Several studies have demonstrated that bariatric surgery is effective for inducing weight loss in obese patients. In addition, the effects of this surgery on multiple associated alterations are well known, including changes in the secretion and activity of hormones involved in appetite regulation, satiety, and energy expenditure, as well as alterations in the gut microbiota composition.

However, in cases of severe obesity, recent data have challenged the prevailing view, as bacterial species associated with low microbial richness (prior to surgery) appear to change only marginally after bariatric surgery, despite significant metabolic improvements.

Our objective is to examine whether gut microbiota and gastrointestinal peptides are further impaired in severe obesity and, additionally, to explore how the microbiota relates to metabolic profile or sex, as well as whether bariatric surgery may differentially correct obesity-related intestinal microbial features.

To this end, we propose a prospective, interventional, translational clinical study involving a cohort of 60 obese patients (BMI > 35 kg/m²) undergoing laparoscopic gastric bypass surgery. Patients will be grouped according to their degree of obesity to assess potential baseline differences and to evaluate the efficacy of the intervention. Furthermore, we will investigate whether these parameters differ according to metabolic profile or sex.

Body composition and nutritional status will be assessed, along with cardiovascular risk factors and comorbidities (hypertension, obstructive sleep apnea syndrome, dyslipidemia, type 2 diabetes mellitus, and insulin resistance). Gastrointestinal hormones (ghrelin, GIP, GLP-1, PYY, CCK, and leptin) will be measured in serum using Luminex XMAP technology. The content and diversity of the gut microbiota will be analyzed (16S rRNA amplicon sequencing and shotgun metagenomic sequencing using Illumina MiSeq technology) in stool samples collected before and 6-12 months after surgery. Additionally, individualized dietary follow-up and assessment of participants' quality of life will be conducted.

Obesity and type 2 diabetes mellitus (T2DM) continue to increase worldwide and are strongly associated with adverse metabolic and cardiovascular outcomes. Bariatric surgery, particularly laparoscopic gastric bypass (LGB), remains the most effective therapeutic strategy for substantial weight reduction, glycemic improvement, and favorable modulation of gastrointestinal hormones, bile acids, and gut microbiota. Nevertheless, the mechanisms explaining heterogeneous metabolic responses-especially in individuals with severe, morbid, or extreme obesity-remain insufficiently elucidated. In particular, gut microbial diversity, microbial gene richness, and related inflammatory pathways appear to contribute to metabolic health, yet their behavior in severe obesity and their evolution following bariatric surgery remain poorly characterized.

To address these gaps, the investigators designed a prospective, interventional, comparative, and translational clinical study including 60 obese individuals scheduled for bariatric surgery at a single tertiary hospital. Participants will be consecutively recruited according to strict inclusion and exclusion criteria and classified by degree of adiposity (severe, morbid, or extreme obesity), metabolic phenotype (metabolically healthy or metabolically abnormal obesity), and sex.

Prior to the surgical intervention, a structured dietary protocol will be implemented. Participants will receive an individualized hypocaloric diet tailored to nutritional requirements, with a macronutrient distribution of 55% carbohydrates, 30% fats, and 15% proteins. During the two weeks preceding surgery, a very low-calorie diet based on Optisource® sachets will be administered in three daily intakes. Postoperative dietary progression will follow standardized clinical guidelines, with close monitoring to support weight reduction and metabolic stabilization. Follow-up evaluations will occur at 1, 6, and 12 months after surgery.

All participants will undergo comprehensive baseline and longitudinal assessments, including: evaluation of nutritional status; anthropometric measurements; body composition and basal metabolism; cardiovascular risk factors; metabolic comorbidities; and exclusion of secondary causes of obesity. Analytical parameters will include fasting plasma glucose, lipid profile (triglycerides, HDL-cholesterol), blood pressure, markers of hepatic and renal function, and additional biochemical indices relevant to obesity and T2DM risk stratification. Gut microbiota profiling will be conducted to evaluate microbial composition, diversity, and gene richness. Gastrointestinal hormone analyses will quantify circulating incretins and appetite-regulating peptides, including GLP-1, PYY, and ghrelin, to explore their association with postoperative metabolic outcomes. Oxidative stress and inflammatory parameters will be also assessed in order to stratify cardiovascular risk before and after induced weight loss after bariatric surgery.

Statistical analyses will be performed using SPSS 17.0. Categorical variables will be summarized with frequency distributions and percentages. Quantitative variables will be described using mean, range, and standard deviation, with normality assessed via the Kolmogorov-Smirnov test. Between-group comparisons (sex, metabolic phenotype) will employ unpaired t-tests or Mann-Whitney U tests; comparisons across obesity categories will use ANOVA. Longitudinal changes following surgery will be evaluated with repeated measures ANOVA or paired t-tests. Correlations will be analyzed using Pearson or Spearman methods, and logistic regression models will be developed to identify predictive factors for specific metabolic responses.

This study aims to determine whether gut microbiota alterations are directly linked to metabolic abnormalities across obesity phenotypes and whether bariatric surgery elicits differential microbial and hormonal responses according to degree of adiposity and sex. It further explores whether gastrointestinal peptides and other peripheral biomarkers can refine obesity phenotyping and enhance individualized clinical management.

Inclusion Criteria:

• Patients with a body mass index (BMI) greater than 40 kg/m² (or > 35 kg/m² with minimum one obesity comorbidity).
• Aged between 18 and 65 years.
• Patients with a known duration of obesity exceeding five years, despite dietary interventions and farmacological treatment.

Exclusion Criteria:

• Due to the nature of the study, patients with acute or chronic inflammatory diseases, established hepatic or renal insufficiency (defined as transaminase levels ±2 SD from the mean and estimated glomerular filtration rate [CKD-EPI formula] >60), neoplasic diseases, or secondary causes of obesity (e.g., hypothyroidism, Cushing's syndrome) will be excluded.