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Detection of the free acid of bimatoprost in aqueous humor samples from human eyes treated with bimatoprost before cataract surgery.

PURPOSE: To determine whether bimatoprost is hydrolyzed to its free acid after topical application in humans in vivo. DESIGN: Prospective, masked, and vehicle controlled. PARTICIPANTS: Thirty-one eyes of 31 patients with cataracts. METHODS: Beginning 7 days before scheduled cataract surgery, one eye of each patient was treated with bimatoprost 0.03% or vehicle once daily, with the last drop administered 2 to 12 hours before anterior chamber paracentesis before cataract surgery. In a masked fashion, aqueous humor specimens were assayed for bimatoprost and its free acid by high-pressure liquid chromatography and mass spectrometry. MAIN OUTCOME MEASURE: Detection of the free acid of bimatoprost in aqueous humor. RESULTS: Aqueous humor concentrations of the free acid of bimatoprost were 22.0+/-7.0 nmol/l (mean +/- standard error of the mean, n = 12) and 7.0+/-4.6 nmol/l (n = 8) at 2 and 12 hours, respectively, and below the limit of detection after vehicle (n = 10). Concentrations of bimatoprost (amide) were 5.7+/-1.4 and 1.1+/-0.4 nmol/l at 2 and 12 hours, respectively, and undetectable after vehicle. CONCLUSION: After topical application of bimatoprost in humans, a sufficient concentration of its free acid, a potent FPprostanoid receptor agonist, is found in the aqueous humor to account for its ability to reduce intraocular pressure.

Administration, Topical↗

[Antibodies to human T-cell lymphotropic virus type 1 in the aqueous humor of HTLV-I associated uveitis].

To elucidate the pathomechanism of human T-cell lymphotropic virus type 1 (HTLV-I) associated uveitis (HAU), we assessed antibodies to HTLV-I and IgG for paired samples of serum and aqueous humor. Seven of 10 samples from HAU patients showed antibodies to HTLV-I in the aqueous humor with antibody titer ranging from 16 to 128, and five of these samples showed significantly high antibody quotients as calculated by reference to the IgG levels in the serum and aqueous humor, hence suggesting local production of antibodies rather than their transfer from the serum. On the other hand, eight of 45 control samples from HTLV-I carriers with senile cataract but without uveitis had antibodies to HTLV-I in the aqueous humor with antibody titer ranging from 8 to 128, but none of these samples showed high antibody quotient. We conclude that, although antibodies to HTLV-I in aqueous humor were detected in those HAU and control cases which showed high antibody titer in the serum, intraocular synthesis of antibodies to HTLV-I may sometimes occur in HAU.

Adult↗

Decreased fibronectin levels in aqueous humor after corneal injury.

Fibronectin plays an important role in wound healing and cell differentiation. The present study measures fibronectin levels in aqueous humor from injured corneas. In response to transcorneal freezing treatments, a thickened scar-like matrix is deposited on the posterior surface of rabbit corneas. This posterior collagenous layer (PCL) is visible grossly and has a uniform diameter. Aqueous humor from eyes with established posterior corneal scarring has significantly less fibronectin compared with control eye aqueous humor. Tissue culture experiments show that corneal endothelial cells from control eyes and PCL-associated cells from injured eyes produce similar amounts of fibronectin. The marked decline in fibronectin levels in aqueous humor in scarred corneas supports the hypothesis that fibronectin plays a role in the pathological events which result from corneal injury.

Animals↗

Diagnostic value of polymerase chain reaction in blood and aqueous humor in immunocompetent patients with ocular toxoplasmosis.

PURPOSE: To evaluate the diagnostic value of polymerase chain reaction (PCR) in blood and aqueous humor samples from immunocompetent patients with reactivated ocular toxoplasmosis. METHODS: Group 1 was composed of seven patients with a clinical diagnosis of reactivated ocular toxoplasmosis. Group 2 consisted of 33 controls. In each subject, blood and aqueous humor samples were obtained for detection of Toxoplasma gondii DNA by means of simple PCR, seminested PCR, and Southern blot hybridization. RESULTS: Group 1: Simple PCR was positive in 3 of 7 blood samples (42%) and in 2 of 7 (28%) aqueous humor samples. Seminested PCR was positive in 4 of 7 (57%) blood samples and in 3 of 7 (42%) aqueous humor samples. Group 2: Simple and seminested PCR were positive in both samples in 2 of 33 (6%) and 4 of 33 (12%), respectively. Sensitivity 57% (18.41-90.10), specificity 87% (71.80-96.60); positive and negative likelihood ratio 4.38 and 0.49, respectively. CONCLUSIONS: Polymerase chain reaction can be useful for confirming the diagnosis of ocular toxoplasmosis, especially in those eyes where fundus examination does not yield conclusive results. The detection of T. gondii DNA in blood suggests that reactivation of ocular toxoplasmosis cannot be considered a local event.

Adult↗

Aqueous humor flow in sleeping humans is unaffected by norepinephrine infusion.

PURPOSE: Intravenous administration of the catecholamine epinephrine is known to have a stimulatory effect on aqueous humor flow in sleeping human subjects, an effect that is augmented by plasma corticosteroids. This study was performed to determine whether the closely related catecholamine norepinephrine has a similar effect on aqueous humor flow. METHODS: Twenty normal subjects were studied. Aqueous flow was measured by fluorophotometry. At night during sleep, norepinephrine or placebo was infused intravenously (i.v.) between midnight and 6 AM. The rate of aqueous flow during the norepinephrine infusion was compared with the rate of flow during placebo infusion, with each subject serving as his/her own control. The urinary excretions of epinephrine and norepinephrine were measured at the end of each infusion period. RESULTS: The norepinephrine infusion caused an 8% increase in systolic blood pressure (P < 0.001), a 15% increase in diastolic blood pressure (P < 0.001), and a 9% decrease in heart rate (P=0.003) compared with the placebo. The rate of aqueous humor flow during sleep from 12 AM to 6 AM was unchanged by norepinephrine. The rate was 1.27+/-0.31 microl/min (mean+/-SD) during i.v. infusion of placebo and 1.30+/-0.27 microl/min during infusion of norepinephrine (P=0.63). CONCLUSIONS: An infusion of norepinephrine during sleep that causes measurable changes in cardiovascular parameters has no measurable effects on the rate of aqueous humor flow. The lack of a measurable effect of a norepinephrine infusion contrasts to the stimulatory effect of an epinephrine infusion.

Adult↗

Aqueous humor levels of topically applied levofloxacin in human eyes.

PURPOSE: To evaluate transcorneal penetration of topically applied 0.5% levofloxacin into the aqueous humor in human eyes. METHODS: Twenty cataract patients (14 females, 6 males) received 3 drops of 0.5% levofloxacin at 15 min intervals from 90 minutes before the surgery. At the beginning of the cataract surgery, 50 microL of aqueous humor was aspirated from the anterior chamber and stored at -80 degrees C until analysis. The drug concentration of the samples was analyzed using high-performance liquid chromatography. RESULTS: A mean aqueous humor level of levofloxacin was 1.00 +/- 0.48 microg/mL (mean +/- standard deviation, n = 20), ranging from 0.30 microg/mL to 2.32 microg/mL. CONCLUSIONS: The mean concentration of levofloxacin in the aqueous humor was higher than the MIC(90) values against some common pathogens of postoperative endophthalmitis, although a great degree of interpatient variability was present.

Administration, Topical↗

Lipid-laden aqueous humor associated with anterior uveitis and concurrent hyperlipemia in two dogs.

Two dogs with cloudy aqueous humor due to accumulation of lipid and lipoprotein had concurrent hyperlipemia and anterior uveitis. In 1 dog, serum and aqueous humor had similar lipoprotein electrophoretic patterns, and in the other dog, the patterns were different. The therapeutic approach included symptomatic treatment of the uveitis and reduction of dietary fat intake. Clearing of fat from aqueous humor followed control of the uveitis.

Animals↗

Laboratory evaluation of aqueous humor in the healthy dog, cat, horse, and cow.

Using routinely available clinical laboratory methods, aqueous humor samples were evaluated from 12 healthy dogs, 15 healthy cats, 7 healthy horses, and 6 healthy cows. Aqueous humor was almost acellular; cells that were present had degenerated beyond recognition. Protein concentration was low; only albumin was detectable on electrophoresis. Creatine phosphokinase and lactate dehydrogenase isoenzymes were not detected. Artifacts induced by sampling were insignificant compared with alterations in aqueous humor composition that occur with ocular diseases.

Albumins↗

Transforming growth factor beta 2 levels in the aqueous humor in different types of glaucoma and the relation to filtering bleb development.

PURPOSE: To investigate the transforming growth factor beta 2 (TGF-beta 2) levels and total protein levels in the aqueous humor of eyes with different types of glaucoma [primary open-angle glaucoma (POAG), pseudoexfoliation glaucoma (PSX), juvenile glaucoma (JG)], and the relation to filtering bleb development after trabeculectomy. METHODS: Aqueous humor was collected at the beginning of surgery from 52 eyes with glaucoma (29 POAG eyes, 17 PSX eyes, 6 JG eyes) and from 29 control eyes that underwent cataract operation. TGF-beta 2 levels (intrinsically activated and total TGF-beta 2) using ELISA methods as well as total protein concentrations of the aqueous humor were determined. All preoperative clinical data of the glaucoma eyes (age, gender, IOP, previous treatment, type of surgery) were compared with the TGF-beta 2 levels. In 40 of these eyes, the postoperative follow-up (filtering bleb development, need for intervention, IOP) was correlated to the preoperatively determined TGF-beta 2 levels. RESULTS: TGF-beta 2 levels were increased in nearly half of the eyes with POAG and in most of the eyes with JG, but in eyes with PSX, TGF-beta 2 levels were within the normal range. No correlation between TGF-beta 2 levels and age, gender, IOP, previous treatment, or type of surgery, or between TGF-beta 2 levels and protein levels in aqueous humor, was found. Correlation between bleb formation and TGF-beta 2 levels revealed that all but two of the POAG eyes with good clinical outcome (type 1 bleb) had normal levels of activated TGF-beta 2. Of the 13 eyes that needed postoperative intervention (type 2 and type 3 bleb), 8 had high and 5 had normal TGF-beta 2 levels. CONCLUSIONS: PSX eyes differ from POAG and JG eyes not only by their clinical or biomicroscopic appearance, but also by their normal TGF-beta 2 levels in aqueous humor. The fact that most of the POAG eyes with favorable bleb development had normal TGF-beta 2 levels indicated that there might be some relationship between bleb formation and TGF-beta 2 levels. On the other hand, the fact that eyes with less favorable bleb development had both low and high TGF-beta 2 levels indicated that other factors are also involved in the scarring of the filtration bleb.

Adolescent↗

[Anaphylatoxin in the aqueous humor of rabbit with endotoxin induced endophthalmitis].

Anaphylatoxin-activated complement fragment is an important mediator of inflammation. In this study, trends of anaphylatoxin in clinical and experimental endophthalmitis was evaluated. The level of anaphylatoxin (C3a, C4a and C5a) and protein of human aqueous humor aspirated from patients with senile cataracts before surgery was measured. As a model of acute endophthalmitis, endotoxin-Lipopolysaccharide from E. coli (LPS) was used intravitreously in albino rabbits. The level of anaphylatoxin of aqueous humor was measured by radioimmunoassay. The level of C3a was 44.0 +/- 13.4 ng/ml in aqueous humor of cataract patients. An increase of cell count and protein was demonstrated in the aqueous humor of rabbits 24 hrs after endotoxin injection. The level of C3a increased remarkably with time. It was suggested that C3a in aqueous humor of the rabbit eye with endotoxin-induced endophthalmitis plays an important role in chemotaxis and in mediating the inflammatory process.

Aged↗

A comparison between the effect of topical and systemic carbonic anhydrase inhibitors on aqueous humor secretion.

We have assessed the onset and duration of decreased intraocular pressure and aqueous humor flow contrasting systemic and topical administration of carbonic anhydrase inhibitors. The relationship between physiological effects and fractional activity of carbonic anhydrase isoenzymes in the eye was also investigated. Experiments were performed in normotensive New Zealand white rabbits. Intraocular pressure was determined manometrically or tonometrically and aqueous humor flow by sulfacetamide clearance. We studied methazolamide (25 mg kg-1), ethoxzolamide (4 mg kg-1), and MK-927 (2% in 0.5% hydroxyethylcellulose, topical, pH 4.8). There is an immediate reduction in intraocular pressure (1.2 and 1.8 mmHg by 2 min) and aqueous flow (33% and 40% by 5 min) following intravenous dosing with either methazolamide or ethoxzolamide. This correlates with rapid appearance of drug in the anterior uvea and very low fractional activity of ocular carbonic anhydrase isoenzymes II (cytosolic) and IV (membrane bound). Peak intraocular pressure reduction averaged 4.2 +/- 0.68 mmHg and 4.5 +/- 0.8 mmHg for methazolamide and ethoxzolamide at 60 and 45 min, respectively. Peak flow reduction was 38% for methazolamide and 40% for ethoxzolamide, at 5 min. Aqueous flow and intraocular pressure returned to baseline at 7 and 4 hr following methazolamide and ethoxzolamide, respectively. This corresponds to decay of drug from ocular tissues and significant increases in fractional activity of carbonic anhydrase isoenzymes. Topical MK-927 resulted in a 1.2 mmHg decrease in pressure by 5 min. This correlated with the early appearance of drug in the anterior uvea prior to its appearance in aqueous humor and very low fractional activity of carbonic anhydrase isoenzymes. Intraocular pressure decreased 3.6 +/- 0.35 mmHg at 1 hr and returned to baseline by 6 hr. Aqueous flow was reduced 12% by 5 min and 35% at 1 hr. The appearance of MK-927 in the anterior uvea prior to detection in aqueous suggests a significant non-corneal route of absorption following topical administration. Topical MK-927 results in a more gradual reduction in intraocular pressure and flow, although peak effects are not statistically different from systemic carbonic anhydrase inhibitors. The time of pressure return to baseline is also comparable to systemic carbonic anhydrase inhibitors. Because the relations between carbonic anhydrase II and carbonic anhydrase IV in the ciliary process are not yet clear and since the drugs have different affinities for the isozymes, the precise degree of fractional inhibition necessary for pharmacological effect is not certain, but based on drug concentration in the anterior uvea, may take 98% inhibition for full intraocular pressure reduction.

Administration, Topical↗

Transcellular aqueous humor outflow: a theoretical and experimental study.

When the aqueous humor passed through the transcellular channels of the inner wall endothelium of Schlemm's canal the flow was estimated to be very slow and viscous (Reynolds' number about 10(-3)). It was found to be accurate to treat this extreme flow situation with conventional hydrodynamic formulae without specific corrections. Four different principal configurations of the transcellular channels were investigated. The theoretical calculations yielded results in good agreement with those obtained from the experimental model studies. Where deviations occurred they were probably explained by a radial flow component, which was not incorporated in the theoretical formulae applied. A spherical channel configuration was considered most similar to the in vivo situation and this yielded experimental resistance values half those of previously applied theoretical methods. Using previous morphological data the present results indicated that less than 5% of the total resistance to aqueous humor outflow was located in the inner wall endothelium in normal human eyes, and about 10% in cynomolgus monkey eyes at normal intraocular pressures.

Animals↗

Elevated levels of calcitonin gene-related peptide in aqueous humor of patients with proliferative vitreoretinopathy.

PURPOSE: To detect the levels of the sensory peptide calcitonin gene-related peptide in aqueous humor of patients with proliferative vitreoretinopathy and to compare them with those of uninflamed eyes (cataract and uncomplicated rhegmatogenous retinal detachment). MATERIALS AND METHODS: Using a highly specific and sensitive radioimmunoassay the concentration of calcitonin gene-related peptide was detected in fresh samples of aqueous humor obtained via paracentesis. Furthermore, calcitonin gene-related peptide-like immunoreactivities were characterized by high-pressure liquid chromatography. RESULTS: The mean level of calcitonin gene-related peptide was 6.11 fmol/ml in cataract controls and 14.77 fmol/ml in uncomplicated rhegmatogenous retinal detachment. In the cataract group, 9 of 18 cases were below the detection limit and in the retinal detachment group, 5 of 16. In proliferative vitreoretinopathy, the peptide averaged 76.92 fmol/ml and none of the samples was below the detection limit. High-pressure liquid chromatography revealed one major peak corresponding to synthetic calcitonin gene-related peptide. CONCLUSION: In recent studies, we found elevated levels of the sensory peptide substance P in aqueous humor of patients with proliferative vitreoretinopathy. This fact and the present result, the elevation of calcitonin gene-related peptide in aqueous humor of eyes with proliferative vitreoretinopathy, clearly point to an involvement of sensory peptides in the pathobiology of the disease. The source of the elevation is not clear, but we hypothesize that it originates from a neurogenic mechanism, i.e. an acceleration of the peptides by their enhanced release from the iris/ciliary body complex subsequent to sensitization of sensory neurons, thus representing a very interesting epiphenomenon of proliferative vitreoretinopathy. Our results constitute novel aspects in the pathophysiological concept of proliferative vitreoretinopathy and extend the knowledge about the pathobiology of the disease process.

Adolescent↗

Pharmacologic and pharmacokinetic assessment of anti-TGFbeta2 aptamers in rabbit plasma and aqueous humor.

PURPOSE: The aim of the study is to determine the bioactivity and effects of PEGylation on the pharmacokinetics in rabbit aqueous humor and plasma of an aptamer directed against TGFbeta2. METHODS: Pharmacological activity of anti-TGFbeta2 aptamer in rabbit ocular fluid was demonstrated using a mink lung epithelial cell proliferation assay. For pharmacokinetic analyses, concentrations of aptamers in plasma and aqueous humor were determined over time following bilateral subconjunctival administration to Dutch-belted rabbits using a hybridization-based pseudo-enzyme-linked immunosorbent assay (ELISA) assay. RESULTS: Anti-TGFbeta2 aptamer (ARC81) binds to human TGFbeta2 with a K(D) of approximately 5 nM and inhibits the activity of human TGFbeta2 in vitro in a cell-based assay with an IC(50) of approximately 100 nM. ARC81 blocks endogenously derived TGFbeta2 in rabbit aqueous humor in vitro with an IC(50) of approximately 200 nM and an IC(90) of approximately 1 microM. In vivo in rabbit, ARC81 [no polyethylene glycol (PEG)] entered systemic circulation rapidly (t(max) = 1 h in plasma) relative to aptamer conjugates ARC117 (20 kDa PEG) and ARC119 (40 kDa PEG), which showed prolonged residence in the subconjunctival space and aqueous compartment (t(max) = 6 and 12 h, respectively, in plasma). Both 20- and 40-kDa aptamer conjugates reached maximal concentrations (C(max)) in aqueous humor of 23-30 nM and remained at or above 1 nM for as long as 12 h. CONCLUSIONS: Pharmacologically active levels of anti-TGFbeta2 aptamers can be sustained in the ocular fluid and local tissue environment over a 12-h period after single administration. Daily subconjunctival administration of PEGylated anti-TGFbeta2 aptamers should allow further pharmacological evaluation of these agents in a rabbit conjunctival scarring model. Perioperative administration, via subconjunctival injection, may prove to be an effective means to deliver therapeutic quantities of TGFbeta2 aptamer conjugates in trabeculectomy procedures.

Animals↗

Aqueous humor as a modulator of growth in fibroblast cultures.

Samples of aqueous humor from rabbits and humans were examined for growth effects on subconfluent monolayer fibroblast cultures from ocular and extraocular tissues. At a concentration of 20% aqueous by medium volume, both homologous and heterologous aqueous stimulated DNA synthesis in fibroblasts from both species. The stimulation was independent of medium serum. When fibroblasts were exposed to 0 to 100% aqueous in serum-free medium or 0 to 98% aqueous in 2% serum-supplemented medium, no concentration of aqueous was observed in which DNA synthesis was inhibited. However, higher concentrations of aqueous led to increasing cell loss resulting in an apparent net inhibition of culture growth. Net growth effects of aqueous humor were, therefore, dependent upon aqueous concentration and were apparently due to the combined effects of stimulation of proliferation and reduction in cell survival.

Animals↗

The corneal contact lens and aqueous humor hypoxia in cats.

The cornea normally receives its oxygen from both the atmosphere and the aqueous humor. When a thick polymethylmethacrylate contact lens is placed on the cat cornea, access of atmospheric oxygen to the cornea is restricted, and the outer part of the cornea becomes hypoxic. This results in an increase in the oxygen flux from the aqueous humor into the cornea, and the oxygen tension in the aqueous humor decreases, as is demonstrated in this study. This increased oxygen flux from the aqueous into the cornea tends to alleviate the hypoxia caused by a corneal contact lens. Thus, the cornea can tolerate a thicker contact lens with less hypoxia than would be expected if only the oxygen supply from the atmosphere were considered.

Animals↗

Demonstration of a considerable amount of mouse epidermal growth factor in aqueous humor.

In a previous study we reported the presence of a considerable amount of mouse epidermal growth factor (mEGF) in abdominal effusion. Using our EIA system for mEGF, we identified a high level of EGF-like immunoreactive material(s) in mouse aqueous humor. This material(s) and mEGF from mouse submaxillary gland were virtually equivalent with respect to molecular weight and antigenicity. Also, on chromatofocusing analysis, the mEGF-like material(s) gave a major peak at pH 4.7 with a minor one at pH 4.2. These results demonstrate that the mEGF-like immunoreactive material(s) found in aqueous humor is a molecule identical to submaxillary gland EGF. Also, no clear difference was observed in the mEGF levels in aqueous humor between male and female. Further, sialoadenectomy did not change dramatically the EGF level in aqueous humor. From these results, it seems that mEGF found in aqueous humor may be synthesized by cells in the eyeball itself or be transported there from some site other than the submaxillary gland.

Animals↗

Systemic aminocaproic acid reduces fibrinolysis in aqueous humor.

Systemically administered aminocaproic acid has been demonstrated to reduce the incidence of secondary hemorrhage in traumatic hyphema. To directly determine the concentration of aminocaproic acid in the aqueous humor, four groups of rabbits were studied following administration of four different regimens of intravenously administered aminocaproic acid. Plasma and aqueous humor samples were assayed for drug content and antifibrinolytic activity. Peak aqueous humor concentrations of aminocaproic acid ranged from 2.5 to 33 mg/dL and varied according to the systemic dose administered. The antifibrinolytic activity paralleled the aminocaproic acid content, with a peak range from 310 to 683 s. These findings indicate a direct relationship between antifibrinolytic activity and the concentration of aminocaproic acid in aqueous humor.

Aminocaproates↗