Smooth muscle cells contain a factor responsible for decreasing big endothelin and endothelin-1 produced by cultured endothelial cells.
The production of big endothelin-1 (big ET-1) and its processing to endothelin (ET-1) were examined in cocultures of bovine pulmonary artery endothelial cells (BPECs) and human bronchiolar smooth muscle cells (HBSMCs). The BPECs produced immunologically reactive ET-1 (ir-ET-1) and big ET-1 (ir-big ET-1) that increased linearly in the cell culture media over a 72 h time course when confluent monolayers were incubated at 37 degrees C in serum-free media. HBSMCs maintained in serum-free media over a 72-h period did not produce any detectable ir-big ET-1 or ir-ET-1. After coculture of these two cell types for 24 h (the monolayers were physically separated), the culture medium contained no detectable ir-big ET-1 or ir-ET-1. When medium conditioned for 24 h with the HBSMCs was transferred to the BPEC monolayers, the accumulation rates of ET-1 and big ET-1 were attenuated 70 and 10%, respectively. When medium conditioned for 24 h with the BPECs was transferred to the HBSM cell monolayers, the amount of ir-ET-1 and ir-big ET-1 in the medium decreased greater than 90% over the next 24 h. Cellular binding and uptake could not account for this decrease as ascertained by quantitation of cellular levels of ET-1 and big ET-1. When conditioned medium from the HBSMCs was added to conditioned medium from the BPECs in a 1:1, 2:1, 3:1, 1:2, or 1:3 ratio (v/v) in the absence of cells, the amount of ET-1 or big ET-1 present in the EC medium did not decrease significantly over the following 24 h. These data imply that HBSMCs contain a factor that is responsible for decreasing the accumulation of ir-big ET-1 and ir-ET-1 in the cell culture media of BPECs. The data also suggest that the most likely mechanism for this effect is a factor that degrades BET-1 and ET-1 at different rates.
Cade
C
C
Department of Pharmacology, Berlex Laboratories, Cedar Knolls, New Jersey 07927.
Ilozue
C V
CV
Rubanyi
G M
GM
Botelho
L H
LH
D000818:Animals; D001980:Bronchi / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D003470:Culture Media; D019332:Endothelin-1; D016232:Endothelins / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D009130:Muscle, Smooth / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism*; D011651:Pulmonary Artery / Q000378:metabolism; D011863:Radioimmunoassay
D003470:Culture Media; D019332:Endothelin-1; D016232:Endothelins; D011498:Protein Precursors