In this study, based on the oxidase activity and photothermal effect of manganese dioxide nanosheets (MnO(2) NSs), with thiamine (TH) as the fluorescence response signal and tris (2,2'-bipyridyl) ruthenium (II) hexahydrate as the reference signal, an enzyme-regulated ratiometric fluorescence and photothermal dual-mode probe was constructed for the quantitative detection of organophosphorus pesticide (OPs) residues. OPs reduced the production of the reductive product thiocholine by inhibiting the activity of acetylcholinesterase, thereby regulating the residual amount of MnO(2) NSs. With the increase of OPs concentration, the color of the probe solution gradually transitioned from red to blue, and the temperature gradually increased. Using dichlorvos and chlorpyrifos as pesticide models, the developed probes exhibited sensitive responses to OPs in a wide linear range of 0.1-8000sng/mL. The detection limits of dichlorvos and chlorpyrifos in fluorescence mode were 1.13sxs10(-3)sng/mL and 0.86sng/mL, respectively. The corresponding detection limits in photothermal mode were 1.01sng/mL and 1.02sng/mL, respectively. The proposed probe displayed excellent anti-interference and reliability in the analysis of OPs residues in real samples. The dual-mode probe with self-verification function is expected to provide more accurate and robust detection results than the single-mode probe, and has a wider application prospect.
Title: Preparation of functional oils rich in diverse medium and long-chain triacylglycerols based on a broadly applicable solvent-free enzymatic strategy Lai Y, Li D, Liu T, Wan C, Zhang Y, Zheng M Ref: Food Res Int, 164:112338, 2023 : PubMed
To address the problems of long reaction times and limited range of adaptation in enzymatic synthesis medium- and long-chain triacylglycerols (MLCTs), a broadly applicable solvent-free enzymatic interesterification strategy was proposed. Candida sp. lipase (CSL) was immobilized on hydrophobic hollow mesoporous silica spheres (HHSS) to construct a biocatalyst designated as CSL@HHSS with a 15.3 % immobilization yield and a loading amount of 94.0 mg/g. The expressed activity and the specific activity were 20.14 U/g and 173.62 U/g, which were 4.6 and 5.6 times higher than that of free CSL, respectively. This biocatalyst demonstrated higher activity, wider applicability, and excellent reusability. Linseed oil, sunflower oil, perilla seed oil, algal oil, and malania oleifera oil were applied as substrates to produce MLCTs with medium-chain triacylglycerols (MCT) catalyzed by CSL@HHSS through interesterification in yields ranging from 69.6 % to 78.0 % within 20 min. Specific fatty acids, including linolenic acid, oleic acid, DHA, and nervonic acid (the first reported), were introduced into MLCT's skeleton, respectively. The structures were finely analyzed and identified by GC and UPLC-MS. The catalytic efficiency value of CSL@HHSS in catalyzing interesterification between linseed oil and MCT (70 degC, 20 min, lipase 6 wt%) is 0.86 g/gmin, which is the highest ever reported. This paper presents an effective and sustainable strategy for functional MLCTs production.
Title: A novel and controllable method for simultaneous preparation of human milk fat substitutes (OPL, OPO and LPL): two-step enzymatic ethanolysis-esterification strategy Li Y, Zhang Y, Zhou Y, Zheng M Ref: Food Res Int, 163:112168, 2023 : PubMed
A novel and effective approach based on the two-step ethanolysis-esterification strategy was proposed for the controllable and simultaneous preparation of 1-oleoyl-2-palmitoyl-3-linoleoylglycerol (OPL), 1,3-dioleoyl-2-palmitoyl-glycerol (OPO) and 1,3-dilinoleoyl-2-palmitoyl-glycerol (LPL) with adjustable proportions. Enzymatic ethanolysis of fractionated palm stearin was carried out to yield 2-monopalmitoylglycerol (79.4s+/-s0.6s%) with over 91.0s% purity at the optimal conditions. The immobilized Candida sp. lipase (CSL) on octyl-functionalized ordered mesoporous silica (OMS-C(8)) was applied to re-esterify 2-monopalmitoylglycerol with oleic acid and linoleic acid for the simultaneous production of OPL, OPO, and LPL. The total content in the final products was 81.5s%, with 91.3s% of palmitic acid (PA) content at the sn-2 position. Besides, OPL/OPO/LPL was conveniently prepared with suitable proportions for worldwide infants by adjusting the ratio of acyl donors. This paper provides a novel and effective two-step ethanolysis-esterification strategy for the development of human milk fat substitutes (HMFS).
Phytochemical investigation on the aerial parts of Tabernaemontana bufalina Lour. (Apocynaceae) led to the identification of four undescribed monoterpenoid indole alkaloids named taberbufamines A-D, an undescribed natural product, and fourteen known indole alkaloids. The structures of the undescribed alkaloids were established by spectroscopic and computational methods, and their absolute configurations were further determined by quantum chemical TDDFT calculations and the experimental ECD spectra. Taberbufamines A and B possessed an uncommon skeleton incorporating an indolizidino [8,7-b]indole motif with a 2-hydroxymethyl-butyl group attached at the pyrrolidine ring. Biosynthetically, Taberbufamines A and B might be derived from iboga-type alkaloid through rearrangement. Vobatensine C showed significant bioactivity against A-549, Bel-7402, and HCT-116 cells with IC(50) values of 2.61, 1.19, and 1.74 microM, respectively. Ervahanine A showed antimicrobial activity against Bacillus subtilis, Mycobacterium smegmatis, and Helicobacter pylori with MIC values of 4, 8, and 16 microg/mL, respectively. 19(S)-hydroxyibogamine was shown as butyrylcholinesterase inhibitor (IC(50) of 20.06 microM) and alpha-glycosidase inhibitor (IC(50) of 17.18 microM), while tabernamine, ervahanine B, and ervadivaricatine B only showed alpha-glycosidase inhibitory activities with IC(50) values in the range of 0.95-4.61 microM.
Title: Design, synthesis, and evaluation of 8-aminoquinoline-melatonin derivatives as effective multifunctional agents for Alzheimer's disease Chen Z, Yu X, Chen L, Xu L, Cai Y, Hou S, Zheng M, Liu F Ref: Ann Transl Med, 10:303, 2022 : PubMed
BACKGROUND: Alzheimer's disease (AD) is thought to be a complex, multifactorial syndrome with many related molecular lesions contributing to its pathogenesis. Thus, multi-target-directed ligands are considered an effective way of treating AD. This study sought to evaluate 8-aminoquinoline-melatonin derivatives as effective multifunctional agents for AD. METHODS: Thioflavin-T fluorescence assays were used to detect the inhibitory potency of 8-aminoquinoline-melatonin hybrids (a1-a5, b1-b5, and c1-c5) on self- and acetylcholinesterase (AChE)-induced amyloid-beta (Abeta) aggregation. The AChE and butyrylcholinesterase (BuChE) inhibitory potency within the compounds was evaluated by Ellman's assays. Methyl thiazolyl tetrazolium (MTT) assays were performed to evaluate the cytotoxicity of the compounds to C17.2 cells. MTT assay was used to detect the cell viability of HT22 cells to evaluate the antioxidant effect of the compounds. Metal chelation property was measured by ultraviolet-visible spectrophotometry. RESULTS: Compounds c3 and c5 had superior inhibitory activity against self-induced Abeta aggregation (with inhibitory rates of 41.4+/-2.1 and 25.5+/-3.2 at 10 microM, respectively) compared to the other compounds. Compounds in the carbamate group (i.e., a4, a5, b4, b5, c4, and c5) showed significant BuChE inhibitory activity and excellent selectivity over AChE. Most of the compounds exhibited low cytotoxicity in the C17.2 cells. Notably, a2, a3, b2, and b3 and series c (c1-c5) exhibited strong protective effects. Additionally, a3 and c1 specifically chelated with copper ions. CONCLUSIONS: Taking all of the promising results together, 8-aminoquinoline-melatonin hybrids can serve as lead molecules in the further development of new multi-functional anti-AD agents.
Title: The physiological, biochemical and transcriptional responses to sulfamethoxazole in the Asian clam, Corbicula fluminea (O. F. Muller, 1774) Liu S, Zhao H, Zheng M, Wang H, Jing C, Zhang W, Hu F Ref: Comparative Biochemistry & Physiology C Toxicol Pharmacol, 260:109406, 2022 : PubMed
Sulfamethoxazole (SMX), a broad-spectrum antibiotic, has been widely used in the treatment and prevention of infection caused by bacteria in recent years. The present study was aimed to evaluate the response mechanisms to SMX stress in gills and digestive gland of Corbicula fluminea (O. F. Mller, 1774). To this end, clams were exposed to environmentally relevant concentrations of SMX (0, 1, 10 and 100 microg/L) for 7 and 28 days, and siphon behavior, tissue-specific enzymatic and transcriptional changes were assayed. Our results showed that exposure to SMX significantly suppressed filtration rate and acetylcholinesterase (AChE) activity, activated antioxidant defense system and elevated transcription of several genes related to cell apoptosis in gills and digestive gland of clams. In general, SMX at environmentally relevant concentrations exhibited a negative impact on siphon behavior and induced neurotoxicology, oxidative stress and cell apoptosis in C. fluminea. The current study will help broaden our understanding of the ecotoxicity of SMX on freshwater bivalves.
This study aimed to identify novel pancreatic lipase (PL) inhibitors using affinity ultrafiltration combined with spectroscopy and molecular docking. Cyanidin-3-O-glucoside (C3G; IC(50): 0.268 mg/mL) and catechin (IC(50): 0.280 mg/mL) were shown to be potent PL inhibitors extracted from black rice and adzuki bean coat extracts. Isobologram analysis revealed that the combined use of C3G and catechin at a ratio of 2:3 had a remarkable synergistic effect (IC(50) of the mixture: 0.201 mg/mL). The inhibitory mechanism of C3G-catechin mixture was of mixed type. The C3G-catechin mixture had a great impact on PL secondary structures. Molecular docking analysis further demonstrated that these polyphenols formed hydrophobic interactions and hydrogen bonds with amino acid residues in the binding pocket of PL. Collectively, C3G and catechin were shown to inhibit PL in a synergistic manner and can be potentially used for the development of food supplements for obesity prevention.
Title: Zeaxanthin remodels cytoplasmic lipid droplets via beta3-adrenergic receptor signaling and enhances perilipin 5-mediated lipid droplet-mitochondrion interactions in adipocytes Xie J, Liu H, Yin W, Ge S, Jin Z, Zheng M, Cai D, Liu M, Liu J Ref: Food Funct, :, 2022 : PubMed
Cytoplasmic lipid droplets (LDs), which are remarkably dynamic, neutral lipid storage organelles, play fundamental roles in lipid metabolism and energy homeostasis. Both the dynamic remodeling of LDs and LD-mitochondrion interactions in adipocytes are effective mechanisms to ameliorate obesity and related comorbidities. Zeaxanthin (ZEA) is a natural carotenoid and has beneficial effects on anti-obesity. However, the underlying mechanisms of ZEA on LD modulation are still unclear. In the present study, ZEA efficiently inhibited LD accumulation and attenuated adipocyte proliferation by arresting the cell cycle. ZEA drove transcriptional alterations to reprogram a lipid oxidative metabolism phenotype in mature 3T3-L1 adipocytes. ZEA significantly decreased the TAG and FA content and modulated the dynamic alterations of LDs by upregulating the expression of lipases and the LD-mitochondrion contact site protein, perilipin 5 (PLIN5), and downregulating the LD fusion protein, fat-specific protein 27 (FSP27). Mechanistically, ZEA stimulated LD remodeling and ameliorated mitochondrial defects caused by large and unilocular LD accumulation by activating beta3-adrenergic receptor (beta3-AR) signaling. Furthermore, the knockdown of PLIN5 impaired the LD-mitochondrion interactions, thereby disrupting the role of ZEA in promoting mitochondrial fatty acid oxidation and respiratory chain operation. Collectively, the present study demonstrates that ZEA induces LD structural and metabolic remodeling by activating beta3-AR signaling and enhances PLIN5-mediated LD-mitochondrion interactions in hypertrophic white adipocytes, thereby enhancing oxidative capacity, and has the potential as a nutritional intervention for the prevention and treatment of obesity and associated metabolic syndrome.
Dinotefuran is a third-generation neonicotinoid pesticide and is increasingly used in agricultural production, which has adverse effects on nontarget organisms. However, the research on the impact of dinotefuran on nontarget organisms is still limited. Here the toxic effects of dinotefuran on an economic lepidopteran model insect, Bombyx mori, were investigated. Exposure to different doses of dinotefuran caused physiological disorders or death. Cytochrome P450, glutathione S-transferase, carboxylesterase, and UDP glycosyl-transferase activities were induced in the fat body at early stages after dinotefuran exposure. By contrast, only glutathione S-transferase activity was increased in the midgut. To overcome the lack of sensitivity of the biological assays at the individual organism level, RNA sequencing was performed to measure differential expressions of mRNA from silkworm larvae after dinotefuran exposure. Differential gene expression profiling revealed that various detoxification enzyme genes were significantly increased after dinotefuran exposure, which was consistent with the upregulation of the detoxifying enzyme. The global transcriptional pattern showed that the physiological responses induced by dinotefuran toxicity involved multiple cellular processes, including energy metabolism, oxidative stress, detoxification, and other fundamental physiological processes. Many metabolism processes, such as carbon metabolism, fatty acid biosynthesis, pyruvate metabolism, and the citrate cycle, were partially repressed in the midgut or fat body. Furthermore, dinotefuran significantly activated the MAPK/CREB, CncC/Keap1, PI3K/Akt, and Toll/IMD pathways. The links between physiological, biochemical toxicity and comparative transcriptomic analysis facilitated the systematic understanding of the integrated biological toxicity of dinotefuran. This study provides a holistic view of the toxicity and detoxification metabolism of dinotefuran in silkworm and other organisms.
Title: Toxic effects of the dinoflagellate Karenia mikimotoi on zebrafish (Danio rerio) larval behavior Niu X, Xu S, Yang Q, Xu X, Zheng M, Li X, Guan W Ref: Harmful Algae, 103:101996, 2021 : PubMed
Karenia mikimotoi is a toxic dinoflagellate that forms harmful blooms in coastal waters, threatening aquaculture worldwide. However, we do not know whether K. mikimotoi has a neurotoxic effect on aquatic animal behavior. Thus, this study investigated potential K. mikimotoi neurotoxicity in zebrafish larvae. Cells of K. mikimotoi were collected at the mid-exponential phase from a batch culture to prepare ruptured cell solutions (RCS). At 6 h post-fertilization (hpf), zebrafish embryos were exposed to different RCS concentrations (0, 10(2), 10(3), 10(4), and 2.5 x 10(4) cells mL(-1)). After 120 hpf, treated larvae were collected to analyze locomotor behavior; activities of acetylcholinesterase (AChE), superoxide dismutase (SOD), catalase (CAT); and expression of genes related to neurodevelopment. We found that RCS did not affect survival rate, but significantly decreased larval locomotion, as well as their AChE, SOD, and CAT activity. Additionally, the examination of the day-night behavioral experiment revealed RCS decreased locomotion only at night. Zebrafish larvae were also significantly hypoactive in response to light and sound stimulations. Of the neurodevelopment genes, three (th, neurog1, and neurod1) were downregulated, while two (bdnf and manf) were upregulated. Our study suggests that K. mikimotoi neurotoxicity occurs through causing oxidative damage, as well as disorders in the cholinergic system and nervous system development. The results provide new insight that K. mikimotoi in low abundance did not cause significant lethal effect but still exhibited significant neurotoxicity on aquatic animals.
Title: An efficient and robust continuous-flow bioreactor for the enzymatic preparation of phytosterol esters based on hollow lipase microarray Xu L, Wang J, Huang F, Zheng M Ref: Food Chem, 372:131256, 2021 : PubMed
In this study, a continuous-flow bioreactor packed with well-organized lipase microarrays was developed for the sustainable synthesis of functional lipid-phytosterol esters (PEs). Hollow mesoporous silicon spheres with a suitable pore size were prepared for lipase immobilization, and the hydrophobic modification endowed the lipase with excellent catalytic activity and stability. The results showed that the condensely packed lipase microarrays offered large specific surface areas and guaranteed the thorough interaction between the lipase and substrates in the continuous-flow bioreactor. Meanwhile, the substrate could pass through the reactor at 1 mL/min with a high conversion of 93.6% due to the hollow structure of the packing spheres. Moreover, the reactors were able to produce 1564 g PEs/g catalyst in a continuous 30-day processing period, which set the highest records for PEs synthesis. This sustainable and highly-converting flow system provided a feasible path for scale-up production of PEs in the food processing area.
Title: Computational biotransformation of polyethylene terephthalate by depolymerase: A QM/MM approach Zheng M, Li Y, Dong W, Feng S, Zhang Q, Wang W Ref: J Hazard Mater, 423:127017, 2021 : PubMed
Despite increasing environmental concerns on ever-lasting Polyethylene Terephthalate (PET), its global production is continuously growing. Effective strategies that can completely remove PET from environment are urgently desired. Here biotransformation processes of PET by one of the most effective enzymes, leaf-branch compost cutinase (LCC), were systematically explored with Molecular Dynamics and Quantum Mechanics/Molecular Mechanics approaches. We found that four concerted steps are required to complete the whole catalytic cycle. The last concerted step, deacylation, was determined as the rate-determining step with Boltzmann-weighted average barrier of 13.6 kcal/mol and arithmetic average of 16.1 +/- 2.9 kcal/mol. Interestingly, unprecedented fluctuations of hydrogen bond length during LCC catalyzed transformation process toward PET were found. This fluctuation was also observed in enzyme IsPETase, indicating that it may widely exist in other catalytic triad (Ser-His-Asp) containing enzymes as well. In addition, possible features (bond, angle, dihedral angle and charge) that influence the catalytic reaction were identified and correlations between activation energies and key features were established. Our results present new insights into catalytic mechanism of hydrolases and shed light on the efficient recycling of the ever-lasting PET.
Title: Hydrolytic Metabolism of Cyanopyrrolidine DPP-4 Inhibitors Mediated by Dipeptidyl Peptidases Kong F, Pang X, Zhao J, Deng P, Zheng M, Zhong D, Chen X Ref: Drug Metabolism & Disposition: The Biological Fate of Chemicals, 47:238, 2019 : PubMed
Nitrile group biotransformation is an unusual or minor metabolic pathway for most nitrile-containing drugs. However, for some cyanopyrrolidine dipeptidyl peptidase 4 (DPP-4) inhibitors (vildagliptin, anagliptin, and besigliptin, but not saxagliptin), the conversion of nitrile group into carboxylic acid is their major metabolic pathway in vivo. DPP-4 was reported to be partly involved in the metabolism. In our pilot study, it was also observed that saxagliptin, a DPP-4 specific inhibitor, decreased the plasma exposures of besigliptin carboxylic acid in rats by only 20%. Therefore, it is speculated that some other enzymes may participate in nitrile group hydrolysis. After incubating gliptins with the cytosol, microsomes, and mitochondria of liver and kidney, carboxylic acid metabolites could all be formed. In recombinant DPP family such as DPP-4, DPP-2, DPP-8, DPP-9, and fibroblast activation protein-alpha, more hydrolytic metabolites were found. Among them, DPP-2 had the highest hydrolytic capacity besides DPP-4, and the DPP-4 inhibitor saxagliptin and DPP-2 inhibitor AX8819 can both inhibit the hydrolysis of gliptins. Western blot results showed that DPP-2 and DPP-4 existed in the aforementioned subcellular organelles at varying amounts. In rats, AX8819 decreased the plasma exposures of besigliptin carboxylic acid by 40%. The amide intermediates of gliptins were detected in vivo and in vitro. When the amide derivatives of gliptins were incubated with DPP-4, they were completely hydrolyzed at a rate far more than that from the parent drug, including saxagliptin-amide. Therefore, it was proposed that gliptins, except saxagliptin, were initially hydrolyzed to their amides by DPPs, which was the rate-limiting step in generating the carboxylic end product.
Title: Characterization of the prognostic values of the NDRG family in gastric cancer Yu C, Hao X, Zhang S, Hu W, Li J, Sun J, Zheng M Ref: Therap Adv Gastroenterol, 12:1756284819858507, 2019 : PubMed
Background: The N-myc downstream-regulated gene (NDRG) family, NDRG1-4, has been involved in a wide spectrum of biological functions in multiple cancers. However, their prognostic values remain sparse in gastric cancer (GC). Therefore, it is crucial to systematically investigate the prognostic values of the NDRG family in GC. Methods: The prognostic values of the NDRG family were evaluated by Kaplan-Meier Plotter and SurvExpress. The mRNA of the NDRG family was investigated in The Cancer Genome Atlas (TCGA). Transcription factors (TFs) and miRNAs associated with the NDRG family were predicted by NetworkAnalysis. The prognostic values of DNA methylation levels were analyzed by MethSurv. The correlation between immune cells and the NDRG family was evaluated by the Tumor Immune Estimation Resource (TIMER) database. Results: High levels of mRNA expression of NDRG2 and NDRG3 were associated with a favorable prognosis in all GCs. In HER2 (-) GC, NDRG1 was significantly associated with a poor prognosis of GC [hazard ratio (HR) = 1.65, 95% confidence interval (CI) = 1.16-2.33, p = 0.0046]. In HER2 (+) GC, NDRG4 showed a poor prognosis (HR = 1.4, 95% CI: 1.06-1.85, p = 0.017). NDRG4 was an independent prognostic factor in recurrence-free survival by TCGA cohort. The low-risk NDRG-signature group displayed a significantly favorable survival outcome than the high-risk group (HR = 1.76, 95% CI: 1.2-2.59, p = 0.00385). The phosphorylated protein NDRG1 (NDRG1_pT346) displayed a favorable overall survival and was significantly associated with HER2 and phosphorylated HER2. Epidermis development was the top biological process (BP) for coexpressed genes associated with NDRG1 and NDRG4, while mitotic nuclear division and mitotic cell processes were the top BPs for NDRG2 and NDRG3, respectively. Overall, 6 CpGs of NDRG1, 4 CpGs of NDRG2, 3 CpGs of NDRG3 and 24 CpGs of NDRG4 were associated with significant prognosis. CD4(+) T-cells showed the highest correlation with NDRG4 (correlation = 0.341, p = 2.14e(-11)). Furthermore, BCL6 in follicular helper T-cells (Tfh) cells showed the highest association with NDRG4 (correlation = 0.438, p = 00e(+)00). Conclusions: This study analyzed the multilevel prognostic values and biological roles of the NDRG family in GC.
BACKGROUND: The molecular mechanisms of Shenxianshengmai (SXSM), a traditional Chinese medicine, on bradycardia have been incompletely understood. The study tried to investigate the gene expression profile and proteomics of bradycardia rabbits' hearts after SXSM treatment. METHODS: Twenty-four adult rabbits were randomly assigned in four groups: sham, model, model plus SXSM treatment, and sham plus SXSM treatment groups. Heart rate was recorded in all rabbits. Then, total RNA of atria and proteins of ventricle were isolated and quantified, respectively. Gene expression profiling was conducted by gene expression chip, and quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was performed to confirm the results of gene expression chip. We used isobaric tags for elative and absolute quantitation and Western blotting to identify altered proteins after SXSM treatment. RESULTS: There was a constant decrease in the mean heart rate (32%, from 238 +/- 6 beats/min to 149 +/- 12 beats/min) after six weeks in model compared with that in sham group. This effect was partially reversed by 4-week SXSM treatment. Complementary DNA microarray demonstrated that the increased acetylcholinesterase and reduced nicotinic receptor were take responsibility for the increased heart rate. In addition, proteins involved in calcium handling and signaling were affected by SXSM treatment. Real-time RT-PCR verified the results from gene chip. Results from proteomics demonstrated that SXSM enhanced oxidative phosphorylation and tricarboxylic acid (TCA) cycle in ventricular myocardium to improve ATP generation. CONCLUSIONS: Long-term SXSM stimulates sympathetic transmission by increasing the expression of acetylcholinesterase and reduces the expression of nicotinic receptor to increase heart rate. SXSM also restored the calcium handling genes and altered genes involved in signaling. In addition, SXSM improves the ATP supply of ventricular myocardium by increasing proteins involved in TCA cycle and oxidation-respiratory chain.
N-myc downstream-regulated gene 1 (NDRG1) has been reported to act as a key regulatory molecule in tumor progression-related signaling pathways, especially in tumor metastasis. However, the related mechanism has not been fully discovered yet. Herein we demonstrated that the novel molecule of cell migration and invasion, caveolin-1, has direct interaction with NDRG1 in human colorectal cancer (CRC) cells. Moreover, we discovered that NDRG1 reduces caveolin-1 protein expression through promoting its ubiquitylation and subsequent degradation via the proteasome in CRC cells. In addition, caveolin-1 mediates the suppressive function of NDRG1 in epithelial-mesenchymal transition, migration and invasion in vitro and metastasis in vivo. These results help to fulfill the potential mechanisms of NDRG1 in anti-metastatic treatment for human colorectal cancer.
Title: N-myc downstream-regulated gene 1 promotes oxaliplatin-triggered apoptosis in colorectal cancer cells via enhancing the ubiquitination of Bcl-2 Yang X, Zhu F, Yu C, Lu J, Zhang L, Lv Y, Sun J, Zheng M Ref: Oncotarget, 8:47709, 2017 : PubMed
N-myc downstream-regulated gene1 (NDRG1) has been identified as a potent tumor suppressor gene. The molecular mechanisms of anti-tumor activity of NDRG1 involve its suppressive effects on a variety of tumorigenic signaling pathways. The purpose of this study was to investigate the role of NDRG1 in the apoptosis of colorectal cancer (CRC) cells. We first collected the clinical data of locally advanced rectal cancer (LARC) patients receiving oxaliplatin-based neoadjuvant chemotherapy in our medical center. Correlation analysis revealed that NDRG1 positively associated with the downstaging rates and prognosis of patients. Then, the effects of over-expression and depletion of NDRG1 gene on apoptosis of colorectal cancer were tested in vitro and in vivo. NDRG1 over-expression promoted apoptosis in colorectal cancer cells whereas depletion of NDRG1 resulted in resistance to oxaliplatin treatment. Furthermore, we observed that Bcl-2, a major anti-apoptotic protein, was regulated by NDRG1 at post-transcriptional level. By binding Protein kinase Calpha (PKCalpha), a classical regulating factor of Bcl-2, NDRG1 enhanced the ubiquitination and degradation of Bcl-2, thus promoting apoptosis in CRC cells. In addition, NDRG1 inhibited tumor growth and promoted apoptosis in mouse xenograft model. In conclusion,NDRG1 promotes oxaliplatin-triggered apoptosis in colorectal cancer. Therefore, colorectal cancer patients can be stratified by the expression level of NDRG1. NDRG1-positive patients may benefit from oxaliplatin-containing chemotherapy regimens whereas those with negative NDRG1 expression should avoid the usage of this cytotoxic drug.
Title: High-level expression and biochemical characterization of a novel cold-active lipase from Rhizomucor endophyticus Duan X, Zheng M, Liu Y, Jiang Z, Yang S Ref: Biotechnol Lett, 38:2127, 2016 : PubMed
OBJECTIVES: To identify novel cold-active lipases from fungal sources and improve their production by heterologous expression in Pichia pastoris. RESULTS: A novel cold-active lipase gene (ReLipB) from Rhizomucor endophyticus was cloned. ReLipB was expressed at a high level in Pichia pastoris using high cell-density fermentation in a 5-l fermentor with the highest lipase activity of 1395 U/ml. The recombinant lipase (RelipB) was purified and biochemically characterized. ReLipB was most active at pH 7.5 and 25 degrees C. It was stable from pH 4.5-9.0. It exhibited broad substrate specificity towards p-nitrophenyl (pNP) esters (C2-C16) and triacylglycerols (C2-C12), showing the highest specific activities towards pNP laurate (231 U/mg) and tricaprylin (1840 U/mg), respectively. In addition, the enzyme displayed excellent stability with high concentrations of organic solvents including cyclohexane, n-hexane, n-heptane, isooctane and petroleum ester and surfactants. CONCLUSIONS: A novel cold-active lipase from Rhizomucor endophyticus was identified, expressed at a high level and biochemically characterized. The high yield and unique enzymatic properties make this lipase of some potential for industrial applications.
Metastasis is a complex process that is regulated by multiple signaling pathways, with the focal adhesion kinase (FAK)/paxillin pathway playing a major role in the formation of focal adhesions and cell motility. N-myc downstream regulated gene-1 (NDRG1) is a potent metastasis suppressor in many solid tumor types, including prostate and colon cancer. Considering the antimetastatic effect of NDRG1 and the crucial involvement of the FAK/paxillin pathway in cellular migration and cell-matrix adhesion, we assessed the effects of NDRG1 on this important oncogenic pathway. In the present study, NDRG1 overexpression and silencing models of HT29 colon cancer and DU145 prostate cancer cells were used to examine the activation of FAK/paxillin signaling and the formation of focal adhesions. The expression of NDRG1 resulted in a marked and significant decrease in the activating phosphorylation of FAK and paxillin, whereas silencing of NDRG1 resulted in an opposite effect. The expression of NDRG1 also inhibited the formation of focal adhesions as well as cell migration and cell-collagen adhesion. Incubation of cells with novel thiosemicarbazones, namely di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone and di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone, that upregulate NDRG1 also resulted in decreased phosphorylation of FAK and paxillin. The ability of these thiosemicarbazones to inhibit cell migration and metastasis could be mediated, at least in part, through the FAK/paxillin pathway.
N-myc downstream regulated gene-1 (NDRG1) is a potent metastasis suppressor that plays a key role in regulating signaling pathways involved in mediating cancer cell invasion and migration, including those derived from prostate, colon, etc. However, the mechanisms and molecular targets through which NDRG1 reduces cancer cell invasion and migration, leading to inhibition of cancer metastasis, are not fully elucidated. In this investigation, using NDRG1 over-expression models in three tumor cell-types (namely, DU145, PC3MM and HT29) and also NDRG1 silencing in DU145 and HT29 cells, we reveal that NDRG1 decreases phosphorylation of a key proto-oncogene, cellular Src (c-Src), at a well-characterized activating site (Tyr416). NDRG1-mediated down-regulation of EGFR expression and activation were responsible for the decreased phosphorylation of c-Src (Tyr416). Indeed, NDRG1 prevented recruitment of c-Src to EGFR and c-Src activation. Moreover, NDRG1 suppressed Rac1 activity by modulating phosphorylation of a c-Src downstream effector, p130Cas, and its association with CrkII, which acts as a "molecular switch" to activate Rac1. NDRG1 also affected another signaling molecule involved in modulating Rac1 signaling, c-Abl, which then inhibited CrkII phosphorylation. Silencing NDRG1 increased cell migration relative to the control and inhibition of c-Src signaling using siRNA, or a pharmacological inhibitor (SU6656), prevented this increase. Hence, the role of NDRG1 in decreasing cell migration is, in part, due to its inhibition of c-Src activation. In addition, novel pharmacological agents, which induce NDRG1 expression and are currently under development as anti-metastatic agents, markedly increase NDRG1 and decrease c-Src activation. This study leads to important insights into the mechanism involved in inhibiting metastasis by NDRG1 and how to target these pathways with novel therapeutics.
Title: The metastasis suppressor NDRG1 modulates the phosphorylation and nuclear translocation of beta-catenin through mechanisms involving FRAT1 and PAK4 Jin R, Liu W, Menezes S, Yue F, Zheng M, Kovacevic Z, Richardson DR Ref: Journal of Cell Science, 127:3116, 2014 : PubMed
N-myc downstream-regulated gene 1 (NDRG1) is a potent metastasis suppressor that has been demonstrated to inhibit the transforming growth factor beta (TGF-beta)-induced epithelial-to-mesenchymal transition (EMT) by maintaining the cell-membrane localization of E-cadherin and beta-catenin in prostate and colon cancer cells. However, the precise molecular mechanism remains unclear. In this investigation, we demonstrate that NDRG1 inhibits the phosphorylation of beta-catenin at Ser33/37 and Thr41 and increases the levels of non-phosphorylated beta-catenin at the plasma membrane in DU145 prostate cancer cells and HT29 colon cancer cells. The mechanism of inhibiting beta-catenin phosphorylation involves the NDRG1-mediated upregulation of the GSK3beta-binding protein FRAT1, which prevents the association of GSK3beta with the Axin1-APC-CK1 destruction complex and the subsequent phosphorylation of beta-catenin. Additionally, NDRG1 is shown to modulate the WNT-beta-catenin pathway by inhibiting the nuclear translocation of beta-catenin. This is mediated through an NDRG1-dependent reduction in the nuclear localization of p21-activated kinase 4 (PAK4), which is known to act as a transporter for beta-catenin nuclear translocation. The current study is the first to elucidate a unique molecular mechanism involved in the NDRG1-dependent regulation of beta-catenin phosphorylation and distribution.
Title: Targeting the Metastasis Suppressor, NDRG1, Using Novel Iron Chelators: Regulation of Stress Fiber-Mediated Tumor Cell Migration via Modulation of the ROCK1/pMLC2 Signaling Pathway Sun J, Zhang D, Zheng Y, Zhao Q, Zheng M, Kovacevic Z, Richardson DR Ref: Molecular Pharmacology, 83:454, 2013 : PubMed
The iron-regulated metastasis suppressor, N-myc downstream-regulated gene 1 (NDRG1), is up-regulated by cellular iron depletion mediated by iron chelators and can inhibit cancer cell migration. However, the mechanism of how NDRG1 achieves this effect remains unclear. In this study, we implemented established and newly constructed NDRG1 overexpression and knockdown models using the DU145, HT29, and HCT116 cancer cell lines to investigate the molecular basis by which NDRG1 exerts its inhibitory effect on cell migration. Using these models, we demonstrated that NDRG1 overexpression inhibits cell migration by preventing actin-filament polymerization, stress fiber assembly and formation. In contrast, NDRG1 knockdown had the opposite effect. Moreover, we identified that NDRG1 inhibited an important regulatory pathway mediated by the Rho-associated, coiled-coil containing protein kinase 1 (ROCK1)/phosphorylated myosin light chain 2 (pMLC2) pathway that modulates stress fiber assembly. The phosphorylation of MLC2 is a key process in inducing stress fiber contraction, and this was shown to be markedly decreased or increased by NDRG1 overexpression or knockdown, respectively. The mechanism involved in the inhibition of MLC2 phosphorylation by NDRG1 was mediated by a significant (P < 0.001) decrease in ROCK1 expression that is a key kinase involved in MLC2 phosphorylation. Considering that NDRG1 is up-regulated after cellular iron depletion, novel thiosemicarbazone iron chelators (e.g., di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone) were demonstrated to inhibit ROCK1/pMLC2-modulated actin-filament polymerization, stress fiber assembly, and formation via a mechanism involving NDRG1. These results highlight the role of the ROCK1/pMLC2 pathway in the NDRG1-mediated antimetastatic signaling network and the therapeutic potential of iron chelators at inhibiting metastasis.
Title: Microsomal epoxide hydrolase (EPHX1) polymorphisms are associated with aberrant promoter methylation of ERCC3 and hematotoxicity in benzene-exposed workers Xing C, Chen Q, Li G, Zhang L, Zheng M, Zou Z, Hou L, Wang QF, Liu X, Guo X Ref: Environmental & Molecular Mutagenesis, 54:397, 2013 : PubMed
Benzene is an important industrial chemical and widespread environmental pollutant known to induce leukemia and other blood disorders. To be carcinogenic, benzene must be metabolized to produce toxic metabolites. To investigate whether single nucleotide polymorphisms (SNPs) in the metabolic enzyme genes are associated with benzene-induced alterations in DNA methylation and hematotoxicity, we genotyped four commonly studied SNPs in three metabolic enzymes genes CYP1A1, EPHX1 and NQO1; and analyzed promoter DNA methylation status in 11 genes which have been reported to be associated with benzene-induced hematotoxicity (BLM, CYP1A1, EPHX1, ERCC3, NQO1, NUDT1, p15, p16, RAD51, TP53 and WRAP53) in 77 benzene-exposed workers and 25 unexposed controls in China. ERCC3, a DNA repair gene, showed a small but statistically significant increase of promoter DNA methylation in the exposed group compared with the unexposed group (mean +/- SD: 4.73 +/- 3.46% vs. 3.63 +/- 1.96%, P = 0.048). We also observed that an increased number of C allele for rs1051740 in EPHX1 was associated with decreased ERCC3 methylation levels in benzene-exposed workers (P(trend) = 0.001), but not in unexposed controls (P(trend) = 0.379). Interestingly, another EPHX1 SNP (rs2234922) was associated with lower white blood cell (WBC) counts (P(trend) = 0.044) in benzene-exposed workers. These associations remained the same when ERCC3 promoter methylation and WBCs were dichotomized according to the 90th percentile (>/=6%) of methylation levels in controls and a leucopenia cutoff (<4 x 10(9) /L), respectively. Our findings suggest that benzene exposure may be associated with hypermethylation in ERCC3, and that genetic variants in EPHX1 may play an important role in epigenetic changes and hematotoxicity among benzene-exposed workers.
BACKGROUND: The mammary gland is a conserved site of lipoprotein lipase expression across species and lipoprotein lipase attachment to the luminal surface of mammary gland vascular endothelial cells has been implicated in the direction of circulating triglycerides into milk synthesis during lactation. PRINCIPAL FINDINGS: Here we report generation of transgenic mice harboring a human lipoprotein lipase gene driven by a mammary gland-specific promoter. Lipoprotein lipase levels in transgenic milk was raised to 0.16 mg/ml, corresponding to an activity of 8772.95 mU/ml. High lipoprotein lipase activity led to a significant reduction of triglyceride concentration in milk, but other components were largely unchanged. Normal pups fed with transgenic milk showed inferior growth performances compared to those fed with normal milk. CONCLUSION: Our study suggests a possibility to reduce the triglyceride content of cow milk using transgenic technology.
Pfam family DUF1023 consists entirely of uncharacterized proteins generated by sequencing the genomes of Actinobacteria (Bateman A., et al., Nucleic Acids Res. 2004;32 Database issue:D138-141.) Utilizing sequence similarity detection methods, we infer homology between DUF1023 and alpha/beta hydrolases. DUF1023 proteins conserve the core secondary structures in alpha/beta hydrolase fold, and share similar catalytic machinery as that of alpha/beta hydrolases. We predict DUF1023 spatial structure and deduce that they function as hydrolases utilizing catalytic Ser-His-Asp triad with the serine as a nucleophile.
Title: [Neurogenetics of the ACh system in Drosophila melanogaster] Zheng M Ref: Sheng Li Ke Xue Jin Zhan, 17:240, 1986 : PubMed
