Glial Cell–Derived Cytokines and Vascular Integrity in Diabetic Retinopathy

Fig. 1. Schematic representation of TJ. (A) Left panel, in this structural model of TJ, there are a number of intercrossing TJ strands (depicted as small dots) and three so-called kissing points of TJ. Right panel, freeze-fracture replica of a TJ. The TJ consists of an anastomosing network of strands that form irregular interstrand compartments and is comprised of a large number of protein components, including membrane proteins such as occludin and claudins, as well as cytoplasmic scaffolding proteins such as ZO-1. Scale bar, 50 nm. (B) In polarized cells, TJs are positioned at the boundary of the apical and basolateral plasma membrane domains to maintain cell polarity by forming a fence. TJs also seal cells together to generate the primary barrier and prevent diffusion of solutes through the paracellular pathway. In addition, a certain type of TJ protein such as occludin is a signaling molecule that has functions in receiving environmental cues and transmitting signals inside the cells.

homeostatic conditions, and this is often destroyed under pathological conditions, such as diabetic retinopathy [7, 8], uveitis [9], and other ocular inflammation and ischemia [10].

In diabetic retinopathy, microvascular complications such as macular edema and retinal neovascularization cause adult blindness of patients with diabetes mellitus [11]. During the pathological progression of diabetic retinopathy, leukocyte binding to the retinal vascular endothelium detected as an initial event results in early BRB breakdown, capillary nonperfusion, and endothelial cell death [12–15]. Also possible are molecular events within the initial stage of the diabetic retinopathy, an increase of vascular permeability caused by the breakdown of BRB, and upregulation of several cytokines and intracellular adhesion molecules. These pathological events merge to contribute to the development of retinal ischemia, diabetic macular edema, and neovascularization. In fact, during this pathological progression of the diabetic retinopathy, several intracellular adhesion molecules, including sICAM-1 and sVCAM-1 [16, 17], and inflammatory cytokines, including TNF-a and IL-1b [18, 19], VEGF [17, 20], GDNF [21, 22], and IL-6 [23], and

others, are induced by high levels of glucose in vitro and in vivo, and high concentrations of these mediators are detected in vitreous or plasma specimens from patients with diabetic retinopathy. Based upon the release profiles of these mediators from pericytes, it was speculated that TNF-a and IL-1b are initially released and trigger the release of intercellular adhesion molecule-1 (ICAM-1) and sVCAM-1, which affect leukostasis, and VEGF, GDNF, and IL-6, which induce vascular permeability during the initial stage of the diabetic retinopathy [24].

In this chapter, to get a better understanding of the pathophysiological roles of glial cell–derived cytokines in the diabetic retinopathy, we focus on the structural and functional aspects of the BRB and its modulation by cytokines derived from glial cells under pathological conditions at an early phase of diabetic retinopathy. In addition, we also describe the possible treatment and prevention of the retinopathy with retinoic acid analogue that affects the glial cell–derived cytokines.

STRUCTURAL AND FUNCTIONAL ASPECTS

OF THE BLOOD-RETINAL BARRIER (BRB)

The BRB Functional Unit Composed of Glial and Endothelial Cells

The endothelial cells of the BRB form near continuous sheets because of impermeable tight junctions (Fig. 1B). They also show no fenestration and few pinocytic vesicles. These distinctive features of the BRB-forming endothelial cells from capillary endothelial cells in other tissues maintain unique microenvironment essential for functions of retinal cells. Thus the tight junction of the BRB-forming endothelial cells is a substantial barrier that strictly regulates the paracellular pathways between the cells. Another unique feature of the endothelial cells that form the BRB and the BBB is that the capillaries forming the barrier are almost all ensheathed by vascular feet of astrocytes [25]. The anatomical relationship between the endothelial cells and astrocytes has prompted some of the researchers to explore a functional relationship between these cells. In fact, the characteristics of endothelial cells of the BBB are induced using chick-quail transplantation.

The transplantation of astrocytes into the avascular space of the anterior eye chamber showed that the capillaries that invaded the chamber were similar in characteristics to the BBB-forming endothelial cells. In vitro, a combination of an astrocyte-conditioned medium and cAMP made a tight junction resistant to the paracellular passage [26–28]. Since paracellular passage between the endothelial cells, mostly provided by the structural organization of tight junctions, astrocytes are strongly suggested to secrete some mediators that regulate paracellular passage, in terms of regulation of the tight junctions between the endothelial cells [29]. These findings strongly suggest an important insight into the permeability of the BBB between endothelial cells and astrocytes. In other words, it is feasible that astrocytes regulate the barrier function of the BRB, in terms of impermeability, in a paracrine manner.

Tight Junctions Between Endothelial Cells Are Substantial Barrier of the BRB

Tight junctions, the most apical component of intercellular junctional complexes, separate the apex from the basolateral cell surface domains to establish cell polarity (performing the function of a fence) [26–29] (Fig. 2). Tight junctions also possess a barrier function, inhibiting the flow of solutes and water through the paracellular space [26–29].

Fig. 2. Glial cell as a main component of BRB. (A) Schematic presentation of BRB. Note that cytoplasm of glial cell associates both with neural cell and capillary endothelium (open circles). (B) BRB is a biological unit comprised of specialized endothelial cells firmly connected by intercellular TJs and the endothelium-surrounding glial cells. Glial cell–derived cytokines such as VEGF and GDNF closely associate with the vascular integrity, which is regulated by modulating the TJ function of capillary endothelium in a paracrine manner. (C) BRB-forming glial cell expresses GDNF in the murine retina. Glial cell is highlighted by red in the retina, which is stained with anti-GFAP, a specific marker for glial cell in central nervous system and retina (a, left panel). GDNF expression shows similar distribution in green, suggesting that glial cell expresses GDNF protein (B, right panel).

They form a particular netlike meshwork of fibrils created by the integral membrane proteins, occludin and claudin, and members of the Ig superfamilies JAM and CAR [30]. Several peripheral membrane proteins related to tight junctions, such as ZO-1, ZO-2, ZO-3, 7H6 antigen, cingulin, symplekin, Rab3B, Ras target AF-6, and ASIP, an atypical protein kinase C-interacting protein, have been reported [3, 25, 31]. Recently, a new integral membrane protein tricellulin was also identified at tricellular contacts, which consist of three epithelial cells and have a barrier function [32].