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Mahesh Shivanna, Gangaraju Rajashekhar, Sangly P. Srinivas; Barrier Dysfunction of the Corneal Endothelium in Response to TNF-α: Role of p38 MAP Kinase. Invest. Ophthalmol. Vis. Sci. 2010;51(3):1575-1582. doi: https://doi.org/10.1167/iovs.09-4343.
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TNF-α is elevated in the cornea and aqueous humor during allograft rejection and anterior uveitis. The authors investigated the involvement of p38 MAP kinase in the TNF-α–induced loss of barrier integrity in monolayers of cultured bovine corneal endothelial cells.
Transendothelial electrical resistance (TER), a measure of barrier integrity, was determined by electrical cell-substrate impedance sensing. Barrier integrity was further assessed in terms of permeability to FITC dextran. Reorganization of the apical junctional complex (AJC) in response to TNF-α was visualized by immunofluorescence. The expression of TNF-α receptors was confirmed by RT-PCR. Activation of p38 MAP kinase in response to TNF-α was determined by Western blot analysis.
Exposure to TNF-α induced a continuous decline in TER that persisted for more than 20 hours. It also led to a significant increase in permeability to FITC dextran. At the AJC, the cytokine caused disassembly of microtubules, disruption of perijunctional actomyosin ring (PAMR), and dislocation of ZO-1 and cadherins. Western blot analysis showed that TNF-α also led to the activation of p38 MAP kinase. All these responses to the cytokine were opposed by treatment with SB-203580, a selective p38 MAP kinase inhibitor. TNFR1, but not TNFR2, was expressed in untreated cells with no change in the expression pattern on treatment with the cytokine.
TNF-α breaks down the barrier integrity of corneal endothelium, concomitant with the disruption of PAMR, remodeling of AJC, and disassembly of microtubules. These effects are mediated by transient activation of p38 MAP kinase. Thus, the TNF-α–induced barrier dysfunction in the corneal endothelium can be suppressed by inhibitors of p38 MAP kinase and agents downstream of the kinase that affect the cytoskeleton.
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