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Mitan R. Gokulgandhi, Megha Barot, Megan Haghnegahdar, Ashim K. Mitra; Fluoroquinolones Mediated Overexpression Of P-glycoprotein And Its Reversal By Rosemary Herbal Extract On Corneal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):325.
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Besides ocular pathogen drug resistance, multidrug resistance (MDR) represents a major obstacle to the success of fluoroquinolones (FQs) therapy. Subtherapeutic concentrations in the cornea can result from MDR associated efflux pumps. These pumps expel antimicrobial agents out of cell, leading to suboptimal eradication of microbes. Here, we have determined whether long term use of FQs (Levofloxacin, Ofloxacin and Gatifloxacin) could modify the MDR phenotype (P-glycoprotein, Pgp) on a corneal epithelial cell line (SIRC). In addition, we also demonstrate that an extract of dietary herb rosemary (Rosemarinus officinalis Labiatae), increases the intracellular accumulation of FQs by inhibiting P-gp expression in SIRC cells.
To study the impact of FQs, four cultures of SIRC cells were simultaneously initiated in Minimum Essential Medium. First culture (control) was maintained without any exposure of FQs, second with ofloxacin, third with levofloxacin and fourth with gatifloxacin at the concentration of 10 µg/ml respectively for 3 weeks. Efflux activity of Pgp was assessed by in vitro uptake studies using fluorescent (Rhodamine123, Rho123) and radiolabeled ([14C]Erythromycin) Pgp substrates. Further assessment of Pgp functional activity was performed by flow cytometry analysis. Moreover, effect of rosemary extract on Pgp expression was also examined by in vitro uptake studies.
From week one, in the presence of all FQs, concomitant enhancement in Pgp expression was observed by fluorescence and radioactive in vitro uptake studies. Significantly lower intracellular accumulation of Rho123 was also observed by flow cytometry analysis. Result further indicates that use of rosemary extract along with FQs may reverse the Pgp expression and can facilitate the entry of these fluoroquinolones more into eye to perform their antimicrobial activity.
This study confirms that long term FQs exposure to corneal cells results in over expression of Pgp efflux transporter. Pgp over expression is common phenomenon in anticancer resistance cell line; however this is first time we are showing over expression of Pgp due to fluoroquinolones resistance. Based on this finding it is suggested that strategies should be developed and implemented not only to reduce ocular pathogen resistance but also for FQs based cellular resistance. With rosemary, already established as an effective antimicrobial agent, we further support its effect on Pgp inhibition.
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