Ectopic and eutopic stromal endometriotic cells have a damaged ceramide signaling pathway to apoptosis

Abstract

Objective: To investigate whether sphingosine analogues, which activate the ceramide signaling pathway to apoptosis, can cause the death of ectopic (EEC) and eutopic stromal endometriotic cells (EEU), as well as healthy eutopic stromal endometrial cells (HEU).

Design: The EEC, EEU, and HEU isolated from fertile and infertile women with endometriosis were cultured for 48 hours in RPMI medium with 10% fetal calf serum (FCS) and with 2.5-10 microM sphingosine analogues.

Setting: A clinic for the treatment of endometriosis and basic research laboratories.

Patient(s): Nineteen women with follicular cyst and 16 women with endometriosis.

Main outcome measure(s): The percentage of proliferating cells was determined by 93-(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (MTT) assay. Apoptosis and cell cycle were detected by fluorescence-activated cell sorter (FACS) Calibur flow cytometer.

Result(s): The viability of EEC after exposure to 10 microM sphingosine analogues was 59.5% +/- 9.7% for D-sphingosine and 77.65 +/- 9.7% for DL-erythro-sphingosine, the viability of EEU was 69.2% +/- 14.2% and 42.0% +/- 15.5%, whereas the viability of comparative HEU was 9.0% +/- 4.8% and 18.8% +/- 8.3%, respectively. The differences were significant using the Mann-Whitney test. The apoptotic level of the cells treated with 10 microM sphingosine analogues for comparative HEU was 42.8% +/- 7.5% for D-sphingosine and 42.5% +/- 10.5% for DL-erythro-sphingosine, whereas for EEC this was 16.7% +/- 5.5% for D-sphingosine and 14.1% +/- 4.4% for DL-erythro-sphingosine and for EEU this was 14.3% +/- 4.7% and 22.9% +/- 8.9%, respectively.

Conclusion(s): Ectopic and eutopic stromal endometrial cells from women with endometriosis have a damaged ceramide-downstream pathway to apoptosis.