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1.
Zhonghua Nan Ke Xue ; 27(11): 995-1000, 2021 Nov.
Artigo em Zh | MEDLINE | ID: mdl-37422871

RESUMO

Objective: To evaluate the effects of different filling method-related sperm counting chambers and the structural factors of Leja counting chambers on sperm motility using computer-assisted sperm analysis (CASA). METHODS: Using drop-filled Makler, capillary-loaded Leja and structurally modified Leja sperm counting chambers, we measured sperm concentration, the percentages of progressively motile sperm (PMS) and non-progressively motile sperm (NPMS), total sperm motility, curvilinear velocity (VCL), average path velocity (VAP), straight line velocity (VSL), beat-cross frequency (BCF), linearity (LIN), wobble (WOB) and straightness (STR) in the semen samples of 76 males by CASA and compared them between different chambers. RESULTS: The drop-filled Makler sperm counting chamber achieved remarkably higher PMS, NPMS, total sperm motility, VCL and VAP than the Leja chambers (P < 0.05). There were no statistically significant differences in VSL, BCF, LIN, WOB and STR between the Makler and Leja chambers (P > 0.05), or in sperm concentration, PMS, NPMS and total sperm motility between the capillary-loaded and structurally modified Leja counting chambers (P > 0.05). The ground edge and thickness of the coverslip of the Leja counting chamber produced no significant inference on the kinetic sperm parameters (P > 0.05). CONCLUSIONS: The drop-filled sperm counting chamber achieves significantly higher sperm motility and kinetic parameters than the capillary-loaded Leja chamber. The structural factors such as the ground edge and thickness of the coverslip of the Leja counting chamber do not influence the analysis of sperm parameters.

2.
Pak J Pharm Sci ; 29(6): 1997-2004, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28375116

RESUMO

The objective of this work is to synthesize indolacin-5-fluorouracil-1-ylmethyl ester and the structure was confirmed by means of UV, IR, 1H-NMR, 13C-NMR and mass spectrometry. The physicochemical parameters of melting point, solubility, apparent partition coefficient were investigated. S180 sarcoma, H22 hapatitic cancer and Lewis-transplanted mice were used to evaluate the anti-tumor activity of indolacini-5-fluorouracil-1-ylmethyl ester compared with 5-fluorouracil in vivo. Anti-inflammatory and analgesic activities were evaluated in mice. The inhibitory ratio of indolacini- 5-fluorouracil-1-ylmethyl ester is comparative to that of 5-fluorouracil. This study indicates that 5-fluorouracil-1-ylmethyl ester may represent a new anticancer predrug of 5-fluorouracil to produce a combined effect of indolacin and 5-fluorouracil for cancer therapy.


Assuntos
Antimetabólitos Antineoplásicos/síntese química , Antimetabólitos Antineoplásicos/farmacologia , Carcinoma Pulmonar de Lewis/tratamento farmacológico , Fluoruracila/análogos & derivados , Fluoruracila/síntese química , Fluoruracila/farmacologia , Ácidos Indolacéticos/síntese química , Ácidos Indolacéticos/farmacologia , Sarcoma 180/tratamento farmacológico , Analgésicos/síntese química , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/farmacologia , Antimetabólitos Antineoplásicos/toxicidade , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Carcinoma Pulmonar de Lewis/patologia , Linhagem Celular Tumoral , Fluoruracila/toxicidade , Ácidos Indolacéticos/toxicidade , Dose Letal Mediana , Espectrometria de Massas , Camundongos , Estrutura Molecular , Espectroscopia de Prótons por Ressonância Magnética , Sarcoma 180/patologia , Solubilidade , Espectrofotometria Infravermelho , Espectrofotometria Ultravioleta , Temperatura de Transição , Carga Tumoral/efeitos dos fármacos
3.
J Pharmacol Sci ; 125(3): 320-8, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24989948

RESUMO

The present study aimed to investigate the effect of 4-methylcyclopentadecanone (4-MCPC) on local cerebral ischemia-reperfusion and the possible mechanisms involved. For this purpose, the focal cerebral ischemia rat model was induced by middle cerebral artery occlusion (MCAO) for 2 h, and the rats were treated with 4-MCPC (4 or 8 mg·kg(-1), p.o.) just 0.5 h before reperfusion. The neurological deficit scores and the ischemic infarct volume were recorded 24 h after the MCAO. In addition, the number of apoptotic cells was measured by TUNEL assay, and the expression of apoptosis-regulatory proteins and the PI3K/Akt neuroprotective signaling pathway were investigated by western blotting. Our results indicated that 4-MCPC (4 or 8 mg·kg(-1)) remarkably alleviated cerebral I/R injury by decreasing infarct volume and neurological deficit scores. 4-MCPC also decreased the number of apoptotic cells, regulated the expression of Bcl-2 and Bax, and increased the ratio of Bcl-2/Bax. Further study revealed that 4-MCPC treatment also increased the level of p-Akt and p-GSK-3ß. Wortmannin (PI3K inhibitor) markedly abolished the effects of 4-MCPC. Taken together, our results suggest that 4-MCPC protects against cerebral I/R injury through the inhibition of apoptosis, and this neuroprotective effect may be partly related to the activation of the PI3K/Akt signal pathway.


Assuntos
Lesões Encefálicas/prevenção & controle , Isquemia Encefálica/prevenção & controle , Cetonas/farmacologia , Cetonas/uso terapêutico , Compostos Macrocíclicos/farmacologia , Compostos Macrocíclicos/uso terapêutico , Fármacos Neuroprotetores , Traumatismo por Reperfusão/prevenção & controle , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Lesões Encefálicas/genética , Lesões Encefálicas/patologia , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Modelos Animais de Doenças , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta , Masculino , Fosfatidilinositol 3-Quinases/fisiologia , Proteínas Proto-Oncogênicas c-akt/fisiologia , Ratos Sprague-Dawley , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/patologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Proteína X Associada a bcl-2/metabolismo
4.
J Pharm Biomed Anal ; 241: 115961, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-38237546

RESUMO

Adiponectin (ADPN), which serum/plasma adiponectin levels are closely associated with insulin resistance and type 2 diabetes, and lower adiponectin levels predict an increased risk of diabetes, is a strong indicator of diabetes risk in people at high risk of diabetes in different races. Using the unique principle and performance advantages of chemiluminescence immunoassay (CLIA), an ADPN-CLIA method with high sensitivity, high specificity and wide detection range was established based on the principle of two-steps method of sandwich-type, with the magnetic particles (MPs) as the solid phase carrier and acridinium ester (AE) as the chemiluminescence reaction system. The selection of the main raw materials required, the preparation conditions of MPs-coated antibodies, the methods of AE-labeled antibodies, sample requirements and reaction modes were optimized and evaluated. AE labeling experiment was successfully performed with the labeling efficiency of 8.366 and the antibody utilization rate of 96.8%. The chemiluminescent immunoassay for ADPN had a good linear relationship from 0 ng/mL to 250 ng/mL (R2 =0.9993), with the detection limit of 0.05 ng/mL. The coefficient of variation (CV) of intra-assay and inter-assay precision were both less than 5% respectively. The recovery rates for accuracy were from 91.26% to 107.46%. The comparison experiment of 80 clinical serum samples between the developed ADPN-CLIA with the immunoturbidimetry showed that the correlation coefficient was 0.956, and the Bland-Altman analysis showed that the limits of agreement were - 0.364 and 0.433.


Assuntos
Adiponectina , Diabetes Mellitus Tipo 2 , Humanos , Luminescência , Imunoensaio/métodos , Magnetismo , Anticorpos , Medições Luminescentes/métodos
5.
J Agric Food Chem ; 72(15): 8618-8631, 2024 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-38569082

RESUMO

Daidzein (DAN) is an isoflavone, and it is often found in its natural form in soybean and food supplements. DAN has poor bioavailability owing to its extremely low water solubility and first-pass metabolism. Herein, we hypothesized that a bioactivatable natural amino acid-bearing carbamate prodrug strategy could increase the water solubility and metabolic stability of DAN. To test our hypothesis, nine amino acid prodrugs of DAN were designed and synthesized. Compared with DAN, the optimal prodrug (daidzein-4'-O-CO-N-isoleucine, D-4'-I) demonstrated enhanced water solubility and improved phase II metabolic stability and activation to DAN in plasma. In addition, unlike the passive transport of DAN, D-4'-I maintained high permeability via organic anion-transporting polypeptide 2B1 (OATP2B1)-mediated transport. Importantly, D-4'-I increased the oral bioavailability by 15.5-fold, reduced the gender difference, and extended the linear absorption capacity in the pharmacokinetics of DAN in rats. Furthermore, D-4'-I exhibited dose-dependent protection against liver injury. Thus, the natural amino acid-bearing carbamate prodrug strategy shows potential in increasing water solubility and improving phase II metabolic stability to enhance the oral bioavailability of DAN.


Assuntos
Isoflavonas , Pró-Fármacos , Animais , Ratos , Administração Oral , Aminoácidos/química , Disponibilidade Biológica , Carbamatos/química , Pró-Fármacos/química , Solubilidade , Água
6.
BMC Microbiol ; 13: 58, 2013 Mar 14.
Artigo em Inglês | MEDLINE | ID: mdl-23497180

RESUMO

BACKGROUND: The aminoglycoside-resistance genes encoding aminoglycoside modifying enzymes and 16S rRNA methyltransferases are main factors contributing to increasing resistance to aminoglycosides. Characterization and distribution of antimicrobial resistance gene profiles provide important information on the potential difficulty of treatment of bacteria. Several molecular methods have been developed to investigate the prevalence of aminoglycoside-resistance genes. These existing methods are time-consuming, labor-intensive, expensive or limited sensitivity in the epidemiological investigation. Therefore, it is necessary to develop a rapid, less-costly and high throughput and sensitive method to investigate the distribution of antimicrobial resistance gene in clinical isolates. RESULTS: In this study, we developed a GeXP analyzer-based multiplex PCR assay to simultaneously detect seven aminoglycoside-resistance genes, including aac(3)-II, aac(6')-Ib, aac(6')-II, ant(3″)-I,aph(3')-VI,armA and rmtB, and to analyze the distribution of these genes in clinical Enterobacteriaceae isolates. Under optimized conditions, this assay achieved a limit-of-detection as low as 10 copies of each of the seven genes. The presented method was applied to analyze the distribution of aminoglycoside-resistance genes in 56 clinical Enterobacteriaceae isolates, and the results were compared with that of the conventional single PCR assay. Kappa values of the two methods for detecting each of the seven resistance genes were 0.831, 0.846, 0.810, 0.909, 0.887, 0.810 and 0.825, respectively. CONCLUSION: This GeXP assay is demonstrated to be a rapid, cost-effective and high throughput method with high sensitivity and specificity for simultaneously detecting seven common aminoglycoside-resistance genes.


Assuntos
Aminoglicosídeos/farmacologia , Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Enterobacteriaceae/efeitos dos fármacos , Genes Bacterianos , Ensaios de Triagem em Larga Escala/métodos , Reação em Cadeia da Polimerase Multiplex/métodos , Enterobacteriaceae/genética , Humanos , Testes de Sensibilidade Microbiana/métodos , Sensibilidade e Especificidade
7.
Electrophoresis ; 34(20-21): 3003-7, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24038030

RESUMO

Three-dimensional (3D) paper-based microfluidics, which is featured with high performance and speedy determination, promise to carry out multistep sample pretreatment and orderly chemical reaction, which have been used for medical diagnosis, cell culture, environment determination, and so on with broad market prospect. However, there are some drawbacks in the existing fabrication methods for 3D paper-based microfluidics, such as, cumbersome and time-consuming device assembly; expensive and difficult process for manufacture; contamination caused by organic reagents from their fabrication process. Here, we present a simple printing-bookbinding method for mass fabricating 3D paper-based microfluidics. This approach involves two main steps: (i) wax-printing, (ii) bookbinding. We tested the delivery capability, diffusion rate, homogeneity and demonstrated the applicability of the device to chemical analysis by nitrite colorimetric assays. The described method is rapid (<30 s), cheap, easy to manipulate, and compatible with the flat stitching method that is common in a print house, making itself an ideal scheme for large-scale production of 3D paper-based microfluidics.


Assuntos
Técnicas Analíticas Microfluídicas/instrumentação , Nitritos/análise , Papel , Colorimetria , Desenho de Equipamento , Impressão/economia , Impressão/métodos
8.
Hypertens Res ; 46(8): 1949-1960, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37258626

RESUMO

We detect the antihypertensive effects of maximakinin (MK) on renal hypertensive rats (RHRs) and further research the influence of MK on vascular smooth muscle cells (VSMCs) to explore its hypotensive mechanism. The effects of MK on arterial blood pressure were observed in RHRs. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) assays were performed to detect the effect of MK on VSMC viability. Western blot and flow cytometry were used to investigate the influence of MK on intracellular Ca2+ levels and protein expression changes in VSMCs. In addition, specific protein inhibitors were applied to confirm the involvement of Ca2+-related signaling pathways induced by MK in VSMCs. MK showed a more significant antihypertensive effect than bradykinin in RHRs. MK significantly decreased intracellular Ca2+ concentrations. Furthermore, MK significantly induced the phosphorylation of signaling molecules, including extracellular signal-regulated kinase 1/2 (ERK1/2), P38, AMP-activated protein kinase (AMPK) and Akt in VSMCs. Moreover, only ERK1/2 inhibitor U0126 and AMPK inhibitor Compound C completely restored the decreased intracellular Ca2+ level induced by MK, and further research demonstrated that AMPK functioned upstream of ERK1/2 following exposure to MK. Finally, HOE-140, an inhibitor of the bradykinin B2 receptors (B2Rs), was applied to investigate the potential targets of MK in VSMCs. HOE-140 significantly blocked the AMPK/ERK1/2 pathway induced by MK, suggesting that the B2Rs might play an important role in MK-induced AMPK and ERK1/2 activation. MK significantly reduces blood pressure in RHRs. MK exerts its antihypertensive effect by activating the B2Rs and downstream AMPK/ERK1/2 pathways, leading to significantly reduced Ca2+ levels in VSMCs.


Assuntos
Proteínas Quinases Ativadas por AMP , Músculo Liso Vascular , Ratos , Animais , Músculo Liso Vascular/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Sistema de Sinalização das MAP Quinases , Anti-Hipertensivos/farmacologia , Bradicinina/farmacologia , Bradicinina/metabolismo , Células Cultivadas , Transdução de Sinais , Fosforilação , Miócitos de Músculo Liso/metabolismo
9.
Biosens Bioelectron ; 234: 115353, 2023 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-37120945

RESUMO

Lateral flow assays (LFAs) are promising points-of-care tests, playing a vital role in diseases screening, diagnosis and surveillance. However, development of portable, cheap, and smart LFAs platform for sensitive and accurate quantification of disease biomarkers in complex media is challenging. Here, a cheap handheld device was developed to realize on-site detection of disease biomarkers by Nd3+/Yb3+ co-doped near-infrared (NIR)-to-NIR downconversion nanoparticles (DCNPs) based LFA. Its sensitivity is at least 8-fold higher for detecting NIR light signal from Nd3+/Yb3+ co-doped nanoparticles than conventional expensive InGaAs camera based detection platform. Additionally, we enhance NIR quantum yield of Nd3+/Yb3+ co-doped nanoparticles up to 35.5% via simultaneous high dopant of sensitizer ions Nd3+ and emitter ions Yb3+. Combination of NIR-to-NIR handheld detection device and ultra-bright NIR emitting NaNbF4:Yb60%@NaLuF4 nanoparticle probe allows the detection sensitivity of SARS-CoV-2 ancestral strain and Omicron variants specific neutralizing antibodies LFA up to the level of commercial enzyme linked immunosorbent assay kit. Furthermore, by this robust method, enhanced neutralizing antibodies against SARS-CoV-2 ancestral strain and Omicron variants are observed in healthy participants with Ad5-nCoV booster on top of two doses of inactivated vaccine. This NIR-to-NIR handheld platform provides a promising strategy for on-site evaluating protective humoral immunity after SARS-CoV-2 vaccination or infection.


Assuntos
Técnicas Biossensoriais , COVID-19 , Humanos , COVID-19/diagnóstico , Vacinas contra COVID-19 , SARS-CoV-2 , Vacinação , Anticorpos Neutralizantes , Biomarcadores , Anticorpos Antivirais
10.
J Clin Microbiol ; 50(2): 288-93, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22116146

RESUMO

Hand, foot, and mouth disease (HFMD) is a contagious enteroviral disease occurring primarily in young children and caused by enterovirus 71 (EV71), coxsackievirus A16 (CVA16), and other serotypes of coxsackievirus and echovirus. In this study, a GeXP analyzer-based multiplex reverse transcription (RT)-PCR assay (GeXP assay) consisting of chimeric primer-based PCR amplification with fluorescent labeling and capillary electrophoresis separation was developed to simultaneously identify nine serotypes of enteroviruses associated with HFMD in China, including EV71, CVA16, CVA4, -5, -9, and -10, and CVB1, -3, and -5. The RNAs extracted from cell cultures of viral isolates and synthetic RNAs via in vitro transcription were used to analyze the specificity and sensitivity of the assay. The GeXP assay detected as little as 0.03 tissue culture infective dose (TCID(50)) of EV71 and CVA16, 10 copies of panenterovirus, EV71, CVA16, CVB1, and CVB5, and 100 copies of 10 (including panenterovirus) premixed RNA templates. A total of 180 stool specimens collected from HFMD patients and persons suspected of having HFMD were used to evaluate the clinical performance of this assay. In comparison with the results of conventional methods, the sensitivities of the GeXP assay for detection of panenterovirus, EV71, and CVA16 were 98.79% (163/165), 91.67% (44/48), and 91.67% (33/36), respectively, and the specificities were 80.00% (12/15), 98.48% (130/132), and 100% (144/144), respectively. The concordance of typing seven other serotypes of enteroviruses with the results of conventional methods was 92.59% (25/27). In conclusion, the GeXP assay is a rapid, cost-effective, and high-throughput method for typing nine serotypes of HFMD-associated enteroviruses.


Assuntos
Técnicas de Laboratório Clínico/métodos , Enterovirus/classificação , Enterovirus/genética , Doença de Mão, Pé e Boca/virologia , Reação em Cadeia da Polimerase Multiplex/métodos , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Virologia/métodos , China , Enterovirus/isolamento & purificação , Humanos , Técnicas de Diagnóstico Molecular/métodos , Sensibilidade e Especificidade
11.
Zhonghua Nan Ke Xue ; 18(9): 803-6, 2012 Sep.
Artigo em Zh | MEDLINE | ID: mdl-23193667

RESUMO

OBJECTIVE: To establish a new method for sperm sorting by imitating the physiological process of sperm-cervical mucus interaction on the microfluidic chip. METHODS: We designed a microfluidic chip to imitate the physiological process of natural sperm sorting in the microchannel based on the interaction between sperm and cervical mucus, and obtained motile sperm after the interaction. Meanwhile, we established an integrated real-time sperm detection reservoir on this chip to determine sperm parameters using the computer-assisted sperm analysis system. We analyzed 30 samples using both microfluidic and swim-up methods, and compared the results with those obtained before sorting. RESULTS: The rate of grade a + b sperm, the rate of morphologically normal sperm, straight-line velocity (VSL), average path velocity (VAP) and straightness (STR) were (29.78 +/- 11.24)%, (8.00 +/- 5.19)%, (18.89 +/- 4.90) microm/s, (26.84 +/- 5.13) microm/s and (70.15 +/- 7.61)%, respectively, before sorting, (71.65 +/- 11.18)%, (14.95 +/- 6.79)%, (24.14 +/- 5.95) microm/s, (32.61 +/- 6.36) microm/s and (73.87 +/- 9.34)%, respectively, after swim-up sorting, and (92.37 +/- 6.33)%, (23.33 +/- 7.67)%, (34.03 +/- 16.78) microm/s, (38.73 +/- 16.40) microm/s and (84.91 +/- 12.56)%, respectively, after sorting on the microfluidic chip. The sperm parameters obtained before sorting showed statistically significant differences from those obtained on the chip (P < 0.01) and by the swim-up method (P < 0.05). CONCLUSION: Imitation of the physiological interaction between sperm and cervical mucus on the microfluidic chip helped the realization of both the natural sorting and real-time analysis of sperm. The quality of the sperm sorted on the microfluidic chip is significantly better than that of the sperm before sorting and sorted by the swim-up method. This has prepared the ground for imitating the fertilization process under the physiological condition on the microfluidic chip.


Assuntos
Técnicas Analíticas Microfluídicas/instrumentação , Análise de Sequência com Séries de Oligonucleotídeos , Análise do Sêmen , Espermatozoides/fisiologia , Movimento Celular , Separação Celular , Muco do Colo Uterino , Humanos , Masculino , Microfluídica/métodos , Motilidade dos Espermatozoides/fisiologia
12.
Zhonghua Nan Ke Xue ; 17(4): 301-4, 2011 Apr.
Artigo em Zh | MEDLINE | ID: mdl-21548204

RESUMO

OBJECTIVE: To investigate the effects of a microfluidic sperm sorter on the routine parameters and DNA integrity of human sperm. METHODS: We divided 40 semen samples into two aliquots and performed sperm sorting using a self-made polydimethylsiloxane microfluidic sperm sorter and the swim-up method, respectively. Then we evaluated and compared the effects of these two methods on the sperm routine parameters and DNA integrity by computer-assisted sperm analysis and sperm chromatin dispersion test. RESULTS: After processing, sperm motility, normal morphology and tail hypoosmotic swelling rate were significantly improved, while sperm DNA damage remarkably decreased (P < 0.01). The microfluidic sperm sorter achieved a significantly lower rate of sperm DNA damage than the swim-up method ([ 8.4 +/- 5.8 ]% vs [16.4 +/- 9.2] %, P < 0.01), but no statistically significant differences were found in all other parameters between the two methods. CONCLUSION: High-quality sperm with less DNA integrity damage could be obtained in sperm sorting with the microfluidic sperm sorter.


Assuntos
Separação Celular/instrumentação , Dano ao DNA , Análise do Sêmen/instrumentação , Espermatozoides , Adulto , Separação Celular/métodos , DNA , Humanos , Infertilidade Masculina/genética , Infertilidade Masculina/fisiopatologia , Masculino , Microfluídica , Pessoa de Meia-Idade , Análise Serial de Proteínas , Análise do Sêmen/métodos , Motilidade dos Espermatozoides
13.
Assay Drug Dev Technol ; 19(3): 176-183, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33784479

RESUMO

Background:To investigate the inverse agonistic effect of a novel type 1 cannabinoid (CB1) receptor antagonist, MJ08, on the gastrointestinal tract (GIT).Methods: In vivo, carbon propulsion within the stomach of mice was undertaken to investigate the effects of MJ08. In vitro, the effects of MJ08 were investigated on the contraction of smooth muscle on the isolated gastric fundus, gastric body, duodenum, jejunum, and ileum.Results:Western blotting results showed that MJ08 (0.62 mg/kg body weight) reversed WIN55,212-2 (1.0 mg/kg)-induced reduction of carbon transit. MJ08 (1.25, 2.5 mg/kg) stimulated carbon transit dose dependently, demonstrating an inverse agonistic effect. In vitro experiments showed that the expression of MJ08 increased the contraction of small intestine, and that its inverse agonistic effect was significantly stronger than that of SR141716A, but no effect was noted on the gastric body. Western blotting showed that the MJ08 increased the expression of CB1 receptor in different GIT segments.Conclusion:MJ08 is not only an antagonist but also an inverse agonist of the CB1 receptor. MJ08 and SR141716A can enhance motility in the small intestine and increase the expression of CB1 receptor in the small intestine.


Assuntos
Antagonistas de Receptores de Canabinoides/farmacologia , Intestino Delgado/efeitos dos fármacos , Piperidinas/farmacologia , Pirazóis/farmacologia , Estômago/efeitos dos fármacos , Animais , Western Blotting , Feminino , Cobaias , Masculino , Camundongos , Camundongos Endogâmicos ICR , Ratos , Ratos Sprague-Dawley , Receptor CB1 de Canabinoide/efeitos dos fármacos , Receptor CB1 de Canabinoide/metabolismo
14.
Hypertens Res ; 44(7): 781-790, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-33707758

RESUMO

We investigated the antihypertensive effects of maximakinin (MK) on spontaneously hypertensive rats (SHRs). The effects of MK on arterial blood pressure in SHRs were observed, and flow cytometry and 4,5-diaminofluorescein-2 staining were used to examine MK-induced nitric oxide (NO) release in human umbilical vein endothelial cells (HUVECs). Western blotting was used to analyze the effects of MK on the expression of AMP-activated protein kinase (AMPK), Akt, Connexin 43, ERK1/2, p38, and p-eNOS in HUVECs. The results showed that MK induced a more significant antihypertensive effect on SHRs than bradykinin (BK). MK induced significant increases in endothelial nitric oxide synthase (eNOS) phosphorylation and NO release in HUVECs. MK also significantly increased the phosphorylation of Akt and AMPK in HUVECs. The AMPK inhibitor compound C blocked the effect of MK on the generation of NO. MK induced the phosphorylation of ERK1/2, p38, and Connexin 43. The expression of p-Connexin 43 was significantly decreased in the presence of the ERK1/2 inhibitor U0126 but not the p38 inhibitor SB203580. The effects of MK on the phosphorylation of AMPK and ERK1/2 were significantly decreased by the BK B2 receptor inhibitor HOE-140. In summary, MK can significantly reduce blood pressure in SHRs. The antihypertensive effect might be mediated through the activation of the BK B2 receptor, while the downstream AMPK/PI3K/Akt/eNOS/NO and ERK1/2/Connexin 43 signaling pathways play additional roles.


Assuntos
Anti-Hipertensivos , Hipertensão , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Anti-Hipertensivos/farmacologia , Conexina 43/farmacologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Hipertensão/tratamento farmacológico , Sistema de Sinalização das MAP Quinases , Óxido Nítrico/farmacologia , Óxido Nítrico Sintase Tipo III/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Endogâmicos SHR , Transdução de Sinais/efeitos dos fármacos
15.
Electrophoresis ; 30(24): 4300-5, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19924696

RESUMO

We developed an on-chip DNA analysis method, in which free-solution transient isotachophoresis (FstITP) were coupled with CGE. Using chloride ions in the sample matrix and HEPES in the background electrolyte, respectively, as the leading and terminating ions, dsDNAs were isotachophoretically preconcentrated in free-solution and then separated in sieving polymer. The coupling of FstITP and CGE enabled higher separation efficiency due to higher preconcentration rate in free-solution. The FstITP-CGE analysis offered adjustable signal intensities by varying sample injection time. With the maximum sample loading volume, the LOD of the FstITP-CGE analysis was determined to be 0.24 ng/mL by confocal laser-induced fluorescence detection. The FstITP-CGE method is simple, robust and flexible, thus well suited to the analysis of highly saline DNA samples at different concentration level.


Assuntos
DNA/isolamento & purificação , Dispositivos Lab-On-A-Chip , Limite de Detecção , Microscopia Confocal
16.
Arch Pharm Res ; 32(4): 527-33, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19407970

RESUMO

Myrica rubra Sieb. et Zucc. leaves are commonly used as an astringent, antidiarrheic, and analgesics in folk medicine in China. In the present study, the analgesic activity of myricetin, a major compound in Myrica rubra Sieb. et Zucc. leaves was evaluated in vivo. The analgesic effect of myricetin was tested by a serial of models, such as acetic acid-induced writhing response, formalin-induced paw licking and hot plate test. The sedative activity was evaluated by pentobarbital-induced sleep time. Platelet aggregation induced by collagen and arachidonic acid was also performed in vitro. Myricetin showed a significant inhibition on chemical nociceptive models such as the acetic acid-induced writhing response and the licking time on the late phase in the formalin test in a dose-dependent manner, but did not manifest a signicant effect in hot plate test. Myricetin was also not able to increase the sleeping time induced by pentobarbital, which further indicated that the analgesic effect of myricetin was unrelated to sedation. In addition, myricetin inhibited the content of PGE2 in the peritoneal fluid and platelet aggregation induced by collagen and arachidonic acid in vitro. These results collectively demonstrated that myricetin possessed potent analgesic activity, which was related with peripheral analgesia, but, not with the opioid system. Myricetin may be a potent COX-1 inhibitor with anti-platelet activity.


Assuntos
Analgésicos/farmacologia , Flavonoides/farmacologia , Myrica , Dor/prevenção & controle , Ácido Acético , Analgésicos/isolamento & purificação , Animais , Comportamento Animal/efeitos dos fármacos , Ciclo-Oxigenase 1/metabolismo , Inibidores de Ciclo-Oxigenase/farmacologia , Dinoprostona/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Flavonoides/isolamento & purificação , Formaldeído , Masculino , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/metabolismo , Camundongos , Myrica/química , Dor/induzido quimicamente , Dor/metabolismo , Medição da Dor , Limiar da Dor/efeitos dos fármacos , Cavidade Peritoneal , Lavagem Peritoneal , Folhas de Planta , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação Plaquetária/farmacologia , Coelhos , Sono/efeitos dos fármacos
17.
Free Radic Res ; 53(7): 714-726, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30947567

RESUMO

The antitumor effects of silibinin are of increasing interest, though its mechanism is not yet clear. The goal of this study was to clarify the mechanism of silibinin-induced cell death in the A431 human epidermoid carcinoma cell line. We used a cell viability assay, flow cytometry, nitric oxide (NO) assay, and western blotting to examine relationships between silibinin, NO generation and apoptosis in A431 cells. Silibinin inhibited A431 cell growth in a dose-dependent manner, inducing mitochondrial damage, and apoptosis at a high dose. At the same time, high dose silibinin increased NO levels in A431 cells and the endothelial nitric oxide synthase (eNOS) inhibitor NG-nitro-L-arginine methylester (L-NAME) attenuated silibinin-induced cell growth inhibition. By western blotting, silibinin caused increased eNOS phosphorylation in the mitochondria. The AMP-activated protein kinase inhibitor compound C significantly decreased p-eNOS expression, while blocking eNOS did not affect p-AMPK levels, suggested that AMPK acted upstream of eNOS. This study showed that silibinin increased NO levels in A431 cells by activating the AMPK-eNOS pathway, leading to mitochondrial dysfunction and apoptosis. In this mechanism of action, mitochondrial eNOS played an important role. The results provided new understanding of the functions of intracellular NO.


Assuntos
Epiderme/metabolismo , Mitocôndrias/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Silibina/uso terapêutico , Apoptose , Humanos , Silibina/farmacologia
18.
Electrophoresis ; 29(24): 4976-83, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19130577

RESUMO

We present a sensitive, simple and robust on-chip transient ITP (TITP) method for the analysis of polymerase chain reaction (PCR) samples. This TITP analysis was performed on a poly (methyl methacrylate) microchip with cross structure. The PCR sample was injected into a 4 mm free-solution buffer channel to increase sample loading. When applied separation voltage, the sample ions were stacked by TITP using chloride ions in the sample matrix as the leading ions and HEPES in the background electrolyte as the terminating ions. The stacked sample plug was then separated by zone electrophoresis (ZE) in hydroxypropylmethyl cellulose polymer. The whole operation was carried out in single background electrolyte. Sample injection, preconcentration / separation were performed continuously with the sequential switching of voltage. By injection of the sample into the free-solution buffer channel, sampling bias was avoided and the injection time was reduced. With this TITP method, the detection sensitivity was improved without loss of resolution, a 20-fold signal enhancement was achieved and the average limit of detection was estimated to be 0.24 pg/microL (S/N = 3). The TITP method is simple, fast and sensitive, thus is well suited to direct analysis of the highly saline PCR samples.


Assuntos
Eletroforese em Microchip/métodos , Reação em Cadeia da Polimerase/métodos , Soluções Tampão , DNA/análise , Desenho de Equipamento , Cinética , Sensibilidade e Especificidade
19.
Artigo em Inglês | MEDLINE | ID: mdl-16787766

RESUMO

The effects of carrageenans' structural features on its interaction with granulocyte colony-stimulating factor (G-CSF) and on the growth and differentiation of a G-CSF dependent leukemia cell line (NFS-60) were studied. lambda, iota, and kappa carrageenans, with decreasing contents of sulfation, bound to G-CSF with binding constants of (6.2+/-0.6) x 10(5)M(-1), (7.4+/-0.5) x 10(5)M(-1) and (6.0+/-0.4) x 10(5)M(-1), and with 27.7+/-0.2, 17.4+/-0.1 and 8.4+/-0.1 binding sites, respectively. However, kappa carrageenan oligosaccharide had no affinity for G-CSF. The three carrageenans significantly inhibited G-CSF-induced growth of NFS-60 cells. The high sulfate content lambda carrageenan could also induce the maturation of the cells, but relatively low sulfate content iota and kappa carrageenans could not. The results suggested that G-CSF-carrageenan bindings were dependent on carrageenans' sulfate contents and chain lengths, which could also affect the growth and differentiation of NFS-60 cells.


Assuntos
Carragenina/metabolismo , Fator Estimulador de Colônias de Granulócitos/metabolismo , Heparina/metabolismo , Configuração de Carboidratos , Sequência de Carboidratos , Carragenina/química , Ciclo Celular , Diferenciação Celular , Linhagem Celular Tumoral , Proliferação de Células , Eletroforese Capilar , Humanos
20.
Artigo em Inglês | MEDLINE | ID: mdl-16899416

RESUMO

The genotyping of hepatitis B virus (HBV) has become recently a valuable tool not only for epidemiological reasons but also for the clinical practice. Conventional methods for HBV genotyping typically include amplification of the target DNA sequences with a two-round nested PCR followed by separation of the amplified fragments by gel electrophoresis. A microfluidic chip that couples isotachophoresis (ITP) preconcentration and zone electrophoresis (ZE) separation may provide great advantages for sensitive, rapid and cost-effective clinical analysis. In this study, an HBV genotyping method with only one amplification round was developed by the application of the ITP-ZE chip. All the analysis steps of the ITP-ZE separation including sample injection, stacking and separation were performed continuously, controlled by sequential high-voltage switching. A 2.1cm sample plug was preconcentrated between discontinuous buffers in ITP process, followed by ZE separation. Sensitivity enhancement was obtained through the increase of sample loading volume. The average LOD value of the ITP-ZE microfluidic chip was determined to be 0.0021pg/muL. In a large-scale HBV genotyping test, single round PCR products were analyzed by ITP-ZE microfluidic chip, and the results were compared with that of the conventional method. Among the 200 cases studied, the classification rate obtained with microfluidic chip was 93%, which was 6% higher than that obtained with the conventional method. Method with ITP-ZE chip analysis provides HBV genotyping information in reduced PCR amplification time with higher detection rate when compared with conventional method. This method holds great potential for extrapolation to the abundance of similar molecular biology-based techniques in clinical diagnosis.


Assuntos
Eletroforese/instrumentação , Vírus da Hepatite B/genética , Microfluídica , Sequência de Bases , Primers do DNA , Eletroforese em Microchip/instrumentação , Genótipo , Humanos , Filogenia , Reação em Cadeia da Polimerase , Sensibilidade e Especificidade
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