Obesity is a global health problem that has reached epidemic proportions, affecting more than one billion people worldwide and significantly increasing the risk of multiple comorbidities, including type 2 diabetes, cardiovascular diseases, and cancer (World Health Organization, 2024). Increasing evidence suggests that chronic low-grade inflammation associated with obesity plays a critical role in the development of obesity-related malignancies, including gastric cancer. Adipose tissue dysfunction in obesity leads to the recruitment and activation of various immune cells, such as macrophages and mast cells, which contribute to a pro-inflammatory microenvironment through the release of cytokines, growth factors, and angiogenic mediators.
Adult patients undergoing bariatric surgery (laparoscopic sleeve gastrectomy).
BMI > 35 kg/m²
All participants underwent preoperative evaluation, including blood tests and assessment by a multidisciplinary team (nutritionist, psychiatrist, endocrinologist, radiologist, anesthesiologist, and surgeon).
Patients with secondary causes of obesity, such as Cushing's syndrome or polycystic ovary syndrome (PCOS).
Patients with malignant gastric conditions or previous gastric surgery.
Patients with systemic inflammatory diseases, autoimmune disorders, or chronic infections that may influence immune cell infiltration.
Patients with incomplete clinical data or poor-quality tissue samples.
Fifty gastric tissue samples will be collected from patients with obesity undergoing bariatric surgery and fifty from lean control patients undergoing endoscopic biopsy for benign or malignant gastric conditions. All participants underwent preoperative evaluation including blood tests. All patients were evaluated by a multidisciplinary team consisting of a nutritionist, psychiatrist, endocrinologist, radiologist, anesthesiologist, and surgeon. Endocrinologic assessment excluded secondary causes of obesity (e.g., Cushing's syndrome, polycystic ovary syndrome). Only patients with a body mass index (BMI) \> 35 kg/m² were included. All surgical procedures were laparoscopic sleeve gastrectomies (LapSG).
Cohort
Prospective
bariatric surgery laparoscopic sleeve gastrectomy
Procedure/SurgeryAdipose tissue macrophages (ATMs), mast cells positive for tryptase (MCPT), and microvascular density (MVD) were assessed by immunohistochemistry. Quantitative assessment was performed using a light microscope. For each GTO and NT tissue section, five highly immunostained areas ("hot spots") were identified at low magnification.
lean control patients undergoing endoscopic biopsy for benign or malignant gastric conditions.
Procedure/SurgeryAdipose tissue macrophages (ATMs), mast cells positive for tryptase (MCPT), and microvascular density (MVD) were assessed by immunohistochemistry. Quantitative assessment was performed using a light microscope. For each GTO and NT tissue section, five highly immunostained areas ("hot spots") were identified at low magnification.
Immunohistochemistry
Compare the 2 groups: Adipose tissue macrophages (ATMs) was assessed by immunohistochemistry using a three-step biotin-avidin-peroxidase detection method. five-micrometer-thick serial sections were cut from formalin-fixed, paraffin-embedded gastric tissue of obese (GTO) and control normal tissue (NT) samples. Antigen retrieval was performed using a microwave oven (500 W for 10 minutes), followed by endogenous peroxidase blocking with a 3% hydrogen peroxide solution. Slides were incubated with the following primary antibodies for 1 hour at room temperature: * Anti-tryptase (clone AA1; Dako, Glostrup, Denmark; 1:100) for mast cell identification, * Anti-CD68 (clone KP1; Dako, Glostrup, Denmark; 1:100) for ATM detection, * Anti-CD31 (clone QB-END 10; Bio-Optica, Milan, Italy; 1:50) as a pan-endothelial marker to assess MVD.
Immunohistochemistry
Compare the 2 groups: Mast cells positive for tryptase (MCPT) was assessed by immunohistochemistry using a three-step biotin-avidin-peroxidase detection method. five-micrometer-thick serial sections were cut from formalin-fixed, paraffin-embedded gastric tissue of obese (GTO) and control normal tissue (NT) samples. Antigen retrieval was performed using a microwave oven (500 W for 10 minutes), followed by endogenous peroxidase blocking with a 3% hydrogen peroxide solution. Slides were incubated with the following primary antibodies for 1 hour at room temperature: * Anti-tryptase (clone AA1; Dako, Glostrup, Denmark; 1:100) for mast cell identification, * Anti-CD68 (clone KP1; Dako, Glostrup, Denmark; 1:100) for ATM detection, * Anti-CD31 (clone QB-END 10; Bio-Optica, Milan, Italy; 1:50) as a pan-endothelial marker to assess MVD.
Immunohistochemistry
Compare the 2 groups: Microvascular density (MVD) was assessed by immunohistochemistry using a three-step biotin-avidin-peroxidase detection method. five-micrometer-thick serial sections were cut from formalin-fixed, paraffin-embedded gastric tissue of obese (GTO) and control normal tissue (NT) samples. Antigen retrieval was performed using a microwave oven (500 W for 10 minutes), followed by endogenous peroxidase blocking with a 3% hydrogen peroxide solution. Slides were incubated with the following primary antibodies for 1 hour at room temperature: * Anti-tryptase (clone AA1; Dako, Glostrup, Denmark; 1:100) for mast cell identification, * Anti-CD68 (clone KP1; Dako, Glostrup, Denmark; 1:100) for ATM detection, * Anti-CD31 (clone QB-END 10; Bio-Optica, Milan, Italy; 1:50) as a pan-endothelial marker to assess MVD.
Morphometric Analysis
Compare the 2 groups: Quantitative assessment was performed using a light microscope. For GTO tissue section, five highly immunostained areas ("hot spots") were identified at low magnification. ATMs, MCPT, and MVD were then quantified at ×40 magnification. The mean value across five hot spots per marker was used for each sample. To evaluate mast cell degranulation, the presence of tryptase-positive granules in the extracellular matrix was examined. Degranulating mast cells were identified by the diffusion of immunoreactive granules outside the cell boundaries, indicating active release of tryptase. This extracellular localization of tryptase provided morphological evidence of mast cell activation and was considered a marker of tissue inflammation and remodeling.
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