Premium
Prostaglandin‐stimulated second messenger signaling in bone‐derived endothelial cells is dependent on confluency in culture
Author(s) -
Ida Rodney,
Lee Austin,
Huang Jason,
Yamaguchi Dean T.,
Brandi Maria Luisa
Publication year - 1994
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041600322
Subject(s) - paracrine signalling , extracellular , second messenger system , endocrinology , osteoblast , cell culture , medicine , microbiology and biotechnology , prostaglandin e , prostaglandin e2 , chemistry , biology , intracellular , in vitro , biochemistry , receptor , genetics
New bone formation is associated with an increase in blood flow by the invasion of capillaries. Endothelial cells that line the capillaries can produce paracrine factors that affect bone growth and development, and in turn, could be affected by products produced by bone cells, in particular the osteoblasts. Since osteoblasts produce prostaglandins E 2 and F 2α (PGE 2 , PGF 2α ), it was investigated if these PGs were agonists to bone‐derived endothelial cells (BBE) by assessing changes in cAMP and free cytosolic calcium concentration ([Ca 2+ ]i) second messenger generation. We found that confluent cultures of BBE cells, a clonal endothelial cell line derived from bovine sternal bone, responded to 1 μM PGE 2 by an increase in cAMP. PGF 2α at the same concentration was less potent in stimulating an increase in cAMP production in confluent BBE cells. Subconfluent cells with a morphology similar to that of fibroblastic cells were not as sensitive to PGE 2 ‐stimulated cAMP generation. PGF 2α failed to elicit any cAMP production in subconfluent cultures. PGE 2 and PGF 2α both stimulated an increase in [Ca 2+ ]i concentration in a dose‐dependent manner. The potency of PGE 2 was similar to that of PGF 2α in stimulating an increase in [Ca 2+ ]i. The Ca 2+ response was mostly independent of extracellular Ca + , was unchanged even with prior indomethacin treatment, was unaffected by caffeine pretreatment, but was abolished subsequent to thapsigargin pretreatment. The PG‐induced increase in [Ca 2+ ]i was also dependent on the confluency of the cells. In a subconfluent state, the responses to PGE 2 or PGF 2α were either negligible, or only small increases in [Ca 2+ ]i were noted with high concentrations of these two PGs. Consistent, dose‐dependent increases in [Ca 2+ ]i were stimulated by these PGs only when the cells were confluent and had a cobblestoned appearance. Since it was previously demonstrated that BBE cells respond to parathyroid hormone (PTH) by the production of cAMP, we tested if bovine PTH(1‐34) amide bPTH(1—34) also increased [Ca 2+ ]i in these cells. No change in [Ca 2+ ]i was found in response to bPTH (1—34), although bPTH (1—34) stimulated a nine to tenfold increase in cAMP. We conclude that BBE cells respond to PGE 2 and PGF 2α but not to bPTH(1—34) by an increase in [Ca 2+ ]i probably secondary to stimulation of phospholipase C and that the cAMP and [Ca 2+ ]i second messenger responses in BBE cells are dependent on the state of confluency of the cells. © 1994 Wiley‐Liss, Inc.