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Several stressors are known to influence epithelial tight junction (TJ) integrity, but the association between DNA damage and TJ integrity remains unclear. Here we examined the effects of daunorubicin and rebeccamycin, two anti-tumor chemicals that induce DNA damage, on TJ integrity in human intestinal epithelial cells. Daunorubicin and rebeccamycin dose-dependently enhanced transepithelial electrical resistance (TER) and decreased flux of the 4 kDa FITC-dextran in Caco-2 cell monolayer. Daunorubicin- or rebeccamycin-induced enhancement of the TJ barrier function partly rescued attenuation of the barrier function by the inflammatory cytokines TNF-α and IFN-γ. Daunorubicin and rebeccamycin increased claudin-5 expression and the product was distributed in the actin cytoskeleton fraction, which was enriched with TJ proteins. Caffeine, which is an inhibitor of ataxia telangiectasia mutated protein (ATM) and ataxia telangiectasia mutated and Rad3-related protein (ATR), and the Chk1 inhibitor inhibited the TER increases induced by daunorubicin and rebeccamycin, whereas a Chk2 inhibitor did not. Treatment with Chk1 siRNA also significantly inhibited the TER increases. Induction of claudin-5 expression was inhibited by Chk1 inhibitor and by siRNA treatment. Our results suggest that Chk1 activation by daunorubicin and rebeccamycin induced claudin-5 expression and enhanced TJ barrier function in Caco-2 cell monolayer, which suggests a link between DNA damage and TJ integrity in the human intestine.

Efficient vaccine delivery to mucosal tissues including mucosa-associated lymphoid tissues is essential for the development of mucosal vaccine. We previously reported that claudin-4 was highly expressed on the epithelium of nasopharynx-associated lymphoid tissue (NALT) and thus claudin-4-targeting using C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE) effectively delivered fused antigen to NALT and consequently induced antigen-specific immune responses. In this study, we applied the C-CPE-based vaccine delivery system to develop a nasal pneumococcal vaccine. We fused C-CPE with pneumococcal surface protein A (PspA), an important antigen for the induction of protective immunity against Streptococcus pneumoniae infection, (PspA-C-CPE). PspA-C-CPE binds to claudin-4 and thus efficiently attaches to NALT epithelium, including antigen-sampling M cells. Nasal immunization with PspA-C-CPE induced PspA-specific IgG in the serum and bronchoalveolar lavage fluid (BALF) as well as IgA in the nasal wash and BALF. These immune responses were sufficient to protect against pneumococcal infection. These results suggest that C-CPE is an efficient vaccine delivery system for the development of nasal vaccines against pneumococcal infection.