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1.
J Org Chem ; 2021 Nov 16.
Artigo em Inglês | MEDLINE | ID: mdl-34783555

RESUMO

The spiro scaffold chiral organocatalyst of 3,2'-pyrrolidinyl spiro-oxindole amine was successfully prepared from racemic spiro-oxindole amine using l-menthol as a chiral pool in 4 steps in 28%-40% overall yields with at least 99% ee in scale-up preparation, and its catalytic activity was evaluated in the enantioselective aldol condensation between 3-(3-hydroxy-1H-pyrazol-1-yl)-oxindole and paraformaldehyde. The spiro organocatalyst showed superior catalytic activity and selectivity compared with its counterparts, and most substrates offered good to excellent results with up to 96% yield in 96% ee.

2.
Org Lett ; 23(6): 2227-2231, 2021 03 19.
Artigo em Inglês | MEDLINE | ID: mdl-33656901

RESUMO

A new and crucial synthon of 3-((diphenylmethylene)-amino)-oxindole was designed and synthesized, for which an organocatalytic and enantioselective Michael/cyclization reaction with a terminal vinyl ketone catalyzed by a cinchona base was disclosed. A wide variety (28 examples) of almost all new chiral spiro[oxindol-3,2'-pyrrols] were prepared in excellent yields (up to 99%) with excellent enantioselectivities (95-99% ee), of which a typical chiral spiro product was further reduced to chiral spiro[oxindole-3,2'-pyrrolidine].


Assuntos
Cinchona/química , Oxindóis/química , Bases de Schiff/química , Compostos de Espiro/química , Ciclização , Cetonas/química , Estrutura Molecular , Estereoisomerismo
3.
Chem Commun (Camb) ; 56(94): 14825-14828, 2020 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-33155600

RESUMO

A cinchona alkaloid squaramide promoted enantioselective [4+2] cyclization between hydroxymaleimides and ortho-hydroxyphenyl p-QMs has been disclosed, and a wide range of chiral hemiketals containing chromane and succinimide frameworks with two adjacent quaternary stereogenic centers have been prepared for the first time with excellent results (up to 99% yield, up to 99 : 1 dr, up to >99% ee) under mild conditions.

4.
Org Biomol Chem ; 18(46): 9511-9515, 2020 Dec 07.
Artigo em Inglês | MEDLINE | ID: mdl-33185640

RESUMO

A new base promoted Michael-Michael domino cycloaddition between isoindigos and α-alkylidene succinimides has been developed for highly efficient and one-step convenient preparation of highly steric bispiroxindoles with two adjacent quaternary carbon centers and four consecutive cycles in excellent yields (up to 96%) and diastereoselectivities (up to >20 : 1) under mild conditions within a few minutes. A series of bisprooxindoles were obtained and the synthetic potential of the protocol was evaluated in a scale-up preparation.

5.
J Org Chem ; 85(14): 9290-9300, 2020 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-32583669

RESUMO

A new enantioselective Michael addition between 3-(3-hydroxy-1H-pyrazol-1-yl)oxindole, a new synthon generated from isatin N,N'-cyclic azomethine imine 1,3-dipole, and ß-nitrostyrene has been disclosed. A series of chiral 3-(3-oxo-2,3-dihydro-1H-pyrazol-1-yl) disubstituted oxindoles were obtained in excellent results (up to 97% yield, up to 94% ee) with moderate to good diastereoselectivities (up to 4.3:1 dr).

6.
Materials (Basel) ; 11(2)2018 Feb 21.
Artigo em Inglês | MEDLINE | ID: mdl-29466288

RESUMO

The ultrasonic transmission spectrum in a double-layered bonded structure is related closely to its interfacial stiffness. Consequently, researching the regularity of the transmission spectrum is of significant interest in evaluating the integrity of the bonded structure. Based on the spring model and the potential function theory, a theoretical model is developed by the transfer matrix method to predict the transmission spectrum in a double-layered bonded structure. Some shift rules of the transmission peaks are obtained by numerical calculation of this model with different substrates. The results show that the resonant transmission peaks move towards a higher frequency with the increase of the normal interfacial stiffness, and each of them has different movement distances with the increasing interfacial stiffness. Indeed, it is also observed that the movement starting points of these peaks are at the specific frequency at which the thickness of either substrate plate equals an integral multiple of half a wavelength. The results from measuring the bonding specimens, which have different interfacial properties and different substrates in this experiment, are utilized to verify the theoretical analysis. Though the theory of "starting points" is not demonstrated effectively, the shift direction and distance exactly match with the result from the theoretical algorithm.

7.
J Huazhong Univ Sci Technolog Med Sci ; 37(6): 895-903, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29270750

RESUMO

Human Nestin (hNestin) has been found to express in melanoma, and its expression is positively correlated with the advanced stage of melanoma. However, the precise role of hNestin in the development of melanoma has not been fully understood. The present study aimed to explore the role of hNestin in the proliferation and invasion of melanoma cells. The lentivirus vector carrying a short hairpin RNAs (shRNAs) targeting hNestin (hNestin-shRNA-LV) was stably infected into human melanoma cells UACC903, which expressed high levels of hNestin. The effects of hNestin knockdown on the proliferation, apoptosis, migration of melanoma cells and the related signaling pathways were investigated by immunofluorence, Western blotting and reverse transcription polymerase chain reaction (RT-PCR), respectively. The results showed that hNestin was expressed in most melanoma specimens and the melanoma cells studied. Knockdown of hNestin expression significantly inhibited the proliferation of melanoma cells, blocked the formation of cell colony, arrested cell cycle at G1/S stage and suppressed the activation of Akt and GSK3ß. hNestin-silent cells also showed a sheet-like appearance with tight cell-cell adhesion, decreased membrane expression of N-cadherin and ß-catenin, and attenuated migration. Furthermore, hNestin silence resulted in the inhibition of tumor growth in vivo. Our study indicates that hNestin knockdown suppresses the proliferation of melanoma cells, which might be through affecting Akt-GSK3ß-Rb pathway-mediated G1/S arrest, and hNestin silence inhibits the migration by selectively modulating the expression of cell adhesion molecules in the process of epithelial-mesenchymal transition.


Assuntos
Regulação Neoplásica da Expressão Gênica , Glicogênio Sintase Quinase 3 beta/genética , Melanoma/genética , Nestina/genética , Proteínas Proto-Oncogênicas c-akt/genética , Proteína do Retinoblastoma/genética , Neoplasias Cutâneas/genética , Antígenos CD/genética , Antígenos CD/metabolismo , Apoptose , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal , Pontos de Checagem da Fase G1 do Ciclo Celular , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Lentivirus/genética , Lentivirus/metabolismo , Melanoma/metabolismo , Melanoma/patologia , Melanoma/terapia , Nestina/antagonistas & inibidores , Nestina/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Proteína do Retinoblastoma/metabolismo , Transdução de Sinais , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/terapia , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genética , beta Catenina/metabolismo
8.
Zhonghua Nan Ke Xue ; 21(6): 532-5, 2015 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-26242044

RESUMO

OBJECTIVE: To investigate the influence of the time interval from the end of semen processing to artificial intrauterine in semination with husband's sperm (AIH-IUI) on the rate of clinical pregnancy. METHODS: This study involved 191 AIH-IUI cycles with the same ovulation induction protocol. After Percoll density gradient centrifugation, we divided the sperm into four groups based on the incubation time: 0-19, 20-39, 40-59, and 60-80 min, and again into another four groups according to the total progressively motile sperm count (TPMC): (0-9), (10-20), (21-30), and > 30 x 10(6). We analyzed the correlation of the clinical pregnancy rate with the time interval from the end of sperm processing to AIH-IUI and with other influencing factors, such as maternal age, infertility duration, and semen quality. RESULTS: The rate of clinical pregnancy was significantly higher in the 20-39 min group (18.3%) than in the 0-19, 40-59, and 60-80 min groups (12.7, 11.4 and 9.1%) (all P < 0.05). The (10-20) x 10(6) group achieved a remarkably higher pregnancy rate (16.7%) than the (0-9), (21-30), and > 30 x 10(6) groups (0, 11.4, and 8.3%) (all P < 0.05). Logistic multivariate analysis showed that the rate of clinical pregnancy was decreased with the increased age of the women (OR 0.89, 95% CI 0.83-0.94) but significantly elevated in the 20-39 min group (OR 2.11, 95% CI 1.34-3.13) and of (10-20) x 10(6) group (OR 2.06, 95% CI 1.32-3.46). CONCLUSION: The time interval from the end of sperm processing to AIH-IUI is a most significant factor influencing the rate of clinical pregnancy of AIH-IUI.


Assuntos
Infertilidade/terapia , Inseminação Artificial Homóloga/estatística & dados numéricos , Taxa de Gravidez , Centrifugação com Gradiente de Concentração , Feminino , Humanos , Masculino , Gravidez , Sêmen , Análise do Sêmen , Contagem de Espermatozoides , Espermatozoides , Fatores de Tempo
9.
Biologia (Bratisl) ; 67(2): 405-410, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-32214412

RESUMO

Herpes simplex virus type 2 (HSV-2) is the major cause of genital herpes in humans. The glycoprotein D of HSV-2 (gD2) is a promising subunit vaccine candidate for the treatment of genital herpes. The aim of the present study was to express a biologically active recombinant gD2 in eukaryotic baculovirus system in quantities sufficient for further studies. Human cDNA encoding a gD2 protein with 393 amino acids was subcloned into the pFastBac HTb vector and the recombinant protein was expressed in Spodoptera frugiperda (Sf9) cells by high-density cell culture. In a stirred bioreactor, the key limiting factors including glucose concentration, glutamine concentration and dissolved oxygen (DO) were optimized for high-density cell growth. The Sf9 cell density could reach 9.6×106 cells/mL and the yield of recombinant gD2 protein was up to 192 mg/L in cell culture under the optimal conditions of 15 mM glucose, 0.4 g/L glutamine and 40% DO. Production of significant amounts of pure, full-length gD2 opened up the possibility to investigate novel functions of gD2. Moreover, the purified recombinant gD2 protein revealed a partial prophylactic immune function in genital herpes of guinea pigs infected with HSV-2.

11.
Zhonghua Yu Fang Yi Xue Za Zhi ; 43(3): 201-5, 2009 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-19534925

RESUMO

OBJECTIVE: To develop a rapid and simple multiplex polymerase chain reaction (PCR) method which discriminates extended-spectrum beta-lactamases (ESBLs) genes in sporadic Shigella isolates from 1998 to 2007 in Hangzhou city, China. METHODS: After ESBLs screening according to the Clinical and Laboratory Standards Institute (CLSI) method, CTX-M, TEM, SHV and OXA-1 encoding genes were detected by using a multiplex PCR method, and the results were verified by 8 single gene PCR amplification. RESULTS: Seventeen isolates harbored ESBLs genes among 195 Shigella isolates (8.72%). Genes encoding CTX-M (17 strains), TEM (2 strains), OXA-1 (10 strains) and SHV (0 strains) were discriminated with multiplex PCR analysis, which coincided with eight single gene PCR analysis at 94.12%. CONCLUSION: Multiplex PCR should be a suitable tool for initial rapid screening and discriminating ESBLs genes in Shigella isolates. With similar trend of national surveillance data, the proportion of sporadic Shigella isolates harbouring ESBLs genes might probably be on increase.


Assuntos
Reação em Cadeia da Polimerase/métodos , Shigella/genética , beta-Lactamases/genética , DNA Bacteriano/análise , Genes Bacterianos , Genótipo , Humanos , Testes de Sensibilidade Microbiana , Shigella/isolamento & purificação
12.
J Antimicrob Chemother ; 63(5): 917-20, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19297378

RESUMO

OBJECTIVES: The aim of this study was to characterize fluoroquinolone-resistant Shigella and determine whether the qnr and aac(6')-Ib-cr genes could contribute to sporadic shigellosis at the clinic in the Hangzhou area of China. METHODS: A total of 202 strains of Shigella (79 Shigella sonnei and 123 Shigella flexneri ) isolated from sporadic cases of shigellosis from 1998 to 2007 were analysed for their antimicrobial susceptibility. The gyrA, gyrB, parC, parE, qnr and aac(6')-Ib-cr genes and the profiles and incompatibility of plasmids were characterized. Chromosomal DNA fingerprinting was determined by XbaI-based digestion and PFGE. RESULTS: All strains of S. sonnei were susceptible to fluoroquinolones (ciprofloxacin and levofloxacin) while 15 out of 123 strains of S. flexneri were resistant. All of the 15 resistant strains displayed common mutations in the gyrA and parC genes and formed eight distinct groups with unique molecular characteristics. Notably, 10 isolates showed mutations at codon 87 of gyrA, and the other 5 were qnrS-positive. Two strains were positive for the aac(6')-Ib-cr gene. Importantly, this is the first report of qnrS- and aac(6')-Ib-cr-positive Shigella in China, the qnrS-positive S. flexneri serotypes 1a, 2a and 4c and the aac(6')-Ib-cr-positive S. flexneri serotypes 2a and 4c worldwide. CONCLUSIONS: The common mutations at position 83 of gyrA and position 80 of parC were crucial for resistance to nalidixic acid in S. flexneri. The mutation at position 87 of gyrA or the presence of the qnrS gene is necessary for high-level resistance to fluoroquinolones in Shigella isolates from China.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Disenteria Bacilar/microbiologia , Fluoroquinolonas/farmacologia , Shigella flexneri/efeitos dos fármacos , Shigella flexneri/isolamento & purificação , Proteínas de Bactérias/genética , Técnicas de Tipagem Bacteriana , China , Análise por Conglomerados , Impressões Digitais de DNA , DNA Girase/genética , DNA Topoisomerase IV/genética , Eletroforese em Gel de Campo Pulsado , Genótipo , Humanos , Testes de Sensibilidade Microbiana , Mutação de Sentido Incorreto , Plasmídeos , Shigella sonnei/efeitos dos fármacos , Shigella sonnei/isolamento & purificação
13.
J Clin Microbiol ; 46(3): 837-41, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18160451

RESUMO

The common respiratory viruses, including influenza A, influenza B, and newly emerging severe acute respiratory syndrome (SARS) viruses, may cause similar clinical symptoms. Therefore, differential diagnosis of these virus pathogens is frequently required for single clinical samples. In addition, there is an urgent need for noninfectious and stable RNA standards and controls for multivirus detection. In this study, reverse transcription-PCR (RT-PCR) targeting of the RNAs of influenza A and influenza B viruses and SARS coronavirus was performed, and the resulting products were spliced into a fragment which was packaged into armored RNA for use as a noninfectious, quantifiable synthetic substitute. Furthermore, in the present study we developed a multiplex real-time RT-PCR assay in which the armored RNA was used as an external positive control and the three RNA viruses could be detected simultaneously in a single reaction mix. The detection limit of the multiplex real-time PCR was 10 copies/microl of armored RNA.


Assuntos
Vírus da Influenza A/isolamento & purificação , Vírus da Influenza B/isolamento & purificação , RNA Viral , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Vírus da SARS/isolamento & purificação , Sequência de Bases , Escherichia coli/genética , Escherichia coli/metabolismo , Escherichia coli/virologia , Humanos , Vírus da Influenza A/genética , Vírus da Influenza B/genética , Influenza Humana/diagnóstico , Levivirus/genética , Levivirus/metabolismo , Dados de Sequência Molecular , RNA Viral/análise , RNA Viral/genética , Padrões de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas , Vírus da SARS/genética , Síndrome Respiratória Aguda Grave/diagnóstico , Montagem de Vírus
14.
Artigo em Chinês | MEDLINE | ID: mdl-18322598

RESUMO

OBJECTIVE: To prepare the armored RNA containing M gene of influenza H3N2. METHODS: The vector pAR-1 was constructed from expression vector pET30b in which the bacteriophage MS2 DNA fragment, containing the genes for maturase and coat protein and the pac site, was inserted. The M gene fragment of influenza A was inserted into the HindIII site downstream of the pac site on the pAR-1, which formed a new recombinant plasmid pAR-2. After the prokaryotic expression was carried out, armored RNA AR-2 containing M gene was obtained. AR-2 was purified, and then was quantified by real time RT-PCR. Moreover, the stability of AR-2 was checked. RESULTS: AR-2 was expressed successfully. AR-2 remained stable under various storage environments. Approximately 8.9 x 10(11) copies of AR-2 particles can be purified from one milliliter of culture. CONCLUSION: It showed that AR-2 was stable and RNase-resistant, which, as a virus surrogate, would be used as RT-PCR standards, controls and training or proficiency samples.


Assuntos
Vírus da Influenza A Subtipo H3N2/genética , RNA Viral/genética , Proteínas da Matriz Viral/genética , Plasmídeos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/normas
15.
Zhonghua Yu Fang Yi Xue Za Zhi ; 39(2): 129-32, 2005 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-15842838

RESUMO

OBJECTIVE: To detect the RNA of severe acute respiratory syndrome virus (SARS-CoV) by using reverse transcription polymerase chain reaction (RT-PCR) targeted for a two loci and a modified nested real-time RT-PCR as to improving the reliability and sensitivity of tests. METHODS: A nested RT-PCR was used for detecting one fragment of SARS-CoV RNA in oropharyngeal swabs from 3 SARS probable patients, 4 SARS suspect patients and other 27 patients with fever in Hangzhou, and the nested RT-PCR product from one SARS probable patient was sequenced. Meanwhile in these 3 SARS probable patients, other three RT-PCR methods, including a hemi-nested RT-PCR targeted for another fragment of SARS-CoV RNA, a real-time RT-PCR and a modified nested real-time RT-PCR, were employed to detect SARS-CoV RNA. RESULTS: Two positives were found in the 3 SARS probable patients, and none positive in 4 SARS suspect patients and other 27 patients with fever, using the nested RT-PCR. The sequence of the nested RT-PCR product from one SARS probable patient was identified with the counterpart of SARS-CoV genomes published in public database. The results of the hemi-nested RT-PCR, the real-time RT-PCR and the modified nested real-time RT-PCR in the 3 SARS patients were consistent with the one of the nested RT-PCR. During detecting specimen with low copies of RNA, a weak positive signal was produced after about 35 cycles in the real-time RT-PCR, but a strong positive signal was found only after 10 cycles in the modified nested real-time RT-PCR. CONCLUSION: It might improve the reliability of test by employing RT-PCR targeted for two or more fragments in SARS-CoV genome. The modified nested real-time RT-PCR might have higher sensitivity than the routine real-time RT-PCR.


Assuntos
RNA Viral/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Vírus da SARS/genética , Síndrome Respiratória Aguda Grave/diagnóstico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Bases , Humanos , Pessoa de Meia-Idade , RNA Viral/metabolismo , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Síndrome Respiratória Aguda Grave/virologia , Adulto Jovem
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