RESUMO
Diabetic cognitive impairment is a common complication in type 2 diabetes. Berberine (BBR) is an isoquinoline alkaloid that has been shown to have neuroprotective effects against diabetes. This study aimed to investigate the effect of BBR on the gray and white matter of the brain by using magnetic resonance imaging (MRI) and to explore the underlying mechanisms. The study used diabetic db/db mice and administered BBR (50 and 100 mg/kg) intragastrically for twelve weeks. Morris water maze was applied to examine cognitive function. T2-weighted imaging (T2WI) was performed to assess brain atrophy, and diffusion tensor imaging (DTI) combined with fiber tracking was conducted to monitor the structural integrity of the white matter, followed by histological immunostaining. Furthermore, the protein expressions of the phosphatidylinositol 3-kinase (PI3K)/ protein kinase B (AKT)/ glycogen synthase kinase-3ß (GSK-3ß) were detected. The results revealed that BBR significantly improved the spatial learning and memory of the db/db mice. T2WI exhibited ameliorated brain atrophy in the BBR-treated db/db mice, as evidenced by reduced ventricular volume accompanied by increased hippocampal volumes. DTI combined with fiber tracking revealed that BBR increased FA, fiber density and length in the corpus callosum/external capsule of the db/db mice. These imaging findings were confirmed by histological immunostaining. Notably, BBR significantly enhanced the protein levels of phosphorylated AKT at Ser473 and GSK-3ß at Ser9. Collectively, this study demonstrated that BBR significantly improved the cognitive function of the diabetic db/db mice through ameliorating brain atrophy and promoting white matter reorganization via AKT/GSK-3ß pathway.
Assuntos
Atrofia , Berberina , Encéfalo , Disfunção Cognitiva , Imageamento por Ressonância Magnética , Substância Branca , Animais , Berberina/farmacologia , Berberina/uso terapêutico , Disfunção Cognitiva/tratamento farmacológico , Disfunção Cognitiva/diagnóstico por imagem , Atrofia/tratamento farmacológico , Camundongos , Masculino , Substância Branca/efeitos dos fármacos , Substância Branca/diagnóstico por imagem , Substância Branca/patologia , Substância Branca/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Encéfalo/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Imagem de Tensor de Difusão , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Glicogênio Sintase Quinase 3 beta/metabolismoRESUMO
The effect of intestinal flora changes on the pharmacokinetics of astragaloside â £ in rats with type 2 diabetes mellitus was explored in this study. The rat model in preliminary experiment was established by high-sugar and high-fat diet combined with the intraperitoneal injection of low-dose streptozotocin(STZ). Rats were divided into model group, astragaloside â £ group, berberine group and combination group(five rats in each group). After two weeks of gavage, the rats' feces was taken for 16 S rRNA sequencing of intestinal flora. Pharmacokinetic experiments were performed on astragaloside â £ in the four groups one day after the preliminary experiment. Plasma samples were precipitated in methanol with ginsenoside Rb_1 as an internal standard, and the plasma concentrations of astragaloside â £ at different time points were determined by UPLC-MS/MS. The chromatographic separation was performed on a Waters Acquity UPLC BEH-C_(18) column(2.1 mm×100 mm, 1.7 µm) via gradient elution. The mobile phase was acetonitrile(A) and 5 mmol·L~(-1) ammonium formate solution with 0.2% formic acid(B). The flow rate was 0.4 mL·min~(-1), the injection volume 5 µL and the column temperature 40 â. The mass spectrometry was carried out with electrospray ionization source(ESI) in multiple reaction monitoring and positive ion modes. The specificity, linearity range, accuracy, precision, stability and dilution effect of the method all met the requirements for the determination of astragaloside â £ in plasma. Plasma concentration-time curves were plotted and relevant pharmacokinetic parameters were calculated by DAS 3.2.8. The results showed that the concentration of absorbed astragaloside â £ increased within 0-3.95 h and began to decline since 3.95 h. After 36 h, the metabolism was complete. The area under the plasma concentration-time curve(AUC_(0-t)) and the peak concentration(C_(max)) of astragaloside â £ were increased in the three administration groups compared with the model group, but without significant difference, which suggested that the pharmacokinetic characteristics of saponin components would not necessarily change after the drug-induced alteration of intestinal flora.
Assuntos
Diabetes Mellitus Tipo 2 , Microbioma Gastrointestinal , Saponinas , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Ratos , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem , TriterpenosRESUMO
Owing to the potential health benefits, anthocyanin-rich teas (Camellia sinensis) have attracted interest over the past decade. Previously, we developed the cultivar 'Ziyan,' which has dark-purple leaves because of the accumulation of a high amount of anthocyanins. In this study, we performed a genetic analysis of this anthocyanin-rich tea cultivar and 176 of its naturally pollinated offspring. For two consecutive years, we quantified the anthocyanins and catechins of 'Ziyan' and the offspring population. While >60% of the offspring accumulated less than half of the amount of anthocyanins of 'Ziyan,' 17 (2018) and 15 (2019) individuals exceeded 'Ziyan' in anthocyanin content. A negative correlation between anthocyanin and total catechin content (r = -0.59, P < 0.001) was observed. The population was genotyped with 131 SSR markers spanning all linkage groups of the C. sinensis genome. Kruskal-Wallis tests identified 10 markers significantly associated with anthocyanins, catechins and their ratios in both years. Quantitative trait locus (QTL) analyses using the interval mapping method detected 13 QTLs, suggesting the dark-purple trait of 'Ziyan' is because of the pyramiding of anthocyanin-promoting alleles on at least five linkage groups. Two genetic loci reversely related to anthocyanin and total catechin contents were identified. This study provides valuable information for genetic improvement of purple tea cultivars and for fine-mapping related genes.
Assuntos
Camellia sinensis/genética , Catequina , Antocianinas , Folhas de Planta/genética , Locos de Características Quantitativas/genéticaRESUMO
To establish HPLC-MS/MS method for simultaneous determination of daphnetin, daphnoretin, and daphneticin in rat plasma after oral and intravenous administration of Daphne giraldii extract, and then use them in the calculation of pharmacokinetic parameters. Six sprague-dawley rats received intragastric administration of D. giraldii extract (daphnetin, daphnoretin and daphneticin were 88.40, 3.24 and 4.28 mgâ¢kg⻹, respectively). Their drug plasma concentration was determined by LC-MS/MS with schisandrin as an internal standard to draw plasma concentration-time curve. The pharmacokinetic parameters were calculated by Kinetica 4.4. The results showed that the linear range was 5-1 000 µgâ¢L⻹ for daphnetin, daphnoretin and daphneticin, and the method ological test showed conformance to the requirements.The intraday and inter-day variable coefficients (RSD) were both less than 15.0%, indicating that both of legitimate precise and accuracy were consistent with the analysis requirements of biological samples. For daphnetin, the pharmacokinetic parameters Tmax, Cmax, AUC0-t, T1/2 and MRT were 4 h, 858.96 µgâ¢L⻹, 10 566.4 µgâ¢L⻹â¢h, 5.19 h and 9.43 h, respectively. For daphnoretin, the pharmacokinetic parameters Tmax, Cmax, AUC0-t, T1/2 and MRT were 2.92 h, 178.00 µgâ¢L⻹, 905.89 µgâ¢L⻹â¢h, 3.50 h and 6.95 h, respectively. For daphneticin, the pharmacokinetic parameters Tmax, Cmax, AUC0-t, T1/2 and MRT were 2 h, 36.67 µgâ¢L⻹, 355.11 µgâ¢L⻹â¢h, 4.95 h and 8.27 h, respectively. The LC-MS/MS analysis method established in this study was proved to be so accurate and sensitive that it can be applied to the pharmacokinetic study of daphnetin, daphnoretin and daphneticin.
Assuntos
Cumarínicos/sangue , Daphne/química , Extratos Vegetais/farmacocinética , Umbeliferonas/sangue , Animais , Cromatografia Líquida , Cumarínicos/farmacocinética , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas em Tandem , Umbeliferonas/farmacocinéticaRESUMO
Ischemic brain injury is a major disease which threatens human health and safety. (3, 5, 6-trimethylpyrazin-2-yl) methyl 3-methoxy-4-[(3, 5, 6-trimethylpyrazin-2-yl) methoxy] benzoate (VA-T), a newly discovered lead compound, is effective for the treatment of ischemic brain injury and its sequelae. But the poor solubility of VA-T leads to poor dissolution and limited clinical application. In order to improve the dissolution of VA-T, the pharmaceutical technology of solid dispersions was used in the present study. VA-T/polyvinylpyrrolidone (PVP) solid dispersion was prepared by the solvent method. The dissolution studies were carried out and solid state characterization was evaluated by differential scanning calorimetry (DSC), infrared spectroscopy (IR), x-ray diffraction (XRD) and scanning electron microscopy (SEM). The dissolution rate of VA-T was significantly improved by solid dispersion compared to that of the pure drug and physical mixture. The results of DSC and XRD indicated that the VA-T solid dispersion was amorphous. The IR spectra showed the possible interaction between VA-T and PVP was the formulation of hydrogen bonding. The SEM analysis demonstrated that there was no VA-T crystal observed in the solid dispersions. The ideal drug-to-PVP ratio was 1:5. In conclusion, the solid dispersion technique can be successfully used for the improvement of the dissolution profile of VA-T.
Assuntos
Benzoatos/química , Sistemas de Liberação de Medicamentos , Povidona/química , Benzoatos/administração & dosagem , Isquemia Encefálica/tratamento farmacológico , Química Farmacêutica/métodos , SolubilidadeRESUMO
AIM: To explore the association of NFKB1 c.-798_-795delATTG (rs28362491), NFKBIA c.-949C>T (rs2233406), IL-8 c.-352A>T (rs4073), IL-10 c.-854T>C (rs1800871), TNF c.-418G>A (rs361525), and TNF c.-488G>A (rs1800629) polymorphisms with breast cancer risk in an East Chinese population. METHODS: We conducted a case-control study including 975 study participants (474 breast cancer patients and 501 female controls without cancer) and genotyped the polymorphisms employing polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Logistic regression was used to assess the association of the polymorphisms with breast cancer risk. RESULTS: We found that the ins/del and del/del genotypes of NFKB1 polymorphism and TT genotype of IL-10 polymorphism significantly increased breast cancer risk (NFKB1 ins/del odds ratio [OR] 1.69, 95% [CI] 1.23-2.33, P=0.001; NFKB1 del/del OR 2.42, 95% CI 1.72-3.42, P<0.001; IL-10 TT OR 2.36, 95% CI 1.58-3.52, P<0.001). On the other hand, the TT genotype of IL-8 polymorphism, GA and AA genotypes of TNF c.-418G>A polymorphism, and GA genotype of TNF c.-488G>A polymorphism significantly reduced breast cancer risk (IL-8 TT OR 0.48, 95% CI 0.33-0.72, P<0.001; TNF c.-418 GA OR 0.58, 95% CI 0.41-0.80, P=0.001; TNF c.-418 AA OR 0.38, 95% CI 0.14-0.98, P=0.044; TNF c.-488 GA OR 0.68, 95% CI 0.48-0.96, P=0.029). When stratified by menopausal status, the CT genotype of NFKBIA polymorphism significantly reduced the risk among pre-menopausal women (OR 0.63, 95% CI 0.40-0.99, P=,043), but not among post-menopausal women. CONCLUSIONS: NFKB1, NFKBIA, IL-8, IL-10, and TNF polymorphisms could serve as useful predictive biomarkers for breast cancer risk among women in East China.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Predisposição Genética para Doença , Proteínas de Neoplasias/genética , Polimorfismo Genético , Estudos de Casos e Controles , Feminino , Genótipo , Humanos , Proteínas I-kappa B/genética , Interleucina-10/genética , Interleucina-8/genética , Pessoa de Meia-Idade , Inibidor de NF-kappaB alfa , Subunidade p50 de NF-kappa B/genética , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Fator de Necrose Tumoral alfaRESUMO
AIM: In order to obtain single domain antibody against surface protective antigen A (SpaA)of Erysipelothrix rhusiopathiae. METHODS: The SpaA-N recombinant protein was used to screen binders from Bactrian camel VHH phage display library. After sequencing, the interested VHH gene fragments were subcloned into pET-30a vector to overexpress the protein in E.coli BL21. The binding specificity of the recombinant VHH with SpaA-N was determined by Western blotting. The thermal stability of single-domain antibody was evaluated by ELISA. RESULTS: By enrichment of screening, 2 clones were selected. Recombinant single domain antibodies purified by Ni-ion affinity chromatography showed a single band at M(r); 29 000, 23 000 on SDS-PAGE. ELISA results showed that VHH can bind its antigen specifically. After thermal denaturation, VHH can restore the antigen binding ability after refolding. Western blotting results showed that the recombinant VHH specific bind surface protective antigen of Erysipelothrix rhusiopathiae at M(r); 66 000. Two VHH single domain antibodies with high thermal stability and good antigen binding specificity were identified by screening Bactrian camel VHH phage display library. CONCLUSION: Two single domain antibodies that specifically aggulated SpaA-N is obtained, which provide the basis for further study in the immune role of single domain antibody against Erysipelothrix rhusiopathiae infection.