RESUMO
Tight junctions (TJs) maintain cellular polarity between the apical and basolateral region of epithelial cells. Claudin, a tetra-transmembrane protein, plays a pivotal role in the barrier function of TJs. We previously found that a claudin modulator, the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE), may be a promising candidate for improving the mucosal absorption of drugs. C-CPE is a fragment of enterotoxin, and putative CPE claudin receptors are highly expressed in liver and kidney. The safety and antigenicity of C-CPE must be evaluated for future clinical application. Therefore, we evaluated whether C-CPE administration in mice leads to tissue injury or production of antibodies. Intravenous administration of C-CPE at 5 mg/kg, which is a more than 25-fold higher dose than that used in a murine mucosal absorption model, did not increase biochemical markers of liver and kidney injury even after 11 injections once a week. Nasal C-CPE administration (2 mg/kg) once a week for 11 administrations also did not increase these biochemical markers, but 6 administrations of C-CPE resulted in elevation of C-CPE-specific serum IgG. These results indicate that development of a less antigenic claudin modulator will be essential for future clinical application of a C-CPE-based mucosal absorption enhancer.
Assuntos
Claudinas/efeitos dos fármacos , Enterotoxinas/toxicidade , Administração Intranasal , Animais , Claudinas/biossíntese , Relação Dose-Resposta a Droga , Enterotoxinas/química , Enterotoxinas/imunologia , Feminino , Imunoglobulina G/biossíntese , Injeções Intravenosas , Rim/efeitos dos fármacos , Rim/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Fragmentos de Peptídeos/farmacologia , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismoRESUMO
A high-performance liquid chromatographic method was developed for the rapid and sensitive determination of histidine. The method is based on separation by reversed-phase ion-pair chromatography followed by highly selective fluorescence derivatization of histidine with o-phthaldialdehyde. A linear calibration curve was obtained over the range of 0.25-200 pmol per injection (10 microl) with the coefficient of variation of 0.9% at 2 pmol (n=10) and with the detection limit (SIN=8) of 25 fmol. The method was applicable to the assay of histidine in human serum. Serum histidine values obtained by the present method were in good agreement with values obtained with an amino acid analyzer.