Evaluating synergy between marbofloxacin and gentamicin in Pseudomonas aeruginosa strains isolated from dogs with otitis externa

The aim of this study was to determine antimicrobial susceptibility of Pseudomonas aeruginosa strains to marbofloxacin and gentamicin, and investigate the possible synergistic, additive, indifferent or antagonistic effects between the two agents. P. aeruginosa strains can develop resistance quickly against certain antibiotics if used alone, thus the need emerges to find synergistic combinations. A total of 68 P. aeruginosa strains isolated from dogs were examined. In order to describe interactions between marbofloxacin and gentamicin the checkerboard microdilution method was utilized. The MICs (minimum inhibitory concentrations) for marbofloxacin and gentamicin were in the range 0.25–64 mg/L and 0.25–32 mg/L, respectively. The combination of marbofloxacin and gentamicin was more effective with a MIC range of 0.031–8 mg/L and a MIC90 of 1 mg/L, compared to 16 mg/L for marbofloxacin alone and 8 mg/L for gentamicin alone. The FIC (fractional inhibitory concentration) indices ranged from 0.0945 (pronounced synergy) to 1.0625 (indifference). Synergy between marbofloxacin and gentamicin was found in 33 isolates. The mean FIC index is 0.546, which represents a partial synergistic/additive effect close to the full synergy threshold. In vitro results indicate that marbofloxacin and gentamicin as partially synergistic agents may prove clinically useful in combination therapy against P. aeruginosa infections. Although marbofloxacin is not used in the human practice, the interactions between fluoroquinolones and aminoglycosides may have importance outside the veterinary field

Gentamicin sulphate permeation through porcine intestinal epithelial cell monolayer

Gentamicin is an aminoglycoside antibiotic widely used in combination with dimethyl sulphoxide (DMSO) in topical drug formulations. It is not known, however, whether DMSO can enhance the permeation of gentamicin through biological membranes, leading to oto- and nephrotoxic side effects. A simple and reliable high-performance liquid chromatographic (HPLC) method was applied for the quantitative determination of gentamicin collected from the apical and basolateral compartments of the porcine intestinal epithelial cell line IPEC-J2 cell monolayer using fluorometric derivatisation of the analyte with fluorenylmethyloxycarbonyl chloride (FMOC) prior to chromatographic run in the presence and absence of 1% DMSO. The lack of change in transepithelial electrical resistance (TER) demonstrated that gentamicin and 1% DMSO did not affect IPEC-J2 cell monolayer integrity via the disruption of cell membranes. Chromatographic data also ascertained that gentamicin penetration across the cell monolayer even in the presence of 1% DMSO was negligible at 6 h after the beginning of apical gentamicin administration. This study further indicates that the addition of this organic solvent does not increase the incidence of toxic effects related to gentamicin permeation

Reinforced Epithelial Barrier Integrity via Matriptase Induction with Sphingosine-1-Phosphate Did Not Result in Disturbances in Physiological Redox Status

Objectives. The relationship among matriptase function, cellular redox status, and maintenance of intestinal barrier integrity has not been established yet. The aim of this study is to reveal if the crosstalk between matriptase activators and intestinal epithelial monolayers can lead to perturbations in physiological redox regulation in vitro. Methods. The effects of suramin and sphingosine-1-phosphate (S1P) were tested on viability of intestinal porcine epithelial IPEC-J2 cells using MTS assay. Measurements of transepithelial electrical resistance (TER) were performed to determine changes in barrier integrity of cell monolayers. Amplex Red assay was used to monitor extracellular hydrogen peroxide production. Occludin distribution pattern was detected prior to and after matriptase activation using immunofluorescent staining technique. Results. TER reduction was observed in suramin-treated IPEC-J2 cell monolayers, which could be attributed to cell cytotoxic properties of 48 hr 50 μM suramin administration. In contrast, S1P treatment increased TER significantly and elevated occludin accumulation in tight junctions. It was also found that extracellular hydrogen peroxide levels were maintained in IPEC-J2 cells exposed to matriptase activators. Discussion. S1P administration not accompanied by redox imbalance might be one of the key strategies in the improvement of barrier function and consequently in the therapy of intestinal inflammations

Expression of claudin-1 in canine peripheral nerve sheath tumours and perivascular wall tumours. Immunohistochemical study

Aims: A peripheral nerve sheath tumour consists of neoplastic Schwann cells or perineurial cells, or a mixture of Schwann cells, perineurial cells and fibroblasts. The first aim of the present study was to characterise the expression of the claudin-1 tight junction protein in canine intact peripheral nerves, canine benign peripheral nerve sheath tumours (cBPNSTs), such as schwannomas, neurofibromas, perineuriomas and canine malignant peripheral nerve sheath tumours (cMPNSTs), and in different other benign and malignant canine spindle cell tumours. The second aim of the present study was to examine whether claudin-1 can help to distinguish the subgroups of canine perivascular wall tumours. Methods and results: The biopsy and necropsy samples (n=203) included 10 intact peripheral nerves, 20 cBPNSTs (4 schwannomas, 8 neurofibromas, 8 perineuriomas), 16 cMPNSTs, 6 psammomatous meningiomas, 6 dermatofibromas, 6 leiomyomas, 6 myxomas, 4 spindle cell hemangiomas, 2 spindle cell lipomas, 6 fibrohistiocytic nodules, 8 fibrosarcomas, 8 leiomyosarcomas, 6 myxosarcomas, 8 hemangiosarcomas, 8 anaplastic sarcomas, 8 amelanotic spindle cell melanomas, 8 histiocytic sarcomas, 8 spindle cell carcinomas, 8 myoepitheliomas, 8 complex carcinomas, 5 cardiac rhabdomyosarcomas, 4 synovial sarcomas, 5 osteosarcomas, 4 chondrosarcomas and 4 liposarcomas; 31 canine perivascular wall tumours: 10 hemangiopericytomas, 8 myopericytomas, 6 angioleiomyomas, 4 angioleiomyosarcomas, 3 angiofibromas. The immunohistochemical panel consisted of humanized antibodies: anti-claudin-1, anti-neuron specific enolase, anti-S-100 protein, anti-α-smooth muscle actin, anti-vimentin, anti-cytokeratin AE1-AE3, anti-claudin-5, anti-Melan-A and anti-heavy caldesmon, anti-calponin and anti-desmin. The intact perineurial cells, all perineuriomas, neurofibromas, cMPNSTs, spindle cell carcinomas and epithelial components of the complex carcinomas, all hemangiopericytomas and myo-pericytomas showed claudin-1 positivity. The schwannomas and other spindle shape cell tumours were negative for claudin-1. Conclusion: Our findings suggest that an antibody against claudin-1, in combination with other antibodies, can be used as a novel diagnostic tool to differentiate canine peripheral nerve sheath tumours from other fusocellular tumours, and anti-claudin-1, together with other antibodies, can also be used to subclassify cBPNSTs. Furthermore, analysis of claudin-1 expression can help to differentiate between subgroups of canine perivascular wall tumour