Claudins in the tight junctions of stria vascularis marginal cells
Section snippets
Materials and methods
Tissue preparation. Healthy, pigmented, adult guinea pigs of both sexes with intact Preyer’s reflex were anesthetized and subsequently killed by inhalation of 100% CO2. The cochleae were opened and the bony walls were micromechanically removed as described previously in detail [14]. The stria vascularis of each cochlear turn was transferred to a cell-tak (BD Biosciences, Bedford, USA) coated coverslip in a culture dish (35 mm diameter; Nalge Nunc International, Denmark). With the culture dish on
Tight junction protein expression
Immunoblot analyses were performed for the tight junction-proteins claudin-1 to -4 and occludin. Crude membrane fractions were used because these proteins are integral membrane proteins forming tight junction strands. All claudins were detected as a band at ∼23 kDa (Fig. 1B). The protein expression of the kidney sample served for reference (100%) since all of the claudins studied are expressed in different nephron segments as well [16].
Western blot analysis (Figs. 1A and B) and subsequent
Discussion
In the cochlea, the barrier function of epithelial tight junctions is essential for sensory transduction and the compartmentalization of the fluid-filled spaces of the cochlea. Thus, mutation of the tight junction-protein claudin-14 can lead to hereditary deafness DFNB29 [12]. There are, however, only few reports on integral tight junction proteins of the mammalian inner ear. In cultured epithelial cells from semicircular canals (without receptor organs), the intracellular tight
Acknowledgements
We thank Anja Fromm for the excellent technical assistance. The work was supported by the Deutsche Forschungsgemeinschaft (DFG) and by Funds of the Medical Faculty, Free University of Berlin.
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