Mogroside V alleviates inflammation response by modulating miR-21-5P/SPRY1 axis.

Mogroside V (MV) is a natural sweetener extracted from the edible plant Siraitia grosvenorii that possesses anti-inflammatory bioactivity. It has been reported that microRNAs (miRNAs) play an important role in the inflammation response suppression by natural agents. However, whether the anti-inflammation effect of mogroside V is related to miRNAs and the underlying mechanism remains unclear. Our study aimed to identify the key miRNAs important for the anti-inflammation effect of MV and reveal its underlying mechanisms. Our results showed that MV effectively alleviated lung inflammation in ovalbumin-induced (OVA-induced) asthmatic mice. miRNA-seq and mRNA-seq combined analysis identified miR-21-5p as an important miRNA for the inflammation inhibition effect of MV and it predicted SPRY1 to be a target gene of miR-21-5p. We found that MV significantly inhibited the production of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-2 (IL-2), interleukin-6 (IL-6), and nitric oxide (NO), as well as the protein expression of p-P65/P65, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) in OVA-induced asthmatic mice and LPS-treated RAW 264.7 cells. Moreover, the release of ROS increased in LPS-stimulated RAW 264.7 cells but was mitigated by MV pretreatment. In the meantime, the expression of miR-21-5p was decreased by MV, leading to an increase in the expression of SPRY1 in RAW 264.7 cells. Furthermore, miR-21-5p overexpression or SPRY1 knockdown reversed MV's protective effect on inflammatory responses. Conversely, miR-21-5p inhibition or SPRY1 overexpression enhanced MV's effect on inflammatory responses in LPS-exposed RAW 264.7 cells. Therefore, the significant protective effect of mogroside V on inflammation response is related to the downregulation of miR-21-5p and upregulation of SPRY1 in vitro and in vivo, MiR-21-5p/SPRY1 may be novel therapeutic targets of MV for anti-inflammation treatment.

ANP32A Knockdown Attenuates the Malignant Biological Behavior of Colorectal Cancer Cells by Suppressing Epithelial-mesenchymal Transition and ERK Activation.

Acidic leucine rich nuclear phosphoprotein-32A (ANP32A) protein has a variety of functions, such as regulating cell differentiation, influencing cell apoptosis and cell cycle progression. Our previous study demonstrated that high expression of ANP32A was found in the tumor tissues of colorectal cancer (CRC) patients and was positively associated with tumor grading. However, the function and underlying mechanisms of ANP32A in CRC metastasis have not been fully explored. In this study, we found that ANP32A knockdown significantly attenuated the migration and invasion, and epithelial-mesenchymal transition (EMT) in cells. Further mechanistic studies revealed that ANP32A knockdown inhibited the expression of ß-catenin and phosphorylated-ERK. The immunofluorescent staining experiment has revealed that ANP32A was expressed in the cell membrane, cytosol and nucleus, and its expression was positively associated with ß-catenin expression levels. Moreover, the ability of cell migration and invasion was inhibited, the expression of E-cadherin was enhanced following ANP32A knockdown, and these affects were abolished by an ERK activator PMA, enhanced by an ERK inhibitor PD98059. Moreover, our animal experiment also demonstrated that silenced ANP32A inhibited CRC cell growth, multi-organ metastasis, ERK activation and EMT progression in vivo. Collectively, these findings demonstrated that ANP32A promotes CRC progression and that may be a promising target for the anti-metastasis treatment of CRC.

RNA-binding domain 2 of nucleolin is important for the autophagy induction of curcumol in nasopharyngeal carcinoma cells.

BACKGROUND & AIMS: Excessive autophagy induces cell death and is regarded as the treatment of cancer therapy. We have confirmed that the anti-cancer mechanism of curcumol is related to autophagy induction. As the main target protein of curcumol, RNA binding protein nucleolin (NCL) interacted with many tumor promoters accelerating tumor progression. However, the role of NCL in cancer autophagy and in curcumol's anti-tumor effects haven't elucidated. The purpose of the study is to identify the role of NCL in nasopharyngeal carcinoma autophagy and reveal the immanent mechanisms of NCL played in cell autophagy. METHODS & RESULTS: In the current study, we have found that NCL was markedly upregulated in nasopharyngeal carcinoma (NPC) cells. NCL overexpression effectively attenuated the level of autophagy in NPC cells, and NCL silence or curcumol treatment obviously aggravated the autophagy of NPC cells. Moreover, the attenuation of NCL by curcumol lead a significant suppression on PI3K/AKT/mTOR signaling pathway in NPC cells. Mechanistically, NCL was found to be directly interact with AKT and accelerate AKT phosphorylation, which caused the activation of the PI3K/AKT/mTOR pathway. Meanwhile, the RNA Binding Domain (RBD) 2 of NCL interacts with Akt, which was also influenced by curcumol. Notably, the RBDs of NCL delivered AKT expression was related with cell autophagy in the NPC. CONCLUSION: The results demonstrated that NCL regulated cell autophagy was related with interaction of NCL and Akt in NPC cells. The expression of NCL play an important role in autophagy induction and further found that was associated with its effect on NCL RNA-binding domain 2. This study may provide a new perspective on the target protein studies for natural medicines and confirm the effect of curcumol not only regulating the expression of its target protein, but also influencing the function domain of its target protein.

Instrumental and transcriptome analysis reveals the chemotherapeutic effects of doxorubicin-loaded black phosphate nanosheets on abiraterone-resistant prostate cancer.

Prostate cancer is the second most common cause of cancer-related deaths in men and is common in most developed countries. Androgen deprivation therapy (ADT) that uses abiraterone acetate (AA) is an effective second-line treatment for prostate cancer. However, approximately 20-40% of patients develop primary resistance to abiraterone post-treatment. In this study, we aimed to understand the molecular mechanisms underlying the development of abiraterone resistance in prostate cancer cells and the potential use of black phosphorus nanosheets (BPNS) for treating abiraterone-resistant prostate cancer. We first established abiraterone-resistant prostate cancer PC-3 cells and found that these cells have higher migration ability than normal prostate cancer cells. Using comparative transcriptomic and bioinformatics analyses between abiraterone-sensitive PC-3 and abiraterone-resistant PC-3 cells, we highlighted the differentially expressed genes (DEGs) involved in the biological processes related to prostate gland morphogenesis, drug response, immune response, angiogenesis. We further studied the therapeutic effects of BPNS. Our results show that BPNS reduced the proliferation and migration of abiraterone-resistant PC-3 cells. Bioinformatics analysis, including gene ontology, Kyoto encyclopedia of genes and genomes enrichment analysis, and ingenuity pathway analysis (IPA) of the DEGs, suggested that BPNS treatment controlled cancer cell proliferation, metastasis, and oncogenic signaling pathways. Furthermore, the IPA gene network highlighted the involvement of the MMP family, ATF, and notch families in the anti-prostate cancer function of BPNS. Our findings suggest that BPNS may have a chemotherapeutic function in treating abiraterone-resistant prostate cancer.

Correction: Curcumol inhibits breast cancer growth via NCL/ERα36 and the PI3K/AKT pathway.

Correction for 'Curcumol inhibits breast cancer growth via NCL/ERα36 and the PI3K/AKT pathway' by Zhou Lu Wei et al., Food Funct., 2023, https://doi.org/10.1039/d2fo02387c.

Osthole Induces Apoptosis and Caspase-3/GSDME-Dependent Pyroptosis via NQO1-Mediated ROS Generation in HeLa Cells.

Osthole is a natural coumarin which has been proved to inhibit growth of cancer cells by inducing cell death, while its mechanism was considered to be just caused by apoptosis. In our study, we found that osthole activated not just apoptosis, but also pyroptosis which is a form of regulated cell death accompanied by loss of cell membrane integrity and lactate dehydrogenase (LDH) release. Caspase-3 is a key protein of apoptosis as well as pyroptosis. The apoptosis and pyroptosis induced by osthole were all inhibited by irreversible caspase-3 inhibitor Z-DEVD-FMK. Meanwhile, knockdown of gasdermin E (GSDME) only reduced the osthole-induced pyroptosis but did not affect the occurrence of apoptosis. Our proteomic analysis revealed that the expression of NAD(P)H: quinone oxidoreductase 1 (NQO1) was decreased in osthole-treated cells. Moreover, NQO1 inhibition by osthole induced the overproduction of reactive oxygen species (ROS), as well as apoptosis and pyroptosis. ROS inhibitor N-Acetyl-L-cysteine (NAC) not only reduced osthole-induced apoptosis but also reversed its effect on the pyroptosis. Meanwhile, knockdown of NQO1 by si-NQO1 or its inhibitor dicoumarol (DIC) not only enhanced ROS generation but also strengthened the GSDME-mediated pyroptosis. Finally, we demonstrated that osthole inhibited tumor growth and the expression of NQO1 in a HeLa xenograft mode. Similar to the results in vitro, osthole stimulated the activation of caspase-3, PARP, and GSDME in vivo. Taken together, all these data suggested that osthole induced apoptosis and caspase-3/GSDME-mediated pyroptosis via NQO1-mediated ROS accumulation.

A Combined Transcriptomic and Proteomic Approach to Reveal the Effect of Mogroside V on OVA-Induced Pulmonary Inflammation in Mice.

Mogroside V is a bioactive ingredient extracted from the natural food Siraitia grosvenorii which possesses functions that stimulate lung humidification and cough relief activities, but its underlying mechanisms were rarely studied. To estimate its potential protective effect on ovalbumin (OVA)-induced pulmonary inflammation and understand its system-wide mechanism, integrated omics was applied in this study. Mogroside V effectively reduced the levels of IgE, TNF-α, and IL-5 in OVA-induced mice. The results of RNA-seq and data-independent acquisition proteomics approach revealed that 944 genes and 341 proteins were differentially expressed in the normal control group (NC) and ovalbumin-induced control group (OC) and 449 genes and 259 proteins were differentially expressed between the OC and the group treated with 50 mg/kg mogroside V (MV). After a combined analysis of the transcriptome and the proteome, 93 major pathways were screened, and we discovered that mogroside V exerts an anti-inflammation effect in the lung via NF-κB and JAK-STAT, both of which are among the signaling pathways mentioned above. In addition, we found that the key regulatory molecules (Igha, Ighg1, NF-κB, Jak1, and Stat1) in the two pathways were activated in inflammation and inhibited by mogroside V. Thus, mogroside V may be the main bioactivity component in S. grosvenorii that exerts lung humidification and cough relief effects.

The liver metabolic features of Mogroside V compared to Siraitia grosvenorii fruit extract in allergic pneumonia mice.

BACKGROUND: For a long time, Siraitia grosvenorii fruit extract (SGFE) and its dominant compounds, mogroside V(MV) were both reported to have therapeutic effects on allergic pneumonia, while previous studies only stay on phenotype and mechanism of the two active ingredients, hardly have any studies compared the two ingredients on the effect of liver metabolic, and revealed the relationship between mechanism and liver metabolism. OBJECTIVE: Here we elucidated and compared the curative mechanisms of SGFE and MV on allergic pneumonia through liver metabolomics. METHODS: We established allergic pneumonia mice using ovalbumin, then treated the mice with SGFE, MV and positive drug of Suhuang Zhike Jiaonang. The effects of the drugs were evaluated by detecting inflammatory cytokines, pathological examination and liver oxidative stress biomarkers. We explored the metabolic features between SGFE and MV through liver metabolomics consequently. RESULTS: At phenotype, we confirmed that MV and SGFE both inhibited the expression of inflammatory cytokines including interleukins-5 (IL-5), IL-13, IL-17 and OVA-induced immunoglobulin E, which can also relieve inflammatory cells infiltration and mesenchymal thickening in lung tissue compared with positive drug. In addition, both of them can alleviate oxidative stress damage in liver, while MV showed a superior effect than SGFE. In metabolomic analysis, the two ingredients were found to ameliorate inflammatory and oxidative reaction mainly in controlling pathways of Riboflavin metabolism and Glutathione metabolism. While SGFE were found to control other metabolic pathways such as Phenylalanine metabolism, Sphingolipid metabolism, Glycerollipid metabolism, Glycine, serine and threonine metabolism and Arginine and proline metabolism. CONCLUSION: From the results we can infer that the minor ingredients except MV in SGFE contribute poor function to the treatment of allergic pneumonia and MV may be the main functional constituent that relieve allergic pneumonia in SGFE. This study will be beneficial to figuring out a systematic theory of Siraitia grosvenorii active ingredients and proposing a guidance for pharmacology development.

Discovery and Characterization of the Key Constituents in Ginkgo biloba Leaf Extract With Potent Inhibitory Effects on Human UDP-Glucuronosyltransferase 1A1.

Human UDP-glucuronosyltransferase 1A1 (hUGT1A1) is one of the most essential phase II enzymes in humans. Dysfunction or strong inhibition of hUGT1A1 may result in hyperbilirubinaemia and clinically relevant drug/herb-drug interactions (DDIs/HDIs). Recently, a high-throughput fluorescence-based assay was constructed by us to find the compounds/herbal extracts with strong inhibition against intracellular hUGT1A1. Following screening of over one hundred of herbal products, the extract of Ginkgo biloba leaves (GBL) displayed the most potent hUGT1A1 inhibition in HeLa-UGT1A1 cells (Hela cells overexpressed hUGT1A1). Further investigations demonstrated that four biflavones including bilobetin, isoginkgetin, sciadopitysin and ginkgetin, are key constituents responsible for hUGT1A1 inhibition in living cells. These biflavones potently inhibit hUGT1A1 in both human liver microsomes (HLM) and living cells, with the IC50 values ranging from 0.075 to 0.41 µM in living cells. Inhibition kinetic analyses and docking simulations suggested that four tested biflavones potently inhibit hUGT1A1-catalyzed NHPN-O-glucuronidation in HLM via a mixed inhibition manner, showing the K i values ranging from 0.07 to 0.74 µM. Collectively, our findings uncover the key constituents in GBL responsible for hUGT1A1 inhibition and decipher their inhibitory mechanisms against hUGT1A1, which will be very helpful for guiding the rational use of GBL-related herbal products in clinical settings.

Synthesis and Evaluation of Chiral Rhodanine Derivatives Bearing Quinoxalinyl Imidazole Moiety as ALK5 Inhibitors.

BACKGROUND: TGF-ß signaling pathway inhibition is considered an effective way to prevent the development of several diseases. In the design and synthesis of TGF-ß inhibitors, a rhodanine compound containing a quinoxalinyl imidazole moiety was found to have strong antimicrobial activity. OBJECTIVE: The purpose of this work was to investigate the antimicrobial activity of other chiral rhodanine TGF-ß inhibitors synthesized. METHODS: Two series of 3-substituted-5-(5-(6-methylpyridin-2-yl)-4-(quinoxalinyl-6-yl)- 1Himidazol- 2-yl)methylene)-2-thioxothiazolin-4-ones (12a-h and 13a-e) were synthesized and evaluated for their ALK5 inhibitory and antimicrobial activity. The structures were confirmed by their 1H NMR, 13C NMR and HRMS spectra. All the synthesized compounds were screened against Grampositive strains, Gram-negative strains, and fungi. RESULTS: Among the synthesized compounds, compound 12h showed the highest activity (IC50 = 0.416 µM) against ALK5 kinase. Compound 12h exhibited a good selectivity index of >24 against p38α MAP kinase and was 6.0-fold more selective than the clinical candidate, compound 2 (LY- 2157299). Nearly all the compounds displayed high selectivity toward both Gram-positive and Gram-negative bacteria. They also showed similar or 2.0-fold greater antifungal activity (minimum inhibitory concentration [MIC] = 0.5 µg/mL) compared with the positive control compounds Gatifloxacin (MIC = 0.5 µg/mL) and fluconazole (MIC = 1 µg/mL). CONCLUSION: The findings suggest that the synthesized rhodanine compounds have good ALK5 inhibitory activity, and merit further research and development as potential antifungal drugs.

Identification of four ene reductases and their preliminary exploration in the asymmetric synthesis of (R)-dihydrocarvone and (R)-profen derivatives.

The ene reductases (ERs) from the old yellow enzymes (OYEs) family have the ability to reduce activated alkenes to generate up to two stereocenters, therefore they have been received extensive attention as powerful biocatalysts. In this study, through gene mining, four ERs were identified from the genomes of Ensifer adhaerens, Pseudomonas fluorescens, and Pseudomonas veronil. The biocatalytic properties of these four ERs were identified, and their applications in the synthesis process of dihydrocarvone and profen derivatives were further evaluated. Among them, three ERs (EaER2, PvER1, and PvER2) belonging to the classic OYEs showed the best catalytic activity at 30 °C and pH 7.0 (100 mM potassium phosphate buffer) and the PfER2, which belongs to the thermophilic-like OYEs exhibited the best catalytic at 40 °C and pH 7.0 (100 mM potassium phosphate buffer). When exploring the influence of organic solvents on the catalytic efficiency, it was found that the four ERs were more sensitive to toluene and had tolerance to several other selected organic solvents. In addition, EaER2, PfER2, PvER1 and PvER2 showed excellent catalytic activity toward carvone, and the stereoselectivity of PvER2 toward carvone could reach up to 88.7 % de. EaER2 and PfER2 can catalyze the synthesis of a variety of profen derivatives with a stereoselectivity over 99 % ee. Moreover, through homology modeling and molecular docking, we preliminarily explained the mechanism of catalytic activity and stereoselectivity of the four ERs, which provided a solid base on the rational design of their stereo-preference in the future. The discovery of EaER2, PfER2, PvER1, and PvER2 provides four new enzyme sources for the study of the OYEs family and enriches the biocatalytic toolbox of ERs. Our exploration of the enzymatic properties of these four ERs will provide the sufficient data basis for future research and industrialization progress.

Mogroside V reduce OVA-induced pulmonary inflammation based on lung and serum metabolomics.

BACKGROUND: Mogroside V, the main ingredient of Siraitia grosvenorii, has been proved to have therapeutic effects on pulmonary diseases. The specific mechanism still remains to be clarified, which hinders the potence of its medicinal value. PURPOSE: Serum and lung metabolomics based on LC-MS analysis were applied to explore the mechanism of mogroside V against lung inflammation. METHOD: In this study, balb/c mice were divided into control, model, mogeoside V and SH groups. We evaluated the protective effects of mogroside V on lung inflammation in asthmatic mice. Suhuang Zhike Jiaonang was used as positive drug. Metabolic profiles of serum and lung samples of mice in control, model and mogroside V groups were analyzed by LC-MS. RESULTS: Administration of mogroside V effectively relieved the expression of biochemical cytokines and lung inflammatory infiltration of asthmatic mice caused by ovalbumin (OVA). And visceral index of mice treated with mogroside V was close to control group. These results indicated that mogroside V ameliorated OVA-induced lung inflammation. LC-MS based metabolomics analysis demonstrated 6 main pathways in asthmatic mice including Vitamin B6 metabolism, Taurine and hypotaurine metabolism, Ascorbate and aldarate metabolism, Histidine metabolism, Pentose and glucuronate interconversions, Citrate cycle (TCA cycle) were regulated after using mogroside V. CONCLUSION: The study firstly elucidates the metabolic pathways regulated by mogroside V on lung inflammation through metabolomics, providing a theoretical basis for more sufficient utilization and compatibility of mogroside V.

Curcumol inhibits EBV-positive Nasopharyngeal carcinoma migration and invasion by targeting nucleolin.

Metastasis is a major cause of recurrence and death in patients with EBV-positive Nasopharyngeal carcinoma (NPC). Previous reports documented that curcumol has both anti-cancer and anti-viral effects, but there is little literature systematically addressing the mechanism of curcumol in EBV-positive tumors. Previously we found that nucelolin (NCL) is a target protein of curcumol in CNE2 cells, an EBV-negative NPC, and in this experiment, we reported a critical role for NCL in promoting migration and invasion of C666-1 cells, an EBV-positive NPC, and found that the expression of NCL determined the level of curcumol's efficacy. Mechanistically, NCL interacted with Epstein-Barr Virus Nuclear Antigen 1 (EBNA1) to activate VEGFA/VEGFR1/PI3K/AKT signaling pathway, which in turn promoted NPC cell invasion and metastasis. Moreover, further study showed that the differential expression of NCL and curcumol intervention only had a regulatory effect on the nuclear accumulation of VEGFR1, which strengthened the anti-cancer effect of curcumol mediated through NCL. Our findings indicated that curcumol exerted anti EBV-positive NPC invasion and metastasis by downregulating EBNA1 and inhibiting VEGFA/VEGFR1/PI3K/AKT signaling by targeting NCL, which provides a novel pharmacological basis for curcumol's clinical use in treating patients with EBV-positive NPC.

Curcumol inhibits the viability and invasion of colorectal cancer cells via miR-30a-5p and Hippo signaling pathway.

MicroRNA-30a-5p (miR-30a-5p), which functions as a tumor suppressor, has been reported to be downregulated in colorectal cancer (CRC) tissues and to be associated with cancer invasion. However, the detailed regulatory mechanism of curcumol in the malignant progression of CRC remains unknown. MTT, Transwell, scratch, western blotting and reverse transcription-quantitative PCR assays were performed to examine how curcumol inhibited CRC cell viability, invasion and migration, and to detect the role of miR-30a-5p and curcumol in the invasion and Hippo signaling pathways of CRC cells. The present study revealed that miR-30a-5p expression was downregulated in human CRC tissues and cells. The results demonstrated that miR-30a-5p downregulation was accompanied by the inactivation of the Hippo signaling pathway, which was demonstrated to promote CRC cell viability, invasion and migration. Curcumol treatment was identified to increase miR-30a-5p expression and to activate the Hippo signaling pathway, which in turn inhibited the invasion and migration of CRC cells. Overexpression of miR-30a-5p enhanced the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. Furthermore, downregulation of miR-30a-5p reversed the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. These findings identified novel signaling pathways associated with miR-30a-5p and revealed the effects of curcumol on miR-30a-5p expression. Therefore, curcumol may serve as a potential therapeutic strategy to delay CRC progression.

5-Aryl-furan derivatives bearing a phenylalanine- or isoleucine-derived rhodanine moiety as potential PTP1B inhibitors.

Two series of 5-aryl-furan derivatives bearing a phenylalanine- or isoleucine-derived rhodanine moiety were identified as competitive protein tyrosine phosphatase 1B (PTP1B) inhibitors. Among the compounds studied, 5g was found to have the best PTP1B inhibitory potency (IC50 = 2.66 ± 0.16 µM) and the best cell division cycle 25 homolog B (CDC25B) inhibitory potency (IC50 = 0.25 ± 0.02 µM). Enzymatic data together with molecular modeling results demonstrated that the introduction of a sec-butyl group at the 2-position of the carboxyl group remarkably improved the PTP1B inhibitory activity.

Isolation, Substrate Specificity, and Subunit Characterization of the Xylanosomes Produced by Oerskovia turbata JCM 3160.

This work aims at functional studies of the multienzyme complexes produced by Oerskovia turbata JCM 3160 and reveal of their subunit structures. The multienzyme complexes were isolated, enzymatic assayed, the whole genome sequence was determined in fine scale, and the subunit structure was identified by Maldi-TOF mass spectrometry. The isolated multienzyme complexes here show similar particle size with the xylanosomes produced by Cellulosimicrobium cellulans F16, have at least two conserved multi-domain proteins, while differ significantly in enzymatic activities and low molecular weight subunit compositions. This is the first report of the enzymatic activities and subunit structures of xylanosome produced by Oerskovia turbata, providing insights into its diverse capability as well as degrading bias on hemicelluloses.

Isolation and subunit structure of the xylanosome complex produced by Actinotalea fermentans JCM9966.

Xylanosomes, also known as hemicellulosomes, are hemicellulose-degrading nano-scale multienzyme complexes produced by some Firmicutes, Actinobacteria, and Fungi. Here we report the isolation of the MECs produced by Actinotalea fermentas JCM9966, as well as the functional studies and subunit structure revealed by proteomic identifications. The isolated MECs here shows similar particle size with the xylanosomes produced by C. cellulans F16, have several conserved multi-domain proteins, while differ significantly in enzymatic activities and low molecular weight subunit compositions, indicating diverse capability as well as bias in degrading hemicelluloses.

Isolation and subunit compositions of the xylanosome complexes produced by Cellulosimicrobium species.

Cellulosimicrobium cellulans, which is type species of the genus Cellulosimicrobium, produces xylanase predominant nanoscale multienzyme complexes, i.e., xylanosomes, when grown on water-insoluble polysaccharides. Here, we report on the isolation of similar multienzyme complexes (MECs) produced by two other species in genus Cellulosimicrobium (Cellulosimicrobium funkei and Cellulosimicrobium terreum). Functional studies and subunit structure identifications using genomic sequencing and proteomic techniques were also performed. When compared with the xylanosomes produced by C. cellulans F16, the isolated MECs showed a larger particle size and shared at least three conserved multidomain proteins. In addition, they also exhibited different enzymatic activities and subunit compositions, which indicates diverse capability and strategies in degrading hemicelluloses.

Inhibition of human carboxylesterases by magnolol: Kinetic analyses and mechanism.

Magnolol, the most abundant bioactive constituent of the Chinese herb Magnolia officinalis, has been found with multiple biological activities, including anti-oxidative, anti-inflammatory and enzyme-regulatory activities. In this study, the inhibitory effects and inhibition mechanism of magnolol on human carboxylesterases (hCEs), the key enzymes responsible for the hydrolytic metabolism of a variety of endogenous esters as well as ester-bearing drugs, have been well-investigated. The results demonstrate that magnolol strongly inhibits hCE1-mediated hydrolysis of various substrates, whereas the inhibition of hCE2 by magnolol is substrate-dependent, ranging from strong to moderate. Inhibition of intracellular hCE1 and hCE2 by magnolol was also investigated in living HepG2 cells, and the results showed that magnolol could strongly inhibit intracellular hCE1, while the inhibition of intracellular hCE2 was weak. Inhibition kinetic analyses and docking simulations revealed that magnolol inhibited both hCE1 and hCE2 in a mixed manner, which could be partially attributed to its binding at two distinct ligand-binding sites in each carboxylesterase, including the catalytic cavity and the regulatory domain. In addition, the potential risk of the metabolic interactions of magnolol via hCE1 inhibition was predicted on the basis of a series of available pharmacokinetic data and the inhibition constants. All these findings are very helpful in deciphering the metabolic interactions between magnolol and hCEs, and also very useful for avoiding deleterious interactions via inhibition of hCEs.

Anthraquinones from Cassiae semen as thrombin inhibitors: in vitro and in silico studies.

Thrombin inhibitor therapy is one of the most effective therapeutic strategies for the prevention and treatment of cardiovascular and thrombotic diseases. Although several marketed direct thrombin inhibitors (DTIs) have been widely used in clinic, the potentially serious complications of these DTIs prompted the researchers to find more DTIs with improved safety profiles. Herein, we report that natural anthraquinones in Cassiae semen (the seed of Cassia obtusifolia L. or C. tora L.), including obtusifolin, obtusin, aurantio-obtusin and chryso-obtusin, display strong to moderate inhibition on human thrombin, with the IC50 values ranging from 9.08 µM to 27.88 µM. Further investigation on the inhibition kinetics demonstrates that these anthraquinones are mixed inhibitors against thrombin-mediated Z-GGRAMC acetate hydrolysis, while obtusifolin and aurantio-obtusin show strong thrombin inhibition capacity, with the Ki values of 9.63 µM and 10.30 µM, respectively. Docking simulations demonstrate that both obtusifolin and aurantio-obtusin can simultaneously bind on the catalytic cavity and the two anion binding exosites (ABE1 and ABE2), while the hydroxyl group at the C-7 site and the methoxyl group at the C-8 site can create key interactions with the amino acids surrounding the catalytic cavity via hydrogen bonding. All these findings suggest that obtusifolin and aurantio-obtusin are strong thrombin inhibitors possessing a unique anthraquinone skeleton, and could be used as lead compounds for the development of new thrombin inhibitors with improved properties.