Embryonic development. HGF and its receptor, Met, are expressed during gastrulation and throughout later phases of vertebrate embryogenesis (Stern et al., 1990; Sonnenberg et al., 1993; Andermarcher et al., 1996). Overlapping expression of both genes persists into the earliest phases of organogenesis in the heart, condensing somites and neural crest cells (Andermarcher et al., 1996), but thereafter HGF is expressed in mesenchymal tissues and Met in the surrounding ectoderm in differentiated somites as well as lungs, liver, placenta, muscle, gut, heart and nervous system (Sonnenberg et al., 1993; Woolf et al., 1995; Andermarcher et al., 1996; Thewke and Seeds, 1996; Birchmeier and Gherardi, 1998; Ishikawa et al., 2001). Studies using tissue explants and cultured cells confirm the suspected role of HGF in epithelial branching morphogenesis, e.g. in the developing lung (Santos et al., 1994; Woolf et al., 1995; Ohmichi et al., 1998).
The expression of HGF and Met genes in ventral motor neurons of the embryonic spinal cord is also consistent with a role in tissue patterning through the regulation of migratory and morphogenic processes, such as axon guidance (Sonnenberg et al., 1993; Ebens et al., 1996; Wong et al., 1997). Functional studies indicate that HGF guides axons of spinal motor neurons to their distant muscle targets in the limbs (Ebens et al., 1996; Wong et al., 1997; Yamamoto et al., 1997) and acts as an essential survival factor for a subpopulation of limb-innervating motoneurons (Wong et al., 1997; Yamamoto et al., 1997). Both HGF and Met are also expressed in the brain and retina during development (E12-13) and in the adult, where signaling supports neuron survival and maturation (Jung et al., 1994; Honda et al., 1995; Yamagata et al., 1995; Hamanoue et al., 1996; Achim et al., 1997; Sun et al., 1999; Thewke and Seeds, 1999).
Loss of HGF or Met function in mice with homozygous gene deletion is embryonic lethal between days E12.5 and E15.5 (Schmidt et al., 1995; Uehara et al., 1995; Bladt et al., 1995). Defects in the proliferation and survival of cells in the liver and placenta result in arrested organogenesis of these and other tissues, underscoring the importance of HGF stimulated mitogenicity and survival in target cells. These models also highlight the importance of HGF as a potent and critical regulator of cell migration. Skeletal muscle progenitor cells that form limb, tongue, and diaphragm musculature normally delaminate from the epithelial dermomyotome of the somites by an epithelial-to-mesenchymal transition and migrate to their final destination where they complete differentiation. Homozygous deletion of Met results in defective delamination and migration of muscle progenitors from the dermomyotome and failure to form the skeletal muscles of the limb and diaphragm (Bladt et al., 1995; Maina et al., 1996; Dietrich et al., 1999; Rosário and Birchmeier, 2003; Christ and Brand-Saberi, 2002). Conversely, HGF overexpression in transgenic mouse embryos induces the inappropriate formation of skeletal muscle in the central nervous system (CNS) through dysregulated migration of Met containing myogenic precursor cells to the neural tube (Takayama et al., 1996).
Mice bearing conditional deletions of HGF or Met also have been used to demonstrate relevance of pathway activation at later developmental stages and in adulthood. Met and epidermal growth factor receptor jointly regulate final nephron number and collecting duct morphology (Ishibe et al., 2009). Mice with a targeted mutation of the gene encoding urokinase plasminogen activator, considered an important HGF activator, have decreased HGF levels and a substantial reduction in neocortical GABAergic interneurons at embryonic and perinatal ages, leading to changes in circuit organization and behavior (Powell et al., 2001; Powell et al., 2003a). Mice with targeted mutation of two critical carboxyl terminal tyrosine residues in Met were found to be phenotypically similar to Met null animals. In contrast, targeting one of those sites and thereby disrupting the consensus for Grb2 binding allowed development to proceed to term, but caused a striking reduction in limb muscle mass and a generalized deficit of secondary fibers, indicating the importance of HGF signaling in late myogenesis (Maina et al., 1996).

Maturity and adult homeostasis. In the developed brain, HGF is expressed in neurons, primarily in the hippocampus, cortex, and the granule cell layer of the cerebellum, as well as in ependymal cells, the chorioid plexus, and the pineal body (Streit et al., 1995). Met is expressed in neurons, preferentially in the CA-1 area of the hippocampus, the cortex, and the septum, as well as in the pons (Jung et al., 1994; Streit et al., 1995; Honda et al., 1995; Yamagata et al., 1995; Thewke and Seeds, 1999). HGF is though to provide a neurotrophic function in the CNS, supporting the survival and reconstruction of specific neurons in response to cerebral injury (Honda et al., 1995). HGF attracts and promotes the growth of cranial motor axons (Caton et al., 2000), induces c-Fos expression and activates the Ras pathway in brain neurons (Streit et al., 1997), stimulates Schwann cell growth (Krasnoselsky et al., 1994) and promotes axon outgrowth of embryonal carcinoma cells (Yang and Park, 1993). HGF stimulates neurite outgrowth in sensory and sympathogenic neurons, as well as enhanced survival and differentiation from progenitors (Maina et al., 1997; Maina et al., 1998).
HGF and Met are expressed in the cerebellum, where development is primarily postnatal and requires extensive cell proliferation and migration. Met is localized in granule cell precursors and cultures of these cells proliferate in response to HGF (Ieraci et al., 2002). HGF also promotes oligodendrocyte progenitor cell proliferation and delays their differentiation into myelinating oligodendrocytes during early postnatal development; subsequent down-regulation of HGF mRNA in the striatum observed between postnatal days 7 to 14 presumably permits differentiation and myelination to proceed (Ohya et al., 2007). Schwann cells, responsible for nerve myelination in the peripheral nervous system, also express Met mRNA (Krasnoselsky et al., 1994). Although Schwann cells are normally quiescent in adulthood, nerve injury and certain diseases such as type 1 neurofibromatosis trigger proliferation through several mitogenic pathways, including that of HGF (Krasnoselsky et al., 1994).
The mammary gland undergoes cyclic morphogenic differentiation during the menstrual cycle, pregnancy and lactation. HGF and Met are expressed and HGF is regulated temporally during mouse mammary development and differentiation (Niranjan et al., 1995; Yang et al., 1995). HGF secreted by fibroblasts acts on mammary myoepithelial and luminal epithelial cells expressing Met, promoting tubulogenesis in underlying myoepithelial cells, branching of the epithelial ductal tree and motogenesis in both cell types (Niranjan et al., 1995; Yang et al., 1995; Yant et al., 1998; Niemann et al., 1998).
HGF production in the adult vascular system is positively regulated by prostaglandins and HGF itself, and negatively regulated by angiotensin II, TGF-beta, glucose and hypoxia (reviewed in Morishita et al., 2002). HGF is induced in cardiac and skeletal muscle in animal models of ischemic injury (Aoki et al., 2000) and serum HGF levels are increased with hypertension, peripheral artery disease and myocardial infarction, consistent with homeostatic and repair functions (reviewed in Morishita et al., 2002).

Wound repair and tissue regeneration. Exogenous administration of the HGF protein or gene promotes angiogenesis without the increased permeability often observed with vascular endothelial cell growth factor (VEGF) treatment (Aoki et al., 2000; Taniyama et al., 2001; Morishita et al., 2004). HGF promotes angiogenesis directly (Sengupta et al., 2003) but also by inducing VEGF expression (Wojta et al., 1999; Gille et al., 1998), and the two factors appear to act synergistically on the vasculature (Van Belle et al., 1998; Xin et al., 2001). These and other findings support the use of HGF for therapeutic angiogenesis to treat peripheral artery disease, myocardial infarction and restenosis after angioplasty. Recent clinical trials indicate that HGF gene therapy is safe and effective for the treatment of critical limb ischemia (Powell et al., 2008; Shigematsu et al., 2010).
HGF signaling supports the natural reconstruction of central and peripheral neuronal networks in response to injury, and/or as a potential therapeutic agent to facilitate wound repair. Both HGF and Met expression are increased in reactive astrocytes in the subacute to chronic stage of spinal cord injury in rats (Shimamura et al., 2007). HGF gene transfer attenuated brain ischemic injury in rats, without cerebral edema, through angiogenic, neuroprotective and neuriotogenic activities, as well as prevention of gliosis (Shimamura et al., 2004; Shimamura et al., 2006). Intrastriatal administration of HGF protein also potently protected hippocampal neurons against postischemic delayed neuronal death (Miyazawa et al., 1996).
Tissue fibrosis is a common pathological consequence of chronic injury to kidneys and lungs. With chronic injury to these organs, the normal production and secretion of growth factors, including HGF, inflammatory cell recruitment, cell proliferation and differentiation, and matrix production and remodeling become increasingly aberrant, leading to matrix overproduction, abnormal organization, fibrotic lesions and scarring. Mice with conditional knockout of Met in the collecting duct of the kidney are more susceptible to interstitial fibrosis and tubular necrosis after unilateral ureteral obstruction, and show a diminished capacity for tubular cell regeneration after release of the obstruction (Ma et al., 2009). Conditional Met knockout targeted to renal podocytes was associated with more severe podocyte apoptosis and albuminurea than in control littermates subjected to nephrotoxic renal damage (Dai et al., 2010). HGF produced in response to injury antagonizes the actions of transforming growth factor-beta (TGF-beta), a critical profibrotic agent, thereby inhibiting fibrosis and preserving normal organ architecture and function (reviewed in Liu, 2004; Mizuno et al., 2008; Crosby and Waters, 2010; Panganiban and Day, 2011). The reciprocal effects of the HGF and TGF-beta signaling pathways occur via direct modulation of intracellular effectors downstream of TGF-beta and HGF receptors in common target cells, as well as by eliciting opposing activities in cells targeted independently (Yo et al., 1998; Gao et al., 2002; Mizuno et al., 2005). TGF-beta induced apoptosis of podocyte, endothelial and tubular epithelial cells, epithelial-to-mesenchymal transition by tubular epithelial cells, and myofibroblastic activation, are critical pathogenic events that are opposed by HGF signaling (reviewed by Böttinger and Bitzer, 2002). An abundance of findings support the therapeutic use of exogenous HGF, the HGF gene, or the induction of endogenous HGF expression, for the treatment of a variety of chronic fibrotic disorders in kidney (Mizuno et al.,1998; Mizuno et al., 2001; Dworkin et al., 2004; Dai et al., 2004; Herrero-Fresneda et al., 2006; reviewed in Liu and Yang, 2006; Mizuno et al., 2008) and lung (Yanagita et al., 1993; Dohi et al., 2000; Mizuno et al., 2005).
HGF signaling is required for liver regeneration (Nakamura et al., 1984; Thaler and Michalopoulos, 1985; Zarnegar and Michalopoulos, 1989; Nakamura et al., 1989; Miyazawa et al., 1989; Okajima et al., 1990). Studies of tissue selective HGF overexpression or Met suppression in genetically engineered animal models confirm and extend earlier studies (Shiota and Kawasaki, 1998; Borowiak et al., 2004; Huh et al., 2004; Paranjpe et al., 2007; Factor et al., 2010). In addition to stimulating the proliferation of mature hepatocytes, HGF contributes to the differentiation and maturation of hepatic progenitor cells (Kamiya et al., 2001). Treatment of animals with exogenous HGF protein or the HGF gene promotes survival in various experimental animal models of acute hepatic failure (Kosai et al., 1998; Nomi et al., 2000) and prevents fibrosis associated with liver cirrhosis (Kaibori et al., 1997; Matsuda et al., 1997). Clinical trials of recombinant human HGF for treatment of patients with fulminant hepatic failure are in progress (Ido and Tsubouchi, 2009).
HGF/Met signaling is required for full-thickness skin wound repair. Damage to the epidermis and dermis of the skin requires reepithelialization of the epidermis and the transient formation of dermal granulation tissue. During reepithelialization, keratinocytes from the wound edge form the hyperproliferative epithelium, which proliferates and migrates over the injured dermis and the granulation tissue. In addition to other important soluble regulators of skin repair such as epidermal and fibroblast growth factor family ligands and transforming growth factor-beta, locally secreted HGF promotes granulation tissue formation and reepithelialization (Yoshida et al., 2003; Chmielowiec et al., 2007). Engineered overexpression or exogenous application of HGF protein, or exogenous HGF gene transfer, to treat full-thickness skin wounds accelerates both processes, as well as vascularization, in rodent models (Toyoda et al., 2001; Yoshida et al., 2003; Bevan et al., 2004; Kunugiza et al., 2006).