requires the CNTFRa receptor subunit [34]. CNTF is primarily localized in Müller cells and is expressed in both the developing and mature retina in the rat [35, 36]. The CNTF receptor is located in the retinal Müller, horizontal, amacrine, and ganglion cells [35].

CNTF has several functions in the retina including, but not limited to, promoting the survival and axonal regeneration of RGCs, promoting green cone cell differentiation, and inhibiting rod cell differentiation [31, 32]. The majority of CNTF’s functions are through the JAK/STAT intracellular signaling pathway [37], although it can also activate the ERK [38] and PI3-K/Akt pathways [39].

CNTF has been shown to aid in the survival of the retinal neurons in several retinal degenerative disorders [31, 40]. Intravitreal injection of recombinant CNTF into a retinal degeneration model led to a short-term rescue of photoreceptors [35, 40]. In another study, injection of an adenovirus expressing CNTF delayed photoreceptor degeneration in retinal degeneration (rd/rd) mice [41, 42]. Future studies are considering the use of an intravitreal implant that would apply a prolonged delivery of CNTF to the retina for longer neuronal protection [43].

ANTI-ANGIOGENIC NEUROTROPHIC FACTORS

Pigment-Epithelium-Derived Factor

Pigment-epithelium-derived factor (PEDF) is a member of the SERPIN gene family [44] and was first isolated from fetal retinal pigment epithelial cells [8, 45]. PEDF’s actions were initially characterized as being primarily involved in neuronal differentiation [46]. However, as more information was gathered about PEDF, its role as an angiogenic inhibitor was revealed [47]. In fact, PEDF and another neurotrophic factor, vascular endothelial growth factor (VEGF), play reciprocal roles in the angiogenic process [8]. In models of oxygen-induced retinopathy (OIR) and DR, as the levels of the proangiogenic factor (VEGF) increase, the levels of PEDF decrease in the aqueous humor and vitreous of the eye [17, 18, 48–51]. This intricate balance between VEGF and PEDF levels is essential in maintaining the BRB integrity through prevention of vascular permeability [47, 52]. However, a reduced level of PEDF in the ischemic, nondiabetic eye has also been observed. This indicates that the reduced level of PEDF observed in DR is due to hypoxia rather than hyperglycemia [17]. In addition to its potent anti-angiogenic properties, recent findings have shown that PEDF is also an anti-inflammatory factor in the eye [53]. PEDF plays a role in inhibiting reactive oxygen species (ROS) as well as the subsequent VEGF increase that is seen in DR [47].

The effects of exogenous PEDF treatments on angiogenesis and other DR-associated symptoms have been studied. For instance, intraperitoneal administration of PEDF was shown to inhibit retinal neovascularization in a neonatal mouse exposed to hypoxic conditions [54]. A second study used an adenovirus expressing PEDF (AAV-PEDF). Intravitreal injection of AAV-PEDF inhibited both retinal and choroidal neovascularization in the mouse [8, 55]. In a third study, retinal vascular permeability and inflammatory factors were reduced in animal models of DR and OIR after intravitreal injection of PEDF [53].

SERPINA3K

SERPINA3K, a member of the SERPIN family, is a specific inhibitor of tissue kallikrein (a serine proteinase) and is often referred to as kallikrein-binding protein (KBP) [56, 57]. The kallikrein-kinin system was originally characterized to have functions in inflammation, local blood flow, and vasodilation regulation [58, 59]. As research continued on SERPINA3K, additional functions were uncovered, including its role as an anti-angiogenic factor [60].

In the STZ-induced diabetic rat model, the retinal levels of KBP are decreased, hinting at an essential role in the progression of DR [61]. In 2008, it was uncovered that SERPINA3K can function in a protective manner in both Müller and retinal neuronal cells against oxidative stress-induced damage, conditions seen in DR [62]. This protective effect occurs through blocking the intracellular calcium overload induced by oxidative stress [62].

THE DOUBLE-EDGED SWORDS: PRO-ANGIOGENIC

NEUROTROPHIC FACTORS

As the knowledge increases about the anti-angiogenic neurotrophic factors in the retina, the relationship between neuronal cell protection and pro-angiogenic factors has broadened. Several pro-angiogenic factors, to be described below, have dual functions in the cell: promoting angiogenesis while promoting neuronal cell maintenance, differentiation, and development.

Brain-Derived Neurotrophic Factor

Brain-derived neurotrophic factor (BDNF) shares a similar structure to the most highly studied neurotrophic factor, NGF, as both are members of the neurotrophin gene family [63]. In the retinal tissue, BDNF targets (and is expressed in) RGCs and Müller glia [64] and has been shown to be important for the survival of RGCs and bipolar cells [11, 65, 66]. In addition, it has been shown that BDNF can prevent amacrine cell death [67, 68]. Upon the degeneration of dopaminergic amacrine cells in the retinas of STZ-induced diabetic rats, there is a reduction in the levels of BDNF in both RGCs and Müller cells [11]. BDNF’s ability to bind to both the Trk and p75-type receptors facilitates its action in both retinal development and survival [69]. However, a recent study has suggested a novel pro-angiogenic role for BDNF in ischemic tissues [70].

The therapeutic potential of BDNF has been examined. Upon intraocular injection, BDNF prevented dopaminergic amacrine cell neurodegeneration [11]. In order to gather more information on BDNF and its usefulness in treating DR-associated pathological phenotypes, more studies remain to be conducted.

Fibroblast Growth Factors

Fibroblast growth factor (FGF) was first characterized as a growth and differentiation factor for mesodermand neuroectoderm-derived cells [71]. However, researchers have isolated two derivatives of the FGF family from the bovine retina, basic FGF (bFGF) and acidic FGF (aFGF) [71, 72]. bFGF is constitutively expressed by the RPE at

considerably higher amounts than aFGF [71, 73]. During retinal ischemia and instances of proliferative DR, the retinal levels of bFGF are increased [22, 71]. In fact, it is speculated that during retinal hemorrhage, infiltrative macrophages in the vitreous may induce an enhanced secretion of bFGF [71, 74].

Although early studies on FGF had showed a link between its elevated expression and angiogenesis, now the primary function of FGF is thought to be neurotrophic and neuroprotective [22, 75]. Although bFGF has not been utilized for DR therapies, injections of bFGF into the eye of rats with either inherited retinal degeneration or ischemic injury led to a delay in the progression of degeneration [76, 77].

Insulin and Insulin-Like Growth Factor 1

Insulin and Insulin-like growth factor 1 (IGF-1) have been shown to prolong the survival of retinal neurons in culture as well as decrease apoptosis and stimulate cell proliferation, differentiation, and maturation [78–80]. In DR, increased levels of IGF-1 were observed in the vitreous of patients; IGF-2 levels do not increase [81, 82].

The use of IGF-1 has been examined as a possible therapeutic agent in the treatment of DR. Exogenous exposure of IGF-1 to cultured retinal neurons led to the enhanced survival of amacrine neurons [83]. Exposure of high levels of either insulin or IGF-2 led to the same effects [83]. Furthermore, upon depletion of these factors, there was an increase in amacrine apoptosis [83].

Erythropoietin

Erythropoietin (EPO) was initially described as a regulator of red blood cell production, or erythropoiesis, throughout the body [84, 85]. However, as the information about EPO broadened, it was found to be expressed in the retina [86]. In the retina, as well as in the brain, EPO is both a neurotrophic factor and an endothelial survival factor [22, 87]. EPO is elevated in the diabetic eye, and although it is neuroprotective in the retina, it has been shown in both in vitro and in vivo studies to stimulate angiogenesis [87]. EPO is regulated by hypoxia-inducible factor (HIF), and oxidative stress stimulates EPO production in the eye [88]. However, EPO’s production is not solely dependent on the presence of oxidative stress because elevated levels of EPO were observed in cases of macular edema, a condition that is not solely dependent on hypoxic conditions [84, 86].

Intravitreal injection of EPO has been found to prevent apoptosis during early stages of DR [89]. In addition, suberythropoietic administration of EPO reduces the unnecessary side effects that can be associated with other potential EPO therapies, such as induction of angiogenesis, oxidative stress, and pericyte loss [87]. Another study explored the possibility of using siRNAs to EPO as a novel therapeutic agent for DR. Intravitreal injections of siRNA to EPO resulted in reduced levels of EPO and subsequent suppression of retinal neovascularization [90]. Although the results from the siRNA study are promising, methods to knock down EPO are risky due to its dual role as both an angiogenic stimulator and a neurotrophic factor in the retina.

Vascular Endothelial Growth Factor

VEGF is constitutively secreted by the retinal pigment epithelium [8, 91]. There are at least five different splice forms of VEGF, and each shows a differing amount of angiogenic activity [92, 93]. A combination of the isoforms can stimulate a higher