Claudins are essential for tight junction formation and paracellular transport, and affect key cellular events including proliferation and migration. The properties of tight junctions are dynamically modulated by a variety of inputs. We previously showed that the inflammatory cytokine Tumor Necrosis Factor-α (TNFα), a major pathogenic factor in kidney disease, alters epithelial permeability by affecting the expression of claudin-1, 2 and 4 in kidney tubular cells. Here we explored the effect of TNFα on claudin-3 (Cldn-3), a ubiquitous barrier-forming protein. We found that TNFα elevated Cldn-3 protein expression in tubular epithelial cells (LLC-PK1 and IMCD3) as early as 3h post-treatment. Bafilomycin A and Bortezomib, inhibitors of lysosomal and proteasomes, respectively, reduced Cldn-3 degradation. However, TNFα caused a strong upregulation of Cldn-3 in the presence of bafilomycin, suggesting an effect independent from lysosomes. Blocking protein synthesis using Cycloheximide prevented Cldn-3 upregulation by TNFα, verifying the contribution of de novo Cldn-3 synthesis. Indeed, TNFα elevated Cldn-3 mRNA levels at early time points. Using pharmacological inhibitors and siRNA-mediated silencing we determined that the effect of TNFα on Cldn-3 was mediated by extracellular signal regulated kinase (ERK)-dependent activation of NFκB and protein kinase A (PKA)-induced activation of CREB1. These two pathways were turned on by TNFα in parallel and both were required for the upregulation of Cldn-3. Since Cldn-3 was suggested to modulate cell migration and epithelial-mesenchymal transition, and TNFα were shown to affect these processes, Cldn-3 upregulation may modulate regeneration of the tubules following injury.
Comparison of the effect of lipopolysaccharide on tumor necrosis factor α (TNF-α) secretion and TNF and TNFR1 mRNA levels in feline endometrium throughout the estrous cycle during pyometra and after medroxyprogesterone acetate treatment