Abstract
Background and Purpose: Novobiocin (NOVO), an ABC transporter inhibitor, decreases intestinal wall permeability of capsaicin (CAP), an ABC transporter substrate. However, the mechanism of this effect is not consistent with the action of NOVO as an ABC transporter inhibitor. We previously found that CAP can also be transported via TRPV1, which was site-specific in the permeability of CAP across the intestine. We explored the regulation by NOVO of TRPV1 in the present study.Methods: Rats and transfected IEC-6 cells were used as the models to assess intestinal permeability and expression of TRPV1. Ussing chamber and intracellular accumulation were used to evaluate the influence of NOVO on the transport of CAP in vitro. The expression of TRPV1 was detected after administration of NOVO by qRT-PCR, western blot and immunofluorescent imaging. In addition, MTT and lactate dehydrogenase (LDH) were used to evaluate the cytotoxicity of NOVO in both rat and cell models. Finally, the effect of NOVO on the absorption of CAP in vivo was studied by LC-MS/MS.Results: In vitro data showed that there existed a dose-dependent relationship in the range of concentration between 5 and 50 μM, and even 5 μM NOVO could decrease intestinal permeability of CAP across the intestine. Meanwhile, cytosolic accumulation of CAP decreased when NOVO was used simultaneously or 24 h in advance. NOVO exhibited an inhibition level similar to that of ruthenium red (RR) or SB-705498, a TRPV1-specific inhibitor. NOVO down-regulated TRPV1 expression in the intestine and in transfected cells in a concentration-dependent fashion, hinting that its inhibition of the permeability of CAP is due to its inhibition of TRPV1 expression. Immunofluorescent imaging data showed that the fluorescence intensity of TRPV1 was reduced after pre-treatment with NOVO and SB-705498. In vivo data further demonstrated that oral co-administration of NOVO decreased Cmax and AUC of CAP in dosage-dependent ways, consistent with its role as a TRPV1 inhibitor.Conclusion: NOVO could be a potential TRPV1 inhibitor by attenuating the expression of TRPV1 and may be used to attenuate permeability of TRPV1 substrates.
Highlights
Background and PurposeNovobiocin (NOVO), an ATP-binding cassette protein family (ABC) transporter inhibitor, decreases intestinal wall permeability of capsaicin (CAP), an ABC transporter substrate
In vivo data further demonstrated that oral co-administration of NOVO decreased maximum plasma concentration (Cmax) and Area Under Curve (AUC) of CAP in dosage-dependent ways, consistent with its role as a Transient receptor potential vanilloid member 1 (TRPV1) inhibitor
In order to determine whether the transport of NOVO across the intestine was related to TRPV1, we evaluated how ruthenium red (RR) affected the transport of NOVO
Summary
Rats and transfected IEC-6 cells were used as the models to assess intestinal permeability and expression of TRPV1. Ussing chamber and intracellular accumulation were used to evaluate the influence of NOVO on the transport of CAP in vitro. The expression of TRPV1 was detected after administration of NOVO by qRT-PCR, western blot and immunofluorescent imaging. MTT and lactate dehydrogenase (LDH) were used to evaluate the cytotoxicity of NOVO in both rat and cell models. The effect of NOVO on the absorption of CAP in vivo was studied by LC-MS/MS. CAP (lot number: 048K5058V), HEPES (lot number: 016K54331), and Trizma base (lot number: 103K5411) were obtained from Sigma–Aldrich Chemical Co. HPLC-grade methanol and acetonitrile were purchased from Merck (Darmstach, Germany). SB-705498 was purchased from Skelleck Chemicals LCC (Houston, TX, United States).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
