Efecto de la LDL-oxidada en la hiperplasia prostática benigna: rol de las vesículas extracelulares en la modulación de la proliferación celular

Abstract

Benign Prostatic Hyperplasia (BPH) is characterized by the uncontrolled proliferation of mainly stromal elements, the result of the conjunction of multiple physiopathogenic factors. Evidence supports that pro-atherogenic environment could contribute to the development and progression of BPH. Likewise, the hyperproliferative state of BPH requires coordinated communication between cellular components that maintain a permissive microenvironment. In this sense, we evaluated in vitro the effect of the pro-atherogenic state, simulated by oxidized-LDL (OxLDL), on human prostatic stromal cells (HPSC) and the role of extracellular vesicles (EVs) in the modulation of cell proliferation.

For this purpose, primary HPSC cultures were established from patient samples (n=4) collected at the Sanatorio Allende in the city of Córdoba, with the approval of the Sanatorio Allende Bioethics Committee. All the tests performed were carried out in triplicate and the Student's t statistical test was applied for the analysis of results. The HPSC were stimulated for 24h with OxLDL [20μg/mL] and it was observed by BrdU incorporation and Ki-67 immunocytochemistry that OxLDL significantly increased the proliferation rate vs. vehicle in HPSC (p<0.001). Likewise, EVs derived from HPSC exposed to OxLDL were isolated by differential ultracentrifugation (150K pellet).

After analysis and quantification by transmission electron microscopy, along with EVs identity confirmation by CD63 immunostaining conjugated with colloidal gold, it was determined that OxLDL increased the release of EVs by a factor of 10; particularly within the 15-60 nm fraction corresponding to exosomes vs. control (p<0.001). Also, the roll of EVs in cell proliferation was studied using as a stimulus the EVs obtained from HPSC previously treated with OxLDL [20μg/mL] for 24h vs. control, on both HPSC and epithelial cells of the PC3 line. Subsequently, the autocrine and paracrine effects of EVs-induced proliferation were assessed. HPSC-derived EVs in pro-atherogenic contexts significantly increased proliferation in HPSC and PC3, as determined by Ki-67 (p<0.01).

These findings confirm the pathogenic effect of OxLDL on the abnormal growth of the prostate, promoting a greater release of EVs and an increase in the cell proliferation rate.