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1.
Braz Oral Res ; 33: e117, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31939498

RESUMO

The aim of this study was to evaluate the effect of mineral trioxide aggregate (MTA) and Brazilian propolis on the cell viability, mineralization, anti-inflammatory ability, and migration of human dental pulp cells (hDPCs). The cell viability was evaluated with CCK-8 kit after 1, 5, 7, and 9 days. The deposition of calcified matrix and the expression of osteogenesis-related genes were evaluated by Alizarin Red staining and real-time PCR after incubation in osteogenic medium for 21 days. The expression of inflammation-related genes in cells was determined after exposure to 1 µg/mL LPS for 3 h. Finally, the numbers of cells that migrated through the permeable membranes were compared during 15 h. Propolis and MTA significantly increased the viability of hDPCscompared to the control group on days 7 and 9. In the propolis group, significant enhancement of osteogenic potential and suppressed expression of IL-1ß and IL-6 was observed after LPS exposure compared to the MTA and control groups. The number of migration cells in the propolis group was similar to that of the control group, while MTA significantly promoted cell migration. Propolis showed comparable cell viability to that of MTA and exhibited significantly higher anti-inflammatory and mineralization promotion effects on hDPCs.


Assuntos
Compostos de Alumínio/farmacologia , Anti-Inflamatórios/farmacologia , Compostos de Cálcio/farmacologia , Polpa Dentária/citologia , Polpa Dentária/efeitos dos fármacos , Óxidos/farmacologia , Própole/farmacologia , Silicatos/farmacologia , Antraquinonas , Brasil , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Combinação de Medicamentos , Humanos , Interleucina-1beta/análise , Interleucina-6/análise , Odontoblastos/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo Real , Reprodutibilidade dos Testes , Estatísticas não Paramétricas , Fator de Necrose Tumoral alfa/análise
2.
Elife ; 82019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31571584

RESUMO

Free heme is cytotoxic as exemplified by hemolytic diseases and genetic deficiencies in heme recycling and detoxifying pathways. Thus, intracellular accumulation of heme has not been observed in mammalian cells to date. Here we show that mice deficient for the heme transporter SLC48A1 (also known as HRG1) accumulate over ten-fold excess heme in reticuloendothelial macrophage lysosomes that are 10 to 100 times larger than normal. Macrophages tolerate these high concentrations of heme by crystallizing them into hemozoin, which heretofore has only been found in blood-feeding organisms. SLC48A1 deficiency results in impaired erythroid maturation and an inability to systemically respond to iron deficiency. Complete heme tolerance requires a fully-operational heme degradation pathway as haplo insufficiency of HMOX1 combined with SLC48A1 inactivation causes perinatal lethality demonstrating synthetic lethal interactions between heme transport and degradation. Our studies establish the formation of hemozoin by mammals as a previously unsuspected heme tolerance pathway.


Assuntos
Heme/toxicidade , Hemeproteínas/metabolismo , Macrófagos/metabolismo , Animais , Heme Oxigenase-1/metabolismo , Hemeproteínas/deficiência , Proteínas de Membrana/metabolismo , Camundongos
3.
J Med Econ ; : 1-7, 2019 Sep 24.
Artigo em Inglês | MEDLINE | ID: mdl-31505982

RESUMO

Aims: To evaluate the risk-of-hospitalization (ROH) models developed at Blue Cross Blue Shield of Louisiana (BCBSLA) and compare this approach to the DxCG risk-score algorithms utilized by many health plans. Materials and Methods: Time zero for this study was December 31, 2016. BCBSLA members were eligible for study inclusion if they were fully insured; aged 80 years or younger; and had continuous enrollment starting on or before June 1, 2016, through time zero. Up to 2 years of historical claims data from time zero per patient was included for model development. Members were excluded if they had cancer, renal failure, or were admitted for hospice. The Blue Cross ROH models were developed using (1) regularized logistic regression and (2) random decision forests (a tree ensemble learning classification method). All models were generated using Scikit-learn: Machine Learning in Python. Prognostic capabilities of DxCG risk-score algorithms were compared to those of the Blue Cross models. Results: When stratifying by the top 0.1% of members with the highest ROH, the Blue Cross logistic regression model had the highest area under the receiving operator characteristics curve (0.862) based on the result of 10-fold cross-validation. The Blue Cross random decision forests model had the highest positive predictive value (49.0%) and positive likelihood ratio (61.4), but sensitivity, specificity, negative predictive values, and negative likelihood ratios were similar across all four models. Limitations: The Blue Cross ROH models were developed and evaluated using BCBSLA data, and predictive power may fluctuate if applied to other databases. Conclusions: The predictability of the Blue Cross models show how member-specific, regional data can be used to accurately identify patients with a high ROH, which may allow healthcare workers to intervene earlier and subsequently reduce the healthcare burden for patients and providers.

4.
Stem Cells Dev ; 28(22): 1514-1526, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31544584

RESUMO

Retinitis pigmentosa (RP) is a hereditary disease characterized by degeneration and the loss of photoreceptors. Stem cell-based therapy has emerged as a promising strategy for treating RP. Stem cells from exfoliated deciduous teeth (SHEDs), a type of mesenchymal stem cell from human exfoliated deciduous teeth, have the potential to differentiate into photoreceptor-like cells under specific induction in vitro. It has been confirmed that through paracrine secreta, SHEDs exert neurotrophic, angiogenic, immunoregulatory, and antiapoptotic functions in injured tissues. This study was designed to determine whether retinal-differentiated SHEDs and the conditioned medium derived from SHEDs (SHEDs-CM) have therapeutic effects in a mouse model of RP. The results showed that both SHEDs and SHEDs-CM improved electroretinogram responses, ameliorated photoreceptor degeneration, and maintained the structure of the outer segments of photoreceptors. The therapeutic effects were related to antiapoptotic activity of SHEDs and SHEDs-CM. Thus, SHEDs may be a promising stem cell source for treating retinal degeneration.

5.
Nat Chem Biol ; 15(1): 27-33, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30510189

RESUMO

Stimulated by thromboxane A2, an endogenous arachidonic acid metabolite, the thromboxane A2 receptor (TP) plays a pivotal role in cardiovascular homeostasis, and thus is considered as an important drug target for cardiovascular disease. Here, we report crystal structures of the human TP bound to two nonprostanoid antagonists, ramatroban and daltroban, at 2.5 Å and 3.0 Å resolution, respectively. The TP structures reveal a ligand-binding pocket capped by two layers of extracellular loops that are stabilized by two disulfide bonds, limiting ligand access from the extracellular milieu. These structures provide details of interactions between the receptor and antagonists, which help to integrate previous mutagenesis and SAR data. Molecular docking of prostanoid-like ligands, combined with mutagenesis, ligand-binding and functional assays, suggests a prostanoid binding mode that may also be adopted by other prostanoid receptors. These insights into TP deepen our understanding about ligand recognition and selectivity mechanisms of this physiologically important receptor.


Assuntos
Receptores de Tromboxano A2 e Prostaglandina H2/química , Receptores de Tromboxano A2 e Prostaglandina H2/metabolismo , Sítios de Ligação , Carbazóis/química , Carbazóis/metabolismo , Cristalografia por Raios X , Dissulfetos/química , Humanos , Ligantes , Simulação de Acoplamento Molecular , Fenilacetatos/química , Fenilacetatos/metabolismo , Conformação Proteica , Receptores de Tromboxano A2 e Prostaglandina H2/antagonistas & inibidores , Sulfonamidas/química , Sulfonamidas/metabolismo
6.
Commun Biol ; 1: 120, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30272000

RESUMO

Dietary iron absorption is regulated by duodenal cytochrome b (Dcytb), an integral membrane protein that catalyzes reduction of nonheme Fe3+ by electron transfer from ascorbate across the membrane. This step is essential to enable iron uptake by the divalent metal transporter. Here we report the crystallographic structures of human Dcytb and its complex with ascorbate and Zn2+. Each monomer of the homodimeric protein possesses cytoplasmic and apical heme groups, as well as cytoplasmic and apical ascorbate-binding sites located adjacent to each heme. Zn2+ coordinates to two hydroxyl groups of the apical ascorbate and to a histidine residue. Biochemical analysis indicates that Fe3+ competes with Zn2+ for this binding site. These results provide a structural basis for the mechanism by which Fe3+ uptake is promoted by reducing agents and should facilitate structure-based development of improved agents for absorption of orally administered iron.

7.
Int J Mol Sci ; 19(7)2018 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-30036949

RESUMO

G protein-coupled receptors represent the largest family of human membrane proteins and are modulated by a variety of drugs and endogenous ligands. Molecular modeling techniques, especially enhanced sampling methods, have provided significant insight into the mechanism of GPCR⁻ligand recognition. Notably, the crucial role of the membrane in the ligand-receptor association process has earned much attention. Additionally, docking, together with more accurate free energy calculation methods, is playing an important role in the design of novel compounds targeting GPCRs. Here, we summarize the recent progress in the computational studies focusing on the above issues. In the future, with continuous improvement in both computational hardware and algorithms, molecular modeling would serve as an indispensable tool in a wider scope of the research concerning GPCR⁻ligand recognition as well as drug design targeting GPCRs.


Assuntos
Desenho de Drogas , Receptores Acoplados a Proteínas-G/química , Receptores Acoplados a Proteínas-G/metabolismo , Algoritmos , Animais , Humanos , Ligantes , Modelos Moleculares , Simulação de Dinâmica Molecular , Ligação Proteica , Relação Estrutura-Atividade
8.
J Chem Theory Comput ; 14(5): 2761-2770, 2018 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-29660291

RESUMO

The crystal structure of P2Y1 receptor (P2Y1R), a class A GPCR, revealed a special extra-helical site for its antagonist, BPTU, which locates in-between the membrane and the protein. However, due to the limitation of crystallization experiments, the membrane was mimicked by use of detergents, and the information related to the binding of BPTU to the receptor in the membrane environment is rather limited. In the present work, we conducted a total of ∼7.5 µs all-atom simulations in explicit solvent using conventional molecular dynamics and multiple enhanced sampling methods, with models of BPTU and a POPC bilayer, both in the absence and presence of P2Y1R. Our simulations revealed that BPTU prefers partitioning into the interface of polar/lipophilic region of the lipid bilayer before associating with the receptor. Then, it interacts with the second extracellular loop of the receptor and reaches the binding site through the lipid-receptor interface. In addition, by use of funnel-metadynamics simulations which efficiently enhance the sampling of bound and unbound states, we provide a statistically accurate description of the underlying binding free energy landscape. The calculated absolute ligand-receptor binding affinity is in excellent agreement with the experimental data (Δ Gb0_theo = -11.5 kcal mol-1, Δ Gb0_exp= -11.7 kcal mol-1). Our study broadens the view of the current experimental/theoretical models and our understanding of the protein-ligand recognition mechanism in the lipid environment. The strategy used in this work is potentially applicable to investigate ligands association/dissociation with other membrane-embedded sites, allowing identification of compounds targeting membrane receptors of pharmacological interest.


Assuntos
Proteínas de Membrana/química , Receptores Purinérgicos P2Y1/química , Sítios de Ligação , Ligações de Hidrogênio , Ligantes , Bicamadas Lipídicas/química , Simulação de Dinâmica Molecular , Fosfatidilcolinas/química , Conformação Proteica , Termodinâmica
9.
J Mol Biol ; 430(8): 1081-1083, 2018 04 13.
Artigo em Inglês | MEDLINE | ID: mdl-29510174

Assuntos
Citocromos c , Heme , Cisteína
10.
Acta Biomater ; 72: 206-216, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29567106

RESUMO

While polysaccharide-based nanocarriers have been recognized for their crucial roles in tumor theranostics, the industrial-scale production of nanotherapeutics still remains a significant challenge. Most current approaches adopt a postpolymerization self-assembly strategy that follows a separate synthetic step and thus suffers from subgram scale yields and a limited range of application. In this study, we demonstrate the kilogram-scale formation of polysaccharide-polyacrylate nanocarriers at concentrations of up to 5 wt% through a one-pot approach - starting from various acrylate monomers and polysaccharides - that combines aspects of hydrophobicity-induced self-assembly with the free radical graft copolymerization of acrylate monomers from polysaccharide backbones into a single process that is thus denoted as a graft copolymerization induced self-assembly. We also demonstrate that this novel approach is applicable to a broad range of polysaccharides and acrylates. Notably, by choosing a crosslinker that bears a disulfide group and two vinyl capping groups to structurally lock the nanocarriers, the products are rendered cleavable in the reducing environments encountered at tumor sites and thus provide ideal candidates for the construction of anticancer nanotherapeutic systems. In vitro and in vivo studies demonstrated that the use of this nanocarrier for the delivery of doxorubicin hydrochloride (DOX) significantly decreased the side effects of DOX and improved the bio-safety of the chemotherapy accordingly. STATEMENT OF SIGNIFICANCE: While polysaccharide-based nanocarriers have been recognized for their crucial roles in tumor theranostics, the industrial-scale production of these nanotherapeutics still remains a significant challenge. Most current approaches adopt a post-polymerization self-assembly strategy which that follows a separate synthetic step, and thus suffers from sub-gram scale yields and a limited range of application. In this study, the hydrophobic effect was combined with free radical polymerization to facilitate the graft copolymerization-induced self-assembly (GISA) of acrylate monomers with various hydrophobicities to construct cleavable polysaccharide-polyacrylate nanocarriers at a high efficiency with excellent potential for industrial-scale production. We envision that these nanocarriers will contribute to the development of tumor nanotheranostics that combine the biological functionalities of polysaccharides with the unmatched application-specific flexibility of nanocarriers.


Assuntos
Doxorrubicina , Portadores de Fármacos , Nanoestruturas , Neoplasias Experimentais/tratamento farmacológico , Polissacarídeos , Animais , Preparações de Ação Retardada/química , Preparações de Ação Retardada/farmacocinética , Preparações de Ação Retardada/farmacologia , Doxorrubicina/química , Doxorrubicina/farmacocinética , Doxorrubicina/farmacologia , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Portadores de Fármacos/farmacologia , Células HeLa , Humanos , Células MCF-7 , Camundongos Nus , Nanoestruturas/química , Nanoestruturas/uso terapêutico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Polissacarídeos/química , Polissacarídeos/farmacocinética , Polissacarídeos/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Anal Chem ; 90(5): 3395-3401, 2018 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-29401392

RESUMO

Heme, a hydrophobic and cytotoxic macrocycle, is an essential cofactor in a large number of proteins and is important for cell signaling. This must mean that heme is mobilized from its place of synthesis or entry into the cell to other parts of the cell where hemoproteins reside. However, the cellular dynamics of heme movement is not well understood, in large part due to the inability to image heme noninvasively in live biological systems. Here, using high-resolution transient absorption microscopy, we showed that heme storage and distribution is dynamic in Caenorhabditis elegans. Intracellular heme exists in concentrated granular puncta which localizes to lysosomal-related organelles. These granules are dynamic, and their breaking down into smaller granules provides a mechanism by which heme stores can be mobilized. Collectively, these direct and noninvasive dynamic imaging techniques provide new insights into heme storage and transport and open a new avenue for label-free investigation of heme function and regulation in living systems.


Assuntos
Caenorhabditis elegans/ultraestrutura , Heme/análise , Animais , Células HEK293 , Humanos , Lisossomos/ultraestrutura , Microscopia/métodos , Imagem Óptica/métodos
12.
J Med Chem ; 60(24): 10231-10244, 2017 12 28.
Artigo em Inglês | MEDLINE | ID: mdl-29193967

RESUMO

Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a promising therapeutic target for atherosclerosis, Alzheimer's disease, and diabetic macular edema. Here we report the identification of novel sulfonamide scaffold Lp-PLA2 inhibitors derived from a relatively weak fragment. Similarity searching on this fragment followed by molecular docking leads to the discovery of a micromolar inhibitor with a 300-fold potency improvement. Subsequently, by the application of a structure-guided design strategy, a successful hit-to-lead optimization was achieved and a number of Lp-PLA2 inhibitors with single-digit nanomolar potency were obtained. After preliminary evaluation of the properties of drug-likeness in vitro and in vivo, compound 37 stands out from this congeneric series of inhibitors for good inhibitory activity and favorable oral bioavailability in male Sprague-Dawley rats, providing a quality candidate for further development. The present study thus clearly demonstrates the power and advantage of integrally employing fragment screening, crystal structures determination, virtual screening, and medicinal chemistry in an efficient lead discovery project, providing a good example for structure-based drug design.


Assuntos
1-Alquil-2-acetilglicerofosfocolina Esterase/antagonistas & inibidores , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Relação Estrutura-Atividade , Sulfonamidas/química , Sulfonamidas/farmacologia , 1-Alquil-2-acetilglicerofosfocolina Esterase/química , Administração Oral , Animais , Disponibilidade Biológica , Células CACO-2 , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos/métodos , Estabilidade de Medicamentos , Inibidores Enzimáticos/administração & dosagem , Inibidores Enzimáticos/farmacocinética , Hepatócitos/efeitos dos fármacos , Humanos , Masculino , Simulação de Acoplamento Molecular , Ratos Sprague-Dawley , Sulfonamidas/administração & dosagem , Sulfonamidas/farmacocinética
13.
Am J Manag Care ; 23(12): e402-e408, 2017 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-29261245

RESUMO

OBJECTIVES: This study aimed to investigate the role of the Quality Blue Primary Care (QBPC) program on healthcare utilization and overall cost among the beneficiaries of Blue Cross and Blue Shield of Louisiana (BCBSLA). STUDY DESIGN: Retrospective observational cohort study using claims data from adults residing in QBPC-implemented regions continuously enrolled through BCBSLA from June 2012 to December 2014 (N = 89,034). METHODS: Controlling for age, gender, and risk score by propensity score weighting, inpatient, outpatient, and corresponding medical expenditures were each compared between the QBPC group and the control group using a difference-in-differences regression model. RESULTS: Average total cost increased in both the QBPC and control groups in 2014, but the increase was significantly less in the intervention group-a difference of $27.09 per member per month (PMPM) (P ≤.001). Savings in total cost were derived largely from a decrease in hospitalizations-a difference of $18.85 PMPM (P = .0023). Furthermore, savings were associated with shifts in healthcare utilization by the intervention group toward proactive management, including increased primary care physician visits (P = .0106) and higher screening rates for diabetes (P = .0019). Inpatient admissions also decreased in the QBPC group, most significantly among those with chronic conditions (P <.05). Conversely, an unexpected increase was observed in emergency department visits. CONCLUSIONS: The QBPC program was associated with a shift in primary care delivery and reductions in overall cost. Savings were achieved largely through reductions in hospitalization costs.


Assuntos
Planos de Seguro Blue Cross Blue Shield/economia , Assistência ao Paciente/economia , Atenção Primária à Saúde/economia , Qualidade da Assistência à Saúde/economia , Estudos de Coortes , Humanos , Louisiana , Reembolso de Incentivo/economia , Estudos Retrospectivos , Estados Unidos
14.
FEBS J ; 284(19): 3278-3301, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28783254

RESUMO

Extracellular hemoglobin, a byproduct of hemolysis, can release its prosthetic heme groups upon oxidation. This produces metabolically active heme that is exchangeable between acceptor proteins, macromolecules and low molecular weight ligands, termed here labile heme. As it accumulates in plasma labile heme acts in a pro-oxidant manner and regulates cellular metabolism while exerting pro-inflammatory and cytotoxic effects that foster the pathogenesis of hemolytic diseases. Here, we developed and characterized a panel of heme-specific single domain antibodies (sdAbs) that together with a cellular-based heme reporter assay, allow for quantification and characterization of labile heme in plasma during hemolytic conditions. Using these approaches, we demonstrate that when generated during hemolytic conditions labile heme is bound to plasma molecules with an affinity higher than 10-7 m and that 2-8% (~ 2-5 µm) of the total amount of heme detected in plasma can be internalized by bystander cells, termed here bioavailable heme. Acute, but not chronic, hemolysis is associated with transient reduction of plasma heme-binding capacity, that is, the ability of plasma molecules to bind labile heme with an affinity higher than 10-7 m. The heme-specific sdAbs neutralize the pro-oxidant activity of soluble heme in vitro, suggesting that these maybe used to counter the pathologic effects of labile heme during hemolytic conditions. Finally, we show that heme-specific sdAbs can be used to visualize cellular heme. In conclusion, we describe a panel of heme-specific sdAbs that when used with other approaches provide novel insights to the pathophysiology of heme.


Assuntos
Anticorpos Monoclonais/química , Eritrócitos/química , Heme/análise , Biblioteca de Peptídeos , Anticorpos de Domínio Único/química , Sequência de Aminoácidos , Anemia Falciforme/sangue , Animais , Anticorpos Monoclonais/biossíntese , Afinidade de Anticorpos , Especificidade de Anticorpos , Biotina/química , Clonagem Molecular , Ensaio de Imunoadsorção Enzimática , Eritrócitos/parasitologia , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/virologia , Expressão Gênica , Heme/química , Heme/imunologia , Heme/metabolismo , Hemoglobinas/química , Hemoglobinas/metabolismo , Hemólise , Humanos , Cinética , Camundongos , Camundongos Endogâmicos C57BL , Oxirredução , Plasmodium falciparum/crescimento & desenvolvimento , Ligação Proteica , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Anticorpos de Domínio Único/biossíntese , Tetrapirróis/química , Tetrapirróis/metabolismo
15.
Sci Rep ; 7(1): 3219, 2017 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-28607438

RESUMO

Acetylcholinesterase, with a deep, narrow active-site gorge, attracts enormous interest due to its particularly high catalytic efficiency and its inhibitors used for treatment of Alzheimer's disease. To facilitate the massive pass-through of the substrate and inhibitors, "breathing" motions to modulate the size of the gorge are an important prerequisite. However, the molecular mechanism that governs such motions is not well explored. Here, to systematically investigate intrinsic motions of the enzyme, we performed microsecond molecular dynamics simulations on the monomer and dimer of Torpedo californica acetylcholinesterase (TcAChE) as well as the complex of TcAChE bound with the drug E2020. It has been revealed that protein-ligand interactions and dimerization both keep the gorge in bulk, and opening events of the gorge increase dramatically compared to the monomer. Dynamics of three subdomains, S3, S4 and the Ω-loop, are tightly associated with variations of the gorge size while the dynamics can be changed by ligand binding or protein dimerization. Moreover, high correlations among these subdomains provide a basis for remote residues allosterically modulating the gorge motions. These observations are propitious to expand our understanding of protein structure and function as well as providing clues for performing structure-based drug design.


Assuntos
Acetilcolinesterase/metabolismo , Donepezila/metabolismo , Proteínas de Peixes/metabolismo , Simulação de Dinâmica Molecular , Acetilcolinesterase/química , Regulação Alostérica , Animais , Biocatálise/efeitos dos fármacos , Domínio Catalítico , Inibidores da Colinesterase/química , Inibidores da Colinesterase/metabolismo , Inibidores da Colinesterase/farmacologia , Donepezila/química , Donepezila/farmacologia , Proteínas de Peixes/química , Movimento (Física) , Ligação Proteica , Domínios Proteicos , Multimerização Proteica , Torpedo/metabolismo
16.
Chemistry ; 23(54): 13387-13403, 2017 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-28657690

RESUMO

Researchers have gained a deeper understanding of DNA-based encryption and its effectiveness in enhancing information security in recent years. However, there are many theoretical and technical issues about DNA-based encryption that need to be addressed before it can be effectively used in the field of security. Currently, the most popular DNA-based encryption schemes are based on traditional cryptography and the integration of existing DNA technology. These schemes are not completely based on DNA computing and biotechnology. Herein, as inspired by nature, encryption based on DNA has been developed, which is, in turn, based on two fundamental biological axioms about DNA sequencing: 1) DNA sequencing is difficult under the conditions of not knowing the correct sequencing primers and probes, and 2) without knowing the correct probe, it is difficult to decipher precisely and sequence the information of unknown and mixed DNA/peptide nucleic acid (PNA) probes, which only differ in nucleotide sequence, arranged on DNA chips (microarrays). In essence, when creating DNA-based encryption by means of biological technologies, such as DNA chips and polymerase chain reaction (PCR) amplification, the encryption method discussed herein cannot be decrypted, unless the DNA/PNA probe or PCR amplification is known. The biological analysis, mathematical analysis, and simulation results demonstrate the feasibility of the method, which provides much stronger security and reliability than that of traditional encryption methods.


Assuntos
DNA/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Sequência de Bases , DNA/química , Primers do DNA/química , Primers do DNA/metabolismo , Sondas de DNA/química , Sondas de DNA/metabolismo , Ácidos Nucleicos Peptídicos/metabolismo , Reação em Cadeia da Polimerase , Software
17.
Nat Cell Biol ; 19(7): 799-807, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28581477

RESUMO

Growing evidence in vertebrates predicts that cellular haem levels in animals are maintained not only by a cell's internal capacity for haem synthesis in a cell-autonomous manner, but also by an inter-organ haem trafficking network through cell-non-autonomous regulation. Using Caenorhabditis elegans, a genetically and optically amenable animal model for visualizing haem-dependent signalling, we show that HRG-7, a protein with homology to aspartic proteases, mediates inter-organ signalling between the intestine and extra-intestinal tissues. Intestinal HRG-7 functions as a secreted signalling factor during haem starvation in extra-intestinal tissues and is regulated through a DBL-1, homologous to BMP5, dependent signal from neurons. Given the evidence that vertebrate homologues exist for each of the components of the HRG-7-mediated signalling pathway, it is conceivable that the cell-non-autonomous signalling framework that we uncovered in C. elegans may have functional relevance for inter-organ regulation of iron and haem metabolism in humans.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Heme/metabolismo , Hemeproteínas/metabolismo , Mucosa Intestinal/metabolismo , Transdução de Sinais , Animais , Animais Geneticamente Modificados , Transporte Biológico , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Regulação da Expressão Gênica , Heme/deficiência , Hemeproteínas/genética , Homeostase , Neurônios/metabolismo , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Interferência de RNA , Fatores de Tempo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo
18.
Biosens Bioelectron ; 92: 563-569, 2017 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-27836591

RESUMO

Electrode materials play a key role in the development of electrochemical sensors, particularly enzyme-based biosensors. Here, a novel NiCo2S4 with reticulated hollow spheres assembled from rod-like structures was prepared by a one-pot solvothermal method and its formation mechanism was discussed. Moreover, comparison of NiCo2S4 materials from different experiment conditions as biosensors was investigated by electrochemical impedance spectroscopy (EIS) and differential pulse voltammetry (DPV), and the best one that was reticulated hollow spheres assembled from rod-like structures NiCo2S4 has been successfully employed as a matrix of AChE immobilization for the special structure, superior conductivity and rich reaction active sites. When using common two kinds of organophosphate pesticides (OPs) as model analyte, the biosensors demonstrated a wide linear range of 1.0×10-12-1.0×10-8gmL-1 with the detection limit of 4.2×10-13gmL-1 for methyl parathion, and 1.0×10-13-1.0×10-10gmL-1 with the detection limit of 3.5×10-14gmL-1 for paraoxon, respectively. The proposed biosensors exhibited many advantages such as acceptable stability and low cost, providing a promising tool for analysis of OPs.


Assuntos
Técnicas Biossensoriais/métodos , Cobalto/química , Metil Paration/análise , Níquel/química , Paraoxon/análise , Praguicidas/análise , Sulfetos/química , Acetilcolinesterase/metabolismo , Inibidores da Colinesterase/análise , Inibidores da Colinesterase/metabolismo , Técnicas Eletroquímicas/métodos , Enzimas Imobilizadas/metabolismo , Limite de Detecção , Metil Paration/metabolismo , Paraoxon/metabolismo , Praguicidas/metabolismo
19.
Proc Natl Acad Sci U S A ; 113(35): E5144-52, 2016 08 30.
Artigo em Inglês | MEDLINE | ID: mdl-27528661

RESUMO

Heme is an essential prosthetic group in proteins that reside in virtually every subcellular compartment performing diverse biological functions. Irrespective of whether heme is synthesized in the mitochondria or imported from the environment, this hydrophobic and potentially toxic metalloporphyrin has to be trafficked across membrane barriers, a concept heretofore poorly understood. Here we show, using subcellular-targeted, genetically encoded hemoprotein peroxidase reporters, that both extracellular and endogenous heme contribute to cellular labile heme and that extracellular heme can be transported and used in toto by hemoproteins in all six subcellular compartments examined. The reporters are robust, show large signal-to-background ratio, and provide sufficient range to detect changes in intracellular labile heme. Restoration of reporter activity by heme is organelle-specific, with the Golgi and endoplasmic reticulum being important sites for both exogenous and endogenous heme trafficking. Expression of peroxidase reporters in Caenorhabditis elegans shows that environmental heme influences labile heme in a tissue-dependent manner; reporter activity in the intestine shows a linear increase compared with muscle or hypodermis, with the lowest heme threshold in neurons. Our results demonstrate that the trafficking pathways for exogenous and endogenous heme are distinct, with intrinsic preference for specific subcellular compartments. We anticipate our results will serve as a heuristic paradigm for more sophisticated studies on heme trafficking in cellular and whole-animal models.


Assuntos
Heme/metabolismo , Hemeproteínas/metabolismo , Espaço Intracelular/metabolismo , Peroxidase/metabolismo , Animais , Animais Geneticamente Modificados , Transporte Biológico , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Membrana Celular/metabolismo , Endocitose , Células HEK293 , Heme/química , Humanos , Microscopia Confocal , Organelas/metabolismo , Peroxidase/química , Peroxidase/genética
20.
FASEB J ; 30(10): 3501-3514, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27363426

RESUMO

Nematodes lack a heme biosynthetic pathway and must acquire heme from exogenous sources. Given the indispensable role of heme, this auxotrophy may be exploited to develop drugs that interfere with heme uptake in parasites. Although multiple heme-responsive genes (HRGs) have been characterized within the free-living nematode Caenorhabditis elegans, we have undertaken the first study of heme transport in Brugia malayi, a causative agent of lymphatic filariasis. Through functional assays in yeast, as well as heme analog, RNAi, and transcriptomic experiments, we have shown that the heme transporter B. malayi HRG-1 (BmHRG-1) is indeed functional in B. malayi In addition, BmHRG-1 localizes both to the endocytic compartments and cell membrane when expressed in yeast cells. Transcriptomic sequencing revealed that BmHRG-1, BmHRG-2, and BmMRP-5 (all orthologs of HRGs in C. elegans) are down-regulated in heme-treated B. malayi, as compared to non-heme-treated control worms. Likely because of short gene lengths, multiple exons, other HRGs in B. malayi (BmHRG-3-6) remain unidentified. Although the precise mechanisms of heme homeostasis in a nematode with the ability to acquire heme remains unknown, this study clearly demonstrates that the filarial nematode B. malayi is capable of transporting exogenous heme.-Luck, A. N., Yuan, X., Voronin, D., Slatko, B. E., Hamza, I., Foster, J. M. Heme acquisition in the parasitic filarial nematode Brugia malayi.


Assuntos
Brugia Malayi , Heme/imunologia , Homeostase/fisiologia , Animais , Caenorhabditis elegans , Interferência de RNA
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