ALTEX :: Alternatives to Animal Experiments

2001, VOLUME 1

Establishment and Characterisation of an Immortalised Human Sebaceous Gland Cell Line (SZ95)*

Holger Seltmann¹, Heidemarie Neitzel², Constantin E. Orfanos¹ and Christos C. Zouboulis¹

¹Department of Dermatology, University Medical Center Benjamin Franklin, The Free University of D-Berlin; ²Institute of Human Genetics, Virchow Clinic, Humboldt University, D-Berlin

* This poster got a prize at the alternatives congress Linz 2000

SUMMARY

There is increasing evidence that sebocytes may play critical roles in the pathophysiologic processes and disorders of the pilosebaceous unit, especially in acne. To date, much of our understanding of the physiology and pathophysiology of the sebaceous gland stems from experimental animal models but no animal model was found predictive in assessing the effects of anti-acne drugs in human beings. The facts that acne is an exclusively human disease and that the secretory activity of the sebaceous gland is remarkably species-specific led to the search for human models.

Therefore, 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 100 times to date, have been cloned, and show no signs of senescence after 5 years in vitro, whereas normal human sebocytes can only be grown for three to 6 passages. The immortalized transfected cells, termed SZ95, expressed the Simian virus-40 large T antigen and exhibited epithelial, polymorphous characteristics with different cell sizes of up to 3?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, 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 to 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 5-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.