Metabolic and Inflammatory Outcomes of the Ketogenic Diet Comparing Saturated and Unsaturated Fat Sources

Study Overview
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Brief Summary
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The goal of this clinical trial is to compare a healthy KETO diet supplemented with canola oil (KETO-Can) compared to a traditional KETO diet high in saturated fat (KETO-Sat) and low-fat diet (LFD) in adults at high risk of or diagnosed with type 2 diabetes. The main question[s] it aims to answer are: Effects on CVD risk factors (plasma cholesterol, TG, ApoB100, glucose, insulin and HbA1C). Effects on systemic inflammation and immune function. Safety and adherence to interventions. Participants will be randomized into 1 of the dietary treatments during which they will follow a Keto or a low-fat diet. Comparisons among groups at 3 and 6 months of intervention will be conducted.

Detailed Description
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The ketogenic diet (KETO) is popular for weight loss and is gaining interest as a treatment for type 2 diabetes (T2D) because it is believed to help manage blood glucose and weight. However, KETO is often high in saturated fats (SFA), which may increase cholesterol and other cardiovascular (CVD) risk factors, such as inflammatory profile. Substituting a heart-healthy oil for SFA may improve these outcomes. The purpose of our study is to investigate the health beneficial effects of a healthy KETO diet supplemented with Canola oil, compared to a traditional Keto Diet and low-fat diet in adults at high risk of type 2 diabetes. Participants will be randomized to one of these three diets and will receive nutrition counselling during 6 months. Each month, participants will receive a 1-month supply of canola oil in the KETO-Can group, butter, coconut and palm oil in the KETO-Sat group and whole grain cereal and/or oatmeal in the LFD group to ensure compliance to key nutrients. Fasting blood samples will be taken at baseline, 3 and 6 months. Anthropometric measurements (weight (BW), waist circumference (WC), BMI), blood pressure (BP), systemic inflammation (CRP, IL-6, TNF-α, IL-18), immune function, cardiometabolic risk factors (TG, cholesterol, glucose, insulin and HbA1C) will be determined at each time point. A total of three 24h-recall questionnaires (2 weekdays and 1 weekend day) will be completed at each time point (baseline, 3 months, 6 months). Once a month (in between study visits) a 24h-recall will be completed before meeting the nutrition expert in order to personalize recommendations according to participants' respective diet groups. As in any nutritional study, compliance is a key factor and will be measured differently during the intervention. Menu examples will be provided for each group to facilitate adherence. Adherence to the study protocol will be assessed by (1) evaluation of monthly 24-h recall data (5 in total). Participants with 4 out of 5 recalls being within meeting dietary objectives will be considered highly compliant, 2 or less would be low compliance; (2) Ketosis state will be measured at each study visit using ketone strips to assess adherence to both KETO diets; (3) Participants will be asked to return any remaining food provided each month to determine the level of consumption. Finally, fatty acid composition in plasma (short-term) and red blood cells (RBCs; reflect the past 3 months) will be assessed to confirm adherence to the diets.

Official Title
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Metabolic and Inflammatory Outcomes of the Ketogenic Diet Comparing Saturated and Unsaturated Fat Sources

Conditions
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PreDiabetes, Diabetes Mellitus, Type 2, Overweight and Obesity

Intervention / Treatment
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* Other: Keto-SAT
* Other: Keto-CAN
* Other: LFD


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: Having overweight or obesity and HbA1C ≥ 5.7% at screening Exclusion Criteria: Individuals with specific nutritional habits preventing them from adhering to nutritional recommendations pregnant women people on dialysis or recommended to follow a low-protein diet (base on glomerular filtration rate) familial hypercholesterolemia or hypertriglyceridemia transitioning trans-gender transitioning menopausal women.

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 70 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: Other
Allocation: Randomized
Intervention Model: Parallel Assignment
Interventional Model Description: Randomized Controlled Trial (3-parallel arm study).
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Experimental: KETO-Can<br>KETO diet supplemented with Canola oil (high in MUFA and omega-3 FA). | Other: Keto-CAN<br>* Nutrition counselling focused on Keto diet (unsaturated fat). 10% TE as carbohydrate (mainly from vegetables and whole grain products), protein between 20-30% TE and fat between 60-70%.<br>|
| Experimental: KETO-Sat<br>KETO diet supplemented with butter, coconut and palm oil (high in SFA). | Other: Keto-SAT<br>* Nutrition counselling focused on Keto diet (saturated fat). 10% TE as carbohydrate (mainly from vegetables and whole grain products), protein between 20-30% TE and fat between 60-70%.<br>|
| Active Comparator: Low fat diet (LFD)<br>Low fat diet supplemented with whole grains and other low-fat foods. | Other: LFD<br>* Nutrition counselling focused on low-fat diet. 30% total energy (TE) as fat, 50% TE as carbohydrates (primarily whole grains) and 17-20% TE protein (mainly lean sources).<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Effect of diets on plasma triglycerides levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess triglycerides levels collecting blood samples in a fasting state. Triglycerides will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on plasma triglycerides levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess triglycerides levels collecting blood samples in a fasting state. Triglycerides will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 
| Effect of diets on LDL-cholesterol levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess LDL-cholesterol levels collecting blood samples in a fasting state. LDL-Cholesterol levels will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on LDL-cholesterol levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess LDL-cholesterol levels collecting blood samples in a fasting state. LDL-Cholesterol levels will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Effect of diets on blood glucose levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess blood glucose levels collecting blood samples in a fasting state. Blood glucose levels will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on blood glucose levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess blood glucose levels collecting blood samples in a fasting state. Blood glucose levels will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 
| Effect of diets on insulin levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess insulin levels collecting blood samples in a fasting state. Insulin levels will be determined using commercially available equipment at a local laboratory (Roche Cobas e801). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on insulin levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess insulin levels collecting blood samples in a fasting state. Insulin levels will be determined using commercially available equipment at a local laboratory (Roche Cobas e801). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 
| Effect of diets on hemoglobin A1C (HbA1c) levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess HbA1c collecting blood samples in a fasting state. HbA1c levels will be determined using commercially available equipment at a local laboratory (COBAS c513) and reported in percentage. Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on hemoglobin A1C (HbA1c) levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess HbA1c collecting blood samples in a fasting state. HbA1c levels will be determined using commercially available equipment at a local laboratory (COBAS c513) and reported in percentage. Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 
| Effect of diets on ApoB100 levels by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess ApoB100 collecting blood samples in a fasting state. ApoB100 levels will be determined using commercially available equipment at a local laboratory (Siemens BNII Nephelometer). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on ApoB100 levels by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess ApoB100 collecting blood samples in a fasting state. ApoB100 levels will be determined using commercially available equipment at a local laboratory (Siemens BNII Nephelometer). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months | 
| Effect of diets on systemic immune function and Inflammation measured by blood Immune Markers by comparing groups at 3 months of intervention and changes over time within each group compared to baseline. | In the fasting state, immune cell subset frequencies will be immunophenotyped by flow cytometry. Immune cells will be stained for T cells (CD3, CD4+, CD8+, CD45RA+, CD45RO+), Tregs (FOXP3+), B cell (CD19, CD20), macrophage (CD14, CD64), NK cell (CD56+) and activation markers (CD11b+, CD25+, CD28+, CD80+, CD127+, CD152+, CD278+). All samples will be acquired by flow cytometry and analyzed according to the relative fluorescence intensity using Flowjo Software. In the fasting and postprandial states, cell proliferation will be measured using alamar blue fluorescence and intracellular cytokines accumulation in different immune cell types will be determined using an intracellular staining flow. | 3 months | 
| Effect of diets on systemic immune function and Inflammation measured by blood Immune Markers by comparing groups at 6 months of intervention and changes over time within each group compared to baseline. | In the fasting state, immune cell subset frequencies will be immunophenotyped by flow cytometry. Immune cells will be stained for T cells (CD3, CD4+, CD8+, CD45RA+, CD45RO+), Tregs (FOXP3+), B cell (CD19, CD20), macrophage (CD14, CD64), NK cell (CD56+) and activation markers (CD11b+, CD25+, CD28+, CD80+, CD127+, CD152+, CD278+). All samples will be acquired by flow cytometry and analyzed according to the relative fluorescence intensity using Flowjo Software. In the fasting and postprandial states, cell proliferation will be measured using alamar blue fluorescence and intracellular cytokines accumulation in different immune cell types will be determined using an intracellular staining flow. | 6 months | 
| Effect of diets on inflammatory markers (C-reactive protein (CRP)) by comparing groups at 3 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 3 months). | Assess levels of acute phase protein (CRP) collecting blood samples in a fasting state. CRP will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 3 months | 
| Effect of diets on inflammatory markers (C-reactive protein (CRP)) by comparing groups at 6 months of intervention (i.e. control low fat diet and the 2 keto groups) and changes over time within each group (i.e. baseline vs. 6 months). | Assess levels of acute phase protein (CRP) collecting blood samples in a fasting state. CRP will be determined using commercially available equipment at a local laboratory (Roche Cobas c503). Measurements will be performed for between group comparison and within groups comparison to understand the effect of diets. | 6 months |