Assessment of rapid morphological changes associated with elevated cAMP levels in human orbital fibroblasts

Orbital fibroblasts exhibit a phenotype distinct from that of other types o f fibroblasts. Addition of prostaglandin E-2 (PGE(2)) to culture medium eli cits a dramatic change in orbital fibroblast morphology, That response is m ediated through the generation of cAMP, Orbital fibroblasts can generate hi gh levels of PGE(2) through induction by proinflammatory cytokines of prost aglandin endoperoxide H synthase-2 (PGHS-2). Here we compare the influence on fibroblast morphology of exogenous PGE(2), forskolin, and 8-br-cAMP to t hat mediated through PGHS-2 induction by a lymphocyte-derived cytokine, Wit hin a few hours, orbital fibroblasts treated with any of these test compoun ds appear under phase-contrast microscopy to exhibit a stellate morphology, When these changes were assessed quantitatively by electric cell-substrate impedance sensing (ECIS), it became evident that 8-br-cAMP, forskolin, and PGE(2) initiated shape changes within 30 min of addition to the culture me dium, while effects of the cytokine were first evident after approximately 3.5 h, Dermal fibroblasts failed to respond to any of these compounds with regard to changes in cellular morphology. Analysis of micromotion, manifest ed as small impedance fluctuations, revealed that orbital fibroblasts treat ed with 8-br-cAMP exhibit less motion than did untreated cells, These resul ts suggest that orbital fibroblast shape can be altered by several compound s known to alter intracellular cAMP levels. They demonstrate the utility of ECIS in the assessment of very rapid and dynamic cellular events associate d with changes in cell morphology, (C) 1998 Academic Press.