| P-gp, encoded by the mdrl gene, was the first mammalian member of ABC proteins discovered by Juliano and Ling in Chinese hamster ovary cells. P-gp has a wide substrate specificity and transports a number of neutral and cationic substances, including antimicrobial, antivirotic, anticancer agents, immunosuppressants, cardiac drug and so on. Except expressed in cancer cells, P-gp is highly expressed in the normal tissues. In these tissues, there is a specific staining of the apical surface of secretion and excretion cells. Drug transporters of the intestine, liver and kidney are major determinants of the pharmacokinetic profile of drugs. The expression and localization of P-gp in these tissues can limit the absorption of orally administered-drug, promote the secretion of drug into the bile and the urine. The presented studies mainly focused on the tissues of human and mouse, the data on farm animals is lacking. In this study, immunohistochemisty, real-time RT PCR and HPLC methods were used to identify the localization and expression of P-gp as well as its role in oral enrofloxacin in piglets.The localization of P-gp in healthy piglets was studied by immunohistochemisty method with mouse and rabbit anti-MDR1antibodies. The result showed that immunostaining could be observed in the bile canalicular membranes of the hepatocytes in the liver. In the kidney, P-gp immunoreactivity is obvious in the proximal and distal tubules. And in the intestines (small and large intestines), P-gp was localized in the apical membranes of the enterocytes and the epithelial cells of intestinal gland. The result of immunohistochemisty shows that the presence of P-gp in piglet tissues is very similar to humans and other animals.The expression of mdr1mRNA has been mainly studied in humans and rodents, however, a comprehensive and comparable set of expression data for the pig is lacking. In this study, real-time RT PCR method was used with gapdh as house-keeping gene to detect the expression level of mdr1mRNA. The mdrl mRNA was detected in all test tissues and the mdrl mRNA level of expression ranked from high to low in all tissues was ileum, colon, liver, jejunum, duodenum, rectum, cecum and kidney. Especially the mdrl mRNA expression level of ileum was obviously higher than those of cecum and kidney (P=0.005, P=0.001) and significantly higher than those of duodenum and rectum (P=0.017, P=0.014). Also the expression level of mdrl mRNA of kidney was significantly lower than those of jejunum, colon and liver respectively (P=0.046, P=0.018, P=0.030).The further study of the effect of P-gp on enrofloxacin pharmacokinetics in piglets was carried out with P-gp inhibitor verapamil. Eight healthy piglets weighting19～21kg were used and randomly divided into oral enrofloxacin treated (10mg/kg) and verapamil (10mg/kg) with oral enrofloxacin treated(10mg/kg) groups. Plasma enrofloxacin concentration was detected by HPLC method. The pharmacokinetic calculation of ENRO was performed using3P97. The results showed that AUC (0-2h), ka and Tpeak of enrofloxacin in verapamil treated piglets were significantly (P<0.05) different with those of enrofloxacin in non-treated piglets, which indicated that verapamil could promote the absorbtion of enrofloxacin.In conclusion, P-gp could be detected in liver, intestines and kidney at mRNA and protein level and it had an effect on enrofloxacin disposition in piglets. It requires further study on the function and modulation of P-gp in piglets. |
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