Fatty Liver in Obesity: Long-lifestyle Follow-up (FLiO)

Study Overview
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Brief Summary
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Non-alcoholic fatty liver disease (NAFLD) is a condition of excessive hepatic lipid accumulation in subjects that consume less than 20g ethanol per day, without other known causes as drugs consumption or toxins exposure. In Western countries, the rate of this disease lies about 30% in the general adult population. The process of developing NAFLD can start from simple steatosis to non-alcoholic steatohepatitis (NASH), which eventually can lead to cirrhosis and hepatocellular carcinoma in the absence of alcohol abuse. Liver biopsy is considered the gold standard of steatosis, fibrosis and cirrhosis. However, it is rarely performed because it is an invasive procedure and investigators are focusing in the application of non-invasive liver damage scores for diagnosis. The pathogenesis of NAFLD is multifactorial and triggered by environmental factors such as unbalanced diets and overnutrition as well as by lack of physical activity in the context of a genetic predisposition. Nowadays, the treatment of NAFLD is based on diet and lifestyle modifications. Weight loss, exercise and healthy eating habits are the main tools to fight NAFLD. Nevertheless, there is no a well characterized dietary pattern and further studies are necessary. With this background, the general aim of this project is to increase the knowledge on the influence of nutritional/lifestyle interventions in obese patients with NAFLD, as well as contribute to identify non-invasive biomarkers/scores to early diagnosis of this pathology in future obese people.

Detailed Description
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This project is framed within the promotion of health and lifestyles and, specifically, in liver disorder linked to obesity (FLiO: Fatty Liver in Obesity). The investigation addresses a randomized, parallel, long-term personalized nutritional intervention with two strategies: 1) Control diet based on American Heart Association (AHA); 2) Fatty Liver in Obesity (FLiO) diet based on previous results (RESMENA project).The diet is based on macronutrient distribution, quality and quantity, and is characterized by a low glycemic load, high adherence to the Mediterranean diet and a high antioxidant capacity, with the inclusion of anti-inflammatory foods. It also takes into account the distribution of food throughout the day, number of meals, portion sizes, timing of meal, individual needs, dietary behavior (behavioral therapy: eat slowly, teach what to buy, what to eat, when to eat). The participants are instructed to follow this strategy. This strategy (RESMENA) was even more effective than AHA after 6 months follow-up, in terms of significant reduction of abdominal fat and blood glucose level. In addition, this diet had beneficial effects for participants who were obese and had values of altered glucose, reducing significantly in RESMENA participants LDL-oxidized marker. These results are very important to apply in the present investigation since that patients with NAFLD are commonly insulin resistant. Both strategies were designed within a hypocaloric dietary pattern (-30%) in order to achieve the American Association for the Study of Liver Diseases (AASLD) recommendations for the management of non-alcoholic liver disease (loss of at least 3-5% of body weight appears necessary to improve steatosis, but a greater weight loss, up to 10%, may be needed to improve necroinflammation). At this time the participants are individually supervised and encouraged to follow with the dietary planning instructions assigned. Furthermore, at baseline, 6, 12 and 24 months anticipated variables are obtained. Both dietary groups receive routine control (weight, body composition, strategy adherence) and dietary advice daily by phone (if they need help) and face to face at the time of routine control. In order to get a integral lifestyle intervention, all participants will be encouraged to follow a healthy lifestyle. Thus, physical activity will be recorded in each dietary group. The specific tasks: To recruit and select patients with the adequate characteristics to validate the conclusions reached. To develop and adequately transmit to each patient a personalized strategy according to the group randomly assigned ( AASLD vs FLiO strategy). To check the degree of adherence to the strategy set by regular monitoring: semiquantitative questionnaires of food consumption frequency, pedometers, accelerometers, weight control, satiety. To assess the effect of each strategy on body composition (weight, waist circumference, body fat, muscle mass, bone mineral density), physical status, general biochemistry (lipid profile, glycaemic profile, albumin, blood count, transaminases), specific biomarkers/metabolites in blood or urine (inflammation, oxidative stress, liver damage, appetite, psychological status), quality of life and related factors (anxiety, depression and sleep). To check the evolution of the liver damage, using non-invasive techniques (ultrasound, elastography and magnetic resonance imaging (MRI), metabolomics analysis) and calculating different validated liver scores from the data obtained with each strategy. To compare the effectiveness of strategies, considering not only the ability to decrease body fat, but also other risk factors present in the NAFLD patient such as insulin resistance and cardiovascular risk, which will result in improvement of liver damage. To analyze SNPs (DNA from oral epithelial cells) and the association with NAFLD (diagnosis and response to the strategies). To study gene expression (mRNAs) and microRNAs in white blood cells for identifying biomarkers of diagnosis and response to dietary strategy. To analyze gene DNA methylation patterns in white blood cells for identifying biomarkers of diagnosis and response to dietary strategy. To describe the intestinal microbiota composition by 16s sequencing at baseline and after nutritional intervention for diagnosis and response.

Official Title
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Non-alcoholic Fatty Liver Disease (NAFLD) in Overweight and Obese People Under Nutritional and Lifestyle Follow-up: a Randomized Controlled Trial

Conditions
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Non-Alcoholic Fatty Liver Disease, Obese, Overweight

Intervention / Treatment
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* Other: Control diet
* Other: FLiO diet


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Overweight or obese Diagnosis of NAFLD Age: 30-80 years Female / Male Exclusion Criteria: Known liver disease (other than NAFLD) Abuse of alcohol (>21 and >14 units of alcohol a week for men and women, respectively, eg 1 unit = 125 mL of wine); Drug treatments: immunosuppressants, cytotoxic agents, systemic corticosteroids, agents potentially causing fatty liver disease or abnormal liver tests or weight modifiers Active cancer or a history of malignancy in the last 5 years Problems of massive edemas Obesity known endocrine origin (except treated hypothyroidism) Surgical procedure for weight loss ≥ 3kg weight loss in the last 3 months Severe psychiatric disorders Lack of autonomy or inability to follow the diet (including food allergies or intolerances) or/and lifestyle recommendations as well as to follow scheduled visits. Consumption of any type of food supplements (antioxidants, prebiotics, probiotics, etc.)

Ages Eligible for Study
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Minimum Age: 30 Years
Maximum Age: 80 Years

Sexes Eligible for Study
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All

Accepts Healthy Volunteers
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No

Study Plan
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How is the study designed?
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Design Details

Primary Purpose: Treatment
Allocation: Randomized
Intervention Model: Parallel Assignment
Interventional Model Description: The participants are randomly assigned to Control or FLiO strategy.
Masking: Single


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Placebo Comparator: Control diet<br>A conventional and balanced diet based on American Heart Association (AHA) guidelines and lifestyle advice to achieve the objective of American Association for the Study of Liver Diseases (AASLD): loss of at least 3-5% of the initial body weight and up to 10% needed to improve necroinflammation. | Other: Control diet<br>* The participants follow a conventional and balanced distribution of macronutrients (30% fat, 15% protein, 55% carbohydrates), adequate fiber (25-30 g/day) and dietary cholesterol (<250 mg/day) intake according to AHA guidelines. This strategy was included within a personalized energy-restricted diet (-30% individual needs) under healthy lifestyle advice in order to achieve the objectives of AASLD (loss of at least 3-5% of the initial body weight and up to 10% needed to improve necroinflammation).<br>* Other names: American Heart Association diet;|
| Experimental: FLiO diet<br>A mediterranean dietary strategy based on macronutrient distribution (quantity and quality), antioxidant capacity, meal frequency, dietary behaviour and lifestyle advice to achieve the objective of AASLD: loss of at least 3-5% of the initial body weight and up to 10% needed to improve necroinflammation. | Other: FLiO diet<br>* The participants follow a strategy based on a distribution of macronutrients 30-35% lipid (extra virgin olive oil and fatty acids Ω3 in detriment of saturated, trans and cholesterol)/ protein 25% (vegetable against animal)/carbohydrates 40-45% (low glycaemic index, fiber 30-35 g/day); high adherence to the Mediterranean diet and natural antioxidants; meal frequency of 7 meals/day; size/composition of the ration suitable for each moment; including traditional foods with no additional economic cost that will allow diet adherence without abandonment; avoid inappropriate mealtimes and the eating manners as the eating rate. The participants are instructed to follow this strategy within a personalized energy-restricted diet (-30%) and under healthy lifestyle advice to achieve AASLD objectives.<br>* Other names: Fatty Liver in Obesity diet;|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change from Baseline Weight at 6 months | Weight will be measured by a digital scale | Baseline and 6 months | 
| Change from 6 month Weight at 12 months | Weight will be measured by a digital scale | 6 months and 12 months | 
| Change from Baseline Weight at 12 months | Weight will be measured by a digital scale | Baseline and 12 months | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Change from Baseline Body fat at 6 months | Fat mass will be measured by Dual X-ray absorptiometry | Baseline and 6 months | 
| Change from 6 month Body fat at 12 months | Fat mass will be measured by Dual X-ray absorptiometry | 6 months and 12 months | 
| Change from Baseline Body fat at 12 months | Fat mass will be measured by Dual X-ray absorptiometry | Baseline and 12 months | 
| Change from Baseline Waist circumference at 6 months | Waist circumference will be measured with a tape measure | Baseline and 6 months | 
| Change from 6 month Waist circumference at 12 months | Waist circumference will be measured with a tape measure | 6 months and 12 months | 
| Change from Baseline Waist circumference at 12 months | Waist circumference will be measured with a tape measure | Baseline and 12 months | 
| Change from Baseline handgrip strength at 6 months | Handgrip strength will be measured with a dynamometer | Baseline and 6 months | 
| Change from 6 month handgrip strength at 12 months | Handgrip strength will be measured with a dynamometer | 6 months and 12 months | 
| Change from Baseline handgrip strength at 12 months | Handgrip strength will be measured with a dynamometer | Baseline and 12 months | 
| Change from Baseline Systolic blood pressure at 6 months | Systolic blood pressure will be measured with a sphygmomanometer | Baseline and 6 months | 
| Change from 6 month Systolic blood pressure at 12 months | Systolic blood pressure will be measured with a sphygmomanometer | 6 months and 12 months | 
| Change from Baseline Systolic blood pressure at 12 months | Systolic blood pressure will be measured with a sphygmomanometer | Baseline and 12 months | 
| Change from Baseline Diastolic blood pressure at 6 months | Diastolic blood pressure will be measured with a sphygmomanometer | Baseline and 6 months | 
| Change from 6 month Diastolic blood pressure at 12 months | Diastolic blood pressure will be measured with a sphygmomanometer | 6 months and 12 months | 
| Change from Baseline Diastolic blood pressure at 12 months | Diastolic blood pressure will be measured with a sphygmomanometer | Baseline and 12 months | 
| Change from Baseline lipid metabolism at 6 months | Serum free fatty acids, triglycerides, total cholesterol, LDL cholesterol and HDL cholesterol concentrations will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month lipid metabolism at 12 months | Serum free fatty acids, triglycerides, total cholesterol, LDL cholesterol and HDL cholesterol concentrations will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline lipid metabolism at 12 months | Serum free fatty acids, triglycerides, total cholesterol, LDL cholesterol and HDL cholesterol concentrations will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline uric acid concentration at 6 months | Serum uric acid will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month uric acid concentration at 12 months | Serum uric acid will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline uric acid concentration at 12 months | Serum uric acid will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline homocysteine concentration at 6 months | Serum homocysteine will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month homocysteine concentration at 12 months | Serum homocysteine will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline homocysteine concentration at 12 months | Serum homocysteine will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline glucose metabolism at 6 months | Serum glucose levels will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month glucose metabolism at 12 months | Serum glucose levels will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline glucose metabolism at 12 months | Serum glucose levels will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline insulin concentration at 6 months | Serum insulin levels will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month insulin concentration at 12 months | Serum insulin levels will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline insulin concentration at 12 months | Serum insulin levels will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline Hemoglobin A1c concentration at 6 months | Serum Hemoglobin A1c will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month Hemoglobin A1c concentration at 12 months | Serum Hemoglobin A1c will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline Hemoglobin A1c concentration at 12 months | Serum Hemoglobin A1c will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline liver function at 6 months | Serum aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltransferase, total bilirubin, direct bilirubin, alkaline phosphatase, creatinine, total protein, albumin, prothrombin will be measured in a fasting state | Baseline and 12 months | 
| Change from 6 month liver function at 12 months | Serum aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltransferase, total bilirubin, direct bilirubin, alkaline phosphatase, creatinine, total protein, albumin, prothrombin will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline liver function at 12 months | Serum aspartate aminotransferase, alanine aminotransferase, gamma-glutamyltransferase, total bilirubin, direct bilirubin, alkaline phosphatase, creatinine, total protein, albumin, prothrombin will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline fibroblast growth factor 21 (FGF21) concentration at 6 months | Plasma FGF21 is a specific biomarker of NAFLD and will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month fibroblast growth factor 21 (FGF21) concentration at 12 months | Plasma FGF21 is a specific biomarker of NAFLD and will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline fibroblast growth factor 21 (FGF21) concentration at 12 months | Plasma FGF21 is a specific biomarker of NAFLD and will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline cytokeratin-18 (CK18) concentration at 6 months | Plasma CK18 is a specific biomarker of NAFLD and will be measured in a fasting state | Baseline and 6 months | 
| Change from 6 month cytokeratin-18 (CK18) concentration at 12 months | Plasma CK18 is a specific biomarker of NAFLD and will be measured in a fasting state | 6 months and 12 months | 
| Change from Baseline cytokeratin-18 (CK18) concentration at 12 months | Plasma CK18 is a specific biomarker of NAFLD and will be measured in a fasting state | Baseline and 12 months | 
| Change from Baseline C-reactive protein (CRP) concentration at 6 months | Plasma CRP will be assessed to determine inflammatory status | Baseline and 6 months | 
| Change from 6 month C-reactive protein (CRP) concentration at 12 months | Plasma CRP will be assessed to determine inflammatory status | 6 months and 12 months | 
| Change from Baseline C-reactive protein (CRP) concentration at 12 months | Plasma CRP will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline interleukin 6 (IL-6) concentration at 6 months | Plasma IL-6 will be assessed to determine inflammatory status | Baseline and 6 months | 
| Change from 6 month interleukin 6 (IL-6) concentration at 12 months | Plasma IL-6 will be assessed to determine inflammatory status | 6 months and 12 months | 
| Change from Baseline interleukin 6 (IL-6) concentration at 12 months | Plasma IL-6 will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline tumor necrosis factor-α (TNFα) concentration at 6 months | Plasma TNF-alpha will be assessed to determine inflammatory status | Baseline and 6 months | 
| Change from 6 month tumor necrosis factor-α (TNFα) concentration at 12 months | Plasma TNF-alpha will be assessed to determine inflammatory status | 6 months and 12 months | 
| Change from Baseline tumor necrosis factor-α (TNFα) concentration at 12 months | Plasma TNF-alpha will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline leptin concentration at 6 months | Plasma leptin will be assessed to determine inflammatory status | Baseline and 6 months | 
| Change from 6 month leptin concentration at 12 months | Plasma leptin will be assessed to determine inflammatory status | 6 months and 12 months | 
| Change from Baseline leptin concentration at 12 months | Plasma leptin will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline adiponectin concentration at 6 months | Plasma leptin will be assessed to determine inflammatory status | Baseline and 6 months | 
| Change from 6 month adiponectin concentration at 12 months | Plasma adiponectin will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline adiponectin concentration at 12 months | Plasma adiponectin will be assessed to determine inflammatory status | Baseline and 12 months | 
| Change from Baseline LDL-oxidized concentration at 6 months | LDL-ox will be assessed to determine oxidative status | Baseline and 6 months | 
| Change from 6 month LDL-oxidized concentration at 12 months | LDL-ox will be assessed to determine oxidative status | 6 months and 12 months | 
| Change from Baseline LDL-oxidized concentration at 12 months | LDL-ox will be assessed to determine oxidative status | Baseline and 12 months | 
| Change from Baseline Malondialdehyde concentration at 6 months | Plasma malondialdehyde will be assessed to determine oxidative status | Baseline and 6 months | 
| Change from 6 month Malondialdehyde concentration at 12 months | Plasma malondialdehyde will be assessed to determine oxidative status | 6 months and 12 months | 
| Change from Baseline Malondialdehyde concentration at 12 months | Plasma malondialdehyde will be assessed to determine oxidative status | Baseline and 12 months | 
| Change from Baseline plasma antioxidant capacity at 6 months | Plasma antioxidant capacity will be assessed by measuring the ferric reducing ability of plasma (FRAP) | Baseline and 6 months | 
| Change from 6 month plasma antioxidant capacity at 12 months | Plasma antioxidant capacity will be assessed by measuring the ferric reducing ability of plasma (FRAP) | 6 months and 12 months | 
| Change from Baseline plasma antioxidant capacity at 12 months | Plasma antioxidant capacity will be assessed by measuring the ferric reducing ability of plasma (FRAP) | Baseline and 12 months | 
| Change from Baseline Hepatic echography at 6 months | Echography will be carried out to analyze liver steatosis | Baseline and 6 months | 
| Change from 6 month Hepatic echography at 12 months | Echography will be carried out to analyze liver steatosis | 6 months and 12 months | 
| Change from Baseline Hepatic echography at 12 months | Echography will be carried out to analyze liver steatosis | Baseline and 12 months | 
| Change from Baseline Hepatic elastography at 6 months | Elastography will be carried out to analyze liver fibrosis | Baseline and 6 months | 
| Change from 6 month Hepatic elastography at 12 months | Elastography will be carried out to analyze liver fibrosis | 6 months and 12 months | 
| Change from Baseline Hepatic elastography at 12 months | Elastography will be carried out to analyze liver fibrosis | Baseline and 12 months | 
| Change from Baseline Hepatic Magnetic Resonance Imaging at 6 months | Magnetic Resonance Imaging will be carried out to analyze liver status | Baseline and 6 months | 
| Change from 6 month Hepatic Magnetic Resonance Imaging at 12 months | Magnetic Resonance Imaging will be carried out to analyze liver status | 6 months and 12 months | 
| Change from Baseline Hepatic Magnetic Resonance Imaging at 12 months | Magnetic Resonance Imaging will be carried out to analyze liver status | Baseline and 12 months | 
| Change from Baseline White blood cell count at 6 months | White blood cell count includes: Leucocytes, Neutrophils, Lymphocytes, Monocytes, Eosinophil, Basophils. | Baseline and 6 months | 
| Change from 6 month White blood cell count at 12 months | White blood cell count includes: Leucocytes, Neutrophils, Lymphocytes, Monocytes, Eosinophil, Basophils. | 6 months and 12 months | 
| Change from Baseline White blood cell count at 12 months | White blood cell count includes: Leucocytes, Neutrophils, Lymphocytes, Monocytes, Eosinophil, Basophils. | Baseline and 12 months | 
| Change from Baseline blood rheological properties at 6 months | Red blood cell count, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, red cell distribution width, platelet count, platelet distribution width, mean platelet volume, plateletcrit | Baseline and 6 months | 
| Change from 6 month blood rheological properties at 12 months | Red blood cell count, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, red cell distribution width, platelet count, platelet distribution width, mean platelet volume, plateletcrit | 6 months and 12 months | 
| Change from Baseline blood rheological properties at 12 months | Red blood cell count, hematocrit, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, red cell distribution width, platelet count, platelet distribution width, mean platelet volume, plateletcrit | Baseline and 12 months | 
| Change from Baseline Physical activity level at 6 months | Physical activity will be assessed by accelerometers | Baseline and 6 months | 
| Change from 6 months Physical activity level at 12 months | Physical activity will be assessed accelerometers | 6 months and 12 months | 
| Change from Baseline Physical activity level at 12 months | Physical activity will be assessed by accelerometers | Baseline and 12 months | 
| Change from Baseline Minnesota Physical Activity test at 6 months | Physical activity assessed by Minnesota Physical Activity test | Baseline and 6 months | 
| Change from 6 month Minnesota Physical Activity test at 12 months | Physical activity assessed by Minnesota Physical Activity test | 6 months and 12 months | 
| Change from Baseline Minnesota Physical Activity test at 12 months | Physical activity assessed by Minnesota Physical Activity test | Baseline and 12 months | 
| Change from Baseline number of steps at 6 months | Physical activity assessed by Pedometers | Baseline and 6 months | 
| Change from 6 month number of steps at 12 months | Physical activity assessed by Pedometers | 6 months and 12 months | 
| Change from Baseline number of steps at 12 months | Physical activity assessed by Pedometers | Baseline and 12 months | 
| Change from Baseline chair test at 6 months | Physical activity assessed by the chair test | Baseline and 6 months | 
| Change from 6 month chair test at 12 months | Physical activity assessed by the chair test | 6 months and 12 months | 
| Change from Baseline chair test at 12 months | Physical activity assessed by the chair test | Baseline and 12 months | 
| Change from Baseline sleep quality at 6 months | Sleep information will be assessed by the Pittsburgh Sleep Quality Index | Baseline and 12 months | 
| Change from 6 month sleep quality at 12 months | Sleep information will be assessed by the Pittsburgh Sleep Quality Index | 6 months and 12 months | 
| Change from Baseline sleep quality at 12 months | Sleep information will be assessed by the Pittsburgh Sleep Quality Index | Baseline and 12 months | 
| Change from Baseline Depressive symptoms at 6 months | Depressive symptoms will be assessed by the Beck Depression Inventory (BDI) | Baseline and 6 months | 
| Change from 6 month Depressive symptoms at 12 months | Depressive symptoms will be assessed by the Beck Depression Inventory (BDI) | 6 months and 12 months | 
| Change from Baseline Depressive symptoms at 12 months | Depressive symptoms will be assessed by the Beck Depression Inventory (BDI) | Baseline and 12 months | 
| Change from Baseline Anxiety symptoms at 6 months | Anxiety symptoms will be assessed by State Anxiety test (STAI) | Baseline and 6 months | 
| Change from 6 month Anxiety symptoms at 12 months | Anxiety symptoms will be assessed by State Anxiety test (STAI) | 6 months and 12 months | 
| Change from Baseline Anxiety symptoms at 12 months | Anxiety symptoms will be assessed by State Anxiety test (STAI) | Baseline and 12 months | 
| Single Nucleotide polymorphisms (SNPs) | Single nucleotide polymorphisms will be determined by Genomic DNA from oral epithelial cells | Baseline | 
| Change from Baseline DNA methylation at 6 months | Epigenetics will be assessed by changes in DNA methylation of genes related with NAFLD development | Baseline and 6 months | 
| Change from 6 month DNA methylation at 12 months | Epigenetics will be assessed by changes in DNA methylation of genes related with NAFLD development | 6 months and 12 months | 
| Change from Baseline DNA methylation at 12 months | Epigenetics will be assessed by changes in DNA methylation of genes related with NAFLD development | Baseline and 12 months | 
| Change from Baseline microRNAs at 6 months | Transcriptomic will be assessed by changes in miRNAs | Baseline and 6 months | 
| Change from 6 month microRNAs at 12 months | Transcriptomic will be assessed by changes in miRNAs | 6 months and 12 months | 
| Change from Baseline microRNAs at 12 months | Transcriptomic will be assessed by changes in miRNAs | Baseline and 12 months | 
| Change from Baseline Gut microbiota composition at 6 months | Gut microbiota composition will be analyzed | Baseline and 6 months | 
| Change from 6 month Gut microbiota composition at 12 month | Gut microbiota composition will be analyzed | 6 months and 12 months | 
| Change from Baseline Gut microbiota composition at 12 month | Gut microbiota composition will be analyzed | Baseline and 12 months | 
| Change from Baseline metabolites composition of urine at 6 months | Metabolites composition of urine will be analyzed | Baseline and 6 months | 
| Change from 6 month metabolites composition of urine at 12 months | Metabolites composition of urine will be analyzed | 6 months and 12 months | 
| Change from Baseline metabolites composition of urine at 12 months | Metabolites composition of urine will be analyzed | Baseline and 12 months | 
| Change from Baseline metabolites composition of serum at 6 months | Metabolites composition of serum will be analyzed | Baseline and 6 months | 
| Change from 6 month metabolites composition of serum at 12 months | Metabolites composition of serum will be analyzed | 6 months and 12 months | 
| Change from Baseline metabolites composition of serum at 12 months | Metabolites composition of serum will be analyzed | Baseline and 12 months | 
| Change from Baseline dietary intake at 6 months | Dietary intake will be assessed by means of food frequency questionnaire | Baseline and 6 months | 
| Change from 6 month dietary intake at 12 months | Dietary intake will be assessed by means of food frequency questionnaire | 6 months and 12 months | 
| Change from Baseline dietary intake at 12 months | Dietary intake will be assessed by means of food frequency questionnaire | Baseline and 12 months | 
| Assessment of dietary adherence at Baseline | Dietary adherence will be assessed by means of 3 day weighed food records | Baseline | 
| Assessment of dietary adherence at 6 months | Dietary adherence will be assessed by means of 3 day weighed food records | 6 months | 
| Assessment of dietary adherence at 12 months | Dietary adherence will be assessed by means of 3 day weighed food records | 12 months | 
| Change from Baseline satiety index at 6 months | Satiety index/appetite will be assessed by using the 100 mm Visual Analogue Scale | Baseline and 6 months | 
| Change from 6 month satiety index at 12 months | Satiety index/appetite will be assessed by using the 100 mm Visual Analogue Scale | 6 months and 12 months | 
| Change from Baseline satiety index at 12 months | Satiety index/appetite will be assessed by using the 100 mm Visual Analogue Scale | Baseline and 12 months | 
| Change from Baseline life quality index at 6 months | Life quality index will be assessed by means of the Short Form 36 (SF-36) questionnaire | Baseline and 6 months | 
| Change from 6 month life quality index at 12 months | Life quality index will be assessed by means of the Short Form 36 (SF-36) questionnaire | 6 months and 12 months | 
| Change from Baseline life quality index at 12 months | Life quality index will be assessed by means of the Short Form 36 (SF-36) questionnaire | Baseline and 12 months | 
| Change from Baseline Ghrelin concentration at 6 months | Serum Active Ghrelin will be determined to assess satiety | Baseline and 6 months | 
| Change from 6 month Ghrelin concentration at 12 months | Serum Active Ghrelin will be determined to assess satiety | 6 months and 12 months | 
| Change from Baseline Ghrelin concentration at 12 months | Serum Active Ghrelin will be determined to assess satiety | Baseline and 12 months | 
| Change from Baseline glucagon-like peptide-1 (GLP-1) concentration at 6 months | Serum active glucagon-like peptide-1 will be determined to assess satiety | Baseline and 6 months | 
| Change from 6 month glucagon-like peptide-1 (GLP-1) concentration at 12 months | Serum active glucagon-like peptide-1 will be determined to assess satiety | 6 months and 12 months | 
| Change from Baseline glucagon-like peptide-1 (GLP-1) concentration at 12 months | Serum active glucagon-like peptide-1 will be determined to assess satiety | Baseline and 12 months | 
| Change from Baseline Dopamine concentration at 6 months | Peripheral Dopamine concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 6 months | 
| Change from 6 month Dopamine concentration at 12 months | Peripheral Dopamine concentration will be analysed using high-performance liquid chromatography (HPLC) | 6 months and 12 months | 
| Change from Baseline Dopamine concentration at 12 months | Peripheral Dopamine concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 12 months | 
| Change from Baseline Dopac concentration at 6 months | Peripheral Dopac concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 6 months | 
| Change from 6 month Dopac concentration at 12 months | Peripheral Dopac concentration will be analysed using high-performance liquid chromatography (HPLC) | 6 months and 12 months | 
| Change from Baseline Dopac concentration at 12 months | Peripheral Dopac concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 12 months | 
| Change from Baseline Serotonin (5-HT) concentration at 6 months | Peripheral Serotonin concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 6 months | 
| Change from 6 month Serotonin (5-HT) concentration at 12 months | Peripheral Serotonin concentration will be analysed using high-performance liquid chromatography (HPLC) | 6 months and 12 months | 
| Change from Baseline Serotonin (5-HT) concentration at 12 months | Peripheral Serotonin concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 12 months | 
| Change from Baseline Noradrenaline concentration at 6 months | Peripheral Noradrenaline concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 6 months | 
| Change from 6 month Noradrenaline concentration at 12 months | Peripheral Noradrenaline concentration will be analysed using high-performance liquid chromatography (HPLC) | 6 months and 12 months | 
| Change from Baseline Noradrenaline concentration at 12 months | Peripheral Noradrenaline concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 12 months | 
| Change from Baseline 5-hydroxyindoleacetic acetic (5-HIAAC) concentration at 6 months | Peripheral 5-hydroxyindoleacetic acetic concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 6 months | 
| Change from 6 month 5-hydroxyindoleacetic acetic (5-HIAAC) concentration at 12 months | Peripheral 5-hydroxyindoleacetic acetic concentration will be analysed using high-performance liquid chromatography (HPLC) | 6 months and 12 months | 
| Change from Baseline 5-hydroxyindoleacetic acetic (5-HIAAC) concentration at 12 months | Peripheral 5-hydroxyindoleacetic acetic concentration will be analysed using high-performance liquid chromatography (HPLC) | Baseline and 12 months | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Obesity, Physical activity, Sedentary behaviours, Sleep quality, Sleep duration, Psychological factors, Nutrigenomics, Nutrigenetics, Transcriptomics, Epigenetics, Metabolomics, miRNAs, Lipidomics, NAFLD, NASH, AASLD, AHA, Energy-restricted diet, Lifestyle intervention, Mediterranean diet, Weight loss, Liver damage, Inflammation, Oxidative stress, Microbiota composition, Eating behaviour, Satiety