past decades, studies on the cultured human RPE cells have shown that HGF protects RPE against various oxidative stresses [59–61]. This chapter further discusses the protective effects of HGF from oxidative stress in RPE cells.

16.2HGF and Its Receptor (MET)

HGF is a heterodimeric protein composed of an a-chain and a b-chain. The biological activity of HGF was first demonstrated in the sera of normal and partially hepatectomized rats and was found to be a potent mitogen of hepatocytes. HGF has been purified, cloned, sequenced, and was found to be identical to the scatter factor, fibro- blast-derived tumor cytotoxic factor, and fibroblast-derived epithelial morphogen [37–41]. The HGF receptor (MET) was identified as the product of the c-met protooncogene. MET was found to be expressed not only in hepatocytes, but also in other types of epithelial cells, mesenchymal cells, and neurons. Signal transduction pathways for HGF in these cells involve tyrosine phosphorylation of MET and subsequent activation of a variety of signal pathways [37–41].

16.2.1Production and Secretion of HGF

The HGF gene is located at the long arm of human chromosome 7 at 7q21.2 and consists of 18 exons and 17 introns. The major HGF-producing cell types are mesenchymal cells, including fibroblasts, vascular smooth muscle cells, glial cells, macrophages, and activated T lymphocytes [37–41]. Most epithelial cells produce little or no HGF, with the exception of a few cell types, including RPE cells [51, 52].

HGF is synthesized as a single chain peptide of 728 amino acid residues, which is present in various tissues as an inactivated form. It can be activated by several enzymes. Latent HGF can be cleaved between amino acids Arg494 and Val495 to induce full biologic activity. Activated HGF consists of two peptide chains, a (approximately 64 kDa) and b (approximately 33 kDa) linked by a disulfide bond. The molecules that have been shown to cleave latent HGF to its active form include plasmin, urokinase-type and tissue-type plasminogen activators, and a Factor XIIlike protein known as HGF activator, all of which belong to the serine protease family of proteins [37–41].

The HGF mRNA and protein are present in both embryonic and adult tissues, including blood, brain, liver, kidney, lung, placenta, skin, spleen, brain, spinal cord, peripheral ganglia, and others [37–41]. HGF can be detected in the blood, with normal levels ranging from 120 to 500 pg/mL [38, 39, 41].

Historically, HGF has been described as a paracrine cytokine because it is produced by fibroblasts and exerts its effects upon epithelial cells. Recently, however, several epithelial cells including RPE cells have been found to produce and secrete HGF. It appears, therefore, that an autocrine function is also present under certain circumstances [37–41].

16.2.2MET and Biological Effects of HGF

MET is the protein product of the c-met protooncogene. It is a transmembrane tyrosine kinase expressed predominately by epithelial cells. The c-met gene is located at human chromosome 7q31, spans more than 120 kb and consists of 21 exons and 20 introns. MET is a 190-kDa glycoprotein consisting of an a chain and a b chain. The 50-kDa a chain is heavily glycosylated and fully present on the cell surface. The 145-kDa membrane-spaning b-chain also has an extracellular portion that is involved in ligand binding, as well as a transmembrane segment and a cytoplasmic tyrosine kinase domain containing multiple phosphorylation sites. Binding of HGF to MET leads to tyrosine phosphorylation of MET which in turn influences the migration, mitosis, survival, and morphology of various cell types [37–41].

MET is expressed in both embryonic and adult tissues. In adults, MET is present at relatively low levels in liver, breast, lung, kidney, intestine, placenta, skin, stomach, thyroid, and others. Although MET is expressed mainly in the epithelial cells, it is also present in melanocytes, vascular endothelial cells, microglial cells, neurons, and hemopoietic cells [37–41].

HGF is a potent mitogen which induces dissociation and migration of many cell types, especially epithelial cells. HGF also acts as a mitogen, promoting the growth of epithelial cells, endothelial cells, and some stromal cells [37–41]. HGF also promotes cell survival, especially in neurons [42, 43]. HGF is also a morphoregulatory agent for various cells.

During embryogenesis, HGF supports organogenesis and morphogenesis of diverse tissues and organs. In adult tissues, HGF plays a role in tissue repair, enhances wound healing process, and supports regeneration of numerous organs, e.g., the liver, kidney, and lung [37–41].

HGF has angiogenic activity and is also involved in hematopoiesis and chondrogenesis. In neoplasia, HGF stimulates tumor cell motility and invasion, and promotes angiogenesis and enhances metastasis of malignant tumors [37–41]. Thus, HGF plays important roles in the regulation of both normal and pathologic physiological processes [37–43].

16.2.3Signaling Pathways of HGF

Following HGF binding to its receptor, the kinase activity of MET is switched on, leading to activation of several downstream signal transduction pathways, including the following:

1. The mitogen-activated protein kinase (MAPK) cascades consisting of three subfamilies, the terminal effectors include (1) extracellular signal-regulated kinases (ERK), (2) Jun amino-terminal kinases (JNKs), and (3) p38 MAPK. Phosphorylation of ERK promotes cell proliferation and survival. Phosphorylation of JNKs and p38 leads to cell differentiation, transformation, and apoptosis.

2. The Phosphoinostitide 3-kinase (PI3K)/Akt signal pathway, which promotes cell proliferation and survival and protects cells from apoptosis.

3. The signal transducer and activator of transcription 3 (STAT3) pathway, which promotes cell proliferation, transformation, epithelial tubulogenesis, and tumorigenesis.

4.The nuclear factor-kB (NF-kB) pathway, which upon activation and nuclear translocation promotes expression of various cytokines and growth factors, and modulates cell proliferation and survival [40, 41].

In addition to these pathways, a variety of other elements (Src, phospholipase C-gamma, SHC, etc.) are also involved in the MET-signaling pathways [40, 41, 61].

16.2.4HGF and MET in Disease States

HGF and MET play a role in the pathogenesis of tumors. Overexpression of HGF and/or elevation of HGF serum levels have been found in various malignant tumors, including bladder, breast, gastric, hepatocellular, and lung cancers, leukemia, glioma, and multiple myeloma. Abnormal elevation of HGF displays a positive correlation with disease progression in several types of tumors, such as lung, breast, gastric, skin, and bladder cancers [40].

c-met is also overexpressed in a variety of tumors, including hepatocellular, breast, bladder, colorectal, esophageal, gastric, prostate, and pancreatic cancers, melanoma, glioma, Kaposi’s sarcoma, chondrosracoma, osteosarcoma, Hodgkin’s disease, and leukemia, especially in later stages with metastasis [40].

HGF stimulates tumor cell migration, proliferation, and invasion. HGF also has a role in angiogenesis that may contribute to the development and metastasis of tumors [37–41].

HGF is present in the blood. Elevated HGF blood levels have been found in many malignant tumors and several nonneoplastic diseases, including liver diseases (hepatitis and liver cirrhosis), advanced hypertension, autoimmune diseases (systemic lupus erythematosus, etc.), and myocardial infarction [37–41, 61–63].

HGF is also a potent angiogenic factor, and application of HGF or HGF gene therapy has been proposed to treat myocardial infarction, peripheral vascular disease, and restenosis after angioplasty [61, 62].

16.3HGF and the Eye

HGF has been identified in many tissues as a paracrine modulator of stromal– epithelial interactions through secretion by fibroblastic cells to regulate epithelial cells functions. However, in the eye, several types of epithelial cells have also been found both to produce and secrete HGF [55, 64–68].

HGF is expressed by a variety of ocular cell types, including keratocytes, corneal endothelial cells, vascular smooth muscle cells, pericytes, and Mueller cells [55, 64–73]. Interestingly, several ocular epithelial cells also express HGF, including RPE cells, iris pigment epithelial cells, lens capsule epithelial cells, and corneal epithelial cells, although the levels expressed in the cornea appear to be very low [51, 64–68].

MET is also expressed in many ocular cell types, including corneal epithelial cells and endothelial cells, trabecular meshwork cells, vascular endothelial cells, lens capsule epithelial cells, uveal melanocytes, iris pigment epithelial cells, RPE cells, and Mueller cells [36–41, 51–61, 64–75].

HGF stimulates migration, mitosis of ocular epithelial cells (corneal epithelial cells, iris pigment epithelial cells, lens capsule epithelial cells, and RPE cells), uveal melanocytes, corneal endothelial cells, trabecular cells, vascular endothelial cells, etc. [51–58, 64–66, 68, 74, 75]. HGF also promotes survival of RPE cells [60, 61]. Lacrimal HGF modulates corneal epithelial cell proliferation, migration, and differentiation [69].

HGF can be detected in the aqueous humor. We found that the aqueous humor levels of HGF in cataract patients were 563 ± 179 pg/mL (mean ± standard deviation) [76]. Total aqueous humor protein in eyes undergoing cataract surgery is within normal range. Therefore, although we cannot exclude the possibility that the presence of cataracts may affect the composition of aqueous humor, this data may represent the closest approximation of normal aqueous humor levels of HGF [76].

We collected aqueous humor and plasma in 24 patients with cataract and glaucoma. Aqueous HGF levels (861 pg/mL) were significantly higher than the plasma HGF levels (564 pg/mL). There was no correlation between HGF levels in the aqueous humor and plasma. These results suggest that HGF in the aqueous humor is produced in the eye locally by cells lining the anterior and posterior chambers, including the corneal endothelium, trabecular meshwork cells, iris pigment epithelium, iris fibroblasts, and lens capsular epithelial cells [76].

Aqueous HGF levels are also increased in diabetic retinopathy and glaucoma, especially in exfoliation glaucoma [76].

HGF also can be detected in the vitreous humor. The vitreous acts as a reservoir for several bioactive substances including growth factors. Vitreous HGF levels averaged 1,500–2,000 pg/mL in patients with idiopathic epiretinal membrane (the closet to normal vitreous HGF levels), which is higher than that in the aqueous humor [65, 77, 79, 80]. The difference between HGF levels in the aqueous humor and the vitreous probably reflects anterior–posterior gradients in the eye and/or the rapid clearance of this growth factor from the anterior chamber [79]. It is unlikely that HGF in the vitreous originates from the serum and that the lens and retina stand out as the most probable sources [65].

Vitreous HGF levels are significantly higher in patients with proliferative diabetic retinopathy (with a mean of 5,700 pg/mL, but levels can be as high as 25,000 pg/mL in extreme cases) and proliferative vitreoretinopathy (average of 3,310–3,940 pg/ mL), but not in the early stages of diabetic retinopathy and retinal detachment [65, 77, 79, 80]. MET is overexpressed in the cellular components of preretinal