| To identify the localization and expression of BCRP in tissues of liver, duodenum, ileum and jejunum of broilers and provide basic theory for more information of the function of BCRP, immunohistochemical, real time RT-PCR and Western blot methods were used to detect the distribution and expression of BCRP at mRNA and protein levels in tissues of broilers. The results showed that BCRP was localized in hepatocytes in liver and in epithelial cells of duodenum, jejunum and ileum. BCRP mRNA and protein could express in all tested tissues with relatively high levels in liver and all intestinal segments except duodenum.The expression of BCRP could be affected by age of the broilers. So in this study, real time RT-PCR and Western blot were used with β-actin as house-keeping gene and house-keeping protein to detect the expression of BCRP in tissues of liver, duodenum, ileum and jejunum among broilers at different ages of0,1,2,4,6and8weeks old. The results showed that the expression level of BCRP mRNA in liver increased from0to2weeks old, and decreased a little thereafter. BCRP mRNA of2and6weeks old was extremely significantly higher than that of0week old(P=0.001, P=0.01), and BCRP mRNA of4week old was significantly up regulated compared with that of0week old(P=0.026).The expression level of BCRP mRNA in duodenum decreased from0week old, and levels of BCRP mRNA of2,4,6and8weeks old were significantly lower than that of0week old (P=0.000, P=0.002, P=0.000, P=0.000).The expression levels of BCRP mRNA in jejunums of1,4,8week-old broilers were extremely higher than that of0week(P=0.000, P=0.01, P=0.000), and the level of BCRP mRNA expression in jejunums of2week-old was significantly higher than that of0week-old broilers (P=0.029). In ileum, the level of BCRP mRNA expression came to peak at1week-old, which was extremely significantly higher than that of0week-old(P=0.004), and showed a decreasing trend from1week-old. While result of Western blot showed that among different ages, there is not significant changes of protein expression of BCRP. And there was no significant correlation between the changes of mRNA and protein of BCRP during the growth of broiders.Also, research about other factors including E.coli infection, coccidian and ENRO were done in this study.The same methods were used to detect mRNA and protein of expression BCRP after the chickens were infected by E.coli, coccidian and feeden with enrofloxacin orally respectively. The result showed that the expression level of BCRP mRNA in ileum was up regulated extremely significantly (P=0.009) without significant changes in liver, duodenum and jejunum. The results of Western blot showed that the expression of BCRP in duodenum was increased significantly (P=0.043) without significant changes in other tissues. BCRP mRNA expression levels in liver, ileum and jejunum were down regulated significantly (P=0.021, P=0.021, P=0.021) in broilers infected with coccidian. Though it was up regulated in duodenum, there is not significant change (P>0.05). Results of Western blot showed that there presence a trend of decreasing of BCRP protein expression level in liver, ileum and jejunum. However, the expression level was increased in duodenum. The tendency of protein expression level is consistent with that of mRNA expression level. BCRP mRNA expression level in duodenum of broiler could be significantly increased after orally administration with ENRO (P=0.009), while no significant changes in other tissues. Result of Western blot showed that ENRO by orally administration did not significantly affect the protein expression level of BCRP in all tested tissues. All results indicated that colibacillosis and coccidiosis as well as ENRO could affect the expression level of BCRP mRNA with different degree, while could not affect the expression level of BCRP protein. |
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