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Background Epigenetic age is an estimator of biological age based on DNA methylation; its discrepancy from chronologic age warrants further investigation. We recently reported that greater polyphenol intake benefitted ectopic fats, brain function, and gut microbiota profile, corresponding with elevated urine polyphenols. The effect of polyphenol-rich dietary interventions on biological aging is yet to be determined. Methods We calculated different biological aging epigenetic clocks of different generations (Horvath2013, Hannum2013, Li2018, Horvath skin and blood2018, PhenoAge2018, PCGrimAge2022), their corresponding age and intrinsic age accelerations, and DunedinPACE, all based on DNA methylation (Illumina EPIC array; pre-specified secondary outcome) for 256 participants with abdominal obesity or dyslipidemia, before and after the 18-month DIRECT PLUS randomized controlled trial. Three interventions were assigned: healthy dietary guidelines, a Mediterranean (MED) diet, and a polyphenol-rich, low-red/processed meat Green-MED diet. Both MED groups consumed 28 g walnuts/day (+ 440 mg/day polyphenols). The Green-MED group consumed green tea (3–4 cups/day) and Mankai (Wolffia globosa strain) 500-ml green shake (+ 800 mg/day polyphenols). Adherence to the Green-MED diet was assessed by questionnaire and urine polyphenols metabolomics (high-performance liquid chromatography quadrupole time of flight). Results Baseline chronological age (51.3 ± 10.6 years) was significantly correlated with all methylation age (mAge) clocks with correlations ranging from 0.83 to 0.95; p < 2.2e − 16 for all. While all interventions did not differ in terms of changes between mAge clocks, greater Green-Med diet adherence was associated with a lower 18-month relative change (i.e., greater mAge attenuation) in Li and Hannum mAge (beta = − 0.41, p = 0.004 and beta = − 0.38, p = 0.03, respectively; multivariate models). Greater Li mAge attenuation (multivariate models adjusted for age, se

Background The capacity of a polyphenol-enriched diet to modulate the epigenome in vivo is partly unknown. Given the beneficial metabolic effects of a Mediterranean (MED) diet enriched in polyphenols and reduced in red/processed meat (green-MED), as previously been proven by the 18-month DIRECT PLUS randomized controlled trial, we analyzed the effects of the green-MED diet on methylome and transcriptome levels to highlight molecular mechanisms underlying the observed metabolic improvements. Methods Our study included 260 participants (baseline BMI = 31.2 kg/m2, age = 5 years) of the DIRECT PLUS trial, initially randomized to one of the intervention arms: A. healthy dietary guidelines (HDG), B. MED (440 mg polyphenols additionally provided by walnuts), C. green-MED (1240 mg polyphenols additionally provided by walnuts, green tea, and Mankai: green duckweed shake). Blood methylome and transcriptome of all study subjects were analyzed at baseline and after completing the 18-month intervention using Illumina EPIC and RNA sequencing technologies. Results A total of 1573 differentially methylated regions (DMRs; false discovery rate (FDR) < 5 %) were found in the green-MED compared to the MED (177) and HDG (377) diet participants. This corresponded to 1753 differentially expressed genes (DEGs; FDR < 5 %) in the green-MED intervention compared to MED (7) and HDG (738). Consistently, the highest number (6 %) of epigenetic modulating genes was transcriptionally changed in subjects participating in the green-MED intervention. Weighted cluster network analysis relating transcriptional and phenotype changes among participants subjected to the green-MED intervention identified candidate genes associated with serum-folic acid change (all P < 1 × 10−3) and highlighted one module including the

BackgroundMediterranean (MED) diet is a rich source of polyphenols, which benefit adiposity by several mechanisms. We explored the effect of the green-MED diet, twice fortified in dietary polyphenols and lower in red/processed meat, on visceral adipose tissue (VAT).MethodsIn the 18-month Dietary Intervention Randomized Controlled Trial PoLyphenols UnproceSsed (DIRECT-PLUS) weight-loss trial, 294 participants were randomized to (A) healthy dietary guidelines (HDG), (B) MED, or (C) green-MED diets, all combined with physical activity. Both isocaloric MED groups consumed 28 g/day of walnuts (+ 440 mg/day polyphenols). The green-MED group further consumed green tea (3–4 cups/day) and Wolffia globosa (duckweed strain) plant green shake (100 g frozen cubes/day) (+ 800mg/day polyphenols) and reduced red meat intake. We used magnetic resonance imaging (MRI) to quantify the abdominal adipose tissues.ResultsParticipants (age = 51 years; 88% men; body mass index = 31.2 kg/m2; 29% VAT) had an 89.8% retention rate and 79.3% completed eligible MRIs. While both MED diets reached similar moderate weight (MED: − 2.7%, green-MED: − 3.9%) and waist circumference (MED: − 4.7%, green-MED: − 5.7%) loss, the green-MED dieters doubled the VAT loss (HDG: − 4.2%, MED: − 6.0%, green-MED: − 14.1%; p < 0.05, independent of age, sex, waist circumference, or weight loss). Higher dietary consumption of green tea, walnuts, and Wolffia globosa; lower red meat intake; higher total plasma polyphenols (mainly hippuric acid), and elevated urine urolithin A polyphenol were significantly related to greater VAT loss (p < 0.05, multivariate models).ConclusionsA green-MED diet, enriched with plant-based polyphenols and lower in red/processed meat, may be a potent intervention to promote visceral adiposity regression.

Background: Polyphenols are secondary metabolites produced by plants to defend themselves from environmental stressors. We explored the effect of Wolffia globosa ‘Mankai’, a novel cultivated strain of a polyphenol-rich aquatic plant, on the metabolomic-gut clinical axis in vitro, in-vivo and in a clinical trial. Methods: We used mass-spectrometry-based metabolomics methods from three laboratories to detect Mankai phenolic metabolites and examined predicted functional pathways in a Mankai artificial-gut bioreactor. Plasma and urine polyphenols were assessed among the 294 DIRECT-PLUS 18-month trial participants, comparing the effect of a polyphenol-rich green-Mediterranean diet (+1240 mg/polyphenols/day, provided by Mankai, green tea and walnuts) to a walnuts-enriched (+440 mg/polyphenols/day) Mediterranean diet and a healthy controlled diet. Results: Approximately 200 different phenolic compounds were specifically detected in the Mankai plant. The Mankai-supplemented bioreactor artificial gut displayed a significantly higher relative-abundance of 16S-rRNA bacterial gene sequences encoding for enzymes involved in phenolic compound degradation. In humans, several Mankai-related plasma and urine polyphenols were differentially elevated in the green Mediterranean group compared with the other groups (p < 0.05) after six and 18 months of intervention (e.g., urine hydroxy-phenyl-acetic-acid and urolithin-A; plasma Naringenin and 2,5-diOH-benzoic-acid). Specific polyphenols, such as urolithin-A and 4-ethylphenol, were directly involved with clinical weight-related changes. Conclusions: The Mankai new plant is rich in various unique potent polyphenols, potentially affecting the metabolomic-gut-clinical axis.

Background A Mediterranean diet is favourable for cardiometabolic risk. Objective To examine the residual effect of a green Mediterranean diet, further enriched with green plant-based foods and lower meat intake, on cardiometabolic risk. Methods For the DIRECT-PLUS parallel, randomised clinical trial we assigned individuals with abdominal obesity/dyslipidaemia 1:1:1 into three diet groups: healthy dietary guidance (HDG), Mediterranean and green Mediterranean diet, all combined with physical activity. The Mediterranean diets were equally energy restricted and included 28 g/day walnuts. The green Mediterranean diet further included green tea (3–4 cups/day) and a Wolffia globosa (Mankai strain; 100 g/day frozen cubes) plant-based protein shake, which partially substituted animal protein. We examined the effect of the 6-month dietary induction weight loss phase on cardiometabolic state. Results Participants (n=294; age 51 years; body mass index 31.3 kg/m2; waist circumference 109.7 cm; 88% men; 10 year Framingham risk score 4.7%) had a 6-month retention rate of 98.3%. Both Mediterranean diets achieved similar weight loss ((green Mediterranean −6.2 kg; Mediterranean −5.4 kg) vs the HDG group −1.5 kg; p<0.001), but the green Mediterranean group had a greater reduction in waist circumference (−8.6 cm) than the Mediterranean (−6.8 cm; p=0.033) and HDG (−4.3 cm; p<0.001) groups. Stratification by gender showed that these differences were significant only among men. Within 6 months the green Mediterranean group achieved greater decrease in low-density lipoprotein cholesterol (LDL-C; green Mediterranean −6.1 mg/dL (−3.7%), −2.3 (-0.8%), HDG −0.2 mg/dL (+1.8%); p=0.012 between extreme groups), diastolic blood pressure (green Mediterranean −7.2 mm Hg, Mediterranean −5.2 mm Hg, HDG −3.4 mm Hg; p=0.005 between extreme groups), and homeostatic model assessment for insulin resistance (green Mediterranean −0.77, Mediterranean −0.46, HDG −0.27; p=0.020 between extreme gr