Abstract
Abstract: :
Purpose:Bradykinin (BK) stimulates B2 kinin receptors to activate multiple signaling pathways in trabecular meshwork (TM) cells. The objective of the present study was to investigate effects of BK signaling on secretion of matrix metalloproteinases (MMP) by TM cells and the consequences of these effects on outflow facility. Methods: Experiments on BK signaling and MMP secretion were performed in primary cultures of TM cells studied at passages 2 and 3. Determinations of outflow facility were conducted in preparations of isolated anterior segments perfused at constant pressure of 10 mmHg. Results:Incubation of bovine or human TM cells with BK produced a concentration–dependent increase in intracellular free Ca2+. The dose–response relationship was similar for bovine and human cells, with the response maximum occurring at 100 nM BK and an EC50 of 3 nM. This effect was blocked by pretreatment of cells with Hoe–140, a B2 receptor antagonist. Bk treatment also produced a 10–fold increase in ERK1/2 phosphorylation that reached a maximum after 10 min of stimulation. Incubation of TM cells with 100 nM BK for 2 hours increased secretion of MMP–2 into the extracellular medium compared to untreated control cells, and this effect was prevented when cells were pretreated with Hoe–140. Perfusion with 100 nM BK increased outflow facility in both bovine( 100%) and human (35 %) anterior segments when compared with control perfusion. In each preparation, BK increase of outflow facility was evident within 30 min and achieved a maximum at about 4 hrs after initiation of treatment. Addition of GM6001, a non–selective inhibitor of MMP activity, to the perfusion media blocked BK–induced increase in outflow facility. Conclusions: These results indicate that BK activates B2 kinin receptors to initiate signaling in TM cells that promotes secretion of constitutively expressed MMP–2. Release and activation of MMP–2 in response to BK then initiates events leading to increased outflow facility. The composite of the data supports the possibility that kinins produced within the eye may contribute to the regulation of trabecular cell function and outflow resistance in the TM. Supported by NIH grants EY–09741, EY–014793 and RPB.
Keywords: trabecular meshwork • signal transduction • outflow: trabecular meshwork