Fig. 1.2 Fundus photograph illustrating intraretinal hemorrhage (double circle), nerve fiber layer infarct (arrow), and intraretinal lipid in a case of severe nonproliferative diabetic retinopathy

1.4Pathophysiology

Typical clinical manifestations of DR reflect damage to retinal blood vessels with sequelae that include retinal hemorrhage (Fig. 1.2), nerve fiber layer infarcts (Fig. 1.2), retinal edema (Fig. 1.3), venous beading (Fig. 1.4), intraretinal microvascular abnormalities (Fig. 1.4), venous loops (Fig. 1.5), retinal vascular occlusion (Figs. 1.4 and 1.6), retinal neovascularization (Figs. 1.6 and 1.7), vitreous hemorrhage (Fig. 1.6 and 1.7), and retinal detachment (Fig. 1.8). The pathophysiology underlying these changes is complex. Recent studies suggest chronic hyperglycemia induces inflammation and oxidative stress, both of which promote many interconnected biochemical processes that ultimately lead to microvascular and neuronal dysfunction [70]. Increased expression of inflammatory genes is present in animal models of diabetes [71].

Oxidative stress may be the fundamental mechanism that triggers diabetesrelated microvascular and neuronal complications. Hyperglycemia can induce oxidative stress by promoting the polyol pathway (extra glucose is converted into sorbitol and fructose); increasing expression of inflammatory genes; activating protein kinase-C (PKC); increasing synthesis and over-activation of the hexosamine pathway, glucose auto-oxidation, accumulation of advanced glycation end products (AGEs), and activation of the receptor for AGEs (RAGEs); and activating cytochrome P450 monooxygenase and nitric oxide synthase [72–74]. AGEs and RAGEs play an important role in activation of Müller cells and production of cytokines that are involved in increasing excitotoxicity and oxidative stress at the cellular level [75, 76]. The exact mechanism of enhanced production of reactive oxygen species in the mitochondria of diabetic patients is unclear. The proteins undergo sequential nonenzymatic glycation with reducing sugar to form AGE in a hyperglycemic environment. This step is accompanied by oxidative, free radical-generating reactions

a

b

Fig. 1.3 (a) Color fundus photograph illustrating moderate nonproliferative diabetic retinopathy associated with macular edema involving the fovea. (b) Optical coherence tomography (OCT) illustrating cystoid macular edema. Upper left panel: red-free photograph. Upper right panel: OCT transverse image through fovea demonstrating cystic change in the outer plexiform layer. Lower panel: graph of retinal thickness in transverse image is 555 μm in the foveal center

[77]. AGE binding to RAGE initiates a chain of events that leads to the development of microvascular complications [78]. Soluble RAGE (sRAGE), an inhibitor of AGE-RAGE, protects against AGE-RAGE-mediated microvascular damage [79].

Leukostasis, adhesion of leukocytes to the vascular endothelium, is a major component of inflammation and is thought to be a major contributing factor to diabetic

Fig. 1.4 Fundus photograph illustrating intraretinal microvascular abnormality (long arrow), occluded retinal arteriole (medium-sized arrow), and venous beading (short arrow) in a case of very severe nonproliferative diabetic retinopathy

Fig. 1.5 Fundus photograph illustrating a venous loop (arrow). Venous beading and nerve fiber layer infarcts (also known as cotton wool spots) are also present

vasculopathy. Leukostasis is associated with increased expression of adhesion molecules (intracellular adhesion molecule-1, ICAM-1) on the retinal endothelium and increased expression of integrins on leukocytes. Leukostasis causes endothelial cell injury and death, capillary nonperfusion, ischemia, and breakdown of blood-retinal barrier with retinal vascular leakage. Reduced retinal capillary damage and vascular permeability are noted in knockout mice that lack intracellular ICAM-1 or leukocyte integrin CD18 [80].

Fig. 1.6 Fluorescein angiogram illustrating extensive areas of retinal vascular nonperfusion (black arrow), retinal neovascularization with associated fluorescein dye leakage (white arrows), and an area of vitreous hemorrhage (double circle) that blocks fluorescence

Fig. 1.7 Fundus photograph illustrating vitreous hemorrhage (arrow) arising from retinal neovascularization at the optic nerve head

Ischemia further accentuates the action of AGEs and promotes release of proinflammatory factors, nitric oxide, VEGF, and angiotensin II. VEGF is the most potent vasoactive cytokine. It upregulates ICAM-1 and causes dysfunction of the tight junctions of the endothelial cells causing increased vascular permeability. The blood-retinal barrier, both inner and outer, is compromised, promoting retinal neovascularization and DME. PDR seems to be directly associated with VEGF levels in the vitreous; DME, however, can be seen in any stage of DR and is not necessarily seen in every PDR case. This fact indicates that other pathways, perhaps those involving inflammation, may be contributing to the severity of edema and to ischemia to a certain extent.

Fig. 1.8 Fundus photograph illustrating traction retinal detachment involving the macula arising from fibrovascular proliferation (arrow) in an untreated patient with proliferative diabetic retinopathy. Note that as the retinal neovascularization matures, it acquires a fibrotic component and contractile properties

Persistent low-grade inflammation in diabetes causes an increase in tumor necrosis factor-alpha (TNF-alpha) levels, which also promotes leukostasis [81]. Increased levels of TNF-alpha is present in the vitreous fluid of diabetic patients [82]. Plasma levels of TNF-alpha correlate with the severity of DR [83]. Blockade of TNF-alpha reduces leukostasis, suppresses blood-retinal barrier breakdown, and reduces ICAM-1 expression [84].

DR traditionally has been considered a microvascular disease; however, new evidence suggests that retinal neurodegenerative changes precede microvascular changes [65]. Glutamate, the major excitatory neurotransmitter in the retina, is abnormally elevated in the extracellular space in diabetes [85]. It leads to “excitotoxicity,” an uncontrolled intracellular calcium response in postsynaptic neurons causing neural apoptosis, especially of the retinal ganglion cells [70, 86–89], resulting in thinning of nerve fiber layer in eyes with no or very minimal DR [88–90]. Also, reactive changes are seen in Müller cells with an aberrant expression of glial fibrillary acidic protein [91].

Some neuro-retinal functional changes (dark adaptation, multifocal electroretinogram, and tritan color defects) can be seen much earlier than the structural microvascular changes such as microaneurysms and dot-blot hemorrhages. It is possible that neuronal and vascular destructions occur in parallel. The inner retina receives a sparse retinal blood supply compared to the outer retina, which is mostly supplied by the choroid circulation. Diabetes-induced perfusion changes thus largely affect the inner retinal neurons.

Chan and associates have demonstrated increased levels of pro-inflammatory mediators, interleukin-1b, TNF-alpha, ICAM-1, and vascular cell adhesion mole- cule-1 in the retina of diabetic rats during poor glycemic control, which fail to return to normal levels even with good glycemic control. This irreversibility of retinal inflammatory mediators is consistent with the clinical observation that DR can remain active even when the blood glucose is brought under control [92].