Human facial sebaceous gland cells were transfected with a PBR-322-based plasmid containing the coding region for the Simian virus-40 large T antigen. The resulting proliferating cell cultures have been passaged over 50 times to date, have been cloned, and show no signs of senescence after 4&DF;1 2 y in vitro, whereas normal human sebocytes can only be grown for three to six passages. The immortalized transfected cells, termed SZ95, expressed the Simian virus-40 large T antigen and presented an hyper-diploid-aneuploid karyotype with a modal chromosome number of 64.5. The SZ95 cell line exhibited epithelial, polymorphous characteristics with different cell sizes of up to 3.25-fold during proliferation and 6-fold at confluence, showing numerous cytoplasmic lipid droplets. The cells showed large cytoplasm profiles with abundant organelles, including vacuoles and myelin figures which indicated lipid synthesis. Lack of or only few desmosomal areas were observed. SZ95 cells expressed molecules typically associated with human sebocytes, such as keratins 7, 13, and 19, and several proteins of the polymorphous epithelial mucin family. Functional studies revealed synthesis of the sebaceous lipids squalene and wax esters as well as of triglycerides and free fatty acids, even after 25-40 passages; active lipid secretion; population doubling times of 52.4 +/- 1.6 h; reduced growth but maintenance of lipid synthesis under serum-free conditions; and retrieval of cell proliferation after addition of 5alpha-dihydrotestosterone. Retinoids significantly inhibited proliferation of certain SZ95 cell clones in the expected magnitude 13-cis-retinoic acid > all-trans-retinoic acid > > acitretin. Thus SZ95 is an immortalized human sebaceous gland cell line that shows the morphologic, phenotypic and functional characteristics of normal human sebocytes.