Oxidized low-density lipoprotein, lipid and calcium aggregates reveal oxidative stress and inflammation in the conjunctiva of glaucoma patients.

PURPOSE: Conjunctival specimens from primary open-angle glaucoma (POAG), exfoliation glaucoma (ExG) patients and controls were histologically analysed for oxidized low-density lipoprotein (ox-LDL), lipid and calcium aggregates. Our goal was to use them as biomarkers of oxidative stress and inflammation and to evaluate their correlation with glaucoma and impact on surgical outcome. METHODS: Conjunctival samples were obtained from POAG (n = 14) and ExG (n = 17) patients and from control subjects (n = 11) operated for macular hole, retinal detachment or strabismus. Immunohistochemistry was performed using the antibody against ox-LDL. Lipids and calcium were analysed by histochemical stainings with Nile red and Alizarin red S, respectively. RESULTS: Immunoreaction for ox-LDL was significantly increased in POAG (p = 0.049) and the number of lipid aggregates was significantly higher in ExG (p = 0.009) when compared to control. When POAG and ExG patients were grouped according to the outcome of deep sclerectomy (DS) surgery, the number of lipid (p < 0.001) and calcium aggregates (p = 0.014) were significantly higher in the conjunctival stroma of patients whose surgery failed within a three-year follow-up period. CONCLUSIONS: The lipid-mediated alterations suggested the presence of oxidative stress and inflammation in the conjunctiva of glaucoma patients. The present data further support the role of oxidative stress and inflammation in the wound healing process leading to excessive scarring and failure in DS surgery.

Conjunctival matrix metalloproteinases and their inhibitors in glaucoma patients.

PURPOSE: Chronic conjunctival inflammation, caused by various reasons, for example long-term use of topical drugs and/or their preservatives, affects the outcome of glaucoma surgery by interfering with wound healing. Matrix metalloproteinases (MMPs) remodel extracellular matrix (ECM) and are involved in the wound healing process. This study was designed to evaluate the conjunctival expression of MMPs and their tissue inhibitors (TIMPs) in the normal eye, primary open-angle glaucoma (POAG) and exfoliation glaucoma (ExG) and whether there is an association between staining intensities and deep sclerectomy outcome. METHODS: Immunohistochemical procedures were performed on conjunctival samples which were obtained from POAG (n=11) and ExG (n=14) patients as well as normal (n=7) subjects. Antibodies against MMPs (MMP-1, -2, -3 and -9) and TIMPs (TIMP-1, -2 and -3) were used. RESULTS: In conjunctival stroma, expression levels of MMP-2 (p=0.047), MMP-3 (p=0.009), MMP-9 (p<0.001), TIMP-1 (p=0.003), TIMP-2 (p<0.001) and TIMP-3 (p<0.001) in ExG and MMP-9 (p=0.008), TIMP-2 (p=0.02) and TIMP-3 (p=0.002) in POAG were significantly increased compared to control. We further found correlations between expression of MMP-1 and MMP-3 and the length of pilocarpine treatment. CONCLUSION: The expression of MMPs and TIMPs is increased in the conjunctiva of POAG and ExG patients having a long history of topical antiglaucoma drops. Antiglaucoma agents and/or their preservatives alter the remodelling balance of ECM in conjunctiva of POAG and ExG eyes. The balance between MMPs and TIMPs may play a crucial role in the conjunctival wound healing process and the outcome of glaucoma surgery.

Conjunctival inflammatory cells and their predictive role for deep sclerectomy in primary open-angle glaucoma and exfoliation glaucoma.

PURPOSE: To investigate the conjunctival inflammatory alterations of patients with primary open-angle glaucoma (POAG) and exfoliation glaucoma (ExG) and correlate the findings with the success of deep sclerectomy (DS) surgery and with the patients' medical history. METHODS: Altogether 25 POAG and ExG patients of the prospective DS study were divided, based on the diagnosis and success of the operation, into 4 groups, POAG S (success), POAG F (failure), ExG S, and ExG F. Controls were obtained from other ophthalmologic surgery patients who did not have glaucoma, and their conjunctiva was examined to be normal. Inflammatory cell subtypes in the conjunctiva were identified and quantified by using immunohistochemistry and monoclonal antibodies: CD3 (T-lymphocyte marker), CD4 (T-helper lymphocyte marker), CD8 (T-cytotoxic lymphocyte marker), CD20 (pan-B cell marker), CD38 (plasma cell marker), CD45RA (naïve T-cell marker), and CD68 (macrophage marker). RESULTS: Higher numbers of inflammatory cells were found in the conjunctiva of the glaucoma patients on medical treatment compared with the normal conjunctiva of the controls. Moreover, T-lymphocytes, T-helper lymphocytes, T-cytotoxic lymphocytes, B cells, plasma cells, and macrophages were found in significantly higher numbers in patients in whom DS failed during the follow-up period of 2.5 years than those with surgical success. CONCLUSIONS: High numbers of cytotoxic and helper T-lymphocytes, plasma cells, and macrophages indicate a chronic inflammatory reaction in the conjunctiva of glaucoma patients. The chronic inflammation is most probably owing to the chronic topical treatment of the patients and seems to be a significant risk factor for DS surgery failure.

Mitomycin C-augmented deep sclerectomy in primary open-angle glaucoma and exfoliation glaucoma: a three-year prospective study.

PURPOSE: To investigate the efficacy and safety of mitomycin C (MMC)-augmented deep sclerectomy with implant (DSCI) in primary open-angle glaucoma (POAG) and exfoliation glaucoma (ExG) patients. METHODS: A total of 68 eyes of 68 patients with POAG and ExG were enrolled consecutively to undergo DSCI with MMC (0.4 mg/ml for 2 min). The intraocular pressure (IOP), number of antiglaucoma medications, neodymium:yttrium-aluminum-garnet (Nd:YAG) laser goniopuncture treatments and complications were compared postoperatively after 36- month follow-up. Surgery was considered as a complete success when IOP was <18 mmHg without antiglaucoma medication. RESULTS: Preoperatively the mean IOPs were 23 ± 6 mmHg and 25 ± 8 mmHg, and 13 ± 4 mmHg and 11 ± 4 mmHg in the POAG and ExG groups, respectively, at 36 months. At 36 months, 74% and 73% of surgeries were a complete success in the POAG and ExG group, respectively [not significant (NS)]. Two patients (8%) of the POAG group and one of the ExG group (3%) were receiving antiglaucoma medication at 36 months (NS). Nd:YAG laser goniopuncture was performed more often in the ExG group (87%) than in the POAG group (61%, p = 0.024). Postoperatively choroidal detachment occurred in 16% of eyes in the POAG group and in 11% of eyes in the ExG group (NS). CONCLUSIONS: DSCI with MMC augmentation appears to be as effective in patients with ExG and POAG in lowering IOP to target levels at medium term with few immediate postoperative complications.

Mitomycin-C-augmented deep sclerectomy in patients with primary open-angle glaucoma and exfoliation glaucoma: a 1-year prospective study.

PURPOSE: To investigate the efficacy and safety of mitomycin C (MMC)-augmented deep sclerectomy with implant (DSCI) in patients with primary open-angle glaucoma (POAG) and exfoliation glaucoma (ExG). METHODS: A total of 68 eyes of 68 patients with POAG and ExG were enrolled consecutively to undergo DSCI with MMC (0.4 mg/ml for 2 min). The intraocular pressure (IOP), number of antiglaucoma medications, neodymium:yttrium-aluminum-garnet (Nd:YAG) laser goniopunctures and complications were compared postoperatively. Surgery was considered as a complete success when IOP was < 18 mmHg without antiglaucoma medication. RESULTS: Preoperatively, the mean IOPs were 23.1 +/- 5.8 and 25.4 +/- 8.3 mmHg, and 13.8 +/- 6.1 and 11.2 +/- 5.6 mmHg in the POAG and ExG groups, respectively, at 12 months. 77.4% and 75.7% of surgeries were a complete success in the POAG and ExG groups, respectively [not significant (NS)]. Five patients (16.1%) in the POAG group but none in the ExG group (0%) were receiving antiglaucoma medication at 12 months (NS). Nd:YAG laser goniopuncture was performed in 29.0% of eyes in the POAG group and in 55.6% of eyes in the ExG group (p = 0.047). Postoperatively, choroidal detachment occurred in 16.1% of eyes in the POAG group and in 10.8% of eyes in the ExG group (NS). We encountered no serious complications related to MMC use. CONCLUSION: DS with MMC augmentation appears to be equally effective in ExG and POAG patients in lowering IOP to target levels, at least in the short term, with few immediate postoperative complications.

The effects of 5-fluorouracil on ocular tissues in vitro and in vivo after controlled release from a multifunctional implant.

PURPOSE: To evaluate the effects of 5-fluorouracil (5-FU) on ocular cells in vitro and the effects of degradable 5-FU-loaded poly(DL-lactide-co-glycolide; PDLGA) 50:50 implant in the rabbit eye in vivo. METHODS: Cytotoxicity was assessed with a tetrazolium salt WST-1 cell proliferation/viability test and a lactate dehydrogenase (LDH) leakage test in rabbit corneal stromal fibroblasts (SIRCs), bovine corneal endothelial cells (BCECs), human conjunctival epithelial cells (IOBA-NHCs), human retinal pigment epithelial cells (ARPE-19), and human corneal epithelial cells (HCECs). The 5-FU-loaded PDLGA implants were surgically placed in rabbit eyes with a deep sclerectomy technique and the histopathology of the eyes was examined. RESULTS: In vitro, 5-FU affected cell proliferation and survival in a time- and dose-dependent manner. In the WST-1 test, adverse effects in serum-free conditions started from 0.0005 mg/mL 5-FU in SIRCS and HCECs, whereas in other cell types, 0.005 mg/mL 5-FU hindered cell proliferation. In serum-free conditions 72-hour 5 mg/mL 5-FU treatment decreased cell viability to 40% in BCECs and to 10% to 15% in other cell types. 5-FU had no or very minor effects on LDH leakage. In vivo, the 5-FU implant showed no signs of toxicity in cornea and retina, whereas in the conjunctival stroma near the implantation site, some inflammatory cells and a marked subepithelial condensation of stromal connective tissue was observed during the postoperative period of 4 weeks. CONCLUSIONS: 5-FU had a broad therapeutic range, and the 5-FU implant showed only minor tissue reactions in conjunctiva at the surgical site. 5-FU is a possible candidate for controlled drug release.

Histopathology of the three implanted degradable biopolymers in rabbit eye.

New straightforward applications of new biopolymers are needed in glaucoma surgery. The aim of this study was to compare biocompatibility of three biomaterials in rabbit eyes after deep sclerectomy; a collagen implant (AquaFlow) represented the "gold standard". A blend of 85:15 poly(L-lactide-co-glycolide) and 70:30 poly(L-lactide-co-1,3-trimethylene carbonate) copolymers in a molar ratio of 70:30 (Bio-1 = Inion GTR membrane) and poly(DL-lactide-co-glycolide with molar compositions of 50:50 (Bio-2) and 85:15 (Bio-3) were inserted into rabbits eyes. Bio-1, Bio-2 or Bio-3 caused very mild eye irritation or tissue response which was comparable to that of the collagen implant. The biodegradation time of Bio-1, Bio-2, and Bio-3 implants was over one year, 3 months and 6 months, respectively. Implant mapping by Fourier transform infrared (FTIR) microscopy revealed a heterogeneous distribution of degradation products throughout Bio-1, Bio-2, and Bio-3. All implants were surrounded by a very fine tissue capsule which was not visible after total degradation of the implants. The FTIR spectrum of tissue capsule around Bio-1 was almost identical to that around Bio-2 whereas significant differences were observed in the spectrum of the tissue capsule around Bio-3. Despite some differences in tissue response, all tested implants represent biologically acceptable materials for drainage devices in glaucoma surgery.

Phospholipases A2 in normal human conjunctiva and from patients with primary open-angle glaucoma and exfoliation glaucoma.

BACKGROUND: Chronic situations like long-term use of topical medications induces conjunctival inflammation and is also a significant risk factor for failure of filtering surgery. We evaluated conjunctival expression of group IIA secretory PLA(2) (sPLA(2)-IIA), group V secretory PLA(2) (sPLA(2)-V), calcium-independent PLA(2) (iPLA(2)) and cytosolic PLA(2) (cPLA(2)). METHODS: Samples were obtained from non-glaucomatous patients (control subjects), and patients with either primary open-angle glaucoma (POAG) or exfoliation glaucoma (ExG). All the glaucoma patients had been treated with antiglaucomatous medication, and underwent deep sclerectomy surgery. Antibodies against sPLA(2)-IIA, sPLA(2)-V, iPLA(2) and cPLA(2) were used for immunohistochemical staining of frozen tissue sections. RESULTS: In the human conjunctiva of non-glaucomatous patients, immunostaining of sPLA(2)-IIA, sPLA(2)-V or cPLA(2) was low and positively stained cells were mainly localized in the surface of the epithelium. In contrast, iPLA(2) was found to predominate in human normal conjunctiva and it demonstrated strong labeling throughout the epithelium. The stromal staining of iPLA(2) was weak. Expression of sPLA(2)-IIA was significantly increased in stromal fibers of patients with POAG or ExG. No changes were found in levels of sPLA(2)-V, iPLA(2) or cPLA(2) between the patient groups and controls. CONCLUSIONS: These findings demonstrate that sPLA(2)-IIA, sPLA(2)-V, iPLA(2) and cPLA(2) are expressed in the conjunctiva of non-glaucomatous patients. In the epithelium, sPLA(2)-IIA, sPLA(2)-V, and cPLA(2) may participate in protection against risks caused by mechanical wear and tear stress whereas iPLA(2) may regulate remodeling and maintenance of membrane phospholipids. sPLA(2)-IIA may also have the important role in the degradation of bacteria. In conjunctival stroma of POAG and ExG patients, sPLA(2)-IIA may play a role in the development of scar tissue after glaucoma filtration surgery.

Phospholipase A2 in chamber angle of normal eyes and patients with primary open angle glaucoma and exfoliation glaucoma.

PURPOSE: Phospholipase A2 (PLA2) is a growing family of lipolytic enzymes that play a key role in various biological processes including general lipid metabolism, membrane homeostasis, and in diseases such as atherosclerosis, arthritis, and acute pancreatitis. Oxidative stress as well as inflammation may be associated with glaucoma pathogenesis. Therefore, our aim was to examine the expression of group IIA secretory PLA2 (sPLA2-IIA), group V secretory PLA2 (sPLA2-V), calcium-independent PLA2 (iPLA2), and cytosolic PLA2 (cPLA2) type in the trabecular meshwork (TM) and the canal of Schlemm in normal eyes and in juxtacanalicular tissue samples from patients with primary open angle glaucoma (POAG) or exfoliation glaucoma (ExG). METHODS: TM tissues were isolated from healthy donor eyes for corneal transplantation. Specimens of inner wall of the Schlemm's canal and the juxtacanalicular tissue were collected during deep sclerectomy from the eyes of patients who had POAG or ExG. Antibodies against PLA2s (sPLA2-IIA, sPLA2-V, iPLA2, and cPLA2) and a standard immunohistochemical procedure were used for the analysis. Quantification of immunoreactions was provided using a Photoshop-based image analysis. Double-staining immunofluorescence of macrophages and sPLA2-IIA was performed by using confocal microscopy. RESULTS: sPLA2-IIA was not present in normal TM. In contrast, sPLA2-IIA levels were significantly higher in glaucoma patients than in controls. Furthermore, sPLA2-IIA expression was much higher in POAG when compared to ExG. iPLA2 was found to predominate in normal human TM, and it demonstrated strong labeling in the uveal and corneoscleral meshwork. The staining of juxtacanalicular meshwork was only moderate in density. In contrast, expression of the enzyme was significantly decreased in glaucoma patients, especially in ExG, when compared to normal controls or to POAG. In addition, strong regional differences were detected in sPLA2-IIA and iPLA2 levels in POAG, whereas immunostaining of these enzymes was much lower and rather uniform throughout ExG sample. In POAG, sPLA2-IIA staining was restricted to certain parts of the trabecular samples where sPLA2-IIA positive macrophages were also present. Immunostaining of sPLA2-V or cPLA2 was low, and no significant changes were found in levels of these enzymes between normal and glaucomatous samples. CONCLUSIONS: sPLA2-IIA, an oxidative stress marker in atherosclerosis, is overexpressed especially in POAG. This result supports the hypothesis that oxidative stress may play a significant role in the pathogenesis of POAG. In ExG, a dramatic decrease in the expression level of iPLA2, a housekeeping enzyme in phospholipid remodeling, may indicate imbalance in phospholipid turnover and also inhibition of normal physiological functions in the TM. These findings may contribute to understanding the pathogenesis of POAG and ExG and may be important for the development of novel therapeutic strategies to different glaucomas.

Matrix metalloproteinases and their inhibitors in the chamber angle of normal eyes and patients with primary open-angle glaucoma and exfoliation glaucoma.

BACKGROUND: In glaucoma, extensive pathological changes occur in the trabecular meshwork (TM) and juxtacanalicular tissue of the chamber angle. Aqueous humor drainage is disturbed due to the accumulation of extracellular matrix (ECM) material in the outflow system. Matrix metalloproteinases (MMPs) remodel ECM material and, thus, they may have a role in regulating outflow facility and intraocular pressure (IOP). This study examined the expression of MMPs and tissue inhibitors of MMPs (TIMPs) in the chamber angle of normal eyes and in primary open-angle glaucoma (POAG) and in exfoliation glaucoma (ExG). METHODS: TM tissues were isolated from healthy donor eyes for corneal transplantation. Specimens of the inner wall of Schlemm's canal and the juxtacanalicular tissue were collected from patients with POAG or ExG during deep sclerectomy operation. Monoclonal antibodies against MMPs (MMP-1, -2, -3, and -9) and antibodies against TIMPs (TIMP-1, -2, and -3) were used for immunohistochemical staining. RESULTS: Immunoreactivity for MMP-2, TIMP-2, or TIMP-3 was observed in human normal TM and in the inner wall of Schlemm's canal. In general, immunoreactions for all of the tested MMPs were more intense in POAG samples than in ExG samples or in the control group. The only exception was the MMP-2 level, which was the highest in the control group. The staining intensity of MMP-1 or MMP-3 was significantly higher in POAG when compared to ExG. TIMP-1 was significantly increased in POAG compared with ExG and there were no marked differences in the levels of TIMP-2 or TIMP-3 between POAG and ExG. The ratios of MMP-1/TIMP-1 and MMP(1+2+3+9) and TIMP(1+2+3) were significantly higher in samples from POAG compared to those of ExG. CONCLUSIONS: Our results reveal an expression imbalance between MMPs and their endogenous tissue inhibitors in tissue samples from patients with POAG and ExG. Differences in immunohistochemical reactions reflect discrete local pathogenic mechanisms involved in POAG and ExG. With respect to the proposed role of MMPs in the remodeling of ECM material, this may point to a weaker reactivity to the accumulation of ECM material in TM in ExG than POAG eyes.

Asymmetric dimethylarginine is not elevated in exfoliation syndrome but symmetric dimethylarginine is related to exfoliative glaucoma.

BACKGROUND: Hyperhomocysteinemia (HH), oxidative stress and endothelial dysfunction are all implicated as possible pathogenetic factors in exfoliation syndrome (XFS) and exfoliative glaucoma (XFG). Asymmetric dimethylarginine (ADMA) is an endogenous inhibitor of endothelial nitric oxide and plasma level of ADMA is often elevated in HH. Thus the present study was undertaken to study plasma levels of ADMA with concomitant measurement of symmetric dimethylarginine (SDMA) and L-arginine (L-Arg) in XFS and XFG. METHODS: This cross-sectional, prospective study involved 36 XFS patients, 11 of them having XFG, and 36 age- and gender-matched controls. Fasting plasma ADMA, SDMA and L-Arg levels of participants were determined. A special view was created how plasma L-Arg, ADMA and SDMA correlate to plasma homocysteine (P-Hcy). In addition, the influence of P-Hey values derived from our previous study on the above mentioned parameters were evaluated by cut-off values of P-Hcy, 12 micromol/l for women and 14.5 micromol/l for men. RESULTS The mean plasma ADMA, SDMA and L-Arg levels were 0.41, 0.49 and 62.9 micromol/l in the XFS/XFG group, and 0.41, 0.44 and 69.7 micromol/l in the control group, respectively. As all parameters within the XFS and control group were compared, no statistical significance was stated. On the other hand, a positive correlation was observed between plasma SDMA and P-Hcy in XFGs (P = 0.002), and additionally, also a statistically significant difference was in plasma SDMA between the two groups sorted by cut-off levels of P-Hcy 0.49 +/- 0.15 vs. 0.36 +/- 0.04 micromol/l, above and below cut-off levels, respectively (P = 0.001), but not between ADMA in a respective assay. The mean values of L-Arg were 64.6 +/- 17.2 vs. 74.8 +/- 13.3 microg/l, respectively (P = 0.031). In the XFS subgroup, on the contrary, there was no positive correlation between P-Hcy and plasma SDMA. CONCLUSIONS: A positive correlation of plasma SDMA in respect to P-Hcy in XFGs and increase of SDMA in mild or intermediate hyperhomocysteinemia may indicate SDMA as a marker of developing XFG in hyperhomocysteinemic

Deep sclerectomy for the treatment of exfoliation and primary open-angle glaucoma.

PURPOSE: To retrospectively compare the efficacy of deep sclerectomy in the treatment of primary open-angle glaucoma (POAG) and exfoliation glaucoma (ExG). METHODS: Deep sclerectomy with either collagen or hyaluronate implants was performed in 31 eyes (45%) with POAG and 38 eyes (55%) with ExG. Pre- and postoperative intraocular pressure (IOP) was recorded, as was the number of glaucoma medications used pre- and postoperatively in each group. The follow-up data referred to a mean period of 18 months (range: 2 weeks to 36 months). RESULTS: At 18 months, complete success had been achieved in 56.3% of POAG eyes and 44.9% of ExG eyes. Qualified success had been achieved in 83.1% and 71.6% of POAG and ExG eyes, respectively. The mean IOP was 18.6 mmHg in POAG eyes and 16.3 mmHg in ExG eyes. YAG-descemetotomies were performed in nine eyes in each group. There were no statistically significant differences between the groups in IOP (except at 1 week postoperatively in favour of POAG; p = 0.05), success rates, need for postoperative glaucoma medication or number of complications. Reoperations were required in three (10%) POAG eyes and seven (18%) ExG eyes. CONCLUSIONS: Deep sclerectomy is equally effective in controlling IOP in both POAG and ExG and has low rates of serious complications, even when the surgeon is inexperienced in the technique. Both survival rates and IOP control were similar between the groups, and there were no serious intra- or postoperative complications.

Penetration of topically applied levofloxacin 0.5% and ofloxacin 0.3% into the vitreous of the non-inflamed human eye.

BACKGROUND: The aim of the study was to evaluate the vitreous penetration of two commercially available ophthalmic fluoroquinolones: ofloxacin and levofloxacin. METHODS: This prospective, double-blind, randomized clinical trial comprised 16 patients scheduled for vitrectomy surgery of one eye for macular hole or macular pucker. The patients were randomly assigned to receive topical ofloxacin 0.3% (n=9) or levofloxacin 0.5% (n=7) the day before, one drop at noon, 4 p.m., 8 p.m. and midnight. The next morning, patients were given their assigned masked antibiotic every 5 min for four doses starting 1 h before surgery. The vitreous humour samples, at least 0.3 ml each, were collected 1 h after the administration of the last dose, at the beginning of the pars plana vitrectomy with infusion disconnected. Samples were assayed for ofloxacin and levofloxacin concentrations by a method using high-performance liquid chromatography (HPLC) coupled with single mass spectrometry with electrospray ionization RESULTS: Equal topical administration of levofloxacin yielded 2.5 times higher vitreal concentration than ofloxacin. The mean vitreous concentrations of ofloxacin and levofloxacin were 5.30+/-3.04 (SD) ng/ml and 13.09+/-5.24 ng/ml, respectively (P=0.002). CONCLUSIONS: Equal dosing with topical administration of levofloxacin 0.5% and ofloxacin 0.3% allows better penetration into the vitreous for levofloxacin, but the levels of mean concentrations of each drug did not exceed the MIC(90) or MIC(50) for most ocular pathogenic bacteria in terms of conventional endophthalmitis therapy.

Plasma and aqueous humour levels of homocysteine in exfoliation syndrome.

BACKGROUND: Recent studies have suggested that the relationship between elevated plasma homocysteine (Hcy) and increased risk of vascular disease holds also for certain diseases of the eye with vascular aetiology. Elevated plasma Hcy levels have been noted among patients with exfoliation syndrome (XFS). The purpose of this study was to establish whether subjects with XFS have higher plasma and aqueous humour Hcy levels values than non-XFS subjects, particularly in relation to vitamin B status. METHODS: Using a cross-sectional study design, 36 subjects with XFS and 36 non-XFS subjects with intraocular pressure (IOP) lower than 23 mmHg, matched by age and gender, were first selected. The participant exclusion criteria included parameters known to alter Hcy metabolism. In the XFS group, 11 subjects had a concurrent diagnosis of exfoliative glaucoma (XFG). Fasting plasma and aqueous humour Hcy samples were collected, along with erythrocyte folate (E-Fol) and serum vitamin B6 and B12 samples. The Hcy samples were analysed using a fluorescence polarization immunoassay method. RESULTS: Plasma Hcy level was significantly higher (P=0.020, after Bonferroni correction for multiple testing) in the XFS group than in the controls. The Hcy concentrations in the aqueous humour did not differ statistically between the two groups. Plasma and aqueous humour Hcy concentrations were not statistically significantly correlated within the groups of exfoliation-positive and -negative subjects. E-Fol, and serum vitamin B6 and B 12 levels did not differ statistically between the XFS group and the control group. CONCLUSIONS: The finding that subjects with XFS are more prone to elevated plasma Hcy emphasizes exfoliation as a clinical sign and a marker of thromboembolic vasculopathies induced by hyperhomocysteinaemia.