Aminoglycosides are clinically essential for treating life-threatening Gram-negative bacterial infections, e.g., meningitis, and in preventing infection in burns and premature babies. However, there is a high incidence of ototoxicity and sensory hair cell death. Sensitive hair cell function is dependent on a structurally and physiologically intact ear. Thus, studies to determine mechanisms of ototoxicity have used intact animals, or inner ear explants excised from animals. There are a number of functional, toxicological, and pharmacological similarities between inner ear cells and cells of the kidney tubules. To exploit these similarities, we have developed cloned sub-lines of currently-available kidney cell lines to determine their validity as in vitro models to study the mechanisms of aminoglycoside transport and toxicity in the inner ear. Using the aminoglycoside gentamicin, we propose to use cloned kidney cells to study two distinct aspects of aminoglycoside ototoxicity: 1) Like hair cells, proximal tubule epithelial cells of the kidney are among the few cell types that are toxicologically sensitive to clinical doses of aminoglycosides. These cell types also share pharmacological sensitivities to several other drugs. We will use a clone of OK cells, a proximal kidney tubule line, to determine the feasibility of using proximal tubule cells as valid in vitro models of aminoglycoside uptake and toxicity by hair cells. 2) Little work has been done to determine the mechanism of drug entry into the endolymph. Research suggests that hair cell uptake of aminoglycosides is across the apical membrane that is exposed only to the highly-regulated endolymph. Thus, pathways of drug entry into the endolymph will be highly-potent sites of intervention to prevent ototoxicity during treatment. The various epithelia surrounding the endolymph are cumbersome to acquire as explants in the quantity and proper configurations to facilitate direct examination of aminoglycoside transit across these epithelia. Therefore, we have developed a kidney distal tubule cell line to use as a model for aminoglycoside transport across the various epithelia enclosing the endolymph compartment. Distal tubule cells share several characteristics with inner ear epithelia such as the marginal cells of the stria vascularis. We will use a clone of MDCK cells, a distal tubule line, to determine the feasibility of using distal tubule cells as valid in vitro models for aminoglycoside transport across the epithelia enclosing the scala media. Results obtained from both studies will periodically be compared with animal models for confirmation. Preliminary data obtained with use of our cloned kidney cell lines have already dramatically changed our understanding of aminoglycoside entry into, distribution within, and transit across epithelia. Both of these in vitro models should provide powerful tools for finding co-therapeutics to reduce or eliminate ototoxicity during aminoglycoside treatment, blocking both toxic events in the hair cells and drug access to the endolymph and, thus, the hair cells.
| Koo, Ja-Won; Quintanilla-Dieck, Lourdes; Jiang, Meiyan et al. (2015) Endotoxemia-mediated inflammation potentiates aminoglycoside-induced ototoxicity. Sci Transl Med 7:298ra118 |
| Wang, Tian; Yang, Yu-Qin; Karasawa, Takatoshi et al. (2013) Bumetanide hyperpolarizes madin-darby canine kidney cells and enhances cellular gentamicin uptake by elevating cytosolic Ca(2+) thus facilitating intermediate conductance Ca(2+)--activated potassium channels. Cell Biochem Biophys 65:381-98 |
| Myrdal, Sigrid E; Steyger, Peter S (2005) TRPV1 regulators mediate gentamicin penetration of cultured kidney cells. Hear Res 204:170-82 |
| Myrdal, Sigrid E; Johnson, Katherine C; Steyger, Peter S (2005) Cytoplasmic and intra-nuclear binding of gentamicin does not require endocytosis. Hear Res 204:156-69 |
