Plasenta Sferoid (Mikro Doku) Modelinde Vortioksetin Etkisinin Araştırılması

Abstract

Maternal-placental transmission of nutrients during pregnancy, maternal nutrition and metabolism, placental transition gradient, utero-placental and feto-placental blood flow, structure, size and transfer capacity of the placental barrier are determined. In recent years, there has been a great increase in the use of chemicals and drugs during pregnancy, but due to ethical restrictions, the need for current placental barrier model studies has increased as tests cannot be performed on humans. Within the scope of this thesis, mono and co-cultures of human umbilical cord vein cells (HUVEC) together with human placental choriocarcinoma cells (BeWo) were made and placental tissue was modeled in two and three dimensional systems. Spheroids were created with three dimensional (3D) Petri Dish® molds in a scaffold-free environment and cell localizations were characterized by fluorescent dyes and diameter measurements. Vortioxetine, the active ingredient of the SSRI type multimodal drug, was exposed to the cultures formed at 7.5, 15 and 30 μM concentrations for 24, 48 and 72 hours. At the end of the incubation, cell viability analyzes (MTS, Calcein-PI-DAPI) were performed. Barrier integrity was measured by transepithelial electrical resistance measurement (TEER) and sodium fluorescence (Na- iv F) transition analysis. The localizations of epithelial (E)-cadherin, filamentous (F)-actin and serotonin transporter (SERT) protein from cytoskeletal elements, which are the connecting proteins involved in the structure of the placental barrier, were demonstrated by immunofluorescence staining. It was determined by measuring changes in the amount of human chorionic gonadotropin (β-hCG) hormone production. Cell viability decreased in 2D groups due to increasing doses of vortioxetine. In the 3D micro-tissues, on the other hand, the dose groups showed resistance to the toxic effect of Vortioxetine and the viability was higher than the control groups. While forming spheroids, diameter length and area increased proportionally with incubation time for BeWo, HUVEC and co-culture groups. Also, cell localization of spheroids was determined by CellTracker staining. Qualitative and quantitative measurements were obtained by comparing calcein-PI fluorescent imaging and Vortioxetine applications with MTS viability analysis. E-cadherin, F-actin and SERT immunofluorescent dyes were applied in BeWo cells, and it was observed that intercellular integrity was impaired, the structure of intracellular skeletal elements was changed, and SERT receptor expression decreased depending on the incubation period at increasing doses. At the same time, the amount of 2D and 3D β-hCG hormones was measured in the BeWo and co-culture groups, and a higher amount of hormone production was observed in the 3D culture groups. This thesis study presents an evaluation in terms of mono and co-cultures by comparing the placental toxicity of vortioxetine, the active ingredient of the antidepressant drug Brintellix, the response of the barrier structure, hormone production in 2D and 3D systems.