Comparative Analysis of Small Extracellular Vesicles in Peritoneal Fluid from Endometriosis and Non-Endometriosis Patients: A Pilot Case-Control Study

Aim: Endometriosis is a chronic inflammatory disorder requiring clinical examination, imaging, and surgical intervention for accurate assessment. The aim of the present study is to identify unique biochemical and biophysical patterns in small extracellular vesicles (sEVs) for advancing innovative approaches in both disease detection and targeted therapy. Methods: sEVs were isolated from the peritoneal fluid of women with endometriosis (n = 7), women without endometriosis who underwent surgery for benign, noninflammatory conditions (n = 7), and follicular fluid samples (n = 3). The tetraspanin profile of the samples was determined using single-particle interferometric reflectance imaging sensing. After size exclusion chromatography and ultracentrifugation, atomic force microscopy (AFM) and Raman spectroscopy were employed to characterize the sEVs. Results: Prominent Raman spectral variations in peritoneal sEV between healthy women and those with endometriosis occur at 891 cm-1 (proteins) and 968 cm-1 (lipids, CH wagging). Statistical analysis of molecular components influencing the Raman signature of sEVs confirms that their lipid composition may be shaped by the pathophysiology of endometriosis, reflecting changes similar to those observed in the eutopic endometrium in previous studies. sEVs isolated from peritoneal fluid and follicular fluid have markedly distinct biochemical compositions and morphometric properties. sEVs from follicular fluid were significantly smaller in size and exhibited a higher rigidity. Conclusion: Distinct differences in sEVs from endometriosis patients highlight their potential as biomarkers, while variations between follicular and peritoneal fluid suggest unique functional roles in reproductive health. These insights provide potential targets for future investigations and advancements in disease understanding.