Identification and radioimmunologic estimation of sexual steroid hormones in aqueous humor and vitreous of rabbit eye

Summary The occurrence of testosterone, estrone, and estradiol-17 in aqueous humor and vitreous body of the rabbit eye was demonstrated. The identity of the steroid hormones was confirmed by their chromatographic properties, microreactions, and radiochemically by crystallization to constant specific activity of their labeled acetates. The levels of testosterone and estrogens in rabbit male and female plasma, aqueous, and vitreous were determined by radioimmunologic assay. The concentrations of the steroid hormones in plasma are higher than in aqueous and vitreous. However, testosterone in aqueous circulates in free form and lacks the binding to sexual hormone-binding globulin as in plasma.

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Zusammenfassung Bericht über das Vorkommen von Testosteron, Oestron und Oestradiol-17 im Kammerwasser und Glasörper des Kaninchenauges. Die Art der Steroide wurde mittels Chromatographie, Mikroreaktionen und radiochemisch ermittelt. Der Testosteron- und Oestrogenspiegel im männlichen und weiblichen Kaninchenplasma, im Kammerwasser und Glaskörper wurde radioimmunologisch bestimmt. Die Plasmakonzentration der Steroidhormone lag höher als diejenige im Kammerwasser oder Glaskörper. Testosteron zirkuliert jedoch im Kammerwasser in freier Form, im Plasma dagegen ist es überwiegend an Sexualhormonebindende Globuline gebunden.

     

Penetration of topical and oral ciprofloxacin into the aqueous and vitreous humor in inflamed eyes.

PURPOSE: To assess the aqueous and vitreous penetration of ciprofloxacin after topical and combined topical and oral administration and investigate the effects of inflammation on drug penetration. METHODS: A standardized penetrating injury was made in the right eyes of 16 rabbits. Intraocular inflammation was induced by intravitreal injection of a suspension of Staphylococcus aureus in these eyes. The animals were divided into two groups according to treatment methodology: topical and topical-oral. The intact left eyes of the animals were maintained as controls. In the topical treatment group, two drops of ciprofloxacin 0.3% were instilled to both eyes every 30 minutes for 4 hours. In the topical-oral treatment group, animals were given two oral 40 mg/kg doses of ciprofloxacin at 12-hour intervals. After the last oral dose, the protocol of the topical group was applied to these eyes. Half an hour after the last drop, 100-microL samples were taken from aqueous and vitreous humor of all eyes. Drug concentrations were measured using high-pressure liquid chromatography. RESULTS: Mean aqueous levels of ciprofloxacin in control eyes were 2.31 microg/mL (range, 1.02-6.27 microg/mL) in the topical group and 5.88 microg/mL (1.52-17.81) in the topical-oral group. Mean aqueous levels in inflamed eyes were 7.36 microg/mL (2.34-17.15) in the topical group and 14.43 microg/mL (2.18-18.66) in the topical-oral group. Mean vitreous levels in control eyes were 0.77 microg/mL (0.09-1.93) in the topical group and 1.01 microg/mL (0.49-1.57) in the topical-oral group. Mean vitreous levels in inflamed eyes were 0.95 microg/mL (0.18-1.27) in the topical group and 1.98 microg/mL (0.51-3.34) in the topical-oral group. There was no significant difference among the groups (P > 0.05). Mean aqueous levels in all eyes and mean vitreous levels in the combined topical and oral group of inflamed eyes were above the 90% minimum inhibitory concentration for most of the common microorganisms causing endophthalmitis. CONCLUSION: There is an increase in both aqueous and vitreous humor concentrations with inflammation and with oral and topical administrations, as opposed to topical only, of ciprofloxacin. Using oral as well as topical treatment may be a beneficial method of antibiotic prophylaxis in ocular trauma once a patient has received intravenous or intravitreal therapy.     

Activated complement in inflamed aqueous humor

Activated complement is an important mediator of inflammation. Radioimmunoassay was used to measure levels of C3a, an activated fragment of C3, in aqueous humor. Additionally, immunoelectrophoresis was performed on aqueous humor to detect Factor B and its conversion product, Bb, as well as C3c, a breakdown product of C3. All six samples of normal aqueous humor had no detectable C3a, C3c, or Factor B. All eight samples of aqueous humor from patients with anterior uveitis had measurable levels of C3a. Factor B and C3c were detected in 3/7 samples of inflamed aqueous humor. Factor B was converted fully to Bb in two of these three samples, suggesting alternative pathway activation of complement. Activated complement fragments are present in the aqueous humor of eyes with anterior uveitis and may help mediate the inflammatory process.     

Polyamines in rabbit aqueous humor after surgical trauma to the eye

The polyamines putrescine, spermidine and spermine are necessary for cellular growth and directly involved in cellular differentiation and cell death. The hypothesis that extracellular polyamine levels in rabbit aqueous humor could be used as biomarkers for trauma after eye surgery was investigated. Changes in polyamine levels in rabbit aqueous humor were measured after anterior chamber lens implantation and compared to normals. The measurements were made by reversed phase HPLC and 9-fluorenylmethyl chloroformate and fluorescence detection. An increase in protein concentration followed by a white blood cell mobilization in the aqueous humor is a response to trauma to the eye. Therefore, the polyamine levels were compared to the aqueous levels of protein and leukocytes. Three days postoperatively a significant increase in spermidine was observed and a significant correlation between elevated protein levels and elevated spermine as well as total polyamines were noticed. No correlation between a high number of leukocytes and high polyamine levels were found. The results suggest that polyamines are evident markers for surgical trauma response, but not necessarily correlated to the postoperative inflammatory phase and the infiltration of inflammatory cells.     

Polypeptide profiles of normal and inflamed rabbit aqueous humor: identification of catabolic products of immunoglobulin G in the normal eye.

The anterior segment of the eye is isolated from the bloodstream by anatomic barriers. These barriers break down during inflammation elicited by muramyl dipeptide (MDP), a component of the cell walls of bacteria. The protein composition of normal aqueous humor and inflamed aqueous were characterized using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. The presence of transferrin, albumin, IgG and IgM were demonstrated in normal and inflamed aqueous. Normal aqueous additionally contained seven polypeptides (35, 31, 29, 25, 22, 21 and 19 kDa, respectively) that were not observed in inflamed aqueous. Four of these peptides (31, 29, 25 and 19 kDa) were identified as fragments of IgG heavy chains based on their reactivity with anti-IgG antibodies and staphylococcal protein A. Immunoglobulin catabolism has been noted to occur in tissues diffusely throughout the body; this is the first report of it occurring in the eye. This phenomenon appears to be circumvented on breakdown of the blood-aqueous barrier. These observations may relate to the immunoprivilege of the eye and its modulation in inflammation.     

Studies of the penetrating ability of fosfomycin into the aqueous humor and vitreous body of the eye

Fosfomycin is an antibiotic with a new simple chemical structure and a broad spectrum of antibacterial activity, especially against staphylococci. In the study reported here, the penetration of fosfomycin into the aqueous humor and vitreous body was investigated after intravenous infusion of 4 and 8 g. The highest average concentration approximately 2 hours after administration was 28.3 micrograms/ml after the 4 g dose and 52 micrograms/ml after the 8 g dose. Both the 4 g and 8 g doses thus produced aqueous humor levels exceeding the minimum inhibitory concentrations required for most gram-positive and gram-negative organisms commonly responsible for bacterial endophthalmitis, including strains of pseudomonas aeruginosa. Extremely high aqueous humor levels of fosfomycin were observed in two patients with inflamed eyes because of a change in the blood-aqueous barrier.     

Effect of Prostaglandin E2 on aqueous humor flow in the rabbit eye

Summary The examinations were carried out in albino rabbits under urethane anaestesia. After intracameral injection of 0.5 g Prostaglandin E₂, dissolved in 10 l 10% ethanol, intraocular pressure increased by 20.5±4.9 mm Hg. Aqueous humor protein concentration in the uninjected control eyes was 0.176±0.03 g%. Thirty minutes after intracameral injection of 10 l 10% ethanol solution the concentration rose to 0.38±0.19 g-%, and after of 0.5 g Prostaglandin E₂ to 1.85 ±0.41 g% These interventions were not followed by significant changes in aqueous humor osmolarity (normal value 313.7±17.9 mOsm, after ethanol 330 5±18. 0, and after prostaglandin E₂ 329.3±6.9).In a special series of experiments the rate of aqueous humor production was determined. In the uninjected control eyes a value of 1.57±0.61 l/min was found, but after intracameral injection of 0.5 g prostaglandin E₂, 5.45±1.99 l/min.The authors draw the conclusion that prostaglandin E₂ increases intraocular tension not by enhancing aqueous humor production but by disrupting the bloodaqueous humor barrier.

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Zusammenfassung Die Untersuchungen wurden an Albinokaninchen in Urethannarkose durchgeführt. Nach Injektion von 0,5 g Prostaglandin E₂ (in 10 l10%iger Äthanollösung) in die Vorderkammer stieg der Augenbinnendruck um 20,5±-4,9 mm Hg an. Der Gesamteiweißgehalt im Kammerwasser der intakten Kontrollaugen betrug 0,176±0,03 g-%. 30 min nach Einspritzung von 10 l 10%iger Äthanollösung in die Vorderkammer erhöhte sich der Gesamteiweißgehalt auf 0,38±0,19 g-%. Diese Einwirkungen hatten keine signifikante Änderung der Kammerwasser-Osmolarität zur Folge (Normalwert 313,7±17,9 mOsm, nach Äthanol 330,5±18,1 und nach Prostaglandin E₂ 329,3±6,9).In einer besonderen Versuchsreihe wurde die Kammerwasserproduktion bestimmt. In den intakten Kontrollaugen ergab sich hierbei ein Wert von 1,57±0,61 (l/m, nach Gabe von 0,5 g Prostaglandin E₂ aber 5,45±1,99 l/min.Die Autoren ziehen den Schluß, daß Prostaglandin E₂ den Augenbinnendruck nicht durch Steigerung der Kammerwasserproduktion, sondern durch Schädigung der Blut-Kammerwasserschranke erhöht.

     

Adenosine 3',5'-monophosphate increases the outflow of aqueous humor from the rabbit eye.

Direct administration of cyclic-AMP into the anterior chamber increases the outflow facility of the eye for aqueous humor. This is consistent with the hypothesis that catecholamines lower the intraocular pressure of the rabbit eye, at least in part, by a cyclic-AMP-mediated mechanism. This mechanism is active in the outflow channels and increases the rate at which aqueous humor leaves the anterior chamber.     

眼球穿孔伤患者早期前房水及玻璃体细菌学培养分析

目的 了解眼球穿孔伤患者早期眼内组织细菌学培养及药物敏感情况.方法 前瞻性地对本院2003年7月至9月收治的受伤时间在24 h以内的45例(45眼)眼球穿孔伤患者的前房水或玻璃体进行细菌培养及药敏实验,并对其结果 进行分析.结果 共15例(33.3%)患者(玻璃体培养8例、房水培养7例)细菌培养结果 阳性,其中10例为表皮葡萄球菌(67.7%).所有细菌均对环丙沙星、氧氟沙星敏感,对头孢哌酮较为敏感,大部分对庆大霉素、氯霉素、利福平耐药.球内有异物存留(χ2=6.544,P=0.016)、伤口累及巩膜(χ2=4.500,P=0.034)者细菌培养结果 阳性率高,而致伤物类型(χ2=1.667,P=0.197)、有无合并晶状体损伤(χ2=1.125,P=0.289)、取材部位(χ2=1.667,P=0.197)对细菌培养结果 无明显影响.结论 表皮葡萄球菌是眼球穿孔伤后进入眼内的主要细菌.对眼球穿孔伤患者预防性用药可首选喹诺酮类药物,次选头孢哌酮.     

Sodium activity in the aqueous humor and corneal stroma of the rabbit.

The present study examined the free sodium concentration of the aqueous humor and corneal stroma of both transparent and non-transparent corneas to assess the transendothelial activity gradient for sodium. In the transparent cornea of the adult rabbit, the sodium activity was higher in the aqueous humor than the stroma. This difference in sodium activity would cause water to diffuse down its concentration gradient from stroma to aqueous humor. In this way corneal transparency and the deturgesced state are maintained. Removal of the corneal endothelium in the adult rabbit produced an opaque swollen cornea. Under these conditions the sodium activity was higher in the stroma than the aqueous humor. However, an osmotic gradient was not produced by the Na+ activity gradient because the endothelium was not present to act as a semi-permeable membrane. The corneal endothelium was no longer present to establish and sustain the activity gradient for sodium that is necessary for corneal transparency in the mature rabbit. The transendothelial sodium activity gradient was also measured in 13-day-old rabbits. At this age, the cornea was not yet transparent, nevertheless the free sodium concentration of the aqueous humor was higher than that of the stroma, similar to the adult transparent cornea. This suggests that forces other than the establishment of the proper transendothelial sodium gradient are responsible for the lack of corneal transparency in the young rabbit.     

Effect of various drugs on pseudofacility and aqueous humor formation in the rabbit eye.

Norepinephrine and epinephrine reduce aqueous humor formation by an amount less than sufficient to reduce the intraocular pressure. Both drugs increase pseudofacility and true outflow facility which both contribute to reduce the pressure. Neither drug changes the value of x_c, the pressure index of the capillaries, which indicates that large hemodynamic changes do not occur. Phenylephrine has no effect on pseudofacility, since, although x_c is reduced, the fluid permeability is increased in a compensatory manner: an increase in true outflow facility is responsible for the small pressure fall.Pilocarpine causes a minor increase in aqueous humor formation, has no effect on pseudofacility yet lowers the intraocular pressure, albeit by only 2 mmHg. In order to lower intraocular pressure, therefore, pilocarpine must increase true outflow facility.Prostaglandin E₁ increases intraocular pressure, aqueous humor formation, x_c and pseudofacility. PGE₁ is known to induce venous congestion and this correlates with the increase in x_c since a secondary rise in capillary pressure would follow. The increase in aqueous humor formation accounts for the increase in intraocular pressure. The increase in pseudofacility accounts for about half of the increase in total outflow facility found with PG's, at least at times up to 2 hr after topical PG application.     

Fibrinolytic activity and localization of plasminogen activator in bovine vitreous body and aqueous humor

The types, activities and concentrations of plasminogen activator (PA) were examined in the bovine vitreous body and aqueous humor. Molecular weights of PA in the vitreous body and aqueous humor were approximately 66 and 110 kilodaltons (kDa), respectively, as determined by fibrinolysis enzymography. These PA activities were neutralized by anti-human tissue PA (t-PA) IgG but not by anti-human urokinase PA (u-PA) IgG. Both findings indicated that the main PA in the vitreous body and aqueous humor is t-PA. The existence of u-PA could not be confirmed in intraocular fluids. Fibrinolytic activities measured by amidolytic assay were 0.12 +/- 0.01 IU/ml in the vitreous body and 0.04 +/- 0.02 IU/ml in the aqueous humor. High concentrations but relatively low fibrinolytic activities suggested that PA and PA inhibitor coexisted in the intraocular fluids. Moreover, unbound PA inhibitor was not detected by reverse fibrinolysis enzymography. The present study suggests that most of the PA inhibitor combines with PA and forms a complex. PA predominates over PA inhibitor in the intraocular fluids, whereas PA inhibitor predominates in the plasma. The results by Todd's fibrinolysis autography and immunohistochemical staining showed that t-PA was present in the corneal endothelial cells, which are in direct contact with the aqueous humor. This suggests that not only the vascular system but the corneal endothelial cells as well might be a source of the PA of the aqueous humor and possibly of the vitreous body also.     

Penetration of imipenem into human aqueous and vitreous humor

Twenty-five patients about to undergo cataract surgery and ten patients about to undergo vitrectomy received a 1-g intravenous dose of imipenem before surgery. Specimens of aqueous or vitreous humor were then obtained and assayed for antibiotic content with a microbiologic disk agar technique. A mean peak aqueous humor level of 2.99 micrograms/ml was found at approximately two hours after administration, and a mean vitreous level of 2.53 micrograms/ml was found from two hours to three hours 15 minutes after administration. These concentrations were well above the minimum inhibitory concentration of imipenem for 90% (MIC90) of Staphylococcus epidermidis, S. aureus, and the Enterobacteriaceae commonly involved in bacterial endophthalmitis.     

Crystals in the aqueous humor of normal eye

An unusual and extremely rare occurrence of the free floating crystals in the aqueous humor of a structurally healthy and functionally normal eye of a 15-year-old white girl is described. The exact nature of these crystals could not be determined but their iridescent nature suggests that they might be cholesterol. This observation leads one to believe, contrary to the current concept, in the very interesting possibility of the crystal formation (or synchysis scintillans) in an apparently healthy eye.