Human ovarian granulosa cells use clathrin-mediated endocytosis for LDL uptake: immunocytochemical and electron microscopic study

Abstract

ABSTRACT The steroidogenic activity of the granulosa cells is important for the reproductive cycle, and lipoproteins are involved in this process. The clathrin-mediated endocytosis pathway for LDL transport is considered to be the main one in eukaryotic cells. However, there are no studies that elucidate LDL internalization in human granulosa cells clarifying whether the clathrin-mediated endocytic pathway is functional in this process. The aim of this study is to investigate the role of clathrin and v-SNARE proteins in the formation of vesicles in human granulosa cells. In this study, the COV434 human granulosa cells were cultured and divided into four groups where in some of the groups Dil-conjugated LDL and Icarugamycin (ICA) a clathrin-mediated endocytosis inhibitor were added. From the collected mediums pregnenolone and progesterone levels were measured using ELISA. Oil red O staining was performed to show the intracellular lipids in the cells. Clathrin-coated vesicles believed to be responsible for carrying LDL, and v-SNARE proteins that direct the vesicles to their target molecules were also labeled and investigated by histological and ultrastructural methods. Our results show that human granulosa cells as well use the LDL cholesterol for steroid biosynthesis and they may prefer the clathrin-mediated endocytotic pathway to internalize it.