63 compounds were purchased and tested in a cell-based functional assay. A total of 15 and 22 compounds were identified as active antagonists for ß2-AR and mAChR M1, respectively. Molecular dynamics simulations and binding free energy analysis were performed to explore the key interactions (e.g., hydrogen bonds and π-π interactions) between those active compounds and their target GPCRs. In summary, our work presents a useful approach to the de novo design of GPCR ligands based on the relevant 3D structural information.

Facile fabrication of nanosheet-assembled MnCoOx hollow flower-like microspheres as highly effective catalysts for the low-temperature selective catalytic reduction of NOx by NH3.

A series of MnCoOx flower-like hollow microspheres with various molecular proportions of reactant were prepared through simple solvothermal method for the ammonia selective catalytic reduction (SCR) at low temperatures. The as-prepared samples have been applied by various characterization techniques to explore the formation process of the morphology and physicochemical properties. The Mn(1)Co(1)Ox presented the optimal intrinsic catalytic performance (95% NOx conversion at 75 °C), favorable thermal stability, and strong SO2 resistance. The excellent properties mainly related to its higher specific surface area and abundant active sites originated from hollow microsphere special structure consists of abundant nanosheets, robust redox properties beneficial for the strong interaction between the manganese and cobalt, larger number of acidic sites and stronger acid strength, etc., which collaboratively dominate its catalytic properties of NH3-SCR at low temperatures.

Fe-modified Ce-MnOx/ACFN catalysts for selective catalytic reduction of NOx by NH3 at low-middle temperature.

A series of MnOx/ACFN, Ce-MnOx/ACFN, and Fe-Ce-MnOx/ACFN catalysts on selective catalytic reduction (SCR) of NOx with NH3 at low-middle temperature had been successfully prepared through ultrasonic impregnation method, and the catalysts were characterized by SEM, XRD, BET, H2-TPR, NH3-TPD, XPS, and FT-IR spectroscopy, respectively. The results demonstrated that the 15 wt% Fe(1)-Ce(3)-MnOx(7)/ACFN catalyst achieved 90% NOx conversion (100~300 °C), good water resistance, and stability (175 °C). The excellent catalytic performance of the Fe(1)-Ce(3)-MnOx(7)/ACFN catalyst was mainly attributed to the interaction among Mn, Ce, and Fe. The doping of Fe promoted the dispersion of Ce and Mn and the formation of more Mn4+ and chemisorbed oxygen on the surface of a catalyst. This work laid a foundation for the successful application of active carbon fiber in the field of industrial denitrification, especially in the aspect of denitrification moving bed. Graphic abstract.

Network Pharmacology and Reverse Molecular Docking-Based Prediction of the Molecular Targets and Pathways for Avicularin Against Cancer.

AIM AND OBJECTIVE: Avicularin has been found to inhibit the proliferation of HepG-2 cells in vitro in the screening of our laboratory. We intended to explain the molecular mechanism of this effect. Therefore, the combined methods of reverse molecular docking and network pharmacology were used in order to illuminate the molecular mechanisms for Avicularin against cancer. MATERIALS AND METHODS: Potential targets associated with anti-tumor effects of Avicularin were screened by reverse molecular docking, then a protein database was established through constructing the drugprotein network from literature mining data, and the protein-protein network was built through an in-depth exploration of the relationships between the proteins, and then the network topology analysis was performed. Additionally, gene function and signaling pathways were analyzed by Go bio-enrichment and KEGG Pathway. RESULTS: The result showed that Avicularin was closely related to 16 targets associated with cancer, and it may significantly influence the pro-survival signals in MAPK signaling pathway that can activate and regulate a series of cellular activities and participate in the regulation of cell proliferation, differentiation, transformation and apoptosis. CONCLUSION: The network pharmacology strategy used herein provided a powerful means for the mechanisms of action for bioactive ingredients.

GPCR structure and function relationship: identification of a biased apelin receptor mutant.

Biased ligands of G protein-coupled receptors (GPCRs) may have improved therapeutic benefits and safety profiles. However, the molecular mechanism of GPCR biased signaling remains largely unknown. Using apelin receptor (APJ) as a model, we systematically investigated the potential effects of amino acid residues around the orthosteric binding site on biased signaling. We discovered that a single residue mutation I109A (I1093.32) in the transmembrane domain 3 (TM3) located in the deep ligand-binding pocket was sufficient to convert a balanced APJ into a G protein signaling biased receptor. APJ I109A mutant receptor retained full capabilities in ligand binding and G protein activation, but was defective in GRK recruitment, ß-arrestin recruitment, and downstream receptor-mediated ERK activation. Based on molecular dynamics simulations, we proposed a molecular mechanism for biased signaling of I109A mutant receptor. We postulate that due to the extra space created by I109A mutation, the phenyl group of the last residue (Phe-13) of apelin rotates down and initiates a cascade of conformational changes in TM3. Phe-13 formed a new cluster of hydrophobic interactions with the sidechains of residues in TM3, including F1103.33 and M1133.36, which stabilizes the mutant receptor in a conformation favoring biased signaling. Interruption of these stabilizing interactions by double mutation F110A/I109A or M113A/I109A largely restored the ß-arrestin-mediated signaling. Taken together, we describe herein the discovery of a biased APJ mutant receptor and provide detailed molecular insights into APJ signaling selectivity, facilitating the discovery of novel therapeutics targeting APJ.

Effects of cadmium stress on the antioxidant system and chlorophyll fluorescence characteristics of two Taxodium clones.

KEY MESSAGE: The T.118 and T.406 seedlings showed strong adaptability under Cd concentrations ≤ 50 µM. The mechanisms of photoprotection in T.118 and T.406 differed in high-Cd concentrations. To explore the physiological response characteristics of Taxodium hybrids to cadmium (Cd) stress and provide basis for screening of Cd-tolerant species, the hydroponic cultivation of T.118 and T.406 seedlings was conducted to demonstrate the effects of Cd stress on seedling growth, antioxidant system, and chlorophyll fluorescence parameters. After 35 days of Cd stress at a concentration ≤ 50 µM, the dry weight biomass of the two clones did not significantly differ from that of the control. T.406 exhibited a significant increase in POD activity compared to T.118 and maintained high SOD activity after exposure to high concentrations of Cd, whereas MDA levels showed little changes. Under low-Cd stress, chlorophyll content and fluorescence parameters remained stable, especially for T.406. Under high-Cd concentration stress, the above parameters were lower than the control, with a more significant decrease in T.118 than in T.406. The non-photochemical quenching coefficient (NPQ) of both clones increased with increasing Cd concentration. T.118 showed a greater increase than T.406, particularly under high-Cd concentration stress. The T.118 and T.406 seedlings adapted to low-Cd concentration stress by enhancing their antioxidant enzyme activity to maintain the balance of reactive oxygen metabolism and reduce cellular damage. The photochemical activity of mesophyll cells remained high to maintain photosynthetic capacity and normal seedling growth. T.406 showed stronger resistance to Cd than T.118. T.406 prevented photodamage by promoting the photochemical utilization of the excitation energy and maintaining a strong antioxidant stress ability. Enhancement of heat dissipation capability may be the main photoprotection mechanism of T.118.