Under ambient UVA exposure, pefloxacin exhibits both immunomodulatory and genotoxic effects via multiple mechanisms.

Pefloxacin (PFLX) is an antibiotic, which shows broad spectrum antimicrobial activities. It is an important derivative of fluoroquinolones (FLQs) group. Ultraviolet radiation (200-400nm) causes major problem for living being which comes at the earth surface naturally through sunlight and increasing regularly due to ozone depletion. PFLX was photodegraded in 5h and forms photoproduct under UVA exposure. At the non photocytotoxic dose PFLX, shows reduced phagocytosis activity, NO (nitric oxide) production, large vacuole formation and down regulated IL-6, TNF-α and IL-1 in BALB/c macrophages at both genes and proteins levels. At higher doses (photocytotoxic doses), PFLX induced a concentration dependent decrease in cell viability of human keratinocyte cell line (HaCaT) and peritoneal macrophages of BALB/c mice. Our molecular docking suggests that PFLX binds only to the cleaved DNA in the DNA-human TOP2A complex. Topoisomerase assay confirmed that PFLX inhibits human topoisomerase by forming an adduct with DNA. Photosensitized PFLX also caused intracellular ROS mediated DNA damage and formation of micronuclei and cyclobutane pyrimidine dimers (CPDs). Increase intracellular ROS leads to apoptosis which was proved through lysosomal destabilization and reduced mitochondrial membrane potential (MMP). Our present study shows that ambient UVA exposure in the presence of PFLX caused immunomodulatory as well as photogenotoxic effects. Therefore, patients under PFLX drug treatment should avoid sunlight exposure, especially during peak hours for their photosafety.

Oral toxicity of pefloxacin, norfloxacin, ofloxacin and ciprofloxacin: comparison of biomechanical and histopathological effects on Achilles tendon in rats.

Four fluoroquinolones (pefloxacin, norfloxacin, ofloxacin and ciprofloxacin) were compared according to their biomechanical and histopathological effects on rat Achilles tendon. Wistar rats were divided into one untreated control and four treatment groups in parallel. Pefloxacin mesylate dihydrate (40 mg/kg), norfloxacin (40 mg/kg), ofloxacin (20 mg/kg) and ciprofloxacin (50 mg/kg) were administered by gavage twice daily for three consecutive weeks. 6 weeks after treatment, the test animals were euthanised and Achilles tendon specimens were collected. A computer monitored tensile testing machine was utilised for biomechanical testing. The mean elastic modulus of the control group was significantly higher than that of the norfloxacin and pefloxacin groups (p<0.05 and p<0.01, respectively). The mean yield force (YF) of the control group was significantly higher than those of ciprofloxacin, norfloxacin and pefloxacin groups (p<0.001, p<0.05 and p<0.01, respectively). The mean ultimate tensile force (UTF) of the control group was significantly higher than of the ciprofloxacin, norfloxacin, and pefloxacin groups (p<0.001, p<0.05 and p<0.01, respectively). Hyaline degeneration and fibre disarrangement were observed in the tendons of the ciprofloxacin, pefloxacin, and ofloxacin treated-groups, whereas myxomatous degeneration was observed only in the ciprofloxacin and pefloxacin groups. In conclusion, these findings in our rat model reveal significant deterioration of biomechanical parameters following fluoroquinolone exposure, and indicate significantly higher biomechanical toxicity for ciprofloxacin and pefloxacin.

Modification of innervation pattern by fluoroquinolone treatment in the rat salivary glands.

Fluoroquinolone antibiotics (FQAs) are widely used in dental and medical therapy. Despite their known severe adverse actions on the central and peripheral nervous system, little attention has been directed toward the potential toxic side effects of these compounds on the oral tissues. As the saliva secretion is controlled by the nervous system and neuropeptides, the neurotoxic effect of pefloxacin (PEF), a representative member of FQAs, was studied in rats in the present work. Previously, we demonstrated a significant weight loss of parotid gland tissue, a marked decrease in 3H-thymidine incorporation, a decreased volume of saliva and amylase activity of the glandular tissue in response to PEF. Animals received intraperitoneal injection of PEF (20 mg/100 g body weight daily) for 3 and 7 days. Normal histology, and neurofilament 200, substance P (SP) and calcitonin gene-related polypeptide (CGRP) containing nerve fibers were detected with immunohistochemical methods. A marked decrease of the weights in salivary glands and the acinar diameters were measured. Similarly, a strong and significant decrease of the number of SP and CGRP containing nerve fibers were detected. These findings suggest that the impaired morphology and innervation pattern of salivary glands is related to the neurotoxic adverse effect of FQA treatment.

Age-dependent effects on redox status, oxidative stress, mitochondrial activity and toxicity induced by fluoroquinolones on primary cultures of rabbit tendon cells.

The age-related difference in fluoroquinolone-induced tendon toxicity was investigated. In vitro tendon cells from juvenile and young adult rabbits, respectively, were incubated with quinolone (nalidixic acid, NA) or fluoroquinolone (ofloxacin, OFX or pefloxacin, PEF) at 0.01 microM to 1 mM for 72 h. Redox status, glutathione (GSH), reactive oxygen species (ROS), and mitochondrial activity were assessed using intracellular fluorescent probes. Fluorescence signal was detected on living adherent tenocytes in microplates using cold-light cytofluorometry. Tendon toxicity differed significantly between the two cell groups and the difference was greatest with highest dose (1 mM). For 72 h, significant (p < 0.001) differences between immature and young adult primary tenocytes were observed for redox status decrease, GSH decrease, and ROS production increase. Mitochondrial activity remained unaltered in immature tenocytes. We confirm two groups of intrinsic tendon toxicity (OFX/NA vs. PEF) associated to oxidative stress (GSH decrease). Our in vitro experimental model confirms the clinical observations of age dependent tenotoxicity. First group (NA, OFX) showed greater intrinsic tenotoxicity for young adult than immature tenocytes, second group (PEF) was highly toxic for immature and young adult cells. The three quinolones do not altered mitochondrial activity in immature tenocytes whereas alteration was observed in young adult tenocytes.

Mechanisms of pefloxacin-induced pain.

In electrophysiological and behavioral experiments on rats we studied the effects of pefloxacin, a member of fluoroquinolone family, on the nociceptive system. Intraperitoneal injection of pefloxacin (80 mg/kg) decreased the thresholds of nociceptive response to noxious stimulation in the hot-plate test. In addition, it decreased the threshold of the late component of nociceptive flexor reflex. Intrathecal application of pefloxacin in a dose of 20 microg provoked allodynia, while the higher dose of 400 microg induced behavioral pattern characteristic of central pain syndrome. It was hypothesized that pain induced by pefloxacin results from disturbances in GABAergic inhibition in the central subdivisions of the nociceptive system.

Extension of the isobolographic approach to interactions studies between more than two drugs: illustration with the convulsant interaction between pefloxacin, norfloxacin, and theophylline in rats.

This work proposes a model to characterize the additivity or the nonadditivity of combinations of more than two agents. Using a Bayesian framework, we modeled the variability between experimental subjects, the errors that occurred during data collection, and the relationship between effects and concentrations of agents at the effect site. The model was used to characterize the additivity (or non-additivity) of norfloxacin, pefloxacin, and theophylline in causing maximal seizures in male Sprague Dawley rats. Animals received the drugs separately or in various combinations. Drug infusion was stopped at the onset of maximal seizures, and cerebrospinal fluid samples were collected for determination of drug concentration by high-performance liquid chromatography. The model was fitted to concentration data using Markov Chain Monte Carlo techniques. Results showed that induction of seizures by mixtures of theophylline and pefloxacin were additive. Seizure induction by mixtures of norfloxacin and pefloxacin or norfloxacin and theophylline were not additive and, given the model, these drugs interacted negatively. There was no triple interaction effect between the drugs. This study demonstrates the ease with which mixtures of more than two drugs can be analyzed with the proposed model.

[Tenotoxic potential of fluoroquinolones in the choice of surgical antibiotic prophylaxis in ophthalmology]. / Prise en compte du potentiel ténotoxique des fluoroquinolones dans le choix d'une antibioprophylaxie chirurgicale en ophtalmologie.

PURPOSE: Fluoroquinolones are mainly used in ophthalmic antibiotic prophylaxis because of their broad spectrum activity and good ocular diffusion. But a single oral dose of fluoroquinolones can result in a serious source of tendinopathy and tendon rupture, especially in patients 60 years and older. It seems very important to investigate tendon toxicity of fluoroquinolones to improve the risk-benefit ratio in ophthalmologic antibiotic prophylaxis. MATERIAL: and methods: The intrinsic tenotoxic potential of four fluoroquinolones (pefloxacin, ofloxacin, ciprofloxacin, levofloxacin) was directly evaluated on living adherent tendon cells in microplates. Cell viability and reactive oxygen species production was evaluated using neutral red, alamar blue, and dichlorofluorescin diacetate tests. RESULTS: Results showed a loss of viability associated with free radical production depending on fluoroquinolone molecules. Pefloxacin appeared more tenotoxic but no study has confirmed its efficacy in surgical antibiotic prophylaxis and its use in the patient who is 60 years and older could be disputed. Ciprofloxacin is highly toxic with a low ocular diffusion and seems to be inappropriate for antibiotic prophylaxis. Ofloxacin and levofloxacin are less cytotoxic, associated with good ocular diffusion and a broad antibacterial spectrum. CONCLUSION: Ofloxacin and levofloxacin seem to be good alternatives for improving the risk-benefit ratio in surgical antibiotic prophylaxis in patients 60 years and older.

Pefloxacin-induced achilles tendon toxicity in rodents: biochemical changes in proteoglycan synthesis and oxidative damage to collagen.

Despite a relatively low incidence of serious side effects, fluoroquinolones and the fluoroquinolone pefloxacin have been reported to occasionally promote tendinopathy that might result in the complication of spontaneous rupture of tendons. In the present study, we investigated in rodents the intrinsic deleterious effect of pefloxacin (400 mg/kg of body weight) on Achilles tendon proteoglycans and collagen. Proteoglycan synthesis was determined by measurement of in vivo and ex vivo radiosulfate incorporation in mice. Collagen oxidative modifications were measured by carbonyl derivative detection by Western blotting. An experimental model of tendinous ischemia (2 h) and reperfusion (3 days) was achieved in rats. Biphasic changes in proteoglycan synthesis were observed after a single administration of pefloxacin, consisting of an early inhibition followed by a repair-like phase. The depletion phase was accompanied by a marked decrease in the endogenous serum sulfate level and a concomitant increase in the level of sulfate excretion in urine. Studies of ex vivo proteoglycan synthesis confirmed the in vivo results that were obtained. The decrease in proteoglycan anabolism seemed to be a direct effect of pefloxacin on tissue metabolism rather than a consequence of the low concentration of sulfate. Pefloxacin treatment for several days induced oxidative damage of type I collagen, with the alterations being identical to those observed in the experimental tendinous ischemia and reperfusion model. Oxidative damage was prevented by coadministration of N-acetylcysteine (150 mg/kg) to the mice. These results provide the first experimental evidence of a pefloxacin-induced oxidative stress in the Achilles tendon that altered proteoglycan anabolism and oxidized collagen.

Adverse testicular effects of some quinolone members in rats.

In the last few years, a marked decrease in male fertility has been reported. Environmental factors were recently suspected for this effect. Among those factors is the misuse of drugs and in particular antibiotics. Quinolones are a group of antibacterial agents with broad-spectrum activity. Testicular impairment of some quinolone members is controversial; a matter which stimulated our attention to investigate the adverse testicular effects of the most familiar quinolone members, namely: ofloxacin, ciprofloxacin and pefloxacin. They were given to rats in doses of 72, 135 and 72 mg kg(-1) day(-1) p.o., respectively, for 15 consecutive days. Ofloxacin was also used to establish a dose-response relationship in doses of 36, 72 and 360 mg kg(-1) day(-1) p.o. for 15 consecutive days. Results revealed that ofloxacin, ciprofloxacin and pefloxacin reduced testicular LDH-X activity by 39.8%, 62.7% and 60.7%, respectively. Moreover, sperm count, motility and daily sperm production were markedly decreased. Ofloxacin induced a dose-dependent decrease in testicular LDH-X activity, sperm count and motility. Furthermore, daily sperm production showed a marked reduction which amounted to 26.1% and 40. 0% following administration of ofloxacin (72, 360 mg kg(-1) day(-1) x 15 days), respectively. Moreover, administration of ofloxacin resulted in marked testicular histopathological changes. It is concluded that, ofloxacin, ciprofloxacin and pefloxacin significantly impaired both testicular function and structure in rats.

A new approach for early assessment of the epileptogenic potential of quinolones.

The epileptogenic potential of pefloxacin and norfloxacin, two quinolone antibiotics, was investigated in vivo in three different animal species by measuring drug concentrations in cerebrospinal fluid (CSF), which is part of the biophase, at the onset of convulsions. Interestingly, the pefloxacin-to-norfloxacin concentration ratios in CSF were virtually constant across the species (7.0, 6.6, and 6.0 in mice, rats, and rabbits, respectively), suggesting that this approach could be used to predict the relative epileptogenic potential of quinolones in humans.

Development of a new quantitative approach for the isobolographic assessment of the convulsant interaction between pefloxacin and theophylline in rats.

PURPOSE: A new mathematical approach was developed to quantify convulsant interaction between pefloxacin and theophylline in rats. METHODS: Animals received each compound separately or in different combination ratios. Infusion was stopped at the onset of maximal seizures. Cerebrospinal fluid (CSF) and plasma samples were collected for HPLC drug determination. The nature and intensity of the pharmacodynamic (PD) interaction between drugs was assessed with a new modeling approach which includes (a) data transformation to create an essentially error-free X-variable and (b) estimation of an interaction parameter a by fitting a nonlinear hyperbolic model to the combination data with unweighted nonlinear regression. RESULTS: Drug disposition to the biophase was linear within the range of administered doses. The estimates of a suggested a Loewe antagonistic interaction between pefloxacin and theophylline at the induction of maximal seizures in rats. Similar intensity of PD interaction was observed at the dose and biophase level (alpha was -0.415 +/- 0.069 and -0.567 +/- 0.079, respectively). CONCLUSIONS: The suitability of the proposed model was assessed by Monte Carlo simulation. This new mathematical approach enabled the characterization of the Loewe antagonistic nature of the PD (convulsant) interaction between pefloxacin and theophylline, whereas previously used methodologies failed to do so.

Interaction between pefloxacin and aminophylline in genetically epilepsy-prone rats.

The effects of a chronic treatment with pefloxacin on aminophylline-induced seizures in genetically epilepsy-prone rat have been investigated. Two series of experiments were performed. In the first, animals received pefloxacin orally twice a day for five days, then were administered aminophylline intraperitoneally and the occurrence of seizures was evaluated. In the second series of experiments, theophylline serum concentration was evaluated in rats subject to the same experimental protocol. Pefloxacin significantly, and in a dose-dependent manner, increased the occurrence of seizure phases induced by aminophylline, but did not influence theophylline serum levels measured at different times after the injection of aminophylline. We suggest that additive neurotoxic effects of both pefloxacin and aminophylline might contribute to the increased severity of seizure score. The possible role of GABA-benzodiazepine, excitatory amino acid and purinergic mechanism, and the role of pharmacokinetic factors are discussed.

Effects of fluoroquinolones on cultured articular chondrocytes flow cytometric analysis of free radical production.

Using flow cytometry, we previously established in an ex vivo model that fluoroquinolones induce a stimulation of the oxidative metabolism in immature chondrocytes. To assess these findings in an in vitro model, primary cultures of immature articular chondrocytes were incubated with four quinolone solutions: ofloxacin, ciprofloxacin, nalidixic acid at 10 micrograms/ml for 24 hr and pefloxacin at 1, 10 and 100 micrograms/ml for various periods of time (2, 4, 6, 12, 24 and 48 hr). Three fluorochromes were used: DCFH-DA, reflecting cellular production of H2O2, rhodamine 123 (Rh123) and 10-N-nonyl-acridine orange (NAO), which are specific for mitochondrial activity and mass, respectively. In immature chondrocyte cultures treated with pefloxacin, ofloxacin and nalidixic acid at 10 micrograms/ml for 24 hr, levels of cellular fluorescent dichlorofluorescein DCF (oxidized form of DCFH-DA) were significantly higher than in control cells. No significant increase could be registered with ciprofloxacin. In the same experimental conditions, incorporation of Rh123 and NAO was not significantly modified. Pefloxacin (10 micrograms/ml, 24 hr) did not induce any significant increase of DCFH-DA processing either in mature chondrocytes or in alveolar macrophages removed from immature rabbits. Quinolones induce in vitro an early stimulation of the oxidative metabolism in immature but not in mature chondrocytes, a phenomenon that could explain juvenile onset of quinolone arthropathy. This in vitro model could be proposed as an easy and reproducible method for screening potential arthrotoxicity of antimicrobial agents, capable of stimulating the formation of H2O2.

Cytofluorometric analysis of chondrotoxicity of fluoroquinolone antimicrobial agents.

To better understand quinolone-related arthropathy, we conceived an experimental ex vivo model using cell cultures of articular chondrocytes issued from pretreated New Zealand White rabbits (NZW). Juvenile (4- to 5-week-old) NZW were orally dosed with ofloxacin or pefloxacin (300 mg/kg of body weight for 1 day) or with pefloxacin (300 mg/kg for 7 days). Adult (5-month-old) NZW were treated with pefloxacin (300 mg/kg for 1 day). Chondrocytes were enzymatically recovered from cartilage and were analyzed by cytofluorometry using 2',7'-dichlorofluorescein diacetate (DCFH-DA) and dihydrorhodamine 123 (DHR), reflecting cellular respiratory-burst activity, and rhodamine 123 (Rh123) and 10-N-nonyl-acridine orange (NAO), specific for the mitochondrial activity and mass, respectively. A significant increase in the respiratory burst was detected by DCFH-DA and DHR in all treated groups of young animals, compared with untreated control groups. No significant increase of respiratory burst was noted in older treated rabbits. The 7-day treatment resulted in a decrease in mitochondrial uptake of Rh123 and an increase in NAO uptake. Fluoroquinolone arthrotoxicity seems to involve in its early phase the respiratory burst of immature articular chondrocytes.

Kinetics and tolerability of intravitreal pefloxacin in rabbits.

In the treatment of patients with bacterial endophthalmitis, the intravitreal administration of antibiotics is suitable for induction therapy since it provides immediate high concentrations in the vitreous humor. Pefloxacin has been shown to have good intraocular penetration when given systemically. In order to extend the potential routes of administration of this agent, we have assessed the kinetics and toxicity of pefloxacin in rabbit phakic eyes following intravitreal instillation. Kinetic parameters were determined for 12 albino and 12 pigmented rabbits after a single injection of 80 micrograms. Pefloxacin was undetectable in the aqueous humor but high concentrations were found in the chorioretina. The vitreal half-life was short (3 h). These results were consistent with posterior elimination via the chorioretina. Pefloxacin concentrations in the iris and chorioretina of pigmented rabbits were two-fold greater than those in albino rabbits, probably because of binding to the pigmentary apparatus. Toxicity studies, including ophthalmological and histopathological investigations, identified a maximum non-toxic dosage of 400 micrograms. Intravitreal pefloxacin may therefore be suitable for induction therapy in patients with endophthalmitis, although further studies in primates are required to confirm the efficacy and tolerability of this route of administration.

Central nervous system toxicity of quinolones: human and animal findings.

Quinolones have been reported to induce CNS reactions in 0.9-2.1% of cases, but severe reactions occur in less than 0.5%. Flumequine and fleroxacin, but not other quinolones, have produced convulsions in animals after systemic administration; by interventricular injection convulsions could be produced by some quinolones, but by pefloxacin only when a dose of 400 micrograms was reached. A possible mechanism for CNS excitation may be the displacement of GABA from receptors. Quinolones may interact with other drugs--theophylline, caffeine, non-steroidal anti-inflammatory drugs--in producing CNS effects.

Cytotoxicity and uptake of pefloxacin, ciprofloxacin, and ofloxacin in primary cultures of rat hepatocytes.

The cytotoxicity and the uptake of three 4-quinolones--pefloxacin, ciprofloxacin, and ofloxacin--were investigated in primary cultures of rat hepatocytes. As assessed by intracellular enzyme release in culture media, pefloxacin at concentration 400 mg/l and ciprofloxacin at 200 mg/l were found to be hepatotoxins. However, concentration ofloxacin up to 400 mg/l were not hepatotoxic. After 48 h incubation, the remaining antibiotic concentrations in the culture medium as determined by HPLC were 55%, 45%, and 35% for ofloxacin, ciprofloxacin, and pefloxacin, respectively. Comparisons of quinolone concentrations in the hepatocyte culture medium determined by HPLC and correspondent antibacterial activities determined by microbiological assays did not reveal any metabolites with antibacterial activity other than those which have been identified from intact animals.