How Healthy Is It to Fortify Cocoa-Based Products with Cocoa Flavanols? A Comprehensive Review

Background: Cocoa’s healthy benefits may be attributed to the potent antioxidant activity of cocoa polyphenols, mainly flavanols, which have been characterised as existing in a high concentration in cocoa. However, the phenolic composition of cocoa and cocoa-derived products is highly variable, and manufacturing processes might significantly reduce their phenolic content. For that reason, the full characterisation of cocoa and cocoa-derived products before evaluating their bioactivity is crucial. The aim of this review is to analyse the available evidence on the effect of flavanol-fortified cocoa-derived products on human health. (2) Methods: Forty-eight clinical trials focused on the health effect of consuming flavanol-fortified drinks, bars and chocolate have been reviewed, with a total of 1523 subjects. (3) Results: Although studies differ widely in methodology, dosage, duration, and target population, beneficial effects of flavanol-rich cocoa consumption have been observed at doses ranging from 45.3 mg/d to 1078 mg/d, especially on cardiovascular health and cognitive function. (4) Conclusions: Considering the high consumption and acceptability of cocoa and cocoa-derived products, the fortification of cocoa products as well as other highly consumed foods with cocoa flavanols could be an effective strategy for health promotionGrant PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/ 501100011033Junta de Andalucía-Consejería de Universidad, Investigación e Innovación—Proyect P21_0077

Estudio del cacao como potenciador de la neurogénesis y la función hipocampal en ratones adultos

Introducción: La formación de nuevas neuronas es un fenómeno neuroplástico esencial que se produce en el cerebro adulto, concretamente en el giro dentado del hipocampo (neurogénesis hipocampal adulta, NHA). Numerosos estudios demuestran que la inhibición de NHA en roedores conduce al deterioro cognitivo, mientras que su aumento potencia la adquisición, consolidación y actualización de los recuerdos dependientes del hipocampo. Estudios recientes destacan el papel de los polifenoles del cacao en la potenciación de la memoria, sin embargo, no existen evidencias concluyentes sobre su efecto en la NHA. Objetivos: En este proyecto, proponemos estudiar la modulación de la NHA mediante la dieta, empleando el cacao como una nueva intervención nutricional que podría potenciar la NHA mediante la acción de los polifenoles. Se emplearán ratones macho y hembra (aproximadamente 3 meses de edad), que recibirán una dieta estándar (grupos “control”) o una dieta enriquecida en cacao (10 %) durante ocho semanas. Se evaluará el efecto del cacao en la función cognitiva y emocional mediante distintas pruebas comportamentales que incluyen tareas para evaluar la ansiedad (laberinto en cruz), la exploración (campo abierto), la indefensión (test de natación forzada) y la memoria (reconocimiento de objeto y laberinto acuático). Se estudiará la NHA mediante la administración de bromodesoxiuridina y técnicas de inmunohistoquímica y microscopía, así como la expresión del factor neurotrófico brain derived neurotrophic factor (BDNF) en el hipocampo mediante western blot. Resultados esperados: Mediante este estudio se espera obtener una mejora del rendimiento cognitivo, debido principalmente a la potenciación de la plasticidad cerebral (incremento de la NHA y BDNF) trasla ingesta de cacao.Proyecto I+D+i PID2020-114374RB-100, financiado por MCIN/AEI/10.13039/501100011033/ Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Exploring the nutritional composition and bioactive compounds in different cocoa powders

Cocoa, the main derivative of the seeds of Theobroma cacao L., has been recognized to have several effects on human health including antioxidant and neuro- and cardio-protective effects, among others. These effects have been attributed mainly to its bioactive compounds. In this context, the aim of this work is to evaluate the nutritional composition, bioactive compounds (i.e., phenolic compounds, procyanidins and methylxanthines) and the antioxidant activity of seven different cocoas (alkalized and non-alkalized) from different origins (Peru, Venezuela, Ivory Coast, Dominican Republic, and West Africa). It represents the first stage of a larger project aiming to find high polyphenol cocoa-based nutritional strategies and related biomarkers that may potentiate brain plasticity and cognitive function. Cocoa powders were extracted by ultrasound-assisted technology, and the total phenolic content (TPC) was measured by Folin–Ciocalteu. Methylxanthines (caffeine and theobromine) and procyanidin contents were determined by HPLC-FLD-DAD, and the antioxidant activity was assessed through DPPH, ABTS and FRAP assays. Non-alkalized cocoas showed higher phenolic and procyanidin contents and higher antioxidant activity compared to the alkalized ones. A strongly significant (p < 0.05) positive correlation between the antioxidant activity and the TPC, especially with the total procyanidin content, but not with methylxanthines was found. In conclusion, the non-alkalized cocoas, especially the one from Peru, were the best candidates in terms of bioactive compounds. The cocoa from Peru had a TPC of 57.4 ± 14.4 mg of gallic acid equivalent/g d.w., 28,575.06 ± 62.37 µg of catechin equivalents/g d.w., and 39.15 ± 2.12 mg/g of methylxanthines. Further studies should be undertaken to evaluate its effect on brain plasticity and cognitive function.This study was funded by Grant PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033 (to C.R.-P. and E.C-O.) and Junta de Andalucía-Consejería de Universidad, Investigación e Innovación—Proyect P21_00777. Partial funding for open access charge: Universidad de Málag

Temozolomide treatment inhibits spontaneous motivation for exploring a complex object in mice: a potential role of adult hippocampal neurogenesis in ‘curiosity’.

Intrinsic exploratory biases are an innate motivation for exploring certain types of stimuli or environments over others, and they may be associated with cognitive, emotional, and even personality-like traits. However, their neurobiological basis has been scarcely investigated. Considering the involvement of the hippocampus in novelty recognition and in spatial and pattern separation tasks, this work researched the role of adult hippocampal neurogenesis (AHN) in intrinsic exploratory bias for a perceptually complex object in mice. Spontaneous object preference tasks revealed that both male and female C57BL/6J mice showed a consistent unconditioned preference for exploring “complex”—irregular—objects over simpler ones. Furthermore, increasing objects’ complexity resulted in an augmented time of object exploration. In a different experiment, male mice received either vehicle or the DNA alkylating agent temozolomide (TMZ) for 4 weeks, a pharmacological treatment that reduced AHN as evidenced by immunohistochemistry. After assessment in a behavioral test battery, the TMZ-treated mice did not show any alterations in general exploratory and anxiety-like responses. However, when tested in the spontaneous object preference task, the TMZ-treated mice did not display enhanced exploration of the complex object, as evidenced both by a reduced exploration time—specifically for the complex object—and a lack of preference for the complex object over the simple one. This study supports a novel role of AHN in intrinsic exploratory bias for perceptual complexity. Moreover, the spontaneous complex object preference task as a rodent model of “curiosity” is discussed.This study was funded by Grant PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033 (to C.R.-P. and E.C.-O.). Author P.R. holds a “Miguel Servet I” research contract from the National System of Health, EU-ERDF-ISCIII (CP19/00068). Authors M.C.M.-P. and S.G.-R. hold predoctoral grants from the Spanish Ministry of Science, Innovation and Universities (FPU17/00276 to M.C.M.-P. and FPU18/00941 to S.G.-R.). The authors acknowledge the IBIMA's common research support structure—ECAI—of animal experimentation and behavior (“Centro de Experimentación y Conducta Animal”; University of Malaga) for maintenance of the mice and the ECAI of Image for the use of the microscope. We are especially thankful to María Visitación Jacinto Hernández and Vanesa Jiménez Gálvez for their valuable contribution to the behavioral experiments and to Lourdes Sánchez Salido and Ana Mar Gálvez Callejón for their technical support. Open access funding provided by University of Málaga CBUA

Reduction of adult neurogenesis by temozolomide inhibits intrinsic preference for exploring complex objects in mice

Póster de congresoIntrinsic exploratory bias is an innate tendency to prefer certain types of stimuli or environments over others. For example, mice would genuinely spent more time exploring perceptually complex objects (i.e. with edges and concavities) than simpler objects without irregularities. Intrinsic exploratory bias are relevant as they may be associated to cognitive, emotional and even personality-like traits. However, their neurobiological basis are scarcely investigated. Adult hippocampal neurogenesis (AHN) is a key neuroplastic phenomenon for the processing of spatial and contextual stimuli in rodents, being involved in novelty recognition, spatial navigation and spatial pattern separation tasks. Therefore, here we studied whether a pharmacological inhibition of AHN influences intrinsic motivation for exploring complex objects. Twenty male young adult C57BL/6J mice (∼3 months old) received vehicle or the DNA alkylating agent temozolomide (TMZ) for four weeks. Bromodeoxyuridine (BrdU) was administered weekly, confirming a reduction of AHN-related markers by TMZ. After the pharmacological treatment, mice were tested for behavior. TMZ did not impair mice’s health nor their general exploratory and anxiety-like responses. Unlike control mice, the TMZ-treated mice did not prefer exploring a complex (i.e. irregular) object over a simple (i.e. non-irregular) object of similar size presented at once. Nevertheless, they were able to discriminate a novel complex object from a familiar complex object. This suggest that the lack of intrinsic preference for complexity could be explained by motivational and not by cognitive variables. Future studies should investigate a new role of AHN in modulating exploratory bias.Universidad de Málaga; Project PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033/ Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Nutrition and adult neurogenesis in the hippocampus: Does what you eat help you remember?

This study was funded by grant PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033 (CR-P and EC-O); Junta de Andalucía-Consejería de Universidad, Investigación e Innovación - Proyecto P21_00777 (CR-P and EC-O); Ministerio de Sanidad, Delegación de Gobierno para el Plan Nacional sobre Drogas grant number 2020/048; Consejería de Universidad, Investigación e Innovación grant number PROYEXCEL_0029. P.R. (CP19/00068) holds a “Miguel Servet I” research contract from the National System of Health, ERDF-EU-ISCIII. MC holds a postdoctoral research contract from Consejeria de Salud – Junta de Andalucia (RH 0081-2021).Neurogenesis is a complex process by which neural progenitor cells (NPCs)/neural stem cells (NSCs) proliferate and differentiate into new neurons and other brain cells. In adulthood, the hippocampus is one of the areas with more neurogenesis activity, which is involved in the modulation of both emotional and cognitive hippocampal functions. This complex process is affected by many intrinsic and extrinsic factors, including nutrition. In this regard, preclinical studies performed in rats and mice demonstrate that high fats and/or sugars diets have a negative effect on adult hippocampal neurogenesis (AHN). In contrast, diets enriched with bioactive compounds, such as polyunsaturated fatty acids and polyphenols, as well as intermittent fasting or caloric restriction, can induce AHN. Interestingly, there is also growing evidence demonstrating that offspring AHN can be affected by maternal nutrition in the perinatal period. Therefore, nutritional interventions from early stages and throughout life are a promising perspective to alleviate neurodegenerative diseases by stimulating neurogenesis. The underlying mechanisms by which nutrients and dietary factors affect AHN are still being studied. Interestingly, recent evidence suggests that additional peripheral mediators may be involved. In this sense, the microbiota-gut-brain axis mediates bidirectional communication between the gut and the brain and could act as a link between nutritional factors and AHN. The aim of this mini-review is to summarize, the most recent findings related to the influence of nutrition and diet in the modulation of AHN. The importance of maternal nutrition in the AHN of the offspring and the role of the microbiota-gut-brain axis in the nutrition-neurogenesis relationship have also been included.Ministerio de Sanidad, Delegación de Gobierno para el Plan Nacional sobre Drogas grant number 2020/048Consejería de Universidad, Investigación e Innovación grant number PROYEXCEL_0029. P.R. (CP19/00068) holds a “Miguel Servet I” research contract from the National System of Health, ERDF-EU-ISCIII. MC holds a postdoctoral research contract from Consejeria de Salud – Junta de Andalucia (RH 0081-2021).Grant PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033 (CR-P and EC-O)Junta de Andalucía-Consejería de Universidad, Investigación e Innovación - Proyecto P21_00777 (CR-P and EC-O

Effects of cocoa-derived polyphenols treatment in regulating the adverse impacts of early-life stress in male and female mice.

Depression is one of the mental disorders that significantly affects today's society and calls for new treatments. In recent years, the gut microbiota has emerged as a key modulator of behaviour and mood due to the existence of the gut-brain microbiota axis. Environmental factors such as early stress or diet can negatively impact the predisposition to develop a mental disorder and the composition of the gut microbiota. Prebiotics are functional foods that stimulate the growth or activity of intestinal bacteria, providing benefits to the host's health. Polyphenols, such as those found in cocoa, possess prebiotic properties. Therefore, the consumption of polyphenols derived from cocoa could be a therapeutic pathway for mental disorders. Our study aimed to evaluate the impact of maternal separation with early weaning (MSEW) on depression and anxiety-like behaviours in male and female mice and neuroinflammatory markers in the brain. We also explore if the modulation of the gut microbiota through cocoa-derived polyphenols could ameliorate these behavioural and molecular alterations induced by early stress. Results showed higher immobility time in the tail suspension test in those mice exposed to MSEW and more depressive-like behaviour in females compared with males. Regarding results in the elevated plus maze, we found reduced travelled distance and time in open arms in MSEW mice. Additionally, we showed higher time in open arms in MSEW female mice treated with polyphenols than MSEW females that received a control diet, suggesting a reduction in anxiety like-behaviour. In conclusion, our findings support that early-life stress increases susceptibility to depression and anxiety-like behaviours in mice. Additionally, sex differences were observed, with females showing more pronounced depressive- and anxiety-like behaviour. Moreover, we highlight that modulation of gut microbiota could be a potential therapeutic target in mood disorders.Proyect PID2020-114374RB-I00 by MCIN/AEI/10.13039/501100011033, Junta de Andalucía POSTDOC21_00365, Universidad de Málaga BI-2022_05, Universidad de Málaga Investigación e Innovación Proyect P21_00777. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Impact of a cocoa-based diet rich in polyphenols on cognitive function, emotional behavior, and adult hippocampal neurogenesis in mice.

Cocoa consumption has long been associated with numerous health benefits due to its rich composition of minerals, fiber, methylxanthines, and polyphenols, particularly flavanols. However, the cognitive effects of cocoa have been scarcely investigated. To further explore the impact of cocoa on cognitive function and emotional behavior, a study was conducted using young adult male and female C57BL/6J mice. The mice were fed a standard diet, a high-polyphenol content cocoa (HPC) diet, or a low-polyphenol content cocoa (LPC) diet for four weeks. A battery of behavioral tests was performed to assess exploratory and emotional behavior and hippocampal-dependent memory. Additionally, adult hippocampal neurogenesis was also evaluated. The results indicated that cocoa-enriched diets did not have a substantial impact on general exploratory activity or anxiety-like behavior. However, the LPC diet led to decreased locomotor activity. The HPC diet improved object recognition memory, while all groups performed similarly in the place memory test. In the forced swimming test, both cocoa-enriched diets increased immobility behavior. Cocoa did not affect spatial memory in the water maze. Moreover, the HPC diet enhanced adult hippocampal neurogenesis. Sex differences did not significantly influence the observed effects. In conclusion, cocoa-enriched diets, particularly the HPC diet, have the potential to improve object memory and promote adult hippocampal neurogenesis in mice. The specific differences observed between the HPC and LPC diets are likely attributed to differing polyphenol compositions. These findings contribute to our understanding of the potential cognitive and neurogenic benefits associated with cocoa consumption. Project PID2020-114374RB-I00 funded by MCIN/AEI/10.13039/501100011033/ and Junta de Andalucía-Consejería de Universidad, Investigación e Innovación—Proyect P21_00777. University of Malaga. University of Malaga.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Inhibition of Adult Neurogenesis in Male Mice after Repeated Exposure to Paracetamol Overdose

Paracetamol, or acetaminophen (N-acetyl-para-aminophenol, APAP), is an analgesic and antipyretic drug that is commonly used worldwide, implicated in numerous intoxications due to overdose, and causes serious liver damage. APAP can cross the blood–brain barrier and affects brain function in numerous ways, including pain signals, temperature regulation, neuroimmune response, and emotional behavior; however, its effect on adult neurogenesis has not been thoroughly investigated. We analyze, in a mouse model of hepatotoxicity, the effect of APAP overdose (750 mg/kg/day) for 3 and 4 consecutive days and after the cessation of APAP administration for 6 and 15 days on cell proliferation and survival in two relevant neurogenic zones: the subgranular zone of the dentate gyrus and the hypothalamus. The involvement of liver damage (plasma transaminases), neuronal activity (c-Fos), and astroglia (glial fibrillar acidic protein, GFAP) were also evaluated. Our results indicated that repeated APAP overdoses are associated with the inhibition of adult neurogenesis in the context of elevated liver transaminase levels, neuronal hyperactivity, and astrogliosis. These effects were partially reversed after the cessation of APAP administration for 6 and 15 days. In conclusion, these results suggest that APAP overdose impairs adult neurogenesis in the hippocampus and hypothalamus, a fact that may contribute to the effects of APAP on brain function