Fluid flow-induced prostaglandin E 2 response of osteoblastic ROS 17/2.8 cells is gap junction-mediated and independent of cytosolic calcium

Abstract

It has been well demonstrated that bone adapts to mechanical loading. To accomplish this at the cellular level, bone cells must be responsive to mechanical loading (mechanoresponsive). This can occur via such mechanisms as direct cell deformation or signal transduction via complex pathways involving chemotransport, hormone response, and/or gene expression, to name a few. Mechanotransduction is the process by which a bone cell senses a biophysical signal and elicits a response. While it has been demonstrated that bone cells can respond to a wide variety of biophysical signals including fluid flow, stretch, and magnetic fields, the exact pathways and mechanisms involved are not clearly understood. We postulated that gap junctions may play an important role in bone cell responsiveness. Gap junctions (GJ) are membrane-spanning channels that physically link cells and support the transport of small molecules and ions in the process of gap junctional intercellular communication (GJIC). In this study we examined the role of GJ and GJIC in mechanically stimulated osteoblastic cells. Following fluid flow stimulation, we quantified prostaglandin E 2 (PGE 2) (oscillatory flow) and cytosolic calcium (Ca 2+) (oscillatory and steady flow) responses in ROS 17/2.8 cells and a derivative of these cells expressing antisense cDNA for the gap junction protein connexin 43 (RCx16) possessing significantly different levels of GJIC. We found that the ROS17/2.8 cells possessing increased GJIC also exhibited increased PGE 2 release to the supernatant following oscillatory fluid flow stimulation in comparison to coupling-decreased RCx16 cells. Interestingly, we found that neither osteoblastic cell line responded to oscillatory or steady fluid flow stimulation with an increase in Ca 2+. Thus, our results suggest that GJ and GJIC may be important in the mechanotransduction mechanisms by which PGE 2 is mechanically induced in osteoblastic cells independent of Ca 2+.