THE ROLE OF NEUROTROPHINS IN GLAUCOMA (BDNF)

 

• The neurons of the central nervous system, and by extension, the fibers of the retinal ganglion cells/optic nerve, are incapable of regeneration. In this regard, glaucoma can be regarded as a neurodegenerative disorder, much like Alzheimer’s or Parkinson’s diseases.

 

• There are a number of concepts which have been propounded for the development of glaucoma. These range from the mechanical to the vascular, biochemical and intraluminal pressure theories among others. Certain risk factors have also been enumerated regarding causation of glaucoma. Among them intra-ocular pressure, increasing age, genetic background, thinner central corneal thickness, vascular dysregulation, such as Flammer’s syndrome, certain systemic diseases such as diabetes and hypertension and also ocular diseases such as myopia and iridocorneal disorders are prominent.

• It is likely that in an individual, several molecular pathways converge and induce retinal ganglion cell (RGC) loss. The cellular, molecular, hormonal signals that promote RGC death in glaucoma are probably accentuated by risk factors, tilting the neuron’s fate towards dysfunction and demise.

• In this post, we take a look at NEUROTROPHINS, and their role in glaucoma.

• Neurotrophins are diffusible trophic molecules that exert a potent survival effect on adult CNS neurons undergoing degeneration. They are a family of small, secreted peptides that include nerve growth factor (NGF), brain derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and neurotrophin-4/5 (NT-4/5) in mammals.

• Apart from cell survival, Neurotrophins mediate several key cellular responses in the developing and mature CNS including proliferation, differentiation, axon growth, as well as dendrite and synapse formation.

• Among the Neurotrophins BDNF is an important factor, because of its role in cell survival. BDNF is strongly expressed in the superior colliculus, and it is retrogradely transported by RGC axons to the retina. Within the retina, BDNF is produced by cells in the ganglion cell layer and inner nuclear layer. Upregulation of BDNF is an early response to axonal injury.

• The biological effects of neurotrophins are mediated by two classes of cell surface receptors:

• i) the tropomyosin related kinase (Trk) family of receptor tyrosine kinases comprising TrkA, the receptor for NGF; TrkB, the receptor for BDNF and NT-4/5; and TrkC, the receptor for NT-3.

• ii) the p75 receptor (p75NTR) which binds all Neurotrophins.

• Activation of Trk receptors is typically associated with cell survival, while p75NTR can stimulate both survival and apoptotic pathways.

• It is now widely accepted that neurotrophic factors promote neuronal survival by inhibiting default apoptotic pathways.

• One way of looking at glaucoma is the slowing down of axonal transport in this disease. As a result, pro-cell survival neurotrophic factor levels and availability is reduced, leading to progressive RGC loss. Supplementation of neurotrophic factors could be a potential strategy to prevent the death of injured RGCs.

• Intraocular injection of exogenous BDNF protein or viral-mediated BDNF gene transfer using adenovirus or adeno-associated virus (AAV) promote robust RGC survival after optic nerve transection or crush.

• The combination of BDNF gene transfer with additional therapies including free radical scavengers and cell-permeable cAMP further increase RGC neuroprotection.

• Paradoxically, in spite of its robust survival effect, BDNF does not stimulate regrowth of injured RGC axons. Studies have reported no significant RGC axon regeneration beyond the lesion site following exogenous BDNF administration. The effect of exogenous BDNF is also temporary. It delays but does not prevent cell death. This short-term activity of BDNF is attributed to the variable level of BDNF receptor TrkB expression in RGCs following injury. This limits the intrinsic capacity of these neurons to respond to neurotrophin stimulation. Unsurprisingly, TrkB gene transfer combined with exogenous BDNF administration markedly increased RGC survival. 76% of RGCs were alive at two weeks after axotomy compared to 10% of neurons that remained in control eyes (66% protection).

 

STRUCTURE VS. FUNCTION DEBATE

 

Ophthalmologists often debate the advantage of structural versus functional studies to diagnose glaucoma. There have also been efforts to combine the two and come up with a structure-function index. The crux of these observations is the need to develop mechanisms which would identify glaucomatous changes at the earliest. Since, a sub-group of patients may manifest structural changes earlier than functional changes and vice-versa, (“the structure-function disassociation”) it has become controversial as to which method is preferable.

The need to develop a singular diagnostic method utilizing structural and functional tests is based on the above mentioned probability of structure-function disassociation. Since, one type of test may be normal in an individual, it is required to perform another type of test to confirm the diagnosis. According to Elliot Kirsten, “Our efforts to coalesce ocular physical measurements with functional values seem to point to our desire to define glaucoma in one simple value. While the effort itself has already shed great benefits to our understanding of this complex disease, it ultimately seems unlikely that we will ever be able to adequately define glaucoma in terms of a singular value”.

A structural measure for glaucoma assessment refers to measurements of the neuroretinal rim, retinal nerve fibre layer (RNFL) and the ganglion cell layer in the macula. The initial methods to study glaucomatous changes at the disc and parapapillary retina started with the development of the direct ophthalmoscope by Hermholtz in 1851. Using this instrument, ophthalmologists were able to study the optic disc and detect changes in its structure. Usually, ophthalmologists resorted to diagrams to denote the cup, disc and other changes. This is still used in a large number of practices. However, these findings are prone to intra- and inter-observer differences. Fundus photography, especially stereo-photographs have a better chance of being consistent among observers. Newer digital platforms are available to capture the images. Study of the disc with instruments such as GDx and Heidelberg Retinal Tomograph (HRT) have brought another dimension to these methods. Unfortunately, these instruments are expensive and their cost-benefit efficiency is probably limited when regarded as a whole in glaucoma diagnostic techniques.

The development of optical coherence tomography (OCT) provided us a vast data set, allowing us to seek out specific retinal measurements that are most highly specific for the diagnosis and observation of glaucomatous progression. OCT provides information regarding the disc, surrounding RNFL and other layers of the retina. Advanced segmentation techniques are giving us important and precise information of the layers most affected in glaucoma. OCT-angiography is also being used to demonstrate areas of retinal loss. Another technique being developed is the Detection of Apoptosing Retinal Cells (DARC). This technique pin-points the apoptosing retinal ganglion cells and gives a true picture of cellular damage in glaucoma patients.

Functional tests are largely based on visual field analysis. Other tests such as Visual Evoked Potential (VEP), pattern electroretinography (pERG), and Relative Afferent Pupillary Defect (RAPDx) have not been much used in common clinical practices and are usually limited to research purposes. The first record of a visual field defect is found in Hippocrates’ description of a hemianopia from the late fifth century B.C. Later, Ptolemy (150 B.C.) quantified the visual field and noted its circular form. Perimetry has now advanced in terms of better hardware and software.

Standard automated perimetry is still the gold standard for detecting functional change. A post-hoc analysis of the Early Manifest Glaucoma Trial (EMGT) by Öhnell et al. has shown that after a median follow-up of 8 years, eyes with a visual field defect at baseline were more than 4 times more likely to have progression first detected as further worsening of the visual field (VF) compared with progression on optic disc photographs. However, since the EMGT was an old study, the findings may not be accurate in the current scenario. Newer imaging systems are providing better documentation of RNFL thinning compared to photographs.

Thus, the question being raised is whether structure or functional tests are more reliable. It is to be highlighted that we need both set of tests to diagnose the complex disease called glaucoma. It is not the deficiency of either type of test in detecting glaucomatous changes, but it is the varied pathogenesis of glaucoma in different individuals which produces this structure-function disassociation. Hence, we may conclude by this sobering thought that no single technique is presently available to diagnose glaucoma with significant sensitivity and specificity in all individuals. 

 

PROSTAGLANDIN ANALOGS: MECHANISM OF ACTION

 

INTRODUCTION:

Anti-glaucoma medications reduce intra-ocular pressure (IOP) by their effects on aqueous humor dynamics.

These agents act by:

• Slowing the production rate of aqueous humor
  
• Decreasing the resistance to flow through the trabecular meshwork
  
• Increasing drainage through the uveoscleral outflow pathway
  
• Or by a combination of these mechanisms

Prostaglandin (PG) F2α analogs reduce IOP by stimulation of aqueous humor drainage primarily through the uveoscleral outflow (non-conventional) pathway. Minor effects on trabecular (conventional) pathway have been reported. Based on most studies, the PG effect on episcleral venous pressure is minimal.

EFFECT ON CONVENTIONAL AQUEOUS OUTFLOW PATHWAY:

Effects on trabecular outflow (Conventional pathway) facility also have been reported. Most studies have found a small (10–15%) increase that may or may not be statistically significant and is not clinically important. Histological analysis of latanoprost-treated anterior segments showed focal loss of Schlemm's canal endothelial cells, separation of inner wall cells from the basal lamina, cell disconnection from the extracellular matrix, and focal loss of extracellular matrix in the juxtacanalicular region.

Studies on EP receptor stimulation has shown that EP2 and EP4 activation results in increased cell contractility of the trabecular meshwork, and decreased cell contractility of the inner wall of Schlemm's canal, mediating IOP through the conventional pathway.

Outflow through the conventional pathway probably does not contribute to any reduction in IOP but an increase in aqueous flow could be considered a healthy side-effect of topical PG analogs because aqueous humor carries essential nutrients and removes waste products, crucial for keeping the avascular tissues of the anterior segment healthy.

EFFECT ON NON-CONVENTIONAL AQUEOUS OUTFLOW PATHWAY:

• Bimatoprost and Latanoprost increase uveoscleral outflow in ocular normotensive and hypertensive subjects. 
• Travoprost increased uveoscleral outflow in monkeys and marginally increased it in ocular hypertensive patients as well.
• Unoprostone, the weakest of the four prescribed PG analogs, is the only one that did not affect uveoscleral outflow in humans despite 5 days of twice-daily dosing.

(For more information on Unoprostone please follow this link:  https://ourgsc.blogspot.com/search?q=unoprostone )

A significant increase in aqueous flow was found at night in young healthy Japanese volunteers treated with Latanoprost, and during the day and at night in healthy predominantly Caucasian volunteers treated with bimatoprost.

 

Prostaglandin analogues elicit their effect by binding to specific receptors localized in the cell membrane and nuclear envelope.

There are 9 prostaglandin receptors: PGE receptor 1–4 (EP1–4), PGD receptor 1–2 (DP1–2), PGIP receptor, PGFP receptor, and thromboxane A2 receptor (TP), their designation based mainly on the prostaglandin for which binding is most specific.

PGF2α binds the FP, EP1, and EP3 receptors with significant affinity, while travoprost binds the FP receptor with highest affinity among the prostaglandin analogues, with minimal affinity for DP, EP1, EP3, EP4, and TP receptors. Pharmacologic and pharmacokinetic data suggest the existence of a unique bimatoprost receptor, distinct from the known FP receptors; however, this receptor is yet to be cloned.

Studies in mice suggest that FP and EP3 are the primary receptors that trigger downstream signaling pathways and the eventual physiologic response following treatment with latanoprost, bimatoprost, and travoprost.

However, in primates, EP2 receptor stimulation has been shown to increase uveoscleral outflow, and EP4 receptor activation reduces IOP by increasing outflow facility without effecting uveoscleral outflow. These results in mice and primates suggest that species-specific mechanisms may exist.

In the ciliary muscle, binding of prostaglandins and prostaglandin analogues to ciliary muscle FP receptors disrupts extracellular matrix turnover. PGF2α and prostaglandin analogues bind to EP and FP receptors in the ciliary muscle, resulting in ciliary muscle relaxation and increased aqueous humor outflow.

Matrix metalloproteinases (MMPs) degrade and remodel the extracellular matrix in the ciliary muscle, iris root, and sclera, reducing outflow resistance to fluid flow. The rate of turnover of the extracellular matrix is dependent on the balance between the molecules that degrade and remodel the extracellular matrix i.e. the MMPs, and their inhibitors [tissue inhibitor of metalloproteinase (TIMPs)].

Treatment with PGF2α and prostaglandin analogues increases the amount of MMPs, while maintaining TIMP expression. This shifts the balance in favor of degradation and remodeling of the extracellular matrix to enhance outflow facility.

Increase in uveoscleral outflow occurs through various mechanisms:

• Remodeling of the extracellular matrix of the ciliary muscle, and sclera causing changes in the permeability of these tissues;
• Widening of the connective tissue-filled spaces among the ciliary muscle bundles, which may be caused in part by relaxation of the ciliary muscle;
• Changes in the shape of ciliary muscle cells as a result of alterations in actin and vinculin localization within the cells.

Remodeling of the extracellular matrix within the ciliary muscle and sclera is the most thoroughly understood effect of PG treatment. Dissolution of collagen types I and III within the connective tissue-filled spaces between the outer longitudinally oriented muscle bundles results from PG-stimulated induction of enzymes MMP1, 2, and 3 in the ciliary muscle and surrounding sclera.

PGF2α- and latanoprost-induced secretion and activation of MMP-2 in ciliary muscle cells were shown to occur via protein kinase C and extracellular signal regulated protein kinase 1/2-dependent pathways.

Inhibition of the latanoprost-induced reduction of IOP in rats by thalidomide suggested that the IOP-lowering response is mediated, in part, through tumor necrosis factor-α-dependent signaling pathways.

PGF2α-isopropyl ester treatment was found to increase MMP-1, -2, and -3 in the sclera, which contributes to outflow.

Studies in FP receptor-deficient mice have shown that the FP receptor is essential for the early IOP lowering response to topical latanoprost, travoprost, bimatoprost, and unoprostone. The involvement of the FP receptor in the IOP reduction with long-term dosing is unknown.

Prostaglandins also alter the production of MMPs in human primary trabecular meshwork cells.

Prostaglandin analogues lower IOP through tissue impedance changes and long-term remodeling of the extracellular matrix within the conventional and unconventional outflow pathways. However, this does not explain the early effects of prostaglandin analogue treatment in cell culture models. IOP was found to be lowered within 2 h of treatment in mice and human anterior segment culture.

SOURCES:

1. Carol B. Toris, B’Ann T. Gabelt, and Paul L. Kaufman. Update on the Mechanism of Action of Topical Prostaglandins for Intraocular Pressure Reduction. Surv Ophthalmol. 2008; 53(SUPPL1): S107–S120. doi:10.1016/j.survophthal.2008.08.010.
2. Winkler NS, Fautsch MP. Effects of prostaglandin analogues on aqueous humor outflow pathways. J Ocul Pharmacol Ther. 2014;30(2-3):102-109. doi:10.1089/jop.2013.0179.