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1.
Anim Reprod Sci ; 213: 106271, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31987324

RESUMO

For vitellogenin (Vtg) absorption to occur, there needs to binding of the glycolipophosphoproteins to the oocyte membrane in oviparous species, including teleosts. The cDNAs encoding homologous Vtg receptors (VgRs) LR8- and Lrp13 were cloned from ovaries of the large yellow croaker (Larimichthys crocea), an economically important species in Chinese aquaculture. The full-length Lc-lr8-/lrp13 cDNAs contained 4266/3760 base pairs (bp) and the deduced Lc-LR8-/Lrp13 proteins had 844/1218 amino acids, respectively. The VgRs comprised a ligand-binding domain, an epidermal growth factor precursor homology domain, YWXD motifs forming a ß-propeller structure, and transmembrane and cytoplasmic domains. There was a marked relative abundance of Lc-lr8-/lrp13 transcripts in the tissues that were evaluated, with the largest abundance in the ovaries at Stage II of development. Furthermore, there was a lesser relative abundance of Lc-lr8-/lrp13 mRNA transcript during ovarian development (Stages II to IV). In situ hybridization technology was used to analyze decreasing relative abundance pattern of Lc-lr8-/lrp13 mRNA transcript during oogenesis in Stage II to IV of ovarian development. By combining mRNA relative abundance with morphological results, a model was developed to explain the reduction in Lc-lr8-/lrp13 mRNA transcript relative abundance during ovarian development. During the early developmental stages, transcription, translation, and differential accumulation of VgRs in previtellogenic and vitellogenic oocytes may occur and result in Vtg absorption in teleost oocytes. Overall, there is preliminary evidence indicating that at least two VgRs (Lc-LR8-/Lrp13) are present in the large yellow croaker and may be important for Vtg transport from the blood into the oocyte during ovarian development.

2.
Insect Biochem Mol Biol ; 117: 103289, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31778795

RESUMO

The Xenopus oocyte and the Human Embryonic Kidney (HEK) 293 cell expression systems are frequently used for functional characterization (deorphanization) of insect odorant receptors (ORs). However, the inherent characteristics of these heterologous systems differ in several aspects, which raises the question of whether the two systems provide comparable results, and how well the results correspond to the responses obtained from olfactory sensory neurons in vivo. Five candidate pheromone receptors were previously identified in the primitive moth Eriocrania semipurpurella (Esem) and their responses were characterized in HEK cells. We re-examined the responses of these five EsemORs in Xenopus oocytes. We showed that in both systems, EsemOR1 specifically responded to the plant volatile ß-caryophyllene. EsemOR3 responded stronger to the pheromone component (S,Z)-6-nonen-2-ol than to its enantiomer (R,Z)-6-nonen-2-ol, the second pheromone component. However, EsemOR3 also responded secondarily to the plant volatile ß-caryophyllene in the oocyte system, but not in the HEK cell system. EsemOR4 was unresponsive in the HEK cells, but responded primarily to (R,Z)-6-nonen-2-ol followed by (S,Z)-6-nonen-2-ol in the oocytes, representing a discovery of a new pheromone receptor in this species. EsemOR5 was broadly tuned in both systems, but the rank order among the most active pheromone compounds and antagonists was different. EsemOR6 showed no response to any compound in either system. We compared the results obtained in the two different heterologous systems with the activity previously recorded in vivo, and performed in situ hybridization to localize the expression of these OR genes in the antennae. In spite of similar results overall, differences in OR responses between heterologous expression systems suggest that conclusions about the function of individual ORs may differ depending on the system used for deorphanization.

3.
Hypertension ; 75(2): 439-448, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31865783

RESUMO

The inhibition of Type II angiotensin II receptor (AT2R) or BK2R (bradykinin type II receptor) stimulates basolateral Kir4.1/Kir5.1 in the distal convoluted tubule (DCT) and activates thiazide-sensitive NCC (Na-Cl cotransporter). The aim of the present study is to examine the role of AT2R and BK2R in mediating the effect of HK (high dietary K+) intake on the basolateral K+ channels, NCC, and renal K+ excretion. Feeding mice (male and female) with HK diet for overnight significantly decreased the basolateral K+ conductance, depolarized the DCT membrane, diminished the expression of pNCC (phosphorylated NCC) and tNCC (total NCC), and decreased thiazide-sensitive natriuresis. Overnight HK intake also increased the expression of cleaved ENaC-α and -γ subunits but had no effect on NKCC2 expression. Pretreatment of the mice (male and female) with PD123319 and HOE140 stimulated the expression of tNCC and pNCC, augmented hydrochlorothiazide-induced natriuresis, and increased the negativity of the DCT membrane. The deletion of Kir4.1 not only decreased the NCC activity but also abolished the stimulatory effect of PD123319 and HOE140 perfusion on NCC activity. Moreover, the effect of overnight HK loading on Kir4.1/Kir5.1 in the DCT and NCC expression/activity was compromised in the mice treated with AT2R/BK2R antagonists. Renal clearance study showed that inhibition of AT2R and BK2R impairs renal K+ excretion in response to overnight HK loading, and the mice pretreated with PD123319 and HOE140 were hyperkalemic during HK intake. We conclude that synergistic activation of AT2R and BK2R is required for the effect of overnight HK diet on Kir4.1/Kir5.1 in the DCT and NCC activity.

4.
Ecotoxicol Environ Saf ; 188: 109930, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31727496

RESUMO

BACKGROUND: For the sake of children's health, iodized salt supply has been stopped in many areas with excessive iodine in the drinking water, but children's iodine nutrition status and thyroid function after terminating the iodized salt supply is unknown. Objective We assessed the iodine nutrition, thyroid function and influencing factors for thyroid abnormalities in children from areas with different concentrations of water iodine; the supply of iodized salt has been stopped in high water iodine areas. This study aimed to evaluate whether the strategy of stopping the supplies of iodized salt alone is enough to avoid thyroid dysfunction in all areas with excess water iodine while still meeting the iodine nutrition needs of children. METHODS: A cross-sectional study was conducted in children from four areas with different drinking water iodine concentrations in Tianjin, China. The drinking water samplings and spot urine samples were collected to estimate the external and internal iodine exposure levels. The thyroid volume was measured, and blood samples were collected to assess thyroid function. Logistic regression analysis was used to analyze risk factors for thyroid abnormalities. A dietary survey was conducted to determine the sources of iodine nutrition among the areas with different iodine concentrations in the drinking water. RESULTS: In the area with a drinking water iodine concentration ≥300 µg/L, the median urinary iodine concentration (UIC) in children was 476.30 (332.20-639.30) µg/L, which was higher than that in other groups (all P < 0.05), and the prevalence of thyroid nodules and the thyroid goiter rate were higher than those in the <100 µg/L, 100-150 µg/L and 150-300 µg/L areas (all P < 0.01). Binary logistic regression analysis indicated that the risk of thyroid abnormalities was significantly increased in the UIC 200-299 µg/L group (OR: 4.534; 95% CI: 1.565, 13.135; bootstrapped 95% CI: 1.689, 21.206, P = 0.004) and in the UIC ≥ 300 µg/L group (OR: 6.962; 95% CI: 2.490, 19.460; bootstrapped 95% CI: 2.838, 32.570, P = 0.001) compared to the 100-199 µg/L group. The iodine contribution rates from water in areas with water iodine concentrations ≥300 µg/L are up to 63.04%. CONCLUSIONS: After termination of the iodized salt supply, the level of iodine nutrition of children in the area with drinking water iodine concentrations ≥300 µg/L is still excessive. The water source needs to be replaced in this area. In the area with a water iodine concentration of 150-300 µg/L, it is proposed that stopping the supply of iodized salt is sufficient to achieve the proper iodine nutrition status in children.


Assuntos
Exposição Dietética/análise , Água Potável/química , Bócio/epidemiologia , Iodo/administração & dosagem , Iodo/análise , Cloreto de Sódio na Dieta/administração & dosagem , Cloreto de Sódio na Dieta/análise , Criança , China/epidemiologia , Estudos Transversais , Feminino , Bócio/urina , Humanos , Iodo/urina , Masculino , Estado Nutricional , Prevalência , Fatores de Risco , Cloreto de Sódio na Dieta/urina , Inquéritos e Questionários
5.
Dalton Trans ; 48(48): 17925-17935, 2019 Dec 28.
Artigo em Inglês | MEDLINE | ID: mdl-31793567

RESUMO

Three novel copper(ii) complexes, Cu(L1)2 (1), Cu(L2)2·2DMF (2), and Cu(L3)2·2DMF (3), were synthesized using three aroylhydrazone ligands, (E)-2-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL1), (E)-3-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL2) and (E)-4-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide (HL3). The complexes were characterized by elemental analysis, infrared (IR), and Ultraviolet-visible light (UV-vis) spectroscopy. The X-ray crystal structures of the complexes all possess a distorted octahedral coordination geometry. Both an absorption spectral titration and a competitive binding assay (ethidium bromide, 4',6-diamidino-2-phenylindole (DAPI), and methyl green) revealed that complexes 2 and 3 bind readily to calf thymus DNA (ctDNA) through intercalative and minor groove binding modes. Complexes 2 and 3 also exhibited oxidative cleavage of supercoiled plasmid DNA (pUC19) in the presence of ascorbic acid as an activator. Cytotoxicity studies showed that complexes 2 and 3 possessed high cytotoxicities toward the HeLa human cervical cancer cell line, but weak toxicities toward the L929 normal mouse fibroblast cell line. We therefore have reason to believe that complexes 2 and 3 both show potential as promising anticancer candidate drugs.

6.
J Inorg Biochem ; 203: 110919, 2019 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-31783217

RESUMO

In this work, three aroylhydrazone ligands ((E)-2-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide, HL1; (E)-3-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide, HL2; and (E)-4-hydroxy-N'-(1-(pyrazin-2-yl)ethylidene)benzohydrazide, HL3) and their complexes with nickel (Ni(L1)2, NiL1; Ni(L2)2∙2DMF, NiL2; Ni(L3)2∙2DMF, NiL3) were prepared. The single crystal X-ray structures analysis of three compounds showed that they were neutral. The ligand adopts tridentate chelating mode. The nickel ion is six-coordinate with two O atoms and four N atoms from two ligands, and forms an octahedral arrangement. The investigation of DNA binding ability by ultraviolet and fluorescence titrations showed that NiL2 and NiL3 exhibit moderate binding affinity toward calf Thymus DNA. Spectroscopy, molecular docking, and molecular dynamics simulation indicated that NiL2 and NiL3 bind at the minor groove of DNA through intercalation.

7.
Environ Microbiol ; 21(12): 4852-4874, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31667948

RESUMO

Verticillium dahliae is a soil-borne fungus that causes vascular wilt on numerous plants worldwide. The fungus survives in the soil for up to 14 years by producing melanized microsclerotia. The protective function of melanin in abiotic stresses is well documented. Here, we found that the V. dahliae tetraspan transmembrane protein VdSho1, a homolog of the Saccharomyces cerevisiae Sho1, acts as an osmosensor, and is required for plant penetration and melanin biosynthesis. The deletion mutant ΔSho1 was incubated on a cellophane membrane substrate that mimics the plant epidermis, revealing that the penetration of ΔSho1 strain was reduced compared to the wild-type strain. Furthermore, VdSho1 regulates melanin biosynthesis by a signalling mechanism requiring a kinase-kinase signalling module of Vst50-Vst11-Vst7. Strains, ΔVst50, ΔVst7 and ΔVst11 also displayed defective penetration and melanin production like the ΔSho1 strain. Defects in penetration and melanin production in ΔSho1 were restored by overexpression of Vst50, suggesting that Vst50 lies downstream of VdSho1 in the regulatory pathway governing penetration and melanin biosynthesis. Data analyses revealed that the transmembrane portion of VdSho1 was essential for both membrane penetration and melanin production. This study demonstrates that Vst50-Vst11-Vst7 module regulates VdSho1-mediated plant penetration and melanin production in V. dahliae, contributing to virulence.

8.
Anal Chem ; 91(24): 15345-15354, 2019 Dec 17.
Artigo em Inglês | MEDLINE | ID: mdl-31714062

RESUMO

With the ever-increasing demand for improved medical diagnosis, safe food supply, advanced biotechnology, and sustainable ecosystems, characterization of the microbial world by sensitive, specific, rapid, and quantitative tools is gaining more and more attentions. Surface-enhanced Raman spectroscopy (SERS) and its flexible integration with other tools, such as microscopes, stable isotope probing, microfluidic analysis, and chemometric analysis, have advanced rapidly and showed great promise for versatile microbial characterization. This perspective provides an overview of the recent advances of SERS and related coupling techniques for microbial diagnosis (e.g., identification and antibiotic resistance testing), phenotypic response profiling, microbial function assessment, in situ biofilm characterization, and multifunctional SERS tags. We further propose future requirements and the direction for SERS methodology to be headed, including standardization of SERS methodology, database generation and management, coupling with omics technology and single-cell sorting, and application in deciphering microbial processes.

9.
Zhongguo Zhong Yao Za Zhi ; 44(15): 3323-3329, 2019 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-31602890

RESUMO

To study the correlation between ultra high performance liquid chromatography( UPLC) fingerprint of Smilax china and its anti-pelvic inflammatory effect,and to explore the pharmacodynamic material basis of S. china against pelvic inflammatory disease.UPLC fingerprints of 10 batches of S. china from different habitats were established,and the values of SOD,MDA,TNF-α,and IL-6 in rats with pelvic inflammation were measured. The weight of each single pharmacodynamics index to the total efficacy was determined by analytic hierarchy process,and the contribution of each peak in fingerprints to the each single pharmacodynamics index and total efficacy was analyzed by the grey relational analysis. Then the structures of chemical constituents at the identified peaks were confirmed by comparing with the reference substance. The 27 common characteristic peaks of UPLC fingerprints were all related to the anti-pelvic inflammation effect of S. china,of which 13 peaks were identified as peak 2( 3,5-dihydroxy-2-methylbenzoic acid-3-O-glucoside),peak 3( chlorogenic acid),peak 5( 2,7,4-trihydroxydihydroflavone-5-O-glucoside),peak 6( 7,4-dihydroxydihydroflavonol-5-O-glucoside),peak 7( taxifolin-7-O-glucoside),peak 9( taxifolin),peak 10( polydatin),peak 11( oxyresveratrol),peak 12( astilbin),peak15( resveratrol),peak 16( quercitrin),peak 18( engeletin) and peak 24( kaempferol). The correlation degree of 21 peaks and the total efficacy was greater than 0. 8,and the top 10 ranked by correlation degree were as follows: peak 1,3,7,19,18,17,4,11,16,and 21. The results showed that the anti-pelvic inflammation effect of S. china was achieved by the combined action of pharmacodynamic substances. In order to control the quality of S. china and its prepared slices more effectively,the index components of content detection should be selected reasonably.


Assuntos
Doença Inflamatória Pélvica/tratamento farmacológico , Extratos Vegetais/farmacologia , Smilax/química , Animais , China , Cromatografia Líquida de Alta Pressão , Feminino , Compostos Fitoquímicos/farmacologia , Ratos
10.
J Am Soc Nephrol ; 30(8): 1425-1438, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31239388

RESUMO

BACKGROUND: The basolateral potassium channel in the distal convoluted tubule (DCT), comprising the inwardly rectifying potassium channel Kir4.1/Kir5.1 heterotetramer, plays a key role in mediating the effect of dietary potassium intake on the thiazide-sensitive NaCl cotransporter (NCC). The role of Kir5.1 (encoded by Kcnj16) in mediating effects of dietary potassium intake on the NCC and renal potassium excretion is unknown. METHODS: We used electrophysiology, renal clearance, and immunoblotting to study Kir4.1 in the DCT and NCC in Kir5.1 knockout (Kcnj16-/- ) and wild-type (Kcnj16+/+ ) mice fed with normal, high, or low potassium diets. RESULTS: We detected a 40-pS and 20-pS potassium channel in the basolateral membrane of the DCT in wild-type and knockout mice, respectively. Compared with wild-type, Kcnj16-/- mice fed a normal potassium diet had higher basolateral potassium conductance, a more negative DCT membrane potential, higher expression of phosphorylated NCC (pNCC) and total NCC (tNCC), and augmented thiazide-induced natriuresis. Neither high- nor low-potassium diets affected the basolateral DCT's potassium conductance and membrane potential in Kcnj16-/- mice. Although high potassium reduced and low potassium increased the expression of pNCC and tNCC in wild-type mice, these effects were absent in Kcnj16-/- mice. High potassium intake inhibited and low intake augmented thiazide-induced natriuresis in wild-type but not in Kcnj16-/- mice. Compared with wild-type, Kcnj16-/- mice with normal potassium intake had slightly lower plasma potassium but were more hyperkalemic with prolonged high potassium intake and more hypokalemic during potassium restriction. CONCLUSIONS: Kir5.1 is essential for dietary potassium's effect on NCC and for maintaining potassium homeostasis.

11.
Front Chem ; 7: 257, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31139610

RESUMO

High-responsivity phototransistors with a structure of perovskite-WS2 nanosheet composite optical absorber and a reduced graphene oxide (rGO) channel layer is demonstrated via a facile and low-cost solution-processing method. The WS2 nanosheets are dispersed within the perovskite matrix, forming the perovskite-WS2 bulk heterojunction (BHJ). The hybrid phototransistor exhibits excellent figures of merit including high photoresponsivity of 678.8 A/W, high specific detectivity of 4.99 × 1011 Jones, high EQE value of 2.04 × 105% and rapid response to photoswitching. The high photoresponsivity could be attributed to the WS2 nanosheets induced photo-generated electron-hole separation promotion effects due to the selective electron trapping effects in the WS2 nanosheets, together with the high carrier mobility of the rGO channel. This work provides a promising platform for constructing high-responsivity photodetectors.

12.
Front Cell Neurosci ; 13: 134, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31110474

RESUMO

Chemical signaling is ubiquitous and employs a variety of receptor types to detect the cacophony of molecules relevant for each living organism. Insects, our most diverse taxon, have evolved unique olfactory receptors with as little as 10% sequence identity between receptor types. We have identified a promiscuous volatile, 2-methyltetrahydro-3-furanone (coffee furanone), that elicits chemosensory and behavioral activity across multiple insect orders and receptors. In vivo and in vitro physiology showed that coffee furanone was detected by roughly 80% of the recorded neurons expressing the insect-specific olfactory receptor complex in the antenna of Drosophila melanogaster, at concentrations similar to other known, and less promiscuous, ligands. Neurons expressing specialized receptors, other chemoreceptor types, or mutants lacking the complex entirely did not respond to this compound. This indicates that coffee furanone is a promiscuous ligand for the insect olfactory receptor complex itself and did not induce non-specific cellular responses. In addition, we present homology modeling and docking studies with selected olfactory receptors that suggest conserved interaction regions for both coffee furanone and known ligands. Apart from its physiological activity, this known food additive elicits a behavioral response for several insects, including mosquitoes, flies, and cockroaches. A broad-scale behaviorally active molecule non-toxic to humans thus has significant implications for health and agriculture. Coffee furanone serves as a unique tool to unlock molecular, physiological, and behavioral relationships across this diverse receptor family and animal taxa.

13.
Mol Plant Pathol ; 20(6): 857-876, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30957942

RESUMO

Improving genetic resistance is a preferred method to manage Verticillium wilt of cotton and other hosts. Identifying host resistance is difficult because of the dearth of resistance genes against this pathogen. Previously, a novel candidate gene involved in Verticillium wilt resistance was identified by a genome-wide association study using a panel of Gossypium hirsutum accessions. In this study, we cloned the candidate resistance gene from cotton that encodes a protein sharing homology with the TIR-NBS-LRR receptor-like defence protein DSC1 in Arabidopsis thaliana (hereafter named GhDSC1). GhDSC1 expressed at higher levels in response to Verticillium wilt and jasmonic acid (JA) treatment in resistant cotton cultivars as compared to susceptible cultivars and its product was localized to nucleus. The transfer of GhDSC1 to Arabidopsis conferred Verticillium resistance in an A. thaliana dsc1 mutant. This resistance response was associated with reactive oxygen species (ROS) accumulation and increased expression of JA-signalling-related genes. Furthermore, the expression of GhDSC1 in response to Verticillium wilt and JA signalling in A. thaliana displayed expression patterns similar to GhCAMTA3 in cotton under identical conditions, suggesting a coordinated DSC1 and CAMTA3 response in A. thaliana to Verticillium wilt. Analyses of GhDSC1 sequence polymorphism revealed a single nucleotide polymorphism (SNP) difference between resistant and susceptible cotton accessions, within the P-loop motif encoded by GhDSC1. This SNP difference causes ineffective activation of defence response in susceptible cultivars. These results demonstrated that GhDSC1 confers Verticillium resistance in the model plant system of A. thaliana, and therefore represents a suitable candidate for the genetic engineering of Verticillium wilt resistance in cotton.

14.
Fish Physiol Biochem ; 45(3): 829-848, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30843140

RESUMO

The large yellow croaker (Larimichthys crocea) is a marine fish that is economically important to Chinese fisheries, and its reproductive and developmental biology have been extensively investigated. However, the molecular mechanism of oogenesis in L. crocea is not clear. Here, we investigated the multiple vitellogenin (Vtg) system in large yellow croaker. Three different vtg cDNA sequences, including vtgAa, vtgAb and vtgC, were cloned, which indicate the existence of multiple Vtg proteins in large yellow croaker (Lc-Vtgs). Subsequently, the vtg cDNA sequences and predicted Vtg protein structures were analysed, and Vtg protein structures were found to be highly conserved. To research the expression of vtgs during the development of the ovaries, we examined ovarian development and oogenesis by histological analysis. Four stages of ovary development - stages II, III, IV and V - were observed and their boundaries were defined. Soon afterwards, the expression of vtgs in the liver (known as the main site of Vtg synthesis in teleosts) and ovary were analysed. The expression of vtgs was detected in the two tissues. Interestingly, in the early stages of development (stages II and III), there is little or no generation of yolk granules and the expression of vtgs in the liver is low. However, in the late stages (stages IV and V), yolk granules are generated rapidly and the expression of vtgs is significantly increased in the liver. These results support the hypothesis that the Vtgs were synthetized by the liver, and absorbed by the growing oocytes to promote oogenesis in large yellow croaker. We also detected the presence of vtg mRNA in the liver cells and oocytes by in situ hybridization, which indicated that vths were expressed both in the liver and ovaries. Importantly, we found that the distribution of vtgAa and vtgAb mRNA was close to the sites of yolk granule formation in oocytes.


Assuntos
Peixes/crescimento & desenvolvimento , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Ovário/crescimento & desenvolvimento , Transcriptoma , Vitelogeninas/genética , Animais , Clonagem Molecular , Feminino , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Peixes/genética , Peixes/metabolismo , Fígado/crescimento & desenvolvimento , Fígado/metabolismo , Ovário/metabolismo , Filogenia , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Vitelogeninas/metabolismo
15.
Biochim Biophys Acta Mol Basis Dis ; 1865(6): 1113-1125, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-30779947

RESUMO

Vasculogenic mimicry (VM) results in the formation of an alternative circulatory system that can improve the blood supply to multiple malignant tumors, including hepatocellular carcinoma (HCC). However, the potential mechanisms of RhoC/ROCK in VM have not yet been investigated in HCC. Here, RhoC expression was upregulated in HCC tissues, especially the VM-positive (VM+) group, compared to noncancerous tissues (P < 0.01), and patients with high expression of RhoC had shorter survival times (P < 0.001). The knockdown of RhoC via short hairpin RNA (shRNA) in SK-Hep-1 cells significantly decreased VM formation and cell motility. In contrast, cell motility and VM formation were remarkably enhanced when RhoC was overexpressed in HepG2 cells. To further assess the potential role of ROCK1 and ROCK2 on VM, we stably knocked down ROCK1 or ROCK2 in MHCC97H cells. Compared to ROCK1 shRNA, ROCK2 shRNA could largely affect VM formation, cell motility and the key VM factors, as well as the epithelial-mesenchymal transition (EMT) markers in vitro and in vivo. Moreover, p-ERK, p-MEK, p-FAK, p-paxillin, MT1-MMP and MMP2 levels were clearly altered following the overexpression of RhoC, but ROCK2 shRNA had little effect on the expression of p-FAK, which indicated that RhoC regulates FAK/paxillin signaling, but not through ROCK2. In conclusion, our results show that RhoC/ROCK2 may have a major effect on VM in HCC via ERK/MMPs signaling and might be a potential therapeutic target for the treatment of HCC.

16.
Artigo em Inglês | MEDLINE | ID: mdl-30766614

RESUMO

Tumor-associated macrophages (TAMs) with M2 phenotype play an essential role in tumor microenvironment (TME) during the progression and development of numerous cancers and associated with poor prognosis. Thus, regulation of TAMs polarization emerged as a new strategy for tumor immune therapy. According to Traditional Chinese Medicine (TCM) theory, herbs with Qi-tonifying character are involved in improving the defense capacity of immune system. In this study, we screened extracts and ingredients from five Qi-tonifying herbs exhibiting an inhibitory effect on M2 polarization of murine macrophages RAW264.7 induced by IL-4 and IL-13. Among these candidates, total flavonoids from Glycyrrhiza Radix et Rhizoma (TFRG) and ethanol extract of Ginseng Radix et Rhizoma significantly inhibited the expression of Arginase-1 (Arg-1) (above 90% at 100µg/mL), one of the phenotype markers of M2 macrophages. The inhibition of total saponins of Ginseng Radix et Rhizoma, ethanol extract of Cordyceps, ethanol extract of Acanthopanacis senticosi Radix et Rhizoma Seu caulis, and ethanol extract of Astragali Radix reached above 50% at 100µg/mL. The inhibition of ingredients including glabridin, isoliquiritin apioside, lysionotin, cordycepin, astragaloside IV, and calycosin reached above 50% at 50µM. Then, we investigated the molecular mechanisms of TFRG. TFRG abolished the migration of murine breast cancer 4T1 stimulated by the conditioned medium from M2 macrophages (M2-CM). In addition to Arg-1, TFRG also antagonized the IL-4/13-mediated mRNA upregulation of the M2 markers including found in inflammatory zone 1 (FIZZ1), chitinase-3-like protein 3 (YM1), and mannose receptor (CD206) and upregulated the expression of inducible nitric oxide synthase (iNOS), one of the M1 markers. The further exploration showed that TFRG decreased the phosphorylation of STAT6 and increased the expression of miR-155. Our study provides a series of potential immune regulating natural products from five Qi-tonifying herbs on M2 phenotype. For instance, TFRG suppressed M2 polarization of macrophages partly by inactivating STAT6 pathway and enhanced the level of miR-155 to regulate the expressions of M1 and M2 markers.

17.
New Phytol ; 222(2): 1012-1029, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30609067

RESUMO

Verticillium dahliae is a broad host-range pathogen that causes vascular wilts in plants. Interactions between three hosts and specific V. dahliae genotypes result in severe defoliation. The underlying mechanisms of defoliation are unresolved. Genome resequencing, gene deletion and complementation, gene expression analysis, sequence divergence, defoliating phenotype identification, virulence analysis, and quantification of V. dahliae secondary metabolites were performed. Population genomics previously revealed that G-LSR2 was horizontally transferred from the fungus Fusarium oxysporum f. sp. vasinfectum to V. dahliae and is exclusively found in the genomes of defoliating (D) strains. Deletion of seven genes within G-LSR2, designated as VdDf genes, produced the nondefoliation phenotype on cotton, olive, and okra but complementation of two genes restored the defoliation phenotype. Genes VdDf5 and VdDf6 associated with defoliation shared homology with polyketide synthases involved in secondary metabolism, whereas VdDf7 shared homology with proteins involved in the biosynthesis of N-lauroylethanolamine (N-acylethanolamine (NAE) 12:0), a compound that induces defoliation. NAE overbiosynthesis by D strains also appears to disrupt NAE metabolism in cotton by inducing overexpression of fatty acid amide hydrolase. The VdDfs modulate the synthesis and overproduction of secondary metabolites, such as NAE 12:0, that cause defoliation either by altering abscisic acid sensitivity, hormone disruption, or sensitivity to the pathogen.

19.
J Cell Physiol ; 234(6): 8908-8917, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30317616

RESUMO

BACKGROUND: Osteoarthritis (OA) is a kind of chronic osteoarthropathy and degenerative joint disease. Epigenetic regulation in the gene expression dynamics has become increasingly important in OA. We performed a combined analysis of two types of microarray datasets (gene expression and DNA methylation) to identify methylation-based key biomarkers to provide a better understanding of molecular biological mechanisms of OA. METHODS: We obtained two expression profiling datasets (GSE55235, GSE55457) and one DNA methylation profiling data set (GSE63695) from the Gene Expression Omnibus. First, differentially expressed genes (DEGs) between patients with OA and controls were identified using the Limma package in R(v3.4.4). Then, function enrichment analysis of DEGs was performed using a DAVID database. For DNA methylation datasets, ChAMP methylation analysis package was used to identify differential methylation genes (DMGs). Finally, a comprehensive analysis of DEGs and DMGs was conducted to identify genes that exhibited differential expression and methylation simultaneously. RESULTS: We identified 112 DEGs and 2,896 DMGs in patients with OA compared with controls. Functional analysis of DEGs obtained that inflammatory responses, immune responses, and positive regulation of apoptosis, tumor necrosis factor (TNF) signaling pathway, and osteoclast differentiation may be involved in the pathogenesis of OA. Cross-analysis revealed 26 genes that exhibited differential expression and methylation in OA. Among them, ADAMTS9, FKBP5, and PFKBF3 were identified as valuable methylation-based biomarkers for OA. CONCLUSION: In summary, our study identified different molecular features between patients with OA and controls. This may provide new clues for clarifying the pathogenetic mechanisms of OA.

20.
Oncogene ; 38(7): 1106-1120, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30209363

RESUMO

Members of microRNA-200 (miRNA-200) family have a regulatory role in epithelial to mesenchymal transition (EMT) by suppressing Zeb1 and Zeb2 expression. Consistent with its role in suppressing EMT, Hsa-miR-200c-3p (miR-200c), a member of miR-200 family is poorly expressed in mesenchymal-like triple-negative breast cancer (TNBC) cells and ectopic miR-200c expression suppresses cell migration. In this study, we demonstrated that miR-200c potently inhibited TNBC cell growth and tumor development in a mechanism distinct from its ability to downregulate Zeb1 and Zeb2 expression, because silencing them only marginally affected TNBC cell growth. We identified phosphodiesterase 7B (PDE7B) as a bona fide miR-200c target. Importantly, miR-200c-led inhibition in cell growth and tumor development was prevented by forcing PDE7B transgene expression, while knockdown of PDE7B effectively inhibited cell growth. These results suggest that miR-200c inhibits cell growth by targeting PDE7B mRNA. To elucidate mechanism underlying miR-200c/PDE7B regulation of TNBC cell growth, we showed that cAMP concentration was lower in TNBC cells compared with estrogen receptor-positive (ER + ) cells, and that both miR-200c and PDE7B siRNAs were able to increase cAMP concentration in TNBC cells. High level of cellular cAMP has been shown to induce cell cycle arrest and apoptosis in TNBC cells. Our observation that ectopic expression of miR-200c triggered apoptosis indicates that it does so by elevating level of cellular cAMP. Analysis of breast tumor gene expression datasets revealed an inverse association between miR-200c and PDE7B expression. Especially, both low miR-200c and high PDE7B expression were correlated with poor survival of breast cancer patients. Our study supports a critical role of miR-200c/PDE7B relationship in TNBC tumorigenesis.


Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 7/biossíntese , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , MicroRNAs/biossíntese , Proteínas de Neoplasias/biossíntese , RNA Neoplásico/biossíntese , Neoplasias de Mama Triplo Negativas/metabolismo , Linhagem Celular Tumoral , AMP Cíclico/genética , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 7/genética , Feminino , Humanos , MicroRNAs/genética , Proteínas de Neoplasias/genética , RNA Neoplásico/genética , Neoplasias de Mama Triplo Negativas/mortalidade , Neoplasias de Mama Triplo Negativas/patologia
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