Takumi Tomono

Contribution of Radixin to P-Glycoprotein Expression and Transport Activity in Mouse Small Intestine In Vivo

The ERM proteins, ezrin, radixin, and moesin, are membrane-cytoskeleton cross-linkers with multiple physiological functions. We previously showed that radixin is involved in posttranslational regulation of P-glycoprotein (P-gp) in human hepatoblastoma HepG2 cells. Here, we investigated the physiological role of radixin in regulating P-gp expression and activity in the small intestine by comparing wild-type- and radixin knockout (Rdx) mice. In intestinal tissue homogenates, P-gp protein levels increased markedly from the upper part to the lower part of the small intestine in both wild-type- and Rdx(-/-) mice. In the membrane fractions, a similar pattern was seen in wild-type mice. However, the membrane expression of P-gp protein remained at the same level from the upper to the lower part of the small intestine in Rdx(-/-) mice. When rhodamine123 (Rho123), a substrate of P-gp, was orally administered to Rdx(-/-) and wild-type mice, the absorption phase of Rho123 was greater in Rdx(-/-) than in wild-type mice, whereas the elimination phase in Rdx(-/-) mice was not different from that of wild-type mice. Our results indicate that radixin plays an important role in regulating P-gp localization and P-gp functional activity at the intestinal membrane.

Entinostat reverses P-glycoprotein activation in snail-overexpressing adenocarcinoma HCC827 cells

Epithelial-to-mesenchymal transition (EMT) in cancer cells facilitates tumor progression by promoting invasion and metastasis. Snail is a transcriptional factor that induces EMT, while P-glycoprotein (P-gp) is an efflux transporter involved in anticancer drug resistance, and P-gp efflux activity is stimulated in Snail-overexpressing lung cancer cells with EMT characteristics. Since the histone deacetylase (HDAC) inhibitor entinostat (Ent) reverses EMT features, our aim in this study was to determine whether Ent also suppresses P-gp activation in Snail-induced cells. First, we confirmed that Ent treatment reduced migration activity, downregulated E-cadherin and upregulated vimentin at the mRNA level in Snail-overexpressing cells, thus inhibiting EMT. Efflux and uptake assays using rhodamine123 (Rho123), a fluorescent P-gp substrate, showed that Ent also inhibited Snail-induced activation of P-gp. Moreover, P-gp activity was more strongly inhibited by Ent in Snail-overexpressing cells than in Mock cells. When we evaluated the uptakes of Rho123 by LLC-PK1 cells and P-gp-overexpressing LLC-GA5COL150 cells, Rho123 accumulation in LLC-GA5COL150 cells was significantly decreased compared with that in LLC-PK1 cells. Coincubation with Ent had no effect on Rho123 accumulation in either of the cell lines. Thus, Ent appears to be an inhibitor, but not a substrate, of P-gp at low concentration. Our results suggest that Ent treatment might suppress not only Snail-induced cancer malignant alteration, but also P-gp-mediated multidrug resistance.

Advances in Studies of P-Glycoprotein and Its Expression Regulators

This review deals with recent advances in studies on P-glycoprotein (P-gp) and its expression regulators, focusing especially on our own research. Firstly, we describe findings demonstrating that the distribution of P-gp along the small intestine is heterogeneous, which explains why orally administered P-gp substrate drugs often show bimodal changes of plasma concentration. Secondly, we discuss the post-translational regulation of P-gp localization and function by the scaffold proteins ezrin, radixin and moesin (ERM proteins), together with recent reports indicating that tissue-specific differences in regulation by ERM proteins in normal tissues might be retained in corresponding cancerous tissues. Thirdly, we review evidence that P-gp activity is enhanced in the process of epithelial-to-mesenchymal transition (EMT), which is associated with cancer progression, without any increase in expression of P-gp mRNA. Finally, we describe two examples in which P-gp critically influences the brain distribution of drugs, i.e., oseltamivir, where low levels of P-gp associated with early development allow oseltamivir to enter the brain, potentially resulting in neuropsychiatric side effects in children, and cilnidipine, where impairment of P-gp function in ischemia allows cilnidipine to enter the ischemic brain, where it exerts a neuroprotective action.

Interaction of Peptide Transporter 1 With d-Glucose and l-Glutamic Acid; Possible Involvement of Taste Receptors

We investigated the influence of sweet and umami (savory) tastants on the intestinal absorption of cephalexin (CEX), a substrate of peptide transporter 1 (PEPT1, SLC15A1) in rats. After oral administration of glucose or mannitol to rats, CEX was administered together with a second dose of glucose or mannitol. Western blot analysis indicated that expression of PEPT1 in rat jejunum membrane was decreased by glucose, compared to mannitol. Furthermore, the maximum plasma concentration (Cmax) of orally administered CEX was reduced by glucose compared to mannitol. The effect of glucose was diminished by nifedipine, a L-type Ca(2+) channel blocker. We also found that Cmax of orally administered CEX was reduced by treatment with L-glutamic acid, compared to D-glutamic acid. Thus, excessive intake of glucose and L-glutamic acid may impair oral absorption of PEPT1 substrates.

Recommended Daily Dose of Sesame Lignans Has No Influence on Oral Absorption of P-Glycoprotein Substrates in Rats.

Sesamin (SM) and episesamin (ESM) are constituents of sesame seeds, which are used in health foods and considered to have various beneficial effects in the prevention of lifestyle-related diseases. P-Glycoprotein (P-gp) is an ATP-binding cassette transporter involved in drug absorption in the human gastrointestinal tract. A recent report indicated that SM influences P-gp-mediated drug transport. In the present study, we investigated whether SM and ESM inhibit P-gp in vitro, using Caco-2 cells and the typical P-gp substrates rhodamine123 (Rho123) and fexofenadine. SM and ESM showed no effect on accumulation of these compounds, indicating that SM and ESM do not influence P-gp function. In addition, an in vivo study using Rho123 indicated that SM and ESM do not affect absorption of P-gp substrates. Overall, these results suggest that health foods containing SM and ESM are unlikely to interact with P-gp substrates.