Cytotoxicity of Ciprofloxacin and Steroids in Mouse Tympanic Membrane Fibroblasts.

Objective Ciprofloxacin, commonly given as eardrops, has been shown to adversely affect tympanic membrane fibroblasts. Dexamethasone potentiates this effect. A newly available eardrop contains ciprofloxacin and fluocinolone, a more potent steroid. We evaluated the cytotoxic effects of this preparation on mouse tympanic membrane fibroblasts. Study Design Prospective, in vitro. Setting Academic laboratory. Subjects and Methods In experiment 1, fibroblasts were exposed to 1:10 dilutions of commercially available 0.3% ofloxacin, 0.3% ciprofloxacin, 0.3% ciprofloxacin + 0.1% dexamethasone, 0.3% ciprofloxacin + 0.025% fluocinolone, or dilute hydrochloric acid (control), twice within 24 hours. In experiment 2, cells were also treated with the dilutions of the pure form of dexamethasone 0.1% or fluocinolone 0.025%, alone and in combination with ofloxacin or ciprofloxacin. Cells were exposed to the solutions for 2 hours each time and were placed back in growth media after the treatments. Cells were observed with phase-contrast microscope until the cytotoxicity assay was performed. Results Survival of fibroblasts treated with ofloxacin was not different from the control. Fibroblasts treated with ciprofloxacin, ciprofloxacin + dexamethasone, or ciprofloxacin + fluocinolone had much lower survival (all P < .0001). Cells treated with ciprofloxacin + fluocinolone had lower survival than ciprofloxacin ( P < .0001) and ciprofloxacin + dexamethasone ( P = .0001). Steroids alone also decreased fibroblast survival compared to control ( P < .0001). The combination of dexamethasone or fluocinolone with ciprofloxacin, but not ofloxacin, further decreased fibroblast survival ( P < .0001). Phase-contrast images mirrored the cytotoxicity findings. Conclusion Tympanic membrane fibroblast cytotoxicity of ciprofloxacin is potentiated by corticosteroids. This effect may be deleterious when treating a healing perforation but beneficial when treating granulation tissue on the tympanic membrane.

Hyalinization of the pyloric stomach in CD-1 mice following oral (dietary) administration of the corticosteroid agonists mometasone furoate, budesonide, and flunisolide.

The purpose of this study was to compare the toxicity of three marketed corticosteroid receptor agonists (mometasone furoate, budesonide, or flunisolide) to the stomach of female CD-1 mice following oral administration via the diet for up to 52 weeks, with a 16-week recovery period (budesonide and flunisolide). A range of tissues was examined by light microscopy, accompanied by clinical pathology measurements to assess anticipated corticosteroid effects as a surrogate marker of systemic drug exposure. Microscopic changes seen in the stomach with each corticosteroid included pyloric hyalinization. This previously unreported finding was investigated using histochemical and immunohistochemical techniques and was found to consist of hyalinized collagen, in association with increased immunohistochemical signal for transglutaminase-2 and osteopontin. The significance of the osteopontin finding is unclear; however, the ability of transglutaminase-2 to facilitate the formation of degradation resistant protein bonds implies this protein may be involved in the pathogenesis of this change. Furthermore, published evidence that transglutaminase-2 may be induced by a corticosteroid agonist raises the possibility that pyloric stomach hyalinization may be a class effect of corticosteroids via the action of this enzyme.

Safety and pharmacokinetics of an intraocular fluocinolone acetonide sustained delivery device.

PURPOSE: To determine the safety and pharmacokinetics of an intraocular fluocinolone acetonide sustained drug delivery device. METHODS: Nonbiodegradable drug delivery devices containing 2 or 15 mg of a synthetic corticosteroid, fluocinolone acetonide, were constructed. The long-term in vitro release rates of these devices were determined in protein-free buffer or buffer containing 50% plasma protein. Fifteen-milligram devices were also implanted into the vitreous cavities of rabbit eyes. Intravitreal drug levels, the amount of drug remaining in explanted devices, and the release rate of explanted devices were determined over a 1-year time period. Drug toxicity was assessed over this same time period by slit lamp examination, indirect ophthalmoscopy, electroretinography, and histologic examination. RESULTS: The drug release rates for the 2-mg device, 1.9 +/- 0.25 microg/d, and for the 15-mg device, 2.2 +/- 0.6 microg/d, remained linear over the 6-month and 45-day testing period, respectively. The release rate increased by approximately 20% when devices were transferred from protein-free buffer to buffer that contained protein (P: < 0.0001). Vitreous levels remained fairly constant (0.10-0.21 microg/ml) over a 1-year period. No drug was present in the aqueous humor during this time period. Based on the device release rates, the predicted life span of the 2- and 15-mg devices are 2.7 and 18.6 years, respectively. There was no evidence of drug toxicity by clinical examination, electroretinography, or histologic examination. CONCLUSIONS: It is feasible to construct a nontoxic fluocinolone acetonide drug delivery device that reproducibly releases fluocinolone acetonide in a linear manner over an extended period. These devices show great promise in the treatment of ocular diseases such as uveitis, which are often managed with chronic corticosteroid therapy.

The effects of topical nasal steroids on rat respiratory mucosa in vivo, with special reference to benzalkonium chloride.

Fifty rats were treated with topical nasal steroids with and without the preservative benzalkonium chloride in their right nostril twice daily for 21 days, while the left nostrils were exposed to 0.9% NaCl. By cutting the noses serially in frontal sections, the structure of the mucosal lining of all parts of the nose could be investigated. Areas with squamous cell metaplasia were observed in all nostrils exposed to topical steroids containing benzalkonium chloride. Such alterations were not observed in any nasal cavities exposed to the topical nasal steroid without the preservative or to 0.9% NaCl. In conclusion, benzalkonium chloride appears to be potentially toxic to the mucosa in vivo.

Effects of immunosuppressive chemicals on lymphoid development in foetal thymus organ cultures.

A murine foetal thymus organ culture system was employed to screen a number of immunotoxic chemicals for direct thymus toxicity. The toxic effects caused by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and its congeners on the system used had previously been shown to be similar to those caused in vivo on lymphoid development. The most potent compound tested was the corticosteroid fluocinolone acetonide, which caused a 50% inhibition of lymphoid development (EC50) at a concentration of 5 x 10(-11) M. The EC50 of TCDD was around 5 x 10(-10) M while that of 4 beta-phorbol 12-myristate 13-acetate (TPA) was ca 10(-7) M. TCDD and its congeners are believed to act via binding to the Ah receptor. Other known or presumed ligands of this receptor, which are potent inducers of P1-450 (P-448) -dependent polysubstrate monooxygenase activities, were considerably less toxic with EC50 levels varying between 10(-5) M (7,12-dimethylbenz(alpha-) antracene, alpha-naphthoflavone, benzo(alpha)pyrene) and 10(-4) M (beta-naphthoflavone and 3-methylcholantrene). Dinaphtho/2,3-b,5,6-b/dioxin and indolo/2,3-b/carbazole showed toxicity at 5 x 10(-6)-10(-5) M and 5 x 10(-5) M respectively. TCDD, TPA, and fluocinolone showed additive effects when added two by two in different combinations. Thus fluocinolone, known to counteract the toxicity and epidermal growth factor (EGF) cell-surface receptor-decreasing activity caused by TPA in other cell types, failed to decrease TPA toxicity in the thymus culture system.

Teratogenicity study of the new glucocorticosteroid budesonide in rabbits.

16 alpha,17 alpha-Butylidenedioxy-11 beta,21-dihydroxy-pregna-1,4-diene-3,20-dione (budesonide), a newly developed glucocorticosteroid, was administered subcutaneously to pregnant rabbits at doses of 0.01, 0.06 and 0.29 mumol/kg during the organogenetic period. As a reference compound the glucocorticosteroid fluocinolone acetonide, at the dose 0.28 mumol/kg, was used. In the group given 0.06 mumol/kg 2 dams out of 15 aborted. All animals receiving 0.29 mumol/kg aborted. All dams receiving fluocinolone acetonide aborted. The food consumption was reduced in all groups given budesonide. The body weight change was dose-dependently reduced for the groups receiving budesonide. The group receiving fluocinolone acetonide also showed a negative body weight change compared with the control group. The affected litter parameters were: reduced litter and fetal weight in the groups receiving 0.01 and 0.06 mumol/kg of budesonide, indicating intrauterine growth retardation. Furthermore, the frequency of fetal abnormalities, mainly skeletal anomalies, was significantly increased in the group receiving 0.06 mumol/kg of budesonide. The anomalies seen were mostly different kinds of delayed development of the bones in the skull and at the vertebra. A direct teratogenic effect cannot be excluded but severe maternal toxic reactions may have contributed to the pregnancy outcome.

Enhancement of beta-propiolactone tumorigenesis in mouse skin by pretreatment with the anti-inflammatory steroid fluocinolone acetonide.

Using a two-step carcinogenesis protocol, SENCAR mice were initiated with 25 micrograms 7,12-dimethylbenz[a] anthracene (DMBA) and were then treated twice weekly with either (a) 0.5 mg beta-propiolactone (BPL) or (b) 1 microgram fluocinolone acetonide (FA) followed in 30 min by 0.5 mg BPL. The tumor incidence for the group receiving FA prior to BPL was significantly greater than for BPL alone (P less than 0.0005). Under these experimental conditions, BPL alone showed neither promoting activity nor complete carcinogenic activity. These results were not anticipated, but the reasons for their occurrence are being explored.