RESUMEN
Spinocerebellar ataxia type 3 (SCA3) is a neurodegenerative disorder characterized by ataxia and other neurological manifestations, with a poor prognosis and a lack of effective therapies. The amyloid aggregation of the ataxin-3 protein is a hallmark of SCA3 and one of the main biochemical events prompting its onset, making it a prominent target for the development of preventive and therapeutic interventions. Here, we tested the efficacy of an aqueous Lavado cocoa extract and its polyphenolic components against ataxin-3 aggregation and neurotoxicity. The combination of biochemical assays and atomic force microscopy morphological analysis provided clear evidence of cocoa flavanols' ability to hinder ATX3 amyloid aggregation through direct physical interaction, as assessed by NMR spectroscopy. The chemical identity of the flavanols was investigated by ultraperformance liquid chromatography-high-resolution mass spectrometry. The use of the preclinical model Caenorhabditis elegans allowed us to demonstrate cocoa flavanols' ability to ameliorate ataxic phenotypes in vivo. To the best of our knowledge, Lavado cocoa is the first natural source whose extract is able to directly interfere with ATX3 aggregation, leading to the formation of off-pathway species.
Asunto(s)
Enfermedad de Machado-Joseph , Animales , Ataxina-3/genética , Ataxina-3/metabolismo , Enfermedad de Machado-Joseph/tratamiento farmacológico , Enfermedad de Machado-Joseph/genética , Enfermedad de Machado-Joseph/metabolismo , Proteínas Amiloidogénicas/metabolismo , Amiloide/metabolismo , Caenorhabditis elegans , Polifenoles/uso terapéutico , Extractos Vegetales/farmacologíaRESUMEN
The metabolic profile of Lavado cocoa was characterized for the first time by NMR spectroscopy, then compared with the profiles of fermented and processed varieties, Natural and commercial cocoa. The significant difference in the contents of theobromine and flavanols prompted us to examine the cocoa varieties to seek correlations between these metabolite concentrations and the anti-amyloidogenic activity reported for cocoa in the literature. We combined NMR spectroscopy, preparative reversed-phase (RP) chromatography, atomic force microscopy, in vitro biochemical and cell assays, to investigate and compare the anti-amyloidogenic properties of extracts and fractions enriched in different metabolite classes. Lavado variety was the most active and the catechins and theobromine were the chemical components of cocoa hindering Aß peptide on-pathway aggregation and toxicity in a human neuroblastoma SH-SY5Y cell line.
Asunto(s)
Cacao/química , Alimentos Fermentados/análisis , Péptidos beta-Amiloides/química , Péptidos beta-Amiloides/metabolismo , Antioxidantes/química , Cacao/metabolismo , Línea Celular Tumoral , Cromatografía Líquida de Alta Presión , Cromatografía de Fase Inversa , Flavanonas/análisis , Humanos , Espectroscopía de Resonancia Magnética , Fragmentos de Péptidos/química , Fragmentos de Péptidos/metabolismo , Extractos Vegetales/química , Extractos Vegetales/farmacología , Agregado de Proteínas/efectos de los fármacos , Teobromina/análisisRESUMEN
We set up an efficient protocol for the rapid analysis of NMR spectra of green and roasted coffee extracts, enabling the automatic identification and quantification of metabolites in approximately two minutes per spectrum. This method allowed for the metabolic profiling and the subsequent evaluation of the content of bioactive compounds and antioxidant activity of coffee samples, depending on their species (Arabica and Robusta), geographical origin and extraction procedure (hydroalcoholic, espresso and moka). The hydroalcoholic extraction is the most efficient method in terms of yields of low molecular weight compounds (in particular chlorogenic acids), while moka extraction provides the highest amounts of melanoidins. Moreover, that the ratio between health-giving compounds (chlorogenic acids, trigonelline and choline) and caffeine is higher in Arabica coffees. The data collected provide useful insights for the selection of coffee raw material to be used in the preparation of coffee-based dietary supplements, nutraceuticals and functional beverages.
Asunto(s)
Cafeína/análisis , Coffea/química , Alcaloides/análisis , Antioxidantes/química , Ácido Clorogénico/análisis , Coffea/metabolismo , Café/química , Humanos , Espectroscopía de Resonancia Magnética , Polímeros/análisis , Análisis de Componente Principal , Semillas/química , Semillas/metabolismo , SonicaciónRESUMEN
To identify food and beverages that provide the regular intake of natural compounds capable of interfering with toxic amyloidogenic aggregates, we developed an experimental protocol that combines NMR spectroscopy and atomic force microscopy, in vitro biochemical and cell assays to detect anti-Aß molecules in natural edible matrices. We applied this approach to investigate the potential anti-amyloidogenic properties of coffee and its molecular constituents. Our data showed that green and roasted coffee extracts and their main components, 5-O-caffeoylquinic acid and melanoidins, can hinder Aß on-pathway aggregation and toxicity in a human neuroblastoma SH-SY5Y cell line. Coffee extracts and melanoidins also counteract hydrogen peroxide- and rotenone-induced cytotoxicity and modulate some autophagic pathways in the same cell line.
Asunto(s)
Péptidos beta-Amiloides/química , Café/química , Manipulación de Alimentos , Extractos Vegetales/química , Extractos Vegetales/farmacología , Multimerización de Proteína/efectos de los fármacos , Línea Celular Tumoral , Color , Humanos , Espectroscopía de Resonancia MagnéticaRESUMEN
Recent epidemiological studies have demonstrated that the consumption of healthy foods that are particularly rich in polyphenols might reduce the incidence of cancer and neurodegenerative diseases. In particular, chlorogenic acids (CGAs) occur ubiquitously in food and represent the most abundant polyphenols in the human diet. A number of beneficial biological effects of CGAs, such as anti-inflammatory activity, anti-carcinogenic activity, and protection against neurodegenerative diseases, have been reported. However, the molecular mechanisms at the base of these biological activities have not yet been investigated in depth. By combining NMR spectroscopy, molecular docking, surface plasmon resonance and ex vivo assays of the Ras-dependent breast cancer cell line MDA-MB-231, we contribute to the elucidation of the molecular basis of the activity of CGAs and natural extracts from green and roasted coffee beans as chemoprotective dietary supplements.