No abstract

Tumor-induced angiogenic responses lead to complex phenotypic changes in vascular endothelial cells, which must coordinate the expression of both proteases and protease inhibitors prior to the proliferation and invasion of surrounding stroma. Matrix metalloproteinase 2 (MMP2), which degrades Type IV collagen, is produced as proMMP2. proMMP2 is activated in part through its interactions with membrane Type 1 MMP (MT1-MMP) and tissue inhibitor of matrix metalloproteinase 2 (TIMP2). In this study, we demonstrate that platelet-activating factor (PAF) is a potent inducer of human umbilical vein endothelial cell (HUVEC) migration and invasion, which is attenuated by PAF receptor antagonists, and that PAF receptor antagonists inhibit the migration and invasion of HUVEC mediated by medium conditioned by a prostatic carcinoma cell line. We confirm that PAF receptor antagonists inhibit proliferation of HUVEC grown in rich growth medium. We show that PAF increases mRNA levels for MT1-MMP and TIMP2, followed by increased temporal conversion of latent proMMP2 to MMP2. Finally, we demonstrate that the ratio of MT1-MMP to TIMP2 in membrane preparations from PAF-stimulated HUVEC is 1.6:1, approximating the hypothesized ideal ratio of 2:1 necessary for the conversion of proMMP2 to MMP2. Our data support the involvement of PAF in vascular endothelial cell migration and invasion.

During corneal injury and inflamation, PAF is an important factor in the activation of proteolytic cascades, which could lead to corneal epithelial defects and ultimately ulceration. One important goal in treating these defects is to modulate the activity of enzymes that destroy the extracellular matrix. Our results suggest that COX-2 induction following PAF stimulation and subsequent eicosanoid release may play a crucial role in the induction of uPA, MMP-1 and MMP-9 enzymes. Specific COX-2 inhibition could therefore block the actions of PAF when inflammation is sustained.

No abstract

No abstract

Activation of protein kinase C (PKC) involves its recruitment to the membrane, where it interacts with its activator(s). Protein kinase C (PKC)1 is a multifunctional family of serine/ threonine protein kinases with 12 different isoforms, whose activities are dependent on Ca 2ϩ , lipid second messengers, and/or protein activators and regulators (1, 2). Among them, four classical PKC isoforms (␣, ␤I, ␤II, and ␥) require Ca 2ϩ , diacylglycerol, and phosphatidylserine for their activation. The novel PKC isoforms (␦, ⑀, , , and ) require diacylglycerol and phosphatidylserine for activation but are independent of Ca 2ϩ . The atypical PKC isoforms (, , and ) are activated by phospholipids. These isoenzymes play crucial roles as transducers of various extracellular receptor-originated signals by hormones, neurotransmitters, and growth factors triggering cell proliferation, differentiation, cytoskeletal alterations, and gene expression (1, 3). They also contribute to cellular signaling through cross-talk with other signaling cascades.Upon various physiologic stimuli, PKCs display differential patterns of subcellular localization (4, 5). The movement of PKC is a prerequisite for its effective response to physiologic activators, such as membrane lipids, and allows it to gain access to its specific substrates and to be selective in their response. The time course and duration of PKC relocation may determine the eventual signal response. Previous studies from our laboratory showed that rabbit corneal epithelium expresses PKC␣, ␥, ⑀, , and , that PKC␣ is activated after corneal injury, and that inhibition of its expression delays epithelial wound healing (6, 7). We previously proposed that PKC␣ plays a role in corneal epithelial proliferation (7). However, the transduction mechanisms underlying PKC␣ activation in cornea are still poorly understood.Growth factors such as EGF, HGF, and KGF increase in response to corneal epithelial injury (8) and are important in homeostasis and wound repair (9 -11). Earlier studies showed that PKC␣ activation requires changes in subcellular localization after phorbol ester (12-O-tetradecanoylphorbol-13-acetate) stimulation (12). However, information about relocalization of PKC␣ as a function of time upon growth factor stimulation is relatively scant.

PAF stimulates corneal neovascularization by a receptor-mediated mechanism. Induction of VEGF expression and stimulation of vascular endothelial cell migration are initial events in PAF-promoted corneal angiogenesis.

No abstract