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1.
Structure ; 32(4): 424-432.e4, 2024 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-38325370

RESUMO

The design of small-molecule-binding proteins requires protein backbones that contain cavities. Previous design efforts were based on naturally occurring cavity-containing backbone architectures. Here, we designed diverse cavity-containing backbones without predefined architectures by introducing tailored restraints into the backbone sampling driven by SCUBA (Side Chain-Unknown Backbone Arrangement), a neural network statistical energy function. For 521 out of 5816 designs, the root-mean-square deviations (RMSDs) of the Cα atoms for the AlphaFold2-predicted structures and our designed structures are within 2.0 Å. We experimentally tested 10 designed proteins and determined the crystal structures of two of them. One closely agrees with the designed model, while the other forms a domain-swapped dimer, where the partial structures are in agreement with the designed structures. Our results indicate that data-driven methods such as SCUBA hold great potential for designing de novo proteins with tailored small-molecule-binding function.


Assuntos
Redes Neurais de Computação , Proteínas , Conformação Proteica , Modelos Moleculares , Proteínas/química
2.
ACS Omega ; 8(37): 33392-33402, 2023 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-37744847

RESUMO

Heat stroke (HS) is a potentially fatal acute condition caused by an interplay of complex events including inflammation, endothelial injury, and coagulation abnormalities that make its pharmacological treatment a challenging problem. The traditional Chinese medicine Xuebijing injection (XBJ) has been shown to reduce inflammatory responses and prevent organ injuries in HS-induced mice. However, the underlying mechanism of XBJ in HS-induced brain injury remains unclear. In this study, HS-induced rat models and cell models were established to elucidate the effects and underlying mechanisms of XBJ injection on HS-induced brain injury in vivo and in vitro. The results revealed that XBJ injection improved the survival outcome of HS rats and attenuated HS-induced brain injury in a concentration-dependent manner. Subsequently, the reduction in viability and proliferation of neurons induced by HS were reversed by XBJ treatment, while the HS-induced increased ROS levels and neuron death were also inhibited by XBJ injection. Mechanistically, HS activated PARP-1/AIF signaling in vitro and in vivo, inducing the translocation of AIF from the cytoplasm to the nucleus, leading to PARP-1-dependent cell death of neurons. Additionally, we compared XBJ injection effects in young and old age rats. Results showed that XBJ also provided protective effects in HS-induced brain injury in aging rats; however, the treatment efficacy of XBJ injection at the same concentration was more significant in the young age rats. In conclusion, XBJ injection attenuates HS-induced brain injury by inhibiting oxidative stress and Parthanatos via the PARP-1/AIF signaling, which might provide a novel therapeutic strategy for HS treatment.

3.
Mol Cell ; 83(14): 2493-2508.e5, 2023 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-37343553

RESUMO

Type IV CRISPR-Cas systems, which are primarily found on plasmids and exhibit a strong plasmid-targeting preference, are the only one of the six known CRISPR-Cas types for which the mechanistic details of their function remain unknown. Here, we provide high-resolution functional snapshots of type IV-A Csf complexes before and after target dsDNA binding, either in the absence or presence of CasDinG, revealing the mechanisms underlying CsfcrRNA complex assembly, "DWN" PAM-dependent dsDNA targeting, R-loop formation, and CasDinG recruitment. Furthermore, we establish that CasDinG, a signature DinG family helicase, harbors ssDNA-stimulated ATPase activity and ATP-dependent 5'-3' DNA helicase activity. In addition, we show that CasDinG unwinds the non-target strand (NTS) and target strand (TS) of target dsDNA from the CsfcrRNA complex. These molecular details advance our mechanistic understanding of type IV-A CRISPR-Csf function and should enable Csf complexes to be harnessed as genome-engineering tools for biotechnological applications.


Assuntos
Proteínas Associadas a CRISPR , DNA , DNA/genética , DNA de Cadeia Simples/genética , Sistemas CRISPR-Cas , Proteínas Associadas a CRISPR/metabolismo
4.
Int J Mol Sci ; 24(2)2023 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-36674645

RESUMO

Arap3, a dual GTPase-activating protein (GAP) for the small GTPases Arf6 and RhoA, plays key roles in regulating a wide range of biological processes, including cancer cell invasion and metastasis. It is known that Arap3 is a PI3K effector that can bind directly to PI(3,4,5)P3, and the PI(3,4,5)P3-mediated plasma membrane recruitment is crucial for its function. However, the molecular mechanism of how the protein recognizes PI(3,4,5)P3 remains unclear. Here, using liposome pull-down and surface plasmon resonance (SPR) analysis, we found that the N-terminal first pleckstrin homology (PH) domain (Arap3-PH1) can interact with PI(3,4,5)P3 and, with lower affinity, with PI(4,5)P2. To understand how Arap3-PH1 and phosphoinositide (PIP) lipids interact, we solved the crystal structure of the Arap3-PH1 in the apo form and complex with diC4-PI(3,4,5)P3. We also characterized the interactions of Arap3-PH1 with diC4-PI(3,4,5)P3 and diC4-PI(4,5)P2 in solution by nuclear magnetic resonance (NMR) spectroscopy. Furthermore, we found overexpression of Arap3 could inhibit breast cancer cell invasion in vitro, and the PIPs-binding ability of the PH1 domain is essential for this function.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Proteínas Ativadoras de GTPase , Fosfatidilinositóis , Humanos , Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Ativadoras de GTPase/química , Invasividade Neoplásica , Fosfatos de Fosfatidilinositol/metabolismo , Ligação Proteica , Domínios Proteicos
5.
J Mol Biol ; 434(19): 167756, 2022 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-35870648

RESUMO

Histone chaperones, which constitute an interaction and functional network involved in all aspects of histone metabolism, have to date been identified only in eukaryotes. The Epstein-Barr virus tegument protein BKRF4 is a histone-binding protein that engages histones H2A-H2B and H3-H4, and cellular chromatin, inhibiting the host DNA damage response. Here, we identified BKRF4 as a bona fide viral histone chaperone whose histone-binding domain (HBD) forms a co-chaperone complex with the human histone chaperone ASF1 in vitro. We determined the crystal structures of the quaternary complex of the BKRF4 HBD with human H3-H4 dimer and the histone chaperone ASF1b and the ternary complex of the BKRF4 HBD with human H2A-H2B dimer. Through structural and biochemical studies, we elucidated the molecular basis for H3-H4 and H2A-H2B recognition by BKRF4. We also revealed two conserved motifs, D/EL and DEF/Y/W, within the BKRF4 HBD, which may represent common motifs through which histone chaperones target H3-H4 and H2A-H2B, respectively. In conclusion, our results identify BKRF4 as a histone chaperone encoded by the Epstein-Barr virus, representing a typical histone chaperone found in a non-eukaryote. We envision that more histone chaperones await identification and characterization in DNA viruses and even archaea.


Assuntos
Proteínas do Capsídeo , Proteínas de Ciclo Celular , Herpesvirus Humano 4 , Chaperonas de Histonas , Proteínas do Capsídeo/química , Proteínas de Ciclo Celular/química , Cromatina/química , Herpesvirus Humano 4/genética , Chaperonas de Histonas/química , Histonas/metabolismo , Humanos , Ligação Proteica , Conformação Proteica
6.
BMC Infect Dis ; 22(1): 662, 2022 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-35907785

RESUMO

BACKGROUND: We aimed to explore the prognostic utilities of C-reactive protein (CRP), procalcitonin (PCT), neutrophil CD64 (nCD64) index, in combination or alone, in septic patients. METHODS: We retrospectively included 349 septic patients (based on Sepsis 3.0 definition). The primary outcome was 28-day all-cause mortality. Cox regression model, receiver-operating characteristic (ROC) curve, reclassification analysis, Kaplan-Meier survival curves were performed to evaluate the predictive efficacy of the above parameters. RESULTS: CRP, nCD64 index were independent predictors of 28-day mortality for sepsis in the Cox regression model [CRP, HR 1.004 (95% CI 1.002-1.006), P < 0.001; nCD64 index, HR 1.263 (95% CI 1.187-1.345, P < 0.001]. Area under the ROC curve (AUC) of CRP, PCT, nCD64 index, nCD64 index plus PCT, nCD64 index plus CRP, were 0.798 (95% CI 0.752-0.839), 0.833 (95% CI 0.790-0.871), 0.906 (95% CI 0.870-0.935), 0.910 (95% CI 0.875-0.938), 0.916 (95% CI 0.881-0.943), respectively. nCD64 plus CRP performed best in prediction, discrimination, and reclassification of the 28-day mortality risk in sepsis. The risk of 28-day mortality increased stepwise as the number of data exceeding optimal cut-off values increased. CONCLUSIONS: nCD64 index combined with CRP was superior to CRP, PCT, nCD64 index and nCD64 index plus PCT in predicting 28-day mortality in sepsis. Multi-marker approach could improve the predictive accuracy and be beneficial for septic patients.


Assuntos
Pró-Calcitonina , Receptores de IgG/sangue , Sepse , Biomarcadores/metabolismo , Proteína C-Reativa/análise , Humanos , Unidades de Terapia Intensiva , Neutrófilos/metabolismo , Prognóstico , Curva ROC , Receptores de IgG/metabolismo , Estudos Retrospectivos , Sepse/diagnóstico , Sepse/metabolismo
7.
Sci Rep ; 12(1): 12752, 2022 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-35882892

RESUMO

Amyloid precursor protein (APP) is a transmembrane protein that plays critical role in the pathogenesis of Alzheimer's disease (AD). It is also involved in many types of cancers. Increasing evidence has shown that the tyrosine phosphorylation site Y682 in the intracellular tail of APP is crucial for APP function. Here, we report that Vav2, a guanine nucleotide exchange factor (GEF) for Rho family GTPase, is a novel interaction partner of APP. We found that Vav2-SH2 domain was able to bind directly to the Y682-phosphorylated intracellular tail of APP through isothermal titration calorimetry and NMR titrating experiments. The crystal structure of Vav2-SH2 in complex with an APP-derived phosphopeptide was determined to understand the structural basis of this recognition specificity. The interaction of APP and Vav2 in a full-length manner was further confirmed in cells by GST pull-down, co-immunoprecipitation and immunofluorescence staining experiments. In addition, we found overexpression of Vav2 could inhibit APP degradation and markedly increase the protein levels of APP and its cleavage productions in 20E2 cells, and this function of Vav2 required a functional SH2 domain.


Assuntos
Precursor de Proteína beta-Amiloide , Fatores de Troca do Nucleotídeo Guanina , Precursor de Proteína beta-Amiloide/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fosforilação , Proteínas rho de Ligação ao GTP/metabolismo , Domínios de Homologia de src
8.
J Mol Cell Biol ; 14(5)2022 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-35512546

RESUMO

In the fission yeast Schizosaccharomyces pombe, Mei2, an RNA-binding protein essential for entry into meiosis, regulates meiosis initiation. Mei2 binds to a specific non-coding RNA species, meiRNA, and accumulates at the sme2 gene locus, which encodes meiRNA. Previous research has shown that the Mei2 C-terminal RNA recognition motif (RRM3) physically interacts with the meiRNA 5' region in vitro and stimulates meiosis in vivo. However, the underlying mechanisms still remain elusive. We first employed an in vitro crosslinking and immunoprecipitation sequencing (CLIP-seq) assay and demonstrated a preference for U-rich motifs of meiRNA by Mei2 RRM3. We then solved the crystal structures of Mei2 RRM3 in the apo form and complex with an 8mer RNA fragment, derived from meiRNA, as detected by in vitro CLIP-seq. These results provide structural insights into the Mei2 RRM3-meiRNA complex and reveal that Mei2 RRM3 binds specifically to the UUC(U) sequence. Furthermore, a structure-based Mei2 mutation, Mei2F644A causes defective karyogamy, suggesting an essential role of the RNA-binding ability of Mei2 in regulating meiosis.


Assuntos
Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Meiose , RNA/genética , RNA/metabolismo , RNA não Traduzido/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/genética , Proteínas de Schizosaccharomyces pombe/metabolismo
9.
Nature ; 602(7897): 523-528, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35140398

RESUMO

A protein backbone structure is designable if a substantial number of amino acid sequences exist that autonomously fold into it1,2. It has been suggested that the designability of backbones is governed mainly by side chain-independent or side chain type-insensitive molecular interactions3-5, indicating an approach for designing new backbones (ready for amino acid selection) based on continuous sampling and optimization of the backbone-centred energy surface. However, a sufficiently comprehensive and precise energy function has yet to be established for this purpose. Here we show that this goal is met by a statistical model named SCUBA (for Side Chain-Unknown Backbone Arrangement) that uses neural network-form energy terms. These terms are learned with a two-step approach that comprises kernel density estimation followed by neural network training and can analytically represent multidimensional, high-order correlations in known protein structures. We report the crystal structures of nine de novo proteins whose backbones were designed to high precision using SCUBA, four of which have novel, non-natural overall architectures. By eschewing use of fragments from existing protein structures, SCUBA-driven structure design facilitates far-reaching exploration of the designable backbone space, thus extending the novelty and diversity of the proteins amenable to de novo design.


Assuntos
Redes Neurais de Computação , Proteínas , Sequência de Aminoácidos , Modelos Moleculares , Conformação Proteica , Proteínas/química
10.
Anticancer Drugs ; 33(1): e186-e197, 2022 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-34387608

RESUMO

Chemo-resistance is considered a major obstacle in the clinical treatment of non-small-cell lung cancer (NSCLC). Circular RNA (circRNA) circ-RNF121 (hsa_circ_0023404) has been identified to be related to the cisplatin (DDP) resistance. However, the role and mechanism of circ-RNF121 in the DDP resistance in NSCLC are still unknown. Real-time quantitative PCR (RT-qPCR) was applied to detect the levels of circ-RNF121, microRNA-646 (miR-646) and SRY-related HMG box transcription factor 4 (SOX4). Cell viability, proliferation, apoptosis, migration, invasion and cell cycle progression were assessed by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT), colony formation, flow cytometry, wound-healing, transwell and flow cytometry assays, severally. The binding relationship between miR-646 and circ-RNF121 or SOX4 was predicted by the circular RNA interactome or Target Scan Human7.2 and then verified by a dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. SOX4 protein level was measured by western blot assay. The biological role of circ-RNF121 on NSCLC tumor growth and drug resistance was examined by the xenograft tumor model in vivo. Circ-RNF121 and SOX4 were increased, and miR-646 was declined in DDP-resistant NSCLC tissues and cells. Furthermore, the circ-RNF121 deficiency could enhance DDP sensitivity by inhibiting cell proliferation, migration, invasion, cell cycle progression and promoting apoptosis in DDP-resistant NSCLC cells in vitro. Mechanically, circ-RNF121 served as a sponge of miR-646 to increase SOX4 expression. Circ-RNF121 knockdown improved the drug sensitivity of NSCLC in vivo. Circ-RNF121 silencing could reduce the DDP resistance of NSCLC cells by regulating SOX4 expression via miR-646. These findings hinted at a promising therapeutic target for NSCLC treatment.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Neoplasias Pulmonares/patologia , MicroRNAs/metabolismo , RNA Circular/metabolismo , Fatores de Transcrição SOXC/metabolismo , Animais , Apoptose/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Sobrevivência Celular/fisiologia , Cisplatino/farmacologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Front Oncol ; 11: 650919, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34150618

RESUMO

Prostate cancer (PCa) is a common aggressive disease worldwide which usually progresses into incurable castration-resistant prostate cancer (CRPC) in most cases after 18-24 months treatment. Androgen receptor (AR) has been considered as a crucial factor involved in CRPC and the study of AR as a potential therapeutic target in CRPC may be helpful in disease control and life-cycle management. In this study, we identified a potent small molecule compound, HG122, that suppressed CRPC cells proliferation and metastasis, and inhibited tumor growth both in subcutaneous and orthotopic tumor model. In addition, HG122 reduced the mRNA expression of PSA and TMPRSS2 which are target genes of AR, resulting in cell growth inhibition and metastasis suppression of CRPC, without affecting the expression of AR mRNA level. Mechanically, HG122 promoted AR protein degradation through the proteasome pathway impairing the AR signaling pathway. In conclusion, HG122 overcomes enzalutamide (ENZ) resistance in CRPC both in vitro and in vivo, thus suggesting HG122 is a potential candidate for the clinical prevention and treatment of CRPC.

12.
Biochem Biophys Res Commun ; 524(2): 490-496, 2020 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-32008746

RESUMO

3' uridylation is an essential modification associated with coding and noncoding RNA degradation in eukaryotes. In Arabidopsis, HESO1 was first identified as the major nucleotidyl transferase that uridylates most unmethylated miRNAs, and URT1 was later reported to play a redundant but important role in miRNA uridylation when HESO1 is absent. Two enzymes work sequentially and collaboratively to tail different forms of the same miRNAs in vivo. For mRNA, however, URT1 becomes the main enzyme to uridylate the majority of mRNA and repairs their deadenylated ends to restore the binding site for Poly(A) Binding Protein (PABP). HESO1, on the other hand, targets mostly the mRNAs with very short oligo(A) tails and fails in fulfilling the same task. To understand the structural basis these two functional homologues possess for their different substrate preferences and catalytic behaviors, we first determined the crystal structures of URT1 in the absence and presence of UTP. Our structures, together with functional assay and sequence analysis, indicated that URT1 has a conserved UTP-recognition mechanism analogue to the terminal uridylyl transferases from other species whereas HESO1 may evolve separately to recognize UTP in a different way. Moreover, URT1 N552 may be an important residue in interacting with 3' nucleotide of RNA substrate. The URT1 structure we determined represents the first structure of uridylyl transferase from plants, shedding light on the mechanisms of URT1/HESO1-dependent RNA metabolism.


Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/química , RNA Nucleotidiltransferases/química , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Domínio Catalítico , Cristalografia por Raios X , Modelos Moleculares , Conformação Proteica , RNA Nucleotidiltransferases/metabolismo , Uridina Trifosfato/metabolismo
13.
Structure ; 27(12): 1798-1810.e3, 2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31653339

RESUMO

In eukaryotes, nucleosome assembly is crucial for genome integrity. The histone chaperone NAP1 plays an important role in histone folding, storage, and transport, as well as histone exchange and nucleosome assembly. At present, the molecular basis of these activities is not fully understood. We have solved high-resolution crystal structures of Caenorhabditis elegans NAP1 (ceNAP1) in complex with its cognate substrates: the C. elegans H2A-H2B dimer (ceH2A-H2B) and the H2A.Z-H2B dimer (ceH2A.Z-H2B). Our structural and biochemical data reveals the acidic concave surface is relevant to tetramerization, and uncovers how a ceNAP1 homodimer uses its concave surface to asymmetrically recognize a ceH2A-H2B or ceH2A.Z-H2B heterodimer. Intriguingly, an "acidic strip" within the concave surface of ceNAP1 is crucial for binding histones, including H2A-H2B, H3-H4, and histone variants. Thus, our results provide insight into the molecular mechanisms of NAP1 histone chaperone activity.


Assuntos
Proteínas de Caenorhabditis elegans/química , Caenorhabditis elegans/genética , Histonas/química , Proteína 1 de Modelagem do Nucleossomo/química , Sequência de Aminoácidos , Animais , Sítios de Ligação , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Clonagem Molecular , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Histonas/genética , Histonas/metabolismo , Modelos Moleculares , Proteína 1 de Modelagem do Nucleossomo/genética , Proteína 1 de Modelagem do Nucleossomo/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Multimerização Proteica , Estrutura Terciária de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade por Substrato
14.
Nat Commun ; 10(1): 4273, 2019 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-31537782

RESUMO

Recognition of specific chromatin modifications by distinct structural domains within "reader" proteins plays a critical role in the maintenance of genomic stability. However, the specific mechanisms involved in this process remain unclear. Here we report that the PHD-Bromo tandem domain of tripartite motif-containing 66 (TRIM66) recognizes the unmodified H3R2-H3K4 and acetylated H3K56. The aberrant deletion of Trim66 results in severe DNA damage and genomic instability in embryonic stem cells (ESCs). Moreover, we find that the recognition of histone modification by TRIM66 is critical for DNA damage repair (DDR) in ESCs. TRIM66 recruits Sirt6 to deacetylate H3K56ac, negatively regulating the level of H3K56ac and facilitating the initiation of DDR. Importantly, Trim66-deficient blastocysts also exhibit higher levels of H3K56ac and DNA damage. Collectively, the present findings indicate the vital role of TRIM66 in DDR in ESCs, establishing the relationship between histone readers and maintenance of genomic stability.


Assuntos
Dano ao DNA/genética , Reparo do DNA/genética , Células-Tronco Embrionárias/citologia , Histonas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Acetilação , Animais , Linhagem Celular , Cromatina/metabolismo , Cristalografia por Raios X , Instabilidade Genômica/genética , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Knockout , Domínios Proteicos/fisiologia , Sirtuínas/metabolismo
15.
J Cell Biochem ; 120(10): 17573-17583, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31111550

RESUMO

Non-small cell lung cancer (NSCLC) is still an unresolved source of tumor-related death internationally. Current studies have discovered that microRNAs (miRNAs) are associated with diverse cancers development, including NSCLC. Our paper focused on the functional character of miR-4286 in NSCLC. miR-4286 level in 68 cases of NSCLC tissues, matched neighboring nontumor tissues and different cancer cell lines were inspected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). The connection concerning miR-4286 expression and clinicopathological features of patients with NSCLC were further determined. After knockdown or overexpression of miR-4286, cell viability, cell cycle, and/or apoptotic cells were examined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assay, respectively. Moreover, the cell cycle- and apoptosis-related proteins were estimated by qRT-PCR and Western blot. In comparison with the matched nontumor tissues, miR-4286 was significantly enhanced in lung malignancy tissues and different cell lines. miR-4286 expression was related with the tumor-node-metastasis stage, lymphatic metastasis, and distant metastasis. Cell viability was ominously weakened by suppression of miR-4286 in A549 cells, whereas was statistically upregulated by overexpression of miR-4286 in NCI-H1299 cells. Additionally, we detected that suppression of miR-4286 tempted cell cycle arrest in G1 stage and fortified apoptosis in A549 cells. Runx3 was recognized as one target gene of miR-4286, and the impacts of suppression of miR-4286 on cell viability and apoptosis were through regulation of Runt-related transcription factor 3. Our study suggests that miR-4286 overexpression represents a tumor promoter role in NSCLC cells.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Subunidade alfa 3 de Fator de Ligação ao Core/genética , MicroRNAs/genética , Prognóstico , Células A549 , Adulto , Idoso , Apoptose/genética , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/epidemiologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia
16.
Sci China Life Sci ; 60(4): 397-403, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28321657

RESUMO

Hemostasis is a major challenge in surgical procedures and traumas. Conventional hemostatic methods have limited efficacy and may cause additional tissue damage. In this study, we designed a novel hemostatic agent based on the in situ gel formation of gelatin cross-linked by a novel microbial transglutaminase (mTGase), in which the amino acid sequences differed from commercial mTGases. The new hemostatic agent showed the same biochemical crosslinking chemistry as the final stages of the blood coagulation cascade while using gelatin as a "structural" protein (rather than fibrin) and a calcium-independent mTGase as the crosslinking catalyst (rather than factor XIIIa). In rat liver hemostasis models, the hemostatic agent not only showed a similar hemostatic effect as that of SURGIFLO® (positive control), but also stronger adhesion strength and elasticity than SURGIFLO®. Therefore, this biomimetic gelatin-mTGase mix hemostatic is a novel and effective surgical sealant.


Assuntos
Gelatina/química , Hemostáticos , Streptomycetaceae/enzimologia , Transglutaminases/química , Sequência de Aminoácidos , Animais , Masculino , Ratos , Ratos Sprague-Dawley , Homologia de Sequência de Aminoácidos
17.
Sci China Life Sci ; 60(2): 202-214, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28194552

RESUMO

Tumor angiogenesis is characterized by abnormal vessel morphology, endowing tumor with highly hypoxia and unresponsive toward treatment. To date, mounting angiogenic factors have been discovered as therapeutic targets in antiangiogenic drug development. Among them, vascular endothelial growth factor receptor 2 (VEGFR2) inhibitors exerts potent antiangiogenic activity in tumor therapy. Therefore, it may provide a valid strategy for cancer treatment through targeting the tumor angiogenesis via VEGFR2 pathway. In this study, we established a high-profile compounds library and certificated a novel compound named N-(N-pyrrolidylacetyl)-9-(4-bromobenzyl)-1,3,4,9-tetrahydro-ß-carboline (YF-452), which remarkably inhibited the migration, invasion and tube-like structure formation of human umbilical vein endothelial cells (HUVECs) with little toxicity invitro. Rat thoracic aorta ring assay indicated that YF-452 significantly blocked the formation of microvascular exvivo. In addition, YF-452 inhibited angiogenesis in chick chorioallantoic membrane (CAM) and mouse corneal micropocket assays. Moreover, YF-452 remarkably suppressed tumor growth in xenografts mice model. Furthermore, investigation of molecular mechanism revealed that YF-452 inhibited VEGF-induced phosphorylation of VEGFR2 kinase and the downstream protein kinases including extracellular signal regulated kinase (ERK), focal adhesion kinase (FAK) and Src. These results indicate that YF-452 inhibits angiogenesis and may be a potential antiangiogenic drug candidate for cancer therapy.


Assuntos
Inibidores da Angiogênese/farmacologia , Antineoplásicos/farmacologia , Carbolinas/química , Neovascularização Patológica/prevenção & controle , Neoplasias da Próstata/tratamento farmacológico , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Animais , Aorta/efeitos dos fármacos , Células COS , Linhagem Celular Tumoral , Embrião de Galinha , Chlorocebus aethiops , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Técnicas In Vitro , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fosforilação , Ratos Sprague-Dawley , Transdução de Sinais , Ensaios Antitumorais Modelo de Xenoenxerto
18.
Saudi J Biol Sci ; 24(8): 1939-1942, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29551948

RESUMO

Objective: To investigate the clinical effect of fast track surgery (FTS) in perioperative nursing of colorectal cancer surgery. Background: In recent years, many complicated surgery began to develop in the direction of low invasion and short hospital time, which provides an unprecedented opportunity for the development of fast track surgery (FTS). Methods: According to different nursing measures, 156 cases of colorectal cancer patients treated in our hospital were divided into FTS nursing group (86 cases) and traditional nursing group (70 cases). FTS nursing care and traditional nursing care were respectively employed to analyze and compare postoperative recovery and complications of the two groups. Results: FTS nursing group was significantly shorter than the traditional care group in terms of the first postoperative exhaust time, the first defecation time, the first eating time, ambulation time and postoperative hospital time, with statistical significance (P < .05); compared with the conventional nursing group, FTS group significantly had lower incidence of postoperative intestinal obstruction, lower limb vein thrombus formation and gastrointestinal discomfort, with statistical significance (P < .05); FTS group has less situations of nausea and vomiting, incision infection, pulmonary infection, urinary tract infection and anastomotic leakage compared to the conventional nursing group. Conclusion: FTS nursing can effectively promote the postoperative recovery of intestinal function for patients with colorectal cancer and reduce the occurrence of postoperative complications, which will relieve postoperative pain and shorten the length of stay, giving patients increased rehabilitation quality.

19.
Biomed Pharmacother ; 84: 1274-1281, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27810784

RESUMO

The microRNAs represent a class of noncoding RNAs with short length and play diverse roles in many biological processes. Despite tremendous effects have been devoted, the role of miR-635 in non-small cell lung cancer (NSCLC) remains elusive. Here we report a potential tumor suppressive function of miR-635 in NSCLC. By microRNA screening based on invasion assays, we identified series of functional miRNAs. The miR-635 was then identified as a potent inhibitor of cell invasion and differentially expressed in normal and cancerous tissues. We further confirmed that Ying Yang 1 (YY1) may be the direct target of miR-635 as miR-635 transfection can significantly decrease endogenous YY1 expression which can mimic the effect of siRNA-mediated YY1 knockdown. Meanwhile, both miR-635 transfection and YY1 silencing can attenuate NSCLC cell invasion. In addition, miR-635 transfection can also significantly inhibit the growth of xenograft tumors. Taken together, our results have suggested a novel tumor suppressive role for miR-635 in NSCLC.


Assuntos
Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , MicroRNAs/genética , Idoso , Animais , Sequência de Bases , Carcinogênese/patologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Progressão da Doença , Feminino , Humanos , Neoplasias Pulmonares/patologia , Masculino , Camundongos , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Invasividade Neoplásica , Fator de Transcrição YY1/genética
20.
Biomed Rep ; 4(6): 687-690, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27284407

RESUMO

Escherichia coli (E. coli) is a common opportunistic pathogen for nosocomial infection. The aim of the study was to examine the phenotype, genotype and epidemiology of plasmid-mediated AmpC ß-lactamases in E. coli. In total, 96 clinical isolates of repeated E. coli were collected from different hospitals between August and October 2012. Using a cefoxitin disk diffusion method to identify the phenotype of AmpC ß-lactamases in E. coli, the plasmid was extracted, and multiplex polymerase chain reaction (PCR) was used to determine the amp gene. The PCR products were purified and sequenced. Of the 96 isolates strains, 43 strains were cefoxitin-resistant. Twelve (12.5%) isolates were detected to produce AmpC ß-lactamases with multiplex PCR, 11 strains carried DHA type ampC-resistant genes, and one strain carried ACC type ampC-resistant genes. In conclusion, the incidence of producing a plasmid-mediated AmpC enzyme of E. coli strains was relatively high. Therefore, antibiotics such as imipenem, a carbapenem, potentially serve as the treatment of choice for the infection.

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