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1.
EMBO J ; 39(6): e103412, 2020 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-32090355

RESUMO

Bacterial cyclic-di-GMP (c-di-GMP) production is associated with biofilm development and the switch from acute to chronic infections. In Pseudomonas aeruginosa, the diguanylate cyclase (DGC) SiaD and phosphatase SiaA, which are co-transcribed as part of a siaABCD operon, are essential for cellular aggregation. However, the detailed functions of this operon and the relationships among its constituent genes are unknown. Here, we demonstrate that the siaABCD operon encodes for a signaling network that regulates SiaD enzymatic activity to control biofilm and aggregates formation. Through protein-protein interaction, SiaC promotes SiaD diguanylate cyclase activity. Biochemical and structural data revealed that SiaB is an unusual protein kinase that phosphorylates SiaC, whereas SiaA phosphatase can dephosphorylate SiaC. The phosphorylation state of SiaC is critical for its interaction with SiaD, which will switch on or off the DGC activity of SiaD and regulate c-di-GMP levels and subsequent virulence phenotypes. Collectively, our data provide insights into the molecular mechanisms underlying the modulation of DGC activity associated with chronic infections, which may facilitate the development of antimicrobial drugs.

2.
J Agric Food Chem ; 68(1): 17-32, 2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31809036

RESUMO

Weeds had caused significant loss for crop production in the process of agriculture. Herbicides have played an important role in securing crop production. However, the high reliance on herbicides has led to environmental issues as well as the evolution of herbicide resistance. Thus, there is an urgent need for new herbicides with safer toxicological profiles and novel modes of action. Actinomycetes produce very diverse bioactive compounds, of which some show potent biopesticidal activity. The herbicidal secondary metabolites from actinomycetes can be classified into several groups, such as amino acids, peptides, nucleosides, macrolides, lactones, amide, amines, etc., some of which have been successfully developed as commercial herbicides. The structure diversity and evolved biological activity of secondary metabolites from actinomycetes can offer opportunities for the development of both directly used bioherbicides and synthetic herbicides with new target sites, and thus, this review focuses on the structure, herbicidal activity, and modes of action of secondary metabolites from actinomycetes.


Assuntos
Actinobacteria/química , Herbicidas/química , Herbicidas/farmacologia , Actinobacteria/metabolismo , Descoberta de Drogas , Herbicidas/metabolismo , Plantas Daninhas/efeitos dos fármacos , Plantas Daninhas/crescimento & desenvolvimento , Metabolismo Secundário , Controle de Plantas Daninhas
3.
Chem Biol Interact ; 316: 108921, 2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31838053

RESUMO

Hyperproliferation and oxidative stress induced by hyperglycemia in mesangial cells plays crucial roles in the pathological process of diabetic nephropathy. Farrerol, isolated from rhododendron leaves, possesses broad anti-oxidative and anti-inflammatory properties towards several diseases, but its role in diabetic neuropathy remains unclear. The aim of this study was to evaluate the effects of farrerol in high glucose induced mesangial cell injury, and to explore underlying molecular mechanisms. Our results showed that high glucose in vitro conditions significantly stimulated cell proliferation, inflammatory cytokine secretion, extracellular matrix deposition, excessive oxidative stress, and NADPH oxidase activity in mesangial cells. Levels of NADPH oxidase 4 (Nox4) expression, ERK1/2 phosphorylation, and TGF-ß1/Smad2 activation were significantly induced by high glucose conditions in mesangial cells. Inversely, farrerol treatments at 40, 60, and 80 µM concentrations, dose-dependently alleviated this molecular damage by high glucose in mesangial cells. We also found that restoration of Nox4 expression abolished the protective effects of farrerol on high glucose-induced proliferation and reactive oxygen species generation. Furthermore, pretreatment with the Nox4 inhibitor diphenyliodonium or the ERK1/2 pathway inhibitor PD98059, displayed similar ameliorated effects of farrerol on high glucose-induced mesangial cell damage. Taken together, these data suggest that farrerol displays protective effects on high glucose induced mesangial cell injury, partly through the Nox4-mediated ROS/ERK1/2 signaling pathway. These observations may provide novel insights into the application of farrerol as a diabetic neuropathy treatment.


Assuntos
Cromonas/farmacologia , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , NADPH Oxidase 4/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Glucose/toxicidade , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Células Mesangiais/citologia , Células Mesangiais/efeitos dos fármacos , Células Mesangiais/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ratos , Fator de Crescimento Transformador beta/metabolismo
4.
J Hazard Mater ; 382: 121103, 2020 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-31479824

RESUMO

An in situ freeze method (in liquid N2) is developed to study the adsorption behaviours of volatile organic compounds (VOCs). Using the proposed method, the adsorption state and adsorption mechanism of adsorbates (pore filling effect or mono/multilayer stacking) in pores of adsorbent can be perceived clearly. We can also acquire the contribution rate of each pore to adsorption capacity. It is found that for toluene adsorption, more pores smaller than around 1.1 nm should be made in activated carbon if it's below 100 ppm, while more pores of around 1.2-5 nm should be made if it's beyond 1000 ppm. This measurement method presents a more comprehensive description on the adsorption state of VOCs in micro- and mesopores, extending the development of adsorbent design and adsorption mechanism in chemical and environmental engineering.

5.
PLoS Pathog ; 15(12): e1008198, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31790504

RESUMO

The type VI secretion system (T6SS) is widely distributed in Gram-negative bacteria, whose function is known to translocate substrates to eukaryotic and prokaryotic target cells to cause host damage or as a weapon for interbacterial competition. Pseudomonas aeruginosa encodes three distinct T6SS clusters (H1-, H2-, and H3-T6SS). The H1-T6SS-dependent substrates have been identified and well characterized; however, only limited information is available for the H2- and H3-T6SSs since relatively fewer substrates for them have yet been established. Here, we obtained P. aeruginosa H2-T6SS-dependent secretomes and further characterized the H2-T6SS-dependent copper (Cu2+)-binding effector azurin (Azu). Our data showed that both azu and H2-T6SS were repressed by CueR and were induced by low concentrations of Cu2+. We also identified the Azu-interacting partner OprC, a Cu2+-specific TonB-dependent outer membrane transporter. Similar to H2-T6SS genes and azu, expression of oprC was directly regulated by CueR and was induced by low Cu2+. In addition, the Azu-OprC-mediated Cu2+ transport system is critical for P. aeruginosa cells in bacterial competition and virulence. Our findings provide insights for understanding the diverse functions of T6SSs and the role of metal ions for P. aeruginosa in bacteria-bacteria competition.


Assuntos
Proteínas de Bactérias/metabolismo , Cobre/metabolismo , Proteínas de Ligação a DNA/metabolismo , Pseudomonas aeruginosa/patogenicidade , Sistemas de Secreção Tipo VI/metabolismo , Virulência/fisiologia , Animais , Camundongos , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/metabolismo
6.
BMC Neurol ; 19(1): 320, 2019 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-31830942

RESUMO

BACKGROUND: X-linked agammaglobulinaemia (XLA) is a rare inherited primary immunodeficiency disease characterized by the B cell developmental defect, caused by mutations in the gene coding for Bruton's tyrosine kinase (BTK), which may cause serious recurrent infections. The diagnosis of XLA is sometimes challenging because a few number of patients have higher levels of serum immunoglobulins than expected. In this study, we reported an atypical case with recurrent meningitis, delayed diagnosis with XLA by genetic analysis at the second episode of meningitis at the age of 8 years. CASE REPORT: An 8-year-old Chinese boy presented with fever, dizziness and recurrent vomiting for 3 days. The cerebrospinal fluid (CSF) and magnetic resonance imaging (MRI) results were suggestive of bacterial meningoencephalitis, despite the negative gram staining and cultures of the CSF. The patient was treated with broad-spectrum antibiotics and responded well to the treatment. He had history of another episode of acute pneumococci meningitis 4 years before. The respective level of Immunoglobulin G (IgG), Immunoglobulin A (IgA) and Immunoglobulin M (IgM) was 4.85 g/L, 0.93 g/L and 0.1 g/L at 1st episode, whereas 1.9 g/L, 0.27 g/L and 0 g/L at second episode. The B lymphocytes were 0.21 and 0.06% of peripheral blood lymphocytes at first and second episode respectively. Sequencing of the BTK coding regions showed that the patient had a point mutation in the intron 14, hemizyous c.1349 + 5G > A, while his mother had a heterozygous mutation. It was a splice site mutation predicted to lead to exon skipping and cause a truncated BTK protein. CONCLUSION: Immunity function should be routinely checked in patients with severe intracranial bacterial infection. Absence of B cells even with normal level of serum immunoglobulin suggests the possibility of XLA, although this happens only in rare instances. Mutational analysis of BTK gene is crucial for accurate diagnosis to atypical patients with XLA.

7.
Oncogenesis ; 8(12): 74, 2019 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-31822656

RESUMO

Postsurgical recurrence within 2 years is the major cause of poor survival of hepatocellular carcinoma (HCC) patients. However, the molecular mechanism underlying HCC recurrence remains unclear. Here, we distinguish the function and mechanism of Sec62 in promoting HCC recurrence. The correlation between Sec62 and early recurrence was demonstrated in 60 HCC samples from a prospective study. HCC cells with Sec62 knockdown (Sec62KD) or overexpression (Sec62OE) were used to determine the potential of Sec62 in cell migration in vitro. Microarray analysis comparing Sec62KD or Sec62OE to their control counterparts was used to explore the mechanisms of Sec62-induced recurrence. A luciferase-labelled orthotopic nude mouse model of HCC with Sec62KD or Sec62OE was used to validate the potential of Sec62 in early HCC recurrence in vivo. We found that high expression of Sec62 was positively correlated with surgical recurrence in clinical HCC samples. Multivariate analysis revealed that Sec62 was an independent prognostic factor for early recurrence in postoperative HCC patients. Moreover, Sec62 promoted migration and invasion of HCC cells in vitro and postsurgical recurrence in vivo. Mechanically, integrinα/CAV1 signalling was identified as one of the targets of Sec62 in cell movement. Overexpression of integrin α partially rescued the Sec62 knockdown-induced inhibition of cell migration. Sec62 is a potentially prognostic factor for early recurrence in postoperative HCC patients and promotes HCC metastasis through integrinα/CAV1 signalling. Sec62 might be an attractive drug target for combating HCC postsurgical recurrence.

8.
J Zhejiang Univ Sci B ; 20(12): 1021-1026, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31749349

RESUMO

Mycoplasma infection is the most prevalent contamination in cell culture. Analysis of cell culture in laboratories from different countries shows that mycoplasma contamination ranges from 15% to 80% and, in some cases, even reaches 100% (Chernov et al., 2014). Whilst mycoplasma infection is not visible to the naked eye in cell culture, the consequences of mycoplasma contamination have been shown to induce a number of cellular changes, for example, increased resistance to chemotherapeutic drugs. Therefore, any results obtained from tissue culture studies, in the presence of mycoplasma contamination, potentially render the data invalid (Kim et al., 2015; Gedye et al., 2016). As such, mycoplasmas are not harmless bystanders and cannot be ignored in in vitro studies.

9.
Food Funct ; 10(11): 7152-7163, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31596288

RESUMO

This study demonstrated different effects of bone morphogenetic protein 4 (BMP4) and retinoic acid (RA) signaling on the induction of germ cell formation in chickens. In vitro, BMP4 significantly promoted primordial germ cell (PGC) formation, while RA promoted spermatogonial stem cell (SSC) formation. Hematoxylin-Eosin (HE) staining of reproductive ridge and testicular slices showed that BMP4 signaling was activated during PGC formation but was inhibited during PGC differentiation into SSC. In contrast, RA signaling was significantly activated during PGC differentiation to SSC. Mechanistically, elevated expression of phosphorylated mothers against decapentaplegic homolog 5 (p-Smad5) activated BMP4 signaling, while inhibition of p-Smad5 significantly reduced the PGC formation. Additionally, BMP4 regulated the PGC formation through histone acetylation and DNA methylation in deleted in azoospermia-like (DAZL) gene. Luciferase report showed RA binding to RARα regulated stimulated by RA 8 (Stra8) promoter activity during SSC formation, while mutations in RAR binding sites inhibited the Stra8 expression and SSC formation. Further, both HAT and HDAC regulated the RARα isoform, and HAT binding to RARα activated the Stra8 transcription. RNA-seq of embryonic stem cells (ESC), PGC, and SSC showed inverse expression of genes related to the BMP4 and RA pathways during PGC and SSC formation. Additionally, Smad5 and Smurf were critical for the interactions between the two pathways. Specifically, through Smurf promotion of Smad5 ubiquitination, RA could inhibit the BMP4 signal transduction. In conclusion, the BMP4 and RA signaling pathways play opposing roles in germ cell formation, driven by epigenetic processes such as phosphorylation, ubiquitination, and histone acetylation.


Assuntos
Células-Tronco Germinativas Adultas/metabolismo , Proteína Morfogenética Óssea 4/metabolismo , Células Germinativas/metabolismo , Tretinoína/metabolismo , Células-Tronco Germinativas Adultas/citologia , Animais , Diferenciação Celular , Galinhas , Feminino , Regulação da Expressão Gênica , Células Germinativas/citologia , Masculino , Receptor alfa de Ácido Retinoico/genética , Receptor alfa de Ácido Retinoico/metabolismo , Transdução de Sinais , Proteína Smad5/genética , Proteína Smad5/metabolismo
10.
Asia Pac Allergy ; 9(3): e28, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31384583

RESUMO

Eosinophilic esophagitis (EoE) is a recently recognized esophageal inflammatory disease with clinical manifestations arising from esophageal dysfunction. The etiology of EoE is currently being clarified and food allergy is evolving as the central cornerstone of EoE disease pathogenesis. Given the large number of eosinophils in the esophagus of people with EoE verified by data from murine models EoE is widely considered as the hallmark T-helper type 2 (Th2) disease of the esophagus. It is also known that some eosinophilic inflammation is controlled by other subsets of T cells such as Th9 or Th17 and control is also exerted by type 2 innate lymphoid cells acting together with basophils. In this paper we review results from molecular studies of mouse models in light of the results from the first clinical trials targeting key cytokines in humans and present in-depth molecular understanding of EoE.

11.
Langmuir ; 35(27): 8927-8934, 2019 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-31199161

RESUMO

The pore surface structure and chemistry of the ordered mesoporous silica adsorbent (KIT-6) were modified for toluene removal, including the substituent groups on the silica surface and the pore size uniformity. Uniform pore size was obtained by low-temperature heat treatment of the template at 200 °C. In the dynamic adsorption process, better channel uniformity led to higher adsorption capacity under no humidity condition because of faster pore diffusion. However, better channel uniformity resulted in poor hydrophobicity under high humidity condition because it favors the adsorption of water vapor to a greater extent. The result of Biot number indicated that triphenyl-grafted KIT-6 had a faster intraparticle mass transfer rate than pure KIT-6. Triphenyl-grafted KIT-6 had a higher adsorption capacity for toluene as compared to phenyl-grafted KIT-6 under no humidity condition because of its higher surface area ( SA). The higher SA was owing to the low modification of phenyl, which was caused by the isolated grafting of silicon triphenyl rather than a more even coverage by silicon phenyl. As a result, triphenyl-grafted KIT-6 exposed more hydrophilic Si-O-Si groups and therefore was less hydrophobic than phenyl-grafted KIT-6.

12.
Artigo em Inglês | MEDLINE | ID: mdl-31157178

RESUMO

Glutathione (GSH) is the most abundant antioxidant in all living organisms. Previously, we have shown that a deletion mutant in the glutathione synthetase gene (ΔgshB) decreases the expression of type III secretion system (T3SS) genes of Pseudomonas aeruginosa. However, the mechanism remains elusive. In this study, a comprehensive transcriptomic analysis of the GSH-deficient mutant ΔgshAΔgshB was used to elucidate the role of GSH in the pathogenesis of P. aeruginosa. The data show that the expression of genes in T3SS, type VI secretion system (T6SS) and some regulatory genes were impaired. ΔgshAΔgshB was attenuated in a mouse model of acute pneumonia, swimming and swarming motilities, and biofilm formation. Under T3SS inducing conditions, GSH enhanced the expression of T3SS in both wild-type PAO1 and ΔgshAΔgshB, but not in Δvfr. Genetic complementation of Δvfr restored the ability of GSH to induce the expression of T3SS genes. Site-directed mutagenesis based substitution of cysteine residues with alanine in Vfr protein abolished the induction of T3SS genes by GSH, confirming that GSH regulates T3SS genes through Vfr. Exposure to H2O2 decreased free thiol content on Vfr, indicating that the protein was sensitive to redox modification. Importantly, GSH restored the oxidized Vfr to reduced state. Collectively, these results suggest that GSH serves as an intracellular redox signal sensed by Vfr to upregulate T3SS expression in P. aeruginosa. Our work provides new insights into the role of GSH in P. aeruginosa pathogenesis.


Assuntos
Proteínas de Bactérias/metabolismo , Proteína Receptora de AMP Cíclico/metabolismo , Glutationa/metabolismo , Pseudomonas aeruginosa/metabolismo , Sistemas de Secreção Tipo III/metabolismo , Animais , Proteínas de Bactérias/genética , Biofilmes/crescimento & desenvolvimento , Proteína Receptora de AMP Cíclico/genética , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Glutationa Sintase/genética , Glutationa Sintase/metabolismo , Peróxido de Hidrogênio/metabolismo , Masculino , Camundongos , Mutagênese Sítio-Dirigida , Mutação , Pneumonia , Infecções por Pseudomonas , Pseudomonas aeruginosa/genética , Sistemas de Secreção Tipo III/genética , Virulência/genética
13.
Front Microbiol ; 10: 560, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30949153

RESUMO

Pseudomonas aeruginosa is an important human pathogen which uses the type III secretion system (T3SS) as a primary virulence factor to establish infections in humans. The results presented in this report revealed that the ATP-binding protein PA4595 (named ArtR, a Regulator that is an ATP-activated Repressor of T3SS) represses T3SS expression in P. aeruginosa. The expression of T3SS genes, including exoS, exoY, exoT, exsCEBA, and exsD-pscB-L, increased significantly when artR was knockout. The effect of ArtR on ExsA is at the transcriptional level, not at the translational level. The regulatory role and cytoplasm localization of ArtR suggest it belongs to the REG sub-family of ATP-binding cassette (ABC) family. Purified GST-tagged ArtR showed ATPase activity in vitro. The conserved aspartate residues in the dual Walker B motifs prove to be essential for the regulatory function of ArtR. The regulation of T3SS by ArtR is unique, which does not involve the known GacS/A-RsmY/Z-RsmA-ExsA pathway or Vfr. This is the first REG subfamily of ATP-binding cassette that is reported to regulate T3SS genes in bacteria. The results specify a novel player in the regulatory networks of T3SS in P. aeruginosa.

14.
Exp Ther Med ; 17(5): 4154-4166, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30988793

RESUMO

The use of bone marrow mesenchymal stem cells (BMSCs) has great potential in cell therapy, particularly in the orthopedic field. BMSCs represent a valuable renewable cell source that have been successfully utilized to treat damaged skeletal tissue and bone defects. BMSCs can be induced to differentiate into osteogenic lineages via the addition of inducers to the growth medium. The present study examined the effects of all-trans retinoic acid (ATRA) and curcumin on the osteogenic differentiation of mouse BMSCs. Morphological changes, the expression levels of the bone-associated gene markers bone morphogenetic protein 2, runt-related transcription factor and osterix during differentiation, an in vitro mineralization assay, and changes in osteocalcin expression revealed that curcumin supplementation promoted the osteogenic differentiation of BMSCs. By contrast, the application of ATRA increased osteogenic differentiation during the early stages, but during the later stages, it decreased the mineralization of differentiated cells. In addition, to the best of our knowledge, the present study is the first to examine the effect of curcumin on the osteogenic potency of mouse embryonic fibroblasts (MEFs) after reprogramming with human lim mineralization protein (hLMP-3), which is a positive osteogenic regulator. The results revealed that curcumin-supplemented culture medium increased hLMP-3 osteogenic potency compared with that of MEFs cultured in the non-supplemented medium. The present results demonstrate that enrichment of the osteogenic culture medium with curcumin, a natural osteogenic inducer, increased the osteogenic differentiation capacity of BMSCs as well as that of MEFs reprogrammed with hLMP-3.

15.
Org Biomol Chem ; 17(14): 3635-3639, 2019 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-30916700

RESUMO

Oxazoles are an important class of biologically active metabolites from nature, and exhibit broad biological activities as the lead for drug discovery. Hinduchelins are a class of unusual natural products with an oxazole unit, isolated from Streptoalloteichus hindustanus, and with a potential iron-chelating ability. These compounds are the first identified naturally occurring unusual oxazole derivatives to possess a catechol unit. However, some of these compounds are not abundant in nature, and thus, the efficient syntheses of these compounds are advantageous in exploring their potential applications. This paper reports the efficient synthesis and bio-evaluation of hinduchelins A-D and their derivatives with convenient procedures and high yields.


Assuntos
Antibacterianos/farmacologia , Antifúngicos/farmacologia , Antineoplásicos/farmacologia , Produtos Biológicos/farmacologia , Inseticidas/farmacologia , Oxazóis/farmacologia , Actinomycetales/química , Animais , Antibacterianos/síntese química , Antibacterianos/química , Antifúngicos/síntese química , Antifúngicos/química , Antineoplásicos/síntese química , Antineoplásicos/química , Afídeos/efeitos dos fármacos , Produtos Biológicos/síntese química , Produtos Biológicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Fungos/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Humanos , Inseticidas/síntese química , Inseticidas/química , Testes de Sensibilidade Microbiana , Mariposas , Oxazóis/síntese química , Oxazóis/química
16.
Eur J Med Chem ; 168: 293-300, 2019 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-30826506

RESUMO

Pityriacitrin is a marine alkaloid with typical ß-carboline scaffold, and which has been proven to exhibit diverse biological functions. During the course of our research for highly active compounds from natural products, the pityriacitrin have also been isolated and identified from a Chinese Burkholderia sp. NBF227. So, in order to explore the potential functional molecules, a series of ß-carboline analogues derived from pityriacitrin were designed and synthesized, and their in vitro cytotoxic activities against SGC-7901, A875, HepG2, and MARC145 cell lines were evaluated. The results demonstrated that some of these ß-carboline derivatives exhibited moderate to good cytotoxic activities, especially, compound 9o with a special sulfonyl group presented the highest inhibitory activities against all tested cell lines with the IC50 values of 6.82 ±â€¯0.98, 8.43 ±â€¯1.93, 7.69 ±â€¯2.17, 7.19 ±â€¯1.43 µM, respectively, which might be used as lead compound for discovery of novel cytotoxic agents.


Assuntos
Alcaloides/farmacologia , Antineoplásicos/farmacologia , Produtos Biológicos/farmacologia , Carbolinas/farmacologia , Alcaloides/química , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Produtos Biológicos/síntese química , Produtos Biológicos/química , Burkholderia/química , Carbolinas/síntese química , Carbolinas/química , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Relação Estrutura-Atividade
17.
J Cell Biochem ; 2019 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-30656739

RESUMO

Recently, the surface marker genes of spermatogonial stem cells (SSCs) were increasingly excavated and verified. However, few studies focused on the key genes involved in the regulation of SSCs differentiation. Our laboratory has screened the Lbc gene (GenBank accession number: XM_429585.3), which is specifically expressed on the SSCs. The aim of this study is to investigate the function of Lbc and its regulatory mechanism for SSCs. The indirect immunofluorescence assay (IFA) showed that Lbc was located in both nucleus and cytoplasm. Lbc was also overexpressed and knocked out both in vitro and in vivo to verify its function in SSCs, respectively. As a result, the overexpressed Lbc could promote the formation of spermatogonial stem cells like cells (SSCs-like), while the deficiency of Lbc blocked the formation of SSCs-like. We also identified the core region of Lbc promoter that located into the upstream of the transcription initiation site -247 to -2bp. Moreover, the activity of Lbc promoter could be increased by histone acetylation which is leading to the higher expression of Lbc. When we mutated the transcription factor HOXA5 and SOX10 that bound to the core region of Lbc promoter, HOXA5 could reduce the transcription activity of Lbc whereas the SOX10 was not. Currently, we found Lbc is a new specific marker of SSCs. This gene can be modified by histone acetylated and promote the formation of chicken SSCs via the transcription factor HOXA5. The present research will lay the foundation for further study on the regulatory mechanism of SSCs.

18.
Phytomedicine ; 54: 365-370, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30217547

RESUMO

BACKGROUND: Considering that the quality control indicators in Chinese medicine (CM) are disconnected from safety and effectiveness, Prof. Chang-xiao Liu et al. has proposed a concept regarding the quality marker (Q-marker) of CM to promote the healthy development of the CM industry and improve the CM quality control method. PURPOSE: In this study, we proposed a strategy to discover and verify the toxicity Q-marker of CM based on network toxicology. METHODS: First, traditional biochemical pathology indicators and sensitive biomarkers were used to predict the toxicity of CM. Next, the chemical composition of toxic CMs and their metabolites were rapidly identified by multidimensional detection techniques. Subsequently, the interaction network between "toxicity - toxic chemical composition - toxic target - effect pathway" was built through network toxicology, and the potential toxicity Q-marker of CM was initially screened. Finally, the chemical properties of toxicity Q-markers were verified by traceability and testability. RESULTS: Based on the predicted results of network toxicology, the toxic compounds of CM were preliminarily identified, and the toxic mechanism was comprehensively interpreted. In the context of definite biological properties and chemical properties, the toxicity Q-marker was finally confirmed. CONCLUSION: This extensive review provides a study method for the toxicity Q-marker of CM, which helps to systemically and thoroughly reveal the internal toxicity mechanism of CM. The in-depth study of the toxicity Q-marker provides the material basis and technical support for the safety evaluation of CM.


Assuntos
Biomarcadores/análise , Medicamentos de Ervas Chinesas/farmacologia , Medicina Tradicional Chinesa/normas , Humanos , Medicina Tradicional Chinesa/efeitos adversos , Controle de Qualidade
19.
J Cell Biochem ; 120(1): 332-342, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30277598

RESUMO

To better understand the mechanisms in transcriptional regulation, we analyzed the promoters of the reprogramming key genes Sox2, c-Myc, and Oct4. Here, we cloned different 5' deletions of the goat Sox2, c-Myc, and Oct4 promoters, and evaluated their functions by green fluorescent protein reporter system and dual-luciferase reporter system. Site-directed mugagenesis and epigenetic modifiers were used to explore the influence of transcription binding sites and epigenetic status on the promoters. The results suggested that the basal promoters were located in the - 109 to 49, - 147 to 1, and - 96 to 30 bp regions of the Sox2, c-Myc, and Oct4 promoters. The transcription factors that identified to influence the Sox2, c-Myc, and Oct4 promoter activities were Elf-1 and activating protein 2 (AP-2), C/EBP and Sp1, and Mzf1 and Sp1, respectively. The epigenetic alternation of the Sox2, c-Myc, and Oct4 promoters by 5-aza-2'-deoxycytidine or/and trichostatin A significantly increased the promoter activities. In conclusion, the result determined the core promoter areas of the Sox2, c-Myc, and Oct4 genes, and identified the transcription factors that influence their promoter activities. We also verified that the Sox2, c-Myc, and Oct4 promoters were hypermethylated and hypoacetylated.


Assuntos
Fator 3 de Transcrição de Octâmero/genética , Proteínas Proto-Oncogênicas c-myc/genética , Fatores de Transcrição SOXB1/genética , Acetilação , Animais , Sítios de Ligação/genética , Linhagem Celular Tumoral , Metilação de DNA/genética , Deleção de Genes , Regulação da Expressão Gênica , Cabras/embriologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Histonas/metabolismo , Camundongos , Microscopia de Fluorescência , Filogenia , Plasmídeos , Regiões Promotoras Genéticas , Vírus 40 dos Símios/genética , Ativação Transcricional , Transfecção
20.
Int J Biochem Cell Biol ; 106: 84-95, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30453092

RESUMO

Large bone defects and bone loss after fractures remain significant challenges for orthopedic surgeons. Our study aims to find an available, applicable and biological treatment for bone regeneration overcoming the limitations in ESC/iPSC technology. We directly reprogrammed the mouse embryonic fibroblast (MEF) into osteoblast cells using different combinations of Yamanaka factors with human lim mineralization protein-3 (hLMP-3). LMP is an intracellular LIM-domain protein acting as an effective positive regulator of the osteoblast differentiation. After transduction, cells were cultured in osteogenic medium, and then examined for osteoblast formation. The expression of osteogenic markers (BMP2, Runx2 and Osterix) during reprogramming and in vitro mineralization assay revealed that the best reprogramming cocktail was (c-Myc - Oct4) with hLMP-3. In addition, both immunofluorescent staining and western blot analysis confirmed that osteocalcin (OCN) expression increased in the cells treated with the c-Myc/Oct4/hLMP3 cocktail than using hLMP-3 alone. Furthermore, this reprogramming cocktail showed efficient healing in an induced femoral bone defect in rat animal model one month after transplantation. In the present study, we reported for the first time the effect of combining Yamanaka factors with hLMP-3 to induce osteoblast cells from MEF both in vitro and in vivo.


Assuntos
Reprogramação Celular , Embrião de Mamíferos/metabolismo , Fibroblastos/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Proteínas com Domínio LIM/biossíntese , Osteoblastos/metabolismo , Animais , Antígenos de Diferenciação/biossíntese , Antígenos de Diferenciação/genética , Técnicas de Reprogramação Celular , Embrião de Mamíferos/citologia , Fibroblastos/citologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas com Domínio LIM/genética , Camundongos , Osteoblastos/citologia
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