myelin-basic-protein and Ocular-Hypertension

Axonal transport blockade is an initial step in retinal ganglion cell (RGC) degeneration in glaucoma and targeting maintenance of normal axonal transport could confer neuroprotection. We present an objective, quantitative method for assessing axonal transport blockade in mouse glaucoma models. Intraocular pressure (IOP) was elevated unilaterally in CD1 mice for 3 days using intracameral microbead injection. Longitudinal sections of optic nerve head (ONH) were immunofluorescently labeled for myelin basic protein (MBP) and amyloid precursor protein (APP), which is transported predominantly orthograde by neurons. The beginning of the myelin transition zone, visualized with the MBP label, was more posterior with elevated IOP, 288.8 ± 40.9 μm, compared to normotensive control eyes, 228.7 ± 32.7 μm (p = 0.030, N = 6 pairs). Glaucomatous regional APP accumulations in retina, prelaminar ONH, unmyelinated ONH, and myelinated optic nerve were identified by objective qualification of pixels with fluorescent intensity greater than the 97.5th percentile value of control eyes (suprathreshold pixels). This method segregated images with APP blockade from those with normal transport of APP. The fraction of suprathreshold pixels was significantly higher following IOP elevation than in normotensive controls in the unmyelinated ONH and myelinated nerve regions (paired analyses, p = 0.02 and 0.003, respectively, N = 12), but not in retina or prelaminar ONH (p = 0.91 and 0.08, respectively). The mean intensity of suprathreshold pixels was also significantly greater in glaucoma than in normotensive controls in prelaminar ONH, unmyelinated ONH and myelinated optic nerve (p = 0.01, 0.01, 0.002, respectively). Using this method, subconjunctival glyceraldehyde, which is known to worsen long-term RGC loss with IOP elevation, also produced greater APP blockade, but not statistically significant compared to glaucoma alone. Systemic losartan, which aids RGC axonal survival in glaucoma, reduced APP blockade, but not statistically significant compared to glaucoma alone. The method provides a short-term assessment of axonal injury for use in initial tests of neuroprotective therapies that may beneficially affect RGC transport in animal models of glaucoma.

Topics: Amyloid beta-Protein Precursor; Animals; Antihypertensive Agents; Axonal Transport; Axons; Disease Models, Animal; Female; Fluorescent Antibody Technique, Indirect; Glyceraldehyde; Intraocular Pressure; Losartan; Mice; Myelin Basic Protein; Nerve Fibers, Myelinated; Nerve Fibers, Unmyelinated; Ocular Hypertension; Optic Disk; Optic Nerve; Tonometry, Ocular

In the current study, it was aimed to evaluate the changes in myelinated and unmyelinated nerve fibers in retinal ischemia‑reperfusion injuries caused by acute ocular hypertension and to determine the sequence of these changes. Adult healthy New Zealand white rabbits were randomized to the hemodynamic group [n=12; used to determine the optimal intraocular pressure (IOP) for the subsequent experiments] and the hypertension group (n=6; 70‑mmHg hypertension induced in one eye). IOP was adjusted using a cannula and saline. Doppler ultrasound was used to measure the velocity of the optic artery under different intraocular pressures. Immunohistochemistry for myelin basic protein (MBP) was performed. Apoptosis of retinal cells was detected by terminal deoxynucleotidyl transferase biotin‑dUTP nick end labeling (TUNEL) assay. Electron microscopy was used to investigate the changes in myelinated and unmyelinated nerve fibers. IOP of the hypertension eyes was maintained at 70.2±1.0 mmHg, while IOP of control eyes was 7‑14 mmHg. Doppler ultrasound demonstrated an obvious decline of peak systolic velocity and an increase of resistance index of retinal bloodstream under a 70‑mmHg IOP. MBP immunohistochemistry and electron microscopy demonstrated obvious injuries to the myelin fibers. TUNEL indicated a significantly higher apoptosis rate in the hypertension eyes compared with control eyes. The apoptosis rate of retinal ganglion cells and bipolar cells in unmyelinated regions was higher than in myelinated regions. In conclusion, an IOP of 70 mmHg led to incomplete retinal ischemia but was the threshold for retinal ischemia, leading to obvious injuries to the myelin fibers.

Topics: Acute Disease; Animals; Disease Models, Animal; Intraocular Pressure; Male; Microscopy, Electron; Myelin Basic Protein; Nerve Fibers, Unmyelinated; Ocular Hypertension; Optic Nerve; Rabbits; Retina; Ultrasonography, Doppler

The aim of this study was to gain more information about the possible immunological mechanisms in glaucoma. We analyzed the complex autoantibody patterns against human optic nerve antigens in sera of patients with glaucoma and tried to identify important antigens.. Sera of 133 patients were included: healthy control subjects (n = 44), primary open-angle glaucoma (n = 44), and normal tension glaucoma patients (n = 45). The sera were tested against Western blots of human optic nerve, and antibody bands were visualized with chloronaphthol. IgG antibody patterns were analyzed by multivariate statistical techniques, and the most significant antigens were identified by mass spectrometry (Maldi-TOFTOF).. All subjects, even healthy ones, showed different and complex antibody patterns. Glaucoma groups showed specific up- and down-regulations of antibody reactivities compared to the control group. The multivariate analysis of discriminance found significant differences (P < 0.05) in IgG antibody profiles against human optic nerve antigens between both glaucoma groups and healthy subjects. The identified antigens include: myelin basic protein (up-regulated in the POAG group), glial fibrillary acidic protein (down-regulated in the glaucoma groups), and vimentin (down-regulated in the glaucoma groups in comparison to controls).. Using human optic nerve antigen, we were able to demonstrate that complex IgG autoantibody patterns exist in sera of patients with glaucoma. Large correlations between the given and our previous studies using bovine optic nerve antigens could be seen. Furthermore, anti-myelin basic protein antibodies, which can also be detected in patients with multiple sclerosis, were found in sera of glaucoma patients.

Topics: Aged; Amino Acid Sequence; Autoantibodies; Autoantigens; Blotting, Western; Female; Glaucoma, Open-Angle; Glial Fibrillary Acidic Protein; Humans; Male; Middle Aged; Molecular Sequence Data; Myelin Basic Protein; Nerve Tissue Proteins; Ocular Hypertension; Optic Nerve; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Transcription Factors; Vimentin

To investigate the antigenic specificity of the immune neuroprotective mechanism that can protect retinal ganglion cells (RGCs) against death caused by high intraocular pressure (IOP).. A unilateral increase in IOP was induced in rats by argon laser photocoagulation of the episcleral veins and limbal plexus. Rats with high IOP were immunized with glatiramer acetate (Cop-1, a synthetic copolymer) or with myelin-derived or uveitogenic peptides. When the steroid drug methylprednisolone was used, it was administered intraperitoneally every other day for 12 days.. Vaccination with myelin-derived peptides that reside in the axons failed to protect RGCs from death caused by high IOP. In contrast, IOP-induced RGC loss was reduced by vaccination with R16, a peptide derived from interphotoreceptor retinoid-binding protein, an immunodominant antigen residing in the eye. The benefit of protection against IOP-induced RGC loss outweighed the cost of the monophasic experimental autoimmune uveitis (EAU) that transiently developed in a susceptible rat strain. Treatment with methylprednisolone alleviated the disease symptoms, but caused further loss of RGCs. Cop-1 vaccination was effective in both EAU-resistant and EAU-susceptible strains.. To benefit damaged neurons, immune neuroprotection should be directed against immunodominant antigens that reside in the site of damage. In a rat model of high IOP, RGCs can benefit from vaccination with peptides derived from proteins that are immunodominant in the eye but not from myelin-associated proteins. This suggests that the site of primary degeneration in IOP-induced RGC loss is in the eye. Cop-1 vaccination apparently circumvents the site-specificity barrier and provides protection without risk of inducing autoimmune disease.

Topics: Animals; Autoimmune Diseases; Cell Survival; Cytoprotection; Disease Models, Animal; Eye Proteins; Glaucoma; Immunodominant Epitopes; Intraocular Pressure; Male; Methylprednisolone; Myelin Basic Protein; Ocular Hypertension; Oligopeptides; Rats; Rats, Inbred Lew; Rats, Sprague-Dawley; Retinal Ganglion Cells; Retinol-Binding Proteins; Uveitis; Vaccination; Vaccines, Synthetic

Our group recently demonstrated that autoimmune T cells directed against central nervous system-associated myelin antigens protect neurons from secondary degeneration. We further showed that the synthetic peptide copolymer 1 (Cop-1), known to suppress experimental autoimmune encephalomyelitis, can be safely substituted for the natural myelin antigen in both passive and active immunization for neuroprotection of the injured optic nerve. Here we attempted to determine whether similar immunizations are protective from retinal ganglion cell loss resulting from a direct biochemical insult caused, for example, by glutamate (a major mediator of degeneration in acute and chronic optic nerve insults) and in a rat model of ocular hypertension. Passive immunization with T cells reactive to myelin basic protein or active immunization with myelin oligodendrocyte glycoprotein-derived peptide, although neuroprotective after optic nerve injury, was ineffective against glutamate toxicity in mice and rats. In contrast, the number of surviving retinal ganglion cells per square millimeter in glutamate-injected retinas was significantly larger in mice immunized 10 days previously with Cop-1 emulsified in complete Freund's adjuvant than in mice injected with PBS in the same adjuvant (2,133 +/- 270 and 1,329 +/- 121, respectively, mean +/- SEM; P < 0.02). A similar pattern was observed when mice were immunized on the day of glutamate injection (1,777 +/- 101 compared with 1,414 +/- 36; P < 0.05), but not when they were immunized 48 h later. These findings suggest that protection from glutamate toxicity requires reinforcement of the immune system by antigens that are different from those associated with myelin. The use of Cop-1 apparently circumvents this antigen specificity barrier. In the rat ocular hypertension model, which simulates glaucoma, immunization with Cop-1 significantly reduced the retinal ganglion cell loss from 27.8% +/- 6.8% to 4.3% +/- 1.6%, without affecting the intraocular pressure. This study may point the way to a therapy for glaucoma, a neurodegenerative disease of the optic nerve often associated with increased intraocular pressure, as well as for acute and chronic degenerative disorders in which glutamate is a prominent participant.

Topics: Adjuvants, Immunologic; Adoptive Transfer; Amino Acid Sequence; Animals; Cell Death; Glatiramer Acetate; Glaucoma; Glutamic Acid; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Molecular Sequence Data; Myelin Basic Protein; Myelin Proteins; Myelin-Associated Glycoprotein; Myelin-Oligodendrocyte Glycoprotein; Ocular Hypertension; Peptides; Rats; Rats, Inbred Lew; Retinal Ganglion Cells; T-Lymphocytes; Vaccination