Induction of angiogenic immediate early genes by activation of FP prostanoid receptors in cultured human ciliary smooth muscle cells

Purpose: Agonists of the F prostanoid receptor for prostaglandin F _2α exert. exert an ocular hypotensive effect that has been attributed to increased aqueous humor outflow through the uveoscleral pathway. Although tissue remodeling of the ciliary muscle has been described, the signaling mechanisms that link activation of the FP receptor to remodeling of the ciliary muscle are poorly understood. Herein, we describe the identification of novel signaling mechanisms that may contribute to this process. Materials and Methods: Cultures of human ciliary smooth muscle cells were established from fetal eye tissue explants. The cells were validated by their expression of α-smooth muscle-actin and functional FP receptors. Cultures were treated with prostaglandin F_2α and examined for the induction of three immediate early genes related to tissue remodeling using Western blot analysis, quantitative real-time polymerase chain reaction, and reporter gene assays. Results: Human ciliary smooth muscle cells express functional FP receptors whose activation up-regulates the expression of early growth response factor-1 and connective tissue growth factor at the mRNA and protein levels. Prostaglandin F_2α stimulation also increases the protein expression of hypoxia-inducible factor-1α and activates luciferase reporter plasmids under the control of the hypoxia response element. Conclusions: Early growth response factor-1 and hypoxia-inducible factor-1α are important transcriptional activators of downstream genes involved in tissue remodeling and angiogenesis, whereas connective tissue growth factor is a secreted growth factor that also contributes to these processes. Thus, stimulation of FP receptors in human ciliary smooth muscle cells up-regulates the expression of immediate early genes that may coordinate the remodeling of the ciliary muscle, thereby facilitating aqueous outflow.