Fecha de publicación: -- Fuente: Digital CSIC Título: The OLTRAD Study (I): Clinical efficacy
Autor: Matamoros-Domínguez, Alejandro; Espinosa-Cabello, Juan M.; García-González, Aída; León-Aranda, Manuel Jesús; Pérez-Muñoz, Gisela; Jiménez-Sánchez, Andrés; Martínez-Ortega, Antonio Jesús; García-Rey, Silvia; Roque-Cuellar, María Del Carmen; Piñar-Gutiérrez, Ana; Ruíz-Trillo, Carmen Amelia; Mangas-Cruz, Miguel Ángel; Pereira-Cunill, José Luis; Cerrillo, Isabel; Yubero-Serrano, Elena María; Perona, Javier S.; García-Luna, Pedro Pablo; Castellano, José María
Descripción: [Description of methods used for collection/generation of data]
Design overview
The OLTRAD study was a randomized, parallel-group, double-blinded, and placebo-controlled superiority trial conducted at the Endocrinology and Nutrition Service of the Virgen del Roc o University Hospital (Seville, Spain) between February 2022 and March 2025. The trial was designed to demonstrate that the regular intake of a functional olive oil enriched in OA may be effective as an adjuvant to the pharmacological therapy with metformin and other conventional antidiabetic drugs. Participants were randomly assigned to one of two study groups: OA-enriched olive oil (intervention group) or the same olive oil not reinforced with the triterpene (control oil) for a 12-month follow-up period.
Settings and Participants
Eligible participants complied with all the following criteria: 1) community-residing men and women aged 18-75 years; 2) Body Mass Index (BMI) between 25.0 and 39.9 kg/m2; 3) diagnosed with T2DM at least six months before enrollment [following the American Diabetes Association (ADA) 2019 criteria]; 4) treated with metformin (stable dose ? 850 mg/day at least three months before recruitment) as monotherapy or in combination with other hypoglycemic agents, except pioglitazone and sulphonylureas; 5) Glycated hemoglobin (HbA1c) 3 times upper limit of reference range; 4) food allergies or intolerances relevant to the intervention; 5) pregnancy or lactation; 6) women of childbearing potential unwilling to use effective contraception; 7) fasting triglycerides >600 mg/dL despite treatment; 8) grade 3 hypertension [systolic blood pressure (SBP)?180 mmHg and/or diastolic blood pressure (DBP) ?110 mmHg]; 9) use of pioglitazone or sulfonylureas; 10) use of weight-loss medications other than GLP-1RA, provided that the GLP-1RA indication was a T2DM diagnosis, the drug use was chronic (defined as 12 months or more prior to enrollment), and the participant's weight remained stable (defined as body weight variations of up to 5% in the 6 months prior to enrollment); 11) chronic systemic corticosteroid use (>14 days) in the previous 6 months (excluding topical/inhaled forms); 12) major cardiovascular events within the past 6 months (e.g., myocardial infarction, unstable angina, NYHA class III IV heart failure, stroke); 13) uncontrolled endocrine disorders (e.g., thyrotoxicosis, adrenal crisis); 14) history of an active or untreated malignancy, or are in remission from a clinically significant malignancy (other than basal or squamous cell skin cancer, in situ carcinomas of the cervix, or in situ prostate cancer) during the last 5 years prior to study entry; 15) participation in another clinical trial in the past 30 days (or longer if prior investigational drug had a long half-life); 16) concurrent enrollment in any other interventional clinical study deemed incompatible by the investigators; 17) hematologic conditions interfering with HbA1c measurement (e.g., hemolytic anemia, sickle cell disease); and 18) any other medical, psychiatric, or substance-related condition that, in the investigator s opinion, could interfere with study adherence or completion.
Intervention and follow-up
At the time of recruitment and quarterly thereafter, participants were subjected to anthropometric, blood pressure, and heart rate measurements. They were also asked about their perception of their own health status, and fill in a questionnaire about lifestyle, medical conditions and use of medication. In addition, body composition analysis was carried out by electrical bioimpedance (Tanita BC 420 S MA; West Drayton, UK). Also, at baseline and every three months afterwards, fasting blood samples were withdrawn from the median cubital vein and collected in sterile plastic tubes with vacuum system. Conventional analytical determinations in blood and urine were performed in the Laboratory of Analysis and Clinical Biochemistry of the Virgen del Roc o University Hospital of Seville following standard operating procedures.
Outcomes
Anthropometric measurements and clinical examination
Participants underwent a comprehensive clinical evaluation including measurement of anthropometric parameters (BMI, height and waist circumference), pulse and blood pressure (SBP and DBP). In addition, body composition analysis was carried out by electrical bioimpedance, and Fat Mass Index (FMI) was calculated as total fat mass (kg) divided by the square of height (m ) and expressed in kg/m .
Blood biochemistry
In samples of cubital blood we realized cell count and determinations related to circulating lipids [total triglycerides (TG), total cholesterol (TC), low density lipoprotein-associated cholesterol (LDL-c), high density lipoprotein-associated cholesterol (HDL-c), total apolipoprotein B]; glucose homeostasis [glycemia, insulinemia, C peptide, glycated haemoglobin (HbA1c)]; oxidative stress and inflammation [ultra-sensitive C-reactive protein], as well as associated to liver and kidney injury [hepatic transaminases (ALT, AST, GGT), and creatinine]. Serum creatinine (sCr)-based estimated glomerular filtration rate (eGFR) was calculated using the CKD-Epi (chronic kidney disease epidemiology collaboration) equation.
Medication use
Concomitant medication, including metformin dosage, as well as the use of GLP-1 receptor agonists and sodium-glucose cotransporter-2 (SGLT2) inhibitors, was obtained from the participants' electronic medical records and self-completed questionnaires, reflecting the individualized pharmacological management of each of them.
Determination of serum OA concentration
The identification and quantification of OA in the serum were performed using a gas chromatography mass spectrometry (GC-MS) system, following a modification of the method previously established and validated in our laboratory [DOI 10.1002/bmc.3480]. Briefly, 15 ?L of betulinic acid (internal standard, 0.1 mg/mL in methanol) were added to 1 mL of serum. After the methanol removal by vacuum, the sample was filtered (0.20 ?m) and extracted with 3.0 mL of 5% isopropyl alcohol in diethyl ether (3 min vortexing and centrifugation at 1620 g, 10 min at 20 0C; repeated 3x). The combined supernatant was dried, and the residue was resuspended and derivatized with BSTFA [N,O-bis-(trimethylsilyl) trifluoroacetamide +1% TMCS (trimethylchlorosilane)] in pyridine (1:1 v/v), assisted by ultrasound (70 W; 40 C, 15 min). A 1 ?L aliquot of the derivatized sample was then injected and analyzed by GC-MS.; [Methods for processing the data]
All statistical analyses were performed using JASP version 0.98.1 (JASP Team, University of Amsterdam, Amsterdam, Netherlands). Continuous variables following a normal distribution were expressed as mean standard deviation, whereas those showing a non-normal, skewed distribution were expressed as the median and interquartile range (25th and 75th percentiles). Categorical variables are expressed as absolute and relative frequencies. Initial comparisons between groups were conducted using Student's t-test or the Mann-Whitney U test for continuous variables, depending on the data distribution, and Fisher's exact test for categorical variables.
The primary outcome for each clinical parameter was defined as the 12-month change from baseline (? = T12 ? T0). The effect of treatment on each ? variable was assessed using linear regression-based analysis of covariance (ANCOVA), adjusted for baseline value, age and sex. Normality of residuals and homoscedasticity were assessed for each model using Shapiro-Wilk and Levene's tests, respectively; both assumptions were adequately met across all models, and no non-parametric alternatives were required.
In addition to the primary analysis, exploratory post-hoc subgroup analyses examined whether the treatment effect was modified by baseline metformin dose, fat mass index (FMI), or concomitant use of GLP-1 RA and iSGLT2, through four separate interaction models, each incorporating a group effect-modifier term alongside the baseline covariates:
Model 1 (metformin dose): ? variable as the dependent variable; baseline value, age, sex, and continuous BMI as covariates; treatment group, metformin dose (