Protease chymotrypsin mediates the endothelial expression of P- and E-selectin, but not ICAM and VCAM, induced by placental trophoblasts from pre-eclamptic pregnancies.

OBJECTIVES: Soluble endothelial-cell adhesion molecules (ICAM, VCAM and PECAM) are markers of endothelial activation, and are elevated in the maternal circulation during pregnancy and even further increased in pregnancies complicated by pre-eclampsia (PE). To identify possible sources of endothelial activators during pregnancy, we addressed whether factors released from placental trophoblast cells (TCs) activate endothelial cells (ECs) to enhance adhesion molecule expression on ECs. We also examined whether proteases released by placental cells induce the endothelial cell surface molecule expression in PE.

METHODS: Confluent ECs were co-cultured with placental TCs derived from normal (n=9) or PE (n=8) pregnancies or with placental conditioned media (CM) derived from PE placental cultures (n=7). ICAM, VCAM, P-selectin and E-selectin were quantified using an enzyme-linked immunosorbent assay (ELISA). The protease inhibitors alpha(2)-macroglobulin (alpha(2)M), thrombin inhibitor (TI) and chymotrypsin inhibitor (CI) were tested in the co-culture system. mRNAs for ICAM, VCAM, P-selectin and E-selectin were determined by RNase protection assay (RPA). NF-kappaB activity in ECs was also determined.

RESULTS: (1) ICAM and VCAM expression was significantly increased on ECs co-cultured with both normal-TCs and PE-TCs, compared to control ECs (P<0.01). ICAM and VCAM expression in ECs co-cultured with normal-TCs did not differ from ECs co-cultured with PE-TCs. (2) E-selectin expression was increased on ECs co-cultured with normal-TCs (P<0.05) and further increased in ECs co-cultured with PE-TCs (P<0.01). (3) P-selectin expression was increased on ECs co-cultured with PE-TCs, but not ECs co-cultured with normal-TCs compared to control ECs (P<0.05). (4) alpha(2)M and TI did not alter the ICAM, VCAM, P-selectin and E-selectin expression on ECs induced by PE-CM. (5) CI blocked the upregulation of P-selectin and E-selectin (P<0.05), but not ICAM and VCAM expression, in ECs cultured with PE-CM. (6) Changes in mRNA for ICAM, VCAM, P-selectin and E-selectin paralleled the increases in protein expression on ECs cultured with PE-CM. (7) NF-kappaB activity was also increased in cells challenged with PE-CM.

CONCLUSIONS: (1) Factor(s) released from both normal-TCs and PE-TCs promote ICAM and VCAM expression on ECs. (2) Factor(s) released from PE-TCs significantly increase EC P-selectin and E-selectin expression. (3) CI blocks the upregulation of P-selectin and E-selectin on ECs induced by factors released from PE placental cells, suggesting that chymotrypsin is responsible for the increased endothelial expression of P-selectin and E-selectin in pre-eclampsia.