Ординатура / Офтальмология / Английские материалы / The Retinal Muller Cell Structure and Function_Sarthy, Ripps_2001

tional diversity of ECM molecules is further broadened by their interactions with other kinds of regulatory molecules. Many cell adhesion molecules like NCAM interact with the ECM. Growth factors such as b-FGF bind to the ECM, and their activity and stability appears to be regulated by interactions with the ECM (Hall and Schachner, 1998).

In the developing nervous system, ECM molecules play a vital role in neuronal migration and axonal growth (Reichardt and Tomaselli, 1991; Letourneau et al., 1994). Many ECM molecules are found in both developing and adult retina (Table 2.2). However, neither the function nor the cellular origin of these molecules is well studied. The ECM molecules are generally prominent at the ILM and could be derived from ganglion cells, astrocytes, Müller cells, or the retinal vasculature, all of which abut or are in close proximity to the ILM. Müller cells appear to be a likely source for some of these molecules. However, in situ hybridization studies demonstrate Müller cells do not synthesize either laminin B1 mRNA or collagen lV mRNA during development or in the adult mouse retina (Fig. 2.13; Sarthy et al., 1991; Sarthy, 1993). Whether other ECM molecules such as merosin, vitronectin, and thrombospondin present at the ILM are derived from Müller cells remains to be established. On the other hand, two inhibitory ECM molecules, EAP-300 (embryonic avian polypeptide of 300 kDa) and clausterin (a 320 kDa keratan sulfate proteoglycan), are expressed by Müller cells in the developing retina (McCabe and Cole, 1992). EAP-300 and clausterin may be involved in retinal stratification (McCabe and Cole, 1992).

As described previously, cells interact with the extracellular matrix via the integrin family of cell surface receptors. Integrins are heterodimers made up of α and β subunits each of which belongs to a large subfamily (Reichardt and Tomiselli, 1991; Powell and Kleinman, 1997). Müller cells have been reported to express some integrins (Elner and Elner, 1997). Although we know very little about integrin function in Müller cells, we might speculate that integrins mediate adhesion of the Müller cell end feet to the vitreous collagen (see Chapter 1) , and may also be involved in Müller cell migration into epiretinal membranes (Elner and Elner, 1997).

2.2.5.Growth and Neurotrophic Factors

It is widely recognized that growth and neurotrophic factors serve as important signaling molecules in development of the nervous system. Many of the factors present in the developing CNS are also found in the developing retina (Tanihara et al., 1997) (Table 2.3). Some factors such as TGF-α regulate cell proliferation while others appear to promote neuronal differentiation and survival (Harris, 1997). Indeed, there is growing evidence that

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Figure 2.13. Immunolocalization and in situ hybridization data for laminin B1 expression in developing mouse retina. Immunostaining shows that laminin is present at the internal limiting membrane in embryonic retina at E12 (A), E-15 (B), and E-20 (C). It is also present, albeit in a smaller quantity, in the adult retina. In situ hybridization data show that laminin B1 mRNA is not present in Müller cells at P3 (D), P-7 (E), or adult retina (F). The data show that laminin is highly expressed at the INL but the site of laminin synthesis is not the Müller cell. Arrows in A, B, C show internal limiting membrane. Arrowhead, RPE. Nb, neuroblast layer; le, lens; and r, retina. Arrows in D, E, F show ganglion cell bodies (Sarthy and Fu, 1990). (Copyright 1990 The Rockefeller University Press, reprinted with permission.)

Müller cells secrete extrinsic factors that can influence neuronal differentiation and survival.

One of the best examples comes from the studies of Neophytou and associates (1997) who observed that serum had a dramatic effect on rod cell differentiation in cultures of neonatal mouse retina. In medium containing 10% fetal calf serum (FCS), very few rods were observed whereas in cultures

R, results with retina; C, data from retinal cell cultures.

with little serum, a large number of rods were found. Moreover, it appeared that when serum was present in the medium, rod development was somehow arrested. Further studies showed that the serum effect was indirect. It was found that serum acted by stimulating Müller cells in the culture to proliferate and release the cytokine, leukemia inhibitory factor (LIF), which in turn arrested rod development (Fig. 2.14). Similar experiments in rat retinal cultures indicate that ciliary neurotrophic factor (CNTF) and LIF inhibit rod development by redirecting rod-precursors to become bipolar cells (Kirsch et al., 1996; Ezzedine et al., 1997). Although these findings point to the exciting possibility that Müller cells regulate rod differentiation, it remains to be shown that CNTF/LIF released by Müller cells has an inhibitory effect in vivo.

There are other situations where Müller cellderived extrinsic factors appear to promote neuronal survival. Müller cell-conditioned medium has

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Figure 2.14. The inhibitory effect of Müller cells on rod differentiation. (A) The histograms show the number of rhodopsin+ cells (rods) is decreased in retinal cultures that contain fetal calf serum (FCS) or Müller cells. (B) There is a dramatic decrease in the number of rods in Müller cell-conditioned medium (MCM) but not in 3T3 conditioned medium (3T3CM).

(C) the suppression of rod generation in FCS can be mimicked by adding transforming growth factor-α (TGFα) which is a known Müller cell mitogen. It appears that inhibition of rod generation by FCS is an indirect effect, and arises from proliferation of Müller cells in FCS containing culture medium. Also, the conditioned medium effect could be blocked by antibodies to leukemia inhibitory factor (data not shown) which suggests that LIF released by Müller cells inhibits rod generation (Neophytou et al., 1997). (Copyright 1997 Company of Biologists Ltd., reprinted with permission.)

been reported to support the survival of ganglion cells (Sarthy et al., 1985; Armson et al., 1987), and recent studies show that a combination of growth factors and neuronal stimulation can mimic the effects of the Müller cellconditioned medium on ganglion cell survival (Meyer-Frank et al., 1995). This exciting observation provides good evidence that Müller cells may produce growth factors that are important for long-term neuronal survival in the retina.

2.2.6. Retinoic Acid

Retinoic acid (RA) has long been recognized as a morphogenetic factor in the developing retina (McCaffrey et al., 1991; Hyatt et al., 1996; Hyatt and Dowling, 1997). Treatment of zebrafish with RA stimulates precocious rod differentiation, and conversely, inhibition of RA synthesis in the eye leads to retarded rod development (Hyatt et al., 1996). In dissociated rat retinal cultures, RA application results in an increase in the number of progenitors that develop as rods (Stenkamp et al., 1993; Kelly et al., 1994). In agreement with these observations, the retinas of double null mice lacking the retinoic acid receptors β2 and γ2 are thinner, and show limited photoreceptor differentiation (Grondona et al., 1996).

The cellular sources of retinoic acid in the developing retina have yet to be identified. Recent studies suggest that Müller cells may be a major source

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are present in the retina before day 15, but the lack of circulating glucocorticoids prevents induction of glutamine synthetase. Moreover, glutamine synthetase induction in this system appears to accompany Müller cell differentiation.

Although the molecular mechanisms underlying glutamine synthetase induction are complex and not completely understood, experimental evidence suggests that Müller cell-specific expression is achieved through the concerted involvement of positive and negative cis elements, a glucocorticoid response element (GRE) and a neural-restrictive silencer (Zhang et al., 1993; Li et al., 1996; Avisar et al., 1999). In addition, it is quite clear that glucocorticoid receptors play an important role in this process (Zhang et al., 1993; Grossman et al., 1994; Gorovits et al., 1994). Results from a recent developmental study indicate that glutamine synthetase expression in chick retina is strongly dependent on the level of a 95 kDa glucocorticoid receptor (Gorovits et al., 1994). At early developmental stages, all retinal cells express the receptor, and consequently all cells express glutamine synthetase. At later developmental stages, however, receptor expression appears to be lost from all retinal cells except Müller cells. As a consequence, glutamine synthetase can be induced only in Müller cells (Fig. 2.17). Interestingly, transcription of the glucocorticoid receptor gene is strongly repressed by the c-Jun protein, which is expressed in proliferating neuroblasts (Vardimon et al., 1999).

Figure 2.18. Glutamate synthetase induction is dependent on cell contact. Northern blots show that GS mRNA induction is higher in cell aggregates and retina than in monolayer

cultures of Müller cells. Note that CAII and H3.3 levels do not change appreciably. The figure to the right shows a proposed model for glutamine synthetase induction in Müller cells (GLIA) (Moscona and Vardimon, 1988). (Copyright 1988 Wiley-Liss, Inc., a subsidiary of John Wiley & Sons, Inc., reprinted with permission.)

A

Proliferation

Migration

Differentiation

B

Mitosis

Inhibition

Gene

Regulation

Figure 2.19. Potential Müller cell–neuron interactions in developing retina. A. Influence of Müller cells on the proliferation, migration, and differentiation of neurons. These processes are mediated by growth factors, cell adhesion molecules, and extracellular matrix molecules; B. Neuronal influence on Müller cell activity. Neurons may regulate the mitosis and gene activity in glial cells.

Perhaps the most remarkable feature of the system is that contact interaction with neurons appears to be essential for glutamine synthetase induction in Müller cells. When retinal cultures containing both neurons and Müller cells are treated with glucocorticoids (cortisol), glutamine synthetase is strongly induced in Müller cells (Moscona, 1983). However, if Müller cells are grown in monolayer cultures in the absence of neurons, hormone treatment fails to induce glutamine synthetase (Fig. 2.18). Although the mechanism underlying this phenomenon is still not established, it appears that disruption of neuron–glia contact leads to activation of the c-Jun signaling pathway which in turn inhibits transcription of glucocorticoid receptor gene (Reisfeld and Vardimon, Vardimon et al., 1999). In the absence of the receptor, glutamine synthetase induction becomes nonresponsive to glucocorticoids.

Neuronal interaction is apparently not a universal requirement for gene expression in Müller cells (Fig. 2.19). Although glutamine synthetase induction is dependent on neuronal contact, synthesis of CA II, filamin, or CRALBP is not affected by the absence of neurons (Linser and Moscona, 1981; Hicks and Courtois, 1986; Lewis et al., 1988; Lemmon, 1986; Sarthy et al., 1998; Roque et al., 1997). Clearly, the expression of many Müller cellspecific proteins does not require contact with neurons.