Primary culture of human omental preadipocytes and study of their biological properties

Abstract

To develop a primary culture method of human omental preadipocytes and to study their biological properties, such as hyperplasia, hypertrophy and endocrine secretion of human visceral adipose tissue. Using enzyme-digesting method, fibroblast-like cells from the human omental adipose tissues were cultured. The morphological changes of the cultured cells were observed and the growth curve was drawn by MTT method. The intracytoplasmic lipid of the cultured cells was determined by oil red O staining. The leptin and adiponectin levels in the culture supernatants were measured by ELISA. The cultured fibroblast-like cells were homogeneous. Proliferation of cells began at the 3 rd day and the cell numbers increased in indicial way from the 3 rd day to the 9 th day. The doubling time of cells was about 60 hours. During the process of induction by conditional medium, the cells became round and larger, and more adipose droplets were aggregated. On the 21 st day, more than 90% of the cells became adipocytes. Leptin secretion was detected at low level in the preadipocytes and continuously increased during differentiation, with a peak on day 17. It remained constant from day 17 onward. Unlike leptin, adiponectin secretion was not detected until day 7 after induction, when differentiated adipocytes had already been observed. Its secretion increased dramatically between days 7 and 17, and reached a maximum level on day 17, but had a significant reduction on day 21. Extraction of intracytoplasmic lipid stained with oil red O and detection of leptin and adiponectin both verified the isolated cells were preadipocytes functioning actively. A human omental preadipocytes model has been established and different secretion patterns of leptin and adiponectin secretion related to preadipocyte differentiation has been characterized. Adiponectin may be proposed as a specific marker for preadipocyte differentiation.